Human cgrp receptor binding antibodies

(56) References cited:

WO-A1-2007/076336 WO-A1-2010/012911

WO-A2-2006/068953 US-A1 - 2005 282 252 • TAYLOR CHRISTOPHER K ET AL: PHARMACOLOGICAL CHARACTERIZATION OF NÖVEL ALPHA-CALCITONIN GENE-RELATED PEPTIDE (CGRP) RECEPTOR PEPTIDE ANTAGONISTS THAT ARE SELECTIVE FÓR HUMÁN CGRP RECEPTORS JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS, AMERICAN SOCIETY FÓR PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS, US, vol. 319, no. 2,1 November 2006 (2006-11-01), pages 749-757, XP009084228 ISSN: 0022-3565

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DAVIS CARL D ET AL: The Tortuous Road to an Ideál CGRP Function Blocker for the Treatment of Migraine CURRENT TOPICS IN MEDICINAL CHEMISTRY, BENTHAM SCIENCE PUBLISHERS LTD, NL, vol. 8, no. 16, 1 November 2008 (2008-11-01), pages 1468-1479, XP008119816 ISSN: 1568-0266

DURHAM PAUL L: CGRP-RECEPTOR ANTAGONISTS—A FRESH APPROACH TO MIGRAINE THERAPY? NEW ENGLAND JOURNAL OF MEDICINE, MASSACHUSETTS MEDICAL SOCIETY, BOSTON, MA, US, vol. 350, no. 11, 11 March 2004 (2004-03-11), pages 1073-1075, XP009082423 ISSN: 1533-4406 • CHAUHAN M ET AL: Studies on the effects of the N-terminal domain antibodies of calcitonin receptor-like receptor and receptor activity-modifying protein 1 on calcitonin gene-related peptide-induced vasorelaxation in rat uterine artery. June 2004(2004-06), BIOLOGY OF REPRODUCTION JUN 2004, VOL. 70, NR. 6, PAGE(S) 1658 - 1663 , XP002571487 ISSN: 0006-3363 the whole document

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Description

BACKGROUND [0001] The calcitonin superfamily of peptides includes at leastfive known members: calcitonin, amylin, adrenomedullin, and two calcitonin gene-related peptides (CGRP), CGRP1 (alsó known as ctCGRP, orCGRP) and CGRP2 (alsó known as pCGRP). CGRP is a 37 amino acid vasoactive neuropeptide expressed in both the Central and peripheral nervous systems, and has been shown to be a potent vasodilator in the periphery, where CGRP-containing neurons are closely associated with blood vessels. CGRP- mediated vasodilatation is alsó associated with neurogenic inflammation, as part ofa Cascade of events that results in extravasation of plasma and vasodialation ofthe microvasculature and is present in migraine. Amylin alsó has specific binding sites in the CNS and is thought to regulate gastric emptying and have a role in carbohydrate metabolism. Adrenomedullin is a potent vasodilator. adrenomedullin has specific receptors on astrocytes and its messenger RNA is upregulated in CNS tissues that are subject to ischemia. (Zimmermann, et al., Identification of adrenomedullin receptors in cultured rat astrocytes and in neuroblastoma glioma hybrid cells (NG108-15), Brain Rés., 724:238-245 (1996); Wang et al., Discovery of adrenomedullin in rat ischemic cortex and evidence for its role in exacerbating focal brain ischemic damage, Proc. Natl. Acad. Sci. USA, 92:11480-11484 (1995)).

[0002] Calcitonin is involved in the control of boné metabolism and is alsó active in the Central nervous system (CNS). The biological activities of CGRP include the regulation of neuromuscular junctions, of antigén presentation within the immuné system, of vascular tone and of sensory neurotransmission. (Poyner, D. R., Calcitonin gene-related peptide: multiple actions, multiple receptors, Pharmacol. Ther., 56:23-51 (1992); Muff et al., Calcitonin, calcitonin gene related peptide, adrenomedullin and amylin: homologous peptides, separate receptors and overlapping biological actions, Eur. J. Endocrinol., 133: 17-20 (1995)). Three calcitonin receptor stimulating peptides (CRSPs) have alsó been identified in a number of mammalian species; the CRSPs may form a newsubfamily in the CGRP family. (Katafuchi, T and Minamino, N, Structure and biological properties of three calcitonin receptor-stimulating peptides, növel members ofthe calcitonin gene-related peptide family, Peptides, 25(11):2039-2045 (2004)).

[0003] The calcitonin superfamily peptides act through seven-transmembrane-domain G-protein-coupled receptors (GPCRs). The calcitonin receptor (CT, CTR or CT receptor) and CGRP receptors are type II (family B) GPCRs, which family includes other GPCRs that recognize regulatory peptides such as secretin, glucagon and vasoactive intestinal polypeptide (VIP). The best characterized splice variants of humán calcitonin receptor differ depending on the presence (formerly CTR_N+ or CTR1, now known as CTor absence (the major splice variant, formerly CTR_N_ or CTR₂, now known as CT(_ap of 16 amino acids in the first intracellular loop. (Gorn et al., Expression of two humán skeletal calcitonin receptor isoforms cloned from a giant cell tumorof boné: the first intracellular domain modulates ligand binding and signal transduction, J. Clin. Invest., 95:2680-2691 (1995); Hay et al., Amylin receptors: molecular composition and pharmacology, Biochem. Soc. Trans., 32:865-867 (2004); Poyner et al., 2002). The existence of at least two CGRP receptor subtypes had been proposed from differential antagonist affinities and agonist potencies in a variety of in vivő and in vitro bioassays. (Dennis et al., CGRP8-37, A calcitonin gene-related peptide antagonist revealing calcitonin generelated peptide receptor heterogeneity in brain and periphery, J. Pharmacol. Exp. Ther., 254:123-128 (1990); Dennis et al., Structure-activity profile of calcitonin gene-related peptide in peripheral and brain tissues. Evidence for multiplicity, J. Pharmacol. Exp. Ther., 251:718-725 (1989); Dumont etal., A potent and selective CGRP2 agonist, [Cys(Et)2,7]hCGRP: comparison in prototypical CGRP1 and CGRP2 in vitro assays, Can. J. Physiol. Pharmacol., 75:671-676 (1997)). [0004] The CGRP₁ receptor subtype was found to be sensitive to the antagonist fragment CGRP(8-37). (Chiba et al., Calcitonin gene-related peptide receptor antagonist humán CGRP-(8-37),Am.J. Physiol., 256: E331-E335 (1989); Dennis et al. (1990); Mimeault et al., Comparative affinities and antagonistic potencies of various humán calcitonin gene-related peptide fragments on calcitonin gene-related peptide receptors in brain and periphery, J. Pharmacol. Exp. Ther., 258:1084-1090 (1991)). By contrast, the CGRP₂ receptor was sensitive to linear humán CGRP (hCGRP) analogs, in which the cysteine residues at positions 2 and 7 were derivatized (e.g., with acetoaminomethyl [Cys(ACM)²·⁷] or ethylamide [Cys(Et)²·⁷]) bút CGRP₂ receptor was insensitive to fragment CGRP(8-37). (Dennis et al. (1989); Dennis et al. (1990); Dumont etal. (1997)).

[0005] Ligand specificity of calcitonin receptor and calcitonin-like receptor (CL, CLR or CRLR) depend on the coexpression of members of a family of accessory proteins called the receptor activity modifying proteins (RAMPs). The RAMP family includes three polypeptides (RAMP1, RAMP2 and RAMP3) that act as receptor modulators that determine the ligand specificity of receptors for the calcitonin family members. RAMPs are type I transmembrane proteins that share about 30% amino acid sequence identity and a common predicted topology, with short cytoplasmic C-termini, one trans-membrane domain and large extracellular N-termini that are responsible forthe specificity. (McLatchie etal., (1998) RAMPs regulate the transport and ligand specificity ofthe calcitonin-receptor-like receptor, Natúré, 393:333-339; Fraser et al., (1999) The amino terminus of receptor activity modifying proteins is a critical determinant of glycosylation state and ligand binding of calcitonin receptor-like receptor, Molecular Pharmacology, 55:1054-1059).

[0006] In 1998, the CGRP₁ receptor was identified as a heterodimer composed of a növel single transmembrane

ΕΡ 2 379 594 Β1 domain accessory protein, receptor activity-modifying protein 1 (RAMP1), and CRLR. (McLatchie et al., supra). Crosslinking experiments suggested the CGRP receptor consisted of a one-to-one stoichiometric arrangement of CRLR and RAMP1 (Hilairet et al. JBC 276, 42182-42190 (2001)), more recent studies using several methodologies such as BRET and BiFC revealed that the functional CGRP receptor complex may be composed of asymmetric homo-oligomer of CRLR and monomer of RAMP1 (Heroux etal. JBC 282, 31610-31620 (2007)).

[0007] A purified CRLR N-terminal domain has been shown to specifically bind ¹²⁵I-CGRP (Chauhan etal. Biochemistry 44, 782 (2005)), confirming the important and direct interaction between the CRLR with CGRP ligand. In particular, Leu 24 and Leu 34 of CRLR are believed to constitute the docking site ofthe C-terminus Phe37 of CGRP (Banerjee et al. BMC Pharmacol. 6, 9 (2006)). Furthermore, Koller et al. (FEBS Lett. 531,464-468 (2002)) obtained evidence that that the N-terminal 18 amino acid residues of CRLR contributes the selective interaction with CGRP or adrenomedullin, and Ittner et al (Biochemistry 44, 5749- 5754 (2005)) suggested that the N-terminal amino acid residues 23- 60 of CRLR mediate association with RAMP 1.

[0008] A structure-function analysis of RAMP1 identified residues 91-103, which correlate to helix 3 (Simms et al. Biophys. J. 91,662 - 669 (2006)), as potentially significant in interaction with CRLR, and residues Trp74 and Phe92 as potentially interacting with the CGRP ligand in connection with its binding to the CGRP receptor complex. Ligand binding studies using a human/rat RAMP1 chimera suggest that the binding site for certain small molecule inhibitors of CGRP R (e.g., BIBN4096BS), is located within a region which includes amino acids 66-102 of RAMP1 (Mallee et al. JBC 277, 14294 - 14298 (2002)).

[0009] CRLR has 55% overall amino acid sequence identity with CTR, although the transmembrane domains are almost 80% identícai. (McLatchie etal. (1998); Poyneret al., International unión of pharmacology. XXXII. The mammalian calcitonin gene-related peptides, adrenomedullin, amylin and calcitonin receptors, Pharmacol. Rév., 54:233-246 (2002)). [0010] CRLR has been shown to form a high affinity receptor for CGRP, when associated with RAMP1, or, to preferentially bind adrenomedullin when associated with RAMP2 or RAMP3. (McLatchie et al. (1998); Sexton et al., Receptor activity modifying proteins, Cellular Signaling, 13:73-83 (2001); Conner et al., Interaction of calcitonin-gene-related peptide with its receptors, Biochemical Society Transactions 30(Part 4): 451-454 (2002)). The glycosylation state of CRLR is associated with its pharmacology. RAMPs 1,2, and 3 transport CRLR to the plasma membráné with similar efficiencies, however RAMP1 presents CRLR as a terminally glycosylated, mature glycoprotein and a CGRP receptor, whereas RAMPs 2 and 3 present CRLR as an immature, core glycosylated adrenomedullin receptor (AM or AMR or AM receptor. (Fraser et al. (1999)). eharacterization ofthe CRLR/RAMP2 and CRLR/RAMP3 receptors in HEK293T cells by radioligand binding (¹²⁵l-adrenomedullin as radioligand), functional assay (cAMP measurement), or biochemical analysis (SDS-polyacrylamide gél eleetrophoresis) revealed them to be indistinguishable, even though RAMPs 2 and 3 share only 30% amino acid sequence identity. (Fraser et al. 1999)). Differences have been observed, however, in the pharmacology for CRLR expressed with RAMP 2 versus RAMP 3. Both CGRP and CGRP8-37, as well as adrenomedullin and the adrenomedullin-derived peptide AM 22-52, are active at the RAMP 3 heterodimer, indicating that this complex may act as both a CGRP and an AM receptor. (Howitt et al., British Journal of Pharmacology, 140:477-486 (2003); Muff et al., Hypertens. Rés., 26:S3-S8 (2003)). Co-expression of humán CRLR with rat RAMP1, and vice versa, suggested that the RAMP1 species determined the pharmacological characteristics of the CRLR/RAMP1 complex with respect to several small molecule CGRP receptor antagonists tested. (Mallee et al., Receptor Activity-Modifying Protein 1 determines the species selectivity of non-peptide CGRP receptor antagonists, J. Bioi. Chem., 277(16):14294-14298 (2002)). Unless associated with a RAMP, CRLR is nőt known to bind any endogenous ligand; it is currently the only GPCR thought to behave this way. (Conner et al., A key role for transmembrane prolines in calcitonin receptor-like agonist binding and signaling: implications for family B G-protein-coupled receptors, Molec. Pharmacol., 67(1):20-31 (2005)). [0011] Calcitonin receptor (CT) has alsó been demonstrated to form heterodimeric complexes with RAMPs, which are known as amylin receptors (AMY, AMY R or AMY receptor). Generally, CT/RAMP1 receptors (referred to as AMY^' orAMY1) have high affinity for salmon calcitonin, amylin and CGRP and lower affinity for mammalian calcitonins. For CT/RAMP2 receptors (AMY₂ or AMY2) and CT/RAMP3 receptors (AMY₃ or AMY3), a similar pattern is principally observed, although the affinity for CGRP is lower and may nőt be significant at physiologically relevant ligand concentrations. The precise receptor phenotype is dependent on cell type and CTR splice variant (CT(_a) or CT(_bp, particularly for RAMP2-generated amylin receptors. For example, a pure population of osteoclast-like cells reportedly expressed RAMP2, CTR, and CRLR, bút nőt RAMP1 or RAMP3. (Hay et al. (2004); Christopoulos et al., Multiple amylin receptors arise from receptor activity-modifying protein interaction with the calcitonin receptor gene product, Molecular Pharmacology, 56:235-242 (1999); Muff et al., An amylin receptor is revealed following co-transfection of a calcitonin receptor with receptor activity modifying proteins-1 or-3, Endocrinology, 140:2924-2927 (1999); Sexton et al. (2001); Leuthauser et al., Receptor-activity-modifying protein 1 forms heterodimers with two G-protein-coupled receptors to define ligand recognition, Biochem. J., 351:347-351 (2000); Tilakaratne et al., Amylin receptor phenotypes derived from humán calcitonin receptor/RAMP co-expression exhibit pharmacological differences dependent on receptor isoform and hőst cell environment, J. Pharmacol. Exp. Ther., 294:61-72 (2000); Nakamura etal., Osteoclast-like cells express receptor activity modifying protein 2: application of laser capture microdissection, J. Molec. Endocrinol., 34:257-261 (2005)).

ΕΡ 2 379 594 Β1 [0012] Table 1, below, summarizes the relationship ofthe receptor components discussed above.

Table 1

<td> Receptor Component</td><td> CRLR (CL)</td><td> CT (calcitonin receptor)</td>

<td> RAMP1</td><td> CGRP receptor</td><td> AMY1 receptor</td>

<td> RAMP2</td><td> AMI receptor</td><td> AMY2 receptor</td>

<td> RAMP3</td><td> AM2 receptor</td><td> AMY3 receptor</td>

[0013] Therapeutic uses of CGRP antagonists have been proposed. Noda et al. described the use of CGRP or CGRP derivatives for inhibiting platelet aggregation and for the treatment or prevention of arteriosclerosis or thrombosis. (EP 0385712 B1). Liu et al. disclosed therapeutic agents that modulate the activity of CTR, including vehicle-conjugated peptides such as calcitonin and humán aCGRP. (WO 01/83526 A2; US 2002/0090646 A1). Vasoactive CGRP peptide antagonists and their use in a method for inhibiting CGRP binding to CGRP receptors were disclosed by Smith et al.; such CGRP peptide antagonists were shown to inhibit CGRP binding to coronary artery membranes and to relax capsaicin-treated pig coronary arteries. (U.S. Pat. No. 6,268,474 B1; and U.S. Pat. No. 6,756,205 B2). Rist et al. disclosed peptide analogs with CGRP receptor antagonist activity and their use in a drug for treatment and prophylaxis ofa variety of disorders. (DE 19732944 A1).

[0014] CGRP is a potent vasodilator that has been implicated in the pathology ofa number ofvasomotor symptoms, such as all forms of vascular headache, including migraines (with or without aura) and cluster headache. Durham, N. Engl. J. Med. 350:1073-1 075, 2004. Migraine pathophysiology involves the activation ofthe trigeminal ganglia, where CGRP is localized, and CGRP levels significantly increase during a migraine attack. This in turn, promotes cranial blood véssél dilation and neurogenic inflammation and sensitization. (Doods, H., Curr. Opin. Investig. Drugs, 2:1261-1268 (2001)). Further, the serum levels of CGRP in the external jugular vein are elevated in patients during migraine headache. Goadsby et al., Ann. Neurol. 28:183-7, 1990. Intravenous administration of humán ci-CGRP induced headache and migraine in patients suffering from migraine without aura, supporting the view that CGRP has a causative role in migraine (Lassen et al, Cephalalgia 22:54-61,2002).

[0015] Migraine is a complex, common neurological condition that is characterized by severe, episodic attacks of headache and associated features, which may include nausea, vomiting, sensitivity to light, sound or movement. In somé patients, the headache is preceded or accompanied by an aura. The headache pain may be severe and may alsó be unilateral in certain patients. Migraine attacks are disruptive to daily life. In US and Western Europe, the overall prevalence of migraine sufferers is 11 % ofthe generál population (6% males; 15-18% females). Furthermore, the médián frequency of attacks in an individual is 1.5/month. While there are a number of treatments available to alleviate or reduce symptoms, preventive therapy is recommended for those patients having more than 3-4 attacks of migraine per month. Goadsby, et al. New Engl. J. Med. 346(4): 257-275, 2002. Somé migraine patients have been treated with topiramate, an anticonvulsant that blocks voltage-dependent sodium channels and certain glutamate receptors (AMPA-kainate), potentiates GABA-A receptor activity, and blocks carbonic anhydrase. The relatively recent success of serotonin 5HTI B/ID and/or 5HT-1 a receptor agonists, such as sumatriptan, in somé patients has led researchers to propose a serotonergic etiology of the disorder. Unfortunately, while somé patients respond well to this treatment, others are relatively resistant to its effects.

[0016] Possible CGRP involvement in migraine has been the basis for the development and testing of a number of compounds that inhibit release of CGRP (e.g., sumatriptan), antagonize atthe CGRP receptor (e.g., dipeptide derivative BIBN4096BS (Boehringer Ingelheim); CGRP(8-37)), or interact with one or more of receptor-associated proteins, such as, RAMP1. Brain, S. et al., Trends in Pharmacological Sciences 23:51-53, 2002. Alpha-2 adrenoceptor subtypes and adenosine Al receptors alsó control (inhibit) CGRP release and trigeminal activation (Goadsby et al., Brain 125:1392401,2002). On the other hand, treatment with compounds that exclusively inhibit neurogenic inflammation (e.g., tachykinin NKI receptor antagonists) or trigeminal activation (e.g., 5HT10 receptor agonists) appears to be relatively ineffective as acute treatments for migraine, leading somé to question whether inhibiting release of CGRP is the basis of effective anti-migraine treatments. Arulmani et al., Eur. J. Pharmacol. 500:315-330, 2004.

[0017] Although the precise pathophysiology of migraine is nőt yet well understood, the therapeutic use of CGRP antagonists and CGRP-targeting aptamers has been proposed for the treatment of migraine and other disorders. (E.g., Ölesen et al., Calcitonin gene-related peptide receptor antagonist BIBN 4096 BS for the acute treatment of migraine,

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New Engl. J. Med., 350:1104-1110 (2004); Perspective: CGRP-receptor antagonists—a fresh approach to migraine, New Engl. J. Med., 350:1075 (2004); Vater et al., Short bioactive Spiegelmers to migraine-associated calcitonin gene-related peptide rapidiy identified by a növel approach: tailored-SELEX, Nuc. Acids Rés., 31(21 e130):1-7 (2003); WO 96/03993). Further, a potent small-molecule CGRP antagonist has been shown to relieve moderate-to-severe migraine attacks, including migraine pain and migraine-associated symptoms, in a recent Phase Ili clinical trial (Connor, et al. Efficacy and Safety of telcagepant (MK-0974), a Növel Órai CGRP Receptor Antagonist, for Acute Migraine Attacks. Poster, European Headache and Migraine Trust International Congress, London, England, September 2008).

[0018] CGRP may alsó be involved in chronic pain syndromes other than migraine. In rodents, intrathecally delivered CGRP induces severe pain, and CGRP levels are enhanced in a numberof pain models. In addition, CGRP antagonists partiaiíy block nociception in acute pancreatitis in rodents (Wick, et al., (2006) Surgery, Volume 139, Issue 2, Pages 197-201). Together, these observations imply that a potent and selective CGRP receptor antagonist can be an effective therapeutic for treatment of chronic pain, including migraine.

[0019] Durham et al, New England Journal of Medicine, Massachusetts Medical Society, Boston, MA, US, 350,11, 2004,1073-1075 discloses CGRP receptor antagonists and the potential use in migraine therapy. Chauhan etal, Biology of Reproduction, 2004, 70, 1658-1663 discloses the production of rat antagonistic anti-rat, CRLR polyclonal antibodies.

SUMMARY [0020] Isolated antibodies, antigen-binding fragments thereof and other isolated antigen-binding proteins that bind CGRP R, particularly primate CGRP R, e.g., humán CGRP R, are described herein. Such isolated antigen-binding proteins may selectively inhibit primate CGRP R (as compared with primate AM1, AM2, CT or amylin receptors) and may bind both the CRLR and RAMP1 components of CGRP R. The CGRP R binding proteins were found to inhibit, interfere with, or modulate at least one of the biological responses related to CGRP R, and as such, are useful for ameliorating the effects of CGRP R-related diseases or disorders. Binding of certain antigen-binding proteins to CGRP R can, therefore, have one or more ofthe following activities: inhibiting, interfering with, or modulating CGRP R, inhibiting vasodialation, decreasing neurogenic inflammation, and alleviating, ameliorating, treating, preventing, or reducing symptoms of chronic pain or migraine.

[0021] Embodiments ofthe invention are:

1. An antibody or antigen-binding fragment thereof that binds humán CGRP receptor comprising a CDRH1, a CDRH2, a CDRH3, a CDRL1, a CDRL2 and a CDRL3, wherein:

(a) CDRL1, CDRL2, and CDRL3 have the sequence of SEQ ID NOs: 42, 43 and 44, respectively, and CDRH1, CDRH2, and CDRH3 have the sequence of SEQ ID NOs: 73, 74 and 75, respectively;

(b) CDRL1, CDRL2, and CDRL3 have the sequence of SEQ ID NOs: 45, 46 and 47, respectively, and CDRH1, CDRH2, and CDRH3 have the sequence of SEQ ID NOs: 76, 77 and 78, respectively;

(c) CDRL1, CDRL2, and CDRL3 have the sequence of SEQ ID NOs: 48, 49 and 50, respectively, and CDRH1, CDRH2, and CDRH3 have the sequence of SEQ ID NOs: 79, 80 and 81, respectively;

(d) CDRL1, CDRL2, and CDRL3 have the sequence of SEQ ID NOs: 51,52 and 53, respectively, and CDRH1, CDRH2, and CDRH3 have the sequence of SEQ ID NOs: 82, 83 and 84, respectively;

(e) CDRL1, CDRL2, and CDRL3 have the sequence of SEQ ID NOs: 54, 55 and 56, respectively, and CDRH1, CDRH2, and CDRH3 have the sequence of SEQ ID NOs: 85, 86 and 87, respectively;

(f) CDRL1, CDRL2, and CDRL3 have the sequence of SEQ ID NOs: 57, 58 and 59, respectively, and CDRH1, CDRH2, and CDRH3 have the sequence of SEQ ID NOs: 88, 89 and 90, respectively;

(g) CDRL1, CDRL2, and CDRL3 have the sequence of SEQ ID NOs: 60, 55 and 56, respectively, and CDRH1, CDRH2, and CDRH3 have the sequence of SEQ ID NOs: 85, 86 and 87, respectively;

(h) CDRL1, CDRL2, and CDRL3 have the sequence of SEQ ID NOs: 45, 61 and 47, respectively, and CDRH1, CDRH2, and CDRH3 have the sequence of SEQ ID NOs: 76, 91 and 78, respectively;

(i) CDRL1, CDRL2, and CDRL3 have the sequence of SEQ ID NOs: 62, 63 and 64, respectively, and CDRH1, CDRH2, and CDRH3 have the sequence of SEQ ID NOs: 92, 93 and 94, respectively;

(j) CDRL1, CDRL2, and CDRL3 have the sequence of SEQ ID NOs: 45, 61 and 47, respectively, and CDRH1, CDRH2, and CDRH3 have the sequence of SEQ ID NOs: 76, 95 and 78, respectively;

(k) CDRL1, CDRL2, and CDRL3 have the sequence of SEQ ID NOs: 65, 55 and 56, respectively, and CDRH1, CDRH2, and CDRH3 have the sequence of SEQ ID NOs: 85, 86 and 87, respectively;

(l) CDRL1, CDRL2, and CDRL3 have the sequence of SEQ ID NOs: 42, 43 and 44, respectively, and CDRH1, CDRH2, and CDRH3 have the sequence of SEQ ID NOs: 73, 74 and 96, respectively;

(m) CDRL1, CDRL2, and CDRL3 have the sequence of SEQ ID NOs: 66, 67 and 68, respectively, and CDRH1, CDRH2, and CDRH3 have the sequence of SEQ ID NOs: 97, 98 and 99, respectively;

ΕΡ 2 379 594 Β1 (η) CDRL1, CDRL2, and CDRL3 have the sequence ofSEQ ID NOs: 69, 70 and 71, respectively, and CDRH1, CDRH2, and CDRH3 have the sequence ofSEQ ID NOs: 100, 101 and 102, respectively; or (o) CDRL1, CDRL2, and CDRL3 have the sequence ofSEQ ID NOs: 69, 70 and 72, respectively, and CDRH1, CDRH2, and CDRH3 have the sequence ofSEQ ID NOs: 100, 101 and 102, respectively.

2. The antibody or antigen-binding fragment thereof according to item 1, wherein the antibody or antigen-binding fragment thereof comprises:

(a) a V_L comprising the sequence of SEQ ID NO: 137 and a V_H comprising the sequence of SEQ ID NO:158;

(b) a V_L comprising the sequence of SEQ ID NO: 138 and a V_H comprising the sequence of SEQ ID NO:159;

(c) a V_L comprising the sequence of SEQ ID NO: 139 and a V_H comprising the sequence of SEQ ID NO:160;

(d) a V_L comprising the sequence of SEQ ID NO: 140 and a V_H comprising the sequence of SEQ ID NO:161;

(e) a V_L comprising the sequence of SEQ ID NO: 141 and a V_H comprising the sequence of SEQ ID NO:162;

(f) a V_L comprising the sequence of SEQ ID NO: 142 and a V_H comprising the sequence of SEQ ID NO:158;

(g) a V_L comprising the sequence of SEQ ID NO: 143 and a V_H comprising the sequence of SEQ ID NO:163;

(h) a V_L comprising the sequence of SEQ ID NO: 144 and a V_H comprising the sequence of SEQ ID NO:162;

(i) a V_L comprising the sequence of SEQ ID NO: 145 and a V_H comprising the sequence of SEQ ID NO:158;

(j) a V_L comprising the sequence of SEQ ID NO: 146 and a V_H comprising the sequence of SEQ ID NO:164;

(k) a V_L comprising the sequence of SEQ ID NO: 147 and a V_H comprising the sequence of SEQ ID NO:165;

(l) a V_L comprising the sequence of SEQ ID NO: 148 and a V_H comprising the sequence of SEQ ID NO:166;

(m) a V_L comprising the sequence ofSEQ ID NO: 148 and a V_H comprising the sequence ofSEQ ID NO:167;

(n) a V_L comprising the sequence of SEQ ID NO: 149 and a V_H comprising the sequence of SEQ ID NO:162;

(o) a V_L comprising the sequence of SEQ ID NO: 150 and a V_H comprising the sequence of SEQ ID NO:168;

(p) a V_L comprising the sequence of SEQ ID NO: 151 and a V_H comprising the sequence of SEQ ID NO:169;

(q) a V_L comprising the sequence ofSEQ ID NO: 152 and a V_H comprising the sequence ofSEQ ID NO:170; or (r) a V_L comprising the sequence of SEQ ID NO: 153 and a V_H comprising the sequence of SEQ ID NO:170.

3. The antibody or antigen-binding fragment thereof according to items 1 or 2, wherein the antibody or antigenbinding fragment is selected from the group consisting of a monoclonal antibody, a Fab fragment, an Fab’ fragment, an F(ab’)₂ fragment, an Fv fragment, a diabody, and a single chain antibody.

4. The antibody or antigen-binding fragment thereof according to item 3, wherein the antibody is a monoclonal antibody selected from the group consisting of a fully humán antibody, a humanized antibody and a chimeric antibody.

5. The antibody or antigen-binding fragment thereof according to item 4, wherein the monoclonal antibody is an lgG1 -, lgG2-, lgG3-, or lgG4-type antibody.

6. The antibody or antigen-binding fragment thereof according to item 5, wherein the monoclonal antibody is an lgG1 or lgG2 antibody.

7. An isolated polynucleotide that encodes an antibody or antigen-binding fragment thereof according to any of items 1-6.

8. An expression vector comprising the isolated polynucleotide of item 7.

9. A cell line transformed with the expression vector according to item 8.

10. A method of making an antibody or antigen-binding fragment thereof according to any of items 1-6, comprising preparing the antibody or antigen-binding fragment thereof from a hőst cell that secretes the antibody or antigenbinding fragment.

11. The antibody or antigen-binding fragment thereof according to item 1, wherein: the antibody or antigen-binding fragment comprises an antibody light chain of SEQ ID NO. 17 and an antibody heavy chain of SEQ ID NO. 29.

12. The antibody or antigen-binding fragment thereof according to item 1, wherein the antibody or antigen-binding fragment comprises an antibody light chain of SEQ ID NO:17 minus any signal sequences and an antibody heavy chain ofSEQ ID NO:29 minus any signal sequences.

ΕΡ 2 379 594 Β1

13. The antibody or antigen-binding fragment thereof according to item 1, wherein the antibody or antigen-binding fragment comprises:

(i) a CDRL1 comprising SEQ ID NO:42, a CDRL2 comprising SEQ ID NO:43, a CDRL3 comprising SEQ ID NO:44, a CDRH1 comprising SEQ ID NO:73, a CDRH2 comprising SEQ ID NO:74 and a CDRH3 comprising SEQ ID NO:75; or (ii) a V_L comprising SEQ ID NO:142 and a V_H comprising SEQ ID NO:158.

14. An antibody or antigen-binding fragment thereof according to any one of items 1 to 6 or 11 to 13 fór use in the treatment of a condition associated with CGRP receptor, wherein the condition is headache.

15. An antibody or antigen-binding fragment thereof according to any one of items 1 to 6 or 11 to 13 fór use in the treatment of a condition associated with CGRP receptor, wherein the condition is migraine.

[0022] The isolated antigen-binding proteins as claimed may selectively inhibit humán CGRP receptor (as compared with the humán AM1, AM2 or amylin receptors). In somé embodiments, the isolated antigén binding protein as claimed may selectively inhibits the humán CGRP receptor with a selectivity ratio of 50 or more, 75 or more, 100 or more, 150 or more, 200 or more, 250 or more, 300 or more, 400 or more, 500 or more, 750 or more or 1,000 or more. The degree of selective inhibition may be determined using any suitable method, e.g., using a cAMP assay as described in the Examples herein. In somé embodiments, the isolated antigén binding protein specifically binds to both humán CRLR and humán RAMP1, and does nőt specifically bind to humán AM1, humán AM2 or a humán amylin receptor (e.g., AMY1 or AMY2). Fór example, the isolated antigén binding protein may specifically bind humán CGRP R with a K_D &lt;1 μΜ, &lt;100 nM, &lt;10 nM, or &lt;5 nM. In somé embodiments, the isolated antigén binding protein specifically binds to humán CGRP R with a K_D &lt;100 nM, &lt;10 nM, or &lt;5 nM as determined using a FACS binding assay and analyzed, fór example, using methods described in Rathanaswami, et al., Biochemical and Biophysical Research Communications 334 (2005) 1004-1013. In somé embodiments, the isolated antigén binding protein has a Ki of &lt;100 nM, &lt;10 nM, &lt;1 nM, &lt;0.5 nM or&lt;0.1 nM in a CGRP binding competition assay. In somé embodiments, the isolated antigén binding protein has a Ki of&lt;100 nM, &lt;50 nM, &lt;20 nM, &lt;10 nM, &lt;1 nM, &lt;0.5 nM or&lt;0.1 nM in a radiolabeled ¹²⁵I-CGRP binding competition assay to membranes from cells expressing humán CGRP R, fór example, the assay described in Example 5 herein. [0023] In another exemplary aspect, the isolated antigen-binding proteins as claimed may compete fór binding to humán CGRP R, e.g., the extracellular portion of CGRP R, with a reference antibody comprising a heavy chain variable region comprising a sequence selected from the group consisting of SEQ ID NO:158-170 and a light chain variable region comprising a sequence selected from the group consisting ofSEQ ID NO: 137-153. In somé embodiments, binding competition is assessed using a binning assays, e.g., using a Biacore analysis, fór example, as described in Example 7 herein. In somé embodiments, the isolated antigén binding protein as claimed may compete fór binding to humán CGRP R with a reference antibody, the reference antibody comprising (i) a heavy chain variable region comprising a sequence selected from the group consisting of SEQ ID NOs: 161, 163, 164, 166 and 168; and (ii) a light chain variable region comprising a sequence selected from the group consisting ofSEQ ID NOs: 140,143,146, 148 and 150. In certain embodiments, the reference antibody comprises (i) a heavy chain defined by a sequence selected from the group consisting ofSEQ ID NOs:32, 34, 35, 37 and 39; and (ii) a light chain defined by a sequence selected from the group consisting ofSEQ ID NOs: 15, 18, 21, 23 and 25. In more specific embodiments, the reference antibody comprises a heavy chain and a light chain defined by one of the following pairs of sequences: (i) SEQ ID NO: 32 and SEQ ID NO: 15; (ii) SEQ ID NO: 34 and SEQ ID NO: 18; (iii) SEQ ID NO: 35 and SEQ ID NO: 21; (iv) SEQ ID NO: 37 and SEQ ID NO: 23; and (v) SEQ ID NO: 39 and SEQ ID NO: 25. In one such embodiment, the reference antibody comprises a heavy chain comprising SEQ ID NO: 32 and a light chain comprising SEQ ID NO: 15. In another such embodiment, the reference antibody comprises a heavy chain comprising SEQ ID NO: 34 and a light chain comprising SEQ ID NO: 18. In another such embodiment, the reference antibody comprises a heavy chain comprising SEQ ID NO: 35 and a light chain comprising SEQ ID NO: 21. In another such embodiment, the reference antibody comprises a heavy chain comprising SEQ ID NO: 37 and a light chain comprising SEQ ID NO: 23. In another such embodiment, the reference antibody comprises a heavy chain comprising SEQ ID NO: 39 and a light chain comprising SEQ ID NO: 25.

[0024] In certain embodiments, the isolated antigen-binding proteins as claimed that compete fór binding to humán CGRP R may alsó selectively inhibit the humán CGRP receptor, e.g., with a selectivity ratio of 100 or more, 250 or more, 500 or more, 750 or more, 1,000 or more, 2,500 or more, 5,000 or more or 10,000 or more, and such selectivity may be determined, e.g., using a cAMP assay as described in the Examples herein. In related embodiments, the isolated antigenbinding proteins as claimed that compete fór binding to humán CGRP R may specifically binds to humán CGRP R with a K_D &lt;1 μΜ, &lt;100 nM, &lt;10 nM, or &lt;5 nM, e.g., as determined using a FACS binding assay and analyzed, fór example, using methods described in Rathanaswami, et al., Biochemical and Biophysical Research Communications 334 (2005) 1004-1013. In related embodiments, the isolated antigen-binding proteins as claimed that compete fór binding to humán

EP 2 379 594 Β1

CGRP R may have a Ki of &lt;100 nM, &lt;10 nM, &lt;1 nM, &lt;0.5 nM or&lt;0.1 nM in a CGRP binding competition assay, e.g., in aradiolabeled ¹²⁵I-CGRP binding competition assay to membranes from cells expressing humán CGRP R, fór example, the assay described in Example 5 herein.

[0025] In any ofthe above-mentioned embodiments, the isolated antigen-binding protein that competes fór binding to humán CGRP R may be, fór example, a monoclonal antibody, a polyclonal antibody, a recombinant antibody, a humán (e.g., fully humán) antibody, a humanized antibody, a chimeric antibody, a multi-specific antibody, or an antigén binding fragment thereof. Further, the antibody fragment of the isolated antigen-binding protein that competes fór binding to humán CGRP R can be a Fab fragment, and Fab’ fragment, an F(ab’)₂ fragment, an Fv fragment, a diabody or a single chain antibody molecule; and may be, fór example, a humán monoclonal antibody, e.g., an lgG1 -, lgG2-, lgG3-, or lgG4type antibody. In certain embodiments, the isolated antigén binding proteins that compete fór binding to humán CGRP R may be neutralizing antigén binding proteins.

[0026] In any ofthe above-mentioned sequence-defined embodiments, the isolated antigen-binding protein may be, fór example, a monoclonal antibody, a polyclonal antibody, a recombinant antibody, a humán (e.g., fully humán) antibody, a humanized antibody, a chimeric antibody, a multi-specific antibody, or an antigén binding fragment thereof. Further, the antibody fragment of the isolated antigen-binding proteins may be a Fab fragment, and Fab’ fragment, an F(ab’)₂fragment, an Fv fragment, a diabody, or a single chain antibody molecule. Fór example, the isolated antigén binding protein may be a humán monoclonal antibody, and may be, e.g., an lgG1 -, lgG2-, lgG3-, or lgG4-type antibody. Further, the isolated antigén binding proteins may be neutralizing antigén binding proteins.

[0027] In any ofthe above-mentioned sequence-defined embodiments, the isolated antigen-binding protein may specifically bind to both humán CRLR and humán RAMP1 and nőt specifically bind to AM1, AM2 óra humán amylin receptor (e.g., AMY1), fór example, the isolated antigén binding protein may specifically bind to humán CGRP R with a K_D &lt;1 μΜ, &lt;100 nM, &lt;10 nM, or &lt;5 nM, e.g., as determined using a FACS binding assay and analyzed, fór example, using methods described in Rathanaswami, et al., Biochemical and Biophysical Research Communications 334 (2005) 1004-1013. In any ofthe above-mentioned sequence-defined embodiments, the isolated antigen-binding protein may selectively inhibit humán CGRP R, relatíve to the humán the AM1, AM2 or AMY1 receptors, e.g., with a selectivity ratio of 100 or more, 250 or more, 500 or more, 750 or more, 1,000 or more, 2,500 or more, 5,000 or more or 10,000 or more, where the degree of selective inhibition may be determined using any suitable method, e.g., using a cAMP assay as described in the Examples herein. In any ofthe above-mentioned sequence-defined embodiments, the isolated antigenbinding protein may have a Ki of &lt;100 nM, &lt;10 nM, &lt;1 nM, &lt;0.5 nM or&lt;0.1 nM in a CGRP binding competition assay, e.g., in a radiolabeled ¹²⁵I-CGRP binding competition assay to membranes from cells expressing humán CGRP R, e.g., the assay described in Example 5 herein.

[0028] Exemplary CDR consensus sequences illustrated herein are as follows:

K1 Consensus [0029] CDR1 RASQGIRX.|DLG (SEQ ID NO:103), wherein X₁ is selected from the group consisting of N and K. [0030] CDR2 X.|ASSLQS (SEQ ID NO:104), wherein X₁ is selected from the group consisting of A and G.

[0031] CDR3 LQYNX₁X₂PWT (SEQ ID NO:105), wherein X₁ is selected from the group consisting of I and S, and X₂is selected from the group consisting of Y and F.

K4 Consensus [0032] CDR3 QQYGNSLX-iR (SEQ ID NO: 106), wherein X₁ is selected from the group consisting of S and C.

K1,4 Consensus [0033] CDR1 RASQX₁X₂X₃X₄GX₅LX₆ (SEQ ID NO:107), wherein X₁ is selected from the group consisting of S and G, X₂ is selected from the group consisting ofV and I, X₃ is selected from the group consisting of S and R, X₄ is selected from the group consisting of S, N and K, X₅ is selected from the group consisting of Y and D, and X₆ is selected from the group consisting of T and G.

[0034] CDR2 X₁ASSX₂X₃X₄ (SEQ ID NO: 108), wherein X₁ is selected from the group consisting of G and A, X₂ is selected from the group consisting of R and L, X₃ is selected from the group consisting of A and Q, and X₄ is selected from the group consisting of T and S.

[0035] CDR3 X.jQYXgXgX^gXgXy (SEQ ID NO:109), wherein X₁ is selected from the group consisting of Q and L, X₂ is selected from the group consisting of G and N, X₃ is selected from the group consisting of N and T, X₄ is selected from the group consisting of S, Y and F, X₅ is selected from the group consisting of L and P, X₆ is selected from the group consisting of C, W and S, and X₇ is selected from the group consisting of R and T.

ΕΡ 2 379 594 Β1

Κ3 Consensus [0036] CDR1 KSSQSLLHSX₁GX₂X₃YLY (SEQ ID ΝΟ:110), wherein Χ₁ is selected from the group consisting ofD and A, X₂ is selected from the group consisting of R and K, and X₃ is selected from the group consisting of N and T.

K2,3 Consensus [0037] CDR1 X₁SSQSLLHSX₂GX₃X₄YLX₅ (SEQ ID NO:111), wherein X₁ is selected from the group consisting of R and K, X₂ is selected from the group consisting of F, D and A, X₃ is selected from the group consisting of Y, R and K, X₄ is selected from the group consisting of N and T, and X₅ is selected from the group consisting of D and Y.

[0038] CDR2 X₁X₂SNRX₃S (SEQ ID NO:112), wherein X₁ is selected from the group consisting of L and E, X₂ is selected from the group consisting of G and V, and X₃ is selected from the group consisting of A and F.

[0039] CDR3 MQX₁X₂X₃X₄PX₅T (SEQ ID NO:113), wherein X₁ is selected from the group consisting of A and S, X₂is selected from the group consisting of L and F, X₃ is selected from the group consisting of Q and P, X₄ is selected from the group consisting of T and L, and X₅ is selected from the group consisting of F and L.

Lm3 Consensus [0040] CDR2 RX.|NQRPS (SEQ ID NO:114), wherein X₁ is selected from the group consisting of N and S.

Lm1,2,3 Consensus [0041] CDR1 SGSSSNIGX₁NX₂VX₃ (SEQ ID NO:115), wherein X₁ is selected from the group consisting of N and S, X₂ is selected from the group consisting of Y and T, and X₃ is selected from the group consisting of S, N and Y.

[0042] CDR2 X₁X₂NX₃RPS (SEQ ID NO:116), wherein X₁ is selected from the group consisting of D, T and R, X₂ is selected from the group consisting of N and S, and X₃ is selected from the group consisting of K and Q.

[0043] CDR3 X₁X₂X₃DX₄XSLX₆X₇VV (SEQ ID NO:117), wherein X₁ is selected from the group consisting of G and A, X₂ is selected from the group consisting of T and A, X₃ is selected from the group consisting of W and R, X₄ is selected from the group consisting of S and D, X₅ is selected from the group consisting of R and S, X₆ is selected from the group consisting of S and N, and X₇ is selected from the group consisting of A and G.

LmAII Consensus [0044] CDR1 X₁GX₂X₃SX₄X₅X₆X₇X₈X_gX₁₀X₁₁ (SEQ ID NO:118), wherein X₁ is selected from the group consisting of S and Q, X₂ is present or absent, and if present, is S, X₃ is selected from the group consisting of S and D, X₄ is present or absent, and if present, is N, X₅ is selected from the group consisting of I and L, X₆ is selected from the group consisting of G and R, X₇ is selected from the group consisting of N and S, X₈ is selected from the group consisting of N and F, X_g is selected from the group consisting of Y and T, X₁₀ is selected from the group consisting of V and A, and X_n is selected from the group consisting of S, N and Y.

[0045] CDR2 X₁X₂NX₃RPS (SEQ ID NO:119), wherein X₁ is selected from the group consisting of D, G, T, and R, X₂is selected from the group consisting of N, K and S, and X₃ is selected from the group consisting of K, N and Q.

[0046] CDR3 X₁X₂X₃DX₄X₅X₆X₇X₈X_gV (SEQ ID NO:120), wherein X₁ is selected from the group consisting of G, N and A, X₂ is selected from the group consisting of T, S and A, X₃ is selected from the group consisting of W and R, X₄is selected from the group consisting of S and D, X₅ is selected from the group consisting of R and S, X₆ is selected from the group consisting of L and V, X₇ is selected from the group consisting of S, Y and N, X₈ is selected from the group consisting of A, H and G, and X_g is selected from the group consisting of V and L.

HC1 Consensus [0047] CDR1 X₁YYMX₂ (SEQ ID NO:121), wherein X₁ is selected from the group consisting of G and D, X₂ is selected from the group consisting of H and Y.

[0048] CDR2 WIX^NSGGTNYAQKFQG (SEQ ID NO:122), wherein X! is selected from the group consisting of N and S.

[0049] CDR3 X₁X₂X₃SX₄X₅X₆X₇X₈GX₉X₁₀X₁₁X₁₂YYX₁₃GMDV (SEQ ID NO:123), wherein X! is selected from the group consisting of D and G, X₂ is selected from the group consisting of Q and G, X₃ is selected from the group consisting of M and Y, X₄ is selected from the group consisting of I and G, X₅ is selected from the group consisting of I and Y, X₆is selected from the group consisting of M and A, X₇ is present or absent, and if present, is L, X₈ is present or absent, and if present, is R, X_g is selected from the group consisting of V and L, X₁₀ is selected from the group consisting of F

EP 2 379 594 Β1 and Y, X_n is selected from the group consisting of P and S, X₁₂ is selected from the group consisting of P and H, and X₁₃ is present or absent, and if present, is Y.

HC2 Consensus [0050] CDR2 RIKSX₁TDGGTTDYX₂APVKG (SEQ ID NO:124), wherein X! is selected from the group consisting of K and T, and X₂ is selected from the group consisting of T and A.

HC3 Consensus [0051] CDR1 X₁YX₂MX₃ (SEQ ID NO:125), wherein X₁ is selected from the group consisting of T and S, X₂ is selected from the group consisting of S and A, and X₃ is selected from the group consisting of N and S.

[0052] CDR2 X₁ISX₂SX₃X₄X₅X₆YYADSVKG (SEQ ID NO:126), wherein X! is selected from the group consisting of S and A, X₂ is selected from the group consisting of S and G, X₃ is selected from the group consisting of S and G, X₄is selected from the group consisting of S and G, X₅ is selected from the group consisting of Y and R, and X₆ is selected from the group consisting of R and T.

[0053] CDR3 X₁X₂X₃X₄X₅X₆X₇PYSX₈X₉WYDYYYGMDV (SEQ ID NO:127), wherein X! is selected from the group consisting of E and D, X₂ is selected from the group consisting of G and Q, X₃ is selected from the group consisting of V and R, X₄ is selected from the group consisting of S and E, X₅ is selected from the group consisting of G and V, X₆is selected from the group consisting of S and G, X₇ is present or absent, and if present, is S, X₈ is selected from the group consisting of I and S, and X₉ is selected from the group consisting of S and G.

HC4 Consensus [0054] CDR1 SX-|GMH (SEQ ID NO:128), wherein X₁ is selected from the group consisting of F and Y.

[0055] CDR2 VISX₁DGSX₂KYX₃X₄DSVKG (SEQ ID NO:129), wherein X! is selected from the group consisting of F and Y, X₂ is selected from the group consisting of I and H, X₃ is selected from the group consisting of S and Y, and X₄is selected from the group consisting of V and A.

[0056] CDR3 X₁RX₂X₃X₄X₅X₆SX₇X₈YYX₉X₁₀X₁₁YYGX₁₂X₁₃V (SEQ ID NO:130), wherein X! is selected from the group consisting of D and E, X₂ is selected from the group consisting of L and K, X₃ is selected from the group consisting of N and R, X₄ is selected from the group consisting of Y and V, X₅ is selected from the group consisting of Y and T, X₆is selected from the group consisting of D and M, X₇ is selected from the group consisting of S and T, X₈ is selected from the group consisting of G and L, X₉ is selected from the group consisting of H and Y, X₁₀ is present or absent, and if present, is Y, X_n is selected from the group consisting of K and F, X₁₂ is selected from the group consisting of M and L, and X₁₃ is selected from the group consisting of A and D.

HCA Consensus [0057] CDR1 X₁X₂X₃MX₄ (SEQ ID NO: 131), wherein X₁ isselectedfrom the group consisting of N and S, X₂ is selected from the group consisting of A, Y and F, X₃ is selected from the group consisting of W, A and G, and X₄ is selected from the group consisting of S and H.

[0058] CDR2 X! IX^X^gXgGX^gXgX^! iXi₂^x13^X14^VKG (SEQ ID NO:132), wherein X! is selected from the group consisting of R, A and V, X₂ is selected from the group consisting of K, S and W, X₃ is selected from the group consisting of S, G, F and Y, X₄ is present or absent, and if present, is selected from the group consisting of K and T, X₅ is present or absent, and if present, isT, X₆ is selected from the group consisting of D and S, X₇ is selected from the group consisting of G and S, X₈ is selected from the group consisting of T, R, I, N and H, X₉ is selected from the group consisting of T and K, X₁₀ is selected from the group consisting of D and Y, X_n is selected from the group consisting of Y and S, X₁₂is selected from the group consisting of T, A and V, X₁₃ is selected from the group consisting of A and D, and X₁₄ is selected from the group consisting of P and S.

[0059] CDR3 X₁X₂X₃X₄X₅X₆X₇X₃X₉X₁₀X₁₁X₁₂X₁₃X₁₄X₁₅X₁₆X₁₇GX₁₃X₁₉V (SEQ ID NO:133), wherein X! is selected from the group consisting of D, A and E, X₂ is selected from the group consisting of R, Q and G, X₃ is selected from the group consisting of T, R, L, G and K, X₄ is selected from the group consisting of G, E, Ν, I and R, X₅ is selected from the group consisting of Υ, V and A, X₆ is selected from the group consisting of S, G, Y, A and T, X₇ is selected from the group consisting of I, P, D, A and M, X₈ is present or absent, and if present, is selected from the group consisting of S and Y, X₉ is present or absent, and if present, is selected from the group consisting of W, S and T, X₁₀ is selected from the group consisting of S, G and L, X_n is selected from the group consisting of S, G, L and Y, X₁₂ is present or absent, and if present, is selected from the group consisting of W and Y, X₁₃ is selected from the group consisting of Y and H, X₁₄ is present or absent, and if present, is selected from the group consisting of Y and D, X₁₅ is selected from the group

ΕΡ 2 379 594 Β1 consisting of Υ, Κ and F, Χ₁₆ is present or absent, and if present, is Y, X₁₇ is present or absent, and if present, is Y, X₁₈is selected from the group consisting of M and L, and X_lg is selected from the group consisting of D and A.

HCB Consensus [0060] CDR1 X.1X2X3X4X5 (SEQ ID NO:134), wherein X₁ is selected from the group consisting of N, G, D, S and A, X₂ is selected from the group consisting of A, F and Y, X₃ is selected from the group consisting of W, Y, A and G, X₄ is selected from the group consisting of M and L, and X₅ is selected from the group consisting of S and H.

[0061] CDR2X₁IX₂X₃X₄X₅X₆X₇X₈X₉X₁₀X₁₁X₁₂X₁3X₁₄X₁₅X₁₆X₁₇G (SEQ ID NO:135), wherein X! is selected from the group consisting of R, W, A, V, S and F, X₂ is selected from the group consisting of K, N, S, W and R, X₃ is selected from the group consisting of S, P, G, F and Y, X₄ is present or absent, and if present, is selected from the group consisting of K, T and R, X₅ is present or absent, and if present, is selected from the group consisting of T and A, X₆ is selected from the group consisting of D, Ν, H, S and Y, X₇ is selected from the group consisting of G and S, X₃ is selected from the group consisting of G and S, X_g is selected from the group consisting of T, G, R, I, Ν, H and Y, X₁₀ is selected from the group consisting of T, K, R and P, X_n is selected from the group consisting of D, Ν, Y and E, X₁₂ is selected from the group consisting of Y and S, X₁₃ is selected from the group consisting of T, A and V, X₁₄ is selected from the group consisting of A, Q and D, X₁₅ is selected from the group consisting of Ρ, K and S, X₁₆ is selected from the group consisting of V and F, and X₁₇ is selected from the group consisting of K and Q.

[0062] CDR3 X₁X₂X₃X₄X₅SX₆X₇X₈X_gX₁₀X₁₁X₁₂X₁₃X₁₄X₁₅X₁₆GX₁₇X₁₈V (SEQ ID NO:136), wherein X! is selected from the group consisting of D, G, A and E, X₂ is selected from the group consisting of R, G and Q, X₃ is selected from the group consisting of T, Μ, Y, R, L, G and K, X₄ is selected from the group consisting of G, S, Ε, Ν, I and R, X₅ is selected from the group consisting of Υ, I, G, V and A, X₆ is selected from the group consisting of S, I, Y, G, A and T, X₇ is selected from the group consisting of I, M, A, P and D, X₃ is present or absent, and if present, is selected from the group consisting of S, L and Y, X_g is present or absent, and if present, is selected from the group consisting of W, R, S and T, X₁₀ is selected from the group consisting of S, G and L, X₁₁ is selected from the group consisting of S, V, L, G and Y, X₁₂ is present or absent, and if present, is selected from the group consisting of F, Y and W, X₁₃ is selected from the group consisting of Y, P, S and H, X₁₄ is present or absent, and if present, is selected from the group consisting of Y, P, D and H, X₁₅ is selected from the group consisting of Y, K and F, X₁₆ is present or absent, and if present, is Y, X₁₇is present or absent, and if present, is Y and X₁₃ is selected from the group consisting of M and L.

[0063] In any ofthe above-mentioned consensus sequence defined embodiments, the isolated antigen-binding protein may be, for example, an AVIMER polypeptide, a monoclonal antibody, a polyclonal antibody, a recombinant antibody, a humán (e.g., fully humán) antibody, a humanized antibody, a chimeric antibody, a multi-specific antibody, oran antigén binding fragmentthereof. Further, the antibody fragment ofthe isolated antigen-binding proteins may be a Fab fragment, and Fab’ fragment, an F(ab’)₂ fragment, an Fv fragment, a diabody, or a single chain antibody molecule. For example, the isolated antigén binding protein may be a humán monoclonal antibody, and may be, e.g., an lgG1 -, lgG2-, lgG3-, or lgG4-type antibody. Further, the isolated antigén binding proteins may be neutralizing antigén binding proteins. [0064] In any ofthe above-mentioned consensus sequence defined embodiments, the isolated antigen-binding protein may specifically bind to both humán CRLR and humán RAMP1 and nőt specifically bind to AM1, AM2 óra humán amylin receptor (e.g., AMY1), for example, the isolated antigén binding protein may specifically bind to humán CGRP R with a K_D &lt;1 μΜ, &lt;100 nM, &lt;10 nM, or &lt;5 nM, e.g., as determined using a FACS binding assay and analyzed, for example, using methods described in Rathanaswami, et al., Biochemical and Biophysical Research Communications 334 (2005) 1004-1013. In any ofthe above-mentioned consensus sequence defined embodiments, the isolated antigen-binding protein may selectively inhibit humán CGRP R, relatíve to the humán the AM1, AM2 or AMY1 receptors, e.g., with a selectivity ratio of 100 or more, 250 or more, 500 or more, 750 or more, 1,000 or more, 2,500 or more, 5,000 or more or 10,000 or more, where the degree of selective inhibition may be determined using any suitable method, e.g., using a cAMP assay as described in the Examples herein. In any of the above-mentioned consensus sequence defined embodiments, the isolated antigen-binding protein may have a Ki of &lt;100 nM, &lt;10 nM, &lt;1 nM, &lt;0.5 nM or &lt;0.1 nM in a CGRP binding competition assay, e.g., in a radiolabeled ¹²⁵I-CGRP binding competition assay to membranes from cells expressing humán CGRP R, e.g., the assay described in Example 5 herein.

[0065] In any ofthe above-mentioned V_L and V_H sequence defined embodiments, the isolated antigen-binding protein may specifically bind to both humán CRLR and humán RAMP1 and nőt specifically bind to AM1, AM2 óra humán amylin receptor (e.g., AMY1), for example, the isolated antigén binding protein may specifically bind to humán CGRP R with a K_D &lt;1 μΜ, &lt;100 nM, &lt;10 nM, or &lt;5 nM, e.g., as determined using a FACS binding assay and analyzed, for example, using methods described in Rathanaswami, et al., Biochemical and Biophysical Research Communications 334 (2005) 1004-1013. In any ofthe above-mentioned V_L and V_H sequence defined embodiments, the isolated antigen-binding protein may selectively inhibit humán CGRP R, relatíve to the humán the AM1, AM2 or AMY1 receptors, e.g., with a selectivity ratio of 100 or more, 250 or more, 500 or more, 750 or more, 1,000 or more, 2,500 or more, 5,000 or more or 10,000 or more, where the degree of selective inhibition may be determined using any suitable method, e.g., using

ΕΡ 2 379 594 Β1 a cAMP assay as described in the Examples herein. In any of the above-mentioned V_L and V_H sequence-defined embodiments, the isolated antigen-binding protein may have a Ki of &lt;100 nM, &lt;10 nM, &lt;1 nM, &lt;0.5 nM or &lt;0.1 nM in a CGRP binding competition assay, e.g., in a radiolabeled ¹²⁵I-CGRP binding competition assay to membranes from cells expressing humán CGRP R, e.g., the assay described in Example 5 herein.

[0066] In any of the above-mentioned light and heavy chain sequence defined embodiments, the isolated antigenbinding protein may comprise the specified heavy and/or light chain sequence, butwith a different signal peptide or with no signal peptide. In any ofthe above-mentioned light and heavy chain sequence defined embodiments, the isolated antigen-binding protein may be, fór example, a monoclonal antibody, a polyclonal antibody, a recombinant antibody, a humán (e.g., fully humán) antibody, a humanized antibody, a chimeric antibody, a multi-specific antibody, or an antigén binding fragment thereof. Further, the antibody fragment ofthe isolated antigen-binding proteins may be a Fab fragment, and Fab’ fragment, an F(ab’)₂ fragment, an Fv fragment, a diabody, or a single chain antibody molecule. Fór example, the isolated antigén binding protein may be a humán monoclonal antibody, and may be, e.g., an lgG1 -, lgG2-, lgG3-, or lgG4-type antibody. Further, the isolated antigén binding proteins may be neutralizing antigén binding proteins. [0067] In any of the above-mentioned light and heavy chain sequence defined embodiments, the isolated antigenbinding protein may specifically bind to both humán CRLR and humán RAMP1 and nőt specifically bind to AM1, AM2 or a humán amylin receptor (e.g., AMY1), fór example, the isolated antigén binding protein may specifically bind to humán CGRP R with a K_D&lt;1 μΜ, &lt;100 nM, &lt;10 nM, or &lt;5 nM, e.g., as determined using a FACS binding assay and analyzed, fór example, using methods described in Rathanaswami, et al., Biochemical and Biophysical Research Communications 334 (2005) 1004-1013. In any ofthe above-mentioned light and heavy chain sequence defined embodiments, the isolated antigen-binding protein may selectively inhibit humán CGRP R, relatíve to the humán the AM1, AM2 or AMY1 receptors, e.g., with a selectivity ratio of 100 or more, 250 or more, 500 or more, 750 or more, 1,000 or more, 2,500 or more, 5,000 or more or 10,000 or more, where the degree of selective inhibition may be determined using any suitable method, e.g., using a cAMP assay as described in the Examples herein. In any ofthe above-mentioned light and heavy chain sequence-defined embodiments, the isolated antigen-binding protein may have a Ki of &lt;100 nM, &lt;10 nM, &lt;1 nM, &lt;0.5 nM or &lt;0.1 nM in a CGRP binding competition assay, e.g., in a radiolabeled ¹²⁵I-CGRP binding competition assay to membranes from cells expressing humán CGRP R, e.g., the assay described in Example 5 herein. [0068] In a further aspect, alsó provided are isolated nucleic acid polynucleotides that encode any of the CGRP R antigen-binding proteins summarized above. In somé instances, the isolated nucleic acid molecules are operably-linked to a controi sequence. In related embodiments, the isolated polynucleotides are incorporated intő an expression vector. [0069] Alsó included are cell lines transformed with expression vectors comprising isolated polynucleotides as described above. In a related aspect, alsó provided are expression vectors and hőst cells transformed or transfected with the expression vectors that comprise the aforementioned isolated nucleic acid molecules that encode CGRP R antigenbinding proteins described above [0070] In another aspect, alsó provided is a method of preparing the antigen-binding proteins that includes the step of preparing the antigén binding protein from a hőst cell that secretes the antigen-binding protein. In somé embodiments, the antigén binding protein isgenerated using an immunogen comprising soluble CGRP receptor. In somé embodiments, such soluble CGRP receptor is obtained by co-expressing and purifying an N-terminal extracellular domain (ECD) of humán CRLR and an ECD of humán RAMP1, e.g., an ECD of humán CRLR comprising SEQ ID NO: 6 and an ECD of RAMP 1 comprising SEQ ID NO: 8, fór example, as described in Examples 1 and 2 herein.

[0071] In yet another aspect, a pharmaceutical composition is provided comprising at least one ofthe antigen-binding proteins summarized above and a pharmaceutically acceptable excipient. In one embodiment, the pharmaceutical composition may comprise an additional active agent that is selected from the group consisting ofa radioisotope, radionuclide, a toxin, or a therapeutic and a chemotherapeutic group.

[0072] In one aspect, the isolated antigén binding protein is effective to inhibit vasodialation and/ordecrease neurogenic inflammation when administered to a patient. In one embodiment, the isolated antigén binding protein is effective to reduce the frequency and/or severity of headaches, fór example, migraine headaches. Fór example, the antigén binding protein may be used as an acute treatment of migraine, and/or as a prophylactic treatment to prevent or reduce the frequency and/or severity of symptoms, particularly pain symptoms, associated with a migraine attack.

[0073] Illustrated are methods fór treating or preventing a condition associated with CGRP R in a patient, comprising administering to a patient an effective amount of at least one isolated antigen-binding protein summarized above. The condition may be a headache, fór example, a migraine headache or a cluster headache or another type of pain, e.g., a chronic pain; in another embodiment it is diabetes mellitus (type II) ; alternatively it is inflammation, particularly neurogenic inflammation; alternatively it is a cardiovascular disorder; alternatively it is a hemodynamic derangement associated with endotoxemia and sepsis; alternatively it is vasodialation.

[0074] Alsó illustrated is a method of inhibiting binding of CGRP to humán CGRP R, e.g., the extracellular portion of CGRP R, in a patient comprising administering an effective amount of at least one antigen-binding protein provided herein and/or summarized above.

[0075] These and other aspects will be described in greater detail herein. Each ofthe aspects provided can encompass

ΕΡ 2 379 594 Β1 various embodiments provided herein. It is therefore anticipated that each ofthe embodiments involving one element or combinations of elements can be included in each aspect described, and all such combinations ofthe above aspects and embodiments are expressly considered. Other features, objects, and advantages ofthe invention are apparent in the detailed description that follows.

BRIEF DESCRIPTION OF THE DRAWINGS [0076]

Fig. 1 shows an alignment of RAMP-1 sequences from humán, cynomolgus monkey and rat.

Fig. 2 shows an alignment of CRLR sequences from humán, cynomolgus monkey and rat.

Figs 3A and 3B show phylogenetically-based sequence alignments of light chain CDRs from the indicated antiCGRP receptor antibody clones having kappa light chains, and certain corresponding consensus sequences.

Fig. 4 shows phylogenetically-based sequence alignments of light chain CDRs from the indicated anti-CGRP receptor antibody clones having lambda light chains, and certain corresponding consensus sequences.

Figs. 5A, 5B, 5C, 5D and 5E show phylogenetically-based sequence alignments of heavy chain CDRs from the indicated anti-CGRP receptor antibody clones, and certain corresponding consensus sequences.

Fig. 5F shows consensussequencesof exemplary anti-CGRP receptor anti body heavy chain CDRs disclosed herein. Fig. 6 is a plot of data from two experiments showing percent inhibition of labeled ligand binding to CGRP R by 1092 anti-CGRP R hybridoma supernatants (diamonds) and 68 negative control supernatants (squares).

Figs. 7A-D show exemplary cAMP assay IC50 data from cells expressing hCGRP receptor (Fig. 7A), hAM1 (Fig. 7B), hAM2 (Fig. 7C) and humán amylin receptors (Fig. 7D) fór three indicated anti-CGRP R mAbs.

Fig. 8 shows an example of ¹²⁵I-CGRP binding data such as may be used to determine the Ki of mAbs to humán CGRP receptor.

Figs. 9A-D show Biacore competition data fór selected antibodies disclosed herein.

Fig. 10 shows a FACS Kd determination of mAb 12G8.

Fig. 11 shows an alignment of cynomolgus, humán, humán chimeras, rat, and rhesus RAMP1 sequences.

Figs. 12A-B show an alignment of humán, cynomolgus, rhesus, rat, humán chimera and consensus CRLR sequences.

Figs. 13A-13C show representative FACS data of different chimeric CGRP receptors binding to anti-CGRP R antibodies.

Fig. 14 shows peptide maps derived from AspN digestions of CGRP R alone (chromatogram A) and from digestion ofa control sample containing CGRP R monoclonal antibody 12G8 (chromatogram B).

Fig. 15 shows AspN digestions of CGRP R in the presence of different concentrations of CGRP R neutralizing antibody.

Fig. 16 shows AspN digestions of CGRP R in the presence of different concentration of CGRP R neutralizing antibody, 4E4.

Fig. 17 shows immunohistochemistry staining intensity of cells expressing various receptor components with antibody 32H7.

DETAILED DESCRIPTION [0077] The seetion headings used herein are fór organizational purposes only and are nőt to be construed as limiting the subject matter described.

[0078] Unless otherwise defined herein, scientific and technical terms used in connection with the present application shall have the meanings that are commonly understood by those of ordinary skill in the art. Further, unless otherwise required by context, singular terms shall include pluralities and plural terms shall include the singular.

[0079] Generally, nomenclatures used in connection with, and techniques of, cell and tissue culture, molecular biology, immunology, microbiology, genetics and protein and nucleic acid chemistry and hybridization described herein are those well known and commonly used in the art. The methods and techniques ofthe present application are generally performed according to conventional methods well known in the art and as described in various generál and more specific references that are cited and discussed throughout the present specification unless otherwise indicated. See, e.g., Sambrook et al., Molecular Cloning: A Laboratory Manual, 3rd ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (2001), Ausubel et al., Current Protocols in Molecular Biology, Greene Publishing Associates (1992), and Harlow and Lane Antibodies: A Laboratory Manual Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (1990). Enzymatic reactions and purification techniques are performed according to manufacturer’s specifications, as commonly accomplished in the art or as described herein. The terminology used in connection with, and the laboratory procedures and techniques of, analytical chemistry, synthetic organic chemistry, and medicinái and pharmaceutical chemistry described

ΕΡ 2 379 594 Β1 herein are those well known and commonly used in the art. Standard techniques can be used for Chemical syntheses, Chemical analyses, pharmaceutical preparation, formulation, and delivery, and treatment of patients.

[0080] It should be understood that this invention is nőt limited to the particular methodology, protocols, and reagents, etc., described herein and as such may vary. The terminology used herein is for the purpose of describing particular embodiments only, and is nőt intended to limit the scope ofthe present invention, which is defined solely by the claims. [0081] Other than in the operating examples, or where otherwise indicated, all numbers expressing quantities of ingredients or reaction conditions used herein should be understood as modified in all instances by the term about. The term about when used in connection with percentages means ±1%.

Definitions [0082] The term polynucleotide or nucleic acid includes both single-stranded and double-stranded nucleotide polymers. The nucleotides comprising the polynucleotide can be ribonucleotides or deoxyribonucleotides or a modified form of either type of nucleotide. Said modifications include base modifications such as bromouridine and inosine derivatives, ribose modifications such as 2’,3’-dideoxyribose, and internucleotide linkage modifications such as phosphorothioate, phosphorodithioate, phosphoroselenoate, phosphorodiselenoate, phosphoroanilothioate, phoshoraniladate and phosphoroamidate.

[0083] The term oligonucleotide means a polynucleotide comprising 200 orfewer nucleotides. In somé embodiments, oligonucleotides are 10 to 60 bases in length. In other embodiments, oligonucleotides are 12, 13, 14, 15, 16, 17, 18, 19, or 20 to 40 nucleotides in length. Oligonucleotides may be single stranded or double stranded, e.g., for use in the construction of a mutant gene. Oligonucleotides may be sense or antisense oligonucleotides. An oligonucleotide can include a label, including a radiolabel, a fluorescent label, a hapten or an antigenic label, for detection assays. Oligonucleotides may be used, for example, as PCR primers, cloning primers or hybridization probes.

[0084] An isolated nucleic acid molecule means a DNA or RNA of genomic, mRNA, cDNA, or synthetic origin or somé combination thereof which is nőt associated with all óra portion ofa polynucleotide in which the isolated polynucleotide is found in natúré, or is linked to a polynucleotide to which it is nőt linked in natúré. For purposes ofthis disclosure, it should be understood that a nucleic acid molecule comprising a particular nucleotide sequence does nőt encompass intact chromosomes. Isolated nucleic acid molecules comprising specified nucleic acid sequences may include, in addition to the specified sequences, coding sequences for up to ten or even up to twenty other proteins or portions thereof, or may include operably linked regulatory sequences that control expression ofthe coding region ofthe recited nucleic acid sequences, and/or may include vector sequences.

[0085] Unless specified otherwise, the left-hand end of any single-stranded polynucleotide sequence discussed herein is the 5’ end; the left-hand direction of double-stranded polynucleotide sequences is referred to as the 5’ direction. The direction of 5’ to 3’ addition of nascent RNA transcripts is referred to as the transcription direction; sequence regions on the DNA strand having the same sequence as the RNA transcript that are 5’ to the 5’ end of the RNA transcript are referred to as upstream sequences; sequence regions on the DNA strand having the same sequence as the RNA transcript that are 3’ to the 3’ end of the RNA transcript are referred to as downstream sequences.

[0086] The term control sequence refers to a polynucleotide sequence that can affect the expression and Processing of coding sequences to which it is ligated. The natúré of such control sequences may depend upon the hőst organism. In particular embodiments, control sequences for prokaryotes may include a promoter, a ribosomal binding site, and a transcription termination sequence. For example, control sequences for eukaryotes may include promoters comprising one or a plurality of recognition sites for transcription factors, transcription enhancer sequences, and transcription termination sequence. Control sequences can include leader sequences and/or fusion partner sequences.

[0087] The term vector means any molecule or entity (e.g., nucleic acid, plasmid, bacteriophage or vírus) used to transfer protein coding information intő a hőst cell.

[0088] The term expression vector or expression construct refers to a vector that is suitable for transformation of a hőst cell and contains nucleic acid sequences that direct and/or control (in conjunction with the hőst cell) expression of one or more heterologous coding regions operatively linked thereto. An expression construct may include, bút is nőt limited to, sequences that affect or control transcription, translation, and, if introns are present, affect RNA splicing of a coding region operably linked thereto.

[0089] As used herein, operably linked means that the components to which the term is applied are in a relationship that allows them to carry out their inherent functions under suitable conditions. For example, a control sequence in a vector that is operably linked to a protein coding sequence is ligated thereto so that expression ofthe protein coding sequence is achieved under conditions compatible with the transcriptional activity ofthe control sequences.

[0090] The term hőst cell means a cell that has been transformed, or is capable of being transformed, with a nucleic acid sequence and thereby expresses a gene of interest. The term includes the progeny of the parent cell, whether or nőt the progeny is identical in morphology or in genetic make-up to the original parent cell, so long as the gene of interest is present.

ΕΡ 2 379 594 Β1 [0091] The term transduction means the transfer ofgenes from one bacterium to another, usually by bacteriophage. Transduction alsó refers to the acquisition and transfer of eukaryotic celiular sequences by replication defective retroviruses.

[0092] The term transfection means the uptake offoreignorexogenous DNA by a cell, and a cell has been transfected when the exogenous DNA has been introduced inside the cell membráné. A number of transfection techniques are well known in the art and are disclosed herein. See, e.g., Graham et al., 1973, Virology 52:456; Sambrook et al., 2001, Molecular Cloning: A Laboratory Manual, supra; Davis et al., 1986, Basic Methods in Molecular Biology, Elsevier; Chu et al., 1981, Gene 13:197. Such techniques can be used to introduce one or more exogenous DNA moieties intő suitable hőst cells.

[0093] The term transformation refers to a change in a cell’s genetic characteristics, and a cell has been transformed when it has been modified to contain new DNA or RNA. Fór example, a cell is transformed where it is genetically modified from its native state by introducing new genetic matéria! via transfection, transduction, or other techniques. Following transfection or transduction, the transforming DNA may recombine with that ofthe cell by physically integrating intő a chromosome of the cell, or may be maintained transiently as an episomal element without being replicated, or may replicate independently as a plasmid. A cell is considered to have been stably transformed when the transforming DNA is replicated with the division ofthe cell.

[0094] The terms polypeptide or protein are used interchangeably herein to referto a polymer of amino acid residues. The terms alsó apply to amino acid polymers in which one or more amino acid residues is an analóg or mimetic of a corresponding naturally occurring amino acid, as well as to naturally occurring amino acid polymers. The terms can alsó encompass amino acid polymers that have been modified, e.g., by the addition of carbohydrate residues to form glycoproteins, or phosphorylated. Polypeptides and proteins can be produced by a naturally-occurring and non-recombinant cell; or it is produced by a genetically-engineered or recombinant cell, and comprise molecules having the amino acid sequence of the native protein, or molecules having deletions from, additions to, and/or substitutions of one or more amino acids of the native sequence. The terms polypeptide and protein specifically encompass antigén binding proteins, e.g., CGRP R antigen-binding proteins, CGRP R binding proteins, antibodies, or sequences that have deletions from, additions to, and/or substitutions ofone or more amino acids ofan antigen-binding protein. The term polypeptide fragment refers to a polypeptide that has an amino-terminal deletion, a carboxyl-terminal deletion, and/or an internál deletion as compared with the full-length protein. Such fragments may alsó contain modified amino acids as compared with the full-length protein. In certain embodiments, fragments are about five to 500 amino acids long. Fór example, fragments may be at least 5, 6, 8, 10, 14, 20, 50, 70, 100, 110, 150, 200, 250, 300, 350, 400, or450 amino acids long. Usefui polypeptide fragments include immunologically functional fragments of antibodies, including binding domains. In the case of a CGRP R-binding antibody, usefui fragments include bút are nőt limited to a CDR region, a variable domain of a heavy or light chain, a portion of an antibody chain or just its variable domain including two CDRs, and the like. TheCGRP receptor, or CGRP R, is understood to comprise RAMP1 and CRLR.

[0095] The term isolated protein (e.g., isolated antigén binding protein), isolated polypeptide or isolated antibody means that a subject protein, polypeptide or antibody (1) is free of at least somé other proteins with which it would normally be found, (2) is essentially free of other proteins from the same source, e.g., from the same species, (3) is expressed by a cell from a different species, (4) has been separated from at least about 50 percent of polynucleotides, lipids, carbohydrates, or other materials with which it is associated in natúré, (5) is operably associated (by covalent or noncovalent interaction) with a polypeptide with which it is nőt associated in natúré, or (6) does nőt occur in natúré. Typically, an isolated protein, isolated polypeptide or isolated antibody constitutes at least about 5%, at least about 10%, at least about 25%, or at least about 50% of a given sample. Genomic DNA, cDNA, mRNA or other RNA, of synthetic origin, or any combination thereof may encode such an isolated protein. Preferabiy, the isolated protein polypeptide or antibody is substantially free from other proteins or other polypeptides or other contaminants that are found in its natural environment that would interfere with its therapeutic, diagnostic, prophylactic, research or other use.

[0096] A variant of a polypeptide (e.g., an antigén binding protein, oran antibody) comprises an amino acid sequence wherein one or more amino acid residues are inserted intő, deleted from and/or substituted intő the amino acid sequence relatíve to another polypeptide sequence. Variants include fusion proteins.

[0097] A derivative of a polypeptide is a polypeptide (e.g., an antigén binding protein, or an antibody) that has been chemically modified in somé manner distinct from insertion, deletion, or substitution variants, e.g., via conjugation to another Chemical moiety.

[0098] The term naturally occurring as used throughout the specification in connection with biological materials such as polypeptides, nucleic acids, hőst cells, and the like, refers to materials which are found in natúré.

[0099] An antigén binding protein as used herein means a protein that specifically binds a specified target antigén, such as CGRP R, particularly primate, e.g., humán CGRP R. A CGRP R antigén binding protein specifically binds the humán CGRP receptor.

[0100] An antigén binding protein is said to specifically bind its target when the dissociation constant (K_D) is &lt;10^-6M. The antibody specifically binds the target antigén with high affinity when the K_D is &lt;1x 10^_θ M. In one embodiment,

ΕΡ 2 379 594 Β1 the antibodies will bind to CGRP R, or humán CGRP R with a K_D&lt;5x 10^-7; in another embodiment the antibodies will bind with a KD &lt;1x 10⁷; in another embodiment the antibodies will bind with a KD &lt;5x 10⁸; in another embodiment the antibodies will bind with a KD &lt;1x 10⁸; in another embodiment the antibodies will bind with a KD &lt;5x 10⁹; in another embodiment the antibodies will bind with a KD &lt;1x 10^-9; in another embodiment the antibodies will bind with a KD &lt;5x 10^_1°; in another embodiment the antibodies will bind with a K_D &lt;1x 10^_1°.

[0101] An antibody, antigén binding fragment thereof or antigén binding protein selectively inhibits a specific receptor relatíve to other receptors when the IC50 of the antibody, antigén binding fragment thereof or antigén binding protein in an inhibition assay of the specific receptor is at least 50-fold lower than the IC50 in an inhibition assay of another reference receptor. The selectivity ratio is the IC50 ofthe reference receptor divided by IC50 ofthe specific receptor. An antibody, antigén binding fragment thereof or antigén binding protein selectively inhibits the humán CGRP receptor if the IC50 ofthe antibody, antigén binding fragment thereof or antigén binding protein in a cAMP assay, e.g., the cAMP inhibition assay as described in Example 4 herein, is at least 50-fold lower than the IC50 of that same antibody, antigén binding fragment thereof or antigén binding protein in an inhibition assay ofthe humán AM1, AM2 or an amylin receptor (e.g., AMY1). By way of non-limiting example, if the IC50 ofa specific anti-CGRP R antibody in a cAMP assay of hCGRP R is, e.g., between 0.1 nM and 20 nM, and the IC50 ofthe same antibody in a cAMP assay ofthe hAM1, hAM2 or humán AMY1 receptor is 1000 nM or more, that antibody selectively inhibits the hCGRP receptor. An antigén binding protein that selectively inhibits a specific receptor is alsó understood to be a neutralizing antigén binding protein with respect to that receptor.

[0102] Antigén binding region means a protein, or a portion of a protein, that specifically binds a specified antigén. Fór example, that portion ofan antigén binding protein that contains the amino acid residues that interact with an antigén and confer on the antigén binding protein its specificity and affinity fór the antigén is referred to as antigén binding region. An antigén binding region typically includes one or more complementary binding regions (CDRs). Certain antigén binding regions alsó include one or more framework regions. A CDR is an amino acid sequence that contributes to antigén binding specificity and affinity. Framework regions can aid in maintaining the proper conformation of the CDRs to promote binding between the antigén binding region and an antigén.

[0103] In certain aspects, recombinant antigén binding proteins that bind CGRP R protein, or humán CGRP R, are provided. In this context, a recombinant protein is a protein made using recombinant techniques, i.e., through the expression ofa recombinant nucleic acid as described herein. Methods and techniques fór the production of recombinant proteins are well known in the art.

[0104] The term antibody refers to an intact immunoglobulin of any isotype, or an antigén binding fragment thereof that can compete with the intact antibody fór specific binding to the target antigén, and includes, fór instance, chimeric, humanized, fully humán, and bispecific antibodies. An antibody as such is a species of an antigén binding protein. An intact antibody generally will comprise at least two full-length heavy chains and two full-length light chains, bút in somé instances may include fewer chains such as antibodies naturally occurring in camelids which may comprise only heavy chains. Antibodies may be derived solely from a single source, or may be chimeric, that is, different portions of the antibody may be derived from two different antibodies as described further below. The antigén binding proteins, antibodies, or binding fragments may be produced in hybridomas, by recombinant DNA techniques, or by enzymatic or Chemical cleavage of intact antibodies. Unless otherwise indicated, the term antibody includes, in addition to antibodies comprising two full-length heavy chains and two full-length light chains, derivatives, variants, fragments, and mutations thereof, examples ofwhich are described below.

[0105] The term light chain includes a full-length light chain and fragments thereof having sufficient variable region sequence to confer binding specificity. A full-length light chain includes a variable region domain, V_L, and a constant region domain, C_L. The variable region domain ofthe light chain is at the amino-terminus ofthe polypeptide. Light chains include kappa chains and lambda chains.

[0106] The term heavy chain includes a full-length heavy chain and fragments thereof having sufficient variable region sequence to confer binding specificity. A full-length heavy chain includes a variable region domain, V_H, and three constant region domains, C_H1, C_H2, and C_H3. The V_H domain is at the amino-terminus of the polypeptide, and the C_Hdomains are at the carboxyl-terminus, with the C_H3 being closest to the carboxy-terminus of the polypeptide. Heavy chains may be of any isotype, including IgG (including lgG1, lgG2, lgG3 and lgG4 subtypes), IgA (including lgA1 and lgA2 subtypes), IgM and IgE.

[0107] The term signal sequence, leader sequence or signal peptide refers to a short (3-60 amino acids long) peptide chain that directs the transport ofa protein. Signal peptides may alsó be called targeting signals, signal sequences, transit peptides, or localization signals. Somé signal peptides are cleaved from the protein by signal peptidase after the proteins are transported, such that the biologically activeform ofthe protein (e.g., an antigén binding protein as described herein) is the cleaved, shorterform. Accordingly, terms such as antibody comprising a heavy chain..., antibody comprising a light chain..., etc., where the antibody is characterized as having a heavy and/or light chain with a particular identified sequence, are understood to include antibodies having the specific identified sequences, antibodies having the specific identified sequences except that the signal sequences are replaced by different signal sequences, as well

ΕΡ 2 379 594 Β1 as antibodies having the identified sequences, minus any signal sequences.

[0108] The term antigén binding fragment (or simply fragment) of an antibody or immunoglobulin chain (heavy or light chain), as used herein, comprises a portion (regardless of how that portion is obtained or synthesized) ofan antibody that lacks at least somé of the amino acids present in a full-length chain bút which is capable of specifically binding to an antigén. Such fragments are biologically active in that they bind specifically to the target antigén and can compete with other antigén binding proteins, including intact antibodies, for specific binding to a given epitope. In one aspect, such a fragment will retain at least one CDR present in the full-length light or heavy chain, and in somé embodiments will comprise a single heavy chain and/or light chain or portion thereof. These biologically active fragments may be produced by recombinant DNA techniques, or may be produced by enzymatic or Chemical cleavage of antigén binding proteins, including intact antibodies. Immunologically functional immunoglobulin fragments include, bút are nőt limited to, Fab, Fab’, F(ab’)₂, Fv, domain antibodies and single-chain antibodies, and may be derived from any mammalian source, including bút nőt limited to humán, mouse, rat, camelid or rabbit. It is contemplated further that a functional portion ofthe antigén binding proteins disclosed herein, for example, one or more CDRs, could be covalently bound to a second protein orto asmall molecule to create a therapeutic agent directed to a particular target in the body, possessing bifunctional therapeutic properties, or having a prolonged serum half-life.

[0109] An Fab fragment is comprised of one light chain and the C_H1 and variable regions of one heavy chain. The heavy chain of a Fab molecule cannot form a disulfide bond with another heavy chain molecule.

[0110] An Fc region contains two heavy chain fragments comprising the C_H1 and C_H2 domains of an antibody. The two heavy chain fragments are held together by two or more disulfide bonds and by hydrophobic interactions ofthe C_H3 domains.

[0111] An Fab’ fragment contains one light chain and a portion of one heavy chain that contains the V_H domain and the C_H1 domain and alsó the region between the C_H1 and C_H2 domains, such that an interchain disulfide bond can be formed between the two heavy chains of two Fab’ fragments to form an F(ab’)₂ molecule.

[0112] An F(ab’)₂ fragment contains two light chains and two heavy chains containing a portion ofthe constant region between the C_H1 and C_H2 domains, such that an interchain disulfide bond is formed between the two heavy chains. A F(ab’)₂ fragment thus is composed of two Fab’ fragments that are held together by a disulfide bond between the two heavy chains.

[0113] The Fv region comprises the variable regions from both the heavy and light chains, bút lacks the constant regions.

[0114] Single-chain antibodies are Fv molecules in which the heavy and light chain variable regions have been connected by a flexible linker to form a single polypeptide chain, which forms an antigen-binding region. Single chain antibodies are discussed in detail in International Patent Application Publication No. WO 88/01649 and United States Patent No. 4,946,778 and No. 5,260,203.

[0115] A domain antibody is an immunologically functional immunoglobulin fragment containing only the variable region of a heavy chain or the variable region ofa light chain. In somé instances, two or more V_H regions are covalently joined with a peptide linker to create a bivalent domain antibody. The two V_H regions of a bivalent domain antibody may target the same or different antigens.

[0116] A bivalent antigén binding protein or bivalent antibody comprises two antigén binding sites. In somé instances, the two binding sites have the same antigén specificities. Bivalent antigén binding proteins and bivalent antibodies may be bispecific, see, infra.

[0117] A multispecific antigén binding protein or multispecific antibody is one that targets more than one antigén or epitope.

[0118] A bispecific, dual-specific or bifunctional antigén binding protein or antibody is a hybrid antigén binding protein or antibody, respectively, having two different antigén binding sites. Bispecific antigén binding proteins and antibodies are a species of multispecific antigén binding protein or multispecific antibody and may be produced by a variety of methods including, bút nőt limited to, fusion of hybridomas or linking of Fab’ fragments. See, e.g., Songsivilai and Lachmann, 1990, Clin. Exp. Immunoi. 79:315-321; Kostelny et al., 1992, J. Immunoi. 148:1547-1553. The two binding sites of a bispecific antigén binding protein or antibody will bind to two different epitopes, which may reside on the same or different protein targets.

[0119] The term neutralizing antigén binding protein or neutralizing antibody refers to an antigén binding protein or antibody, respectively, that binds to a ligand, prevents binding ofthe ligand to its binding partner and interrupts the biological response that otherwise would result from the ligand binding to its binding partner. In assessing the binding and specificity of an antigén binding protein, e.g., an antibody or immunologically functional antigén binding fragment thereof, an antibody or fragment will substantially inhibit binding of a ligand to its binding partner when an excess of antibody reduces the quantity of binding partner bound to the ligand by at least about 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 95%, 97%, 99% or more (as measured in an in vitro competitive binding assay). In the case ofa CGRP R binding protein, such a neutralizing molecule will diminish the ability of CGRP R to bind CGRP.

[0120] The term compete, when used in the context of antigén binding proteins that may bind the same region on a

ΕΡ 2 379 594 Β1 target antigén, means competition between antigén binding proteins is determined by an assay in which the antigén binding protein (e.g., antibody or immunologically functional antigén binding fragment thereof) under test prevents or inhibits specific binding of a reference antigén binding protein (e.g., a ligand, or a reference antibody) to a common antigén (e.g., CGRP R or an antigén binding fragment thereof). Any of a number of competitive binding assays can be used, fór example: solid phase direct or indirect radioimmunoassay (RIA), solid phase direct or indirect enzyme immunoassay (EIA), sandwich competition assay (see, e.g., Stahli et al., 1983, Methods in Enzymology 9:242-253); solid phase direct biotin-avidin EIA (see, e.g., Kirkland et al., 1986, J. Immunoi. 137:3614-3619) solid phase direct labeled assay, solid phase direct labeled sandwich assay (see, e.g., Harlow and Lane, 1988, Antibodies, A Laboratory Manual, Cold Spring Harbor Press); solid phase direct label RIA using 1-125 label (see, e.g., Morei et al., 1988, Molec. Immunoi. 25:7-15); solid phase direct biotin-avidin EIA (see, e.g., Cheung, et al., 1990, Virology 176:546-552); and direct labeled RIA (Moldenhauer et al., 1990, Scand. J. Immunoi. 32:77-82). Such an assay may involve the use of purified antigén bound to a solid surface or cells bearing either of these, an unlabelled test antigén binding protein and a labeled reference antigén binding protein. Competitive inhibition may measured by determining the amount of label bound to the solid surface or cells in the presence ofthe test antigén binding protein. Antigén binding proteins identified by competition assay (competing antigén binding proteins) include antigén binding proteins binding to the same epitope as the reference antigén binding proteins and antigén binding proteins binding to an adjacent epitope sufficiently proxímal to the epitope bound by the reference antigén binding protein fór stearic hindrance to occur. Usually, when a competing antigén binding protein is present in excess, it will inhibit specific binding of a reference antigén binding protein to a common antigén by at least 40%, 45%, 50%, 55%, 60%, 65%, 70% or 75%. In somé instance, binding is inhibited by at least 80%, 85%, 90%, 95%, or 97% or more. Competitive inhibition may alsó be measured by immobilizing a reference antigén binding protein to a substrate, e.g., a sensor chip, capturing antigén on the substrate via binding to the reference antibody, and assaying whether a different antigén binding protein (a competing antigén binding protein) can additionally bind to the antigén. An example ofthe latter competitive binding assay employs a Biacore analysis, and is described in Example 7 herein.

[0121] The term antigén or immunogen refers to a molecule or a portion of a molecule capable of being bound by a selective binding agent, such as an antigén binding protein (including, e.g., an antibody or immunological functional antigén binding fragment thereof), and additionally capable of being used in an animal to produce antibodies capable of binding to that antigén. An antigén may possess one or more epitopes that are capable of interacting with different antigén binding proteins, e.g., antibodies.

[0122] The term epitope is the portion of a molecule that is bound by an antigén binding protein (fór example, an antibody). The term includes any determinant capable of specifically binding to an antigén binding protein, such as an antibody or to a T-cell receptor. An epitope can be contiguous or non-contiguous (e.g., (i) in a single-chain polypeptide, amino acid residues that are nőt contiguous to one another in the polypeptide sequence bút that within in context of the molecule are bound by the antigén binding protein, or (ii) in a multimeric receptor, e.g., CGRP R, comprising two or more individual components, e.g., RAMP1 and CRLR, amino acid residues present on two orrnore ofthe individual components, bút that within the context ofthe multimeric receptor are bound by the antigén binding protein). In certain embodiments, epitopes may be mimetic in that they comprise a three dimensional structure that is similarto an epitope used to generate the antigén binding protein, yet comprise nőne or only somé of the amino acid residues found in that epitope used to generate the antigén binding protein. Most often, epitopes reside on proteins, bút in somé instances may reside on other kinds of molecules, such as nucleic acids. Epitope determinants may include chemically active surface groupings of molecules such as amino acids, sugár side chains, phosphoryl or sulfonyl groups, and may have specific three dimensional structural characteristics, and/or specific charge characteristics. Generally, antibodies specific fór a particular target antigén will preferentially recognize an epitope on the target antigén in a complex mixture of proteins and/or macromolecules.

[0123] The term identity refers to a relationship between the sequences of two or more polypeptide molecules or two or more nucleic acid molecules, as determined by aligning and comparing the sequences. Percent identity means the percent of identical residues between the amino acids or nucleotides in the compared molecules and is calculated based on the size of the smallest of the molecules being compared. Fór these calculations, gaps in alignments (if any) must be addressed by a particular mathematical model or computer program (i.e., an algorithm). Methods that can be used to calculate the identity ofthe aligned nucleic acids or polypeptides include those described in Computational Molecular Biology, (Lesk, A. M., ed.), 1988, New York: Oxford University Press; Biocomputing Informatics and Genome Projects, (Smith, D. W., ed.), 1993, New York: Academic Press; Computer Analysis of Sequence Data, Part I, (Griffin, A. M., and Griffin, H. G., eds.), 1994, New Jersey: Humana Press; von Heinje, G., 1987, Sequence Analysis in Molecular Biology, New York: Academic Press; Sequence Analysis Primer, (Gribskov, M. and Devereux, J., eds.), 1991, New York: M. Stockton Press; and Carillo et al., 1988, SIAM J. Applied Math. 48:1073.

[0124] In calculating percent identity, the sequences being compared are aligned in away that gives the largest match between the sequences. The computer program used to determine percent identity is the GCG program package, which includes GAP (Devereux et al., 1984, Nucl. Acid Rés. 12:387; Genetics Computer Group, University of Wisconsin,

ΕΡ 2 379 594 Β1

Madison, Wl). The computer algorithm GAP is used to align the two polypeptides orpolynucleotides for which the percent sequence identity is to be determined. The sequences are aligned for optimál matching of their respective amino acid or nucleotide (the matched span, as determined by the algorithm). A gap opening penalty (which is calculated as 3x the average diagonal, wherein the average diagonal is the average of the diagonal of the comparison mátrix being used; the diagonal is the score or number assigned to each perfect amino acid match by the partieuiar comparison mátrix) and a gap extension penalty (which is usually 1/10 times the gap opening penalty), as well as a comparison mátrix such as PAM 250 or BLOSUM 62 are used in conjunction with the algorithm. In certain embodiments, a standard comparison mátrix (see, Dayhoff et al., 1978, Atlas of Protein Sequence and Structure 5:345-352 for the PAM 250 comparison mátrix; Henikoff et al., 1992, Proc. Natl. Acad. Sci. U.S.A. 89:10915-10919 for the BLOSUM 62 comparison mátrix) is alsó used by the algorithm.

[0125] Recommended parameters for determining percent identity for polypeptides or nucleotide sequences using the GAP program are the following:

Algorithm: Needleman etal., 1970, J. Mól. Bioi. 48:443-453;

Comparison mátrix: BLOSUM 62 from Henikoff et al., 1992, supra;

Gap Penalty: 12 (bút with no penalty for end gaps)

Gap Length Penalty: 4 Threshold of Similarity: 0 [0126] Certain alignment schemes for aligning two amino acid sequences may result in matching of only a short region of the two sequences, and this small aligned region may have very high sequence identity even though there is no significant relationship between the two full-length sequences. Accordingly, the selected alignment method (GAP program) can be adjusted if so desired to result in an alignment that spans at least 50 contiguous amino acids of the target polypeptide.

[0127] As used herein, substantially pure means that the described species of molecule is the predominant species present, that is, on a molar basis it is more abundant than any other individual species in the same mixture. In certain embodiments, a substantially pure molecule is a composition wherein the objeet species comprises at least 50% (on a molar basis) of all macromolecular species present. In other embodiments, a substantially pure composition will comprise at least 80%, 85%, 90%, 95%, or 99% of all macromolecular species present in the composition. In other embodiments, the objeet species is purified to essential homogeneity wherein contaminating species cannot be detected in the composition by conventional detection methods and thus the composition eonsists of a single detectable macromolecular species.

[0128] The term treating refers to any indicia of success in the treatment or amelioration of an injury, pathology or condition, including any objective or subjective paraméter such as abatement; remission; diminishing of symptoms or making the injury, pathology or condition more tolerable to the patient; slowing in the rate of degeneration or decline; making the final point of degeneration less debilitating; improving a patient’s physical or mentái well-being. The treatment or amelioration of symptoms can be based on objective or subjective parameters; including the results of a physical examination, neuropsychiatric exams, and/or a psychiatric evaluation. For example, certain methods presented herein successfully treat migraine headaches either prophylactically or as an acute treatment, decreasing the frequency of migraine headaches, decreasing the severity of migraine headaches, and/or ameliorating a symptom associated with migraine headaches.

[0129] An effective amount is generally an amount sufficient to reduce the severity and/or frequency of symptoms, eliminate the symptoms and/or underlying cause, prevent the occurrence of symptoms and/or their underlying cause, and/or improve orremediate the damage that results from oris associated with migraine headache. In somé embodiments, the effective amount is a therapeutically effective amount or a prophylactically effective amount. A therapeutically effective amount is an amount sufficient to remedy a disease state (e.g. migraine headache) or symptoms, particularly a state or symptoms associated with the disease state, or otherwise prevent, hinder, retard or reverse the progression ofthe disease state or any other undesirable symptom associated with the disease in any way whatsoever. A prophylactically effective amount is an amount ofa pharmaceutical composition that, when administered to a subject, will have the intended prophylactic effect, e.g., preventing or delaying the onset (or reoccurrence) of migraine headache, or reducing the likelihood ofthe onset (or reoccurrence) of migraine headache or migraine headache symptoms. The full therapeutic or prophylactic effect does nőt necessarily occur by administration of one dose, and may occur only after administration ofa series of doses. Thus, a therapeutically or prophylactically effective amount may be administered in one or more administrations.

[0130] Amino acid includes its normál meaning in the art. The twenty naturally-occurring amino acids and their abbreviations follow conventional usage. See, Immunology-A Synthesis, 2nd Edition, (E. S. Golub and D. R. Green, eds.), Sinauer Associates: Sunderland, Mass. (1991). Stereoisomers (e.g., D-amino acids) ofthe twenty conventional amino acids, unnatural amino acids such as α-,α-disubstituted amino acids, N-alkyl amino acids, and other unconventional

ΕΡ 2 379 594 Β1 amino acids may alsó be suitable components fór polypeptides and are included in the phrase amino acid. Examples of unconventional amino acids include: 4-hydroxyproline, γ-carboxyglutamate, ε-Ν,Ν,Ν-trimethyllysine, ε-Ν-acetyllysine, O-phosphoserine, N-acetylserine, N-formylmethionine, 3-methylhistidine, 5-hydroxylysine, σ-Ν-methylarginine, and other similar amino acids and imino acids (e.g., 4-hydroxyproline). In the polypeptide notation used herein, the left-hand direction is the amino terminál direction and the right-hand direction is the carboxyl-terminal direction, in accordance with standard usage and convention.

General OverView [0131] Antigen-binding proteins that bind CGRP R protein, including humán CGRP R (hCGRP R) protein are provided herein. The antigén binding proteins provided are polypeptides intő which one or more complementary determining regions (CDRs), as described herein, are embedded and/or joined. In somé antigén binding proteins, the CDRs are embedded intő a framework region, which orients the CDR(s) such that the proper antigén binding properties ofthe CDR(s) is achieved. In generál, antigén binding proteins that are provided can interfere with, block, reduce or modulate the interaction between CGRP and CGRP R.

[0132] Certain antigén binding proteins described herein are antibodies or are derived from antibodies. In certain embodiments, the polypeptide structure ofthe antigén binding proteins is based on antibodies, including, bút nőt limited to, monoclonal antibodies, bispecific antibodies, minibodies, domain antibodies, synthetic antibodies (sometimes referred to herein as antibody mimetics), chimeric antibodies, humanized antibodies, humán antibodies, anti body fusions (sometimes referred to herein as antibody conjugates), and fragments thereof. The various structures are further described herein below.

[0133] The antigén binding proteins provided herein have been demonstrated to bind to CGRP R, in particular humán CGRP R. As described further in the examples below, certain antigén binding proteins were tested and found to bind to epitopes different from those bound by a numberof other antibodies directed against one orthe other ofthe components of CGRP R. The antigén binding proteins that are provided compete with CGRP and thereby prevent CGRP from binding to its receptor. As a consequence, the antigén binding proteins provided herein are capable of inhibiting CGRP R activity. In particular, antigén binding proteins binding to these epitopes can have one or more of thefollowing activities: inhibiting, inter alia, induction of CGRP R signal transduction pathways, inhibiting vasodialation, causing vasoconstriction, decreasing inflammation, e.g., neurogenic inflammation, and other physiological effects induced by CGRP R upon CGRP binding. [0134] The antigén binding proteins that are disclosed herein have a variety of Utilities. Somé ofthe antigén binding proteins, fór instance, are useful in specific binding assays, affinity purification of CGRP R, in particular hCGRP R or its ligands and in screening assays to identify other antagonists of CGRP R activity. Somé of the antigen-binding proteins are useful fór inhibiting binding of CGRP to CGRP R.

[0135] The antigen-binding proteins can be used in a variety of treatment applications, as explained herein. Fór example, certain CGRP R antigen-binding proteins are useful fortreating conditions associated with CGRP R mediated signaling, such as reducing, alleviating, or treating the frequency and/or severity of migraine headache, reducing, alleviating, or treating cluster headache, reducing, alleviating, ortreating chronic pain, alleviating ortreating diabetes mellitus (type II), reducing, alleviating, ortreating cardiovasculardisorders, and reducing, alleviating, ortreating hemodynamic derangements associated with endotoxemia and sepsis in a patient. Other uses fór the antigén binding proteins include, fór example, diagnosis of CGRP R-associated diseases or conditions and screening assays to determine the presence or absence of CGRP R. Somé ofthe antigén binding proteins described herein are useful in treating consequences, symptoms, and/or the pathology associated with CGRP R activity. These include, bút are nőt limited to, various types of migraine headaches.

CGRP Receptor [0136] The antigén binding proteins disclosed herein bind to CGRP R, in particular humán CGRP R. CGRP R is a multimer that includes both CRLR and RAMP1. The nucleotide sequence of humán CRLR is provided herein as SEQ ID NO:1. The amino acid sequence of humán CRLR is provided herein as SEQ ID NO:2. The nucleotide sequence of humán RAMP1 is provided herein as SEQ ID NO:3. The amino acid sequence of humán RAMP1 is provided herein as SEQ ID NO:4. The antigén binding proteins described herein bind the extracellular portion of CGRP R, which comprises the extracellular portions of CRLR and RAMP1. An exemplary extracellular domain (ECD) of humán CRLR is encoded by the nucleotide sequence presented as SEQ ID NO:5, and has the amino acid sequence presented as SEQ ID NO:6. This sequence includes a signal peptide; an exemplary mature (minus the signal peptide) CRLR ECD has the amino acid sequence presented asSEQ ID NO: 10. An exemplary ECD of humán RAMP1 is encoded bythe nucleotide sequence presented as SEQ ID NO:7, and has the amino acid sequence presented as SEQ ID NO:8. This sequence includes a signal peptide; an exemplary mature (minus the signal peptide) RAMP1 ECD has the amino acid sequence presented as SEQ ID NO:11. As described below, CGRP R proteins may alsó include fragments. As used herein, the terms are

ΕΡ 2 379 594 Β1 used interchangeablyto mean a receptor, in particular, unless otherwise specified, a humán receptorthat binds specifically to CGRP.

[0137] The term CGRP R alsó includes post-translational modifications ofthe CGRP R amino acid sequence, fór example, possible N-linked glycosylation sites. Thus, the antigén binding proteins may bind to or be generated from proteins glycosylated at one or more ofthe positions.

CGRP Receptor Binding Proteins [0138] A variety of selective binding agents useful fór regulating the activity of CGRP R are provided. These agents include, fór instance, antigén binding proteins that contain an antigén binding domain (e.g., single chain antibodies, domain antibodies, immunoadhesions, and polypeptides with an antigén binding region) and specifically bind to CGRP R, in particular humán CGRP R. Somé of the agents, fór example, are useful in inhibiting the binding of CGRP to CGRP R, and can thus be used to inhibit, interfere with or modulate one or more activities associated with CGRP R signaling. [0139] In generál, the antigén binding proteins that are provided typically comprise one or more CDRs as described herein (e.g., 1,2,3,4,5 or 6). In somé instances, the antigén binding protein comprises (a) a polypeptide strueture and (b) one or more CDRs that are inserted intő and/or joined to the polypeptide strueture. The polypeptide strueture can take a variety of different forms. Fór example, it can be, or comprise, the framework of a naturally occurring antibody, or fragment or variant thereof, or may be completely synthetic in natúré. Examples of various polypeptide structures are further described below.

[0140] In certain embodiments, the polypeptide strueture ofthe antigén binding proteins is an antibody or is derived from an antibody, including, bút nőt limited to, monoclonal antibodies, bispecific antibodies, minibodies, domain antibodies, synthetic antibodies (sometimes referred to herein as antibody mimetics), chimeric antibodies, humanized antibodies, antibody fusions (sometimes referred to as antibody conjugates), and portions or fragments of each, respectively. In somé instances, the antigén binding protein is an immunological fragment ofan antibody (e.g., a Fab, a Fab’, a F(ab’)₂, or a scFv). The various structures are further described and defined herein.

[0141] Certain ofthe antigén binding proteins as provided herein specifically bind to humán CGRP R. In a specific embodiment, the antigén binding protein specifically binds to humán CGRP R protein comprising humán CRLR having the amino acid sequence of SEQ ID NO:2 and humán RAMP1 having the amino acid sequence of SEQ ID NO:4. [0142] In embodiments where the antigén binding protein is used fór therapeutic applications, an antigén binding protein can inhibit, interfere with or modulate one or more biological activities of CGRP R. In this case, an antigén binding protein binds specifically and/or substantially inhibits binding of humán CGRP R to CGRP when an excess of antibody reduces the quantity of humán CGRP R bound to CGRP, or vice versa, by at least about 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 95%, 97%, 99% ormore (fór example by measuring binding in an/'n vitro competitive binding assay).

Naturally Occurring Antibody Strueture [0143] Somé ofthe antigén binding proteins that are provided have the strueture typically associated with naturally occurring antibodies. The structural units of these antibodies typically comprise one or more tetramers, each composed of two identical couplets of polypeptide chains, though somé species of mammals alsó produce antibodies having only a single heavy chain. In a typical antibody, each pair or couplet includes one full-length light chain (in certain embodiments, about 25 kDa) and one full-length heavy chain (in certain embodiments, about 50-70 kDa). Each individual immunoglobulin chain is composed of several immunoglobulin domains, each consisting of roughly 90 to 110 amino acids and expressing a characteristic folding pattern. These domains are the basic units of which antibody polypeptides are composed. The amino-terminal portion of each chain typically includes a variable domain that is responsible fór antigén recognition. The carboxy-terminal portion is more conserved evolutionarily than the other end ofthe chain and is referred to as the constant region or C region. Humán light chains generally are classified as kappa and lambda light chains, and each of these contains one variable domain and one constant domain. Heavy chains are typically classified as mu, delta, gamma, alpha, or epsilon chains, and these define the antibody’s isotype as IgM, IgD, IgG, IgA, and IgE, respectively. IgG has several subtypes, including, bút nőt limited to, IgG 1, lgG2, lgG3, and lgG4. IgM subtypes include IgM, and lgM2. IgA subtypes include lgA1 and lgA2. In humans, the IgA and IgD isotypes contain four heavy chains and four light chains; the IgG and IgE isotypes contain two heavy chains and two light chains; and the IgM isotype contains five heavy chains and five light chains. The heavy chain C region typically comprises one or more domains that may be responsible fór effector function. The number of heavy chain constant region domains will depend on the isotype. IgG heavy chains, fór example, each contain three C region domains known asC_H1, C_H2 and C_H3. The antibodies that are provided can have any of these isotypes and subtypes. In certain embodiments, the CGRP R antibody is ofthe IgG 1, lgG2, or lgG4 subtype.

[0144] In full-length light and heavy chains, the variable and constant régions are joined by a J region of about twelve or more amino acids, with the heavy chain alsó including a D region of about ten more amino acids. See, e.g., Fun22

ΕΡ 2 379 594 Β1 damental Immunology, 2nd ed., Ch. 7 (Paul, W., ed.) 1989, New York: Raven Press. The variable regions of each light/heavy chain pair typically form the antigén binding site.

[0145] One example ofan lgG2 heavy constant domain ofan exemplary CGRP R monoclonal antibody has the amino acid sequence:

ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFP

AVLQSSGLYSLSSVVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPPCPAP

PVAGPSVFLFPPKPKDTLMISRTPEVTCWVDVSHEDPEVQFNWYVDGVEVHNAKTK

PREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLPAPIEKTISKTKGQPREPQV

YTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFL YSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (the last 326 residues of the sequence shown as SEQ. ID NO:29).

[0146] One example of a kappa light Constant domain of an exemplary CGRP R monoclonal antibody has the amino acid sequence:

RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQ

ESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (the last 107 residues ofthe sequence shown as SEQ ID NO:14).

[0147] Variable regions of immunoglobulin chains generally exhibit the same overall structure, comprising relatively conserved framework regions (FR) joined by three hypervariable regions, more often called complementarity determining regions or CDRs. The CDRs from the two chains of each heavy chain/light chain pair mentioned above typically are aligned by the framework regions to form a structure that binds specifically with a specific epitope on the target protein (e.g., CGRP R). From N-terminal to C-terminal, naturally-occurring light and heavy chain variable regions both typically conform with the following order of these elements: FR1, CDR1, FR2, CDR2, FR3, CDR3 and FR4. A numbering system has been devised for assigning numbers to amino acids that occupy positions in each of these domains. This numbering system is defined in Kábát Sequences of Proteins of Immunological Interest (1987 and 1991, NIH, Bethesda, MD), or Chothia &amp; Lesk, 1987, J. Mól. Bioi. 196:901-917; Chothia et al., 1989, Natúré 342:878-883.

[0148] The various heavy chain and light chain variable regions provided herein are depicted in Table 3. Each of these variable regions may be attached to the above heavy and light chain constant regions to form a complete antibody heavy and light chain, respectively. Further, each ofthe so generated heavy and light chain sequences may be combined to form a complete antibody structure. lt should be understood that the heavy chain and light chain variable regions provided herein can alsó be attached to other constant domains having different sequences than the exemplary sequences listed above.

[0149] Specific examples of somé of the full length light and heavy chains of the antibodies that are illustrated and their corresponding amino acid sequences are summarized in Tables 2A and 2B. Table 2A shows exemplary light chain sequences, and Table 2B shows exemplary heavy chain sequences.

EP 2 379 594 B1

Table 2A - Exemplary Antibody Light Chain Amino Acid Sequences

<td> Sequence</td><td> MDMRVPAQLLGLLLLWLRGARCQSVLTQPPSVSEAPGQKVTISC SGSSSNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSGIPDRFSGS KSGTSATLGITGLQTGDEADYYCGTWDSRLSAVVFGGGTKLTVL GQPKANPTVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKAD GSPVKAGVETTKPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQ VTHEGSTVEKTVAPTECS</td><td> MDMRVPAQLLGLLLLWLRGARCQSVLTQPPSASGTPGQRVTISC SGSSSNIGSNYVYWYQQLPGAAPKLLIFRSNQRPSGVPDRFSGS KSGTSASLAISGLRSEDEADYYCAAWDDSLSGWVFGGGTKLTVL GQPKANPTVTLFPPSSEELQANKATLVCUSDFYPGAVTVAWKAD GSPVKAGVETTKPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQ VTHEGSTVEKTVAPTECS</td><td> MDMRVPAQLLGLLLLWLRGARCDIQMTQSPSSLSASVGDRVTIT CRASQGIRNDLGWFQQKPGKAPKRLIYAASSLQSGVPSRFSGS GSGTEFTLTISSLQPEDLATYYCLQYNIYPWTFGQGTKVEIKRTVA APSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQ SGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC</td>

<td> Contained in Clone</td><td> 01E11 LC</td><td> 01H7LC</td><td> o _l UJ CM O</td>

<td> Designation D</td><td> _l</td><td> L2</td><td> CO _l</td>

<td> SEQ ID NO:</td><td> CM</td><td> CO</td><td> ’xt</td>

ΕΡ 2 379 594 Β1

<td> Sequence</td><td> MDMRVPAQLLGLLLLWLRGARCSSELTQDPTVSVALGQTVKITC QGDSLRSFYASWYQQKPGQAPVLVFYGKNNRPSGIPDRFSGSS SGNTASLTITGAQAEDEADYYCNSRDSSVYHLVLGGGTKLTVLG QPKANPTVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADG SPVKAGVETTKPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQV THEGSTVEKTVAPTECS</td><td> MDMRVPAQLLGLLLLWLRGARCDIILAQTPLSLSVTPGQPASISC KSSQSLLHSAGKTYLYWYLQKPGQPPQLLIYEVSNRFSGVPDRF SGSGSGTDFTLKISRVEAEDVGIYYCMQSFPLPLTFGGGTKVEIK RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVD NALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACE VTHQGLSSPVTKSFNRGEC</td><td> MDMRVPAQLLGLLLLWLRGARCQSVLTQPPSVSAAPGQKVTISC</td><td> SGSSSNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSGIPDRFSGS KSGTSTTLGITGLQTGDEADYYCGTWDSRLSAVVFGGGTKLTVL GQPKANPTVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKAD GSPVKAGVETTKPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQ VTHEGSTVEKTVAPTECS</td>

<td> Contained in Clone</td><td> 03B6 LC</td><td> o _l 00 O co o</td><td> o _l 'xt LU 'χίΟ</td><td></td>

<td> Designation D</td><td> L4</td><td> in _l</td><td> CD _l</td><td></td>

<td> SEQ ID NO:</td><td> in</td><td> CD</td><td></td><td></td>

ΕΡ 2 379 594 Β1

<td> Sequence</td><td> MDMRVPAQLLGLLLLWLRGARCDIVMTQSPLSLPVTPGEPASISC RSSQSLLHSFGYNYLDWYLQKPGQSPQLLIYLGSNRASGVPDRF SGSGSGTDFTLKISRVEAEDVGVYYCMQALQTPFTFGPGTKVDI KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKV DNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYAC EVTHQGLSSPVTKSFNRGEC</td><td> MDMRVPAQLLGLLLLWLRGARCDIILTQTPLSLSVTPGQPASISC KSSQSLLHSDGKTYLYWYLQKPGQPPQLLIYEVSNRFSGEPDRF SGSGSGTDFTLKISRVEAEDVGTYYCMQSFPLPLTFGGGTKVEIK RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVD NALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACE VTHQGLSSPVTKSFNRGEC</td><td> MDMRVPAQLLGLLLLWLRGARCQSVLTQPPSVSAAPGQKVTISC SGSSSNIGNNYVSWYQQFPGTAPKLLIYDNNKRPSGIPDRFSGS KSGTSATLGITGLQTGDEADYYCGTWDSRLSAVVFGGGTKLTVL GQPKANPTVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKAD GSPVKAGVETTKPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQ VTHEGSTVEKTVAPTECS</td>

<td> Contained in Clone</td><td> 04H6 LC</td><td> 05F5 LC</td><td> 09D4 LC</td>

<td> Designation D</td><td> _l</td><td> 00 _l</td><td> O) _l</td>

<td> SEQ ID NO:</td><td> 00</td><td> O)</td><td> 20</td>

EP 2 379 594 Β1

<td> Sequence</td><td> MDMRVPAQLLGLLLLWLRGARCQSVLTQSPSASGTPGQRVTISC SGSSSNIGSNYVYWYQQLPGAAPKLLILRNNQRPSGVPDRFSGS KSGTSASLTISGLRSEDEADYYCAAWDDSLSGWVFGGGTKLTVL GQPKANPTVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKAD GSPVKAGVETTKPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQ VTHEGSTVEKTVAPTECS</td><td> MDMRVPAQLLGLLLLWLRGARCQSVLTQPPSASGTPGQRVTISC SGSSSNIGSNTVNWYQQLPGTAPKLLIYTNNQRPSGVPDRFSGS KSGTSASLAISGLQSEDEADFYCAARDESLNGVVFGGGTKLTVL GQPKANPTVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKAD GSPVKAGVETTKPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQ</td><td> VTHEGSTVEKTVAPTECS</td><td> MDMRVPAQLLGLLLLWLRGARCQSVLTQPPSASGTPGQRVTISC SGSSSNIGSNYVYWYQQLPGAAPKLLIFRNNQRPSGVPDRFSGS KSGTSASLAISGLRSEDEADYYCAAWDDSLSGWVFGGGTKLTVL GQPKANPTVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKAD GSPVKAGVETTKPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQ VTHEGSTVEKTVAPTECS</td>

<td> Contained in Clone</td><td> 09F5 LC</td><td> o _l 'xt LU O</td><td></td><td> 11D11 HL 11H9LC</td>

<td> Designation D</td><td> L10</td><td> _l</td><td></td><td> L12</td>

<td> SEQ ID NO:</td><td> CM</td><td> 22</td><td></td><td> 23</td>

EP 2 379 594 Β1

<td> Sequence</td><td> MDMRVPAQLLGLLLLWLRGARCDITLTQTPLSLSVSPGQPASISC KSSQSLLHSDGRNYLYWYLQKPGQPPQLLIYEVSNRFSGLPDRF SGSGSGTDFTLKISRVEAEDVGIYYCMQSFPLPLTFGGGTKVEIK RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVD NALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACE VTHQGLSSPVTKSFNRGEC</td><td> MDMRVPAQLLGLLLLWLRGARCQSVLTQPPSVSAAPGQKVTISC SGSSSNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSGIPDRFSGS KSGTSATLGITGLQTGDEADYYCGTWDSRLSAVVFGGGTKLTVL GQPKANPTVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKAD GSPVKAGVETTKPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQ VTHEGSTVEKTVAPTECS</td><td> MDMRVPAQLLGLLLLWLRGARCDIQMTQSPSSLSASVGDRVTIT CRASQGIRKDLGWYQQKPGKAPKRLIYGASSLQSGVPSRFSGS GSGTEFTLTISSLQPEDFATYYCLQYNSFPWTFGQGTKVEIKRTV AAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNAL QSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTH QGLSSPVTKSFNRGEC</td>

<td> Contained in Clone</td><td> 12E8LC</td><td> 12G8 HL</td><td> 13H2 LC</td>

<td> Designation D</td><td> L13</td><td> L14</td><td> L15</td>

<td> SEQ ID NO:</td><td> 24</td><td> 25</td><td> 26</td>

ΕΡ 2 379 594 Β1

<td> Sequence</td><td> METPAQLLFLLLLWLPDTTGEIVLTQSPGTLSLSPGERATLSCRA SQSVSSGYLTWYQQKPGQAPRLLIYGASSRATGIPDRFSGSGSG TDFTLTISRLEPEDFAVYYCQQYGNSLCRFGQGTKLEIKRTVAAP SVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSG NSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGL SSPVTKSFNRGEC</td><td> METPAQLLFLLLLWLPDTTGEIVLTQSPGTLSLSPGERATLSCRA SQSVSSGYLTWYQQKPGQAPRLUYGASSRATGIPDRFSGSGSG TDFTLTISRLEPEDFAVYYCQQYGNSLSRFGQGTKLEIKRTVAAP</td><td> SVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSG NSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGL SSPVTKSFNRGEC</td>

<td> Contained in Clone</td><td> 32H7 LC</td><td> 32H7 CS LC</td><td></td>

<td> Designation D</td><td> L16</td><td> L17</td><td></td>

<td> SEQ ID NO:</td><td> 27</td><td> 28</td><td></td>

ΕΡ 2 379 594 Β1

Table 2Β - Exemplary Antibody Heavy Chain Amino Acid Sequences

<td> SEQ IDNO:</td><td> Design ation</td><td> Contained in Clone</td><td> Sequence</td>

<td> 29</td><td> H1</td><td> 01E11 HC 04E4 HC 09D4 HC</td><td> MDMRVPAQLLGLLLLWLRGARCQVQLVESGGGVVQPGRSLRLS CAASGFTFSSFGMHWVRQAPGKGLEWVAVISFDGSIKYSVDSVK GRFTISRDNSKNTLFLQMNSLRAEDTAVYYCARDRLNYYDSSGY YHYKYYGMAVWGQGTTVTVSSASTKGPSVFPLAPCSRSTSEST AALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLS SVVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPPCP APPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFN WYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEY KCKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVS LTCLVKGFYPSDIAVEWESNGQPENNYKKPPMLDSDGSFFLYS KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK</td>

<td> 30</td><td> H2</td><td> 01 H7 HC</td><td> MDMRVPAQLLGLLLLWLRGARCEVQLVESGGGLVKPGGSLRLS CAASGFTFSNAWMSWVRQAPGKGLEWVGRIKSKDGGTTDYAA PVKGRFTISRDDSKNTLYLQMNSLKTEDTAVYYCHDRTGYSISW SSYYYYYGMDVWGQGTTVTVSSASTKGPSVFPLAPCSRSTSES TAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSL SSVVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPPC PAPPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQF NWYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKE YKCKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQV SLTCLVKGFYPSDIAVEWESNGQPENNYKKPPMLDSDGSFFLY SKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK</td>

<td> 31</td><td> H3</td><td> 02E7 HC</td><td> MDMRVPAQLLGLLLLWLRGARCEVQLLESGGGLVQPGESLRLS CAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGRTYYADSV KGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKDQREVGPYSS GWYDYYYGMDVWGQGTTVTVSSASTKGPSVFPLAPCSRSTSES TAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSL</td>

ΕΡ 2 379 594 Β1 (continued)

<td> SEQ IDNO:</td><td> Design ation</td><td> Contained in Clone</td><td> Sequence</td>

<td></td><td></td><td></td><td> SSVVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPPC PAPPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQF NWYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKE YKCKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQV SLTCLVKGFYPSDIAVEWESNGQPENNYKKPPMLDSDGSFFLY SKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK</td>

<td> 32</td><td> H4</td><td> 03B6 HC</td><td> MDMRVPAQLLGLLLLWLRGARCQVQLVQSGAEVKKPGASVKVS CKASGYTFTGYYMHWVRQAPGQGLEWMGWINPNSGGTNYAQK FQGRVTMTRDTSISTAYMELSRLRSDDTAVYFCARDQMSIIMLRG VFPPYYYGMDVWGQGTTVTVSSASTKGPSVFPLAPCSRSTSEST AALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLS SVVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPPCP APPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFN WYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEY KCKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVS LTCLVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYS KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK</td>

<td> 33</td><td> H5</td><td> 03C8 HC 05F5 HC 12E8 HC</td><td> MDMRVPAQLLGLLLLWLRGARCQVQLVESGGGVVQPGRSLRLS CAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSHESYADSV KGRFTISRDISKNTLYLQMNSLRAEDTAVYFCARERKRVTMSTLY YYFYYGMDVWGQGTTVTVSSASTKGPSVFPLAPCSRSTSESTAA LGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSV VTVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPPCPAP PVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNW YVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYK CKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSL TCLVKGFYPSDIAVEWESNGQPENNYKKPPMLDSDGSFFLYSK LTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK</td>

ΕΡ 2 379 594 Β1 (continued)

<td> SEQ IDNO:</td><td> Design ation</td><td> Contained in Clone</td><td> Sequence</td>

<td> 34</td><td> H6</td><td> 04H6 HC</td><td> MDMRVPAQLLGLLLLWLRGARCEVQLVESGGGLVKPGRSLRLS CTASGFTFGDYAMSWFRQAPGKGLEWIGFIRSRAYGGTPEYAAS VKGRFTISRDDSKTIAYLQMNSLKTEDTAVYFCARGRGIAARWDY WGQGTLVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYF PEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSNFG TQTYTCNVDHKPSNTKVDKTVERKCCVECPPCPAPPVAGPSVFL</td>

<td></td><td></td><td></td><td> FPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGVEVHN AKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLPA PIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPS DIAVEWESNGQPENNYKKPPMLDSDGSFFLYSKLTVDKSRWQ QGNVFSCSVMHEALHNHYTQKSLSLSPGK</td>

<td> 35</td><td> H7</td><td> 09F5 HC</td><td> MDMRVPAQLLGLLLLWLRGARCEVQLVESGGGLVKPGGSLRLS CAASGFTFSNAWMSWVRQAPGKGLEWVGRIKSKTDGGTTDYTA PVKGRFTISRDDSKNTLYLQMNSLKAEDTAVYYCTTDRTGYSISW SSYYYYYGMDVWGQGTTVTVSSASTKGPSVFPLAPCSRSTSES TAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSL SSVVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPPC PAPPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQF NWYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKE YKCKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQV SLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLY SKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK</td>

ΕΡ 2 379 594 Β1 (continued)

<td> SEQ IDNO:</td><td> Design ation</td><td> Contained in Clone</td><td> Sequence</td>

<td> 36</td><td> H8</td><td> 10E4 HC</td><td> MDMRVPAQLLGLLLLWLRGARCQVQLVQSGAEVKKPGASVKVS CKASGYTFTDYYMYWVRQAPGQGLEWMGWISPNSGGTNYAQK FQGRVTMTRDTSISTAYMELSRLRSDDTAVYYCVRGGYSGYAGL YSHYYGMDVWGQGTTVTVSSASTKGPSVFPLAPCSRSTSESTA ALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSS VVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPPCPA PPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFN WYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEY KCKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVS LTCLVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYS KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK</td>

<td> 37</td><td> H9</td><td> 11D11 HC</td><td> MDMRVPAQLLGLLLLWLRGARCEVQLVESGGGLVKPGGSLRLS CAASGFTFGNAWMSWVRQAPGKGLEWVGRIKSKTDGGTTDYA ΑΡνκορπιεροοεκΝτίΥίαΜΝεικτΕοτΑνγροπορτθΥειε WSSYYYYYGMDVWGQGTTVTVSSASTKGPSVFPLAPCSRSTSE STAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYS LSSVVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPP CPAPPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQ</td>

<td></td><td></td><td></td><td> FNWYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGK EYKCKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQ VSLTCLVKGFYPSDIAVEWESNGQPENNYKHPPMLDSDGSFFL YSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK</td>

ΕΡ 2 379 594 Β1 (continued)

<td> SEQ IDNO:</td><td> Design ation</td><td> Contained in Clone</td><td> Sequence</td>

<td> 38</td><td> H10</td><td> 11H9HC</td><td> MDMRVPAQLLGLLLLWLRGARCEVQLVESGGGLVKPGGSLRLS CAASGFTFGNAWMSWVRQAPGKGLEWVGRIKSKTDGGKDYA APVKGRFTISRDDSKNTLYLQMNSLKTEDTAVYYCTTDRTGYSIS WSSYYYYYGMDVWGQGTTVTVSSASTKGPSVFPLAPCSRSTSE STAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYS LSSVVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPP CPAPPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQ FNWYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGK EYKCKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQ VSLTCLVKGFYPSDIAVEWESNGQPENNYKKPPMLDSDGSFFL YSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK</td>

<td> 39</td><td> H11</td><td> 12G8 HC</td><td> MDMRVPAQLLGLLLLWLRGARCQVQLVESGGGVVQPGRSLRLS CAASGFTFSSFGMHWVRQAPGKGLEWVAVISFDGSIKYSVDSVK GRFTISRDNSKNTLFLQMNSLRAEDTAVYYCARDRLNYYDSSGY YHYKYYGLAVWGQGTTVTVSSASTKGPSVFPLAPCSRSTSESTA ALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSS VVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPPCPA PPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFN WYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEY KCKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVS LTCLVKGFYPSDIAVEWESNGQPENNYKKPPMLDSDGSFFLYS KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK</td>

<td> 40</td><td> H12</td><td> 13H2 HC</td><td> MDMRVPAQLLGLLLLWLRGARCEVQLVESGGGLVKPGGSLRLS CAASGYTFSTYSMNWVRQAPGKGLEWVSSISSSSSYRYYADSV KGRFTISRDNAKNSLYLQMSSLRAEDTAVYYCAREGVSGSSPYSI SWYDYYYGMDVWGQGTTVTVSSASTKGPSVFPLAPCSRSTSES TAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSL SSVVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPPC PAPPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQF NWYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKE</td>

ΕΡ 2 379 594 Β1 (continued)

<td> SEQ IDNO:</td><td> Design ation</td><td> Contained in Clone</td><td> Sequence</td>

<td></td><td></td><td></td><td> YKCKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQV SLTCLVKGFYPSDIAVEWESNGQPENNYKKPPMLDSDGSFFLY SKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK</td>

<td> 41</td><td> H13</td><td> 32H7 HC</td><td> MDMRVPAQLLGLLLLWLRGARCQVQLVESGGGVVQPGRSLRLS CAASGFTFSSYGMHWVRQAPGKGLEWVAVIWYDGSNKYYADS VKGRFIISRDKSKNTLYLQMNSLRAEDTAVYYCARAGGIAAAGLY YYYGMDVWGQGTTVTVSSASTKGPSVFPLAPCSRSTSESTAAL GCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVV TVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPPCPAPP VAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWY VDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKC KVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLT CLVKGFYPSDIAVEWESNGQPENNYKKPPMLDSDGSFFLYSKL TVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK</td>

[0150] The first 22 amino acids of each of the light chain sequences in Table 2A, except 32H7 and 32H7 CS, is a signal sequence. In the case of 32H7 and 32H7 CS, the signal sequence is 20 amino acids. Similarly, the first 22 amino acids of each ofthe heavy chain sequences in Table 2B is a signal sequence. The signal peptides may be changed to signal peptides having different sequences, e.g., for more optimál expression in certain hőst cells. lt will be therefore be understood that the invention alsó includes antibodies having the light and/or heavy chain sequences as specified in Tables 2A and 2B, bút with different signal sequences.

[0151] Again, each ofthe exemplary heavy chains (H1, H2, H3 etc.) listed in Table 2B can be combined with any of the exemplary light chains shown in Table 2A to form an antibody. Examples of such combinations include H1 combined with any of L1 through L17; H2 combined with any of L1 through L17; H3 combined with any of L1 through L17, and so on. In somé instances, the antibodies include at least one heavy chain and one light chain from those listed in Tables 2A and 2B. In somé instances, the antibodies comprise two different heavy chains and two different light chains listed in Tables 2A and 2B. In other instances, the antibodies contain two identícai light chains and two identícai heavy chains. As an example, an antibody or immunologically functional fragment may include two H1 heavy chains and two L1 light chains, ortwo H2 heavy chains and two L2 light chains, ortwo H3 heavy chains and two L3 light chains and other similar combinations of pairs of light chains and pairs of heavy chains as listed in Tables 2A and 2B.

[0152] Other antigén binding proteins that are illustrated are variants of antibodies formed by combination ofthe heavy and light chains shown in Tables 2A and 2B and comprise light and/or heavy chains that each have at least 70%, 75%, 80%, 85%, 90%, 95%, 97% or 99% identity to the amino acid sequences of these chains. In somé instances, such antibodies include at least one heavy chain and one light chain, whereas in other instances the variant forms contain two identícai light chains and two identícai heavy chains.

Variable Domains of Antibodies [0153] Alsó illustrated are antigén binding proteins that contain an antibody heavy chain variable region selected from the group consisting ofV_H1, V_H2, V_H3, V_H4, V_H5, V_H6, V_H7, V_H8, V_H9, V_H10, V_H11, V_H12, and V_H13, and/or an antibody light chain variable region selected from the group consisting of V_L1, V_L2, V_L3, V_L4, V_L5, V_L6, V_L7, V_L8, V_L9, V_L10, V_L11, V_L12, V_l13, V_l14, V_l15, V_l16, and V_L17, as shown in Table 3 below, and immunologically functional fragments, derivatives, muteins and variants of these light chain and heavy chain variable regions.

ΕΡ 2 379 594 Β1 [0154] Sequence alignments ofthe various heavy and light chain variable regions, respectively, are provided in Figs. 1A and 1B.

[0155] Antigén binding proteins of this type can generally be designated by the formula V_Hx/ V_Ly, where x corresponds to the numberof heavy chain variable regions and y corresponds to the number ofthe light chain variable regions.

Table 3: Exemplary V_H and V_L Chain Amino Acid Sequences

<td> Contained in Clone</td><td> Designation</td><td> SEQ ID NO.</td><td> Amino Acid Sequence</td>

<td> 1E11</td><td> V_L1</td><td> 137</td><td> QSVLTQPPSVSEAPGQKVTISCSGSSSNIGNNYVSWYQQLP GTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATLGITGLQTGDE ADYYCGTWDSRLSAVVFGGGTKLTVL</td>

<td> 1H7</td><td> V_l2</td><td> 138</td><td> QSVLTQPPSASGTPGQRVTISCSGSSSNIGSNYVYWYQQLP GAAPKLLIFRSNQRPSGVPDRFSGSKSGTSASLAISGLRSED EADYYCAAWDDSLSGWVFGGGTKLTVL</td>

<td> 2E7</td><td> V_l3</td><td> 139</td><td> DIQMTQSPSSLSASVGDRVTITCRASQGIRNDLGWFQQKPG KAPKRLIYAASSLQSGVPSRFSGSGSGTEFTLTISSLQPEDLA TYYCLQYNIYPWTFGQGTKVEIK</td>

<td> 3B6</td><td> V_l4</td><td> 140</td><td> SSELTQDPTVSVALGQTVKITCQGDSLRSFYASWYQQKPGQ APVLVFYGKNNRPSGIPDRFSGSSSGNTASLTITGAQAEDEA DYYCNSRDSSVYHLVLGGGTKLTVL</td>

<td> 3C8</td><td> V_l5</td><td> 141</td><td> DIILAQTPLSLSVTPGQPASISCKSSQSLLHSAGKTYLYWYLQ KPGQPPQLLIYEVSNRFSGVPDRFSGSGSGTDFTLKISRVEA EDVGIYYCMQSFPLPLTFGGGTKVEIK</td>

<td> 4E4</td><td> V_l6</td><td> 142</td><td> QSVLTQPPSVSAAPGQKVTISCSGSSSNIGNNYVSWYQQLP GTAPKLLIYDNNKRPSGIPDRFSGSKSGTSTTLGITGLQTGDE ADYYCGTWDSRLSAVVFGGGTKLTVL</td>

<td> 4H6</td><td> V_l7</td><td> 143</td><td> DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSFGYNYLDWYL QKPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVE AEDVGVYYCMQALQTPFTFGPGTKVDIK</td>

<td> 5F5</td><td> V_l8</td><td> 144</td><td> DIILTQTPLSLSVTPGQPASISCKSSQSLLHSDGKTYLYWYLQ KPGQPPQLLIYEVSNRFSGEPDRFSGSGSGTDFTLKISRVEA EDVGTYYCMQSFPLPLTFGGGTKVEIK</td>

<td> 9D4</td><td> V_l9</td><td> 145</td><td> QSVLTQPPSVSAAPGQKVTISCSGSSSNIGNNYVSWYQQFP GTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATLGITGLQTGDE ADYYCGTWDSRLSAVVFGGGTKLTVL</td>

ΕΡ 2 379 594 Β1 (continued)

<td> Contained in Clone</td><td> Designation</td><td> SEQ ID NO.</td><td> Amino Acid Sequence</td>

<td> 9F5</td><td> V_L10</td><td> 146</td><td> QSVLTQSPSASGTPGQRVTISCSGSSSNIGSNYVYWYQQLP GAAPKLLILRNNQRPSGVPDRFSGSKSGTSASLTISGLRSED EADYYCAAWDDSLSGWVFGGGTKLTVL</td>

<td> 10E4</td><td> V_L11</td><td> 147</td><td> QSVLTQPPSASGTPGQRVTISCSGSSSNIGSNTVNWYQQLP GTAPKLLIYTNNQRPSGVPDRFSGSKSGTSASLAISGLQSED EADFYCAARDESLNGVVFGGGTKLTVL</td>

<td> 11D1111H9</td><td> V_l12</td><td> 148</td><td> QSVLTQPPSASGTPGQRVTISCSGSSSNIGSNYVYWYQQLP GAAPKLLIFRNNQRPSGVPDRFSGSKSGTSASLAISGLRSED EADYYCAAWDDSLSGWVFGGGTKLTVL</td>

<td> 12E8</td><td> V_l13</td><td> 149</td><td> DITLTQTPLSLSVSPGQPASISCKSSQSLLHSDGRNYLYWYL QKPGQPPQLLIYEVSNRFSGLPDRFSGSGSGTDFTLKISRVE AEDVGIYYCMQSFPLPLTFGGGTKVEIK</td>

<td> 12G8</td><td> V_l14</td><td> 150</td><td> QSVLTQPPSVSAAPGQKVTISCSGSSSNIGNNYVSWYQQLP GTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATLGITGLQTGDE ADYYCGTWDSRLSAVVFGGGTKLTVL</td>

<td> 13H2</td><td> V_l15</td><td> 151</td><td> DIQMTQSPSSLSASVGDRVTITCRASQGIRKDLGWYQQKPG KAPKRLIYGASSLQSGVPSRFSGSGSGTEFTLTISSLQPEDFA TYYCLQYNSFPWTFGQGTKVEIK</td>

<td> 32H7</td><td> V_l16</td><td> 152</td><td> EIVLTQSPGTLSLSPGERATLSCRASQSVSSGYLTWYQQKP GQAPRLLIYGASSRATGIPDRFSGSGSGTDFTLTISRLEPEDF AVYYCQQYGNSLCRFGQGTKLEIK</td>

<td> 32H7 CS</td><td> V_l17</td><td> 153</td><td> EIVLTQSPGTLSLSPGERATLSCRASQSVSSGYLTWYQQKP GQAPRLLIYGASSRATGIPDRFSGSGSGTDFTLTISRLEPEDF AVYYCQQYGNSLSRFGQGTKLEIK</td>

<td> 32H8</td><td> V_l18</td><td> 154</td><td> DIVMTQSPDSLAVSLGERATINCKSSQSILDSSNNDNYLAWY QQKPGQPPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSL QAEDVAVYYCQQYYNTPFTFGPGTKVDIK</td>

ΕΡ 2 379 594 Β1 (continued)

<td> Contained in Clone</td><td> Designation</td><td> SEQ ID NO.</td><td> Amino Acid Sequence</td>

<td> 33B5</td><td> V_l19</td><td> 155</td><td> DIQMTQSPSSLSASVGDRVTITCRASQGIRNDLGWYQQKPG KAPKRLIYVASSLQSGVPSRFSGSGSGTEFTLTISSLQPEDFA TYYCLQYNTYPLTFGGGTKVEIK</td>

<td> 33E4</td><td> V_l20</td><td> 156</td><td> EIVMTQSPATLSVSPGERATLSCRASQSVRSNLAWYQQKPG QAPRLLIHDASPRTAGIPARFSGSGSGTEFTLTINSLQSEDFA VYYCQQYNYWTPITFGQGTRLEIK</td>

<td> 34E3</td><td> V_l21</td><td> 157</td><td> QSVLTQPPSMSAAPGQKVTISCSGSSSNIGNNYVSWYQQLP GTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATLGITGLQTGDE ANYCCGTWDIGLSVWVFGGGTKLTVL</td>

<td> 4E4 9D4 1E11</td><td> V_H1</td><td> 158</td><td> QVQLVESGGGVVQPGRSLRLSCAASGFTFSSFGMHWVRQA PGKGLEWVAVISFDGSIKYSVDSVKGRFTISRDNSKNTLFLQ MNSLRAEDTAVYYCARDRLNYYDSSGYYHYKYYGMAVWGQ GTTVTVSS</td>

<td> 1H7</td><td> V_H2</td><td> 159</td><td> EVQLVESGGGLVKPGGSLRLSCAASGFTFSNAWMSWVRQA PGKGLEWVGRIKSKDGGKDYAAPVKGRFTISRDDSKNTLY LQMNSLKTEDTAVYYCTTDRTGYSISWSSYYYYYGMDVWG QGTTVTVSS</td>

<td> 2E7</td><td> V_h3</td><td> 160</td><td> EVQLLESGGGLVQPGESLRLSCAASGFTFSSYAMSWVRQA PGKGLEWVSAISGSGGRTYYADSVKGRFTISRDNSKNTLYL QMNSLRAEDTAVYYCAKDQREVGPYSSGWYDYYYGMDVW GQGTTVTVSS</td>

<td> 3B6</td><td> V_h4</td><td> 161</td><td> QVQLVQSGAEVKKPGASVKVSCKASGYTFTGYYMHWVRQA PGQGLEWMGWINPNSGGTNYAQKFQGRVTMTRDTSISTAY MELSRLRSDDTAVYFCARDQMSIIMLRGVFPPYYYGMDVWG QGTTVTVSS</td>

<td> 3C8 12E8 5F5</td><td> V_h5</td><td> 162</td><td> QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQA PGKGLEWVAVISYDGSHESYADSVKGRFTISRDISKNTLYLQ MNSLRAEDTAVYFCARERKRVTMSTLYYYFYYGMDVWGQG TTVTVSS</td>

ΕΡ 2 379 594 Β1 (continued)

<td> Contained in Clone</td><td> Designation</td><td> SEQ ID NO.</td><td> Amino Acid Sequence</td>

<td> 4H6</td><td> V_h6</td><td> 163</td><td> EVQLVESGGGLVKPGRSLRLSCTASGFTFGDYAMSWFRQA PGKGLEWIGFIRSRAYGGTPEYAASVKGRFTISRDDSKTIAYL QMNSLKTEDTAVYFCARGRGIAARWDYWGQGTLVTVSS</td>

<td> 9F5</td><td> V_h7</td><td> 164</td><td> EVQLVESGGGLVKPGGSLRLSCAASGFTFSNAWMSWVRQA PGKGLEWVGRIKSKTDGGTTDYTAPVKGRFTISRDDSKNTLY LQMNSLKAEDTAVYYCTTDRTGYSISWSSYYYYYGMDVWG QGTTVTVSS</td>

<td> 10E4</td><td> V_h8</td><td> 165</td><td> QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYYMYWVRQA PGQGLEWMGWISPNSGGTNYAQKFQGRVTMTRDTSISTAY MELSRLRSDDTAVYYCVRGGYSGYAGLYSHYYGMDVWGQ GTTVTVSS</td>

<td> 11D11</td><td> V_h9</td><td> 166</td><td> EVQLVESGGGLVKPGGSLRLSCAASGFTFGNAWMSWVRQA PGKGLEWVGRIKSKTDGGTTDYAAPVKGRFTISRDDSKNTLY LQMNSLKTEDTAVYFCTTDRTGYSISWSSYYYYYGMDVWG QGTTVTVSS</td>

<td> 11H9</td><td> V_H10</td><td> 167</td><td> EVQLVESGGGLVKPGGSLRLSCAASGFTFGNAWMSWVRQA PGKGLEWVGRIKSKTDGGTTDYAAPVKGRFTISRDDSKNTLY LQMNSLKTEDTAVYYCTTDRTGYSISWSSYYYYYGMDVWG QGTTVTVSS</td>

<td> 12G8</td><td> V_H11</td><td> 168</td><td> QVQLVESGGGVVQPGRSLRLSCAASGFTFSSFGMHWVRQA PGKGLEWVAVISFDGSIKYSVDSVKGRFTISRDNSKNTLFLQ MNSLRAEDTAVYYCARDRLNYYDSSGYYHYKYYGLAVWGQ GTTVTVSS</td>

<td> 13H2</td><td> V_h12</td><td> 169</td><td> EVQLVESGGGLVKPGGSLRLSCAASGYTFSTYSMNWVRQA PGKGLEWVSSISSSSSYRYYADSVKGRFTISRDNAKNSLYLQ MSSLRAEDTAVYYCAREGVSGSSPYSISWYDYYYGMDVWG QGTTVTVSS</td>

ΕΡ 2 379 594 Β1 (continued)

<td> Contained in Clone</td><td> Designation</td><td> SEQ ID NO.</td><td> Amino Acid Sequence</td>

<td> 32H7</td><td> V_h13</td><td> 170</td><td> QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQA PGKGLEWVAVIWYDGSNKYYADSVKGRFIISRDKSKNTLYLQ MNSLRAEDTAVYYCARAGGIAAAGLYYYYGMDVWGQGTTV TVSS</td>

<td> 32H8</td><td> V_h14</td><td> 171</td><td> QVQLVQSGAEVKKPGASVKVSCKASGYTFTAYYLHWVRQA PGQGLEWMGWINPHSGGTNYAQKFQGRVTMTRDTSISTAY MELSRLRSDDTAVFYCARGRQWLGFDYWGQGTLVTVSS</td>

<td> 33E4</td><td> V_h15</td><td> 172</td><td> QVQLQQWGAGLLKPSETLSLSCAVYGGSFGGYYWSWIRQP PGKGLEWIGEINHSGGTKYNPSLKSRVTISVDTSKNQFSLKL SSVTAADTAVYFCARGDVVGFFDYWGQGTLVTVSS</td>

<td> 33B5</td><td> V_h16</td><td> 173</td><td> QVQLVQSGAEVKKSGASVKVSCKASGYTFTGYYMHWVRQA PGQGLEWMGWINPNSGGTNYVQKFQGRVTMTRDTSISTAY MELSRLRSDDTAVYYCARNEYSSAWPLGYWGQGTLVTVSS</td>

<td> 34E3</td><td> V_h17</td><td> 174</td><td> QITLKESGPTLVKPTQTLTLTCTFSGFSLSTSGVGVAWIRQPP GKALEWLAUYWTDDKRYSPSLKSRLTITKDTSKNQVVLRMT NMDPLDTATYFCAHRPGGWFDPWGQGTLVTVSS</td>

[0156] Each ofthe heavy chain variable regions listed in Table 3 may be combined with any ofthe light chain variable regions shown in Table 3 to form an antigén binding protein. Examples of such combinations include V_H1 combined with any ofV_L1, V_L2, V_L3, V_L4, V_L5, V_L6, V_L7, V_L8, V_L9, V_L10, V_L11, V_L12, V_L13, V_L14, V_L15, V_L16, or V_L17; V_H2 combined with any of V_L1, V_L2, V_L3, V_L4, V_L5, V_L6, V_L7, V_L8, V_L9, V_L10, V_L11, V_L12, V_L13, V_L14, V_L15, V_L16, or V_L17; V_h3 combined with any ofV_L1, V_L2, V_L3, V_L4, V_L5, V_L6, V_L7, V_L8, V_L9, V_L10, V_L11, V_L12, V_L13, V_L14, V_L15, V_L16, or V_L17; and so on.

[0157] In somé instances, the antigén binding protein includes at least one heavy chain variable region and/or one light chain variable region from those listed in Table 3. In somé instances, the antigén binding protein includes at least two different heavy chain variable regions and/or light chain variable regions from those listed in Table 3. An example of such an antigén binding protein comprises (a) one V_H1, and (b) one of V_H2, V_H3, V_H4, V_H5, V_H6, V_H7, V_H8, V_H9, V_H10, V_H11, V_H12, or V_h13. Another example comprises (a) one V_H2, and (b) one of V_H1, V_H3, V_H4, V_H5, V_H6, V_H7, V_h8, V_h9, V_H10, V_H11, V_H12, or V_h13. Again another example comprises (a) one V_H3, and (b) one of V_H1, V_H2, V_H4, V_h5, V_h6, V_h7, V_h8, V|_|9, V_H10, V_H11, V_H12, or V_H13, etc. Again another example of such an antigén binding protein comprises (a) one V_L1, and (b) one ofV_L2, V_L3, V_L4, V_L5, V_L6, V_L7, V_L8, V_L9, V_L10, V_L11, V_L12, V_L13, V_L14, V_L15, V_L16, or V_l17, V_l18. V_l19, V_l20, or V_l21 . Again another example of such an antigén binding protein comprises (a) one V_L2, and (b) one ofV_L1, V_L3, V_L4, V_L5, V_L6, V_L7, V_L8, V_L9, V_L10, V_L11, V_L12, V_L13, V_L14, V_L15, V_L16, V_L17, V_L18, V_L19, V_l20. or V_L21. Again another example of such an antigén binding protein comprises (a) one V_L3, and (b) one of V_L1, V_L2, V_l4, V_l5, V_l6, V_l7, V_l8, V_l9, V_l1 0, V_L11, V_L12, V_L13, V_L14, V_L15, V_L16, V_L17, V_L18, V_L19, V_L20, or V_L21, etc. [0158] The various combinations of heavy chain variable regions may be combined with any ofthe various combinations of light chain variable regions as is apparent to one of skill in the art.

[0159] In other instances, the antigén binding protein contains two identical light chain variable regions and/or two identical heavy chain variable regions. As an example, the antigén binding protein may be an antibody or immunologically functional fragment that includes two light chain variable regions and two heavy chain variable regions in combinations

ΕΡ 2 379 594 Β1 of pairs of light chain variable régions and pairs of heavy chain variable régions as listed in Table 3.

[0160] Somé antigén binding proteins that are illustrated comprise a heavy chain variable domain comprising a sequence of amino acids that differs from the sequence of a heavy chain variable domain selected from V_H1, V_H2, V_H3, V_H4, V_h5, V_h6, V_h7, V_h8, V_h9, V_h10, V_h11, V_h12, and V_H13 at only 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 or 15 amino acid residues, wherein each such sequence difference is independently either a deletion, insertion or substitution of one amino acid, with the deletions, insertions and/orsubstitutions resulting in no more than 15 amino acid changes relatíve to the foregoing variable domain sequences. The heavy chain variable region in somé antigén binding proteins comprises a sequence of amino acids that has at least 70%, 75%, 80%, 85%, 90%, 95%, 97% or99% sequence identity to the amino acid sequences ofthe heavy chain variable region ofV_H1, VH₂, V_H3, V_H4, V_H5, V_H6, V_H7, V_H8, V_H9, V_H10, V_H11, V_h12, and V_H13.

[0161] Certain antigén binding proteins comprise a light chain variable domain comprising a sequence of amino acids that differs from the sequence of a light chain variable domain selected from V_L1, V_L2, V_L3, V_L4, V_L5, V_L6, V_L7, V_L8, V_l9, V_l10, V_l11, V_l12, V_l13, V_l14, V_l15, V_l16, orV_L17 atonly 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 or 15 amino acid residues, wherein each such sequence difference is independently either a deletion, insertion or substitution of one amino acid, with the deletions, insertions and/orsubstitutions resulting in no more than 15 amino acid changes relatíve to the foregoing variable domain sequences. The light chain variable region in somé antigén binding proteins comprises a sequence of amino acids that has at least 70%, 75%, 80%, 85%, 90%, 95%, 97% or 99% sequence identity to the amino acid sequences of the light chain variable region of V, 1, V, 2, V, 3, V, 4, V, 5, V, 6, V, 7, V, 8, V, 9, V, 10, V, 11, V_l12, V_l13, V_l14, V_l15, V_l16, orV_L17.

[0162] In additional instances, antigén binding proteins comprise the following pairings of light chain and heavy chain variable domains: VL1 with VH1, VL2 with VH2, VL3 with VH3, VL4 with VH4, VL5 with VH5, VL6 with VH1, VL7 with VH6, VL8 with VH5, VL9 with VH1, VL10 with VH7, VL11 with , H8, VL12 with VH9, VL12 with VH10, VL13 with VH5, VL14 with VH11, VL15 with VH12, VL16 with VH13, and VL17 with VH13. In somé instances, the antigén binding proteins in the above pairings may comprise amino acid sequences that have 70%, 75%, 80%, 85%, 90%, 95%, 97% or 99% sequence identity with the specified variable domains.

[0163] Still other antigén binding proteins, e.g., antibodies or immunologically functional fragments, include variant forms of a variant heavy chain and a variant light chain as just described.

CDRs [0164] The antigén binding proteins disclosed herein are polypeptides intő which one or more CDRs are grafted, inserted and/or joined. An antigén binding protein can have 1,2, 3, 4, 5 or 6 CDRs. An antigén binding protein thus can have, fór example, one heavy chain CDR1 (CDRH1), and/or one heavy chain CDR2 (CDRH2), and/or one heavy chain CDR3 (CDRH3), and/or one light chain CDR1 (CDRL1), and/or one light chain CDR2 (CDRL2), and/or one light chain CDR3 (CDRL3). Somé antigén binding proteins include both a CDRH3 and a CDRL3. Specific heavy and light chain CDRs are identified in Tables 4A and 4B, respectively.

[0165] Complementarity determining régions (CDRs) and framework régions (FR) of a given antibody may be identified using the system described by Kábát et al. in Sequences of Proteins of Immunological Interest, 5th Ed., US Dept. of Health and Humán Services, PHS, NIH, NIH Publication no. 91-3242, 1991. Certain antibodies that are disclosed herein comprise one or more amino acid sequences that are identical or have substantial sequence identity to the amino acid sequences of one or more ofthe CDRs presented in Table 4A (CDRHs) and Table 4B (CDRLs).

Table 4A: Exemplary Heavy Chain CDR Amino Acid Sequences

<td> Alt Num</td><td> SEQ ID NO:</td><td> Contained in Reference</td><td> Designation</td><td> Sequence</td>

<td> 42</td><td> 73</td><td> 1E11HCDR1 4E4HCDR1 9D4HCDR1 12G8HCDR1</td><td> CDRH 1-1</td><td> SFGMH</td>

<td> 43</td><td> 76</td><td> 1H7HCDR1 9F5HCDR1 11D11HCDR1 11H9HCDR1</td><td> CDRH 1-2</td><td> NAWMS</td>

<td> 44</td><td> 79</td><td> 2E7HCDR1</td><td> CDRH 1-3</td><td> SYAMS</td>

<td> 45</td><td> 82</td><td> 3B6HCDR1</td><td> CDRH 1-4</td><td> GYYMH</td>

ΕΡ 2 379 594 Β1 (continued)

<td> Alt Num</td><td> SEQ ID NO:</td><td> Contained in Reference</td><td> Designation</td><td> Sequence</td>

<td> 46</td><td> 85</td><td> 3C8HCDR1 5F5HCDR1 12E8HCDR1</td><td> CDRH 1-5</td><td> SYGMH</td>

<td> 47</td><td> 88</td><td> 4H6HCDR1</td><td> CDRH 1-6</td><td> DYAMS</td>

<td> 48</td><td> 92</td><td> 10E4HCDR1</td><td> CDRH 1-7</td><td> DYYMY</td>

<td> 49</td><td> 97</td><td> 13H2HCDR1</td><td> CDRH 1-8</td><td> TYSMN</td>

<td> 50</td><td> 100</td><td> 32H7HCDR1</td><td> CDRH 1-9</td><td> SYGMH</td>

<td> 51</td><td> 74</td><td> 1E11HCDR2 4E4HCDR2 9D4HCDR2 12G8HCDR2</td><td> CDRH 2-1</td><td> VISFDGSIKYSVDSVKG</td>

<td> 52</td><td> 77</td><td> 1H7HCDR2</td><td> CDRH 2-2</td><td> RIKSTTDGGTTDYAAPVKG</td>

<td> 53</td><td> 80</td><td> 2E7HCDR2</td><td> CDRH 2-3</td><td> AISGSGGRTYYADSVKG</td>

<td> 54</td><td> 83</td><td> 3B6HCDR2</td><td> CDRH 2-4</td><td> WINPNSGGTNYAQKF4G</td>

<td> 55</td><td> 86</td><td> 3C8HCDR2 5F5HCDR2 12E8HCDR2</td><td> CDRH 2-5</td><td> VISYDGSHESYADSVKG</td>

<td> 56</td><td> 89</td><td> 4H6HCDR2</td><td> CDRH 2-6</td><td> FIRSRAYGGTPEYAASVKG</td>

<td> 57</td><td> 91</td><td> 9F5HCDR2</td><td> CDRH 2-7</td><td> RIKSKTDGGTTDYTAPVKG</td>

<td> 58</td><td> 93</td><td> 10E4HCDR2</td><td> CDRH 2-8</td><td> WISPNSGGTNYAQKFQG</td>

<td> 59</td><td> 95</td><td> 11D11HCDR2 11H9HCDR2</td><td> CDRH 2-9</td><td> RIKSKTDGGTTDYAAPVKG</td>

<td> 60</td><td> 98</td><td> 13H2HCDR2</td><td> CDRH 2-10</td><td> SISSSSSYRYYADSVKG</td>

<td> 61</td><td> 101</td><td> 32H7HCDR2</td><td> CDRH 2-11</td><td> VIWYDGSNKYYADSVKG</td>

<td> 62</td><td> 75</td><td> 1E11HCDR3 4E4HCDR3 9D4HCDR3</td><td> CDRH 3-1</td><td> DRLNYYDSSGYYHYKYYGMAV</td>

<td> 63</td><td> 78</td><td> 1H7HCDR3 9F5HCDR3 11D11HCDR3 11H9HCDR3</td><td> CDRH 3-2</td><td> DRTGYSISWSSYYYYYGMDV</td>

<td> 64</td><td> 81</td><td> 2E7HCDR3</td><td> CDRH 3-3</td><td> DQREVG PYSSGWYDYYYG M DV</td>

<td> 65</td><td> 84</td><td> 3B6HCDR3</td><td> CDRH 3-4</td><td> DQMSIIMLRGVFPPYYYGMDV</td>

<td> 66</td><td> 87</td><td> 3C8HCDR3 5F5HCDR3 12E8HCDR3</td><td> CDRH 3-5</td><td> ERKRVTMSTLYYYFYYGMDV</td>

<td> 67</td><td> 90</td><td> 4H6HCDR3</td><td> CDRH 3-6</td><td> GRGIAARWDY</td>

<td> 68</td><td> 94</td><td> 10E4HCDR3</td><td> CDRH 3-7</td><td> GGYSGYAGLYSHYYGMDV</td>

<td> 69</td><td> 96</td><td> 12G8HCDR3</td><td> CDRH 3-8</td><td> DRLNYYDSSGYYHYKYYGLAV</td>

<td> 70</td><td> 99</td><td> 13H2HCDR3</td><td> CDRH 3-9</td><td> EGVSGSSPYSISWYDYYYGMDV</td>

<td> 71</td><td> 102</td><td> 32H7HCDR3</td><td> CDRH 3-10</td><td> AG GIAAAG L YYYYG M DV</td>

ΕΡ 2 379 594 Β1

Table 4Β: Exemplary Light Chain CDR Amino Acid Sequences

<td> Alt Num</td><td> SEQIDNO:</td><td> Contained in Reference</td><td> Designation</td><td> Sequence</td>

<td> 72</td><td> 42</td><td> 1E11LCD1 4E4LCD1 9D4LCD1 12G8LCD1</td><td> CDRL1-1</td><td> SGSSSNIGNNYVS</td>

<td> 73</td><td> 45</td><td> 1H7LCD1 9F5LCD1 11D11LC1 11H9LCD1</td><td> CDRL 1-2</td><td> SGSSSNIGSNYVY</td>

<td> 74</td><td> 48</td><td> 2E7LCD1</td><td> CDRL 1-3</td><td> RASQGIRNDLG</td>

<td> 75</td><td> 51</td><td> 3B6LCD1</td><td> CDRL 1-4</td><td> QGDSLRSFYAS</td>

<td> 76</td><td> 54</td><td> 3C8LCD1</td><td> CDRL 1-5</td><td> KSSQSLLHSAGKTYLY</td>

<td> 77</td><td> 57</td><td> 4H6LCD1</td><td> CDRL 1-6</td><td> RSSQSLLHSFGYNYLD</td>

<td> 78</td><td> 60</td><td> 5F5LCD1</td><td> CDRL 1-7</td><td> KSSQSLLHSDGKTYLY</td>

<td> 79</td><td> 62</td><td> 10E4LCD1</td><td> CDRL 1-8</td><td> SGSSSNIGSNTVN</td>

<td> 80</td><td> 65</td><td> 12E8LCD1</td><td> CDRL 1-9</td><td> KSSQSLLHSDGRNYLY</td>

<td> 81</td><td> 66</td><td> 13H2LCD1</td><td> CDRL1-10</td><td> RASQGIRKDLG</td>

<td> 82</td><td> 69</td><td> 32H7LCD1 32H7mLCD1</td><td> CDRL 1-11</td><td> RASQSVSSGYLT</td>

<td> 83</td><td> 43</td><td> 1E11LCD2 4E4LCD2 9D4LCD2 12G8LCD2</td><td> CDRL 2-1</td><td> DNNKRPS</td>

<td> 84</td><td> 46</td><td> 1H7LCD2</td><td> CDRL 2-2</td><td> RSNQRPS</td>

<td> 85</td><td> 49</td><td> 2E7LCD2</td><td> CDRL 2-3</td><td> AASSLQS</td>

<td> 86</td><td> 52</td><td> 3B6LCD2</td><td> CDRL 2-4</td><td> GKNNRPS</td>

<td> 87</td><td> 55</td><td> 3C8LCD2 5F5LCD2 12E8LCD2</td><td> CDRL 2-5</td><td> EVSNRFS</td>

<td> 88</td><td> 58</td><td> 4H6LCD2</td><td> CDRL 2-6</td><td> LGSNRAS</td>

<td> 89</td><td> 61</td><td> 9F5LCD2 11D11LC2 11H9LCD2</td><td> CDRL 2-7</td><td> RNNQRPS</td>

<td> 90</td><td> 63</td><td> 10E4LCD2</td><td> CDRL 2-8</td><td> TNNQRPS</td>

<td> 91</td><td> 67</td><td> 13H2LCD2</td><td> CDRL 2-9</td><td> GASSLQS</td>

<td> 92</td><td> 70</td><td> 32H7 LCD2 32H7m LCD2</td><td> CDRL 2-10</td><td> GASSRAT</td>

<td> 93</td><td> 44</td><td> 1E11LCD3 4E4LCD3 9D4LCD3 12G8LCD3</td><td> CDRL 3-1</td><td> GTWDSRLSAVV</td>

<td></td><td></td><td> 1H7LCD3 9F5LCD3</td><td></td><td></td>

EP 2 379 594 Β1 (continued)

<td> Alt Num</td><td> SEQIDNO:</td><td> Contained in Reference</td><td> Designation</td><td> Sequence</td>

<td> 94</td><td> 47</td><td> 11D11LC3 11H9LCD3</td><td> CDRL 3-2</td><td> AAWDDSLSGWV</td>

<td> 95</td><td> 50</td><td> 2E7LCD3</td><td> CDRL 3-3</td><td> LQYNIYPWT</td>

<td> 96</td><td> 53</td><td> 3B6LCD3</td><td> CDRL 3-4</td><td> NSRDSSVYHLV</td>

<td> 97</td><td> 56</td><td> 3C8LCD3 5F5LCD3 12E8LCD3</td><td> CDRL 3-5</td><td> MQSFPLPLT</td>

<td> 98</td><td> 59</td><td> 4H6LCD3</td><td> CDRL 3-6</td><td> MQALQTPFT</td>

<td> 99</td><td> 64</td><td> 10E4LCD3</td><td> CDRL 3-7</td><td> AARDESLNGVV</td>

<td> 100</td><td> 68</td><td> 13H2LCD3</td><td> CDRL 3-8</td><td> LQYNSFPWT</td>

<td> 101</td><td> 71</td><td> 32H7 LCD3</td><td> CDRL 3-9</td><td> QQYGNSLCR</td>

<td> 102</td><td> 72</td><td> 32H7m LCD3</td><td> CDRL 3-10</td><td> QQYGNSLSR</td>

[0166] The structure and properties of CDRs within a naturally occurring antibody has been described, supra. Briefly, in a traditional antibody, the CDRs are embedded within a framework in the heavy and light chain variable region where they constitute the regions responsible fór antigén binding and recognition. A variable region comprises at least three heavy or light chain CDRs, see, supra (Kábát et al., 1991, Sequences of Proteins of Immunological Interest, Public Health Service N.I.H., Bethesda, MD; see alsó Chothia and Lesk, 1987, J. Mól. Bioi. 196:901-917; Chothia et al., 1989, Natúré 342: 877-883), within a framework region (designated framework regions 1-4, FR1, FR2, FR3, and FR4, by Kábát et al., 1991, supra', see alsó Chothia and Lesk, 1987, supra). The CDRs provided herein, however, may nőt only be used to define the antigén binding domain of a traditional antibody structure, bút may be embedded in a variety of other polypeptide structures, as described herein.

[0167] The CDRs illustrated are (a) a CDRH selected from the group consisting of (i) a CDRH1 selected from the group consisting of SEQ ID NO:73, 76, 79, 82, 85, 88, 92, 97, and 100; (ii) a CDRH2 selected from the group consisting of SEQ ID NO:74, 77, 80, 83, 86, 89, 91,93, 95, 98, 101, and 129; (iii) a CDRH3 selected from the group consisting of SEQ ID NO:75, 78, 81,84, 87, 90, 96, 99, 102, and 123; and (iv) a CDRH of (i), (ii) and (iii) that contains one or more, e.g., one, two, three, four or more amino acid substitutions (e.g., conservative amino acid substitutions), deletions or insertions of no more than five, four, three, two, or one amino acids; (B) a CDRL selected from the group consisting of (i) a CDRL1 selected from the group consisting of SEQ ID NO:42, 45, 48, 51,54, 57, 62, 65, 66, and 69; (ii) a CDRL2 selected from the group consisting of SEQ ID NO:43, 46, 49, 52, 55, 58, 61,63, 67, and 70; (iii) a CDRL3 selected from the group consisting of SEQ ID NO:44, 47, 50, 53, 56, 59, 64, 68, 71, and 72; and (iv) a CDRL of (i), (ii) and (iii) that contains one or more, e.g., one, two, three, four or more amino acid substitutions (e.g., conservative amino acid substitutions), deletions or insertions of no more than five, four, three, two, orone amino acids amino acids.

[0168] An antigén binding protein may include 1,2, 3,4, 5, or 6 variant forms ofthe CDRs listed in Tables 4A and 4B, each having at least 80%, 85%, 90% or 95% sequence identity to a CDR sequence listed in Tables 4A and 4B. Somé antigén binding proteins include 1,2, 3, 4, 5, or 6 ofthe CDRs listed in Tables 4A and 4B, each differing by no more than 1,2, 3, 4 or 5 amino acids from the CDRs listed in these tables.

[0169] The CDRs disclosed herein include consensus sequences derived from groups of related monoclonal antibodies. As described herein, a consensus sequence refers to amino acid sequences having conserved amino acids common among a number of sequences and variable amino acids that vary within a given amino acid sequences. The CDR consensus sequences illustrated include CDRs corresponding to each of CDRH1, CDRH2, CDRH3, CDRL1, CDRL2 and CDRL3.

[0170] An antigén binding protein may includes the following associations of CDRL1, CDRL2 and CDRL3: SEQ ID NOs: 42, 43, and 44; SEQ ID NOs: 45, 46, and 47; SEQ ID NOs: 48, 49, and 50; SEQ ID NOs: 51,52, and 53; SEQ ID NOs: 54, 55, and 56; SEQ ID NOs: 57, 58, and 59; SEQ ID NOs: 60, 55, and 56; SEQ ID NOs: 45, 61, and 47; SEQ ID NOs: 62, 63, and 64; SEQ ID NOs: 65, 55, and 56; SEQ ID NOs: 66, 67, and 68; SEQ ID NOs: 69, 70, and 71; and SEQ ID NOs: 69, 70, and 72.

[0171] An antigén binding protein may include the following associations of CDRH1, CDRH2 and CDRH3: SEQ ID NOs: 73, 74, and 75; SEQ ID NOs: 76, 77, and 78; SEQ ID NOs: 79, 80, and 81; SEQ ID NOs: 82, 83, and 84; SEQ ID NOs: 85, 86, and 87; SEQ ID NOs: 88, 89, and 90; SEQ ID NOs: 76, 91, and 78; SEQ ID NOs: 92, 93, and 94; SEQ ID

ΕΡ 2 379 594 Β1

NOs: 76, 95, and 78; SEQ ID NOs: 73, 74, and 96; SEQ ID NOs: 97, 98, and 99; and SEQ ID NOs: 100, 101, and 102. [0172] An antigén binding protein may include the following associations of CDRL1, CDRL2 and CDRL3 with CDRH1, CDRH2 and CDRH3: SEQ ID NOs: 42, 43, and 44 with SEQ ID NOs: 73, 74, and 75; SEQ ID NOs: 45,46, and 47 with SEQ ID NOs: 76, 77, and 78; SEQ ID NOs: 48,49, and 50 with SEQ ID NOs: 79, 80, and 81; SEQ ID NOs: 51,52, and 53 with SEQ ID NOs: 82, 83, and 84; SEQ ID NOs: 54, 55, and 56 with SEQ ID NOs: 85, 86, and 87; SEQ ID NOs: 57, 58, and 59 with SEQ ID NOs: 88, 89, and 90; SEQ ID NOs: 60, 55, and 56 with SEQ ID NOs: 85, 86, and 87; SEQ ID NOs: 45, 61, and 47 with SEQ ID NOs: 76, 91, and 78; SEQ ID NOs: 62, 63, and 64 with SEQ ID NOs: 92,93, and 94; SEQ ID NOs: 45, 61, and 47 with SEQ ID NOs: 76, 95, and 78; SEQ ID NOs: 65, 55, and 56 with SEQ ID NOs: 85, 86, and 87; SEQ ID NOs: 42, 43, and 44 with SEQ ID NOs: 73, 74, and 96; SEQ ID NOs: 66, 67, and 68 with SEQ ID NOs: 97, 98, and 99; SEQ ID NOs: 69, 70, and 71 with SEQ ID NOs: 100, 101, and 102; and SEQ ID NOs: 69, 70, and 72 with SEQ ID NOs: 100, 101, and 102.

[0173] Consensus sequences were determined using standard phylogenic analyses ofthe CDRs corresponding to the V_H and V_Lof anti-CGRP R antibodies. The consensus sequences were determined by keeping the CDRs contiguous within the same sequence corresponding to a V_H or V_L.

[0174] As illustrated in Figs. 3A, 3B, 4, 5A, 5B, 5C, 5D and 5E, lineage analysis of a variety of the antigén binding proteins illustrated herein resulted in groups of related sequences, designated as light chain CDR groups K1, K2, K3, and K4 (Figs. 3A and 3B), light chain CDR groups L1, L2, L3, and L4 (Fig. 4), and heavy chain CDR groups HC1 (Fig. 5A), HC2 (Fig. 5B), HC3 (Fig. 5C), HC4 (Fig. 5C), HC5 (Fig. 5D) and HC6 (Fig. 5E). Somé of the above groups were used to generate additional consensus sequences, as illustrated in Figs. 3A, 3B, 4, and 5F, to yield light chain CDR groups K1,4 (Fig. 3A), K2,3 (Fig. 3B), L1,2,3 (Fig, 4), and LA11 (Fig. 4), and heavy chain CDR groups HCA and HCB (Fig.5F).

[0175] The consensus sequences ofthe various CDR region groups are provided below:

K1 Consensus [0176] CDR1 RASQGIRX-iDLG (SEQ ID NO:103), wherein X₁ is selected from the group consisting of N and K. [0177] CDR2 X-|ASSLQS (SEQ ID NO:104), wherein X₁ is selected from the group consisting of A and G.

[0178] CDR3 LQYNX₁X₂PWT (SEQ ID NO: 105), wherein X₁ is selected from the group consisting of I and S, and X₂is selected from the group consisting of Y and F.

K4 Consensus [0179] CDR3 QQYGNSLX.|R (SEQ ID NO:106), wherein X₁ is selected from the group consisting of S and C.

K1,4 Consensus [0180] CDR1 RASQX₁X₂X₃X₄GX₅LX₆ (SEQ ID NO:107), wherein X₁ is selected from the group consisting of S and G, X₂ is selected from the group consisting ofV and I, X₃ is selected from the group consisting of S and R, X₄ is selected from the group consisting of S, N and K, X₅ is selected from the group consisting of Y and D, and X₆ is selected from the group consisting of T and G.

[0181] CDR2 X₁ASSX₂X₃X₄ (SEQ ID NO:108), wherein X₁ is selected from the group consisting of G and A, X₂ is selected from the group consisting of R and L, X₃ is selected from the group consisting of A and Q, and X₄ is selected from the group consisting of T and S.

[0182] CDR3 X^ QYX₂X₃X₄X₃X₃X₇ (SEQ ID NO:109), wherein X₁ is selected from the group consisting of Q and L, X₂ is selected from the group consisting of G and N, X₃ is selected from the group consisting of N and T, X₄ is selected from the group consisting of S, Y and F, X₅ is selected from the group consisting of L and P, X₆ is selected from the group consisting of C, W and S, and X₇ is selected from the group consisting of R and T.

K3 Consensus [0183] CDR1 KSSQSLLHSX₁GX₂X₃YLY (SEQ ID NO:110), wherein X₁ is selected from the group consisting ofD and A, X₂ is selected from the group consisting of R and K, and X₃ is selected from the group consisting of N and T.

K2,3 Consensus [0184] CDR1 X₁SSQSLLHSX₂GX₃X₄YLX₅ (SEQ ID NO:111), wherein X₁ is selected from the group consisting of R and K, X₂ is selected from the group consisting of F, D and A, X₃ is selected from the group consisting of Y, R and K, X₄ is selected from the group consisting of N and T, and X₅ is selected from the group consisting of D and Y.

EP 2 379 594 Β1 [0185] CDR2 X₁X₂SNRX₃S (SEQ ID NO:112), wherein X₁ is selected from the group consisting of L and E, X₂ is selected from the group consisting of G and V, and X₃ is selected from the group consisting of A and F.

[0186] CDR3 MQX₁X₂X₃X₄PX₅T (SEQ ID NO:113), wherein X₁ is selected from the group consisting of A and S, X₂is selected from the group consisting of L and F, X₃ is selected from the group consisting of Q and P, X₄ is selected from the group consisting of T and L, and X₅ is selected from the group consisting of F and L.

Lm3 Consensus [0187] CDR2 RX-|NQRPS (SEQ ID NO:114), wherein X₁ is selected from the group consisting of N and S.

Lm1,2,3 Consensus [0188] CDR1 SGSSSNIGX₁NX₂VX₃ (SEQ ID NO:115), wherein X₁ is selected from the group consisting of N and S, X₂ is selected from the group consisting of Y and T, and X₃ is selected from the group consisting of S, N and Y.

[0189] CDR2 X₁X₂NX₃RPS (SEQ ID NO:116), wherein X₁ is selected from the group consisting of D, T and R, X₂ is selected from the group consisting of N and S, and X₃ is selected from the group consisting of K and Q.

[0190] CDR3 X₁X₂X₃DX₄X₅LX₆X₇VV (SEQ ID NO:117), wherein X₁ is selected from the group consisting of G and A, X₂ is selected from the group consisting of T and A, X₃ is selected from the group consisting of W and R, X₄ is selected from the group consisting of S and D, X₅ is selected from the group consisting of R and S, X₆ is selected from the group consisting of S and N, and X₇ is selected from the group consisting of A and G.

LA11 Consensus [0191] CDR1 X₁GX₂X₃SX₄X₅X₆X₇X₃X_gX₁₀X₁₁ (SEQ ID NO:118), wherein X₁ is selected from the group consisting of S and Q, X₂ is present or absent, and if present, is S, X₃ is selected from the group consisting of S and D, X₄ is present or absent, and if present, is N, X₅ is selected from the group consisting of I and L, X₆ is selected from the group consisting of G and R, X₇ is selected from the group consisting of N and S, X₃ is selected from the group consisting of N and F, X₉ is selected from the group consisting of Y and T, X₁₀ is selected from the group consisting of V and A, and X_n is selected from the group consisting of S, N and Y.

[0192] CDR2 X₁X₂NX₃RPS (SEQ ID NO:119), wherein X₁ is selected from the group consisting of D, G, T, and R, X₂is selected from the group consisting of Ν, K and S, and X₃ is selected from the group consisting of K, N and Q.

[0193] CDR3 X₁X₂X₃DX₄X₅X₆X₇X₃X₉V (SEQ ID NO:120), wherein X₁ is selected from the group consisting of G, N and A, X₂ is selected from the group consisting of T, S and A, X₃ is selected from the group consisting of W and R, X₄is selected from the group consisting of S and D, X₅ is selected from the group consisting of R and S, X₆ is selected from the group consisting of L and V, X₇ is selected from the group consisting of S, Y and N, X₃ is selected from the group consisting of A, H and G, and X₉ is selected from the group consisting of V and L.

HC1 Consensus [0194] CDR1 X₁YYMX₂ (SEQ ID NO:121), wherein X₁ is selected from the group consisting of G and D, X₂ is selected from the group consisting of H and Y.

[0195] CDR2 WDqPNSGGTNYAQKFQG (SEQ ID NO:122), wherein X! is selected from the group consisting of N and S.

[0196] CDR3 X₁X₂X₃SX₄X₅X₆X₇X₃GX₉X₁₀X₁₁X₁₂YYX₁₃GMDV (SEQ ID NO:123), wherein X! is selected from the group consisting of D and G, X₂ is selected from the group consisting of Q and G, X₃ is selected from the group consisting of M and Y, X₄ is selected from the group consisting of I and G, X₅ is selected from the group consisting of I and Y, X₆is selected from the group consisting of M and A, X₇ is present or absent, and if present, is L, X₃ is present or absent, and if present, is R, X₉ is selected from the group consisting of V and L, X₁₀ is selected from the group consisting of F and Y, X_n is selected from the group consisting of P and S, X₁₂ is selected from the group consisting of P and H, and X₁₃ is present or absent, and if present, is Y.

HC2 Consensus [0197] CDR2 RIKSX₁TDGGTTDYX₂APVKG (SEQ ID NO:124), wherein X! is selected from the group consisting of K and T, and X₂ is selected from the group consisting of T and A.

ΕΡ 2 379 594 Β1

HC3 Consensus [0198] CDR1 Χ₁ΥΧ₂ΜΧ₃ (SEQ ID ΝΟ:125), wherein Χ₁ is selected from the group consisting of T and S, X₂ is selected from the group consisting of S and A, and X₃ is selected from the group consisting of N and S.

[0199] CDR2 X₁ISX₂SX₃X₄X₅X₆YYADSVKG (SEQ ID NO:126), wherein X! is selected from the group consisting of S and A, X₂ is selected from the group consisting of S and G, X₃ is selected from the group consisting of S and G, X₄is selected from the group consisting of S and G, X₅ is selected from the group consisting of Y and R, and X₆ is selected from the group consisting of R and T.

[0200] CDR3 X₁X₂X₃X₄X₅X₆X₇PYSX₈X₉WYDYYYGMDV (SEQ ID NO:127), wherein X! is selected from the group consisting of E and D, X₂ is selected from the group consisting of G and Q, X₃ is selected from the group consisting of V and R, X₄ is selected from the group consisting of S and E, X₅ is selected from the group consisting of G and V, X₆is selected from the group consisting of S and G, X₇ is present or absent, and if present, is S, X₈ is selected from the group consisting of I and S, and X_g is selected from the group consisting of S and G.

HC4 Consensus [0201] CDR1 SX.|GMH (SEQ ID NO:128), wherein X₁ is selected from the group consisting of F and Y.

[0202] CDR2 VISX₁DGSX₂KYX₃X₄DSVKG (SEQ ID NO:129), wherein X! is selected from the group consisting of F and Y, X₂ is selected from the group consisting of I and H, X₃ is selected from the group consisting of S and Y, and X₄is selected from the group consisting of V and A.

[0203] CDR3 X₁RX₂X3X₄X₅X₆SX₇X₈YYX₉X₁₀X₁₁YYGX₁2X₁₃V (SEQ ID NO:130), wherein X! is selected from the group consisting of D and E, X₂ is selected from the group consisting of L and K, X₃ is selected from the group consisting of N and R, X₄ is selected from the group consisting of Y and V, X₅ is selected from the group consisting of Y and T, X₆is selected from the group consisting of D and M, X₇ is selected from the group consisting of S and T, X₈ is selected from the group consisting of G and L, X₉ is selected from the group consisting of H and Y, X₁₀ is present or absent, and if present, is Y, X_n is selected from the group consisting of K and F, X₁₂ is selected from the group consisting of M and L, and X₁₃ is selected from the group consisting of A and D.

HCA Consensus [0204] CDR1 X₁X₂X₃MX₄ (SEQ ID NO: 131), wherein X₁ is selected from the group consisting of N and S, X₂ is selected from the group consisting of A, Y and F, X₃ is selected from the group consisting of W, A and G, and X₄ is selected from the group consisting of S and H.

[0205] CDR2 X₁IX₂X₃X₄X₅X₆GX₇X₃X₉X₁₀X₁ ^X^₂X^₃X^₄VKG (SEQ ID NO:132), whereinX^ is selected from the group consisting of R, A and V, X₂ is selected from the group consisting of K, S and W, X₃ is selected from the group consisting of S, G, F and Y, X₄ is present or absent, and if present, is selected from the group consisting of K and T, X₅ is present or absent, and if present, isT, X₆ is selected from the group consisting of D and S, X₇ is selected from the group consisting of G and S, X₈ is selected from the group consisting of T, R, I, N and H, X₉ is selected from the group consisting of T and K, X₁₀ is selected from the group consisting of D and Y, X_n is selected from the group consisting of Y and S, X₁₂is selected from the group consisting of T, A and V, X₁₃ is selected from the group consisting of A and D, and X₁₄ is selected from the group consisting of P and S.

[0206] CDR3 X₁X₂X₃X₄X₅X₆X₇X₃X₉X₁₀X₁₁X₁₂X₁₃X₁₄X₁₅X₁₆X₁₇GX₁₃X₁₉V (SEQ ID NO:133), wherein X! is selected from the group consisting of D, A and E, X₂ is selected from the group consisting of R, Q and G, X₃ is selected from the group consisting of T, R, L, G and K, X₄ is selected from the group consisting of G, Ε, Ν, I and R, X₅ is selected from the group consisting of Υ, V and A, X₆ is selected from the group consisting of S, G, Y, A and T, X₇ is selected from the group consisting of I, P, D, A and M, X₈ is present or absent, and if present, is selected from the group consisting of S and Y, X₉ is present or absent, and if present, is selected from the group consisting of W, S and T, X₁₀ is selected from the group consisting of S, G and L, X_n is selected from the group consisting of S, G, L and Y, X₁₂ is present or absent, and if present, is selected from the group consisting of W and Y, X₁₃ is selected from the group consisting of Y and H, X₁₄ is present or absent, and if present, is selected from the group consisting of Y and D, X₁₅ is selected from the group consisting of Y, K and F, X₁₆ is present or absent, and if present, is Y, X₁₇ is present or absent, and if present, is Y, X₁₈is selected from the group consisting of M and L, and X₁₉ is selected from the group consisting of D and A.

HCB Consensus [0207] CDR1 X₁X₂X₃X₄X₅ (SEQ ID NO:134), wherein X₁ is selected from the group consisting of N, G, D, S and A, X₂ is selected from the group consisting of A, F and Y, X₃ is selected from the group consisting of W, Y, A and G, X₄ is selected from the group consisting of M and L, and X₅ is selected from the group consisting of S and H.

ΕΡ 2 379 594 Β1 [0208] CDR2X₁IX₂X₃X₄X₅X₆X₇X₈X_gX₁₀X₁₁X₁₂X₁3X₁₄X₁5^Xi6^Xi7^G (^{seq id} NO:135), wherein X! is selected from the group consisting of R, W, A, V, S and F, X₂ is selected from the group consisting of K, N, S, W and R, X₃ is selected from the group consisting of S, P, G, F and Y, X₄ is present or absent, and if present, is selected from the group consisting of K, T and R, X₅ is present or absent, and if present, is selected from the group consisting of T and A, X₆ is selected from the group consisting of D, Ν, H, S and Y, X₇ is selected from the group consisting of G and S, X₈ is selected from the group consisting of G and S, X_g is selected from the group consisting of T, G, R, I, Ν, H and Y, X₁₀ is selected from the group consisting of Τ, K, R and P, X_n is selected from the group consisting of D, Ν, Y and E, X₁₂ is selected from the group consisting of Y and S, X₁₃ is selected from the group consisting of T, A and V, X₁₄ is selected from the group consisting of A, Q and D, X₁₅ is selected from the group consisting of P, K and S, X₁₆ is selected from the group consisting of V and F, and X₁₇ is selected from the group consisting of K and Q.

[0209] CDR3 X₁X₂X₃X₄X₅SX₆X₇X₈X₉X₁₀X₁₁X₁₂X₁₃X₁₄X₁₅X₁₆GX₁₇X₁₈V (SEQ ID NO:136), wherein X! is selected from the group consisting of D, G, A and E, X₂ is selected from the group consisting of R, G and Q, X₃ is selected from the group consisting of Τ, Μ, Y, R, L, G and K, X₄ is selected from the group consisting of G, S, Ε, Ν, I and R, X₅ is selected from the group consisting of Υ, I, G, V and A, X₆ is selected from the group consisting of S, I, Y, G, A and T, X₇ is selected from the group consisting of I, Μ, A, P and D, X₈ is present or absent, and if present, is selected from the group consisting of S, L and Y, X_g is present or absent, and if present, is selected from the group consisting of W, R, S and T, X₁₀ is selected from the group consisting of S, G and L, X₁₁ is selected from the group consisting of S, V, L, G and Y, X₁₂ is present or absent, and if present, is selected from the group consisting of F, Y and W, X₁₃ is selected from the group consisting of Y, P, S and H, X₁₄ is present or absent, and if present, is selected from the group consisting of Y, P, D and H, X₁₅ is selected from the group consisting of Y, K and F, X₁₆ is present or absent, and if present, is Y, X₁₇is present or absent, and if present, is Y, and X₁₈ is selected from the group consisting of M and L.

[0210] In somé cases the antigén binding protein comprises at least one heavy chain CDR1, CDR2, or CDR3 having one of the above consensus sequences. In somé cases, the antigén binding protein comprises at least one light chain CDR1, CDR2, or CDR3 having one of the above consensus sequences. In other cases, the antigén binding protein comprises at least two heavy chain CDRs according to the above consensus sequences, and/or at least two light chain CDRs according to the above consensus sequences. In still other cases, the antigén binding protein comprises at least three heavy chain CDRs according to the above consensus sequences, and/or at least three light chain CDRs according to the above consensus sequences.

Exemplary Antigén Binding Proteins [0211] Illustrated is an isolated antigen-binding protein that binds CGRP R comprising (A) one or more heavy chain complementary determining regions (CDRHs) selected from the group consisting of: (i) a CDRH1 selected from the group consisting of SEQ ID NO:73, 76, 79, 82, 85, 88, 92, 97, and 100; (ii) a CDRH2 selected from the group consisting of SEQ ID NO:74, 77, 80, 83, 86, 89, 91,93, 95, 98, 101, and 129; (iii) a CDRH3 selected from the group consisting of SEQ ID NO:75, 78, 81,84, 87, 90, 96, 99, 102, and 123; and (ív) a CDRH of (i), (ii) and (iii) that contains one or more, e.g., one, two, three, fouror more amino acid substitutions, deletions or insertions of no more than five, four, three, four, two or one amino acids; (B) one or more light chain complementary determining regions (CDRLs) selected from the group consisting of: (i) a CDRL1 selected from the group consisting of SEQ ID NO:42, 45, 48, 51, 54, 57, 62, 65, 66, and 69; (ii) a CDRL2 selected from the group consisting of SEQ ID NO:43, 46, 49, 52, 55, 58, 61,63, 67, and 70; (iii) a CDRL3 selected from the group consisting of SEQ ID NO:44, 47, 50, 53, 56, 59, 64, 68, 71, and 72; and (ív) a CDRL of (i), (ii) and (iii) that contains one or more, e.g., one, two, three, four or more amino acid substitutions, deletions or insertions of no more than five, four, three, four, two or one amino acids; or (C) one or more heavy chain CDRHs of (A) and one or more light chain CDRLs of (B).

[0212] The isolated antigen-binding protein may comprise (A) a CDRH selected from the group consisting of (i) a CDRH1 selected from the group consisting of SEQ ID NO:73, 76, 79, 82, 85, 88, 92, 97, and 100; (ii) a CDRH2 selected from the group consisting of SEQ ID NO:74, 77, 80, 83, 86, 89, 91,93, 95, 98, 101, and 129; and (iii) a CDRH3 selected from the group consisting of SEQ ID NO:75, 78, 81, 84, 87, 90, 96, 99, 102, and 123; (B) a CDRL selected from the group consisting of (i) a CDRL1 selected from the group consisting of SEQ ID NO:42, 45, 48, 51,54, 57, 62, 65, 66, and 69; (ii) a CDRL2 selected from the group consisting of SEQ ID NO:43, 46, 49, 52, 55, 58, 61,63, 67, and 70; and (iii) a CDRL3 selected from the group consisting of SEQ ID NO:44, 47, 50, 53, 56, 59, 64, 68, 71, and 72; or (C) one or more heavy chain CDRHs of (A) and one or more light chain CDRLs of (B). In one embodiment, the isolated antigen-binding protein may include (A) a CDRH1 of SEQ ID NO:73, 76, 79, 82, 85, 88, 92, 97, and 100, a CDRH2 of SEQ ID NO:74, 77, 80, 83, 86, 89, 91, 93, 95, 98, 101, and 129, and aCDRH3ofSEQ ID NO:75, 78, 81,84, 87, 90, 96, 99, 102, and 123, and (B) a CDRL1 ofSEQ ID NO:42, 45, 48, 51,54, 57, 62, 65, 66, and 69, a CDRL2 ofSEQ ID NO:43, 46, 49, 52, 55, 58, 61,63, 67, and 70, and a CDRL3 of SEQ ID NO:44, 47, 50, 53, 56, 59, 64, 68, 71, and 72.

[0213] Alsó illustrated is an isolated antigén binding protein that specifically binds to an epitope formed of amino acid residues from both the CRLR and RAMP1 components ofthe CGRP R.

ΕΡ 2 379 594 Β1 [0214] The first amino acid sequence ofthe isolated antigen-binding protein may include the CDRH3 of SEQ ID NO:75, 78, 81, 84, 87, 90, 96, 99, 102, and 123, CDRH2ofSEQ ID NO:74, 77, 80, 83, 86, 89, 91, 93, 95, 98, 101, and 129, and CDRH1 of SEQ ID NO:73, 76, 79, 82, 85, 88, 92, 97, and 100, and/or the second amino acid sequence of the isolated antigén binding protein may comprise the CDRL3 of SEQ ID NO:44, 47, 50, 53, 56, 59, 64, 68, 71, and 72, CDRL2 of SEQ ID NO:43, 46, 49, 52, 55, 58, 61,63, 67, and 70, and CDRL1 of SEQ ID NO:42, 45, 48, 51,54, 57, 62, 65, 66, and 69.

[0215] The antigén binding protein may comprise at least two CDRH sequences of heavy chain sequences Η1, H2, H3, H4, H5, H6, H7, H8, H9, H10, H11, H12, or H13, as shown in Table 5A. Alsó, the antigén binding protein may comprise at least two CDRL sequences of light chain sequences L1, L2, L3, L4, L5, L6, L7, L8, L9, L10, L11, L12, L13, L14,L15,L16,orL17, as shown in Table 5B. Alsó, the antigén binding protein may comprise at least two CDRH sequences of heavy chain sequences H1, H2, H3, H4, H5, H6, H7, H8, H9, H10, H11, H12, orH13, as shown in Table 5A , and at least two CDRLsof light chain sequences L1, L2, L3, L4, L5, L6, L7, L8, L9, L10, L11, L12, L13, L14, L15, L16,orL17, as shown in Table 5B.

[0216] Aslo, the antigén binding protein may comprise the CDRH1, CDRH2, and CDRH3 sequences of heavy chain sequences H1,H2,H3, H4,H5,H6, H7,H8,H9,H10,H11,H12,orH13,as shown in Table 5A. Alsó, the antigén binding protein may comprise the CDRL1, CDRL2, and CDRL3 sequences of light chain sequences L1, L2, L3, L4, L5, L6, L7, L8, L9, L10, L11, L12, L13, L14, L15, L16, or L17, as shown in Table 5B.

[0217] Alsó, the antigén binding protein may comprise all six CDRs of L1 and H1, or L2 and H2, or L3 and H3, or L4 and H4, or L5 and H5, or L6 and H1, or L7 and H6, or L8 and H5, or L9 and H1, or L10 and H7, or L11 and H8, or L12 and H9, or L12 and H10, or L13 and H5, or L14 and H11, or L15 and H12, or L16 and H13, or L17 and H13, as shown in Tables 5A and 5B.

ΕΡ 2 379 594 Β1

Table 5Α - Exemplary Heavy Chain Amino Acid Sequence Regions

<td> Reference</td><td> Full Heavy Chain Group</td><td> Full Heavy Chain SEQ ID NO</td><td> Heavy Chain Variable Region Group</td><td> Heavy Chain Variable Region SEQ IDNO</td><td> CDRH1 SEQ ID NO</td><td> CDRH2 SEQ ID NO</td><td> CDRH3 SEQ ID NO</td>

<td> 1E11</td><td> H1</td><td> 29</td><td> V_H1</td><td> 158</td><td> 73</td><td> 74</td><td> 75</td>

<td> 1H7</td><td> H2</td><td> 30</td><td> V_h2</td><td> 159</td><td> 76</td><td> 77</td><td> 78</td>

<td> 2E7</td><td> H3</td><td> 31</td><td> Vh3</td><td> 160</td><td> 79</td><td> 80</td><td> 81</td>

<td> 3B6</td><td> H4</td><td> 32</td><td> Vh4</td><td> 161</td><td> 82</td><td> 83</td><td> 84</td>

<td> 3C8</td><td> H5</td><td> 33</td><td> V_h5</td><td> 162</td><td> 85</td><td> 86</td><td> 87</td>

<td> 4E4</td><td> H1</td><td> 29</td><td> Vh1</td><td> 158</td><td> 73</td><td> 74</td><td> 75</td>

<td> 4H6</td><td> H6</td><td> 34</td><td> V_h6</td><td> 163</td><td> 88</td><td> 89</td><td> 90</td>

<td> 5F5</td><td> H5</td><td> 33</td><td> Vh5</td><td> 162</td><td> 85</td><td> 86</td><td> 87</td>

<td> 9D4</td><td> H1</td><td> 29</td><td> Vh1</td><td> 158</td><td> 73</td><td> 74</td><td> 75</td>

<td> 9F5</td><td> H7</td><td> 35</td><td> V_h7</td><td> 164</td><td> 76</td><td> 91</td><td> 78</td>

<td> 10E4</td><td> H8</td><td> 36</td><td> Vh8</td><td> 165</td><td> 92</td><td> 93</td><td> 94</td>

ΕΡ 2 379 594 Β1

<td> Reference</td><td> Full Heavy Chain Group</td><td> Full Heavy Chain SEQ ID NO</td><td> Heavy Chain Variable Region Group</td><td> Heavy Chain Variable Region SEQIDNO</td><td> CDRH1 SEQ ID NO</td><td> CDRH2 SEQ ID NO</td><td> CDRH3 SEQ ID NO</td>

<td> 11D11</td><td> H9</td><td> 37</td><td> V_h9</td><td> 166</td><td> 76</td><td> 95</td><td> 78</td>

<td> 11Η9</td><td> H10</td><td> 38</td><td> V_h10</td><td> 167</td><td> 76</td><td> 95</td><td> 78</td>

<td> 12Ε8</td><td> H5</td><td> 33</td><td> Vh5</td><td> 162</td><td> 85</td><td> 86</td><td> 87</td>

<td> 12G8</td><td> H11</td><td> 39</td><td> V_h11</td><td> 168</td><td> 73</td><td> 74</td><td> 96</td>

<td> 13Η2</td><td> H12</td><td> 40</td><td> V_h12</td><td> 169</td><td> 97</td><td> 98</td><td> 99</td>

<td> 32Η7</td><td> H13</td><td> 41</td><td> V_h13</td><td> 170</td><td> 100</td><td> 101</td><td> 102</td>

<td> 32Η7 CS</td><td> H13</td><td> 41</td><td> V_h13</td><td> 170</td><td> 100</td><td> 101</td><td> 102</td>

<td> 32Η8</td><td></td><td></td><td> V_h14</td><td> 171</td><td></td><td></td><td></td>

<td> 33Β5</td><td></td><td></td><td> V_h15</td><td> 172</td><td></td><td></td><td></td>

<td> 33Ε4</td><td></td><td></td><td> V_h16</td><td> 173</td><td></td><td></td><td></td>

<td> 34Ε3</td><td></td><td></td><td> V_h17</td><td> 174</td><td></td><td></td><td></td>

ΕΡ 2 379 594 Β1

Table 5Β - Exemplary Light Chain Amino Acid Sequence Regions

<td> Reference</td><td> Full Light Chain Group</td><td> Full Light Chain SEQ ID NO</td><td> Light Chain Variabíe Region Group</td><td> Light Chain Variabíe Region SEQ ID NO</td><td> CDRL1 SEQ ID NO</td><td> CDRL2 SEQ ID NO</td><td> CDRL3 SEQ ID NO</td>

<td> 1E11</td><td> L1</td><td> 12</td><td> V_L1</td><td> 137</td><td> 42</td><td> 43</td><td> 44</td>

<td> 1H7</td><td> L2</td><td> 13</td><td> V_l2</td><td> 138</td><td> 45</td><td> 46</td><td> 47</td>

<td> 2E7</td><td> L3</td><td> 14</td><td> V_l3</td><td> 139</td><td> 48</td><td> 49</td><td> 50</td>

<td> 3B6</td><td> L4</td><td> 15</td><td> V_l4</td><td> 140</td><td> 51</td><td> 52</td><td> 53</td>

<td> 3C8</td><td> L5</td><td> 16</td><td> V_l5</td><td> 141</td><td> 54</td><td> 55</td><td> 56</td>

ΕΡ 2 379 594 Β1

<td> Reference</td><td> Full Light Chain Group</td><td> Full Light Chain SEQ ID NO</td><td> Lignt cnain Variable Region Group</td><td> Light Chain Variable Region SEQ ID NO</td><td> CDRL1 SEQ ID NO</td><td> CDRL2 SEQ ID NO</td><td> CDRL3SEQ ID NO</td>

<td> 4E4</td><td> L6</td><td> 17</td><td> V_l6</td><td> 142</td><td> 42</td><td> 43</td><td> 44</td>

<td> 4H6</td><td> L7</td><td> 18</td><td> V_l7</td><td> 143</td><td> 57</td><td> 58</td><td> 59</td>

<td> 5F5</td><td> L8</td><td> 19</td><td> V_l8</td><td> 144</td><td> 60</td><td> 55</td><td> 56</td>

<td> 9D4</td><td> L9</td><td> 20</td><td> V_l9</td><td> 145</td><td> 42</td><td> 43</td><td> 44</td>

<td> 9F5</td><td> L10</td><td> 21</td><td> V_l10</td><td> 146</td><td> 45</td><td> 61</td><td> 47</td>

<td> 10E4</td><td> L11</td><td> 22</td><td> V_L11</td><td> 147</td><td> 62</td><td> 63</td><td> 64</td>

<td> 11D11</td><td> L12</td><td> 23</td><td> V_l12</td><td> 148</td><td> 45</td><td> 61</td><td> 47</td>

<td> 11H9</td><td> L12</td><td> 23</td><td> V_l12</td><td> 148</td><td> 45</td><td> 61</td><td> 47</td>

<td> 12E8</td><td> L13</td><td> 24</td><td> V_l13</td><td> 149</td><td> 65</td><td> 55</td><td> 56</td>

<td> 12G8</td><td> L14</td><td> 25</td><td> V_l14</td><td> 150</td><td> 42</td><td> 43</td><td> 44</td>

<td> 13H2</td><td> L15</td><td> 26</td><td> V_l15</td><td> 151</td><td> 66</td><td> 67</td><td> 68</td>

<td> 32H7</td><td> L16</td><td> 27</td><td> V_l16</td><td> 152</td><td> 69</td><td> 70</td><td> 71</td>

<td> 32H7 CS</td><td> L17</td><td> 28</td><td> V_l17</td><td> 153</td><td> 69</td><td> 70</td><td> 72</td>

<td> 32H8</td><td></td><td></td><td> V_l18</td><td> 154</td><td></td><td></td><td></td>

<td> 33B5</td><td></td><td></td><td> V_l19</td><td> 155</td><td></td><td></td><td></td>

<td> 33E4</td><td></td><td></td><td> V_l20</td><td> 156</td><td></td><td></td><td></td>

<td> 34E3</td><td></td><td></td><td> V_l21</td><td> 157</td><td></td><td></td><td></td>

[0218] In one aspect, the isolated antigen-binding proteins provided herein can be a monoclonal antibody, a polyclonal antibody, a recombinant antibody, a humán antibody, a humanized antibody, a chimeric antibody, a multispecific antibody, or an antibody antigén binding fragment thereof.

[0219] In another embodiment, the antibody fragment ofthe isolated antigen-binding proteins provided herein can be a Fab fragment, a Fab’ fragment, an F(ab j₂ fragment, an Fv fragment, a diabody, or a single chain antibody molecule. [0220] In a further embodiment, the isolated antigén binding protein provided herein is a humán antibody and can be ofthe IgG 1-, lgG2- lgG3- or lgG4-type.

[0221] In another embodiment, the antigén binding protein consists of a just a light or a heavy chain polypeptide as set forth in Tables 5A-5B. In somé embodiments, the antigén binding protein consists just of a light chain variable or

ΕΡ 2 379 594 Β1 heavy chain variable domain such as those listed in Tables 5A-5B. Such antigén binding proteins can be pegylated with one or more PEG molecules.

[0222] In yet another aspect, the isolated antigen-binding protein provided herein can be coupled to a labeling group and can compete for binding to the extracellular portion of humán CGRP R with an antigén binding protein of one of the isolated antigen-binding proteins provided herein. In one embodiment, the isolated antigén binding protein provided herein can reduce monocyte chemotaxis, inhibit monocyte migration intő tumors or inhibit accumulation and function of tumor associated macrophage in a tumor when administered to a patient.

[0223] As will be appreciated by those in the art, for any antigén binding protein with more than one CDR from the depicted sequences, any combination of CDRs independently selected from the depicted sequences is useful. Thus, antigén binding proteins with one, two, three, four, fiveorsixof independently selected CDRs can be generated. However, as will be appreciated by those in the art, specific embodiments generally utilize combinations of CDRs that are nonrepetitive, e.g., antigén binding proteins are generally nőt made with two CDRH2 regions, etc.

[0224] Somé ofthe antigén binding proteins provided are discussed in more detail below.

Antigén Binding Proteins And Binding Epitopes and Binding Domains as claimed [0225] When an antigén binding protein is said to bind an epitope, such as one or both components of CGRP R, or the extracellular domain of CGRP R, for example, what is meant is that the antigén binding protein specifically binds to a specified portion of CGRP R, which may be on CRLR, RAMP 1, or span portions of both CRLR and RAMP 1. In cases where the antigén binding protein binds only CRLR (and nőt RAMP1), the antigén binding protein would nőt be expected to selectively bind CGRP R because CRLR is shared, interalia, with AM1 and AM1 receptors. Similarly, in cases where the antigén binding protein binds only RAMP1 (and nőt CRLR), the antigén binding protein would nőt be expected to selectively bind CGRP R because RAMP1 is shared, inter alia, with AMY1 receptor. In cases where the antigén binding protein interacts with both CRLR and RAMP1, the antigén binding protein is expected to bind residues or sequences of residues, or regions in both CRLR and RAMP1. In nőne ofthe foregoing embodiments is an antigén binding protein expected to contact every residue within CRLR or RAMP1. Similarly, nőt every amino acid substitution or deletion within CRLR, RAMP1 or the extracellular domains thereof is expected to significantly affect binding affinity.

[0226] Methods detailed, e.g., in Example 10, maybe used to assess what regions of multimeric receptors, such as CGRP R, may be involved in binding to selected antigén binding proteins.

Competing Antigén Binding Proteins as claimed [0227] In another aspect, antigén binding proteins are provided that compete with one ofthe exemplified, or reference antibodies or functional fragments binding to the epitope described above for specific binding to CGRP R. Such antigén binding proteins may alsó bind to the same epitope as one of the herein exemplified antigén binding proteins, or an overlapping epitope. Antigén binding proteins and fragments that compete with or bind to the same epitope as the exemplified or reference antigén binding proteins are expected to show similar functional properties. The exemplified antigén binding proteins and fragments include those with the heavy and light chains, variable region domains V_L1- V17 and V_H1 - V_H13, and CDRs included in Tables 2A, 2B, 3, 4A, 4B, 5A and 5B. Thus, as a specific example, the antigén binding proteins that are provided include those that compete with an antibody having: (a) all 6 ofthe CDRs listed for an antibody listed in Tables 5A and 5B; (b) a V_H and a V_L selected from V_L1- V_L17 and V_H1- V_H13 and listed for an antibody listed in Tables 5A and 5B; or (c) two light chains and two heavy chains as specified for an antibody listed in Tables 5A and 5B. Other examples of suitable reference antibodies include those that have a heavy chain variable region having a sequence eorresponding to any ofthe sequences identified as SEQ ID NO:158-170 and a light chain variable region having a sequence eorresponding to any ofthe sequences identified as SEQ ID NO:137-153.

[0228] Binding competition may be assessed, for example, using a binning assays, such as the Biacore assay described in Example 7, below. In that example, 19 antibodies described herein were tested against each of six reference antibodies — five neutralizing antibodies (11D11,3B6, 4H6, 12G8, and 9F5) and one non-neutralizing antibody (34E3). The assay results, shown in Table 13, indicate that all of the tested neutralizing antibodies (1E11,1 H7,2E7,3B6,3C8,4E4,4H6,5F5,9D4,9F5,10E4,11D11,11 H9,12E8,12G8,13H2 and 32H7) bind to essentially the same region of CGRP R, which is distinct from the region of CGRP R that is bound by the non-neutralizing antibodies tested (32H8, 33B5, 33E4 and 34E3). Based on these data, any ofthe neutralizing antibodies would make exemplary reference antigén binding proteins in a competition assay, particularly any of the neutralizing antibodies that were immobilized in the assay described in Example 7 — 11D11,3B6, 4H6, 12G8, and 9F5.

Monoclonal Antibodies [0229] The antigén binding proteins that are provided include monoclonal antibodies that bind to CGRP R. Monoclonal

ΕΡ 2 379 594 Β1 antibodies may be produced using any technique known in the art, e.g., by immortalizing spleen cells harvested from the transgenic animal after completion ofthe immunization schedule. The spleen cells can be immortalized using any technique known in the art, e.g., by fusing them with myeloma cells to produce hybridomas. Myeloma cells for use in hybridoma-producing fusion procedures preferably are non-antibody-producing, have high fusion efficiency, and enzyme deficiencies that renderthem incapable of growing in certain selective média which support the growth ofonly the desired fused cells (hybridomas). Examples of suitable cell lines for use in mouse fusions include Sp-20, P3-X63/Ag8, P3-X63Ag8.653, NS1/1.Ag 4 1, Sp210-Ag14, FO, NSO/U, MPC-11, MPC11-X45-GTG 1.7 and S194/5XXO Bul; examples of cell lines used in rat fusions include R210.RCY3, Y3-Ag 1.2.3, IR983F and 4B210. Other cell lines useful for cell fusions are U-266, GM1500-GRG2, LICR-LON-HMy2 and UC729-6. An exemplary method of preparing monoclonal antibodies is described in Example 2, below.

[0230] In somé instances, a hybridoma cell line is produced by immunizing an animal (e.g., a transgenic animal having humán immunoglobulin sequences) with a CGRP R immunogen; harvesting spleen cells from the immunized animal; fusing the harvested spleen cells to a myeloma cell line, thereby generating hybridoma cells; establishing hybridoma cell lines from the hybridoma cells, and identifying a hybridoma cell line that produces an antibody that binds CGRP R (e.g., as described in Examples 1-3, below). Such hybridoma cell lines, and anti-CGRP R monoclonal antibodies produced by them, are aspects ofthe present application.

[0231] Monoclonal antibodies secreted by a hybridoma cell line can be purified using any technique known in the art. Hybridomas or mAbs may be further screened to identify mAbs with particular properties, such as the ability to bind cells expressing CGRP, ability to block or interfere the binding of the CGRP ligand or CGRP₈_₃₇ peptide, or the ability to functionally block the receptor, e.g., using a cAMP assay, e.g., as described below.

Chimeric and Humanized Antibodies [0232] Chimeric and humanized antibodies based upon the foregoing sequences are alsó provided. Monoclonal antibodies for use as therapeutic agents may be modified in various ways prior to use. One example is a chimeric antibody, which is an antibody composed of protein segments from different antibodies that are covalently joined to produce functional immunoglobulin light or heavy chains or immunologically functional portions thereof. Generally, a portion of the heavy chain and/or light chain is identical with or homologous to a corresponding sequence in antibodies derived from a particular species or belonging to a particular antibody class or subclass, while the remainder of the chain(s) is/are identical with or homologous to a corresponding sequence in antibodies derived from another species or belonging to another antibody class or subclass. For methods relating to chimeric antibodies, see, for example, United States Patent No. 4,816,567; and Morrison et al., 1985, Proc. Natl. Acad. Sci. USA 81:6851-6855. CDR grafting is described, for example, in United States Patent No. 6,180,370, No. 5,693,762, No. 5,693,761, No. 5,585,089, and No. 5,530,101. [0233] Generally, the goal of making a chimeric antibody is to create a chimera in which the number of amino acids from the intended patient species is maximized. One example is the CDR-grafted antibody, in which the antibody comprises one or more complementarity determining regions (CDRs) from a particular species or belonging to a particular antibody class or subclass, while the remainder of the antibody chain(s) is/are identical with or homologous to a corresponding sequence in antibodies derived from another species or belonging to another antibody class or subclass. For use in humans, the variable region or selected CDRs from a rodent antibody often are grafted intő a humán antibody, replacing the naturally-occurring variable regions or CDRs ofthe humán antibody.

[0234] One useful type of chimeric antibody is a humanized antibody. Generally, a humanized antibody is produced from a monoclonal antibody raised initially in a non-human animal. Certain amino acid residues in this monoclonal antibody, typically from non-antigen recognizing portions ofthe antibody, are modified to be homologous to corresponding residues in a humán antibody of corresponding isotype. Humanization can be performed, for example, using various methods by substituting at least a portion of a rodent variable region for the corresponding regions of a humán antibody (see, e.g., United States Patent No. 5,585,089, and No. 5,693,762; Jones et al., 1986, Natúré 321.522-525', Riechmann et al., 1988, Natúré 332:323-27; Verhoeyen et al., 1988, Science 239:1534-1536), [0235] In one aspect, the CDRs ofthe light and heavy chain variable regions ofthe antibodies provided herein (see, Table 4) are grafted to framework regions (FRs) from antibodies from the same, or a different, phylogenetic species. For example, the CDRs of the heavy and light chain variable regions V_H1, V_H2, V_H3, V_H4, V_H5, V_H6, V_H7, V_H8, V_H9, V_H10, V_H11, V_H12, and V_H13, and/or V_L1, V_L2, V_L3, V_L4, V_L5, V_L6, V_L7, V_L8, V_L9, V_L10, V_L11, V_L12, V_L13, V_L14, V_L15, V_L16, and V_L17 can be grafted to consensus humán FRs. To create consensus humán FRs, FRs from several humán heavy chain or light chain amino acid sequences may be aligned to identify a consensus amino acid sequence. In other embodiments, the FRs of a heavy chain or light chain disclosed herein are replaced with the FRs from a different heavy chain or light chain. In one aspect, rare amino acids in the FRs ofthe heavy and light chains of anti-CGRP R antibody are nőt replaced, while the rest ofthe FR amino acids are replaced. A rare amino acid is a specific amino acid that is in a position in which this particular amino acid is nőt usually found in an FR. Alternatively, the grafted variable regions from the one heavy or light chain may be used with a constant region that is different from the constant region of that

ΕΡ 2 379 594 Β1 particular heavy or light chain as disclosed herein. In otherembodiments, the grafted variable regions are part ofa single chain Fv antibody.

[0236] In certain embodiments, constant regions from species other than humán can be used along with the humán variable region(s) to produce hybrid antibodies.

Fully Humán Antibodies [0237] Fully humán antibodies are alsó provided. Methods are available for making fully humán antibodies specific for a given antigén without exposing humán beings tothe antigén (fully humán antibodies). One specific means provided for implementing the production of fully humán antibodies is the humanization ofthe mouse humorai immuné system. Introduction of humán immunoglobulin (lg) loci intő mice in which the endogenous lg genes have been inactivated is one means of producing fully humán monoclonal antibodies (mAbs) in mouse, an animal that can be immunized with any desirable antigén. Using fully humán antibodies can minimize the immunogenic and allergic responses that can sometimes be caused by administering mouse or mouse-derived mAbs to humans as therapeutic agents.

[0238] Fully humán antibodies can be produced by immunizing transgenic animals (usually mice) that are capable of producing a Tepertőire of humán antibodies in the absence of endogenous immunoglobulin production. Antigens for this purpose typically have six or more contiguous amino acids, and optionally are conjugated to a carrier, such as a hapten. See, e.g., Jakobovits et al., 1993, Proc. Natl. Acad. Sci. USA 90:2551-2555; Jakobovits et al., 1993, Natúré 362:255-258; and Bruggermann etal., 1993, Year in Immunoi. 7:33. In one exampleof such a method, transgenic animals are produced by incapacitating the endogenous mouse immunoglobulin loci encoding the mouse heavy and light immunoglobulin chains therein, and inserting intő the mouse genome large fragments of humán genome DNA containing loci that encode humán heavy and light chain proteins. Partially modified animals, which have less than the full complement of humán immunoglobulin loci, are then cross-bred to obtain an animal having all ofthe desired immuné system modifications. When administered an immunogen, these transgenic animals produce antibodies that are immunospecific for the immunogen bút have humán rather than murine amino acid sequences, including the variable regions. For further details of such methods, see, for example, WO96/33735 and W094/02602. Additional methods relating to transgenic mice for making humán antibodies are described in United States Patent No. 5,545,807; No. 6,713,610; No. 6,673,986; No. 6,162,963; No. 5,545,807; No. 6,300,129; No. 6,255,458; No. 5,877,397; No. 5,874,299 and No. 5,545,806; in PCT publications WO91/10741, W090/04036, and in EP 546073B1 and EP 546073A1.

[0239] The transgenic mice described above, referred to herein as HuMab mice, contain a humán immunoglobulin gene minilocus that encodes unrearranged humán heavy ([mu] and [gamma]) and [kappa] light chain immunoglobulin sequences, together with targeted mutations that inactivate the endogenous [mu] and [kappa] chain loci (Lonberg etal.,

1994, Natúré 368:856-859). Accordingly, the mice exhibit reduced expression of mouse IgM or [kappa] and in response to immunization, and the introduced humán heavy and light chain transgenes undergo eláss switching and somatic mutation to generate high affinity humán IgG [kappa] monoclonal antibodies (Lonberg etal., supra.', Lonberg and Huszar,

1995, Intern. Rév. Immunoi. 13: 65-93; Harding and Lonberg, 1995, Ann. N.YAcad. Sci. 764:536-546). The preparation of HuMab mice is described in detail in Taylor et al., 1992, Nucleic Acids Research 20:6287-6295; Chen et al., 1993, International Immunology 5:647-656; Tuaillon et al., 1994, J. Immunoi. 152:2912-2920; Lonberg et al., 1994, Natúré 368:856-859; Lonberg, 1994, Handbook of Exp. Pharmacology 113:49-101; Taylor etal., 1994, International Immunology 6:579-591; Lonberg and Huszar, 1995, Intern. Rév. Immunoi. 13:65-93; Harding and Lonberg, 1995, Ann. N.Y Acad. Sci. 764:536-546; Fishwild et al., 1996, Natúré Biotechnology 14:845-851. See, further United States Patent No. 5,545,806; No. 5,569,825; No. 5,625,126; No. 5,633,425; No. 5,789,650; No. 5,877,397; No. 5,661,016; No. 5,814,318; No. 5,874,299; and No. 5,770,429; as well as United States Patent No. 5,545,807; International Publication Nos. WO 93/1227; WO 92/22646; and WO 92/03918. Technologies utilized for producing humán antibodies in these transgenic mice are disclosed alsó in WO 98/24893, and Mendez et al., 1997, Natúré Genetics 15:146-156. Förexample, the HCo7 and HCo12 transgenic mice strains can be used to generate anti-CGRP R antibodies. Further details regarding the production of humán antibodies using transgenic mice are provided in the examples below.

[0240] Using hybridoma technology, antigen-specific humán mAbs with the desired specificity can be produced and selected from the transgenic mice such as those described above. Such antibodies may be cloned and expressed using a suitable vector and hőst cell, or the antibodies can be harvested from cultured hybridoma cells.

[0241] Fully humán antibodies can alsó be derived from phage-display libraries (as disclosed in Hoogenboom et al., 1991, J. Mól. Bioi. 227:381; and Marks et al., 1991, J. Mól. Bioi. 222:581). Phage display techniques mimic immuné selection through the display of antibody repertoires on the surface of filamentous bacteriophage, and subsequent selection of phage by their binding to an antigén of choice. One such technique is described in PCT Publication No. WO 99/10494, which deseribes the isolation of high affinity and functional agonistic antibodies for MPL- and msk-receptors using such an approach.

EP 2 379 594 Β1

Bispecific Or Bifunctional Antigén Binding Proteins [0242] The antigén binding proteins that are provided alsó include bispecific and bifunctional antibodies that include one or more CDRs or one or more variable regions as described above. A bispecific or bifunctional antibody in somé instances is an artificial hybrid antibody having two different heavy/light chain pairs and two different binding sites. Bispecific antibodies may be produced by a variety of methods including, bút nőt limited to, fusion of hybridomas or linking of Fab’ fragments. See, e.g., Songsivilai and Lachmann, 1990, Clin. Exp. Immunoi. 79:315-321; Kostelny et al., 1992, J. Immunoi. 148:1547-1553.

Various Other Forms [0243] Somé ofthe antigén binding proteins that are provided are variantforms ofthe antigén binding proteins disclosed above (e.g., those having the sequences listed in Tables 2-5). For instance, somé ofthe antigén binding proteins have one or more conservative amino acid substitutions in one or more ofthe heavy or light chains, variable regions or CDRs listed in Tables 2-5.

[0244] Naturally-occurring amino acids may be divided intő classes based on common side chain properties:

1) hydrophobic: norleucine, Met, Ala, Val, Leu, Ile;

2) neutral hydrophilie: Cys, Ser, Thr, Asn, Gin;

3) acidic: Asp, Glu;

4) basic: His, Lys, Arg;

5) residues that influence chain orientation: Gly, Pro; and

6) aromatic: Trp, Tyr, Phe.

[0245] Conservative amino acid substitutions may involve exchange ofa member ofone of these classes with another member of the same eláss. Conservative amino acid substitutions may encompass non-naturally occurring amino acid residues, which are typically incorporated by Chemical peptide synthesis rather than by synthesis in biological systems. These include peptidomimetics and other reversed or inverted forms of amino acid moieties.

[0246] Non-conservative substitutions may involve the exchange of a member of one of the above classes for a memberfrom another eláss. Such substituted residues may be introduced intő regions ofthe antibody that are homologous with humán antibodies, or intő the non-homologous regions ofthe molecule.

[0247] In making such changes, according to certain embodiments, the hydropathic index of amino acids may be considered. The hydropathic profile ofa protein is calculated by assigning each amino acid a numerical value (hydropathy index) and then repetitively averaging these values along the peptide chain. Each amino acid has been assigned a hydropathic index on the basis of its hydrophobicity and charge characteristics. They are: isoleucine (+4.5); valine (+4.2); leucine (+3.8); phenylalanine (+2.8); cysteine/cystine (+2.5); methionine (+1.9); alanine (+1.8); glycine (-0.4); threonine (-0.7); serine (-0.8); tryptophan (-0.9); tyrosine (-1.3); proline (-1.6); histidine (-3.2); glutamate (-3.5); glutamine (-3.5); aspartate (-3.5); asparagine (-3.5); lysine (-3.9); and arginine (-4.5).

[0248] The importance ofthe hydropathic profile in conferring Interactive biological function on a protein is understood in the art (see, e.g., Kyte et al., 1982, J. Mól. Bioi. 157:105-131). It is known that certain amino acids may be substituted for other amino acids having a similar hydropathic index or score and still retain a similar biological activity. In making changes based upon the hydropathic index, in certain embodiments, the substitution of amino acids whose hydropathic indices are within ±2 is included. In somé aspects, those which are within ± 1 are included, and in other aspects, those within ±0.5 are included.

[0249] It is alsó understood in the art that the substitution of like amino acids can be made effectively on the basis of hydrophilicity, particularly where the biologically functional protein or peptide thereby created is intended for use in immunological embodiments, as in the present case. In certain embodiments, the greatest local average hydrophilicity ofa protein, as governed bythe hydrophilicity of its adjacent amino acids, correlates with its immunogenicity and antigenbinding or immunogenicity, that is, with a biological property of the protein.

[0250] The following hydrophilicity values have been assigned to these amino acid residues: arginine (+3.0); lysine (+3.0); aspartate (+3.0±1); glutamate (+3.0± 1); serine (+0.3); asparagine (+0.2); glutamine (+0.2); glycine (0); threonine (-0.4); proline (-0.5±1); alanine (-0.5); histidine (-0.5); cysteine (-1.0); methionine (-1.3); valine (-1.5); leucine (-1.8); isoleucine (-1.8); tyrosine (-2.3); phenylalanine (-2.5) and tryptophan (-3.4). In making changes based upon similar hydrophilicity values, in certain embodiments, the substitution of amino acids whose hydrophilicity values are within ±2 is included, in other embodiments, those which are within ±1 are included, and in still other embodiments, those within ±0.5 are included. In somé instances, one may alsó identify epitopes from primary amino acid sequences on the basis of hydrophilicity. These regions are alsó referred to as epitopic core regions.

[0251] Exemplary conservative amino acid substitutions are set forth in Table 6.

ΕΡ 2 379 594 Β1

Table 6: Conservative Amino Acid Substitutions

<td> Original Residue</td><td> Exemplary Substitutions</td>

<td> Alá</td><td> Ser</td>

<td> Arg</td><td> Lys</td>

<td> Asn</td><td> Gin, His</td>

<td> Asp</td><td> Glu</td>

<td> Cys</td><td> Ser</td>

<td> Gin</td><td> Asn</td>

<td> Glu</td><td> Asp</td>

<td> Gly</td><td> Pro</td>

<td> His</td><td> Asn, Gin</td>

<td> Ile</td><td> Leu, Val</td>

<td> Leu</td><td> Ile, Val</td>

<td> Lys</td><td> Arg, Gin, Glu</td>

<td> Met</td><td> Leu, Ile</td>

<td> Phe</td><td> Met, Leu, Tyr</td>

<td> Ser</td><td> Thr</td>

<td> Thr</td><td> Ser</td>

<td> Trp</td><td> Tyr</td>

<td> Tyr</td><td> Trp, Phe</td>

<td> Val</td><td> Ile, Leu</td>

[0252] A skilled artisan will be able to determine suitable variants of polypeptides as set forth herein using well-known techniques. One skilled in the art may identify suitable areas ofthe molecule that may be changed without destroying activity by targeting regions nőt believed to be important fór activity. The skilled artisan alsó will be able to identify residues and portions ofthe molecules that are conserved among similar polypeptides. In further embodiments, even areas that may be important fór biological activity orfor structure may be subject to conservative amino acid substitutions without destroying the biological activity or without adversely affecting the polypeptide structure.

[0253] Additionally, one skilled in the art can review structure-function studies identifying residues in similar polypeptides that are important fór activity or structure. In view of such a comparison, one can predict the importance of amino acid residues in a protein that correspond to amino acid residues important fór activity or structure in similar proteins. One skilled in the art may opt fór chemically similar amino acid substitutions fór such predicted important amino acid residues.

[0254] One skilled in the art can alsó analyze the 3-dimensional structure and amino acid sequence in relation to that structure in similar polypeptides. In view of such information, one skilled in the art may predict the alignment of amino acid residues of an antibody with respect to its three dimensional structure. One skilled in the art may choose nőt to make radical changes to amino acid residues predicted to be on the surface ofthe protein, since such residues may be involved in important interactions with other molecules. Moreover, one skilled in the art may generate test variants containing a single amino acid substitution at each desired amino acid residue. These variants can then be sereened using assays fór CGRP R neutralizing activity, (see examples below) thus yielding information regarding which amino acids can be changed and which must nőt be changed. In other words, based on information gathered from such routine experiments, one skilled in the art can readily determine the amino acid positions where further substitutions should be avoided either alone or in combination with other mutations.

[0255] A number of scientific publications have been devoted to the prediction of secondary structure. See, Moult, 1996, Curr. Op. in Biotech. 7:422-427; Chou et al., 1974, Biochem. 13:222-245; Chou et al., 1974, Biochemistry 113:211-222; Chou et al., 1978, Adv. Enzymol. Relat. Areas Mól. Bioi. 47:45-148; Chou et al., 1979, Ann. Rév. Biochem. 47:251-276; and Chou et al., 1979, Biophys. J. 26:367-384. Moreover, computer programs are currently available to

ΕΡ 2 379 594 Β1 assist with predicting secondary strueture. One method of predicting secondary strueture is based upon homology modeling. Fór example, two polypeptides or proteins that have a sequence identity of greater than 30%, or similarity greater than 40% can have similar structural topologies. The recent growth ofthe protein structural database (PDB) has provided enhanced predictability of secondary strueture, including the potential number of folds within a polypeptide’s or protein’s strueture. See, Hóim et al., 1999, Nucl. Acid. Rés. 27:244-247. It has been suggested (Brenner et al., 1997, Curr. Op. Struct. Bioi. 7:369-376) that there are a limited number of folds in a given polypeptide or protein and that once a critical number of structures have been resolved, structural prediction will become dramatically more accurate. [0256] Additional methods of predicting secondary strueture include threading (Jones, 1997, Curr. Opin. Struct. Bioi. 7:377-387; Sippl et al., 1996, Strueture 4:15-19), profile analysis (Bowie et al., 1991, Science 253:164-170; Gribskov et al., 1990, Meth. Enzym. 183:146-159; Gribskov et al., 1987, Proc. Nat. Acad. Sci. 84:4355-4358), and evolutionary linkage (See, Hóim, 1999, supra', and Brenner, 1997, supra).

[0257] In somé embodiments, amino acid substitutions are made that: (1) reduce susceptibility to proteolysis, (2) reduce susceptibilityto oxidation, (3) altér binding affinity fór forming protein complexes, (4) altér ligand or antigén binding affinities, and/or (4) confer or modify other physicochemical or functional properties on such polypeptides. Fór example, single or multiple amino acid substitutions (in certain embodiments, conservative amino acid substitutions) may be made in the naturally-occurring sequence. Substitutions can be made in that portion of the antibody that lies outside the domain(s) forming intermolecular contacts). In such embodiments, conservative amino acid substitutions can be used that do nőt substantially change the structural characteristics of the parent sequence (e.g., one or more replacement amino acids that do nőt disrupt the secondary strueture that characterizes the parent or native antigén binding protein). Examples of art-recognized polypeptide secondary and tertiary structures are described in Proteins, Structures and Molecular Principles (Creighton, Ed.), 1984, W. H. New York: Freeman and Company; Introduction to Protein Strueture (Branden and Tooze, eds.), 1991, New York: Garland Publishing; and Thornton et al., 1991, Natúré 354:105.

[0258] Additional preferred antibody variants include cysteine variants wherein one or more cysteine residues in the parent or native amino acid sequence are deleted from or substituted with another amino acid (e.g., serine). Cysteine variants are useful, inter alia when antibodies must be refolded intő a biologically active conformation. Cysteine variants may have fewer cysteine residues than the native antibody, and typically have an even number to minimize interactions resulting from unpaired cysteines.

[0259] The heavy and light chains, variable régions domains and CDRs that are disclosed can be used to prepare polypeptides that contain an antigén binding region that can specifically bind to CGRP R. Fór example, one or more of the CDRs listed in Tables 4 and 5 can be incorporated intő a molecule (e.g., a polypeptide) covalently or noncovalently to make an immunoadhesion. An immunoadhesion may incorporate the CDR(s) as part of a larger polypeptide chain, may covalently link the CDR(s) to another polypeptide chain, or may incorporate the CDR(s) noncovalently. The CDR(s) enable the immunoadhesion to bind specifically to a particular antigén of interest (e.g., CGRP R or epitope thereof). [0260] Mimetics (e.g., peptide mimetics or peptidomimetics) based upon the variable region domains and CDRs that are described herein are alsó provided. These analogs can be peptides, non-peptides or combinations of peptide and non-peptide régions. Fauchere, 1986, Adv. Drug Rés. 15:29; Veber and Freidinger, 1985, TINS p. 392; and Evans et al., 1987, J. Med. Chem. 30:1229. Peptide mimetics that are structurally similar to therapeutically useful peptides may be used to produce a similar therapeutic or prophylactic effect. Such compounds are often developed with the aid of computerized molecular modeling. Generally, peptidomimetics are proteins that are structurally similar to an antibody displaying a desired biological activity, such as here the ability to specifically bind CGRP R, bút have one or more peptide linkages optionally replaced by a linkage selected from: -CH₂NH-, -CH₂S-, -CH₂-CH₂-, -CH-CH-(cis and trans), -COCH₂-, -CH(OH)CH₂-, and -CH₂SO-, by methods well known in the art. Systematic substitution of one or more amino acids of a consensus sequence with a D-amino acid ofthe same type (e.g., D-lysine in piacé of L-lysine) may be used in certain embodiments to generate more stable proteins. In addition, constrained peptides comprising a consensus sequence or a substantially identical consensus sequence variation may be generated by methods known in the art (Rizo and Gierasch, 1992, Ann. Rév. Biochem. 61:387)), fór example, by adding internál cysteine residues capable of forming intramolecular disulfide bridges which cyclize the peptide.

[0261] Derivatives ofthe antigén binding proteins that are described herein are alsó provided. The derivatized antigén binding proteins can comprise any molecule or substance that imparts a desired property to the antibody or fragment, such as increased half-life in a particular use. The derivatized antigén binding protein can comprise, fór example, a detectable (or labeling) moiety (e.g., a radioactive, colorimetric, antigenic or enzymatic molecule, a detectable bead (such as a magnetic or eleetrodense (e.g., gold) bead), or a molecule that binds to another molecule (e.g., biotin or streptavidin)), a therapeutic or diagnostic moiety (e.g., a radioactive, cytotoxic, or pharmaceutically active moiety), or a molecule that increases the suitability ofthe antigén binding protein fór a particular use (e.g., administration to a subject, such as a humán subject, or other in vivő or in vitro uses). Examples of molecules that can be used to derivatize an antigén binding protein include albumin (e.g., humán serum albumin) and polyethylene glycol (PEG). Albumin-linked and PEGylated derivatives of antigén binding proteins can be prepared using techniques well known in the art. Certain antigén binding proteins include a pegylated single chain polypeptide as described herein. In one embodiment, the

ΕΡ 2 379 594 Β1 antigén binding protein is conjugated or otherwise linked to transthyretin (TTR) óra TTR variant. The TTR orTTR variant can be chemically modified with, for example, a Chemical selected from the group consisting of dextran, poly(n-vinyl pyrrolidone), polyethylene glycols, propropylene glycol homopolymers, polypropylene oxide/ethylene oxide co-polymers, polyoxyethylated polyols and polyvinyl alcohols.

[0262] Other derivatives include covalent or aggregative conjugates of CGRP R binding proteins with other proteins or polypeptides, such as by expression of recombinant fusion proteins comprising heterologous polypeptides fused to the N-terminus orC-terminus ofa CGRP R binding protein. For example, the conjugated peptide may be a heterologous signal (or leader) polypeptide, e.g., the yeast alpha-factor leader, or a peptide such as an epitope tag. CGRP antigén binding protein-containing fusion proteins can comprise peptides added to facilitate purification or identification ofthe CGRP R binding protein (e.g., poly-His). A CGRP R binding protein alsó can be linked to the FLAG peptide as described in Hopp et al., 1988, Biol/Technology 6:1204; and United States Patent No. 5,011,912. The FLAG peptide is highly antigenic and provides an epitope reversibly bound by a specific monoclonal antibody (mAb), enabling rapid assay and facile purification of expressed recombinant protein. Reagents useful for preparing fusion proteins in which the FLAG peptide is fused to a given polypeptide are commercially available (Sigma, St. Louis, MO).

[0263] Oligomers that contain one or more CGRP R binding proteins may be employed as CGRP R antagonists. Oligomers may be in the form of covalently-linked or non-covalently-linked dimers, trimers, or higher oligomers. Oligomers comprising two or more CGRP R binding proteins are contemplated for use, with one example being a homodimer. Other oligomers include heterodimers, homotrimers, heterotrimers, homotetramers, heterotetramers, etc.

[0264] One embodiment is directed to oligomers comprising multiple CGRP R-binding polypeptides joined via covalent or non-covalent interactions between peptide moieties fused to the CGRP R binding proteins. Such peptides may be peptide linkers (spacers), or peptides that have the property of promoting oligomerization. Leucine zippers and certain polypeptides derived from antibodies are among the peptides that can promote oligomerization of CGRP R binding proteins attached thereto, as described in more detail below.

[0265] In particular embodiments, the oligomers comprise from two to four CGRP R binding proteins. The CGRP R binding protein moieties of the oligomer may be in any of the forms described above, e.g., variants or fragments. Preferably, the oligomers comprise CGRP R binding proteins that have CGRP R binding activity.

[0266] In one embodiment, an oligomer is prepared using polypeptides derived from immunoglobulins. Preparation of fusion proteins comprising certain heterologous polypeptides fused to various portions ofantibody-derived polypeptides (including the Fc domain) has been described, e.g., by Ashkenazi et al., 1991, Proc. Natl. Acad. Sci. USA 88:10535j_ Byrn et al., 1990, Natúré 344:677; and Hollenbaugh et al., 1992 Construction of Immunoglobulin Fusion Proteins, in Current Protocols in Immunology, Suppl. 4, pages 10.19.1-10.19.11.

[0267] One embodiment is directed to a dimer comprising two fusion proteins created by fusing a CGRP R binding protein to the Fc region of an antibody. The dimer can be made by, for example, inserting a gene fusion encoding the fusion protein intő an appropriate expression vector, expressing the gene fusion in hőst cells transformed with the recombinant expression vector, and allowing the expressed fusion protein to assemble much like antibody molecules, whereupon interchain disulfide bonds form between the Fc moieties to yield the dimer.

[0268] The term Fc polypeptide as used herein includes native and mutein forms of polypeptides derived from the Fc region of an antibody. Truncated forms ofsuch polypeptides containing the hinge region that promotes dimerization alsó are included. Fusion proteins comprising Fc moieties (and oligomers formed therefrom) offer the advantage of facile purification by affinity chromatography over Protein A or Protein G columns.

[0269] One suitable Fc polypeptide, described in PCT application WO 93/10151 and United States Patent. No. 5,426,048 and No. 5,262,522, is a single chain polypeptide extending from the N-terminal hinge region to the native Cterminus ofthe Fc region of a humán IgG 1 antibody. Another useful Fc polypeptide is the Fc mutein described in United States Patent No. 5,457,035, and in Baum et al., 1994, EMBO J. 13:3992-4001. The amino acid sequence ofthis mutein is identical to that of the native Fc sequence presented in WO 93/10151, except that amino acid 19 has been changed from Leu to Alá, amino acid 20 has been changed from Leu to Glu, and amino acid 22 has been changed from Gly to Alá. The mutein exhibits reduced affinity for Fc receptors.

[0270] In other embodiments, the variable portion of the heavy and/or light chains of a CGRP R binding protein such as disclosed herein may be substituted for the variable portion of an antibody heavy and/or light chain.

[0271] Alternatively, the oligomer is a fusion protein comprising multiple CGRP R binding proteins, with or without peptide linkers (spacer peptides). Among the suitable peptide linkers are those described in United States Patent. No. 4,751,180 and No. 4,935,233.

[0272] Another method for preparing oligomeric CGRP R binding protein derivatives involves use of a leucine zipper. Leucine zipper domains are peptides that promote oligomerization of the proteins in which they are found. Leucine zippers were originally identified in several DNA-binding proteins (Landschulz et al., 1988, Science 240:1759^ and have since been found in a variety ofdifferent proteins. Among the known leucine zippers are naturally occurring peptides and derivatives thereof that dimerize or trimerize. Examples of leucine zipper domains suitable for producing soluble oligomeric proteins are described in PCT application WO 94/10308, and the leucine zipper derived from lung surfactant

ΕΡ 2 379 594 Β1 protein D (SPD) described in Hoppé et al., 1994, FEBS Letters 344:191. The use ofa modified leucine zipper that allows for stable trimerization of a heterologous protein fused thereto is described in Fanslow et al., 1994, Sémin. Immunoi. 6:267-278. In one approach, recombinant fusion proteins comprising a CGRP R binding protein fragment or derivative fused to a leucine zipper peptide are expressed in suitable hőst cells, and the soluble oligomeric CGRP R binding protein fragments or derivatives that form are recovered from the culture supernatant.

[0273] In certain embodiments, the antigén binding protein has a K_D (equilibrium binding affinity) of less than 1 pM, 10 pM, 100 pM, 1 nM, 2 nM, 5 nM, 10 nM, 25 nM or 50 nM.

[0274] Another aspect provides an antigen-binding protein having a half-life of at least one day in vitro or in vivő (e.g., when administered to a humán subject). In one embodiment, the antigén binding protein has a half-life of at least three days. In anotherembodiment, the antibody or portion thereof has a half-life of four days or longer. In anotherembodiment, the antibody or portion thereof has a half-life of eight days or longer. In another embodiment, the antibody or antigenbinding portion thereof is derivatized or modified such that it has a longer half-life as compared to the underivatized or unmodified antibody. In another embodiment, the antigén binding protein contains point mutations to increase serum half life, such as described in WO 00/09560, published Feb. 24, 2000.

Glycosylation [0275] The antigen-binding protein may have a glycosylation pattern that is different or altered from that found in the native species. As is known in the art, glycosylation patterns can depend on both the sequence ofthe protein (e.g., the presence or absence of particular glycosylation amino acid residues, discussed below), or the hőst cell or organism in which the protein is produced. Particular expression systems are discussed below.

[0276] Glycosylation of polypeptides is typically either N-linked or O-linked. N-linked refers to the attachment of the carbohydrate moiety to the side chain of an asparagine residue. The tri-peptide sequences asparagine-X-serine and asparagine-X-threonine, where X is any amino acid except proline, are the recognition sequences for enzymatic attachment ofthe carbohydrate moiety to the asparagine side chain. Thus, the presence of either of these tri-peptide sequences in a polypeptide creates a potential glycosylation site. O-linked glycosylation refers to the attachment of one ofthe sugars N-acetylgalactosamine, galactose, or xylose, to a hydroxyamino acid, most commonly serine or threonine, although 5hydroxyproline or 5-hydroxylysine may alsó be used.

[0277] Addition of glycosylation sites to the antigén binding protein is conveniently accomplished by altering the amino acid sequence such that it contains one or more ofthe above-described tri-peptide sequences (for N-linked glycosylation sites). The alteration may alsó be made by the addition of, or substitution by, one or more serine or threonine residues to the starting sequence (for O-linked glycosylation sites). For ease, the antigén binding protein amino acid sequence may be altered through changes at the DNA level, particularly by mutating the DNA encoding the target polypeptide at preselected bases such that codons are generated that will translate intő the desired amino acids.

[0278] Another means of increasing the numberof carbohydrate moieties on the antigén binding protein is by Chemical or enzymatic coupling of glycosides to the protein. These procedures are advantageous in that they do nőt require production ofthe protein in a hőst cell that has glycosylation capabilities for N- and O-linked glycosylation. Depending on the coupling mode used, the sugar(s) may be attached to (a) arginine and histidine, (b) free carboxyl groups, (c) free sulfhydryl groups such as those of cysteine, (d)free hydroxyl groups such as those of serine, threonine, or hydroxyproline, (e) aromatic residues such as those of phenylalanine, tyrosine, or tryptophan, or (f) the amidé group of glutamine. These methods are described in WO 87/05330 published Sep. 11, 1987, and in Aplin and Wriston, 1981, CRC Crit. Rév, Biochem., pp. 259-306.

[0279] Removal of carbohydrate moieties present on the starting antigén binding protein may be accomplished chemically or enzymatically. Chemical deglycosylation requires exposure ofthe protein to the compound trifluoromethanesulfonic acid, or an equivalent compound. This treatment results in the cleavage of most or all sugars except the linking sugár (N-acetylglucosamine or N-acetylgalactosamine), while leaving the polypeptide intact. Chemical deglycosylation is described by Hakimuddin et al., 1987, Arch. Biochem. Biophys. 259:52 and by Edge et al., 1981, Anal. Biochem. 118:131. Enzymatic cleavage of carbohydrate moieties on polypeptides can be achieved bythe use ofa variety of endoand exo-glycosidases as described by Thotakura et al., 1987, Meth. Enzymol. 138:350. Glycosylation at potential glycosylation sites may be prevented by the use ofthe compound tunicamycin as described by Duskin et al., 1982, J. Bioi. Chem. 257:3105. Tunicamycin blocks the formation of protein-N-glycoside linkages.

[0280] Hence, aspects include glycosylation variants ofthe antigén binding proteins wherein the number and/or type of glycosylation site(s) has been altered compared to the amino acid sequences ofthe parent polypeptide. In certain embodiments, antibody protein variants comprise a greater or a lesser number of N-linked glycosylation sites than the native antibody. An N-linked glycosylation site is characterized by the sequence: Asn-X-Ser or Asn-X-Thr, wherein the amino acid residue designated as X may be any amino acid residue except proline. The substitution of amino acid residues to create this sequence provides a potential new site for the addition of an N-linked carbohydrate chain. Alternatively, substitutions that eliminate or altér this sequence will prevent addition of an N-linked carbohydrate chain

ΕΡ 2 379 594 Β1 present in the native polypeptide. Fór example, the glycosylation can be reduced by the deletion of an Asn or by substituting the Asn with a different amino acid. In other embodiments, one or more new N-linked sites are created. Antibodies typically have a N-linked glycosylation site in the Fc region.

Labels and Effector Groups [0281] In somé embodiments, the antigen-binding comprises one or more labels. The term labeling group or label means any detectable label. Examples of suitable labeling groups include, bút are nőt limited to, the following: radioisotopes or radionuclides (e.g., ³H, ¹⁴C, ¹⁵N, ³⁵S, ⁹⁰Y, Te, ¹¹¹ In, ¹²⁵l, ¹³¹1), fluorescent groups (e.g., FITC, rhodamine, lanthanide phosphors), enzymatic groups (e.g., horseradish peroxidase, β-galactosidase, luciferase, alkaline phosphatase), chemiluminescent groups, biotinyl groups, or predetermined polypeptide epitopes recognized by a secondary reporter (e.g., leucine zipper pair sequences, binding sites fór secondary antibodies, metál binding domains, epitope tags). In somé embodiments, the labeling group is coupled to the antigén binding protein via spacer arms of various lengths to reduce potential steric hindrance. Various methods fór labeling proteins are known in the art and may be used as is seen fit.

[0282] The term effector group means any group coupled to an antigén binding protein that acts as a cytotoxic agent. Examples fór suitable effector groups are radioisotopes or radionuclides (e.g., ³H, ¹⁴C, ¹⁵N, ³⁵S, Y, Te, ¹¹¹ln, ¹²⁵l, ¹³¹1). Other suitable groups include toxins, therapeutic groups, or chemotherapeutic groups. Examples of suitable groups include calicheamicin, auristatins, geldanamycin and maytansine. In somé embodiments, the effector group is coupled to the antigén binding protein via spacer arms of various lengths to reduce potential steric hindrance.

[0283] In generál, labels fali intő a variety of classes, depending on the assay in which they are to be detected: a) isotopic labels, which may be radioactive or heavy isotopes; b) magnetic labels (e.g., magnetic particles); c) redox active moieties; d) optical dyes; enzymatic groups (e.g. horseradish peroxidase, β-galactosidase, luciferase, alkaline phosphatase); e) biotinylated groups; and f) predetermined polypeptide epitopes recognized by a secondary reporter (e.g., leucine zipper pair sequences, binding sites fór secondary antibodies, metál binding domains, epitope tags, etc.). In somé embodiments, the labeling group is coupled to the antigén binding protein via spacer arms of various lengths to reduce potential steric hindrance. Various methods fór labeling proteins are known in the art.

[0284] Specific labels include optical dyes, including, bút nőt limited to, chromophores, phosphors and fluorophores, with the latter being specific in many instances. Fluorophores can be either small molecule fluores, or proteinaceous fluores.

[0285] By fluorescent label is meant any molecule that may be detected via its inherent fluorescent properties. Suitable fluorescent labels include, bút are nőt limited to, fluorescein, rhodamine, tetramethylrhodamine, eosin, erythrosin, coumarin, methyl-coumarins, pyrene, Malacite green, stilbene, Lucifer Yellow, Cascade BlueJ, Texas Red, IAEDANS, EDANS, BODIPY FL, LC Red 640, Cy 5, Cy 5.5, LC Red 705, Oregon green, the Alexa-Fluor dyes (Alexa Fluor 350, Alexa Fluor 430, Alexa Fluor 488, Alexa Fluor 546, Alexa Fluor 568, Alexa Fluor 594, Alexa Fluor 633, Alexa Fluor 647, Alexa Fluor 660, Alexa Fluor 680), Cascade Blue, Cascade Yellow and R-phycoerythrin (PE) (Molecular Probes, Eugene, OR), FITC, Rhodamine, and Texas Red (Pierce, Rockford, IL), Cy5, Cy5.5, Cy7 (Amersham Life Science, Pittsburgh, PA). Suitable optical dyes, including fluorophores, are described in MOLECULAR PROBES HANDBOOK by Richard P. Haugland.

[0286] Suitable proteinaceous fluorescent labels alsó include, bút are nőt limited to, green fluorescent protein, including a Renilla, Ptilosarcus, or Aequorea species of GFP (Chalfie et al., 1994, Science 263:802-805), EGFP (Clontech Labs., Inc., Genbank Accession Number U55762), blue fluorescent protein (BFP, Quantum Biotechnologies, Inc., Quebec, Canada; Stauber, 1998, Biotechniques 24:462-471; Heim etal., 1996, Curr. Bioi. 6:178-182), enhanced yellow fluorescent protein (EYFP, Clontech Labs., Inc.), luciferase (lehiki et al., 1993, J. Immunoi. 150:5408-5417), β galactosidase (Nolan et al., 1988, Proc. Natl. Acad. Sci. U.S.A. 85:2603-2607) and Renilla (WO92/15673, WO95/07463, WO98/14605, WO98/26277, WO99/49019, U nited States Patents No. 5292658, No. 5418155, No. 5683888, No. 5741668, No. 5777079, No. 5804387, No. 5874304, No. 5876995, No. 5925558).

Nucleic Acid Sequences Encoding CGRP Antigén Binding Proteins ^{* 100} [0287] Nucleic acids that encode forthe antigén binding proteins described herein, or portions thereof, are alsó provided, including nucleic acids encoding one or both chains ofan antibody, or a fragment, derivative, mutein, or variant thereof, polynucleotides encoding heavy chain variable regions or only CDRs, polynucleotides sufficient fór use as hybridization probes, PCR primers or sequencing primers fór identifying, analyzing, mutating or amplifying a polynucleotide encoding a polypeptide, anti-sense nucleic acids fór inhibiting expression of a polynucleotide, and complementary sequences of the foregoing. The nucleic acids can be any length. They can be, fór example, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 75,

100, 125, 150, 175, 200, 250, 300, 350, 400, 450, 500, 750, 1,000, 1,500 or more nucleotides in length, and/or can comprise one or more additional sequences, fór example, regulatory sequences, and/or be part of a larger nucleic acid,

ΕΡ 2 379 594 Β1 fór example, a vector. The nucleic acids can be single-stranded or double-stranded and can comprise RNA and/or DNA nucleotides, and artificial variants thereof (e.g., peptide nucleic acids).

[0288] Table 7 shows exemplary nucleic acid sequences encoding an lgG2 heavy chain constant region, a kappa light chain constant region and a lambda hCL-1 light chain constant region. Any variable region provided herein may be attached to these constant regions to form complete heavy and light chain sequences. However, it should be understood that these constant regions sequences are provided as specific examples only — one of skill in the art may employ other constant regions, including lgG1 heavy chain constant region, lgG3 or lgG4 heavy chain constant regions, any ofthe seven lambda light chain constant regions, including hCL-1, hCL-2, hCL-3 and hCL-7; constant regions that have been modified fór improved stability, expression, manufacturability or other desired characteristics, and the like. In somé embodiments, the variable region sequences are joined to other constant region sequences that are known in the art. Exemplary nucleic acid sequences encoding heavy and light chain variable regions are provided in Table 8.

Table 7: Exemplary Heavy And Light Chain Constant Region Nucleic Acid Sequences

<td> Type</td><td> Nucleic Acid Sequence/SEQ ID NO.</td>

<td> lgG2 heavy chain</td><td> gctagcaccaagggcccatcggtcttccccctggcgccctgctccaggagcacctccgagagcacagcggcc ctgggctgcctggtcaaggactacttccccgaaccggtgacggtgtcgtggaactcaggcgctctgaccagcgg cgtgcacaccttcccagctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgccctccagc aacttcggcacccagacctacacctgcaacgtagatcacaagcccagcaacaccaaggtggacaagacagt tgagcgcaaatgttgtgtcgagtgcccaccgtgcccagcaccacctgtggcaggaccgtcagtcttcctcttcccc ccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacgtgcgtggtggtggacgtgagccac gaagaccccgaggtccagttcaactggtacgtggacggcgtggaggtgcataatgccaagacaaagccacg ggaggagcagttcaacagcacgttccgtgtggtcagcgtcctcaccgttgtgcaccaggactggctgaacggc aaggagtacaagtgcaaggtctccaacaaaggcctcccagcccccatcgagaaaaccatctccaaaaccaa agggcagccccgagaaccacaggtgtacaccctgcccccatcccgggaggagatgaccaagaaccaggtc agcctgacctgcctggtcaaaggcttctaccccagcgacatcgccgtggagtgggagagcaatgggcagccg gagaacaactacaagaccacacctcccatgctggactccgacggctccttcttcctctacagcaagctcaccgt ggacaagagcaggtggcagcaggggaacgtcttctcatgctccgtgatgcatgaggctctgcacaaccactac acgcagaagagcctctccctgtctccgggtaaatga [SEQ ID NO:259]</td>

<td> lgG2 kappa light chain</td><td> cgtacggtggctgcaccatctgtcttcatcttcccgccatctgatgagcagttgaaatctggaactgcctctgttgtgt gcctgctgaataacttctatcccagagaggccaaagtacagtggaaggtggataacgccctccaatcgggtaa ctcccaggagagtgtcacagagcaggacagcaaggacagcacctacagcctcagcagcaccctgacgctg agcaaagcagactacgagaaacacaaagtctacgcctgcgaagtcacccatcagggcctgagctcgcccgt cacaaagagcttcaacaggggagagtgttag [SEQ ID NO:260]</td>

<td> lgG2 lambda hCL-1 light chain</td><td> ggtcagcccaaggccaaccccactgtcactctgttcccgccctcctctgaggagctccaagccaacaaggcca cactagtgtgtctgatcagtgacttctacccgggagctgtgacagtggcctggaaggcagatggcagccccgtc aaggcgggagtggagaccaccaaaccctccaaacagagcaacaacaagtacgcggccagcagctacctg agcctgacgcccgagcagtggaagtcccacagaagctacagctgccaggtcacgcatgaagggagcaccgt ggagaagacagtggcccctacagaatgttcatag [SEQ ID NO:261 ]</td>

ΕΡ 2 379 594 Β1 [0289] Table 8 shows exemplary nucleic acid sequences encoding heavy chain and light chain variable regions, in which the various CDRL1, CDRL2 and CDRL3, or CDRH1, CDRH2 and CDRH3, sequences are embedded.

Table 8: Exemplary Light and Heavy Chain Variable Region Nucleic Acid Sequences

<td> Reference</td><td> SEQIDNO.</td><td> Nucleic Acid Sequence</td>

<td> 2E7 V_L</td><td> 175</td><td> gacatccagatgacccagtctccatcctccctgtctgcatctgtaggagacagagtcaccatcactt gccgggcaagtcagggcattagaaatgatttaggctggtttcagcagaaaccagggaaagccc ctaagcgcctgatctatgctgcatccagtttgcaaagtggggtcccatcaaggttcagcggcagtg gatctgggacagaattcactctcacaatcagcagcctgcagcctgaagatttagcaacttattactg tctacagtataatatttacccgtggacgttcggccaagggaccaaggtggaaatcaaa</td>

<td> 13H2 V_L</td><td> 176</td><td> gacatccagatgacccagtctccatcctccctgtctgcatctgtaggagacagagtcaccatcactt gccgggcaagtcagggcattagaaaggatttaggctggtatcagcagaaaccagggaaagcc cctaagcgcctgatctatggagcatccagtttgcaaagtggggtcccatcaaggttcagcggcagt ggatctgggacagaattcactctcacaatcagcagcctgcagcctgaagattttgcaacttattact gtctacagtataatagtttcccgtggacgttcggccaagggaccaaggtggaaatcaaa</td>

<td> 33B5 V_L</td><td> 177</td><td> aggtgcagctggtgcagtctggggctgaggtgaagaagtctggggcctcagtgaaggtctcctgc aaggcttctggatacaccttcaccggctactatatgcactgggtgcgacaggcccctggacaagg gcttgagtggatgggatggatcaaccctaacagtggtggcacaaactatgtacagaagtttcagg gcagggtcaccatgaccagggacacgtccatcagcacagcctacatggagctgagcaggctg agatctgacgacacggccgtgtattactgtgcgagaaatgagtatagcagtgcctggcccttggg gtattggggccagggaaccctggtcaccgtctctagt</td>

<td> 4H6 V_L</td><td> 178</td><td> gatattgtgatgactcagtctccactctccctgcccgtcacccctggagagccggcctccatctcctg caggtctagtcagagcctcctgcatagttttgggtacaactatttggattggtacctgcagaagccag ggcagtctccacagctcctgatctatttgggttctaatcgggcctccggggtccctgacaggttcagt ggcagtggatcaggcacagattttacactgaaaatcagcagagtggaggctgaggatgttgggg tttattactgcatgcaagctctacaaactccattcactttcggccctgggaccaaagtggatatcaaa</td>

<td> 3C8 V_L</td><td> 179</td><td> gatattatactggcccagactccactttctctgtccgtcacccctggacagccggcctccatctcctg caagtctagtcagagcctcctgcacagtgctggaaagacctatttgtattggtacctgcagaagcc aggccagcctccacagctcctgatctatgaagtttccaaccggttctctggagtgccagataggttc agtggcagcgggtcagggacagatttcacactgaaaatcagccgggtggaggctgaggatgttg ggatttattactgcatgcaaagttttccgcttccgctcactttcggcggagggaccaaggtggagatc aaa</td>

EP 2 379 594 Β1 (continued)

<td> Reference</td><td> SEQIDNO.</td><td> Nucleic Acid Sequence</td>

<td> 5F5 V_L</td><td> 180</td><td> gatattattctgacccagactccactttctctgtccgtcacccctggacagccggcctccatctcctgc aagtctagtcagagcctcctgcacagtgatggaaagacctatttgtattggtacctgcagaagccc ggccagcctccacagctcctgatctatgaagtttccaaccggttctctggagagccagataggttca gtggcagcgggtcagggacagatttcacactgaaaatcagccgggtggaggctgaggatgttgg gacttattattgcatgcaaagttttccgcttccgctcactttcggcggagggaccaaggtggagatca aa</td>

<td> 12E8 V_L</td><td> 181</td><td> gatattacactgacccagactccactttctctgtccgtctcccctggacagccggcctccatctcctg caagtctagtcagagcctcctgcacagtgatggaaggaactatctgtattggtacctgcagaagcc aggccagcctccacagctcctgatctatgaagtgtccaaccggttctctggactgccagataggttc agtggcagcgggtcagggacagatttcacactgaaaatcagccgggtggaggctgaggatgttg ggatttattactgcatgcaaagttttccgcttccgctcactttcggcggagggaccaaggtggagatc aaa</td>

<td> 32H7 V_L</td><td> 182</td><td> gaaattgtgttgacgcagtctccaggcaccctgtctttgtctccaggggaaagagccaccctctcct gcagggccagtcagagtgttagcagcggctacttaacctggtaccagcagaaacctggccagg ctcccaggctcctcatctatggtgcatccagcagggccactggcatcccagacaggttcagtggc agtgggtctgggacagacttcactctcaccatcagcagactggagcctgaagattttgcagtgtatt actgtcagcagtatggtaactcactgtgcaggtttggccaggggaccaagctggagatcaaa</td>

<td> 32H7CSV_l</td><td> 183</td><td> gaaattgtgttgacgcagtctccaggcaccctgtctttgtctccaggggaaagagccaccctctcct gcagggccagtcagagtgttagcagcggctacttaacctggtaccagcagaaacctggccagg ctcccagactcctcatctatggtgcatccagcagggccactggcatcccagacaggttcagtggc agtgggtctgggacggacttcactctcaccatcagcagactggagcctgaagattttgcagtgtatt actgtcagcagtatggtaactcactgagcaggtttggccaggggaccaagctggagatcaaa</td>

<td> 33E4 V_L</td><td> 184</td><td> gaaatagtgatgacgcagtctccagccaccctgtctgtgtctccaggggaaagagccaccctctc ctgtagggccagtcagagtgttcgcagcaatttagcctggtaccagcagaaacctggccaggctc ccaggctcctcattcatgatgcatcccccaggaccgctggtatcccagccaggttcagtggcagtg gatctgggacagaattcactctcaccatcaacagcctgcagtctgaagattttgcagtttattactgtc agcagtataattactggactccgatcaccttcggccaagggacacgactggagattaaa</td>

ΕΡ 2 379 594 Β1 (continued)

<td> Reference</td><td> SEQIDNO.</td><td> Nucleic Acid Sequence</td>

<td> 32Η8 V_L</td><td> 185</td><td> gacatcgtgatgacccagtctccagactccctggctgtgtctctgggcgagagggccaccatcaa ctgcaagtccagccagagtattttagacagctccaacaatgataactacttagcttggtaccagca gaaaccaggacagcctcctaaactgctcatttactgggcatctacccgggaatccggggtccctg accgattcagtggcagcgggtctgggacagatttcactctcaccatcagcagcctgcaggctgaa gatgtggcagtttattactgtcagcaatattataatactccattcactttcggccctgggaccaaagtg gatatcaaa</td>

<td> 1E11 V_L</td><td> 186</td><td> cagtctgtgttgacgcagccgccctcagtgtctgaggccccaggacagaaggtcaccatctcctg ctctggaagcagctccaacattgggaataattatgtatcctggtaccagcagctcccaggaacag cccccaaactcctcatttatgacaataataagcgaccctcagggattcctgaccgattctctggctc caagtctggcacgtcagccaccctgggcatcaccggactccagactggggacgaggccgattat tactgcggaacatgggatagccgcctgagtgctgtggttttcggcggagggaccaagctgaccgt ccta</td>

<td> 4E4 V_L</td><td> 187</td><td> cagtctgtgttgacgcagccgccctcagtgtctgcggccccaggacagaaggtcaccatctcctg ctctggaagcagctccaacattgggaataattatgtatcctggtaccagcagctcccaggaacag cccccaaactcctcatttatgacaataataagcgaccctcagggattcctgaccgattctctggctc caagtctggcacgtcaaccaccctgggcatcaccggactccagactggggacgaggccgattat tactgcggaacatgggatagccgcctgagtgctgtggttttcggcggagggaccaagctgaccgt ccta</td>

<td> 9D4 V_L</td><td> 188</td><td> cagtctgtgttgacgcagccgccctcagtgtctgcggccccaggacagaaggtcaccatctcctg ctctggaagcagctccaacattgggaataattatgtatcctggtaccagcagttcccaggaacagc ccccaaactcctcatttatgacaataataagcgaccctcagggattcctgaccgattctctggctcc aagtctggcacgtcagccaccctgggcatcaccggactccagactggggacgaggccgattatt actgcggaacatgggatagccgcctgagtgctgtggttttcggcggagggaccaagctgaccgtc cta</td>

<td> 12G8 V_L</td><td> 189</td><td> cagtctgtgttgacgcagccgccctcagtgtctgcggccccaggacagaaggtcaccatctcctg ctctggaagcagctccaacattgggaataattatgtatcctggtaccagcagctcccaggaacag cccccaaactcctcatttatgacaataataagcgaccctcagggattcctgaccgattctctggctc caagtctggcacgtcagccaccctgggcatcaccggactccagactggggacgaggccgattat tactgcggaacatgggatagccgcctgagtgctgtggttttcggcggagggaccaagctgaccgt ccta</td>

ΕΡ 2 379 594 Β1 (continued)

<td> Reference</td><td> SEQ IDNO.</td><td> NucleicAcid Sequence</td>

<td> 34Ε3 V_L</td><td> 190</td><td> cagtctgtgttgacgcagccgccctcaatgtctgcggccccaggacagaaggtcaccatctcctg ctctggaagcagctccaacattgggaataattatgtatcctggtaccagcagctcccaggaacag cccccaaactcctcatttatgacaataataagcgaccctcagggattcctgaccgattctctggctc caagtctggcacgtcagccaccctgggcatcaccggactccagactggggacgaggccaatta ctgctgcggaacatgggatatcggcctgagtgtttgggtgttcggcggagggaccaaactgaccg tccta</td>

<td> 10E4 V_L</td><td> 191</td><td> cagtctgtgctgactcagccaccctcagcgtctgggacccccgggcagagggtcaccatctcttgt tctggaagcagttccaatatcggaagtaatactgtgaactggtaccagcagctcccaggaacggc ccccaaactcctcatctatactaataatcagcggccctcaggggtccctgaccgattctctggctcc aagtctggcacctcagcctccctggccatcagtggactccagtctgaggatgaggctgatttttact gtgcagcgcgggatgagagcctgaatggtgtggtattcggcggagggaccaagctgaccgtcct a</td>

<td> 11D11 V_L11H9 V_L</td><td> 192</td><td> cagtctgtgctgactcagccaccctcagcgtctgggacccccgggcagagagtcaccatctcttgt tctggaagcagctccaacatcggcagtaattatgtatactggtaccagcagctcccaggagcggc ccccaaactcctcatctttaggaataatcagcggccctcaggggtccctgaccgcttctctggctcc aagtctggcacctcagcctccctggccatcagtgggctccggtccgaggatgaggctgattattact gtgcagcatgggatgacagcctgagtggttgggtgttcggcggagggaccaagctgaccgtcct a</td>

<td> 1H7 V_L</td><td> 193</td><td> cagtctgtgctgactcagccaccctcagcgtctgggacccccgggcagagagtcaccatctcttgt tctggaagcagctccaacatcggcagtaattatgtatactggtaccagcagctcccaggagcggc ccccaaactcctcatctttaggagtaatcagcggccctcaggggtccctgaccgattctctggctcc aagtctggcacctcagcctccctggccatcagtgggctccggtccgaggatgaggctgattattact gtgcagcatgggatgacagcctgagtggttgggtgttcggcggagggaccaagctgaccgtcct a</td>

<td> 9F5 V_L</td><td> 194</td><td> cagtctgtgctgactcagtcaccctcagcgtctgggacccccgggcagagagtcaccatctcttgtt ctggaagcagctccaacatcggcagtaattatgtatactggtaccagcagctcccaggagcggc ccccaaactcctcatccttaggaataatcagcggccctcaggggtccctgaccgattctctggctcc aagtctggcacctcagcctccctgaccatcagtgggctccggtccgaggatgaggctgactattatt gtgcagcatgggatgacagcctgagtggttgggtgttcggcggagggaccaagctgaccgtcct a</td>

ΕΡ 2 379 594 Β1 (continued)

<td> Reference</td><td> SEQ IDNO.</td><td> Nucleic Acid Sequence</td>

<td> 3Β6 V_L</td><td> 195</td><td> tcttctgagctgactcaggaccctactgtgtctgtggccttgggacagacagtcaaaatcacatgcc aaggagacagcctcagaagtttttatgcaagctggtaccagcagaagccaggacaggcccctgt acttgtcttctatggtaaaaacaaccggccctcagggatcccagaccgattctctggctccagctca ggaaacacagcttccttgaccatcactggggctcaggcggaagatgaggctgactattattgtaat tcccgggacagcagtgtttaccatctggtactcggcggagggaccaagctgaccgtccta</td>

<td> 3B6 V_H</td><td> 196</td><td> caggtgcagttggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctg caaggcttctggatacaccttcaccggctactatatgcactgggtgcgacaggcccctggacaag ggcttgagtggatgggatggatcaaccctaacagtggtggcacaaactatgcacagaagtttcag ggcagggtcaccatgaccagggacacgtccatcagcacagcctacatggagctgagcaggct gagatctgacgacacggccgtgtatttctgtgcgagagatcaaatgagtattattatgcttcgggga gtttttcccccttactattacggtatggacgtctggggccaagggaccacggtcaccgtctctagt</td>

<td> 10E4 V_H</td><td> 197</td><td> caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcct gcaaggcttctggatacaccttcaccgactactatatgtactgggtgcgacaggcccctggacaa gggcttgagtggatgggatggatcagccctaatagtggtggcacaaactatgcccagaagtttca gggcagggtcaccatgaccagggacacgtctatcagcacagcctacatggagctgagtaggct gagatctgacgacacggccgtgtattactgtgtgagaggaggatatagtggctacgctgggctcta ctcccactactacggtatggacgtctggggccaagggaccacggtcaccgtctctagt</td>

<td> 32H8 V_H</td><td> 198</td><td> caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcct gcaaggcttctggatacaccttcaccgcctactatttacactgggtgcgacaggcccctggacaag ggcttgagtggatgggatggatcaaccctcacagtggtggcacaaactatgcacagaagtttcag ggcagggtcaccatgaccagggacacgtccatcagcacagcctacatggagctgagcaggct gagatctgacgacacggccgtgttctactgtgcgagaggaaggcagtggctgggctttgactact ggggccagggaaccctggtcaccgtctctagt</td>

<td> 33B5 V_H</td><td> 199</td><td> gacatccagatgacccagtctccatcctccctgtctgcatctgtaggagacagagttaccattacttg ccgggcaagtcagggcattagaaatgatttaggctggtatcagcagaaaccagggaaagcccc taagcgcctgatctatgttgcatccagtttgcaaagtggggtcccatcaaggttcagcggcagtgga tctgggacagaattcactctcacaatcagcagcctgcagcctgaagattttgcaacttattactgtct acagtataacacttacccgctcactttcggcggagggaccaaggtggagatcaag</td>

ΕΡ 2 379 594 Β1 (continued)

<td> Reference</td><td> SEQIDNO.</td><td> Nucleic Acid Sequence</td>

<td> 11D11 V_H</td><td> 200</td><td> gaggtacagctggtggagtctgggggaggcttggtaaagcctggggggtccctcagactctcctg tgcagcctctggattcactttcggtaacgcctggatgagctgggíccgccaggctccagggaagg ggctggagtgggttggccgtattaaaagcaaaactgatggtgggacaacagactacgctgcacc cgtgaaaggcagattcaccatctcaagagatgattcaaaaaacacgctgtatctgcaaatgaac agcctgaaaaccgaggacacagccgtgtatttctgtaccacagatcggaccgggtatagcatca gctggtctagttactactactactacggtatggacgtctggggccaagggaccacggtcaccgtct ctagt</td>

<td> 9F5 V_H</td><td> 201</td><td> gaggtgcagctggtggagtctgggggaggcttggtaaagcctggggggtcccttagactctcctgt gcagcctctggattcactttcagtaacgcctggatgagctgggtccgccaggctccagggaaggg gctggagtgggttggccgtattaaaagcaaaactgatggtgggacaacagactacactgcaccc gtgaaaggcagattcaccatctcaagagatgattcaaaaaacacgctgtatctgcaaatgaatag cctgaaagccgaggacacagccgtgtattactgtaccacagatcggaccgggtatagcatcagc tggtctagttactactactactacggtatggacgtctggggccaagggaccacggtcaccgtctcta gt</td>

<td> 11H9 V_H</td><td> 202</td><td> gaggtacagctggtggagtctgggggaggcttggtaaagcctggggggtcccttagactctcctgt gcagcctctggattcactttcggtaacgcctggatgagctgggtccgccaggctccagggaaggg gctggagtgggttggccgtattaaaagcaaaactgatggtgggacaacagactacgctgcaccc gtgaaaggcagattcaccatctcaagagatgattcaaaaaacacgctgtatctgcaaatgaaca gcctgaaaaccgaggacacagccgtgtattactgtaccacagatcggaccgggtatagcatcag ctggtctagttactactactactacggtatggacgtctggggccaagggaccacggtcaccgtctct agt</td>

<td> 1H7 V_H</td><td> 203</td><td> gaggtgcagctggtggagtctgggggaggcttggtaaagcctggggggtcccttagactctcctgt gcagcctctggattcactttcagtaacgcctggatgagctgggtccgccaggctccagggaaggg gctggagtgggttggccgtattaaaagcacaactgatggtgggacaacagactacgctgcaccc gtgaaaggcagattcaccatctcaagagatgattcaaaaaacacgctgtatctgcaaatgaaca gcctgaaaaccgaggacacagccgtgtattactgtaccacagatcggaccggatatagcatcag ctggtctagttactactactactacggtatggacgtctggggccaagggaccacggtcaccgtctct agt</td>

ΕΡ 2 379 594 Β1 (continued)

<td> Reference</td><td> SEQ IDNO.</td><td> Nucleic Acid Sequence</td>

<td> 13Η2 V_H</td><td> 204</td><td> gaggtgcagctggtggagtctgggggaggcctggtcaagcctggggggtccctgagactctcctg tgcagcctctggatacaccttcagtacctatagcatgaactgggtccgccaggctccagggaagg ggctggagtgggtctcatccattagtagtagtagtagttacagatattacgcagactcagtgaaggg ccgattcaccatctccagagacaacgccaagaactcactgtatctgcaaatgagtagcctgaga gccgaggacacggctgtgtattactgtgcgagagaaggggtgtctggcagttcgccgtatagcat cagctggtacgactactattacggtatggacgtctggggccaagggaccacggtcaccgtctcta gt</td>

<td> 2E7 V_H</td><td> 205</td><td> gaggtgcagctattggagtctgggggaggcttggtacagcctggggagtccctgagactctcctgt gcagcctctgggttcacctttagcagctatgccatgagctgggtccgccaggctccagggaaggg gctggagtgggtctcagctattagtggtagtggtggtcgcacatactacgcagactccgtgaaggg ccggttcaccatctccagagacaattccaagaacacgctgtatctgcaaatgaatagcctgagag ccgaggacacggccgtatattactgtgcgaaagatcaaagggaggtagggccgtatagcagtg gctggtacgactactactacggtatggacgtctggggccaagggaccacggtcaccgtctctagt</td>

<td> 3C8 V_H 12E8 V_H 5F5 V_H</td><td> 206</td><td> caggtgcagctggtggagtctgggggaggcgtggtccagcctgggaggtccctgagactctcctg tgcagcctctggattcaccttcagtagctatggcatgcactgggtccgccaggctccaggcaaggg gctggagtgggtggcagttatttcatatgatggaagtcatgaatcctatgcagactccgtgaagggc cgattcaccatctccagagacatttccaagaacacgctgtatctgcaaatgaacagcctgagagc tgaggacacggctgtgtatttctgtgcgagagagaggaaacgggttacgatgtctaccttatattact acttctactacggtatggacgtctggggccaagggaccacggtcaccgtctctagt</td>

<td> 4E4 V_H 9D4 V_H 1E11 V_H</td><td> 207</td><td> caggtgcagctggtggaatctgggggaggcgtggtccagcctgggaggtccctgagactctcctg tgcagcctctggattcaccttcagtagctttggcatgcactgggtccgccaggctccaggcaaggg gctggagtgggtggcagttatatcatttgatggaagtattaagtattctgtagactccgtgaagggcc gattcaccatctccagagacaattcaaagaacacgctgtttctgcaaatgaacagcctgcgagcc gaggacacggctgtgtattactgtgcgagagatcggctcaattactatgatagtagtggttattatca ctacaaatactacggtatggccgtctggggccaagggaccacggtcaccgtctctagt</td>

<td> 12G8 V_H</td><td> 208</td><td> caggtgcagctggtggaatctgggggaggcgtggtccagcctgggaggtccctgagactctcctg tgcagcctctggattcaccttcagtagctttggcatgcattgggtccgccaggctccaggcaaggg gctggagtgggtggcagttatatcatttgatggaagtattaagtactctgtagactccgtgaagggcc gattcaccatctccagagacaattcaaagaacacgctgtttctgcaaatgaacagcctgcgagcc gaggacacggctgtgtattactgtgcgagagatcggctcaattactatgatagtagtggttattatca ctacaaatactacggtctggccgtctggggccaagggaccacggtcaccgtctctagt</td>

ΕΡ 2 379 594 Β1 (continued)

<td> Reference</td><td> SEQIDNO.</td><td> Nucleic Acid Sequence</td>

<td> 4H6 V_H</td><td> 209</td><td> gaggtgcagctggtggagtctgggggaggcttggtaaagccagggcggtccctgagactctcct gtacagcttctggattcacctttggtgattatgctatgagctggttccgccaggctccagggaagggg ctggagtggataggtttcattagaagcagagcttatggtgggacaccagaatacgccgcgtctgtg aaaggcagattcaccatctcaagagatgattccaaaaccatcgcctatctgcaaatgaacagcct gaaaaccgaggacacagccgtgtatttctgtgctagaggacggggtattgcagctcgttgggact actggggccagggaaccctggtcaccgtctctagt</td>

<td> 32H7 V_H</td><td> 210</td><td> caggtgcagctggtggagtctgggggaggcgtggtccagcctgggaggtccctgagactctcctg tgcagcgtctggattcaccttcagtagctatggcatgcactgggtccgccaggctccaggcaagg ggctggagtgggtggcagttatatggtatgatggaagtaataaatactatgcagactccgtgaagg gccgattcatcatctccagagataaatccaagaacacgctgtatctgcaaatgaacagcctgaga gccgaggacacggctgtgtattactgtgcgagagcggggggtatagcagcagctggcctctacta ctactacggtatggacgtctggggccaagggaccacggtcaccgtctctagt</td>

<td> 33E4 V_H</td><td> 211</td><td> caggtgcagttacagcagtggggcgcaggactgttgaagccttcggagaccctgtccctcagctg cgctgtctatggtgggtccttcggtggttactactggagctggatccgccagcccccagggaaggg gctggagtggattggggaaatcaatcatagtggaggcaccaagtacaacccgtccctcaagagt cgagtcaccatatcagtagacacgtccaagaaccagttctccctgaagctgagctctgtgaccgc cgcggacacggctgtgtatttctgtgcgagaggcgatgtagtaggtttctttgactattggggccagg gaaccctggtcaccgtctctagt</td>

[0290] Table 9 shows the SEQ ID NOs of exemplary nucleic acid sequences encoding complete heavy and light chains, as well as heavy and light chain variable regions, of exemplary isolated antigen-binding proteins, specifically, hCGRP R binding proteins, disclosed herein.

EP 2 379 594 Β1

Exemplary HC, LC, Vh and Vl Nucleic Acid Sequence SEQ ID NOs

<td> tr</td><td> Variable Light SEQ ID NO.</td><td> Variable Heavy SEQ ID NO.</td><td> Full Light SEQ ID NO.</td><td> Full Heavy SEQ ID NO</td>

<td> 2E7</td><td> 175</td><td> 205</td><td> 226</td><td> 244</td>

<td> 13H2</td><td> 176</td><td> 204</td><td> 239</td><td> 257</td>

<td> 4H6</td><td> 178</td><td> 209</td><td> 230</td><td> 248</td>

<td> 3C8</td><td> 179</td><td> 206</td><td> 228</td><td> 246</td>

<td> 5F5</td><td> 180</td><td> 206</td><td> 231</td><td> 249</td>

<td> 12E8</td><td> 181</td><td> 206</td><td> 237</td><td> 255</td>

<td> 1E11</td><td> 186</td><td> 207</td><td> 224</td><td> 242</td>

<td> 4E4</td><td> 187</td><td> 207</td><td> 229</td><td> 247</td>

<td> 9D4</td><td> 188</td><td> 207</td><td> 232</td><td> 250</td>

<td> 12G8</td><td> 189</td><td> 208</td><td> 238</td><td> 256</td>

<td> 10E4</td><td> 191</td><td> 197</td><td> 234</td><td> 252</td>

<td> 11D11</td><td> 192</td><td> 200</td><td> 235</td><td> 253</td>

<td> 11H9</td><td> 192</td><td> 202</td><td> 236</td><td> 254</td>

<td> 1H7</td><td> 193</td><td> 203</td><td> 225</td><td> 243</td>

<td> 9F5</td><td> 194</td><td> 201</td><td> 233</td><td> 251</td>

<td> 3B6</td><td> 195</td><td> 196</td><td> 227</td><td> 245</td>

<td> 32H7</td><td> 182</td><td> 210</td><td> 240</td><td> 258</td>

ΕΡ 2 379 594 Β1

<td> ω QC</td><td> Variable Light SEQ ID NO.</td><td> Variable Heavy SEQ ID NO.</td><td> Full Light SEQ ID NO.</td><td> Full Heavy SEQ ID NO</td>

<td> 32H7 CS</td><td> 183</td><td> 210</td><td> 241</td><td> 258</td>

<td> 32H8</td><td> 185</td><td> 198</td><td></td><td></td>

<td> 33B5</td><td> 177</td><td> 199</td><td></td><td></td>

<td> 33E4</td><td> 184</td><td> 211</td><td></td><td></td>

<td> 34E3</td><td> 190</td><td> 212</td><td></td><td></td>

[0291] Nucleic acids encoding certain antigén binding proteins, or portions thereof (e.g., full length antibody, heavy or light chain, variable domain, or CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, or CDRL3) may be isolated from B-cells of mice that have been immunized with CGRP R or immunogenic components thereof, e.g., by immunizing with fulllength CGRP R (comprising both CRLR and RAMP1), with the extracellular domain of CGRP R (comprising extracellular domains of CRLR and RAMP1), with whole cells expressing CGRP R, with membranes prepared from cells expressing CGRP R, with fusion proteins, e.g., Fc fusions, comprising CRLR, RAMP1 (or extracellular domains thereof) fused to Fc, and other methods known in the art, fór example, as described in the Examples 1-3 herein. The nucleic acid may be isolated by conventional procedures such as polymerase chain reaction (PCR). Phage display is another example of a known technique whereby derivatives of antibodies and other antigén binding proteins may be prepared. In one approach, polypeptides that are components of an antigén binding protein of interest are expressed in any suitable recombinant expression system, and the expressed polypeptides are allowed to assemble toform antigén binding protein molecules.

[0292] The nucleic acids provided in Tables 7-9 are exemplary only. Due to the degeneracy ofthe genetic code, each of the polypeptide sequences listed in Tables 2-5 or otherwise depicted herein are alsó encoded by a large number of other nucleic acid sequences besides those provided. One of ordinary skill in the art will appreciate that the present application thus provides adequate written description and enablement fór each degenerate nucleotide sequence encoding each antigén binding protein.

[0293] An aspect further provides nucleic acids that hybridize to other nucleic acids (e.g., nucleic acids comprising a nucleotide sequence listed in Table 7, Table 8, Table 9 and/or SEQ ID NOs:224-258) under particular hybridization conditions. Methods fór hybridizing nucleic acids are well-known in the art. See, e.g., Current Protocols in Molecular Biology, John Wiley &amp; Sons, N.Y. (1989), 6.3.1-6.3.6. As defined herein, a moderately stringent hybridization condition uses a prewashing solution containing 5x sodium chloride/sodium citrate (SSC), 0.5% SDS, 1.0 mM EDTA (pH 8.0), hybridization buffer of about 50% formamide, 6x SSC, and a hybridization temperature of 55°C (or other similar hybridization Solutions, such as one containing about 50% formamide, with a hybridization temperature of 42°C), and washing conditions of 60°C, in 0.5x SSC, 0.1% SDS. A stringent hybridization condition hybridizes in 6x SSC at 45°C, followed by one or more washes in O.IxSSC, 0.2% SDSat68°C. Furthermore, one of skill in the art can manipulatethe hybridization and/or washing conditions to increase or decrease the stringency of hybridization such that nucleic acids comprising nucleotide sequences that are at least 65%, 70%, 75%, 80%, 85%, 90%, 95%, 98% or 99% identical to each other typically remain hybridized to each other.

[0294] The basic parameters affecting the choice of hybridization conditions and guidance fór devising suitable conditions are set forth by, fór example, Sambrook, Fritsch, and Maniatis (2001, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., supra: and Current Protocols in Molecular Biology, 1995, Ausubel et al., eds., John Wiley &amp; Sons, Inc., sections 2.10 and 6.3-6.4), and can be readily determined by those

ΕΡ 2 379 594 Β1 having ordinary skill in the art based on, e.g., the length and/or base composition ofthe nucleic acid.

[0295] Changes can be introduced by mutation intő a nucleic acid, thereby leading to changes in the amino acid sequence of a polypeptide (e.g., an antibody or antibody derivative) that it encodes. Mutations can be introduced using any technique known in the art. In one embodiment, one or more particular amino acid residues are changed using, fór example, a site-directed mutagenesis protocol. In another embodiment, one or more randomly selected residues is changed using, fór example, a random mutagenesis protocol. However it is made, a mutant polypeptide can be expressed and screened fór a desired property.

[0296] Mutations can be introduced intő a nucleic acid without significantly altering the biological activity ofa polypeptide that it encodes. Fór example, one can make nucleotide substitutions leading to amino acid substitutions at non-essential amino acid residues. Alternatively, one or more mutations can be introduced intő a nucleic acid that selectively changes the biological activity ofa polypeptide that it encodes. Fór example, the mutation can quantitatively or qualitatively change the biological activity. Examples of quantitative changes include increasíng, reducing oreliminating the activity. Examples of qualitative changes include changing the antigén specificity ofan antibody. In one embodiment, a nucleic acid encoding any antigén binding protein described herein can be mutated to altér the amino acid sequence using molecular biology techniques that are well-established in the art.

[0297] Another aspect provides nucleic acid molecules that are suitable fór use as primers or hybridization probes fór the detection of nucleic acid sequences. A nucleic acid molecule can comprise oniy a portion of a nucleic acid sequence encoding a full-length polypeptide, fór example, a fragment that can be used as a probe or primeror a fragment encoding an active portion (e.g., a CGRP R binding portion) ofa polypeptide.

[0298] Probes based on the sequence of a nucleic acid can be used to detect the nucleic acid or similar nucleic acids, fór example, transcripts encoding a polypeptide. The probe can comprise a label group, e.g., a radioisotope, a fluorescent compound, an enzyme, oran enzyme co-factor. Such probes can be used to identify a cell that expresses the polypeptide. [0299] Another aspect provides vectors comprising a nucleic acid encoding a polypeptide or a portion thereof (e.g., a fragment containing one or more CDRs or one or more variable region domains). Examples of vectors include, bút are nőt limited to, plasmids, viral vectors, non-episomal mammalian vectors and expression vectors, fór example, recombinant expression vectors. The recombinant expression vectors can comprise a nucleic acid in a form suitable fór expression ofthe nucleic acid in a hőst cell. The recombinant expression vectors include oneor more regulatory sequences, selected on the basis of the hőst cells to be used fór expression, which is operably linked to the nucleic acid sequence to be expressed. Regulatory sequences include those that direct constitutive expression of a nucleotide sequence in many types of hőst cells (e.g., SV40 eariy gene enhancer, Rous sarcoma vírus promoter and cytomegalovirus promoter), those that direct expression ofthe nucleotide sequence oniy in certain hőst cells (e.g., tissue-specific regulatory sequences, see, Voss et al., 1986, Trends Biochem. Sci. 11:287, Maniatis et al., 1987, Science 236:1237), and those that direct inducible expression of a nucleotide sequence in response to particular treatment or condition (e.g., the metallothionin promoter in mammalian cells and the tet-responsive and/or streptomycin responsive promoter in both prokaryotic and eukaryotic systems (see, id.). It will be appreciated by those skilled in the art that the design of the expression vector can depend on such factors as the choice of the hőst cell to be transformed, the level of expression of protein desired, etc. The expression vectors can be introduced intő hőst cells to thereby produce proteins or peptides, including fusion proteins or peptides, encoded by nucleic acids as described herein.

[0300] Another aspect provides hőst cells intő which a recombinant expression vector has been introduced. A hőst cell can be any prokaryotic cell (fór example, E. coli) or eukaryotic cell (fór example, yeast, insect, or mammalian cells (e.g., CHO cells)). Vector DNA can be introduced intő prokaryotic or eukaryotic cells via conventional transformation or transfection techniques. Fór stable transfection of mammalian cells, it is known that, depending upon the expression vector and transfection technique used, oniy a small fraction of cells may integrate the foreign DNA intő their genome. In order to identify and select these integrants, a gene that encodes a selectable marker (e.g., fór resistance to antibiotics) is generally introduced intő the hőst cells along with the gene of interest. Preferred selectable markers include those which confer resistance to drugs, such as G418, hygromycin and methotrexate. Cells stably transfected with the introduced nucleic acid can be identified by drug selection (e.g., cells that have incorporated the selectable marker gene will survive, while the other cells die), among other methods.

Preparing Of Antigén Binding Proteins [0301] Non-human antibodies that are provided can be, fór example, derived from any antibody-producing animal, such as mouse, rat, rabbit, goat, donkey, or non-human primate (such as monkey (e.g., cynomolgus or rhesus monkey) or ape (e.g., chimpanzee)). Non-human antibodies can be used, fór instance, in in vitro cell culture and cell-culture based applications, or any other application where an immuné response to the antibody does nőt occur or is insignificant, can be prevented, is nőt a concern, or is desired. In certain embodiments, the antibodies may be produced by immunizing animals using methods known in the art, as described above and/or in Examples 1-3 below. The examples describe the generation of anti CGRP R antibodies using three different immunogen preparations - (i) whole cells expressing full74

ΕΡ 2 379 594 Β1 length versions oftwo major components of CGRP R- RAMP1 and CRLR; (ii) membráné extracts from such cells; and (iii) soluble CGRP R obtained by co-expressing and purifying the N-terminal extracellular domains of CRLR and RAMP1. The antibodies may be polyclonal, monoclonal, or may be synthesized in hőst cells by expressing recombinant DNA. Fully humán antibodies may be prepared as described above by immunizing transgenic animals containing humán immunoglobulin loci or by selecting a phage display library that is expressing a repertoire of humán antibodies.

[0302] The monoclonal antibodies (mAbs) can be produced by a variety of techniques, including conventional monoclonal antibody methodology, e.g., the standard somatic cell hybridization technique of Kohler and Milstein, 1975, Natúré 256:495. Alternatively, other techniques for producing monoclonal antibodies can be employed, for example, the viral or oncogenic transformation of B-lymphocytes. One suitable animal system for preparing hybridomas is the murine system, which is a very well established procedure. Immunization protocols and techniques for isolation of immunized splenocytes for fusion are known in the art and illustrative approaches are described in the Examples, below. For such procedures, B cells from immunized mice are typically fused with a suitable immortalized fusion partner, such as a murine myeloma cell line. If desired, rats or other mammals besides can be immunized instead of mice and B cells from such animals can be fused with the murine myeloma cell line to form hybridomas. Alternatively, a myeloma cell line from a source other than mouse may be used. Fusion procedures for making hybridomas alsó are well known.

[0303] The single chain antibodies that are provided may be formed by linking heavy and light chain variable domain (Fv region) fragments via an amino acid bridge (short peptide linker), resulting in a single polypeptide chain. Such singlechain Fvs (scFvs) may be prepared by fusing DNA encoding a peptide linker between DNAs encoding the two variable domain polypeptides (V_L and V_H). The resulting polypeptides can fold back on themselves to form antigen-binding monomers, or they can form multimers (e.g., dimers, trimers, or tetramers), depending on the length of a flexible linker between the two variable domains (Kortt et al., 1997, Prot. Eng. 10:423; Kortt et al., 2001, Biomol. Eng. 18:95-108). By combining different V_L and V_H -comprising polypeptides, one can form multimeric scFvs that bind to different epitopes (Kriangkum et al., 2001, Biomol. Eng. 18:31-40). Techniques developed for the production of single chain antibodies include those described in U.S. Pat. No. 4,946,778; Bírd, 1988, Science 242:423; Huston et al., 1988, Proc. Natl. Acad. Sci. U.S.A. 85:5879; Ward etal., 1989, Natúré 334:544, de Graaf et al., 2002, Methods Mól Bioi. 178:379-387. Single chain antibodies derived from antibodies provided herein include, bút are nőt limited to scFvs comprising the variable domain combinations of the heavy and light chain variable regions depicted in Table 3, or combinations of light and heavy chain variable domains which include CDRs depicted in Tables 4A and 4B.

[0304] Antibodies provided herein that are of one subclass can be changed to antibodies from a different subclass using subclass switching methods. Thus, IgG antibodies may be derived from an IgM antibody, for example, and vice versa. Such techniques allow the preparation of new antibodies that possess the antigén binding properties of a given antibody (the parent antibody), bút alsó exhibit biological properties associated with an antibody isotype or subclass different from that of the parent antibody. Recombinant DNA techniques may be employed. Cloned DNA encoding particular antibody polypeptides may be employed in such procedures, e.g., DNA encoding the constant domain of an antibody ofthe desired isotype. See, e.g., Lantto et al., 2002, Methods Mól. Bioi. 178:303-316.

[0305] Accordingly, the antibodies that are provided include those comprising, for example, the variable domain combinations described, supra., having a desired isotype (for example, IgA, IgG 1, lgG2, lgG3, lgG4, IgE, and IgD) as well as Fab or F(ab’)₂ fragments thereof. Moreover, if an lgG4 is desired, it may alsó be desired to introduce a point mutation (CPSCP->CPPCP) in the hinge region as described in Bloom et al., 1997, Protein Science 6:407 to alleviate a tendency to form intra-H chain disulfide bonds that can lead to heterogeneity in the lgG4 antibodies.

[0306] Moreover, techniques for deriving antibodies having different properties (i.e., varying affinities for the antigén to which they bind) are alsó known. One such technique, referred to as chain shuffling, involves displaying immunoglobulin variable domain gene repertoires on the surface of filamentous bacteriophage, often referred to as phage display. Chain shuffling has been used to prepare high affinity antibodies to the hapten 2-phenyloxazol-5-one, as described by Marks et al., 1992, BioTechnology 10:779.

[0307] Conservative modifications may be made to the heavy and light chain variable regions described in Table 3, orthe CDRs described in Tables 4A and 4B (and corresponding modifications to the encoding nucleic acids) to produce a CGRP R binding protein having certain desirable functional and biochemical characteristics. Methods for achieving such modifications are described above.

[0308] CGRP antigén binding proteins may be further modified in various ways. For example, if they are to be used for therapeutic purposes, they may be conjugated with polyethylene glycol (pegylated) to prolong the serum half-life or to enhance protein delivery. Alternatively, the V region ofthe subject antibodies or fragments thereof may be fused with the Fc region of a different antibody molecule. The Fc region used for this purpose may be modified so that it does nőt bind complement, thus reducing the likelihood of inducing cell lysis in the patient when the fusion protein is used as a therapeutic agent. In addition, the subject antibodies or functional fragments thereof may be conjugated with humán serum albumin to enhance the serum half-life of the antibody or antigén binding fragment thereof. Another useful fusion partner for the antigén binding proteins or fragments thereof is transthyretin (TTR). TTR has the capaeity to form a tetramer, thus an antibody-TTR fusion protein can form a multivalent antibody which may increase its binding avidity.

ΕΡ 2 379 594 Β1 [0309] Alternatively, substantial modifications in the functional and/or biochemical characteristics ofthe antigén binding proteins described herein may be achieved by creating substitutions in the amino acid sequence ofthe heavy and light chains that differ significantly in their effect on maintaining (a) the structure ofthe molecular backbone in the area ofthe substitution, for example, as a sheet or helical conformation, (b) the charge or hydrophobicity of the molecule at the target site, or (c) the bulkiness ofthe side chain. A conservative amino acid substitution may involve a substitution of a native amino acid residue with a nonnative residue that has little or no effect on the polarity or charge ofthe amino acid residue at that position. See, Table 4, supra. Furthermore, any native residue in the polypeptide may alsó be substituted with alanine, as has been previously described for alanine scanning mutagenesis.

[0310] Amino acid substitutions (whether conservative or non-conservative) ofthe subject antibodies can be implemented by those skilled in the art by applying routine techniques. Amino acid substitutions can be used to identify important residues ofthe antibodies provided herein, orto increase or decrease the affinity of these antibodies for humán CGRP R orfor modifying the binding affinity of other antigen-binding proteins described herein.

Methods Of Expressing Antigén Binding Proteins [0311] Expression systems and constructs in the form of plasmids, expression vectors, transeription or expression cassettes that comprise at least one polynucleotide as described above are alsó provided herein, as well hőst cells comprising such expression systems or constructs.

[0312] The antigén binding proteins provided herein may be prepared by any ofa number of conventional techniques. For example, CGRP R antigén binding proteins may be produced by recombinant expression systems, using any technique known in the art. See, e.g., Monoclonal Antibodies, Hybridomas: A New Dimension in Biological Analyses, Kennet et al. (eds.) Plenum Press, New York (1980); and Antibodies: A Laboratory Manual, Harlow and Lane (eds.), Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (1988).

[0313] Antigén binding proteins can be expressed in hybridoma cell lines (e.g., in partieuiar antibodies may be expressed in hybridomas) or in cell lines other than hybridomas. Expression constructs encoding the antibodies can be used to transform a mammalian, insect or microbial hőst cell. Transformation can be performed using any known method for introducing polynucleotides intő a hőst cell, including, for example packaging the polynucleotide in a vírus or bacteriophage and transducing a hőst cell with the construct by transfection procedures known in the art, as exemplified by United States Patent No. 4,399,216; No. 4,912,040; No. 4,740,461; No. 4,959,455. The optimál transformation procedure used will depend upon which type of hőst cell is being transformed. Methods for introduction of heterologous polynucleotides intő mammalian cells are well known in the art and include, bút are nőt limited to, dextran-mediated transfection, calcium phosphate precipitation, polybrene mediated transfection, protoplast fusion, eleetroporation, encapsulation of the polynucleotide(s) in liposomes, mixing nucleic acid with positively-charged lipids, and direct microinjection of the DNA intő nuclei.

[0314] Recombinant expression constructs typically comprise a nucleic acid molecule encoding a polypeptide comprising one or more of the following: one or more CDRs provided herein; a light chain constant region; a light chain variable region; a heavy chain constant region (e.g., C_H1, C_H2 and/or C_H3); and/or another scaffold portion of a CGRP R antigén binding protein. These nucleic acid sequences are ínserted intő an appropriate expression vector using standard ligation techniques. In one embodiment, the heavy or light chain constant region is appended to the C-terminus ofthe anti-CGRP R-specific heavy or light chain variable region and is ligated intő an expression vector. The vector is typically selected to be functional in the partieuiar hőst cell employed (i.e., the vector is compatible with the hőst cell machinery, permitting amplification and/or expression ofthe gene can occur). In somé embodiments, vectors are used that employ protein-fragment complementation assays using protein reporters, such as dihydrofolate reductase (see, for example, U.S. Pat. No. 6,270,964 ). Suitable expression vectors can be purchased, for example, from Invitrogen Life Technologies or BD Biosciences (formerly Clontech). Other useful vectors for cloning and expressing the antibodies and fragments include those described in Bianchi and McGrew, 2003, Biotech. Biotechnoi. Bioeng. 84:439-44. Additional suitable expression vectors are discussed, for example, in Methods Enzymol., vol. 185 (D. V. Goeddel, ed.), 1990, New York: Academic Press.

[0315] Typically, expression vectors used in any ofthe hőst cells will contain sequences for plasmid maintenance and for cloning and expression of exogenous nucleotide sequences. Such sequences, collectively referred to as flanking sequences in certain embodiments will typically include one or more ofthe following nucleotide sequences: a promoter, one or more enhancer sequences, an origin of replication, a transcriptional termination sequence, a complete intron sequence containing a donor and acceptor splice site, a sequence encoding a leader sequence for polypeptide secretion, a ribosome binding site, a polyadenylation sequence, a polylinker region for inserting the nucleic acid encoding the polypeptide to be expressed, and a selectable marker element. Each of these sequences is discussed below.

[0316] Optionally, the vector may contain a tag-encoding sequence, i.e., an oligonucleotide molecule located at the 5’ or 3’ end ofthe CGRP R binding protein coding sequence; the oligonucleotide sequence encodes polyHis (such as hexaHis), or another tag such as FLAG®, HA (hemaglutinin influenza vírus), or myc, for which commercially available

ΕΡ 2 379 594 Β1 antibodies exist. This tag is typically fused to the polypeptide upon expression of the polypeptide, and can serve as a means for affinity purification or detection ofthe CGRP R binding protein from the hőst cell. Affinity purification can be accomplished, for example, by column chromatography using antibodies against the tag as an affinity mátrix. Optionally, the tag can subsequently be removed from the purified CGRP R binding protein by various means such as using certain peptidases for cleavage.

[0317] Flanking sequences may be homologous (i.e., from the same species and/or strain as the hőst cell), heterologous (i.e., from a species other than the hőst cell species or strain), hybrid (i.e., a combination of flanking sequences from more than one source), synthetic or native. As such, the source of a flanking sequence may be any prokaryotic or eukaryotic organism, any vertebrate or invertebrate organism, or any plánt, provided that the flanking sequence is functional in, and can be activated by, the hőst cell machinery.

[0318] Flanking sequences useful in the vectors may be obtained by any of several methods well known in the art. Typically, flanking sequences useful herein will have been previously identified by mapping and/or by restriction endonuclease digestion and can thus be isolated from the propertissue source using the appropriate restriction endonucleases. In somé cases, the full nucleotide sequence of a flanking sequence may be known. Here, the flanking sequence may be synthesized using the methods described herein for nucleic acid synthesis or cloning.

[0319] Whether all or only a portion of the flanking sequence is known, it may be obtained using polymerase chain reaction (PCR) and/or by screening a genomic library with a suitable probe such as an oligonucleotide and/or flanking sequence fragment from the same or another species. Where the flanking sequence is nőt known, a fragment of DNA containing a flanking sequence may be isolated from a larger piece of DNA that may contain, for example, a coding sequence or even another gene or genes. Isolation may be accomplished by restriction endonuclease digestion to produce the proper DNA fragment followed by isolation using agarose gél purification, Oiagen® column chromatography (Chatsworth, CA), or other methods known to the skilled artisan. The selection of suitable enzymes to accomplish this purpose will be readily apparent to one of ordinary skill in the art.

[0320] An origin of replication is typically a part of those prokaryotic expression vectors purchased commercially, and the origin aids in the amplification ofthe vector in a hőst cell. Ifthe vector of choice does nőt contain an origin of replication site, one may be chemically synthesized based on a known sequence, and ligated intő the vector. For example, the origin of replication from the plasmid pBR322 (New England Biolabs, Beverly, MA) is suitable for most gram-negative bacteria, and various viral origins (e.g., SV40, polyoma, adenovirus, vesicularstomatitus vírus (VSV), or papillomaviruses such as HPV or BPV) are useful for cloning vectors in mammalian cells. Generally, the origin of replication component is nőt needed for mammalian expression vectors (for example, the SV40 origin is often used only because it alsó contains the vírus early promoter).

[0321] A transcription termination sequence is typically located 3’ to the end ofa polypeptide coding region and serves to terminate transcription. Usually, a transcription termination sequence in prokaryotic cells is a G-C rich fragment followed by a poly-T sequence. While the sequence is easily cloned from a library or even purchased commercially as part ofa vector, itcan alsó be readily synthesized using methods for nucleic acid synthesis such as those described herein. [0322] A selectable marker gene encodes a protein necessary for the survival and growth of a hőst cell grown in a selective culture médium. Typical selection marker genes encode proteins that (a) confer resistance to antibiotics or other toxins, e.g., am pici Ilin, tetracycline, or kanamycin for prokaryotic hőst cells; (b) complement auxotrophicdeficiencies of the cell; or (c) supply critical nutrients nőt available from complex or defined média. Specific selectable markers are the kanamycin resistance gene, the ampicillin resistance gene, and the tetracycline resistance gene. Advantageously, a neomycin resistance gene may alsó be used for selection in both prokaryotic and eukaryotic hőst cells.

[0323] Other selectable genes may be used to amplify the gene that will be expressed. Amplification is the process wherein genes that are required for production of a protein critical for growth or cell survival are reiterated in tandem within the chromosomes of successive generations of recombinant cells. Examples of suitable selectable markers for mammalian cells include dihydrofolate reductase (DHFR) and promoterless thymidine kinase genes. Mammalian cell transformants are placed under selection pressure wherein only the transformants are uniquely adapted to survive by virtue of the selectable gene present in the vector. Selection pressure is imposed by culturing the transformed cells under conditions in which the concentration of selection agent in the médium is successively increased, thereby leading to the amplification of both the selectable gene and the DNA that encodes another gene, such as an antigén binding protein that binds to CGRP R. As a result, increased quantities ofa polypeptide such as an antigén binding protein are synthesized from the amplified DNA.

[0324] A ribosome-binding site is usually necessary for translation initiation of mRNA and is characterized by a ShineDalgarno sequence (prokaryotes) or a Kozák sequence (eukaryotes). The element is typically located 3’ to the promoter and 5’ to the coding sequence ofthe polypeptide to be expressed.

[0325] In somé cases, such as where glycosylation is desired in a eukaryotic hőst cell expression system, one may manipulate the various pre-or pro-sequences to improve glycosylation or yield. For example, one may altér the peptidase cleavage site of a particular signal peptide, or add prosequences, which alsó may affect glycosylation. The final protein product may have, in the -1 position (relatíve to the first amino acid ofthe mature protein), one or more additional amino

ΕΡ 2 379 594 Β1 acids incident to expression, which may nőt have been totally removed. Fór example, the final protein product may have one ortwo amino acid residues found in the peptidase cleavage site, attached to the amino-terminus. Alternatively, use of somé enzyme cleavage sites may result in a slightly truncated form ofthe desired polypeptide, if the enzyme cuts at such area within the mature polypeptide.

[0326] Expression and cloning will typically contain a promoter that is recognized by the hőst organism and operably linked to the molecule encoding a CGRP R binding protein. Promoters are untranscribed sequences located upstream (i.e., 5j to the start codon of a structural gene (generally within about 100 to 1000 bp) that control transcription of the structural gene. Promoters are conventionally grouped intő one of two classes: inducible promoters and constitutive promoters. Inducible promoters initiate increased levels of transcription from DNA under their control in response to somé change in culture conditions, such as the presence or absence ofa nutrient óra change in temperature. Constitutive promoters, on the other hand, uniformly transcribe a gene to which they are operably linked, that is, with little or no control over gene expression. A large number of promoters, recognized by a variety of potential hőst cells, are well known. A suitable promoter is operably linked to the DNA encoding heavy chain or light chain comprising a CGRP R binding protein by removing the promoterfrom the source DNA by restriction enzyme digestion and inserting the desired promoter sequence intő the vector.

[0327] Suitable promoters fór use with yeast hosts are alsó well known in the art. Yeast enhancers are advantageously used with yeast promoters. Suitable promoters fór use with mammalian hőst cells are well known and include, bút are nőt limited to, those obtained from the genomes of viruses such as polyoma vírus, fowlpox vírus, adenovirus (such as Adenovirus 2), bovine papilloma vírus, avian sarcoma vírus, cytomegalovirus, retroviruses, hepatitis-B vírus, and Simian Vírus 40 (SV40). Other suitable mammalian promoters include heterologous mammalian promoters, fór example, heatshock promoters and the actin promoter.

[0328] Additional promoters which may be of interest include, bút are nőt limited to: SV40 early promoter (Benoist and Chambon, 1981, Natúré 290:304-310); CMV promoter (Thornsen et al., 1984, Proc. Natl. Acad. U.S.A. 81:659-663); the promoter contained in the 3’ long terminál repeat of Rous sarcoma vírus (Yamamoto et al., 1980, Cell 22:787-797); herpes thymidine kinase promoter (Wagner et al., 1981, Proc. Natl. Acad. Sci. U.S.A. 78:1444-1445); promoter and regulatory sequences from the metallothionine gene (Prinsteretal., 1982, Natúré 296:39-42); and prokaryotic promoters such as the beta-lactamase promoter (Villa-Kamaroff et al., 1978, Proc. Natl. Acad. Sci. U.S.A. 75:3727-3731); or the tac promoter (DeBoer et al., 1983, Proc. Natl. Acad. Sci. U.S.A. 80:21-25). Alsó of interest are the following animal transcriptional control regions, which exhibit tissue specificity and have been utilized in transgenic animals: the elastase I gene control region that is active in pancreatic acinar cells (Swift et al., 1984, Cell 38:639-646; Ornitz et al., 1986, Cold Spring Harbor Symp. Quant. Bioi. 50:399-409; MacDonald, 1987, Hepatology 7:425-515); the insulin gene control region that is active in pancreatic béta cells (Hanahan, 1985, Natúré 315:115-122); the immunoglobulin gene control region that is active in lymphoid cells (Grosschedl et al., 1984, Cell 38:647-658; Adames et al., 1985, Natúré 318:533-538; Alexander et al., 1987, Mól. Cell. Bioi. 7:1436-1444); the mouse mammary tumor vírus control region that is active in testicular, breast, lymphoid and mást cells (Leder et al., 1986, Cell 45:485-495); the albumin gene control region that is active in liver (Pinkert et al., 1987, Genes and Devel. 1 :268-276); the alpha-feto-protein gene control region that is active in liver (Krumlauf et al., 1985, Mól. Cell. Bioi. 5:1639-1648; Hammer et al., 1987, Science 253:53-58); the alpha 1antitrypsin gene control region that is active in liver (Kelsey et al., 1987, Genes and Devel. 1:161-171); the beta-globin gene control region that is active in myeloid cells (Mogram et al., 1985, Natúré 315:338-340; Kollias et al., 1986, Cell 46:89-94); the myelin basic protein gene control region that is active in oligodendrocyte cells in the brain (Readhead et al., 1987, Cell 48:703-712); the myosin light chain-2 gene control region that is active in skeletal muscle (Sani, 1985, Natúré 314:283-286); and the gonadotropic releasing hormoné gene control region that is active in the hypothalamus (Mason et al., 1986, Science 234:1372-1378).

[0329] An enhancer sequence may be inserted intő the vector to increase transcription of DNA encoding light chain or heavy chain comprising a humán CGRP R binding protein by higher eukaryotes. Enhancers are cis-acting elements of DNA, usually about 10-300 bp in length, that act on the promoter to increase transcription. Enhancers are relatively orientation and position independent, having been found at positions both 5’ and 3’ to the transcription unit. Several enhancer sequences available from mammalian genes are known (e.g., globin, elastase, albumin, alpha-feto-protein and insulin). Typically, however, an enhancer from a vírus is used. The SV40 enhancer, the cytomegalovirus early promoter enhancer, the polyoma enhancer, and adenovirus enhancers known in the art are exemplary enhancing elements fór the activation of eukaryotic promoters. While an enhancer may be positioned in the vector either 5’ or 3’ to a coding sequence, it is typically located at a site 5’ from the promoter. A sequence encoding an appropriate native or heterologous signal sequence (leader sequence or signal peptide) can be incorporated intő an expression vector, to promote extracellular secretion ofthe antibody. The choice of signal peptide or leader depends on the type of hőst cells in which the antibody is to be produced, and a heterologous signal sequence can replace the native signal sequence. Examples of signal peptides that are functional in mammalian hőst cells include the following: the signal sequence fór interleukin-7 (IL-7) described in US Patent No. 4,965,195; the signal sequence fór interleukin-2 receptor described in Cosman et al., 1984, Natúré 312:768; the interleukin-4 receptor signal peptide described in EP Patent No. 0367 566;

ΕΡ 2 379 594 Β1 the type I interleukin-1 receptor signal peptide deseribed in U.S. Patent No. 4,968,607; the type II interleukin-1 receptor signal peptide deseribed in EP Patent No. 0 460 846.

[0330] The expression vectors that are provided may be constructed from a starting vector such as a commercially available vector. Such vectors may or may nőt contain all of the desired flanking sequences. Where one or more of the flanking sequences deseribed herein are nőt already present in the vector, they may be individually obtained and ligated intő the vector. Methods used fór obtaining each of the flanking sequences are well known to one skilled in the art. [0331] After the vector has been constructed and a nucleic acid molecule encoding light chain, a heavy chain, or a light chain and a heavy chain comprising a CGRP R antigén binding sequence has been inserted intő the proper site of the vector, the completed vector may be inserted intő a suitable hőst cell fór amplification and/or polypeptide expression. The transformation ofan expression vector fór an antigen-binding protein intő a selected hőst cell may be accomplished by well known methods including transfection, infection, calcium phosphate co-precipitation, eleetroporation, microinjection, lipofection, DEAE-dextran mediated transfection, or other known techniques. The method selected will in part be a function ofthe type of hőst cell to be used. These methods and other suitable methods are well known to the skilled artisan, and are set forth, fór example, in Sambrook et al., 2001, supra.

[0332] A hőst cell, when cultured under appropriate conditions, synthesizes an antigén binding protein that can subsequently be collected from the culture médium (ifthe hőst cell secretes it intő the médium) or dírectiy from the hőst cell producing it (if it is nőt secreted). The selection of an appropriate hőst cell will depend upon various factors, such as desired expression levels, polypeptide modifications that are desirabie or necessary fór activity (such as glycosylation or phosphorylation) and ease of folding intő a biologically active molecule.

[0333] Mammalian cell lines available as hosts fór expression are well known in the art and include, bút are nőt limited to, immortalized cell lines available from the American Type Culture Collection (ATCC), including bút nőt limited to Chinese hamster ovary (CHO) cells, HeLa cells, baby hamster kidney (BHK) cells, monkey kidney cells (COS), humán hepatocellular carcinoma cells (e.g., Hep G2), and a number of other cell lines. In certain embodiments, cell lines may be selected through determining which cell lines have high expression levels and constitutively produce antigén binding proteins with CGRP R binding properties. In another embodiment, a cell line from the B cell lineage that does nőt make its own antibody bút has a capacity to make and secrete a heterologous antibody can be selected.

Use of Humán CGRP Antigén Binding Proteins fór Diagnostic and Therapeutic Purposes [0334] Antigén binding proteins are useful fór detecting CGRP R in biological samples and identification of cells or tissues that produce CGRP R. Fór instance, the CGRP R antigén binding proteins can be used in diagnostic assays, e.g., binding assays to detect and/or quantify CGRP R expressed in a tissue or cell. Antigén binding proteins that specifically bind to CGRP R can alsó be used in treatment of diseases related to CGRP R in a patient in need thereof. In addition, CGRP R antigén binding proteins can be used to inhibit CGRP R from forming a complex with its ligand CGRP, thereby modulating the biological activity of CGRP R in a cell or tissue. Examples of activities that can be modulated include, bút are nőt limited to, inhibiting vasodialation and/or decrease neurogenic inflammation. Antigén binding proteins that bind to CGRP R thus can modulate and/or block interaction with other binding compounds and as such may have therapeutic use in ameliorating diseases related to CGRP R.

Indications [0335] A disease or condition associated with humán CGRP R includes any disease or condition whose onset in a patient is caused by, at least in part, the interaction of CGRP R with its ligand, CGRP. The severity of the disease or condition can alsó be increased or decreased by the interaction of CGRP R with CGRP. Examples of diseases and conditions that can be treated with the antigén binding proteins deseribed herein include headaches, such as cluster headaches, migraine, including migraine headaches, chronic pain, type II diabetes mellitus, inflammation, e.g., neurogenic inflammation, cardiovasculardisorders, and hemodynamiederangement associated with endotoxemia and sepsis. [0336] In particular, antigén binding proteins deseribed herein can be used to treat migraine, either asan acute treatment commencing after a migraine attack has commenced, and/or as a prophylactic treatment administered, e.g., daily, weekly, biweekly, monthly, bimonthly, biannually, etc.) to prevent or reduce the frequency and/or severity of symptoms, e.g., pain symptoms, associated with migraine attacks.

Diagnostic Methods [0337] The antigén binding proteins deseribed herein can be used fór diagnostic purposes to detect, diagnose, or monitor diseases and/or conditions associated with CGRP R. Alsó provided are methods fór the detection ofthe presence of CGRP R in a sample using classical immunohistological methods known to those of skill in the art (e.g., Tijssen, 1993, Practice and Theory of Enzyme Immunoassays, Vol 15 (Eds R.H. Burdon and P.H. van Knippenberg, Elsevier, Amster79

EP 2 379 594 Β1 dam); Zola, 1987, Monoclonal Antibodies: A Manual of Techniques, pp. 147-158 (CRC Press, Inc.); Jalkanen et al., 1985, J. Cell. Bioi. 101:976-985; Jalkanen et al., 1987, J. Cell Bioi. 105:3087-3096). The detection of CGRP R can be performed in vivő or in vitro.

[0338] Diagnostic applications provided herein include use of the antigén binding proteins to detect expression of CGRP R and binding of the ligands to CGRP R. Examples of methods useful in the detection of the presence of CGRP R include immunoassays, such as the enzyme linked immunosorbent assay (ELISA) and the radioimmunoassay (RIA). [0339] Fordiagnostic applications, the antigén binding protein typically will be labeled with a detectable labeling group. Suitable labeling groups include, bút are nőt limited to, the following: radioisotopes or radionuclides (e.g., ³H, ¹⁴C, ¹⁵N, ³⁵S, ⁹⁰Y, Te, ¹¹¹ln, ¹²⁵l, ¹³¹l), fluorescent groups (e.g., FITC, rhodamine, lanthanide phosphors), enzymatic groups (e.g., horseradish peroxidase, β-galactosidase, luciferase, alkaline phosphatase), chemiluminescent groups, biotinyl groups, or predetermined polypeptide epitopes recognized by a secondary reporter (e.g., leucine zipper pair sequences, binding sites fór secondary antibodies, metál binding domains, epitope tags). In somé embodiments, the labeling group is coupled to the antigén binding protein via spacer arms of various lengths to reduce potential steric hindrance. Various methods fór labeling proteins are known in the art and may be used.

[0340] In another aspect, an antigén binding protein can be used to identify a cell or cells that express CGRP R. In a specific embodiment, the antigén binding protein is labeled with a labeling group and the binding ofthe labeled antigén binding protein to CGRP R is detected. In a further specific embodiment, the binding of the antigén binding protein to CGRP R detected in vivő. In a further specific embodiment, the CGRP R antigén binding protein is isolated and measured using techniques known in the art. See, fór example, Harlow and Lane, 1988, Antibodies: A Laboratory Manual, New York: Cold Spring Harbor (ed. 1991 and periodic supplements); John E. Coligan, ed., 1993, Current Protocols In Immunology New York: John Wiley &amp; Sons.

[0341] Another aspect provides fór detecting the presence of a test molecule that competes fór binding to CGRP R with the antigén binding proteins provided. An example of one such assay would involve detecting the amount of free antigén binding protein in a solution containing an amount of CGRP R in the presence or absence of the test molecule. An increase in the amount of free antigén binding protein (i.e., the antigén binding protein nőt bound to CGRP R) would indicate that the test molecule is capable of competing fór CGRP R binding with the antigén binding protein. In one embodiment, the antigén binding protein is labeled with a labeling group. Alternatively, the test molecule is labeled and the amount of free test molecule is monitored in the presence and absence of an antigén binding protein.

Methods of Treatment: Pharmaceutical Formulations, Routes of Administration [0342] Methods of using the antigén binding proteins are alsó illustrated. In somé methods, an antigén binding protein is provided to a patient. The antigén binding protein inhibits binding of CGRP to humán CGRP R.

[0343] Pharmaceutical compositions that comprise a therapeutically effective amount of one óra píuraíity ofthe antigén binding proteins and a pharmaceutically acceptable diluent, carrier, solubilizer, emulsifier, preservative, and/or adjuvant are alsó provided. In addition, methods of treating a patient, e.g., fór migraine, by administering such pharmaceutical composition are included. The term patient includes humán patients.

[0344] Acceptable formulation materials are nontoxic to recipients at the dosages and concentrations employed. In specific embodiments, pharmaceutical compositions comprising a therapeutically effective amount of humán CGRP R antigén binding proteins are provided.

[0345] In certain embodiments, acceptable formulation materials preferably are nontoxic to recipients at the dosages and concentrations employed. In certain embodiments, the pharmaceutical composition may contain formulation materials fór modifying, maintaining or preserving, fór example, the pH, osmolarity, viscosity, clarity, color, isotonicity, odor, sterility, stability, rate of dissolution or release, adsorption or penetration of the composition. In such embodiments, suitable formulation materials include, bút are nőt limited to, amino acids (such as glycine, glutamine, asparagine, arginine or lysine); antimicrobials; antioxidants (such as ascorbic acid, sodium sulfite or sodium hydrogen-sulfite); buffers (such as borate, bicarbonate, Tris-HCI, citrates, phosphates or other organic acids); bulking agents (such as mannitol or glycine); cheiating agents (such as ethylenediamine tetraacetic acid (EDTA)); complexing agents (such as caffeine, polyvinyl pyrrolidone, beta-cyclodextrinorhydroxypropyl-beta-cyclodextrin); fillers; monosaccharides; disaccharides; and other carbohydrates (such as glucose, mannose or dextrins); proteins (such as serum albumin, gelatin or immunoglobulins); coloring, flavoring and diluting agents; emulsifying agents; hydrophilic polymers (such as polyvinylpyrrolidone); low molecular weight polypeptides; salt-forming counterions (such as sodium); preservatives (such as benzalkonium chloride, benzoic acid, salicylic acid, thimerosal, phenethyl alcohol, methylparaben, propylparaben, chlorhexidine, sorbic acid or hydrogen peroxide); solvents (such as glycerin, propylene glycol or polyethylene glycol); sugár alcohols (such as mannitol or sorbitol); suspending agents; surfactants or wetting agents (such as pluronics, PEG, sorbitan esters, polysorbates such as polysorbate 20, polysorbate, triton, tromethamine, lecithin, cholesterol, tyloxapal); stability enhancing agents (such as sucrose or sorbitol); tonicity enhancing agents (such as alkali metál halides, preferably sodium or potassium chloride, mannitol sorbitol); delivery vehicles; diluents; excipients and/or pharmaceutical adjuvants. See,

ΕΡ 2 379 594 Β1

REMINGTON’S PHARMACEUTICAL SCIENCES, 18 Edition, (A.R. Genrmo, ed.), 1990, Mack Publishing Company. [0346] In certain embodiments, the optimál pharmaceutical composition will be determined by one skilled in the art depending upon, for example, the intended route of administration, delivery formát and desired dosage. See, for example, REMINGTON’S PHARMACEUTICAL SCIENCES, supra. In certain embodiments, such compositions may influence the physical state, stability, rate of in vivő release and rate of in vivő clearance of the antigén binding proteins disclosed. In certain embodiments, the primary vehicle or carrier in a pharmaceutical composition may be either aqueous or nonaqueous in natúré. For example, a suitable vehicle or carrier may be water for injection, physiological saline solution or artificial cerebrospinal fluid, possibly supplemented with other materials common in compositions for parenteral administration. Neutral buffered saline or saline mixed with serum albumin are further exemplary vehicles. In specific embodiments, pharmaceutical compositions comprise Tris buffer of about pH 7.0-8.5, or acetate buffer of about pH 4.0-5.5, and may further include sorbitol óra suitable substitute. In certain embodiments, humán CGRP R antigén binding protein compositions may be prepared for storage by mixing the selected composition having the desired degree of purity with optional formulation agents (REMINGTON’S PHARMACEUTICAL SCIENCES, supra) in the form ofa lyophilized cake or an aqueous solution. Further, in certain embodiments, the humán CGRP R antigén binding protein may be formulated as a lyophilizate using appropriate excipients such as sucrose.

[0347] The pharmaceutical compositions can be selected for parenteral delivery. Alternatively, the compositions may be selected for inhalation orfor delivery through the digestive tract, such as orally. Preparation of such pharmaceutically acceptable compositions is within the skill ofthe art.

[0348] The formulation components are present preferably in concentrations that are acceptable to the site of administration. In certain embodiments, buffers are used to maintain the composition at physiological pH or at a slíghtiy lower pH, typically within a pH rangé offrom about 5 to about 8.

[0349] When parenteral administration is contemplated, the therapeutic compositions may be provided in the form of a pyrogen-free, parenterally acceptable aqueous solution comprising the desired humán CGRP R binding protein in a pharmaceutically acceptable vehicle. A particularly suitable vehicle for parenteral injection is sterilé distilled water in which the humán CGRP R antigén binding protein is formulated as a sterilé, isotonic solution, properly preserved. In certain embodiments, the preparation can involve the formulation ofthe desired molecule with an agent, such as injectable microspheres, bio-erodible particles, polymeric compounds (such as polylactic acid or polyglycolic acid), beads or liposomes, that may provide controlled or sustained release ofthe product which can be delivered via depót injection. In certain embodiments, hyaluronic acid may alsó be used, having the effect of promoting sustained duration in the circulation. In certain embodiments, implantable drug delivery devices may be used to introduce the desired antigén binding protein.

[0350] Certain pharmaceutical compositions are formulated for inhalation. In somé embodiments, humán CGRP R antigén binding proteins are formulated as a dry, inhalable powder. In specific embodiments, humán CGRP R antigén binding protein inhalation Solutions may alsó be formulated with a propellant for aerosol delivery. In certain embodiments, Solutions may be nebulized. Pulmonary administration and formulation methods therefore are further described in International Patent Application No. PCT/US94/001875, which deseribes pulmonary delivery of chemically modified proteins. Somé formulations can be administered orally. Humán CGRP R antigén binding proteins that are administered in this fashion can be formulated with or without carriers customarily used in the compounding of solid dosage forms such as tablets and capsules. In certain embodiments, a capsule may be designed to release the active portion ofthe formulation atthe point in thegastrointestinal tract when bioavailability is maximized and pre-systemic degradation is minim ized. Additional agents can be included to facilitate absorption of the humán CGRP R antigén binding protein. Diluents, flavorings, low meiting point waxes, vegetable oils, lubricants, suspending agents, tablet disintegrating agents, and binders may alsó be employed.

[0351] Somé pharmaceutical compositions comprise an effective quantity of one or a plurality of humán CGRP R antigén binding proteins in a mixture with non-toxic excipients that are suitable for the manufacture of tablets. By dissolving the tablets in sterilé water, or another appropriate vehicle, Solutions may be prepared in unit-doseform. Suitable excipients include, bút are nőt limited to, inért diluents, such as calcium carbonate, sodium carbonate or bicarbonate, lactose, or calcium phosphate; or binding agents, such as starch, gelatin, or acacia; or lubricating agents such as magnesium stearate, stearic acid, or talc.

[0352] Additional pharmaceutical compositions will be evident to those skilled in the art, including formulations involving humán CGRP R antigén binding proteins in sustained- or controlled-delivery formulations. Techniques for formulating a variety of other sustained- or controlled-delivery means, such as liposome carriers, bio-erodible microparticles or porous beads and depót injections, are alsó known to those skilled in the art. See, for example, International Patent Application No. PCT/US93/00829, which and deseribes controlled release of porous polymeric microparticles for delivery of pharmaceutical compositions. Sustained-release preparations may include semipermeable polymer matrices in the form of shaped articles, e.g., films, or microcapsules. Sustained release matrices may include polyesters, hydrogels, polylactides (as disclosed in U.S. Patent No. 3,773,919 and European Patent Application Publication No. EP 058481), copolymers of L-glutamic acid and gamma ethyl-L-glutamate (Sidman et al., 1983, Biopolymers 2:547-556), poly (281

ΕΡ 2 379 594 Β1 hydroxyethyl-inethacrylate) (Langer etal., 1981, J. Biomed. Mater. Rés. 15:167-277 and Langer, 1982, Chem. Tech. 12:98-105), ethylene vinyl acetate (Langer et al., 1981, supra) or poly-D(-)-3-hydroxybutyric acid (European Patent Application Publication No. EP 133,988). Sustained release compositions may alsó include liposomes that can be prepared by any of several methods known in the art. See, e.g., Eppstein et al., 1985, Proc. Natl. Acad. Sci. U.S.A. 82:3688-3692; European Patent Application Publication Nos. EP 036,676; EP 088,046 and EP 143,949.

[0353] Pharmaceutical compositions used for in vivő administration are typically provided as sterilé preparations. Sterilization can be accomplished by filtration through sterilé filtration membranes. When the composition is lyophilized, sterilization using this method may be conducted either prior toorfollowing lyophilization and reconstitution. Compositions for parenteral administration can be stored in lyophilized form or in a solution. Parenteral compositions generally are placed intő a Container having a sterilé access port, for example, an intravenous solution bag or vial having a stopper pierceable by a hypodermic injection needle.

[0354] In certain embodiments, cells expressing a recombinant antigén binding protein as disclosed herein is encapsulated for delivery (see, Invest. Ophthalmol Vis Sci 43:3292-3298, 2002 and Proc. Natl. Acad. Sciences 103:3896-3901, 2006).

[0355] In certain formulations, an antigén binding protein has a concentration of at least 10 mg/ml, 20 mg/ml, 30 mg/ml, 40 mg/ml, 50 mg/ml, 60 mg/ml, 70 mg/ml, 80 mg/ml, 90 mg/ml, 100 mg/ ml or 150 mg/ml. Somé formulations contain a buffer, sucrose and polysorbate. An example ofa formulation is one containing 50-100 mg/ml of antigén binding protein,

5-20 mM sodium acetate, 5-10% w/v sucrose, and 0.002 - 0.008% w/v polysorbate. Certain, formulations, for instance, contain 65-75 mg/ml of an antigén binding protein in 9-11 mM sodium acetate buffer, 8-10% w/v sucrose, and 0.005-0.006% w/v polysorbate. The pH of certain such formulations is in the rangé of 4.5-6. Other formulations have a pH of 5.0-5.5 (e.g., pH of 5.0, 5.2 or 5.4).

[0356] Once the pharmaceutical composition has been formulated, it may be stored in sterilé vials as a solution, suspension, gél, emulsion, solid, crystal, or as a dehydrated or lyophilized powder. Such formulations may be stored either in a ready-to-use form or in a form (e.g., lyophilized) that is reconstituted prior to administration. Kits for producing a single-dose administration unit are alsó provided. Certain kits contain a first Container having a dried protein and a second Container having an aqueous formulation. In certain embodiments, kits containing single and multi-chambered pre-filled syringes (e.g., liquid syringes and lyosyringes) are provided. The therapeutically effective amountof a humán CGRP R antigén binding protein-containing pharmaceutical composition to be employed will depend, for example, upon the therapeutic context and objectives. One skilled in the art will appreciate that the appropriate dosage levels for treatment will vary depending, in part, upon the molecule delivered, the indication forwhich the humán CGRP R antigén binding protein is being used, the route of administration, and the size (body weight, body surface or organ size) and/or condition (the age and generál health) of the patient. In certain embodiments, the clinician may titer the dosage and modify the route of administration to obtain the optimál therapeutic effect.

[0357] A typical dosage may rangé from about 1 μg/kg to up to about 30 mg/kg or more, depending on the factors mentioned above. In specific embodiments, the dosage may rangé from 10 μg/kg up to about 30 mg/kg, optionally from 0.1 mg/kg up to about 30 mg/kg, alternatively from 0.3 mg/kg up to about 20 mg/kg. In somé applications, the dosage is from 0.5 mg/kg to 20 mg/kg. In somé instances, an antigén binding protein is dosed at 0.3 mg/kg, 0.5mg/kg, 1 mg/kg, 3 mg/kg, 10 mg/kg, or20 mg/kg. The dosage schedule in somé treatment regimes is at a dose of 0.3 mg/kg qW, 0.5mg/kg qW, 1 mg/kg qW, 3 mg/kg qW, 10 mg/kg qW, or20 mg/kg qW.

[0358] Dosing frequency will depend upon the pharmacokinetic parameters ofthe particular humán CGRP R antigén binding protein in the formulation used. Typically, a clinician administers the composition until a dosage is reached that achieves the desired effect. The composition may therefore be administered as a single dose, or as two or more doses (which may or may nőt contain the same amount of the desired molecule) over time, or as a continuous infusion via an implantation device or catheter. Appropriate dosages may be ascertained through use of appropriate dose-response data. In certain embodiments, the antigén binding proteins can be administered to patients throughout an extended time period. Chronic administration ofan antigén binding protein minimizes the adverse imm une or allergie response commonly associated with antigén binding proteins that are nőt fully humán, for example an antibody raised against a humán antigén in a non-human animal, for example, a non-fully humán antibody or non-human antibody produced in a nonhuman species.

[0359] The route of administration of the pharmaceutical composition is in accord with known methods, e.g., orally, through injection by intravenous, intraperitoneal, intracerebral (intra-parenchymal), intracerebroventricular, intramuscular, intra-ocular, intraarterial, intraportal, or intralesional routes; by sustained release systems or by implantation devices. In certain embodiments, the compositions may be administered by bolus injection or continuously by infusion, or by implantation device.

[0360] The composition alsó may be administered locally via implantation ofa membráné, sponge or another appropriate matéria! onto which the desired molecule has been absorbed or encapsulated. In certain embodiments, where an implantation device is used, the device may be implanted intő any suitable tissue or organ, and delivery ofthe desired molecule may be via diffusion, timed-release bolus, or continuous administration.

ΕΡ 2 379 594 Β1 [0361] It alsó may be desirable to use humán CGRP R antigén binding protein pharmaceutical compositions ex vivő. In such instances, cells, tissues or organs that have been removed from the patient are exposed to humán CGRP R antigén binding protein pharmaceutical compositions after which the cells, tissues and/or organs are subsequently implanted back intő the patient.

[0362] In particular, humán CGRP R antigén binding proteins can be delivered by implanting certain cells that have been genetically engineered, using methods such as those described herein, to express and secrete the polypeptide. In certain embodiments, such cells may be animal or humán cells, and may be autologous, heterologous, orxenogeneic. In certain embodiments, the cells may be immortalized. In other embodiments, in order to decrease the chance of an immunological response, the cells may be encapsulated to avoid infiltration of surrounding tissues. In further embodiments, the encapsulation materials are typically biocompatible, semi-permeable polymeric enclosures or membranes that allow the release ofthe protein product(s) bút prevent the destruction ofthe cells by the patient’s immuné system or by other detrimental factors from the surrounding tissues.

EXAMPLE 1

GENERATION OF CGRP RECEPTOR AS ANTIGENS

A. Molecular cloning of humán CRLR and RAMP1 [0363] Humán CRLR cDNA (Gén Bank Accession No. U17473; SEQ ID NO:1) and RAMP1 cDNA (Gén Bank Accession No. AJ001014; SEQ ID NO:3) were cloned intő the mammalian cell expression vectors pcDNA3.1-Zeo and pcDNA3.1Hyg (Invitrogen, Carlsbad, CA), respectively, fór transfections of HEK 293EBNA cells (Invitrogen) as described below. The hCRLR cDNA and hRAMPI cDNA were alsó cloned intő the pDSRa24 vector (Kim, Η. Y. et al. J. Inv. Derm. Symp. Proc. (2007) 12: 48-49) fór transfections of AM-1 CHO cells (U.S. Patent Number 6,210,924).

B. Stably-Transfected Cell Lines

1. Stable expression of humán CGRP R in 293EBNA cells [0364] HEK 293EBNA cells (available from ATCC or Invitrogen) were seeded at a density of 1.5x10^{* 6} cells per 100mm dish. After 24 hours, the cells were co-transfected with 6μg linearized DNAs of huRAMP1/pcDNA3.1-Hyg and huCRLR/pcDNA3.1-Zeo with FuGene6 (Invitrogen, Carlsbad, CA) following instructions supplied by Invitrogen. After two days, the cells were trypsinized and subcultured intő growth médium containing 400μg/ml hygromycin + 250μg/ml zeocin. After two weeks, the resulting drug resistant colonies were trypsinized and combined intő pools. The pools were subjected to four rounds of FACS sorting an Alexa 647-labeled CGRP₈_₃₇ peptide analóg (described below). The highest 5% of expressing cells were collected at each round.

2. Stable expression of humán CGRP R in AM-1 CHO cells [0365] AM-1 CHO cells (a serum-free growth media-adapted variant from the CHO DHFR-deficient cell line described in Urlaub and Chasin, Proc. Natl. Acad. Sci. 77, 4216 (1980), were seeded at 1.5x10⁶ cells per 100mm dish. After 24 hours, the cells were co-transfected with linearized 4 μg DNAs each of pDSRa24/huRAMP1 and pDSRa24/huCRLR with FuGene6 (Invitrogen, Carlsbad, CA) following instructions supplied by Invitrogen. The transfected cells were trypsinized 2 days after transfection and seeded intő CHO DHFR seleetive growth médium containing 10% dialyzed FBS and without hypoxanthine/thymidine supplement. After 2 weeks, the resulting transfected colonies were trypsinized and pooled. The pools were subjected to FACS sorting analysis.

3. Stable expression of humán adrenomedullin (AM1) in HEK 293EBNA cells [0366] 293EBNA cells were seeded in 100mm dishes at 1.5x10⁶ cells/dish in DMEM (high glucose) + 5% FBS + 1% MÉM non-essential amino acids + 1% sodium pyruvate. Thefollowing day the cells were co-transfected using FuGENE transfection reagent (Roche) with pcDNA3.1/zeocin/huCRLR plus pcDNA3.1/hygromycin/huRAMP2. Both DNA constructs were linearized with Fspl. After 48 hours the cells were subcultured intő 100mm dishes at 3 cell densities (8x10⁵, 3.2x10⁵, and 8x10⁴ cells/dish) in growth médium containing 200μg/ml zeocin. The médium was changed twice weekly. After one week the plates were fed with médium containing 20C^g/ml hygromycin + 20C^g/ml zeocin. After two weeks, 96 colonies were isolated with cloning rings. The remaining colonies were collected intő a single pool culture. The clones and pools were assayed fór their response to stimulation by receptor agonist orforskolin. Several clones showed a good response, and one was selected fór use in subsequent experiments.

ΕΡ 2 379 594 Β1

4. Stable expression of cyno CGRP R in HEK 293EBNA cells [0367] 293EBNA cells were seeded in 100mm dishes at 1.5x10⁶ cells/dish in DMEM (high glucose) + 5% FBS + 1% MÉM non-essential amino acids + 1% sodium pyruvate. The following day the cells were co-transfected using FuGENE 6 with pcDNA3.1/zeocin/cynoCRLR plus pcDNA3.1/hygromycin/cynoRAMP1. Both constructs were linearized with Fspl. After 48 hours the cells were subcultured intő growth médium containing 20C^g/ml zeocin + 40C^g/ml hygromycin at dilutions of 1:20, 1:40, 1:100, and 1:200. The médium was changed twice weekly. After two weeks, 96 transfected colonies were isolated using cloning rings. The clones were assayed fór their response to stimulation by CGRP ligand. Several clones showed similar high levels of response and one was selected fór use in subsequent experiments.

C. Isolation of high-expressing CGRP receptor cells [0368] A CGRP₈_₃₇ peptide analóg was synthesized (Midwest Bio-Tech Inc. Fishers, IN) with the sequence below:

Ac-WVTHRLAGLLSRSGGVVRCNFVPTDVGPFAF-_NH2 (SEQ ID NO:9) [0369] The peptide was labeled with Alexa 647-NHS following the manufacturer’s instructions (Molecular Probes, Inc. Cat A 2006). The Alexa 647-labeled CGRP₈_₃₇ showed specific staining of CGRP receptor transfected cells and nőt the non-transfected parental cells and was used as the FACS reagent.

[0370] The huCGRP receptor-transfected 293EBNA and AM-1 CHO cell pools (generated as above) were sorted repeatedly up to four times pools using with Alexa 647-labeled CGRP₈_₃₇ peptide. High expressing cells were collected at each sort, expanded and after the final sorting frozen intő vials. The AM-1 CHO/huCGRP R cells were used fór immunization as described below, and the 293EBNA/huCGRP R cells were used fór titering mouse sera after immunization and in binding sereens of the hybridoma supernatants.

D. Generation of soluble CGRP receptor [0371] Soluble CGRP receptor polypeptides containing the N-terminal extracellular domains (ECDs) of humán CRLR (SEQ ID NO:6) and humán RAMP1 (SEQ ID NO:8) were generated by transiently co-transfecting 293 6E cells (Durocher, et al., Nucleic Acids Rés. 30:E9 (2002)) with vectors containing the corresponding cDNAs (SEQ ID NO:5 or SEQ ID NO:7) as described below. Commonly used tags (polyHis, Flag, HA and/or Fc) were employed to facilitate secretion and/or subsequent purification.

[0372] A soluble heterodimeric CGRP R ECD fused to Fc was prepared by PCR cloning with the appropriate primers intő the transient expression vector pTT5 (Durocher, et al., supra). The CRLR N-terminal ECD-Fc consisted of the Nterminal extracellular domain of CRLR (SEQ ID NO:6) fused to humán lgG1 Fc. The RAMP1 ECD-Fc contains the extracellular domain of RAMP1 (SEQ ID NO:8) fused to humán IgG 1 Fc. In both cases, there was a linker consisting of five consecutive glycines between the ECD domain and Fc.

[0373] The soluble heterodimeric CGRP receptor was expressed by co-transfecting the two constructs as follows. 293-6E cells at 1x10⁶ cells/ml in shake flasks were transfected with 0.5mg/L DNA (hCRLR N-ter ECD-Fc/pTT5 and huRAMPI ECD-Fc/pTT5) with 3ml PEl/mg DNA in FreeStyle 293 média (Invitrogen). Cells were grown in suspension in FreeStyle 293 expression médium supplemented with 0.1% Pluronic F68 and 50 μg/ml Geneticin fór 7 days and harvested fór purification.

[0374] Purifications from conditioned média (CM) were performed by buffering the CM with the addition of 50mM Tris, 400mM sodium citrate, and adjusting the pH to 8.5. The buffered CM was then passed over a Protein A affinity column equilibrated in 50mM Tris, 400mM sodium citrate and pH adjusted to pH 8.5. The Protein A column was washed with PBS and the Fc fusion protein eluted with 0.1 N HOAc. The eluted peak contained both CRLR and RAMP1 components when tested by western biot using individual antibodies specific to either CRLR or RAMP1. Further LC-MS and N-terminal sequencing confirmed the presence of both CRLR:RAMP1 heterodimer and CRLR:CRLR homodimer in approximately (2:3) ratio. This soluble CGRP receptor was shown to compete in Alexa647 labeled CGRP₈_₃₇ binding to CGRP receptor expressing recombinant cells in the FMAT analysis, although it failed to bind CGRP ligand as determined using Biacore testing. The matériái was used as an immunogen as described in Example, despite, inter alia, its heterogeneity and lack of CGRP ligand binding.

E. Generation of membráné extracts from recombinant CGRP receptor expressing cells [0375] Membráné extracts were prepared from CGRP receptor expressing cells using a method described by Bőssé, R. et al., (Journal of Biomolecular Screening, 3(4): 285-292 (1998)). Briefly, approximately 5 grams of cell pásté were pelleted in 50 ml of PBS at 3,000 rpm fór 10 min at 4°C and re-suspended in 30 ml of cold lysis buffer (25 mM HEPES,

ΕΡ 2 379 594 Β1 pH 7.4, 3 mM MgCI₂ plus one Roche protease inhibitor cocktail tablet/ 50mL). The lysate was homogenized with GlasCol (Teflon-glass homogenizer) with ~20 strokes at 5,000 rpm and spun in a JA21 rotor at 20,000 rpm for 15 min at 4°C. This process was repeated once more and the final pellet was re-suspended ion ~1-5 ml ’final pellet’ buffer (25 mM HEPES, pH 7.4, 3 mM MgCI₂,10 % (w/v) sucrose plus one Roche protease inhibitor cocktail tablet/ 50mL). The membráné extracts were sheared by passing through 16 G and 25 G needles 2-3 times. Totál membráné protein concentration was determined with a Microplate BCA Protein Assay (Pierce).

EXAMPLE 2

GENERATION OF ANTIBODIES TO CGRP RECEPTOR

A. Immunization [0376] Immunizations were conducted using the following forms of CGRP receptor antigens, prepared as described in Example 1:

(i) AM-1 CHO transfectants expressing full length humán CRLR and RAMP1 at the cell surface, obtained by cotransfecting CHO cells with humán full length CRLR cDNA (SEQ ID NO:1) encoding a polypeptide having the sequence SEQ ID NO:2, and RAMP1 cDNA (SEQ ID NO:3) encoding a polypeptide having the sequence SEQ ID NO:4 (ii) membráné extract from the cells described in (i) above; and (iii) soluble CGRP receptor obtained by co-expressing and purifying the N-terminal ECD of CRLR (SEQ ID NO:6) and the extracellular domain (ECD) of RAMP1 (SEQ ID NO:8) as described in Example 1.

[0377] XENOMOUSE animals were immunized with purified soluble CGRP receptor protein and purified CGRP R membranes prepared from AM-1 CHO cells stably expressing CGRP R in the same manner using doses of 10 μg/mouse and 150 μg/mouse respectively. CGRP membranes were prepared using methods described above.

[0378] Subsequent boosts were administered at doses often μg/mouse of soluble CGRP Ror 75 μg of purified CGRP R membranes. XENOMOUSE animals were alsó immunized with CGRP receptor-expressing cells using doses of 3.4 χ 10⁶ CGRP R transfected cells/mouse and subsequent boosts were of 1.7 χ 10⁶ CGRP R transfected cells/mouse. Injection sites used were combinations of subcutaneous base-of-tail and intraperitoneal. Immunizations were performed in accordance with methods disclosed in U.S. Patent Number 7,064,244, filed February 19, 2002. Adjuvants TiterMax Gold (Sigma; cat. # T2684), Alum (E.M. Sergent Pulp and Chemical Co., Clifton, NJ, cat. # 1452-250) were prepared according to manufacturers’ instructions and mixed in a 1:1 ratio of adjuvant emulsion to antigén solution.

[0379] Sera were collected 4-6 weeks after the first injection and specific titers were determined by FACs staining of recombinant CGRP receptor-expressing 293EBNA cells.

[0380] Mice were immunized with either cells/membranes expressing full length CGRP R cells or soluble CGRP R extracellular domain, with a rangé of 11 - 17 immunizations over a period of approximately one to three and one-half months. Mice with the highest sera titer were identified and prepared for hybridoma generation. The immunizations were performed in groups of multiple mice, typically ten. Popliteal and inguinal lymph nodes and spleen tissues were typically pooled from each group for generating fusions.

B. Préparation of Monoclonal Antibodies [0381] Animals exhibiting suitable titers were identified, and lymphocytes were obtained from draining lymph nodes and, if necessary, pooled for each cohort. Lymphocytes were dissociated from lymphoid tissue in a suitable médium (for example, Dulbecco’s Modified Eagle Médium; DMEM; obtainable from Invitrogen, Carlsbad, CA) to release the cells from the tissues, and suspended in DMEM. B cells were selected and/or expanded using a suitable method, and fused with suitable fusion partner, for example, nonsecretory myeloma P3X63Ag8.653 cells (American Type Culture Collection CRL 1580; Kearney et al, J. Immunoi. 123, 1979, 1548-1550).

[0382] Lymphocytes were mixed with fusion partner cells at a ratio of 1:4. The cell mixture was gently pelleted by centrifugation at 400 x g for 4 minutes, the supernatant decanted, and the cell mixture gently mixed by using almi pipette. Fusion was indueed with PEG/DMSO (polyethylene glycol/dimethyl sulfoxide; obtained from Sigma-Aldrich, St. Louis MO; 1 ml per millión of lymphocytes). PEG/DMSO was slowly added with gentle agitation over one minute followed, by one minute of mixing. IDMEM (DMEM without glutamine; 2 ml per millión of B cells), was then added over 2 minutes with gentle agitation, followed by additional IDMEM (8 ml per millión B-cells) which was added over 3 minutes.

[0383] The fused cells were gently pelleted (400 xg 6 minutes) and resuspended in 20 ml Selection média (for example, DMEM containing Azaserine and Hypoxanthine [HA] and other supplemental materials as necessary) per millión B-cells.

ΕΡ 2 379 594 Β1

Cells were incubated fór 20-30 minutes at 37°C and then resuspended in 200 ml Selection média and cultured fór three to four days in T175 flasks prior to 96-well plating.

[0384] Cells were distributed intő 96-well plates using standard techniques to maximize clonality of the resulting colonies. After several days of culture, the hybridoma supernatants were collected and subjected to screening assays as detailed in the examples below, including confirmation of binding to humán CGRP receptor, identification of blocking antibodies by a ligand binding competition assay and evaluation of cross-reactivity with other receptors related to CGRP receptor (fór example, humán Adrenomedullin receptor). Positive cells were further selected and subjected to standard cloning and subcloning techniques. Clonal lines were expanded in vitro, and the secreted humán antibodies obtained fór analysis.

C. Sequence Analysis of Selected Monoclonal Antibodies [0385] Selected subcloned monoclonal antibodies were sequenced using standard RT-PCR methods. Table 2A shows the amino acid sequences ofthe light chains of exemplary antibodies disclosed herein. Table 2B shows the amino acid sequences ofthe heavy chains of exemplary antibodies disclosed herein.

[0386] Amino acid sequences corresponding to CDR regions of sequenced antibodies were aligned and the alignments were used to group the clones by similarity.

[0387] Sequence alignments of light chain CDRs from clones having kappa light chains, and certain corresponding consensus sequences, are shown in Figs 3A and 3B.

[0388] Sequence alignments of light chain CDRs from clones having lambda light chains, and certain corresponding consensus sequences, are shown in Fig. 4.

[0389] Sequencealignmentsofheavy chain CDRs of exemplary antibodiesdisclosed herein, and certain corresponding consensus sequences, are shown in Figs. 5A, 5B, 5C, 5D and 5E.

[0390] Certain consensus sequences of exemplary heavy chain CDRs disclosed herein are shown in Fig. 5F.

EXAMPLE 3

IDENTIFICATION OF CGRP RECEPTOR SPECIFIC ANTIBODIES

A. Selection of CGRP receptor specific binding antibodies by FMAT [0391] After 14 days of culture, hybridoma supernatants were sereened fór CGRP R-specific monoclonal antibodies by Fluorometric Microvolume Assay Technology (FMAT) (Applied Biosystems, Foster City, CA). The supernatants were sereened against either the AM-1 CHO huCGRP R cells or recombinant HEK 293 cells that were transfected with humán CGRP R and counter-screened against parental HEK293 cells (prepared as described in Example 1).

[0392] Briefly, the cells in Freestyle média (Invitrogen, Carlsbad, CA) were seeded intő 384-well FMAT plates in a volume of 50 jj_L/well at a density of approximately 4000 cells/well fór the stable transfectants, and at a density of approximately 16,000 cells/well fór the parental cells, and cells were incubated overnight at 37°C. Then, 10 jj_L/well of supernatant was added and plates were incubated fór approximately one hour at 4°C, after which 10 jj_L/well of antihuman lgG-Cy5 secondary antibody (Jackson Immunoresearch, West Grove, PA) was added at a concentration of 2.8 μg/ml (400ng/ml final concentration). Plates were then incubated fór one hour at 4°C, and fluorescence was read using an FMAT macroconfocal scanner (Applied Biosystems, Foster City, CA).

[0393] Fór counter sereens, the parental AM-1 CHO cells or HEK 293 cells were seeded similarly and supernatants sereened by FMAT on these cells in parallel to differentiate and eliminate hybridomas binding to cellular proteins, bút nőt to the CGRP receptor.

B. Identification of blocking antibodies by ligand binding competition assay through FMAT [0394] A ligand binding competition method was developed to identify antibodies (in the hybridoma supernatants) that bind CGRP receptor and block CGRP ligand binding. 384-wells plates (Corning Costar, Cat:#3712) were prepared with 5,000 AM-1 huCGRP R Pool 2 cells and 20,000 untransfected CHO-S cells in each well. 20μΙ of anti-CGRP R hybridoma supernatant were added to each well, and the plates were incubated fór 1 hr at room temperature. 10μΙ of 2.8pg/ml Alexa647-CGRP₈_₃₇ peptide were then added to each well and the plates were incubated fór a further 3 hours at room temperature. The amount of Alexa647-CGRP₈_₃₇ bound to the cells was assayed on a FMAT 8200 Cellular Detection System (Applied Biosystems). Output data were both a numerical FL1 value of signal intensity (higher FL1 values indicate higher signal intensity) and alsó an image of the cells.

[0395] The experiments included negative control hybridoma supernatants. The average FL1 value observed in these negative control experiments was adopted as the maximum possibie signal fór the assay. Experimental supernatants

ΕΡ 2 379 594 Β1 were compared to this maximum signal and a percent inhibition was calculated fór each well (% Inhibition = (1-(FL1 of the anti-CGRP R hybridoma supernatant/Maximum FL1 signal)).

[0396] An overview of the data is shown in Fig. 6. In this experiment, 1092 anti-CGRP R supernatants were tested using the receptor ligand assay. The data were ránk ordered using the average percent inhibition. Ninety supernatants had >25% average inhibition, 31 of these were >50% and 7 were >70% average inhibition.

[0397] An abbreviated data set is shown in Table 10, below. Sample ID Nos. 1-5 illustrate examples of anti-CGRP R hybridoma supernatants which inhibited the binding Alexa647-CGRP₈_₃₇ peptide to CGRP receptor and Sample ID Nos 536 - 540 illustrate examples of anti-CGRP R hybridoma supernatants which did nőt inhibit the binding ofthe Alexa647CGRP₈_₃₇ peptide to the CGRP receptor.

Table 10 - Exemplary data from FMAT ligand binding competition assay

<td> Sample</td><td colspan="4"> Expt. #1</td><td colspan="3"> Expt. #2</td>

<td> ID</td><td> FL1</td><td colspan="2"> Image</td><td> % Inhibition</td><td> FL1</td><td> Image</td><td> % Inhibition</td>

<td> 1</td><td> 2264</td><td></td><td></td><td> 84%</td><td> 2585</td><td></td><td> 88%</td>

<td> 2</td><td> 3007</td><td> H</td><td></td><td> 77%</td><td> 2804</td><td> E3</td><td> 85%</td>

<td> 3</td><td> 3460</td><td></td><td></td><td> 72%</td><td> 2929</td><td></td><td> 84%</td>

<td> 4</td><td> 3650</td><td> il</td><td></td><td> 70%</td><td> 3294</td><td></td><td> 79%</td>

<td> 5</td><td> 3764</td><td> löl</td><td></td><td> 69%</td><td> 3246</td><td></td><td> 80%</td>

<td> 536</td><td> 10412</td><td> n</td><td></td><td> 0%</td><td> 11142</td><td> n</td><td> -17%</td>

<td> 537</td><td> 10413</td><td> ü</td><td></td><td> 0%</td><td> 9388</td><td></td><td> 5%</td>

<td> 538</td><td> 10414</td><td> m</td><td></td><td> 0%</td><td> 9420</td><td></td><td> 4%</td>

<td> 539</td><td> 10415</td><td> n</td><td></td><td> 0%</td><td> 10943</td><td></td><td> -14%</td>

<td> 540</td><td> 10415</td><td colspan="2"> n</td><td> 0%</td><td> 10561</td><td> H</td><td> -10%</td>

ΕΡ 2 379 594 Β1 [0398] Based on the binding competition assays, approximately 30 supernatants were selected for further characterization.

EXAMPLE 4

ACTIVITY OF CGRP RECEPTOR SPECIFIC BLOCKING MONOCLONAL ANTIBODIES IN A cAMP FUNCTIONAL ASSAY

A. CGRP receptor antibody activity.

[0399] Selected CGRP receptor antibodies were screened in an in vitro CGRP receptor mediated cAMP assay to determine intrinsic potency. The in vitro cAMP assay employed a humán neuroblastoma-derived cell line (SK-N-MC; Spengler, et al., (1973) In Vitro 8: 410) obtained from ATCC (ATCC Number HTB-10; HTB-10 cells). HTB-10 cells express CRLR and RAMP1, which form CGRP receptor (L. M. McLatchie etal, 1998). A 293EBNA cell line expressing recombinant cynomolgus CGRP R was generated as described in Example 1, and a rat L6 cell line expressing rat CGRP receptor was obtained from the ATCC (CRL-1458).

[0400] The LÁNCÉ cAMP assay kit (PerkinElmer, Boston, MA) was used in the screening. The assays were performed in white 96-well plates in a totál volume of 60 μι. Briefly, on the day ofthe assay, the frozen HTB-10 cells were thawed at 37°C, cells were washed once with assay buffer and 12 μι of cell suspension containing 10000 cells mixed with Alexa-labeled anti-cAMP antibody was added intő 96 half-area white plates. After adding 12μί CGRP receptor antibody, the mixture was incubated for 30 min at room temperature. Then 12μί CGRP receptor agonist humán α-CGRP (1nM final concentration) was added and further incubated for 15 min at room temperature. After humán α-CGRP stimulation, 24 μι of detection mix was added and incubated for 60 minutes at room temperature and the plates were red on EnVision instrument (PerkinElmer, Boston, MA) at Em665nM. Data were processed and analyzed by Prizm (GraphPad Software Inc.) or ActivityBase (IDBS).

[0401] Fig. 7A shows exemplary data obtained as described above using the hCGRP receptor-expressing cell line HTB-10 for three antibodies - 3C8,13H2 and 1E11. The data are plotted as percentage over control (POC) as a function of antibody (3C8, 13H2 or 1E11) concentration, and are fitted with standard nonlinear regression curves to yield the IC50 vaíues shown at the bottom of the figure.

B. Lack of antibody activity in related receptors.

[0402] Cells expressing related receptors AM1 (HEK 293 cells expressing hCRLR+hRAMP2; D. R. Poyner, et al, Pharmacological review, 54:233-246, 2002), AM2 (CHO cells expressing hCRLR+hRAMP3; D. R. Poyner, et al, Pharmacological review, 54:233-246, 2002) or humán amylin AMY1 receptor (MCF-7 cells hCTR+hRAMP1; Wen-Ji Chen, et al, Molecular pharmacology, 52: 1164-1175, 1997) were used to determine the selectivity ofthe tested antibodies. The AM1-expressing HEK 293 cell line was generated as described in Example 1, above. The AM2-expressing CHO cell line was purchased from EuroScreen (now PerkinElmer, Inc.); and the humán amylin AMY1 receptor-expressing MCF-7 cell line (Zimmermann, et al, Journal of Endocrinology, 423-431,1997), was obtained from the ATCC (HTB-22). Exemplary results, plotted as described above, are shown in Figs. 7B (hAM1-HEK cells), 7C (hAM2-CHO cells) and 7D (hAMY-MCF-7 cells). Note that nőne ofthe tested antibodies had significant inhibitory activity against hAM1, hAM2 or hAMY1 receptors over the rangé tested.

[0403] Similar experiments were performed using recombinant HEK cells expressing cynomolgus CGRP receptors and rat L6 cells expressing rat CGRP receptor (ATCC). Data from these studies, as well as additional IC50 data obtained as described in part A of this Example, are shown in the cAMP columns in Table 11, below. Note that the IC50 vaíues against the humán and cyno CGRP receptors are in the nanomolar rangé, whereas activities against rat CGRP receptor, and humán AM1, AM2 and AMY1 receptors, as well as MCF7 cells expressing calcitonin (data nőt shown) are all greater than 1 micromolar. The difference in IC50 between humán CGRP receptor and humán AM1, AM2, amylin and calcitonin receptors illustrates the high selectivity of the these antibodies for the CGRP receptor over related receptors formed in part ofthe same receptor components. IC50 obtained using humán and cynomolgus CGRP receptors were similar, whereas the tested antibodies did nőt appear to cross-react with rat CGRP receptor.

EP 2 379 594 Β1

Table 11

<td rowspan="2"> Clone</td><td colspan="6"> cAMP assay</td><td>¹²⁵l assay</td>

<td> hCGRPR IC50 (nM)</td><td> Cyno CGRPR IC50 (nM)</td><td> Rat CGRPR IC50 (nM)</td><td> hAmylin 1 IC50 (nM)</td><td> hAM1 IC50 (nM)</td><td> hAM2IC50 (nM)</td><td> Humán CGRP Ki (nM)</td>

<td> 01E11.2</td><td> 1.77</td><td> 2.79</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.030</td>

<td> 01H7.2</td><td> 3.27</td><td> 4.74</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.079</td>

<td> 02A10.1</td><td> 11.81</td><td> 17.6</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.291</td>

<td> 02E7.2</td><td> 6.30</td><td> 5.51</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.117</td>

<td> 03A5.1</td><td> 9.89</td><td> 28.9</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.093</td>

<td> 03B6.2</td><td> 2.74</td><td> 2.22</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.033</td>

<td> 03C8.2</td><td> 6.66</td><td> 5.32</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.044</td>

<td> 03H8.2</td><td> 10.84</td><td> 10.6</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.111</td>

<td> 04E4.2</td><td> 2.38</td><td> 3.52</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.015</td>

<td> 04H6.1</td><td> 3.78</td><td> 5.59</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.052</td>

<td> 05F5.1</td><td> 4.79</td><td> 4.78</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.147</td>

<td> 07B2.1</td><td> 8.96</td><td> 27.7</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.116</td>

<td> 07B3.1</td><td> 10.2</td><td> 14.1</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.127</td>

<td> 07F1.1</td><td> 8.92</td><td> 10.5</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.140</td>

<td> 08B11.2</td><td> 10.7</td><td> 17.0</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.118</td>

<td> 09D4.2</td><td> 1.40</td><td> 2.46</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.023</td>

<td> 09F5.2</td><td> 3.06</td><td> 4.44</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.043</td>

<td> 10E4.2</td><td> 3.08</td><td> 3.23</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.100</td>

<td> 11A9.1</td><td> 16.1</td><td> 47.8</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.157</td>

<td> 11D11.1</td><td> 4.93</td><td> 3.85</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.044</td>

<td> 11H9.1</td><td> 4.56</td><td> 5.07</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.057</td>

<td> 12E8.2</td><td> 2.93</td><td> 4.13</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.097</td>

<td> 12G8.2</td><td> 2.14</td><td> 2.74</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.017</td>

<td> 13D6.2</td><td> 8.23</td><td> 11.8</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.055</td>

<td> 13E2.2</td><td> 18.3</td><td> 49.2</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.128</td>

<td> 13H2.2</td><td> 1.95</td><td> 8.41</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td> 0.033</td>

<td> 32H7.1G</td><td></td><td> 1.93</td><td></td><td> &gt;1000</td><td> &gt;1000</td><td> &gt;1000</td><td></td>

EXAMPLE 5

RADIOLIGAND CGRP BINDING ASSAY FOR Ki DETERMINATION RECEPTOR BLOCKING ANTIBODIES [0404] ¹²⁵l-labeled CGRP (Amersham Biosciences, Piscataway, NJ) and cell membranes from HTB-10 cells (PerkinElmer Inc., Waltham, Massachusetts) were used for radioligand binding experiment in the presence of various concentrations ofthe test antibodies to determine the corresponding Ki values. The CGRP binding assay was set up at room temperature in 96-well plates containing: 110 μΙ binding buffer (20 mM Tris-HCI, pH7.5, 5.0 mM MgSO4, 0.2% BSA (Sigma), 1 tablet of CompleteTM/50 ml buffer (a protease inhibitor)); 20 test compound (10X); 20 μΙ ¹²⁵l-haCGRP (Amersham Biosciences; 10X); and 50 μΙ humán neuroblastoma cell (HTB-10) membráné suspension (10 μg perwell,

ΕΡ 2 379 594 Β1

PerkinElmer). The plates were incubated at room temperature fór 2 hours with shaking at 60 rpm, and then the contents of each well were filtered over 0.5% polyethyleneimine (PEI)-treated (fór at least one hour) GF/C 96-well filter plates. The GF/C filter plates were washed six times with ice-cold 50 mM Tris, pH 7.5 and dried in an oven at 55°C fór 1 hour. The bottoms ofthe GF/C plates were then sealed. 40 μΙ Microscint™ 20 was added to each well, the tops ofthe GF/C plates were sealed with TopSeal™-A(a press-on adhesive sealing film), and the GF/C plateswere counted with TopCount NXT (Packard). The data were analyzed using Prizm (GraphPad Software Inc.) [0405] Exemplary data and Ki values obtained using antibodies 3C8, 12H2 and 1E11 are shown in Fig. 8.

[0406] The right-most column in Table 11, above, lists the Ki values ofthe indicated mAbs in the radiolabeled ¹²⁵lCGRP competition binding assay to HTB-10 cell membranes. The data demonstrate that the CGRP receptor antibodies were highly competitive (all sub-nanomolar rangé) against CGRP binding.

EXAMPLE 6

FACS BINDING ASSAY FÓR KD DETERMINATION OF CGRP RECEPTOR BLOCKING ANTIBODIES [0407] The affinities of anti-CGRP R mAbs fór CGRP receptors expressed on cells were determined using a FACS method. Briefly, AM-1 CHO huCGRP R-expressing cells, prepared as described above, were plated in 96-well plates at densities of 16,000 or 160,000 cells per well in DMEM médium containing 10%FBS, NEAA, PS, L Glut, NaPyr and 0.05% sodium azide. CGRP receptor antibodies were titrated in the same médium from 50 nM to 1 pM and incubated with cells. After an overnight incubation at 4°C in a totál volume of 120 μΙ, on a plate shaker, the cells were washed 2X with PBS+2%FBS, centrifuging and discarding supernatant each time. 100 μΙ/well of G anti-Hu FcCy5 (5μg/mL; Jackson ImmunoResearch Laboratories Inc., West Grove, PA, USA) containing 7AAD ^jJ/well)was then added and incubated at 4°C fór 40 min. The cells were washed 2X with PBS+2%FBS, centrifuging and discarding the supernatant each time. 100 μΙ PBS+2%FBS buffer was then added and analyzed by FACS to determined the binding geomean. The Kd was calculated using KinExA software by taking the negative geomean at each antibody concentration as the amount of free Ab present Rathanaswami, et al., Biochemical and Biophysical Research Communications 334 (2005) 1004-1013. The data obtained at the two different cell concentrations were analyzed by n-curve analysis to determine the Kd and the 95% confidence interval as described in Rathanaswami, et al., Biochemical and Biophysical Research Communications 334 (2005) 1004-1013.

[0408] Exemplary data with corresponding curve fits are shown in Fig. 10 fór antibody 12G8.2. The data of eight blocking antibodies generated in support of the present disclosure are shown in Table 12. One of the antibodies (3B6) was analyzed on two different days. The ratio of 0.9 obtained fór the experiment with 16K cells indicates that the antigén concentration is predicted as 0.9X the Kd and hence the curve obtained by this experiment is a Kd controlled curve. It can be appreciated that the Kd values obtained in this manner were in the low single-digit nanomolar rangé fór all tested antibodies.

Table 12

<td></td><td colspan="5"> N curve analysis</td>

<td></td><td> Kd(nM)</td><td> Kd Low(nM)</td><td> Kd High (nM)</td><td> % error</td><td> Rádió 16K</td>

<td> 1H7</td><td> 1.9</td><td> 1.5</td><td> 3</td><td> 3.8</td><td> 0.001</td>

<td> 2E7</td><td> 1.5</td><td> 0.7</td><td> 3.4</td><td> 6.3</td><td> 0.19</td>

<td> 3B6 (a)</td><td> 1.7</td><td> 1.1</td><td> 2.7</td><td> 5.3</td><td> 0.060</td>

<td> 3B6 (b)</td><td> 2.0</td><td> 1.6</td><td> 2.6</td><td> 3.2</td><td> 0.21</td>

<td> 4E4</td><td> 1.3</td><td> 0.9</td><td> 2.05</td><td> 3.9</td><td> 0.16</td>

<td> 4H6</td><td> 2.4</td><td> 1.78</td><td> 4.35</td><td> 3.8</td><td> 0.070</td>

<td> 9D4</td><td> 2.5</td><td> 1.8</td><td> 4.39</td><td> 4.3</td><td> 0.060</td>

<td> 12E8</td><td> 2.3</td><td> 1.58</td><td> 3.36</td><td> 3.7</td><td> 0.55</td>

<td> 12G8</td><td> 1.4</td><td> 0.92</td><td> 2.21</td><td> 3.6</td><td> 0.94</td>

ΕΡ 2 379 594 Β1

EXAMPLE 7

BINNING OF CGRP RECEPTOR BLOCKING ANTIBODIES BY BICORE BINDING COMPETITION [0409] Biacore analyses (Karlsson, R. et al., Methods; A Companion to Methods in Enzymology, 6: 99-110 (1994) were carried out as follows. Immobilization of anti-CGRP receptor antibodies to the CM5 sensor chip surface was performed according to manufacturer’s instructions, using a continuous flow of 10mM HEPES, 0.15M NaCl, 3.4mM EDTA, 0.005% P-20, pH 7.4 (HBS-EP buffer). Carboxyl groups on the sensor chip surfaces were activated by injecting 60 μι ofa mixture containing 0.2 M N-ethyl-N’ (dimethylaminopropyl)carbodiimide (EDC) and 0.05 M N-hydroxysuccinimide (NHS). Specific surfaces were obtained by injecting 180 μΙ of anti-CGRP receptor antibody diluted in 10 mM acetate, pH 4.0 at a concentration of 30 μg/mL. Excess reactive groups on the surfaces were deactivated by injecting 60 μι of 1 M ethanolamine. Final immobilized levels fór the individual antibodies were as follows:

<td> Antibody</td><td> Resonance Units (RU)</td>

<td> 11D11</td><td> -5,900</td>

<td> 3B6</td><td> -7,200</td>

<td> 4H6</td><td> -8,000</td>

<td> 12G8</td><td> -7,800</td>

<td> 9F5</td><td> -6,600</td>

<td> 34E3</td><td> -3,700</td>

A blank, mock-coupled reference surface was alsó prepared on the sensor chip. Soluble huCGRP receptor at a concentration of 100nM was captured on sensor chips having one of the six immobilized antibodies referenced above (11D11,3B6, 4H6, 12G8, 9F5 or 34E3). Each ofthe 20 test anti-CGRP R antibodies was then injected over the captured huCGRP receptor. If the injected antibody recognized a distinct epitope relatíve to that recognized by the immobilized antibody, a second binding event would be observed. If the antibodies recognize the same or very similar epitopes, only the binding ofthe huCGRP receptor would be observed.

[0410] Exemplary data obtained using a sensor chip coated with immobilized antibody 3B6 are shown in Fig. 9A. The four traces are data obtained using antibodies 1E11, 4E4, 2E7 and 12G8 in the injected solution. Events during the experiment are represented by letters, with A corresponding to injection of huCGRP R-Fc, B corresponding to end of the huCGRP R-Fc injection, C corresponding to injection of second mAb, and D corresponding to end second mAb injection and start of the buffer wash. Note that there is no indication of any binding signal from any of the injected antibody on the immobilized antibody surface, indicating that the four injected antibodies apparently recognize the same or very similar epitope(s) as the immobilized antibody. Essentially the same results were observed with all tested blocking antibodies washed over each the five immobilized neutralizing antibody surfaces, indicating that all tested anti-huCGRP receptor blocking antibodies recognize the same or very similar and strongly overlapping epitope(s).

[0411] In contrast, as shown in part in Figs. 9B, 9C and 9D, the four tested non-blocking, CGRP receptor specific antibodies 32H8, 33B5, 33E4 and 34E3 failed to compete with 11D11 (data nőt shown), 3B6 (Fig. 9B), 12G8 (Fig. 9C) and 9F5 (data nőt shown) although 34E3 was able to compete with 4H6 (Fig. 9D) and weakly with 32H7 (data nőt shown). 32H8failed to compete with 3B6, 4H6,12G8, 9F5 orthe non-blocking antibody 34E3, bút 33B5 and 33E4 could compete with the non-blocking antibody 34E3. The data fór all blocking and non-blocking antibodies are summarized in Table 13, below. NB indicates no binding; + indicates significant binding; and Weak indicates weak binding.

Table 13

<td></td><td colspan="6"> Immobilized Antibodies</td>

<td> Ab in Solution</td><td> 11D11</td><td> 3B6</td><td> 4H6</td><td> 12G8</td><td> 9F5</td><td> 34E3</td>

<td> 1E11</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> +</td>

<td> 1H7</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> +</td>

<td> 2E7</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> +</td>

<td> 3B6</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> +</td>

<td> 3C8</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> +</td>

<td> 4E4</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> +</td>

ΕΡ 2 379 594 Β1 (continued)

<td></td><td colspan="6"> Immobilized Antibodies</td>

<td> Ab in Solution</td><td> 11D11</td><td> 3B6</td><td> 4H6</td><td> 12G8</td><td> 9F5</td><td> 34E3</td>

<td> 4H6</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td>

<td> 5F5</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> +</td>

<td> 9D4</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> +</td>

<td> 9F5</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> +</td>

<td> 10E4</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> +</td>

<td> 11D11</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> +</td>

<td> 11H9</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> +</td>

<td> 12E8</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> +</td>

<td> 12G8</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> +</td>

<td> 13H2</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> +</td>

<td> 32H7</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> NB</td><td> Weak</td>

<td> 32H8</td><td> +</td><td> +</td><td> +</td><td> +</td><td> +</td><td> +</td>

<td> 33B5</td><td> +</td><td> +</td><td> +</td><td> +</td><td> +</td><td> NB</td>

<td> 33E4</td><td> +</td><td> +</td><td> +</td><td> +</td><td> +</td><td> NB</td>

[0412] As can be appreciated from the data, all the tested blocking or neutralizing antibodies bind to the same region as the five immobilized blocking antibodies; i.e., all of the tested neutralizing antibodies bind the same region of the CGRP R molecule. On the other hand, the non-blocking antibodies did nőt generally compete with the immobilized blocking antibodies, indicating that the non-blocking antibodies primarily bind a different region of CGRP R.

EXAMPLE 8

BINDING OF CGRP RECEPTOR ANTIBODIES TO SOLUBLE CGRP RECEPTOR IN WESTERN BLOT [0413] Three representative CGRP receptor blocking antibodies were tested using Western blots for binding to a soluble CGRP receptor-muFc fusion protein.

[0414] 100ng of purified CGRP R-muFc (produced and purified as described above for the CGRP R-huFc except the mouse Fcwas used and the linker between RAMP1 orCRLR ECD and muFcwas changed to GGGGGVDGGGGGV (SEQ ID NO:213)) was diluted in PBS with PAGE sample buffer with (reduced) or without (non-reduced) beta-mercaptoethanol (βΜΕ) at 13.3% concentration. The sample containing βΜΕ was then boiled for 4 min. Reduced and nonreduced samples were loaded onto separate 4-20% Tris-glycine gels (Invitrogen) with alternating lanes of CGRP R-Fc protein and molecular weight markers (Invitrogen). Gels were electroblotted onto 0.2μΐτι nitrocellulose filters (Invitrogen). The blots were washed with Tris-buffered saline + 1% Tween 20 (TBST) and then blocked with TBST + 5% powered dry milk for 30min. The blots were cut intő strips along the molecular weight marker lanes. One strip each with reduced and non-reduced CGRP R-muFc were incubated with purified huCGRP R antibodies 4E4, 9F5, or 3B6 (1:500 dilution in TBST + 5% milk), goat anti-huRAMP1 N-20 (1:500; Santa Cruz Biotechnology, Inc), rabbit anti-mouse IgG-Fc-HRP (1:10,000) (Pierce), or goat anti-human IgG-Fc-HRP (1:10,000) (Pierce). Blots were incubated with the antibodies for one hour followed by 3x10min washes with TBST + 1 % milk. The blots treated with the huCGRP R antibodies were then incubated with goat anti-mouse IgG-Fc-HRP (1:10,000 in TBST + 1 % milk) and the blots treated with anti-huRAMP1 (N-20 anti-RAMP1 goat polyclonal antibody, Santa Cruz Biotech, CA) were incubated with rabbit anti-goat IgG-Fc-HRP (1:10,000) for 20min. Blots were washed 3x15min with TBST. The huCGRP R and anti-huRAMP1 antibody blots were treated with Pierce Supersignal West Pico Detection reagent, and the anti-mouse and anti-human IgG-Fc-HRP blots were treated with Pierce standard Detection Reagent (1 min.). Blots were then exposed with Kodak Biomax MS X-ray film. [0415] All of the three CGRP receptor antibodies, 4E4, 9F5 and 3B6 were able to detect the soluble CGRP R-muFc (containing RAMP1-ECD and CRLR ECD) under non-reduced condition bút nőt under reduced condition indicating that the binding epitope of these CGRP R antibodies was conformational and sensitive to the disulfide linkages (3 pairs in RAMP1-ECD and 3 pairs in CRLR N-ter ECD). In contrast, the commercial anti-RAMP1 antibody N-20 (Santa Cruz

ΕΡ 2 379 594 Β1

Biotech) bound RAMP1 under both reduced and no reduced conditions indicating that the binding site fór N-20 antibody was primarily linear and nőt sensitive to disulfide linkages.

EXAMPLE 9

BINDING OF CGRP RECEPTOR BLOCKING ANTIBODIES TO CHIMERIC RECEPTORS [0416] CGRP receptors formed of either native RAMP1 with chimeric CRLR, or native CRLR with chimeric RAMP1, were used to identify CGRP receptor sequences involved in antibody binding. Since all ofthe humán CGRP receptor blocking antibodies tested failed to show functional activity to the rat CGRP receptor, the chimeric components contained regions of rat sequence in a humán sequence background. The following chimeras were generated fór binding analysis by FACS:

RAMP1 chimera#1 (Q28 to A34): SEQ ID NO:217 [0417] Amino acid residues Q28 to A34 in the humán RAMP1 were replaced with the corresponding sequences from rat RAMP1. This stretch included five amino acid residues that are different between humán and rat RAMP1.

RAMP1 chimera#2 (Q43 to E53): SEQ ID NO:218 [0418] Amino acid residues Q43 to E53 in the humán RAMP1 were replaced with the corresponding sequences from rat RAMP1. This stretch included six amino acid residues that are different between humán and rat RAMP 1.

RAMP1 chimera#3 (R67 to E78): SEQ ID NO:219 [0419] Amino acid residues R67 to E78 in the humán RAMP were replaced with the corresponding sequences from rat RAMP1. This stretch included seven amino acid residues that are different between humán and rat RAMP1.

CRLR chimera#1 (L24 to Q33); SEQ ID NO:223 [0420] Amino acid residues L24 to Q33 in the humán CRLR were replaced with the corresponding sequences from rat CRLR. This stretch included eight amino acid residues that are different between humán and rat CRLR.

[0421] Fig. 11 shows an alignment of RAMP1 amino acid sequences from cynomolgus monkey (SEQ ID NO:215), humán (SEQ ID NO:4), rat (SEQ ID NO:214) and rhesus monkey (SEQ ID NO:216), togetherwith sequences of RAMP1 chimera #1 (SEQ ID NO:217),chimera #2 (SEQ ID NO:218)and chimera#3 (SEQ ID NO:219). Figs. 12Aand 12Bshow an alignment of CRLR amino acid sequences from humán (SEQ ID NO:2), cynomolgus monkey (SEQ ID NO:221), rhesus monkey (SEQ ID NO:222), rat (SEQ ID NO:220) and, as well as the amino acid sequence of CRLR chimera #1 (SEQ ID NO:223).

[0422] 293-6E cells were transiently transfected with CGRP R chimera DNA constructs (CRLR wt + RAMP1 Q28-A34;

CRLR wt + RAMP1 Q43-E53; CRLR wt + RAMP1 R67-E78; CRLR L24-Q33 + RAMP wt; CRLR wt + RAMP1 wt; pTT5 vector controi). Cells were harvested after 72hr, washed with PBS + 0.5% BSA, and counted. Each transfected cell line was resuspended at a dilution of 5x10⁵ cells per 100μΙ PBS/BSA. 100μΙ of cell suspension was aliquot per well in a 96well round-bottom piate (Falcon). The cells were pelleted at 1200rpm fór 5 min. The supernatant was removed and replaced with 100μΙ containing 0.5pg purified huCGRP R antibodies 1H7,2E7, 3B6, 9F5, 4H6,12G8, 3C8,10E4,11D11, 32H8, or 33B5. Controi wells were treated with anti-DNP hulgG2 (0.5pg), Alexa647-CGRP peptide (0.5pg), or PBS/BSA alone. Cells were incubated on ice fór 1 hr. and then washed twice with PBS/BSA. The cells were resuspended in ΙΟΟμΙ/well PBS/BSA containing anti-hug-Fc-FITC (0.5pg) (except fór Alexa647-CGRP treated cells). Cells were incubated on ice in the dark fór 1 hr. and then washed twice with PBS/BSA. Cells were resuspended in 200μΙ PBS/BSA and analyzed using a FACS Calibur.

[0423] Ten representative blocking antibodies (3B6, 9F5, 4H6, 12G8, 3C8, 10E4, 32H7, 4E4, 11D11 and 1H7) and two non-blocking antibody (32H8 and 33B5) were tested. Representative data (9F5 antibody) are shown in Figs. 13A, 13B and 13C. Fig. 13A shows binding to the wild-type CGRP receptor; Fig. 13B show binding to CGRP receptors containing the CRLR L24-Q33 chimera, and Fig. 13C show binding to CGRP receptors containing the RAMP1 Q28-A34 chimera. The FACS analysis showed that all 12 antibodies bind wild type humán CGRP receptor controi as expected. All 12 antibodies showed significantly reduced binding to any ofthe three RAMP1 chimera RAMP1 (Q28-A34), (Q43E53) and (R67 - E78). This could resuít from (1) the expression level ofthe chimera receptor was much lower, (2) the RAMP1 chimera impaired the folding with humán CRLR and altered the conformation of the receptor complex, and/or (3) these three selected regions on RAMP1 are directly involved in the binding of these antibodies to CGRP receptor.

ΕΡ 2 379 594 Β1 [0424] When the FACS tracing were gated to include only the very small expressing cell populations, the non-blocking antibodies 33B5 and 32H8 appeared to consistently bind less well (lower Geo Means) to the RAMP1 O43-E53 chimera as compared to the blocking antibodies, suggesting binding to the RAMP1 O43-E53 region may be more important for the non-blocking antibodies. On the other hand, 33B5 and 32H8 consistently bound better to the RAMP1 R67-E78 chimera than the blocking antibodies, suggesting the RAMP1 R67-E78 sequence may be more importantforthe blocking antibodies.

[0425] All CGRP receptor antibodies tested bound reasonably well to the CRLR chimera (L24-O33), suggesting this site is nőt essential for binding of blocking antibodies.

[0426] In summary, the data show that three discontinuous regions on RAMP1 — (O28-A34), (043 - E53) and (R67E78) -- could be involved in CGRP receptor antibody binding, with (R67-E78) more important to the blocking antibodies. The N-terminal sequences (L24-O33) of CRLR did nőt appear to be critically involved in binding for the CGRP receptor antibodies as analyzed by this method. This approach does nőt rule out additional binding sites that share identical or similar sequences between humán and rat CGRP receptors which were nőt targeted in the analysis.

EXAMPLE 10

IDENTIFICATION OF HUMÁN CGRP R EPITOPES FOR ΑΝΤΙ-CGRP R NEUTRALIZING ANTIBODIES BY PROTEASE PROTECTION ASSAY [0427] The CRLR portion in the mature form ofthe CRLR-Fc fusion molecule (with signal peptide removed; disclosed herein as SEC ID NO:10) contains 116 amino acids (preceding the glycine linker) and has three large loop structures created by formation of three disulfide bonds. The three disulfide bonds in CRLR are Cys1 at sequence position 26 (all CRLR sequence positions listed in this paragraph are with respect to the mature sequence presented as SEC ID NO: 10) linked to Cys3 at sequence position 52 (referred to as CRLR C1-C3), Cys2 at sequence position 43 linked to Cys5 at sequence.position 83 (referred to as CRLR C2-C5), Cys4 at sequence position 66 linked to Cys6 at sequence position 105 (referred to as CRLR C4-C6). RAMP1 portion in mature form of RAMP1-Fc fusion molecule contains 91 amino acids (SEC ID NO:11) preceding the glycine linker, which alsó forms three intramolecular disulfide bonds. The three disulfide bonds in RAMP1 are Cys1 at sequence position 1 (all RAMP1 sequence positions listed in this paragraph are with respect to the mature sequence presented as SEC ID NO:11) linked to Cys5 at sequence position 56 (referred to as RAMP1 C1-C5), Cys2 at sequence position 14 linked to Cys4 at sequence position 46 (referred to as RAMP1 C2-C4), Cys3 at sequence position 31 linked to Cys6 at sequence position 78 (referred to as RAMP1 C3-C6).

[0428] Regions ofthe humán CGRP receptor protein bound by anti-CGRP neutralizing monoclonal antibodies were identified by fragmenting h CGRP R intő peptides with specific proteases, and determining the sequence ofthe resulting h CGRP R peptides (i.e., both disulfide- and non-disulfide-containing peptide fragments for CRLR and RAMP1 portions). A protease protection assay was then performed to determine the proteolytic digestion of hCGRP R in the presence of binding monoclonal antibodies. The generál principle of this assay is that binding of a mAb to CGRP R can result in protection of certain specific protease cleavage sites and this information can be used to determine the region or portion of CGRP R where the mAb binds to.

[0429] Briefly, the peptide digests were subjected to HPLC peptide mapping; the individual peaks were collected, and the peptides identified and mapped by on-line electrospray ionization LC-MS (ESI-LC-MS) analyses and/or by N-terminal sequencing. All HPLC analyses for these studies were performed using a narrow boré reverse-phase C18 column (2.1 mm i.d. x 15 cm length; Zorbax 300SB, 5 μη, Agilent Technologies) for off-line analysis and using a capillary reverse phase C18 column (0.5mm i.d. x 25 cm Vydac C18 MS, 5 μιτι; The Separation Group) for LC-MS. HPLC peptide mapping was performed with a linear gradient from 0.05% trifluoroacetic acid (mobile phase A) to 90% acetonitrile in 0.05% trifluoroacetic acid. Columns were developed over 90 minutes at a flow rate of 0.25 ml/min for narrow boré HPLC for off-line or on-line LC-MS analyses, and 0.018 ml/min for capillary HPLC for on-line LC-MS analyses.

[0430] Mature form humán CGRP R was digested with AspN (which cleaves after aspartic acid and somé glutamic acid residues at the amino end) by incubating about 100 μg of CGRP R at 1.0 mg/ml in 0.1M sodium phosphate (pH 6.5) for 20 hrs at 37°C with 2 μg of AspN.

[0431] HPLC chromatography ofthe AspN digests generated a peptide profile as shown in Fig. 14 (each sample 30μg injected), chromatogram labeled A for CGRP R alone (concentration 1 mg/ml), while a control digestion with a similar amount of CGRP R neutralizing antibody, clone 12G8, shows that the antibody is essentially resistant to AspN endoproteinase (chromatogram labeled B; CGRP R:antibody ratio, 100:2; 100:7; 100:20,weight by weight, respectively). Sequence analyses were conducted by on-line LC-MS/MS and by Edman sequencing on the peptide peaks recovered from HPLC. On-line ESI LC-MS analyses of the peptide digest were performed to determine the precise mass and sequence ofthe peptides that were separated by HPLC. The identifies of several peptides present in the peptide peaks from the AspN digestion were thus determined (indicated as numbered peaks in Fig. 14). Table 14, below, shows the locations of these peptide sequences in the corresponding component (CRLR or RAMP1) of the hCGRP R. A Capital

ΕΡ 2 379 594 Β1 letter C followed by a number or X represents a peptide identified as a CRLR peptide; a Capital letter R followed by a number or X is a RAMP1 peptide and Fc represents the large, undigested Fc fragment released from the CRLR-Fc and RAMP1-Fc fusion molecules.

Table 14

CRLR and RAMP1 peptides identified by peptide mapping of CGRP R AspN digestion

<td> Peptide</td><td> Sequence location</td><td> Disulfide (#)</td><td> Intact mass</td><td> Origin</td>

<td> C1</td><td> E111-V122</td><td> 0</td><td> 1059</td><td> CRLR</td>

<td> C2</td><td> D33-A38</td><td> 0</td><td> 670</td><td> CRLR</td>

<td> C3</td><td> D55-M63</td><td> 0</td><td> 880</td><td> CRLR</td>

<td> C4</td><td> D68-Q71</td><td> 0</td><td> 571</td><td> CRLR</td>

<td> C5</td><td> D55-P67/D86-H110</td><td></td><td> n.d.</td><td> CRLR</td>

<td> C6</td><td> D8-Y24</td><td> 0</td><td> 1938</td><td> CRLR</td>

<td> C7</td><td> E25-Q32/D48-N54</td><td> 1</td><td> 1933</td><td> CRLR</td>

<td> C-X</td><td></td><td> 3</td><td> n.d.</td><td> CRLR</td>

<td> R1</td><td> D32-A44</td><td> 0</td><td> 1622</td><td> RAMP1</td>

<td> R2</td><td> E12-V20/D45-A51 1</td><td></td><td> 1939</td><td> RAMP1</td>

<td> R-X</td><td> C1-R86</td><td> 3</td><td> 10049</td><td> RAMP1</td>

<td> Fc</td><td></td><td></td><td> 20500</td><td> RAMP1/CRLR</td>

[0432] Fig. 15 shows a comparison ofan AspN digestion experiment (each sample 3C^g injected) with CGRP R alone (chromatogram labeled A) with one performed in the presence of neutralizing antibody 12G8 (chromatogram labeled B). The weight ratio of CGRP R:antibody was 1:1. Several peaks (C5, C6, and C7) show a decreased in peak height in chromatogram B relatíve to chromatogram A, while two other peaks (C-X and R-X) show an increase in peak height in chromatogram B relatíve to chromatogram A. A similar peptide map pattern was alsó observed if a different neutralizing anti-CGRP R antibody (10E4, 3B6, 3C8 or 4E4) at a similar quantity was present in the digestion sample as seen in the chromatogram labeled C. Both C6 and C7 are disulfide linked peptides which cover a major portion ofthe CRLR molecule while C5 is a CRLR non-disulfide containing peptide residing at the N-terminal end ofthe molecule and is penultimate to the C7 disulfide peptide. Peak C-X contains three CRLR disulfide bonds with multiple sequences, indicating at least two to three peptides are linked together by disulfide bonds. The fact that peak C-X has increased peak height in a CGRP R digest in the presence of CGRP neutralizing antibody indicates that the antibody has protected CGRP R from AspN digestion at several cleavage sites related to Glu25 andAsp55. The antibody does nőt appearto have a significant protective effect on Asp33 and Asp72 as peak intensity for peptides C2 and C4 did nőt decrease at all. Therefore the antibody appears to bind to a region of CRLR which includes the CRLR C1-C3 and CRLR C4-C6 disulfide region together with the loop region between Cys53 and Cys66.

[0433] The AspN mapping of hCGRP R alsó identified a RAMP1 disulfide peptide (R2) and a RAMP1 non-disulfide peptide (R1) (see Table 14 and Fig. 14). In the presence of any ofthe above-mentioned neutralizing antibodies (12G8, 10E4, 4E4, 3B6 or 3C8), peptide R-X was recovered at a significantly higher peak intensity than what was obtained from the digestion with no antibody in the sample. Mass and sequence analyses showed that R-x contains a single polypeptide chain eorresponding to the RAMP1 sequence between Cys1 and Arg86. These experiments indicate that CGRP R neutralizing antibody can protect a significant region of RAMP1 from AspN proteolytic digestion.

[0434] To assess whether the protective effect of CGRP R AspN proteolysis is specific to CGRP R-neutralizing (blocking) antibodies (as compared with anti-CGRP R non-neutralizing antibodies), an AspN digestion of CGRP R was performed in the presence of an unrelated control monoclonal antibody which does nőt neutralize CGRP R activity. The results are shown in Fig. 15 in chromatogram D. The non-neutralizing antibody does nőt show any significant blocking effect on CGRP R AspN proteolysis; indeed, the peptide map profile (chromatogram D) is nearly indistinguishable in the relevant aspects to the profile derived from digestion of CGRP R alone (chromatogram A).

[0435] The proteolysis protection effect was dependent on the concentration added to the digestion sample. As seen in Fig. 16, a fixed CGRP R quantity in the sample (10C^g) with variable amounts of anti-CGRP R neutralizing antibody 4E4 (CGRP R: antibody ratio in micrograms, 100:2; 100:7; 100:20,weight by weight, respectively) was performed for Aspen proteolysis. The protection profile can be observed and the protection is antibody concentration-dependent. [0436] Taken together, these data demonstrate that blocking or neutralizing anti-CGRP R antibodies disclosed herein can block CGRP R (on both CRLR and RAMP1 components) from AspN proteolysis, suggesting that the blocking

ΕΡ 2 379 594 Β1 antibodies bind to both CRLR and RAMP1 when these antibodies bind to the CGRP receptor. Further, the protection effect is antibody- concentration dependent. These results alsó indicate that CGRP R neutralizing antibodies bind to common regions on humán CGRP R which are close the Asp N cleavage sites.

EXAMPLE 11

COMMERCIALLY-AVAILABLE ANTI-RAMP1 AND ΑΝΤΙ-CRLR ANTIBODIES IN A cAMP FUNCTIONAL ASSAY [0437] Commercially-available antibodies directed against one or the other components (RAMP1 or CRLR) of the humán CGRP receptor were sereened in the CGRP receptor mediated cAMP assay using HTB-10 cells as described in Example 4, above, to determine whether the antibodies had biological activity. The data are presented in Table 15, below. The antibodies had either no detectable (ND), very weak (VW) or weak (W) biological activity over a concentration rangé where the exemplary antibodies disclosed herein had strong biological activity.

Table 15 Commercially-available antibody activity

<td> Name</td><td> Source</td><td> Antigén or epitope</td><td> Vendor</td><td> HTB-10 activity</td>

<td> CRLR antibody (ab13164)</td><td> Rabbit polyclonal Ab</td><td> N-terminal ECD of hCRLR</td><td> Abcam Inc., Cambridge, MA</td><td> ND</td>

<td> CALCRL antibody</td><td> Rabbit polyclonal Ab</td><td> N-terminal ECD of hCRLR</td><td> GenWay Biotech, Inc., San Diego, CA</td><td> ND</td>

<td> CRLR(N-18)</td><td> Goat polyclonal Ab</td><td> epitope mapping near Nterminus of hCRLR</td><td> Santa Cruz Biotech</td><td> VW</td>

<td> CRLR (H-42)</td><td> Rabbit polyclonal Ab</td><td> aa23-64 of hCRLR</td><td> Santa Cruz Biotech</td><td> ND</td>

<td> CALCRL Antibody (A01)</td><td> Mouse polyclonal Ab</td><td> aa23-133 of hCRLR</td><td> Novus Biologicals, Inc.</td><td> VW</td>

<td> RAMP1 (N-20)</td><td> Goat polyclonal Ab</td><td> epitope mapping at Nterminus of hCRLR</td><td> Santa Cruz Biotech</td><td> ND</td>

<td> RAMP1 Antibody</td><td> Mouse polyclonal Ab</td><td> aa27-118of hRAMPI</td><td> Novus Biologicals,</td><td> W</td>

<td> (M01)</td><td></td><td></td><td> Inc., Littleton, CO</td><td></td>

<td> RAMP1 Antibody (1F1)</td><td> Mouse monoclonal Ab</td><td> aa27-118of hRAMPI</td><td> Novus Biologicals, Inc.</td><td> ND</td>

<td> RAMP1 antibody (ab67151)</td><td> Mouse polyclonal Ab</td><td> full-length of hRAMPI</td><td> Abcam, Inc.</td><td> W</td>

<td> RAMP1 (FL-148)</td><td> Rabbit polyclonal Ab</td><td> full length hRAMPI</td><td> Santa Cruz Biotech, Santa Cruz, CA</td><td> ND</td>

EXAMPLE 12

Immunohistochemistry Staining of Cells Expressing Different Receptor Components [0438] 2-4x10⁶ cells were injected per collá plug (Integra LifeSciencesCo., Plainsboro, NJ). Collá plugs were embedded in OCT médium (Sakura Finetek Inc., Torrance, CA), frozen at - 20°C and cut intő 20 μπι sections using a cryostat. Sections were fixed with 4% paraformaldehyde for 1 hour at room temperature (RT) and subsequently washed in phosphate-buffered saline (PBS). Endogenous peroxidase was blocked with 3% H₂O₂/PBS for 15 min and sections were incubated in blocking solution (PBS with 3% normál goat serum (Vector Labs, Burlingame, CA) and 0.3% triton X100) for 1 hour. Subsequently, sections were incubated in humán anti-CGRP receptor primary antibody (32H7, 0.03 0.1 μg/ml) at 4°C over night, washed in PBS and incubated in secondary antibody (biotinylated goat anti-human IgG Fc fragment, 1:800, Jackson Immunoresearch, West Grove, PA) in 1 % normál goat serum/PBS for 1 hour at RT. Immunoreactivity was amplified using the Vector Elité Kit according to the manufacturer’s instructions (Vector Labs, Burlingame,

ΕΡ 2 379 594 Β1

CA) and staining was developed using 3,3’-diaminobenzidine-nickel as chromogen (Sigma-Aldrich, St. Louis, MO). Sections were cleared with xylene and coverslipped with Permount (Fisher Chemicals, Fair Lawn, NJ). Immunoreactivity was analyzed using a Nikon E-800 microscope and associated software (Nikon, Melville, NY).

[0439] Data from cells expressing different receptor com ponents(as identified below) using antibody 32H7 as deseribed above revealed pronounced staining of CHO cells expressing recombinant humán CGRP receptor (CRLR+RAMP1; CHO/CGRP R cells) and weaker staining of SK-N-MC cells that endogenously express CGRP receptors (due to much lower receptor density). No staining was observed in the parent CHO cell line, CHO cells expressing an unrelated recombinant protein (TRPM8), CHO/CGRP R cells after preabsorption with the corresponding 32H7 antigén, CHO cells expressing recombinant humán adrenomedullin receptor 2 (CRLR+RAMP3), MCF-7 cells endogenously expressing amylin receptors, HEK cells expressing recombinant humán adrenomedullin receptor 1 (CRLR+RAMP2), orthe parent HEK cells. The data from these experiments are summarized in Table 16, below.

Table 16 Immunohistochemical staining intensity of indicated cells

<td> Cell line</td><td> Staining intensity (visual score)</td>

<td> CGRP/CHO</td><td> 4+</td>

<td> SK-N-MC</td><td> 1 +</td>

<td> CHO</td><td> 0</td>

<td> TRPM8/CHO</td><td> 0</td>

<td> CGRP/CHO preadsorbed</td><td> 0</td>

<td> AM2/CHO</td><td> 0</td>

<td> MCF-7</td><td> 0</td>

<td> AM1/HEK</td><td> 0</td>

<td> HEK</td><td> 0</td>

SEQUENCE LISTING [0440] <110> AMGEN INC.

<120> HUMÁN CGRP RECEPTOR BINDING PROTEINS <130> A-1472-WO-PCT <140>

<141 >

<150> 61/264,622 <151> 2009-11-25 <150> 61/203,569 <151> 2008-12-23 <160> 261 <170> Patentln version 3.5 <210> 1 <211> 1419 <212> DNA <213> Homo sapiens <400> 1

ΕΡ 2 379 594 Β1

<td> atgttataca</td><td> gcatatttca</td><td> ttttggctta</td><td> atgatggaga</td><td> aaaagtgtac</td><td> cctgtatttt</td>

<td> ctggttctct</td><td> tgcctttttt</td><td> tatgattctt</td><td> gttacagcag</td><td> aattagaaga</td><td> gagtcctgag</td>

<td> gactcaattc</td><td> agttgggagt</td><td> tactagaaat</td><td> aaaatcatga</td><td> cagctcaata</td><td> tgaatgttac</td>

<td> caaaagatta</td><td> tgcaagaccc</td><td> cattcaacaa</td><td> gcagaaggcg</td><td> tttactgcaa</td><td> cagaacctgg</td>

<td> gatggatggc</td><td> tctgctggaa</td><td> cgatgttgca</td><td> gcaggaactg</td><td> aatcaatgca</td><td> gctctgccct</td>

<td> gattactttc</td><td> aggactttga</td><td> tccatcagaa</td><td> aaagttacaa</td><td> agatctgtga</td><td> ccaagatgga</td>

<td> aactggttta</td><td> gacatccagc</td><td> aagcaacaga</td><td> acatggacaa</td><td> attataccca</td><td> gtgtaatgtt</td>

<td> aacacccacg</td><td> agaaagtgaa</td><td> gactgcacta</td><td> aatttgtttt</td><td> acctgaccat</td><td> aattggacac</td>

<td> ggattgtcta</td><td> ttgcatcact</td><td> gcttatctcg</td><td> cttggcatat</td><td> tcttttattt</td><td> caagagccta</td>

<td> agttgccaaa</td><td> ggattacctt</td><td> acacaaaaat</td><td> ctgttcttct</td><td> catttgtttg</td><td> taactctgtt</td>

<td> gtaacaatca</td><td> ttcacctcac</td><td> tgcagtggcc</td><td> aacaaccagg</td><td> ccttagtagc</td><td> cacaaatcct</td>

<td> gttagttgca</td><td> aagtgtccca</td><td> gttcattcat</td><td> ctttacctga</td><td> tgggctgtaa</td><td> ttacttttgg</td>

<td> atgctctgtg</td><td> aaggcattta</td><td> cctacacaca</td><td> ctcattgtgg</td><td> tggccgtgtt</td><td> tgcagagaag</td>

<td> caacatttaa</td><td> tgtggtatta</td><td> ttttcttggc</td><td> tggggatttc</td><td> cactgattcc</td><td> tgcttgtata</td>

<td> catgccattg</td><td> ctagaagctt</td><td> atattacaat</td><td> gacaattgct</td><td> ggatcagttc</td><td> tgatacccat</td>

<td> ctcctctaca</td><td> ttatccatgg</td><td> cccaatttgt</td><td> gctgctttac</td><td> tggtgaatct</td><td> ttttttcttg</td>

<td> ttaaatattg</td><td> tacgcgttct</td><td> catcaccaag</td><td> ttaaaagtta</td><td> cacaccaagc</td><td> ggaatccaat</td>

<td> ctgtacatga</td><td> aagctgtgag</td><td> agctactctt</td><td> atcttggtgc</td><td> cattgcttgg</td><td> cattgaattt</td>

<td> gtgctgattc</td><td> catggcgacc</td><td> tgaaggaaag</td><td> attgcagagg</td><td> aggtatatga</td><td> ctacatcatg</td>

<td> cacatcctta</td><td> tgcacttcca</td><td> gggtcttttg</td><td> gtctctacca</td><td> ttttctgctt</td><td> ctttaatgga</td>

<td> gaggttcaag</td><td> caattctgag</td><td> aagaaactgg</td><td> aatcaataca</td><td> aaatccaatt</td><td> tggaaacagc</td>

<td> ttttccaact</td><td> cagaagctct</td><td> tcgtagtgcg</td><td> tcttacacag</td><td> tgtcaacaat</td><td> cagtgatggt</td>

<td> ccaggttata</td><td> gtcatgactg</td><td> tcctagtgaa</td><td> cacttaaatg</td><td> gaaaaagcat</td><td> ccatgatatt</td>

<td> gaaaatgttc</td><td> tcttaaaacc</td><td> agaaaattta</td><td> tataattga</td><td></td><td></td>

<210> 2 <211 > 472 <212> PRT <213> Homo sapiens <400> 2

ΕΡ 2 379 594 Β1

Met Leu Tyr Ser Ile Phe His Phe Gly Leu Met Met Glu Lys Lys Cys 15 10 15

Thr Leu Tyr Phe Leu Val Leu Leu Pro Phe Phe Met Ile Leu Val Thr 20 25 30

Ala Glu Leu Glu Glu Ser Pro Glu Asp Ser Ile Gin Leu Gly Val Thr 35 40 45

Arg Asn Lys Ile Met Thr Ala Gin Tyr Glu Cys Tyr Gin Lys Ile Met 50 55 60

Gin Asp Pro Ile Gin Gin Ala Glu Gly Val Tyr Cys Asn Arg Thr Trp 65 70 75 80

Asp Gly Trp Leu Cys Trp Asn Asp Val Ala Ala Gly Thr Glu Ser Met 85 90 95

Gin Leu Cys Pro Asp Tyr Phe Gin Asp Phe Asp Pro Ser Glu Lys Val 100 105 110

Thr Lys Ile Cys Asp Gin Asp Gly Asn Trp Phe Arg His Pro Ala Ser 115 120 125

Asn Arg Thr Trp Thr Asn Tyr Thr Gin Cys Asn Val Asn Thr His Glu 130 135 140

Lys Val Lys Thr Ala Leu Asn Leu Phe Tyr Leu Thr Ile Ile Gly His 145 150 155 160

Gly Leu Ser Ile Ala Ser Leu Leu Ile Ser Leu Gly Ile Phe Phe Tyr 165 170 175

Phe Lys Ser Leu Ser Cys Gin Arg Ile Thr Leu His Lys Asn Leu Phe 180 185 190

ΕΡ 2 379 594 Β1

Phe Ser Phe Val Cys Asn Ser Val Val Thr Ile Ile His Leu Thr Alá 195 200 205

Val Alá Asn Asn Gin Alá Leu Val Alá Thr Asn Pro Val Ser Cys Lys 210 215 220

Val Ser Gin Phe Ile His Leu Tyr Leu Met Gly Cys Asn Tyr Phe Trp 225 230 235 240

Met Leu Cys Glu Gly Ile Tyr Leu His Thr Leu Ile Val Val Alá Val 245 250 255

Phe Alá Glu Lys Gin His Leu Met Trp Tyr Tyr Phe Leu Gly Trp Gly 260 265 270

Phe Pro Leu Ile Pro Alá Cys Ile His Alá Ile Alá Arg Ser Leu Tyr 275 280 285

Tyr Asn Asp Asn Cys Trp Ile Ser Ser Asp Thr His Leu Leu Tyr Ile 290 295 300

Ile His Gly Pro Ile Cys Alá Alá Leu Leu Val Asn Leu Phe Phe Leu 305 310 315 320

Leu Asn Ile Val Arg Val Leu Ile Thr Lys Leu Lys Val Thr His Gin 325 330 335

Alá Glu Ser Asn Leu Tyr Met Lys Alá Val Arg Alá Thr Leu Ile Leu 340 345 350

Val Pro Leu Leu Gly Ile Glu Phe Val Leu Ile Pro Trp Arg Pro Glu 355 360 365

Gly Lys Ile Alá Glu Glu Val Tyr Asp Tyr Ile Met His Ile Leu Met 370 375 380

His Phe Gin Gly Leu Leu Val Ser Thr Ile Phe Cys Phe Phe Asn Gly 385 390 395 400

Glu Val Gin Alá Ile Leu Arg Arg Asn Trp Asn Gin Tyr Lys Ile Gin 405 410 415

Phe Gly Asn Ser Phe Ser Asn Ser Glu Alá Leu Arg Ser Alá Ser Tyr 420 425 430

Thr Val Ser Thr Ile Ser Asp Gly Pro Gly Tyr Ser His Asp Cys Pro 435 440 445

Ser Glu His Leu Asn Gly Lys Ser Ile His Asp Ile Glu Asn Val Leu

100

ΕΡ 2 379 594 Β1

450

Leu Lys Pro Glu Asn 5 465 <210>3 <211 > 447 <212> DNA <213> Homo sapiens <400> 3

455 460

Leu Tyr Asn

470

<td> 15</td><td> atggcccggg</td><td> ccctgtgccg</td><td> cctcccgcgg</td>

<td></td><td> ctcttcatga</td><td> ccactgcctg</td><td> ccaggaggct</td>

<td></td><td> ctcacccagt</td><td> tccaggtaga</td><td> catggaggcc</td>

<td> 20</td><td> aggaccatca</td><td> ggagctacag</td><td> ggagctggcc</td>

<td></td><td> ggctgcttct</td><td> ggcccaatgc</td><td> agaggtggac</td>

<td rowspan="2"> 25</td><td> ttcaggagct</td><td> gccccatctc</td><td> aggcagggcc</td>

<td> cccttcatcg</td><td> tggtccccat</td><td> cacggtgacc</td>

<td></td><td> agcaagcgca</td><td> ctgagggcat</td><td> tgtgtag</td>

<td> cgcggcctct</td><td> ggctgctcct</td><td> ggcccatcac</td><td> 60</td>

<td> aactacggtg</td><td> ccctcctccg</td><td> ggagctctgc</td><td> 120</td>

<td> gtcggggaga</td><td> cgctgtggtg</td><td> tgactggggc</td><td> 180</td>

<td> gactgcacct</td><td> ggcacatggc</td><td> ggagaagctg</td><td> 240</td>

<td> aggttcttcc</td><td> tggcagtgca</td><td> tggccgctac</td><td> 300</td>

<td> gtgcgggacc</td><td> cgcccggcag</td><td> catcctctac</td><td> 360</td>

<td> ctgctggtga</td><td> cggcactggt</td><td> ggtctggcag</td><td> 420</td>

447 <210>4 <211> 148 <212> PRT <213> Homo sapiens <400> 4

101

ΕΡ 2 379 594 Β1

Met Alá Arg 1

Alá Leu Cys Arg Leu Pro Arg Arg Gly Leu Trp Leu Leu 5 10 15

Leu Alá His

Gly Alá Leu 35

Glu Alá Val 50

Ser Tyr Arg 65

Gly Cys Phe

His Gly Arg

Asp Pro Pro

115

His Leu Phe Met Thr Thr Alá Cys Gin Glu Alá Asn Tyr 20 25 30

Leu Arg Glu Leu Cys Leu Thr Gin Phe Gin Val Asp Met 40 45

Gly Glu Thr Leu Trp Cys Asp Trp Gly Arg Thr Ile Arg 55 60

Glu Leu Alá Asp Cys Thr Trp His Met Alá Glu Lys Leu 70 75 80

Trp Pro Asn Alá Glu Val Asp Arg Phe Phe Leu Alá Val 85 90 95

Tyr Phe Arg Ser Cys Pro Ile Ser Gly Arg Alá Val Arg 100 105 110

Gly Ser Ile Leu Tyr Pro Phe Ile Val Val Pro Ile Thr

120

125

Val Thr Leu 130

Leu Val Thr Alá Leu Val Val Trp Gin Ser Lys Arg Thr 135 140

Glu Gly Ile 145 <210> 5 <211> 414 <212> DNA <213> Homo sapiens <400> 5

Val

102

ΕΡ 2 379 594 Β1

<td> atggagaaaa</td><td> agtgtaccct</td><td> gtattttctg</td><td> gttctcttgc</td><td> ctttttttat</td><td> gattcttgtt</td><td> 60</td>

<td> acagcagaat</td><td> tagaagagag</td><td> tcctgaggac</td><td> tcaattcagt</td><td> tgggagttac</td><td> tagaaataaa</td><td> 120</td>

<td> atcatgacag</td><td> ctcaatatga</td><td> atgttaccaa</td><td> aagattatgc</td><td> aagaccccat</td><td> tcaacaagca</td><td> 180</td>

<td> gaaggcgttt</td><td> actgcaacag</td><td> aacctgggat</td><td> ggatggctct</td><td> gctggaacga</td><td> tgttgcagca</td><td> 240</td>

<td> ggaactgaat</td><td> caatgcagct</td><td> ctgccctgat</td><td> tactttcagg</td><td> actttgatcc</td><td> atcagaaaaa</td><td> 300</td>

<td> gttacaaaga</td><td> tctgtgacca</td><td> agatggaaac</td><td> tggtttagac</td><td> atccagcaag</td><td> caacagaaca</td><td> 360</td>

<td> tggacaaatt</td><td> atacccagtg</td><td> taatgttaac</td><td> acccacgaga</td><td> aagtgaagac</td><td> tgca</td><td> 414</td>

<210>6 <211> 138 <212> PRT <213> Homo sapiens <400> 6

Met Glu Lys Lys Cys Thr Leu Tyr Phe Leu Val Leu Leu Pro Phe Phe 15 10 15

Met Ile Leu Val Thr Alá Glu Leu Glu Glu Ser Pro Glu Asp Ser Ile 20 25 30

Gin Leu Gly Val Thr Arg Asn Lys Ile Met Thr Alá Gin Tyr Glu Cys

40 45

Tyr Gin Lys Ile Met Gin Asp Pro Ile Gin Gin Alá Glu Gly Val Tyr 50 55 60

Cys Asn Arg Thr Trp Asp Gly Trp Leu Cys Trp Asn Asp Val Alá Alá 65 70 75 80

Gly Thr Glu Ser Met Gin Leu Cys Pro Asp Tyr Phe Gin Asp Phe Asp 85 90 95

Pro Ser

Glu Lys Val Thr Lys 100

Arg His

Pro Alá Ser Asn Arg 115

Val Asn 130

Thr His Glu Lys Val 135

Ile

Thr

120

Lys

Cys

105

Trp

Thr

Asp Gin

Thr Asn

Alá

Asp

Tyr

Gly Asn 110

Thr Gin 125

Trp Phe

Cys Asn <210> 7 <211> 351 <212> DNA <213> Homo sapiens

103

EP 2 379 594 Β1 <400>7

<td> atggcccggg</td><td> ccctgtgccg</td><td> cctcccgcgg</td><td> cgcggcctct</td><td> ggctgctcct</td><td> ggcccatcac</td><td> 60</td>

<td> ctcttcatga</td><td> ccactgcctg</td><td> ccaggaggct</td><td> aactacggtg</td><td> ccctcctccg</td><td> ggagctctgc</td><td> 120</td>

<td> ctcacccagt</td><td> tccaggtaga</td><td> catggaggcc</td><td> gtcggggaga</td><td> cgctgtggtg</td><td> tgactggggc</td><td> 180</td>

<td> aggaccatca</td><td> ggagctacag</td><td> ggagctggcc</td><td> gactgcacct</td><td> ggcacatggc</td><td> ggagaagctg</td><td> 240</td>

<td> ggctgcttct</td><td> ggcccaatgc</td><td> agaggtggac</td><td> aggttcttcc</td><td> tggcagtgca</td><td> tggccgctac</td><td> 300</td>

<td> ttcaggagct</td><td> gccccatctc</td><td> aggcagggcc</td><td> gtgcgggacc</td><td> cgcccggcag</td><td> c</td><td> 351</td>

<210> 8 <211> 117 <212> PRT <213> Homo sapiens <400> 8

Met Alá Arg Alá Leu Cys Arg Leu Pro Arg Arg Gly Leu Trp Leu Leu 15 10 15

Leu Alá His His Leu Phe Met Thr Thr Alá Cys Gin Glu Alá Asn Tyr 20 25 30

Gly Alá Leu Leu Arg Glu Leu Cys Leu Thr Gin Phe Gin Val Asp Met 35 40 45

Glu Alá Val Gly Glu Thr Leu Trp Cys Asp Trp Gly Arg Thr Ile Arg 50 55 60

Ser Tyr Arg Glu Leu Alá Asp Cys Thr Trp His Met Alá Glu Lys Leu 65 70 75 80

Gly Cys Phe Trp Pro Asn Alá Glu Val Asp Arg Phe Phe Leu Alá Val 85 90 95

His Gly Arg Tyr Phe Arg Ser Cys Pro Ile Ser Gly Arg Alá Val Arg 100 105 110

Asp Pro Pro Gly Ser 115 <210>9 <211> 31 <212> PRT <213> Homo sapiens <400>9

104

ΕΡ 2 379 594 Β1

Trp Val Thr His Arg 1 5

Leu

Ala Gly Leu

Leu

Ser Arg

Ser Gly

Gly Val 15

Val Arg Cys Asn Phe Val Pro Thr Asp Val Gly Pro Phe Ala Phe 20 25 30 <210> 10 <211> 116 <212> PRT <213> Homo sapiens <400> 10

Glu Leu Glu Glu Ser Pro Glu Asp Ser Ile Gin Leu Gly Val Thr Arg 15 10 15

Asn Lys

Ile Met 20

Thr Ala Gin

Tyr Glu Cys 25

Tyr Gin Lys

Ile Met Gin 30

Asp Pro

Ile

Gin Gin Ala Glu

Gly Val 40

Tyr Cys

Asn Arg 45

Thr

Trp Asp

Gly Trp 50

Leu Cys

Trp Asn Asp 55

Val

Ala Ala

Gly

Thr Glu 60

Ser Met

Gin

Leu Cys 65

Pro Asp

Tyr Phe 70

Gin Asp

Phe Asp

Pro

Ser Glu Lys

Val

Thr

Lys

Ile

Cys Asp

Gin Asp 85

Gly Asn

Trp

Phe Arg His 90

Pro Ala

Ser Asn 95

Arg Thr Trp

Thr Asn 100

Tyr

Thr Gin Cys Asn Val 105

Asn

Thr His 110

Glu Lys

Val Lys Thr Ala 115 <210> 11 <211> 91 <212> PRT <213> Homo sapiens <400> 11

Cys Gin Glu Ala Asn Tyr Gly Ala Leu Leu Arg Glu Leu Cys Leu Thr 15 10 15

Gin Phe Gin Val 20

Asp Met

Glu

Ala

Val

Gly Glu

Thr Leu

Trp Cys Asp 30

105

ΕΡ 2 379 594 Β1

<td> Trp</td><td> Gly</td><td> Arg 35</td><td> Thr</td><td> Ile</td><td> Arg</td><td> Ser</td><td> Tyr 40</td><td> Arg</td><td> Glu</td><td> Leu</td><td> Alá</td><td> Asp 45</td><td> Cys</td><td> Thr</td><td> Trp</td>

<td> His</td><td> Met 50</td><td> Alá</td><td> Glu</td><td> Lys</td><td> Leu</td><td> Gly 55</td><td> Cys</td><td> Phe</td><td> Trp</td><td> Pro</td><td> Asn 60</td><td> Alá</td><td> Glu</td><td> Val</td><td> Asp</td>

<td> Arg 65</td><td> Phe</td><td> Phe</td><td> Leu</td><td> Alá</td><td> Val 70</td><td> His</td><td> Gly</td><td> Arg</td><td> Tyr</td><td> Phe 75</td><td> Arg</td><td> Ser</td><td> Cys</td><td> Pro</td><td> Ile 80</td>

<td> Ser</td><td> Gly</td><td> Arg</td><td> Alá</td><td> Val</td><td> Arg</td><td> Asp</td><td> Pro</td><td> Pro</td><td> Gly</td><td> Ser</td><td></td><td></td><td></td><td></td><td></td>

90 <210> 12 <211> 238 <212> PRT <213> Artificial Sequence

<td colspan="10"> &lt;220&gt; &lt;221 &gt; source &lt;223&gt; /note=Description of Artificial Sequence: Synthetic polypeptide</td>

<td colspan="2"> &lt;400&gt; 12</td><td rowspan="2"> Pro</td><td rowspan="2"> Alá Gin Leu</td><td rowspan="2"> Leu 10</td><td rowspan="2"> Gly Leu</td><td rowspan="2"> Leu</td><td rowspan="2"> Leu</td><td rowspan="2"> Leu 15</td><td rowspan="2"> Trp</td>

<td> Met Asp Met Arg 1</td><td> Val 5</td>

<td> Leu Arg Gly Alá 20</td><td> Arg</td><td> Cys</td><td> Gin Ser Val 25</td><td> Leu</td><td> Thr Gin</td><td> Pro</td><td> Pro 30</td><td> Ser</td><td> Val</td>

<td> Ser Glu Alá Pro 35</td><td> Gly</td><td> Gin</td><td> Lys Val Thr 40</td><td> Ile</td><td> Ser Cys</td><td> Ser 45</td><td> Gly</td><td> Ser</td><td> Ser</td>

<td> Ser Asn Ile Gly 50</td><td> Asn</td><td> Asn</td><td> Tyr Val Ser 55</td><td> Trp</td><td> Tyr Gin 60</td><td> Gin</td><td> Leu</td><td> Pro</td><td> Gly</td>

<td> Thr Alá Pro Lys 65</td><td> Leu</td><td> Leu 70</td><td> Ile Tyr Asp</td><td> Asn</td><td> Asn Lys 75</td><td> Arg</td><td> Pro</td><td> Ser</td><td> Gly 80</td>

<td> Ile Pro Asp Arg</td><td> Phe 85</td><td> Ser</td><td> Gly Ser Lys</td><td> Ser 90</td><td> Gly Thr</td><td> Ser</td><td> Alá</td><td> Thr 95</td><td> Leu</td>

<td> Gly Ile Thr Gly 100</td><td> Leu</td><td> Gin</td><td> Thr Gly Asp 105</td><td> Glu</td><td> Alá Asp</td><td> Tyr</td><td> Tyr 110</td><td> Cys</td><td> Gly</td>

<td> Thr Trp Asp Ser 115</td><td> Arg</td><td> Leu</td><td> Ser Alá Val 120</td><td> Val</td><td> Phe Gly</td><td> Gly 125</td><td> Gly</td><td> Thr</td><td> Lys</td>

<td> Leu Thr Val Leu 130</td><td> Gly</td><td> Gin</td><td> Pro Lys Alá 135</td><td> Asn</td><td> Pro Thr 140</td><td> Val</td><td> Thr</td><td> Leu</td><td> Phe</td>

106

ΕΡ 2 379 594 Β1

<td> Pro</td><td> Pro</td><td> Ser</td><td> Ser</td><td> Glu</td><td> Glu</td><td> Leu</td><td> Gin</td><td> Alá</td><td> Asn</td><td> Lys</td><td> Alá</td><td> Thr</td><td> Leu</td><td> Val</td><td> Cys</td>

<td> 145</td><td></td><td></td><td></td><td></td><td> 150</td><td></td><td></td><td></td><td></td><td> 155</td><td></td><td></td><td></td><td></td><td> 160</td>

<td> Leu</td><td> Ile</td><td> Ser</td><td> Asp</td><td> Phe</td><td> Tyr</td><td> Pro</td><td> Gly</td><td> Alá</td><td> Val</td><td> Thr</td><td> Val</td><td> Alá</td><td> Trp</td><td> Lys</td><td> Alá</td>

<td></td><td></td><td></td><td></td><td> 165</td><td></td><td></td><td></td><td></td><td> 170</td><td></td><td></td><td></td><td></td><td> 175</td><td></td>

<td> Asp</td><td> Gly</td><td> Ser</td><td> Pro</td><td> Val</td><td> Lys</td><td> Alá</td><td> Gly</td><td> Val</td><td> Glu</td><td> Thr</td><td> Thr</td><td> Lys</td><td> Pro</td><td> Ser</td><td> Lys</td>

<td></td><td></td><td></td><td> 180</td><td></td><td></td><td></td><td></td><td> 185</td><td></td><td></td><td></td><td></td><td> 190</td><td></td><td></td>

<td> Gin</td><td> Ser</td><td> Asn</td><td> Asn</td><td> Lys</td><td> Tyr</td><td> Alá</td><td> Alá</td><td> Ser</td><td> Ser</td><td> Tyr</td><td> Leu</td><td> Ser</td><td> Leu</td><td> Thr</td><td> Pro</td>

<td></td><td></td><td> 195</td><td></td><td></td><td></td><td></td><td> 200</td><td></td><td></td><td></td><td></td><td> 205</td><td></td><td></td><td></td>

<td> Glu</td><td> Gin</td><td> Trp</td><td> Lys</td><td> Ser</td><td> His</td><td> Arg</td><td> Ser</td><td> Tyr</td><td> Ser</td><td> Cys</td><td> Gin</td><td> Val</td><td> Thr</td><td> His</td><td> Glu</td>

<td></td><td> 210</td><td></td><td></td><td></td><td></td><td> 215</td><td></td><td></td><td></td><td></td><td> 220</td><td></td><td></td><td></td><td></td>

<td> Gly</td><td> Ser</td><td> Thr</td><td> Val</td><td> Glu</td><td> Lys</td><td> Thr</td><td> Val</td><td> Alá</td><td> Pro</td><td> Thr</td><td> Glu</td><td> Cys</td><td> Ser</td><td></td><td></td>

225 230 235 <210> 13 <211> 238 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 13

107

ΕΡ 2 379 594 Β1

Met Asp Met Arg Val Pro Alá Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Alá Arg Cys Gin Ser Val Leu Thr Gin Pro Pro Ser Alá 20 25 30

Ser Gly Thr Pro Gly Gin Arg Val Thr Ile Ser Cys Ser Gly Ser Ser 35 40 45

Ser Asn Ile Gly Ser Asn Tyr Val Tyr Trp Tyr Gin Gin Leu Pro Gly 50 55 60

Alá Alá Pro Lys Leu Leu Ile Phe Arg Ser Asn Gin Arg Pro Ser Gly 65 70 75 80

Val Pro Asp Arg Phe Ser Gly Ser Lys Ser Gly Thr Ser Alá Ser Leu 85 90 95

Alá Ile Ser Gly Leu Arg Ser Glu Asp Glu Alá Asp Tyr Tyr Cys Alá 100 105 110

Alá Trp Asp Asp Ser Leu Ser Gly Trp Val Phe Gly Gly Gly Thr Lys 115 120 125

Leu Thr Val Leu Gly Gin Pro Lys Alá Asn Pro Thr Val Thr Leu Phe 130 135 140

Pro Pro Ser Ser Glu Glu Leu Gin Alá Asn Lys Alá Thr Leu Val Cys 145 150 155 160

Leu Ile Ser Asp Phe Tyr Pro Gly Alá Val Thr Val Alá Trp Lys Alá 165 170 175

Asp Gly Ser Pro Val Lys Alá Gly Val Glu Thr Thr Lys Pro Ser Lys 180 185 190

Gin Ser Asn Asn Lys Tyr Alá Alá Ser Ser Tyr Leu Ser Leu Thr Pro 195 200 205

Glu Gin Trp Lys Ser His Arg Ser Tyr Ser Cys Gin Val Thr His Glu 210 215 220

Gly

225

Ser Thr Val

Glu Lys 230

Thr Val Alá

Pro

Thr Glu Cys Ser 235 <210> 14 <211> 236

108

EP 2 379 594 B1 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 14

Met Asp Met Arg Val Pro Alá 1 5

Leu Arg Gly Alá Arg Cys Asp 20

Leu Ser Alá Ser Val Gly Asp 35

Gin Gly Ile Arg Asn Asp Leu 50 55

Alá Pro Lys Arg Leu Ile Tyr 65 70

Pro Ser Arg Phe

Ser Gly 85

Gin Leu Leu Gly Leu Leu Leu Leu Trp 10 15

Ile Gin Met Thr Gin Ser Pro Ser Ser 25 30

Arg Val Thr Ile Thr Cys Arg Alá Ser 40 45

Gly Trp Phe Gin Gin Lys Pro Gly Lys 60

Alá Alá Ser Ser Leu Gin Ser Gly Val 75 80

Ser Gly

Ser

Gly Thr Glu Phe 90

Thr Leu Thr 95

109

ΕΡ 2 379 594 Β1

<td rowspan="2"> Ile</td><td rowspan="2"> Ser</td><td colspan="3" rowspan="2"> Ser Leu Gin 100</td><td rowspan="2"> Pro</td><td colspan="8"> Glu Asp Leu Alá Thr Tyr Tyr Cys</td><td colspan="2" rowspan="2"> Leu Gin</td>

<td colspan="3"> 105</td><td colspan="5"> 110</td>

<td> Tyr</td><td> Asn</td><td> Ile</td><td> Tyr</td><td> Pro</td><td> Trp</td><td> Thr</td><td> Phe</td><td> Gly</td><td> Gin</td><td> Gly</td><td> Thr</td><td> Lys</td><td> Val</td><td> Glu</td><td> Ile</td>

<td></td><td></td><td> 115</td><td></td><td></td><td></td><td></td><td> 120</td><td></td><td></td><td></td><td></td><td> 125</td><td></td><td></td><td></td>

<td> Lys</td><td> Arg</td><td> Thr</td><td> Val</td><td> Alá</td><td> Alá</td><td> Pro</td><td> Ser</td><td> Val</td><td> Phe</td><td> Ile</td><td> Phe</td><td> Pro</td><td> Pro</td><td> Ser</td><td> Asp</td>

<td></td><td> 130</td><td></td><td></td><td></td><td></td><td> 135</td><td></td><td></td><td></td><td></td><td> 140</td><td></td><td></td><td></td><td></td>

<td> Glu</td><td> Gin</td><td> Leu</td><td> Lys</td><td> Ser</td><td> Gly</td><td> Thr</td><td> Alá</td><td> Ser</td><td> Val</td><td> Val</td><td> Cys</td><td> Leu</td><td> Leu</td><td> Asn</td><td> Asn</td>

<td> 145</td><td></td><td></td><td></td><td></td><td> 150</td><td></td><td></td><td></td><td></td><td> 155</td><td></td><td></td><td></td><td></td><td> 160</td>

<td> Phe</td><td> Tyr</td><td> Pro</td><td> Arg</td><td> Glu</td><td> Alá</td><td> Lys</td><td> Val</td><td> Gin</td><td> Trp</td><td> Lys</td><td> Val</td><td> Asp</td><td> Asn</td><td> Alá</td><td> Leu</td>

<td></td><td></td><td></td><td></td><td> 165</td><td></td><td></td><td></td><td></td><td> 170</td><td></td><td></td><td></td><td></td><td> 175</td><td></td>

<td> Gin</td><td> Ser</td><td> Gly</td><td> Asn</td><td> Ser</td><td> Gin</td><td> Glu</td><td> Ser</td><td> Val</td><td> Thr</td><td> Glu</td><td> Gin</td><td> Asp</td><td> Ser</td><td> Lys</td><td> Asp</td>

<td></td><td></td><td></td><td> 180</td><td></td><td></td><td></td><td></td><td> 185</td><td></td><td></td><td></td><td></td><td> 190</td><td></td><td></td>

<td> Ser</td><td> Thr</td><td> Tyr</td><td> Ser</td><td> Leu</td><td> Ser</td><td> Ser</td><td> Thr</td><td> Leu</td><td> Thr</td><td> Leu</td><td> Ser</td><td> Lys</td><td> Alá</td><td> Asp</td><td> Tyr</td>

<td></td><td></td><td> 195</td><td></td><td></td><td></td><td></td><td> 200</td><td></td><td></td><td></td><td></td><td> 205</td><td></td><td></td><td></td>

<td> Glu</td><td> Lys</td><td> His</td><td> Lys</td><td> Val</td><td> Tyr</td><td> Alá</td><td> Cys</td><td> Glu</td><td> Val</td><td> Thr</td><td> His</td><td> Gin</td><td> Gly</td><td> Leu</td><td> Ser</td>

<td></td><td> 210</td><td></td><td></td><td></td><td></td><td> 215</td><td></td><td></td><td></td><td></td><td> 220</td><td></td><td></td><td></td><td></td>

<td> Ser</td><td> Pro</td><td> Val</td><td> Thr</td><td> Lys</td><td> Ser</td><td> Phe</td><td> Asn</td><td> Arg</td><td> Gly</td><td> Glu</td><td> Cys</td><td></td><td></td><td></td><td></td>

225 230 235 <210> 15 <211> 236 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 15

110

ΕΡ 2 379 594 Β1

Met Asp Met Arg Val Pro Alá Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Alá Arg Cys Ser Ser Glu Leu Thr Gin Asp Pro Thr Val 20 25 30

Ser Val Alá Leu Gly Gin Thr Val Lys Ile Thr Cys Gin Gly Asp Ser 35 40 45

Leu Arg Ser Phe Tyr Alá Ser Trp Tyr Gin Gin Lys Pro Gly Gin Alá 50 55 60

Pro Val Leu Val Phe Tyr Gly Lys Asn Asn Arg Pro Ser Gly Ile Pro

70 75 80

Asp Arg Phe Ser Gly Ser Ser Ser Gly Asn Thr Alá Ser Leu Thr Ile

90 95

Thr Gly Alá Gin Alá Glu Asp Glu Alá Asp Tyr Tyr Cys Asn Ser Arg 100 105 110

Asp Ser Ser Val Tyr His Leu Val Leu Gly Gly Gly Thr Lys Leu Thr 115 120 125

Val Leu Gly Gin Pro Lys Alá Asn Pro Thr Val Thr Leu Phe Pro Pro 130 135 140

Ser Ser Glu Glu Leu Gin Alá Asn Lys Alá Thr Leu Val Cys Leu Ile 145 150 155 160

Ser Asp Phe Tyr Pro Gly Alá Val Thr Val Alá Trp Lys Alá Asp Gly 165 170 175

Ser Pro Val Lys Alá Gly Val Glu Thr Thr Lys Pro Ser Lys Gin Ser 180 185 190

Asn Asn Lys Tyr Alá Alá Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gin 195 200 205

Trp Lys Ser His Arg Ser Tyr Ser Cys Gin Val Thr His Glu Gly Ser 210 215 220

Thr Val Glu Lys Thr Val Alá Pro Thr Glu Cys Ser 225 230 235 <210> 16 <211> 241 <212> PRT

111

ΕΡ 2 379 594 Β1 <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 16

Met Asp Met Arg Val Pro Alá 1 5

Leu Arg Gly Alá Arg Cys Asp 20

Leu Ser Val Thr Pro Gly Gin 35

Gin Leu Leu Gly Leu Leu Leu Leu Trp 10 15

Ile Ile Leu Alá Gin Thr Pro Leu Ser 25 30

Pro Alá Ser Ile Ser Cys Lys Ser Ser 40 45

112

ΕΡ 2 379 594 Β1

<td> Gin</td><td> Ser 50</td><td> Leu</td><td> Leu</td><td> His</td><td> Ser</td><td> Alá 55</td><td> Gly</td><td> Lys</td><td> Thr</td><td> Tyr</td><td> Leu 60</td><td> Tyr</td><td> Trp</td><td> Tyr</td><td> Leu</td>

<td> Gin 65</td><td> Lys</td><td> Pro</td><td> Gly</td><td> Gin</td><td> Pro 70</td><td> Pro</td><td> Gin</td><td> Leu</td><td> Leu</td><td> Ile 75</td><td> Tyr</td><td> Glu</td><td> Val</td><td> Ser</td><td> Asn 80</td>

<td> Arg</td><td> Phe</td><td> Ser</td><td> Gly</td><td> Val 85</td><td> Pro</td><td> Asp</td><td> Arg</td><td> Phe</td><td> Ser 90</td><td> Gly</td><td> Ser</td><td> Gly</td><td> Ser</td><td> Gly 95</td><td> Thr</td>

<td> Asp</td><td> Phe</td><td> Thr</td><td> Leu 100</td><td> Lys</td><td> Ile</td><td> Ser</td><td> Arg</td><td> Val 105</td><td> Glu</td><td> Alá</td><td> Glu</td><td> Asp</td><td> Val 110</td><td> Gly</td><td> Ile</td>

<td> Tyr</td><td> Tyr</td><td> Cys 115</td><td> Met</td><td> Gin</td><td> Ser</td><td> Phe</td><td> Pro 120</td><td> Leu</td><td> Pro</td><td> Leu</td><td> Thr</td><td> Phe 125</td><td> Gly</td><td> Gly</td><td> Gly</td>

<td> Thr</td><td> Lys 130</td><td> Val</td><td> Glu</td><td> Ile</td><td> Lys</td><td> Arg 135</td><td> Thr</td><td> Val</td><td> Alá</td><td> Alá</td><td> Pro 140</td><td> Ser</td><td> Val</td><td> Phe</td><td> Ile</td>

<td> Phe 145</td><td> Pro</td><td> Pro</td><td> Ser</td><td> Asp</td><td> Glu 150</td><td> Gin</td><td> Leu</td><td> Lys</td><td> Ser</td><td> Gly 155</td><td> Thr</td><td> Alá</td><td> Ser</td><td> Val</td><td> Val 160</td>

<td> Cys</td><td> Leu</td><td> Leu</td><td> Asn</td><td> Asn 165</td><td> Phe</td><td> Tyr</td><td> Pro</td><td> Arg</td><td> Glu 170</td><td> Alá</td><td> Lys</td><td> Val</td><td> Gin</td><td> Trp 175</td><td> Lys</td>

<td> Val</td><td> Asp</td><td> Asn</td><td> Alá 180</td><td> Leu</td><td> Gin</td><td> Ser</td><td> Gly</td><td> Asn 185</td><td> Ser</td><td> Gin</td><td> Glu</td><td> Ser</td><td> Val 190</td><td> Thr</td><td> Glu</td>

<td> Gin</td><td> Asp</td><td> Ser 195</td><td> Lys</td><td> Asp</td><td> Ser</td><td> Thr</td><td> Tyr 200</td><td> Ser</td><td> Leu</td><td> Ser</td><td> Ser</td><td> Thr 205</td><td> Leu</td><td> Thr</td><td> Leu</td>

<td> Ser</td><td> Lys 210</td><td> Alá</td><td> Asp</td><td> Tyr</td><td> Glu</td><td> Lys 215</td><td> His</td><td> Lys</td><td> Val</td><td> Tyr</td><td> Alá 220</td><td> Cys</td><td> Glu</td><td> Val</td><td> Thr</td>

<td> His 225</td><td> Gin</td><td> Gly</td><td> Leu</td><td> Ser</td><td> Ser 230</td><td> Pro</td><td> Val</td><td> Thr</td><td> Lys</td><td> Ser 235</td><td> Phe</td><td> Asn</td><td> Arg</td><td> Gly</td><td> Glu 240</td>

Cys <210> 17 <211> 238 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 17

113

ΕΡ 2 379 594 Β1

Met Asp Met Arg Val 1 5

Pro Ala Gin Leu Leu 10

Gly Leu Leu Leu Leu Trp 15

Leu Arg Gly Ala Arg 20

Cys Gin Ser Val Leu 25

Thr Gin Pro Pro Ser Val 30

Ser Ala Ala Pro Gly 35

Gin Lys Val Thr Ile 40

Ser Cys Ser Gly Ser Ser 45

Ser Asn Ile Gly Asn 50

Thr Ala Pro Lys Leu 65

Ile Pro Asp Arg Phe 85

Gly Ile Thr Gly Leu 100

Thr Trp Asp Ser Arg 115

Leu Thr Val Leu Gly 130

Pro Pro Ser Ser Glu 145

Leu Ile Ser Asp Phe 165

Asp Gly Ser Pro Val 180

Gin Ser Asn Asn Lys 195

Glu Gin Trp Lys Ser 210

Gly Ser Thr Val Glu 225 <210> 18 <211> 241 <212> PRT

Asn Tyr Val Ser Trp 55

Leu Ile Tyr Asp Asn 70

Ser Gly Ser Lys Ser 90

Gin Thr Gly Asp Glu 105

Leu Ser Ala Val Val 120

Gin Pro Lys Ala Asn 135

Glu Leu Gin Ala Asn 150

Tyr Pro Gly Ala Val 170

Lys Ala Gly Val Glu 185

Tyr Ala Ala Ser Ser 200

His Arg Ser Tyr Ser 215

Lys Thr Val Ala Pro 230

Tyr Gin Gin Leu Pro Gly 60

Asn Lys Arg Pro Ser Gly 75 80

Gly Thr Ser Thr Thr Leu 95

Ala Asp Tyr Tyr Cys Gly 110

Phe Gly Gly Gly Thr Lys 125

Pro Thr Val Thr Leu Phe 140

Lys Ala Thr Leu Val Cys 155 160

Thr Val Ala Trp Lys Ala 175

Thr Thr Lys Pro Ser Lys 190

Tyr Leu Ser Leu Thr Pro 205

Cys Gin Val Thr His Glu 220

Thr Glu Cys Ser 235

114

ΕΡ 2 379 594 Β1 <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 18

115

ΕΡ 2 379 594 Β1

Met Asp Met Arg Val Pro Alá Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Alá Arg Cys Asp Ile Val Met Thr Gin Ser Pro Leu Ser 20 25 30

Leu Pro Val Thr Pro Gly Glu Pro Alá Ser Ile Ser Cys Arg Ser Ser 35 40 45

Gin Ser Leu Leu His Ser Phe Gly Tyr Asn Tyr Leu Asp Trp Tyr Leu 50 55 60

Gin Lys Pro Gly Gin Ser Pro Gin Leu Leu Ile Tyr Leu Gly Ser Asn 65 70 75 80

Arg Alá Ser Gly Val Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr 85 90 95

Asp Phe Thr Leu Lys Ile Ser Arg Val Glu Alá Glu Asp Val Gly Val 100 105 110

Tyr Tyr Cys Met Gin Alá Leu Gin Thr Pro Phe Thr Phe Gly Pro Gly 115 120 125

Thr Lys Val Asp Ile Lys Arg Thr Val Alá Alá Pro Ser Val Phe Ile 130 135 140

Phe Pro Pro Ser Asp Glu Gin Leu Lys Ser Gly Thr Alá Ser Val Val 145 150 155 160

Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Alá Lys Val Gin Trp Lys 165 170 175

Val Asp Asn Alá Leu Gin Ser Gly Asn Ser Gin Glu Ser Val Thr Glu 180 185 190

Gin Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu 195 200 205

Ser Lys Alá Asp Tyr Glu Lys His Lys Val Tyr Alá Cys Glu Val Thr 210 215 220

His Gin Gly Leu Ser Ser Pro Val Thr Lys Ser Phe Asn Arg Gly Glu 225 230 235 240

Cys

116

ΕΡ 2 379 594 Β1 <210> 19 <211> 241 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 19

Met Asp Met Arg Val 1 5

Leu Arg Gly Alá Arg 20

Leu Ser Val Thr Pro 35

Gin Ser Leu Leu His 50

Gin Lys Pro Gly Gin 65

Arg Phe Ser Gly Glu 85

Asp Phe Thr Leu Lys 100

Tyr Tyr Cys Met Gin 115

Thr Lys Val Glu Ile 130

Phe Pro Pro Ser Asp 145

Cys Leu Leu Asn Asn 165

Val Asp Asn Alá Leu 180

Gin Asp Ser Lys Asp 195

Pro Alá Gin Leu Leu Gly 10

Cys Asp Ile Ile Leu Thr 25

Gly Gin Pro Alá Ser Ile 40

Ser Asp Gly Lys Thr Tyr 55

Pro Pro Gin Leu Leu Ile 70 75

Pro Asp Arg Phe Ser Gly 90

Ile Ser Arg Val Glu Alá 105

Ser Phe Pro Leu Pro Leu 120

Lys Arg Thr Val Alá Alá 135

Glu Gin Leu Lys Ser Gly 150 155

Phe Tyr Pro Arg Glu Alá 170

Gin Ser Gly Asn Ser Gin 185

Ser Thr Tyr Ser Leu Ser 200

Leu Leu Leu Leu Trp 15

Gin Thr Pro Leu Ser 30

Ser Cys Lys Ser Ser 45

Leu Tyr Trp Tyr Leu 60

Tyr Glu Val Ser Asn 80

Ser Gly Ser Gly Thr 95

Glu Asp Val Gly Thr 110

Thr Phe Gly Gly Gly 125

Pro Ser Val Phe Ile 140

Thr Alá Ser Val Val 160

Lys Val Gin Trp Lys 175

Glu Ser Val Thr Glu 190

Ser Thr Leu Thr Leu 205

117

ΕΡ 2 379 594 Β1

<td> Ser</td><td> Lys 210</td><td colspan="2"> Alá Asp</td><td colspan="3"> Tyr Glu Lys 215</td><td> His</td><td colspan="2"> Lys Val</td><td> Tyr</td><td> Alá 220</td><td> Cys</td><td> Glu</td><td> Val</td><td> Thr</td>

<td> His</td><td> Gin</td><td> Gly</td><td> Leu</td><td> Ser</td><td> Ser</td><td> Pro</td><td> Val</td><td> Thr</td><td> Lys</td><td> Ser</td><td> Phe</td><td> Asn</td><td> Arg</td><td> Gly</td><td> Glu</td>

<td> 225</td><td></td><td></td><td></td><td></td><td> 230</td><td></td><td></td><td></td><td></td><td> 235</td><td></td><td></td><td></td><td></td><td> 240</td>

<td> Cys</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<210> 20 <211> 238 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 20

118

EP 2 379 594 Β1

Met Asp Met Arg Val Pro Alá Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Alá Arg Cys Gin Ser Val Leu Thr Gin Pro Pro Ser Val 20 25 30

Ser Alá Alá Pro Gly Gin Lys Val Thr Ile Ser Cys Ser Gly Ser Ser 35 40 45

Ser Asn Ile Gly Asn Asn Tyr Val Ser Trp Tyr Gin Gin Phe Pro Gly 50 55 60

Thr Alá Pro Lys Leu Leu Ile Tyr Asp Asn Asn Lys Arg Pro Ser Gly 65 70 75 80

Ile Pro Asp Arg Phe Ser Gly Ser Lys Ser Gly Thr Ser Alá Thr Leu 85 90 95

Gly Ile Thr Gly Leu Gin Thr Gly Asp Glu Alá Asp Tyr Tyr Cys Gly 100 105 110

Thr Trp Asp Ser Arg Leu Ser Alá Val Val Phe Gly Gly Gly Thr Lys 115 120 125

Leu Thr Val Leu Gly Gin Pro Lys Alá Asn Pro Thr Val Thr Leu Phe 130 135 140

Pro Pro Ser Ser Glu Glu Leu Gin Alá Asn Lys Alá Thr Leu Val Cys 145 150 155 160

Leu Ile Ser Asp Phe Tyr Pro Gly Alá Val Thr Val Alá Trp Lys Alá

165

170

175

Asp Gly Ser Pro Val Lys Alá Gly Val Glu Thr Thr Lys Pro Ser Lys 180 185 190

Gin Ser Asn Asn Lys Tyr Alá Alá Ser Ser Tyr Leu Ser Leu Thr Pro 195 200 205

Glu Gin Trp Lys Ser His Arg Ser Tyr Ser Cys Gin Val Thr His Glu 210 215 220

Gly Ser Thr Val Glu Lys Thr Val Alá Pro Thr Glu Cys Ser 225 230 235 <210>21

119

EP 2 379 594 B1 <211 > 238 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 21

Met Asp Met Arg Val Pro Alá Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Alá Arg Cys Gin Ser Val Leu Thr Gin Ser Pro Ser Alá 20 25 30

Ser Gly Thr Pro Gly Gin Arg Val Thr Ile Ser Cys Ser Gly Ser Ser 35 40 45

Ser Asn Ile Gly Ser Asn Tyr Val Tyr Trp Tyr Gin Gin Leu Pro Gly 50 55 60

Alá Alá Pro Lys Leu Leu Ile Leu Arg Asn Asn Gin Arg Pro Ser Gly 65 70 75 80

Val Pro Asp Arg Phe Ser Gly Ser Lys Ser Gly Thr Ser Alá Ser Leu 85 90 95

Thr Ile Ser Gly Leu Arg Ser Glu Asp Glu Alá Asp Tyr Tyr Cys Alá 100 105 110

Alá Trp Asp Asp Ser Leu Ser Gly Trp Val Phe Gly Gly Gly Thr Lys 115 120 125

Leu Thr Val 130

Leu Gly Gin

Pro

135

Lys

Alá Asn Pro

Thr Val

Thr Leu Phe

140

120

ΕΡ 2 379 594 Β1

<td> Pro</td><td> Pro</td><td> Ser</td><td> Ser</td><td> Glu</td><td> Glu</td><td> Leu</td><td> Gin</td><td> Alá</td><td> Asn</td><td> Lys</td><td> Alá</td><td> Thr</td><td> Leu</td><td> Val</td><td> Cys</td>

<td> 145</td><td></td><td></td><td></td><td></td><td> 150</td><td></td><td></td><td></td><td></td><td> 155</td><td></td><td></td><td></td><td></td><td> 160</td>

<td> Leu</td><td> Ile</td><td> Ser</td><td> Asp</td><td> Phe</td><td> Tyr</td><td> Pro</td><td> Gly</td><td> Alá</td><td> Val</td><td> Thr</td><td> Val</td><td> Alá</td><td> Trp</td><td> Lys</td><td> Alá</td>

<td></td><td></td><td></td><td></td><td> 165</td><td></td><td></td><td></td><td></td><td> 170</td><td></td><td></td><td></td><td></td><td> 175</td><td></td>

<td> Asp</td><td> Gly</td><td> Ser</td><td> Pro</td><td> Val</td><td> Lys</td><td> Alá</td><td> Gly</td><td> Val</td><td> Glu</td><td> Thr</td><td> Thr</td><td> Lys</td><td> Pro</td><td> Ser</td><td> Lys</td>

<td></td><td></td><td></td><td> 180</td><td></td><td></td><td></td><td></td><td> 185</td><td></td><td></td><td></td><td></td><td> 190</td><td></td><td></td>

<td> Gin</td><td> Ser</td><td> Asn</td><td> Asn</td><td> Lys</td><td> Tyr</td><td> Alá</td><td> Alá</td><td> Ser</td><td> Ser</td><td> Tyr</td><td> Leu</td><td> Ser</td><td> Leu</td><td> Thr</td><td> Pro</td>

<td></td><td></td><td> 195</td><td></td><td></td><td></td><td></td><td> 200</td><td></td><td></td><td></td><td></td><td> 205</td><td></td><td></td><td></td>

<td> Glu</td><td> Gin</td><td> Trp</td><td> Lys</td><td> Ser</td><td> His</td><td> Arg</td><td> Ser</td><td> Tyr</td><td> Ser</td><td> Cys</td><td> Gin</td><td> Val</td><td> Thr</td><td> His</td><td> Glu</td>

<td></td><td> 210</td><td></td><td></td><td></td><td></td><td> 215</td><td></td><td></td><td></td><td></td><td> 220</td><td></td><td></td><td></td><td></td>

<td> Gly</td><td> Ser</td><td> Thr</td><td> Val</td><td> Glu</td><td> Lys</td><td> Thr</td><td> Val</td><td> Alá</td><td> Pro</td><td> Thr</td><td> Glu</td><td> Cys</td><td> Ser</td><td></td><td></td>

225 230 235 <210> 22 <211> 238 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 22

121

ΕΡ 2 379 594 Β1

Met Asp Met Arg Val Pro Alá Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Alá Arg Cys Gin Ser Val Leu Thr Gin Pro Pro Ser Alá 20 25 30

Ser Gly Thr Pro Gly Gin Arg Val Thr Ile Ser Cys Ser Gly Ser Ser 35 40 45

Ser Asn Ile Gly Ser Asn Thr Val Asn Trp Tyr Gin Gin Leu Pro Gly 50 55 60

Thr Alá Pro Lys Leu Leu Ile Tyr Thr Asn Asn Gin Arg Pro Ser Gly 65 70 75 80

Val Pro Asp Arg Phe Ser Gly Ser Lys Ser Gly Thr Ser Alá Ser Leu 85 90 95

Alá Ile Ser Gly Leu Gin Ser Glu Asp Glu Alá Asp Phe Tyr Cys Alá 100 105 110

Alá Arg Asp Glu Ser Leu Asn Gly Val Val Phe Gly Gly Gly Thr Lys

115

120

125

Leu Thr Val Leu Gly Gin Pro Lys Alá Asn Pro Thr Val Thr Leu Phe 130 135 140

Pro Pro Ser Ser Glu Glu Leu Gin Alá Asn Lys Alá Thr Leu Val Cys 145 150 155 160

Leu Ile Ser Asp Phe Tyr Pro Gly Alá Val Thr Val Alá Trp Lys Alá 165 170 175

Asp Gly Ser Pro Val Lys Alá Gly Val Glu Thr Thr Lys Pro Ser Lys 180 185 190

Gin Ser Asn Asn Lys Tyr Alá Alá Ser Ser Tyr Leu Ser Leu Thr Pro 195 200 205

Glu Gin Trp Lys Ser His Arg Ser Tyr Ser Cys Gin Val Thr His Glu 210 215 220

Gly Ser Thr Val Glu Lys Thr Val Alá Pro Thr Glu Cys Ser 225 230 235

122

ΕΡ 2 379 594 Β1 <210> 23 <211> 238 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 23

Met Asp Met Arg Val Pro Alá 1 5

Leu Arg Gly Alá Arg Cys Gin 20

Ser Gly Thr Pro Gly Gin Arg 35

Ser Asn Ile Gly Ser Asn Tyr 50 55

Alá Alá Pro Lys Leu Leu Ile 65 70

Val

Pro Asp Arg Phe 85

Ser

Gly

Gin Leu Leu Gly Leu Leu Leu Leu Trp 10 15

Ser Val Leu Thr Gin Pro Pro Ser Alá 25 30

Val Thr Ile Ser Cys Ser Gly Ser Ser 40 45

Val Tyr Trp Tyr Gin Gin Leu Pro Gly 60

Phe Arg Asn Asn Gin Arg Pro Ser Gly 75 80

Ser Lys

Ser Gly Thr Ser Alá 90

Ser Leu 95

123

ΕΡ 2 379 594 Β1

<td> Alá</td><td> Ile</td><td colspan="2"> Ser Gly 100</td><td> Leu</td><td> Arg</td><td> Ser</td><td colspan="2"> Glu Asp 105</td><td> Glu</td><td colspan="2"> Alá Asp</td><td> Tyr</td><td colspan="2"> Tyr Cys 110</td><td> Alá</td>

<td> Alá</td><td> Trp</td><td> Asp</td><td> Asp</td><td> Ser</td><td> Leu</td><td> Ser</td><td> Gly</td><td> Trp</td><td> Val</td><td> Phe</td><td> Gly</td><td> Gly</td><td> Gly</td><td> Thr</td><td> Lys</td>

<td></td><td></td><td> 115</td><td></td><td></td><td></td><td></td><td> 120</td><td></td><td></td><td></td><td></td><td> 125</td><td></td><td></td><td></td>

<td> Leu</td><td> Thr</td><td> Val</td><td> Leu</td><td> Gly</td><td> Gin</td><td> Pro</td><td> Lys</td><td> Alá</td><td> Asn</td><td> Pro</td><td> Thr</td><td> Val</td><td> Thr</td><td> Leu</td><td> Phe</td>

<td></td><td> 130</td><td></td><td></td><td></td><td></td><td> 135</td><td></td><td></td><td></td><td></td><td> 140</td><td></td><td></td><td></td><td></td>

<td> Pro</td><td> Pro</td><td> Ser</td><td> Ser</td><td> Glu</td><td> Glu</td><td> Leu</td><td> Gin</td><td> Alá</td><td> Asn</td><td> Lys</td><td> Alá</td><td> Thr</td><td> Leu</td><td> Val</td><td> Cys</td>

<td> 145</td><td></td><td></td><td></td><td></td><td> 150</td><td></td><td></td><td></td><td></td><td> 155</td><td></td><td></td><td></td><td></td><td> 160</td>

<td> Leu</td><td> Ile</td><td> Ser</td><td> Asp</td><td> Phe</td><td> Tyr</td><td> Pro</td><td> Gly</td><td> Alá</td><td> Val</td><td> Thr</td><td> Val</td><td> Alá</td><td> Trp</td><td> Lys</td><td> Alá</td>

<td></td><td></td><td></td><td></td><td> 165</td><td></td><td></td><td></td><td></td><td> 170</td><td></td><td></td><td></td><td></td><td> 175</td><td></td>

<td> Asp</td><td> Gly</td><td> Ser</td><td> Pro</td><td> Val</td><td> Lys</td><td> Alá</td><td> Gly</td><td> Val</td><td> Glu</td><td> Thr</td><td> Thr</td><td> Lys</td><td> Pro</td><td> Ser</td><td> Lys</td>

<td></td><td></td><td></td><td> 180</td><td></td><td></td><td></td><td></td><td> 185</td><td></td><td></td><td></td><td></td><td> 190</td><td></td><td></td>

<td> Gin</td><td> Ser</td><td> Asn</td><td> Asn</td><td> Lys</td><td> Tyr</td><td> Alá</td><td> Alá</td><td> Ser</td><td> Ser</td><td> Tyr</td><td> Leu</td><td> Ser</td><td> Leu</td><td> Thr</td><td> Pro</td>

<td></td><td></td><td> 195</td><td></td><td></td><td></td><td></td><td> 200</td><td></td><td></td><td></td><td></td><td> 205</td><td></td><td></td><td></td>

<td> Glu</td><td> Gin</td><td> Trp</td><td> Lys</td><td> Ser</td><td> His</td><td> Arg</td><td> Ser</td><td> Tyr</td><td> Ser</td><td> Cys</td><td> Gin</td><td> Val</td><td> Thr</td><td> His</td><td> Glu</td>

<td></td><td> 210</td><td></td><td></td><td></td><td></td><td> 215</td><td></td><td></td><td></td><td></td><td> 220</td><td></td><td></td><td></td><td></td>

<td> Gly</td><td> Ser</td><td> Thr</td><td> Val</td><td> Glu</td><td> Lys</td><td> Thr</td><td> Val</td><td> Alá</td><td> Pro</td><td> Thr</td><td> Glu</td><td> Cys</td><td> Ser</td><td></td><td></td>

225 230 235 <210> 24 <211> 241 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 24

124

ΕΡ 2 379 594 Β1

Met Asp Met Arg Val Pro Alá Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Alá Arg Cys Asp Ile Thr Leu Thr Gin Thr Pro Leu Ser 20 25 30

Leu Ser Val Ser Pro Gly Gin Pro Alá Ser Ile Ser Cys Lys Ser Ser 35 40 45

Gin Ser Leu Leu His Ser Asp Gly Arg Asn Tyr Leu Tyr Trp Tyr Leu 50 55 60

Gin Lys Pro Gly Gin Pro Pro Gin Leu Leu Ile Tyr Glu Val Ser Asn

125

ΕΡ 2 379 594 Β1

<td colspan="4"> 65</td><td colspan="4"> 70</td><td colspan="6"> 75</td><td colspan="2"> 80</td>

<td> Arg</td><td> Phe</td><td> Ser</td><td> Gly</td><td> Leu</td><td> Pro</td><td> Asp</td><td> Arg</td><td> Phe</td><td> Ser</td><td> Gly</td><td> Ser</td><td> Gly</td><td> Ser</td><td> Gly</td><td> Thr</td>

<td></td><td></td><td></td><td></td><td> 85</td><td></td><td></td><td></td><td></td><td> 90</td><td></td><td></td><td></td><td></td><td> 95</td><td></td>

<td> Asp</td><td> Phe</td><td> Thr</td><td> Leu</td><td> Lys</td><td> Ile</td><td> Ser</td><td> Arg</td><td> Val</td><td> Glu</td><td> Alá</td><td> Glu</td><td> Asp</td><td> Val</td><td> Gly</td><td> Ile</td>

<td></td><td></td><td></td><td> 100</td><td></td><td></td><td></td><td></td><td> 105</td><td></td><td></td><td></td><td></td><td> 110</td><td></td><td></td>

<td> Tyr</td><td> Tyr</td><td> Cys</td><td> Met</td><td> Gin</td><td> Ser</td><td> Phe</td><td> Pro</td><td> Leu</td><td> Pro</td><td> Leu</td><td> Thr</td><td> Phe</td><td> Gly</td><td> Gly</td><td> Gly</td>

<td></td><td></td><td> 115</td><td></td><td></td><td></td><td></td><td> 120</td><td></td><td></td><td></td><td></td><td> 125</td><td></td><td></td><td></td>

<td> Thr</td><td> Lys</td><td> Val</td><td> Glu</td><td> Ile</td><td> Lys</td><td> Arg</td><td> Thr</td><td> Val</td><td> Alá</td><td> Alá</td><td> Pro</td><td> Ser</td><td> Val</td><td> Phe</td><td> Ile</td>

<td></td><td> 130</td><td></td><td></td><td></td><td></td><td> 135</td><td></td><td></td><td></td><td></td><td> 140</td><td></td><td></td><td></td><td></td>

<td> Phe</td><td> Pro</td><td> Pro</td><td> Ser</td><td> Asp</td><td> Glu</td><td> Gin</td><td> Leu</td><td> Lys</td><td> Ser</td><td> Gly</td><td> Thr</td><td> Alá</td><td> Ser</td><td> Val</td><td> Val</td>

<td> 145</td><td></td><td></td><td></td><td></td><td> 150</td><td></td><td></td><td></td><td></td><td> 155</td><td></td><td></td><td></td><td></td><td> 160</td>

<td> Cys</td><td> Leu</td><td> Leu</td><td> Asn</td><td> Asn</td><td> Phe</td><td> Tyr</td><td> Pro</td><td> Arg</td><td> Glu</td><td> Alá</td><td> Lys</td><td> Val</td><td> Gin</td><td> Trp</td><td> Lys</td>

<td></td><td></td><td></td><td></td><td> 165</td><td></td><td></td><td></td><td></td><td> 170</td><td></td><td></td><td></td><td></td><td> 175</td><td></td>

<td> Val</td><td> Asp</td><td> Asn</td><td> Alá</td><td> Leu</td><td> Gin</td><td> Ser</td><td> Gly</td><td> Asn</td><td> Ser</td><td> Gin</td><td> Glu</td><td> Ser</td><td> Val</td><td> Thr</td><td> Glu</td>

<td></td><td></td><td></td><td> 180</td><td></td><td></td><td></td><td></td><td> 185</td><td></td><td></td><td></td><td></td><td> 190</td><td></td><td></td>

<td> Gin</td><td> Asp</td><td> Ser</td><td> Lys</td><td> Asp</td><td> Ser</td><td> Thr</td><td> Tyr</td><td> Ser</td><td> Leu</td><td> Ser</td><td> Ser</td><td> Thr</td><td> Leu</td><td> Thr</td><td> Leu</td>

<td></td><td></td><td> 195</td><td></td><td></td><td></td><td></td><td> 200</td><td></td><td></td><td></td><td></td><td> 205</td><td></td><td></td><td></td>

<td> Ser</td><td> Lys</td><td> Alá</td><td> Asp</td><td> Tyr</td><td> Glu</td><td> Lys</td><td> His</td><td> Lys</td><td> Val</td><td> Tyr</td><td> Alá</td><td> Cys</td><td> Glu</td><td> Val</td><td> Thr</td>

<td></td><td> 210</td><td></td><td></td><td></td><td></td><td> 215</td><td></td><td></td><td></td><td></td><td> 220</td><td></td><td></td><td></td><td></td>

<td> His</td><td> Gin</td><td> Gly</td><td> Leu</td><td> Ser</td><td> Ser</td><td> Pro</td><td> Val</td><td> Thr</td><td> Lys</td><td> Ser</td><td> Phe</td><td> Asn</td><td> Arg</td><td> Gly</td><td> Glu</td>

<td> 225</td><td></td><td></td><td></td><td></td><td> 230</td><td></td><td></td><td></td><td></td><td> 235</td><td></td><td></td><td></td><td></td><td> 240</td>

<td> Cys</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<td> &lt;210&gt; 25 &lt;211&gt; 238 &lt;212&gt; PRT</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<td colspan="3"> &lt;213&gt; Artificial Sequence</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 25

126

ΕΡ 2 379 594 Β1

Met Asp Met Arg Val 1 5

Pro Alá Gin Leu Leu 10

Gly Leu Leu Leu Leu Trp 15

Leu Arg Gly Alá Arg 20

Cys Gin Ser Val Leu 25

Thr Gin Pro Pro Ser Val 30

Ser Alá Alá Pro Gly 35

Gin Lys Val Thr Ile 40

Ser Cys Ser Gly Ser Ser 45

Ser Asn Ile Gly Asn 50

Thr Alá Pro Lys Leu 65

Ile Pro Asp Arg Phe 85

Gly Ile Thr Gly Leu 100

Thr Trp Asp Ser Arg 115

Leu Thr Val Leu Gly 130

Pro Pro Ser Ser Glu 145

Leu Ile Ser Asp Phe 165

Asp Gly Ser Pro Val 180

Gin Ser Asn Asn Lys 195

Glu Gin Trp Lys Ser 210

Asn Tyr Val Ser Trp 55

Leu Ile Tyr Asp Asn 70

Ser Gly Ser Lys Ser 90

Gin Thr Gly Asp Glu 105

Leu Ser Alá Val Val 120

Gin Pro Lys Alá Asn 135

Glu Leu Gin Alá Asn 150

Tyr Pro Gly Alá Val 170

Lys Alá Gly Val Glu 185

Tyr Alá Alá Ser Ser 200

His Arg Ser Tyr Ser 215

Lys Thr Val Alá Pro 230

Tyr Gin Gin Leu Pro Gly 60

Asn Lys Arg Pro Ser Gly 75 80

Gly Thr Ser Alá Thr Leu 95

Alá Asp Tyr Tyr Cys Gly 110

Phe Gly Gly Gly Thr Lys 125

Pro Thr Val Thr Leu Phe 140

Lys Alá Thr Leu Val Cys 155 160

Thr Val Alá Trp Lys Alá 175

Thr Thr Lys Pro Ser Lys 190

Tyr Leu Ser Leu Thr Pro 205

Cys Gin Val Thr His Glu 220

Thr Glu Cys Ser 235

Gly Ser Thr Val Glu 225 <210> 26 <211> 236 <212> PRT

127

EP 2 379 594 Β1 <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 26

Met Asp Met Arg Val Pro Ala Gin Leu Leu Gly Leu Leu Leu Leu Trp

128

ΕΡ 2 379 594 Β1

10

Thr Gin Ser Pro Ser Ser

Leu Arg Gly Alá 20

Arg Cys Asp Ile Gin Met 25

Leu Ser Alá Ser 35

Val Gly Asp Arg Val Thr 40

Ile Thr Cys Arg Alá Ser 45

Gin Gly Ile Arg 50

Lys Asp Leu Gly Trp Tyr 55

Gin Gin Lys Pro Gly Lys 60

Alá Pro Lys Arg 65

Leu Ile Tyr Gly Alá Ser 70

Ser Leu Gin Ser Gly Val 75 80

Pro Ser Arg Phe

Ser Gly Ser Gly Ser Gly 85 90

Thr Glu Phe Thr Leu Thr 95

Ile Ser Ser Leu 100

Gin Pro Glu Asp Phe Alá 105

Thr Tyr Tyr Cys Leu Gin 110

Tyr Asn Ser Phe 115

Pro Trp Thr Phe Gly Gin 120

Gly Thr Lys Val Glu Ile 125

Lys Arg Thr Val 130

Alá Alá Pro Ser Val Phe 135

Ile Phe Pro Pro Ser Asp 140

Glu Gin Leu Lys 145

Ser Gly Thr Alá Ser Val 150

Val Cys Leu Leu Asn Asn 155 160

Phe Tyr Pro Arg

Glu Alá Lys Val Gin Trp 165 170

Lys Val Asp Asn Alá Leu 175

Gin Ser Gly Asn 180

Ser Gin Glu Ser Val Thr 185

Glu Gin Asp Ser Lys Asp 190

Ser Thr Tyr Ser 195

Leu Ser Ser Thr Leu Thr 200

Leu Ser Lys Alá Asp Tyr 205

Glu Lys His Lys 210

Val Tyr Alá Cys Glu Val 215

Thr His Gin Gly Leu Ser 220

Lys Ser Phe Asn Arg Gly 230

Glu Cys 235

Ser Pro Val Thr 225 <210> 27 <211> 235 <212> PRT

129

ΕΡ 2 379 594 Β1 <213> Artificial Sequence <220>

<221> source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 27

Met Glu Thr Pro Alá Gin Leu Leu Phe Leu Leu Leu Leu Trp Leu Pro

10 15

Asp Thr Thr Gly Glu Ile Val Leu Thr Gin Ser Pro Gly Thr Leu Ser 20 25 30

Leu Ser Pro Gly Glu Arg Alá Thr Leu Ser Cys Arg Alá Ser Gin Ser 35 40 45

Val Ser Ser Gly Tyr Leu Thr Trp Tyr Gin Gin Lys Pro Gly Gin Alá 50 55 60

Pro Arg Leu Leu Ile Tyr Gly Alá Ser Ser Arg Alá Thr Gly Ile Pro 65 70 75 80

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile 85 90 95

Ser Arg Leu Glu Pro Glu Asp Phe Alá Val Tyr Tyr Cys Gin Gin Tyr 100 105 110

Gly Asn Ser Leu Cys Arg Phe Gly Gin Gly Thr Lys Leu Glu Ile Lys 115 120 125

Arg Thr Val Alá Alá Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu 130 135 140

Gin Leu Lys Ser Gly Thr Alá Ser Val Val Cys Leu Leu Asn Asn Phe 145 150 155 160

Tyr Pro Arg Glu Alá Lys Val Gin Trp Lys Val Asp Asn Alá Leu Gin 165 170 175

Ser Gly Asn Ser Gin Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser 180 185 190

Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Alá Asp Tyr Glu 195 200 205

Lys His Lys Val Tyr Alá Cys Glu Val Thr His Gin Gly Leu Ser Ser 210 215 220

Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys 225 230 235 <210> 28 <211> 235 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 28

130

ΕΡ 2 379 594 Β1

Met

Asp

Leu

Val

Pro

Asp

Ser

Gly

Arg

Gin

145

Tyr

Ser

Thr

Lys <210> 29 <211> 478 <212> PRT

Glu Thr Pro Alá Gin Leu Leu Phe Leu Leu Leu Leu Trp Leu Pro 5 10 15

Thr Thr Gly Glu Ile Val Leu Thr Gin Ser Pro Gly Thr Leu Ser 20 25 30

Ser Pro Gly Glu Arg Alá Thr Leu Ser Cys Arg Alá Ser Gin Ser 35 40 45

Ser Ser Gly Tyr Leu Thr Trp Tyr Gin Gin Lys Pro Gly Gin Alá 50 55 60

Arg Leu Leu Ile Tyr Gly Alá Ser Ser Arg Alá Thr Gly Ile Pro 70 75 80

Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile 85 90 95

Arg Leu Glu Pro Glu Asp Phe Alá Val Tyr Tyr Cys Gin Gin Tyr 100 105 110

Asn Ser Leu Ser Arg Phe Gly Gin Gly Thr Lys Leu Glu Ile Lys 115 120 125

Thr Val Alá Alá Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu 130 135 140

Leu Lys Ser Gly Thr Alá Ser Val Val Cys Leu Leu Asn Asn Phe 150 155 160

Pro Arg Glu Alá Lys Val Gin Trp Lys Val Asp Asn Alá Leu Gin 165 170 175

Gly Asn Ser Gin Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser 180 185 190

Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Alá Asp Tyr Glu 195 200 205

His Lys Val Tyr Alá Cys Glu Val Thr His Gin Gly Leu Ser Ser 210 215 220

Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys 225 230 235

131

ΕΡ 2 379 594 Β1 <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 29

Met Asp Met Arg Val Pro Alá Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Alá Arg Cys Gin Val Gin Leu Val Glu Ser Gly Gly Gly 20 25 30

Val Val Gin Pro Gly Arg Ser Leu Arg Leu Ser Cys Alá Alá Ser Gly 35 40 45

Phe Thr Phe Ser Ser Phe Gly Met His Trp Val Arg Gin Alá Pro Gly 50 55 60

Lys Gly Leu Glu Trp Val Alá Val Ile Ser Phe Asp Gly Ser Ile Lys 65 70 75 80

Tyr Ser Val Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn 85 90 95

Ser Lys Asn Thr Leu Phe Leu Gin Met Asn Ser Leu Arg Alá Glu Asp 100 105 110

Thr Alá Val Tyr Tyr Cys Alá Arg Asp Arg Leu Asn Tyr Tyr Asp Ser 115 120 125

Ser Gly Tyr Tyr His Tyr Lys Tyr Tyr Gly Met Alá Val Trp Gly Gin 130 135 140

Gly Thr Thr Val Thr Val Ser Ser Alá Ser Thr Lys Gly Pro Ser Val 145 150 155 160

Phe Pro Leu Alá Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Alá Alá 165 170 175

Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser 180 185 190

132

ΕΡ 2 379 594 Β1

Trp Asn Ser Gly Alá Leu Thr Ser Gly Val His Thr Phe Pro Alá Val 195 200 205

Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro 210 215 220

Ser Ser Asn Phe Gly Thr Gin Thr Tyr Thr Cys Asn Val Asp His Lys 225 230 235 240

Pro Ser Asn Thr Lys Val Asp Lys Thr Val Glu Arg Lys Cys Cys Val 245 250 255

Glu Cys Pro Pro Cys Pro Alá Pro Pro Val Alá Gly Pro Ser Val Phe 260 265 270

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro 275 280 285

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val 290 295 300

Gin Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Alá Lys Thr 305 310 315 320

Lys Pro Arg Glu Glu Gin Phe Asn Ser Thr Phe Arg Val Val Ser Val 325 330 335

Leu Thr Val Val His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys 340 345 350

Lys Val Ser Asn Lys Gly Leu Pro Alá Pro Ile Glu Lys Thr Ile Ser 355 360 365

Lys Thr Lys Gly Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro 370 375 380

Ser Arg Glu Glu Met Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val 385 390 395 400

Lys Gly Phe Tyr Pro Ser Asp Ile Alá Val Glu Trp Glu Ser Asn Gly 405 410 415

Gin Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Met Leu Asp Ser Asp 420 425 430

Gly

Ser Phe Phe 435

Leu

Tyr

Ser Lys 440

Leu

Thr Val Asp

Lys

445

Ser Arg Trp

Gin Gin Gly Asn Val Phe Ser Cys Ser Val Met His Glu Alá Leu His

133

ΕΡ 2 379 594 Β1

460

450

Ser Leu Ser Leu Ser Pro Gly Lys

475

455

Asn His Tyr Thr Gin Lys 465 470 <210> 30 <211> 479 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 30

134

ΕΡ 2 379 594 Β1

Met Asp Met Arg Val Pro Alá Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Alá Arg Cys Glu Val Gin Leu Val Glu Ser Gly Gly Gly 20 25 30

Leu Val Lys Pro Gly Gly Ser Leu Arg Leu Ser Cys Alá Alá Ser Gly 35 40 45

Phe Thr Phe Ser Asn Alá Trp Met Ser Trp Val Arg Gin Alá Pro Gly 50 55 60

Lys Gly Leu Glu Trp Val Gly Arg Ile Lys Ser Thr Thr Asp Gly Gly 65 70 75 80

Thr Thr Asp Tyr Alá Alá Pro Val Lys Gly Arg Phe Thr Ile Ser Arg 85 90 95

Asp Asp Ser Lys Asn Thr Leu Tyr Leu Gin Met Asn Ser Leu Lys Thr 100 105 110

Glu Asp Thr Alá Val Tyr Tyr Cys Thr Thr Asp Arg Thr Gly Tyr Ser 115 120 125

Ile Ser Trp Ser Ser Tyr Tyr Tyr Tyr Tyr Gly Met Asp Val Trp Gly 130 135 140

Gin Gly Thr Thr Val Thr Val Ser Ser Alá Ser Thr Lys Gly Pro Ser 145 150 155 160

Val Phe Pro Leu Alá Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Alá 165 170 175

Alá Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val 180 185 190

135

EP 2 379 594 Β1

Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala 195 200 205

Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val 210 215 220

Pro Ser Ser Asn Phe Gly Thr Gin Thr Tyr Thr Cys Asn Val Asp His 225 230 235 240

Lys Pro Ser Asn Thr Lys Val Asp Lys Thr Val Glu Arg Lys Cys Cys 245 250 255

Val Glu Cys Pro Pro Cys Pro Ala Pro Pro Val Ala Gly Pro Ser Val 260 265 270

Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr 275 280 285

Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu 290 295 300

Val Gin Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys 305 310 315 320

Thr Lys Pro Arg Glu Glu Gin Phe Asn Ser Thr Phe Arg Val Val Ser 325 330 335

Val Leu Thr Val Val His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys 340 345 350

Cys Lys Val Ser Asn Lys Gly Leu Pro Ala Pro Ile Glu Lys Thr Ile 355 360 365

Ser Lys Thr Lys Gly Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro 370 375 380

Pro Ser Arg Glu Glu Met Thr Lys Asn Gin Val Ser Leu Thr Cys Leu 385 390 395 400

Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn 405 410 415

Gly Gin Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Met Leu Asp Ser 420 425 430

Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg 435 440 445

136

ΕΡ 2 379 594 Β1

Trp Gin Gin Gly Asn Val 450

Phe Ser Cys Ser Val 455

Met His Glu Alá Leu 460

His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu Ser Pro Gly Lys 465 470 475 <210> 31 <211> 478 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 31

137

EP 2 379 594 Β1

Met Asp Met Arg Val Pro Alá Gin 1 5

Leu Leu Gly Leu Leu Leu Leu Trp 10 15

Leu Arg Gly Alá Arg Cys Glu Val 20

Gin Leu Leu Glu Ser Gly Gly Gly 25 30

Leu Val Gin Pro Gly Glu Ser Leu 35 40

Arg Leu Ser Cys Alá Alá Ser Gly 45

Phe Thr Phe Ser Ser Tyr Alá Met 50 55

Ser Trp Val Arg Gin Alá Pro Gly 60

Lys Gly Leu Glu Trp Val Ser Alá 65 70

Ile Ser Gly Ser Gly Gly Arg Thr 75 80

Tyr Tyr Alá Asp Ser Val Lys Gly 85

Arg Phe Thr Ile Ser Arg Asp Asn 90 95

Ser Lys Asn Thr Leu Tyr Leu Gin 100

Met Asn Ser Leu Arg Alá Glu Asp 105 110

Thr Alá Val Tyr Tyr Cys Alá Lys 115 120

Asp Gin Arg Glu Val Gly Pro Tyr 125

Ser Ser Gly Trp Tyr Asp Tyr Tyr 130 135

Tyr Gly Met Asp Val Trp Gly Gin 140

Gly Thr Thr Val Thr Val Ser Ser 145 150

Alá Ser Thr Lys Gly Pro Ser Val 155 160

Phe Pro Leu Alá Pro Cys Ser Arg 165

Ser Thr Ser Glu Ser Thr Alá Alá 170 175

Leu Gly Cys Leu Val Lys Asp Tyr 180

Phe Pro Glu Pro Val Thr Val Ser 185 190

138

ΕΡ 2 379 594 Β1

Trp Asn Ser Gly Alá Leu Thr Ser Gly Val His Thr Phe Pro Alá Val 195 200 205

Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro 210 215 220

Ser Ser Asn Phe Gly Thr Gin Thr Tyr Thr Cys Asn Val Asp His Lys 225 230 235 240

Pro Ser Asn Thr Lys Val Asp Lys Thr Val Glu Arg Lys Cys Cys Val 245 250 255

Glu Cys Pro Pro Cys Pro Alá Pro Pro Val Alá Gly Pro Ser Val Phe 260 265 270

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro 275 280 285

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val 290 295 300

Gin Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Alá Lys Thr 305 310 315 320

Lys Pro Arg Glu Glu Gin Phe Asn Ser Thr Phe Arg Val Val Ser Val 325 330 335

Leu Thr Val Val His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys 340 345 350

Lys Val Ser Asn Lys Gly Leu Pro Alá Pro Ile Glu Lys Thr Ile Ser 355 360 365

Lys Thr Lys Gly Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro 370 375 380

Ser Arg Glu Glu Met Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val 385 390 395 400

Lys

Gly Phe

Tyr Pro 405

Ser Asp

Ile Alá Val 410

Glu Trp Glu

Ser Asn Gly 415

Gin Pro Glu Asn Asn Tyr Lys 420

Thr Thr Pro

Pro Met

425

Leu Asp 430

Ser Asp

Gly

Ser Phe Phe 435

Leu

Tyr

Ser

Lys

440

Leu

Thr Val Asp

Lys

445

Ser Arg Trp

139

ΕΡ 2 379 594 Β1

Gin Gin Gly Asn Val Phe 450

Ser Cys Ser Val Met 455

His Glu Alá Leu His 460

Asn His Tyr Thr Gin Lys Ser Leu Ser Leu Ser Pro Gly Lys 465 470 475 <210> 32 <211> 478 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 32

140

ΕΡ 2 379 594 Β1

Met Asp Met Arg Val Pro Alá Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Alá Arg Cys Gin Val Gin Leu Val Gin Ser Gly Alá Glu 20 25 30

Val Lys Lys Pro Gly Alá Ser Val Lys Val Ser Cys Lys Alá Ser Gly 35 40 45

Tyr Thr Phe Thr Gly Tyr Tyr Met His Trp Val Arg Gin Alá Pro Gly 50 55 60

Gin Gly Leu Glu Trp Met Gly Trp Ile Asn Pro Asn Ser Gly Gly Thr 65 70 75 80

Asn Tyr Alá Gin Lys Phe Gin Gly Arg Val Thr Met Thr Arg Asp Thr 85 90 95

Ser Ile Ser Thr Alá Tyr Met Glu Leu Ser Arg Leu Arg Ser Asp Asp 100 105 110

Thr Alá Val Tyr Phe Cys Alá Arg Asp Gin Met Ser Ile Ile Met Leu 115 120 125

Arg Gly Val Phe Pro Pro Tyr Tyr Tyr Gly Met Asp Val Trp Gly Gin 130 135 140

Gly Thr Thr Val Thr Val Ser Ser Alá Ser Thr Lys Gly Pro Ser Val 145 150 155 160

Phe Pro Leu Alá Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Alá Alá 165 170 175

Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser

141

ΕΡ 2 379 594 Β1

180 185 190

Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val 195 200 205

Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro 210 215 220

Ser Ser Asn Phe Gly Thr Gin Thr Tyr Thr Cys Asn Val Asp His Lys 225 230 235 240

Pro Ser Asn Thr Lys Val Asp Lys Thr Val Glu Arg Lys Cys Cys Val 245 250 255

Glu Cys Pro Pro Cys Pro Ala Pro Pro Val Ala Gly Pro Ser Val Phe 260 265 270

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro 275 280 285

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val 290 295 300

Gin Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr 305 310 315 320

Lys Pro Arg Glu Glu Gin Phe Asn Ser Thr Phe Arg Val Val Ser Val 325 330 335

Leu Thr Val Val His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys 340 345 350

Lys Val Ser Asn Lys Gly Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser 355 360 365

Lys Thr Lys Gly Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro 370 375 380

Ser Arg Glu Glu Met Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val 385 390 395 400

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly 405 410 415

Gin Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Met Leu Asp Ser Asp 420 425 430

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp 435 440 445

142

ΕΡ 2 379 594 Β1

Gin Gin Gly Asn Val Phe Ser Cys Ser Val Met His Glu Alá Leu His 450 455 460 ⁵ Asn His Tyr Thr Gin Lys Ser Leu Ser Leu Ser Pro Gly Lys

465 470 475 <210> 33 <211> 477 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 33

<td> 20</td><td> Met</td><td> Asp</td><td> Met</td><td> Arg</td><td> Val</td><td> Pro</td><td> Alá</td><td> Gin</td><td> Leu</td><td> Leu</td><td> Gly</td><td> Leu</td><td> Leu</td><td> Leu</td><td> Leu</td><td> Trp</td>

<td></td><td> 1</td><td></td><td></td><td></td><td> 5</td><td></td><td></td><td></td><td></td><td> 10</td><td></td><td></td><td></td><td></td><td> 15</td><td></td>

<td></td><td> Leu</td><td> Arg</td><td> Gly</td><td> Alá</td><td> Arg</td><td> Cys</td><td> Gin</td><td> Val</td><td> Gin</td><td> Leu</td><td> Val</td><td> Glu</td><td> Ser</td><td> Gly</td><td> Gly</td><td> Gly</td>

<td></td><td></td><td></td><td></td><td> 20</td><td></td><td></td><td></td><td></td><td> 25</td><td></td><td></td><td></td><td></td><td> 30</td><td></td><td></td>

<td> 25</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<td></td><td> Val</td><td> Val</td><td> Gin</td><td> Pro</td><td> Gly</td><td> Arg</td><td> Ser</td><td> Leu</td><td> Arg</td><td> Leu</td><td> Ser</td><td> Cys</td><td> Alá</td><td> Alá</td><td> Ser</td><td> Gly</td>

<td></td><td></td><td></td><td> 35</td><td></td><td></td><td></td><td></td><td> 40</td><td></td><td></td><td></td><td></td><td> 45</td><td></td><td></td><td></td>

<td> 30</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<td></td><td> Phe</td><td> Thr</td><td> Phe</td><td> Ser</td><td> Ser</td><td> Tyr</td><td> Gly</td><td> Met</td><td> His</td><td> Trp</td><td> Val</td><td> Arg</td><td> Gin</td><td> Alá</td><td> Pro</td><td> Gly</td>

<td></td><td></td><td> 50</td><td></td><td></td><td></td><td></td><td> 55</td><td></td><td></td><td></td><td></td><td> 60</td><td></td><td></td><td></td><td></td>

<td> 35</td><td> Lys</td><td> Gly</td><td> Leu</td><td> Glu</td><td> Trp</td><td> Val</td><td> Alá</td><td> Val</td><td> Ile</td><td> Ser</td><td> Tyr</td><td> Asp</td><td> Gly</td><td> Ser</td><td> His</td><td> Glu</td>

<td></td><td> 65</td><td></td><td></td><td></td><td></td><td> 70</td><td></td><td></td><td></td><td></td><td> 75</td><td></td><td></td><td></td><td></td><td> 80</td>

<td></td><td> Ser</td><td> Tyr</td><td> Alá</td><td> Asp</td><td> Ser</td><td> Val</td><td> Lys</td><td> Gly</td><td> Arg</td><td> Phe</td><td> Thr</td><td> Ile</td><td> Ser</td><td> Arg</td><td> Asp</td><td> Ile</td>

<td></td><td></td><td></td><td></td><td></td><td> 85</td><td></td><td></td><td></td><td></td><td> 90</td><td></td><td></td><td></td><td></td><td> 95</td><td></td>

<td> 40</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<td></td><td> Ser</td><td> Lys</td><td> Asn</td><td> Thr</td><td> Leu</td><td> Tyr</td><td> Leu</td><td> Gin</td><td> Met</td><td> Asn</td><td> Ser</td><td> Leu</td><td> Arg</td><td> Alá</td><td> Glu</td><td> Asp</td>

<td></td><td></td><td></td><td></td><td> 100</td><td></td><td></td><td></td><td></td><td> 105</td><td></td><td></td><td></td><td></td><td> 110</td><td></td><td></td>

<td> 45</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<td></td><td> Thr</td><td> Alá</td><td> Val</td><td> Tyr</td><td> Phe</td><td> Cys</td><td> Alá</td><td> Arg</td><td> Glu</td><td> Arg</td><td> Lys</td><td> Arg</td><td> Val</td><td> Thr</td><td> Met</td><td> Ser</td>

<td></td><td></td><td></td><td> 115</td><td></td><td></td><td></td><td></td><td> 120</td><td></td><td></td><td></td><td></td><td> 125</td><td></td><td></td><td></td>

<td></td><td> Thr</td><td> Leu</td><td> Tyr</td><td> Tyr</td><td> Tyr</td><td> Phe</td><td> Tyr</td><td> Tyr</td><td> Gly</td><td> Met</td><td> Asp</td><td> Val</td><td> Trp</td><td> Gly</td><td> Gin</td><td> Gly</td>

<td> 50</td><td></td><td> 130</td><td></td><td></td><td></td><td></td><td> 135</td><td></td><td></td><td></td><td></td><td> 140</td><td></td><td></td><td></td><td></td>

<td></td><td> Thr</td><td> Thr</td><td> Val</td><td> Thr</td><td> Val</td><td> Ser</td><td> Ser</td><td> Alá</td><td> Ser</td><td> Thr</td><td> Lys</td><td> Gly</td><td> Pro</td><td> Ser</td><td> Val</td><td> Phe</td>

<td></td><td> 145</td><td></td><td></td><td></td><td></td><td> 150</td><td></td><td></td><td></td><td></td><td> 155</td><td></td><td></td><td></td><td></td><td> 160</td>

<td> 55</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<td></td><td> Pro</td><td> Leu</td><td> Alá</td><td> Pro</td><td> Cys</td><td> Ser</td><td> Arg</td><td> Ser</td><td> Thr</td><td> Ser</td><td> Glu</td><td> Ser</td><td> Thr</td><td> Alá</td><td> Alá</td><td> Leu</td>

<td></td><td></td><td></td><td></td><td></td><td> 165</td><td></td><td></td><td></td><td></td><td> 170</td><td></td><td></td><td></td><td></td><td> 175</td><td></td>

143

ΕΡ 2 379 594 Β1

Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp 180 185 190

Asn Ser Gly Alá Leu Thr Ser Gly Val His Thr Phe Pro Alá Val Leu 195 200 205

Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser 210 215 220

Ser Asn Phe Gly Thr Gin Thr Tyr Thr Cys Asn Val Asp His Lys Pro 225 230 235 240

Ser Asn Thr Lys Val Asp Lys Thr Val Glu Arg Lys Cys Cys Val Glu 245 250 255

Cys Pro Pro Cys Pro Alá Pro Pro Val Alá Gly Pro Ser Val Phe Leu 260 265 270

Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu 275 280 285

Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Gin 290 295 300

Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Alá Lys Thr Lys 305 310 315 320

Pro Arg Glu Glu Gin Phe Asn Ser Thr Phe Arg Val Val Ser Val Leu 325 330 335

Thr Val Val His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys 340 345 350

Val Ser Asn Lys Gly Leu Pro Alá Pro Ile Glu Lys Thr Ile Ser Lys 355 360 365

Thr Lys Gly Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser 370 375 380

Arg Glu Glu Met Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys 385 390 395 400

Gly Phe Tyr Pro Ser Asp Ile Alá Val Glu Trp Glu Ser Asn Gly Gin 405 410 415

Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Met Leu Asp Ser Asp Gly 420 425 430

Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin

144

ΕΡ 2 379 594 Β1

445

Glu Alá Leu His Asn

460

440

435

Gin Gly Asn Val Phe Ser 450

Cys Ser Val Met His 455

His Tyr Thr Gin Lys Ser Leu Ser Leu Ser Pro Gly Lys 465 470 475 <210> 34 <211> 469 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 34

145

ΕΡ 2 379 594 Β1

Met Asp Met Arg Val Pro Alá Gin 1 5

Leu Leu Gly Leu Leu Leu Leu Trp 10 15

Leu Arg Gly Alá Arg Cys Glu Val 20

Gin Leu Val Glu Ser Gly Gly Gly 25 30

Leu Val Lys Pro Gly Arg Ser Leu 35 40

Arg Leu Ser Cys Thr Alá Ser Gly 45

Phe Thr Phe Gly Asp Tyr Alá Met 50 55

Ser Trp Phe Arg Gin Alá Pro Gly 60

Lys Gly Leu Glu Trp Ile Gly Phe 65 70

Ile Arg Ser Arg Alá Tyr Gly Gly 75 80

Thr Pro Glu Tyr Alá Alá Ser Val 85

Lys Gly Arg Phe Thr Ile Ser Arg 90 95

Asp Asp Ser Lys Thr Ile Alá Tyr 100

Leu Gin Met Asn Ser Leu Lys Thr 105 110

Glu Asp Thr Alá Val Tyr Phe Cys 115 120

Alá Arg Gly Arg Gly Ile Alá Alá 125

Arg Trp Asp Tyr Trp Gly Gin Gly 130 135

Thr Leu Val Thr Val Ser Ser Alá 140

Ser Thr Lys Gly Pro Ser Val Phe 145 150

Pro Leu Alá Pro Cys Ser Arg Ser 155 160

Thr Ser Glu Ser Thr Alá Alá Leu 165

Gly Cys Leu Val Lys Asp Tyr Phe 170 175

146

ΕΡ 2 379 594 Β1

Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Alá Leu Thr Ser Gly 180 185 190

Val His Thr Phe Pro Alá Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu 195 200 205

Ser Ser Val Val Thr Val Pro Ser Ser Asn Phe Gly Thr Gin Thr Tyr 210 215 220

Thr Cys Asn Val Asp His Lys Pro Ser Asn Thr Lys Val Asp Lys Thr 225 230 235 240

Val Glu Arg Lys Cys Cys Val Glu Cys Pro Pro Cys Pro Alá Pro Pro 245 250 255

Val Alá Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr 260 265 270

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val 275 280 285

Ser His Glu Asp Pro Glu Val Gin Phe Asn Trp Tyr Val Asp Gly Val 290 295 300

Glu Val His Asn Alá Lys Thr Lys Pro Arg Glu Glu Gin Phe Asn Ser 305 310 315 320

Thr Phe Arg Val Val Ser Val Leu Thr Val Val His Gin Asp Trp Leu 325 330 335

Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Gly Leu Pro Alá 340 345 350

Pro Ile Glu Lys Thr Ile Ser Lys Thr Lys Gly Gin Pro Arg Glu Pro 355 360 365

Gin Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gin 370 375 380

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Alá 385 390 395 400

Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn Asn Tyr Lys Thr Thr 405 410 415

Pro Pro Met Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu 420 425 430

147

ΕΡ 2 379 594 Β1

Thr Val Asp Lys Ser 435

Arg Trp Gin Gin Gly 440

Asn Val

Phe Ser Cys Ser 445

Val Met His 450

Glu Alá

Leu His 455

Asn His Tyr

Thr Gin 460

Lys Ser Leu Ser

Leu Ser Pro Gly Lys 465 <210> 35 <211> 479 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 35

148

ΕΡ 2 379 594 Β1

Met Asp Met Arg Val Pro Alá Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Alá Arg Cys Glu Val Gin Leu Val Glu Ser Gly Gly Gly 20 25 30

Leu Val Lys Pro Gly Gly Ser Leu Arg Leu Ser Cys Alá Alá Ser Gly 35 40 45

Phe Thr Phe Ser Asn Alá Trp Met Ser Trp Val Arg Gin Alá Pro Gly 50 55 60

Lys Gly Leu Glu Trp Val Gly Arg Ile Lys Ser Lys Thr Asp Gly Gly 65 70 75 80

Thr Thr Asp Tyr Thr Alá Pro Val Lys Gly Arg Phe Thr Ile Ser Arg 85 90 95

Asp Asp Ser Lys Asn Thr Leu Tyr Leu Gin Met Asn Ser Leu Lys Alá 100 105 110

Glu Asp Thr Alá Val Tyr Tyr Cys Thr Thr Asp Arg Thr Gly Tyr Ser 115 120 125

Ile Ser Trp Ser Ser Tyr Tyr Tyr Tyr Tyr Gly Met Asp Val Trp Gly 130 135 140

Gin Gly Thr Thr Val Thr Val Ser Ser Alá Ser Thr Lys Gly Pro Ser 145 150 155 160

Val Phe Pro Leu Alá Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Alá 165 170 175

149

ΕΡ 2 379 594 Β1

Alá Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val 180 185 190

Ser Trp Asn Ser Gly Alá Leu Thr Ser Gly Val His Thr Phe Pro Alá 195 200 205

Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val 210 215 220

Pro Ser Ser Asn Phe Gly Thr Gin Thr Tyr Thr Cys Asn Val Asp His 225 230 235 240

Lys Pro Ser Asn Thr Lys Val Asp Lys Thr Val Glu Arg Lys Cys Cys 245 250 255

Val Glu Cys Pro Pro Cys Pro Alá Pro Pro Val Alá Gly Pro Ser Val 260 265 270

Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr 275 280 285

Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu 290 295 300

Val Gin Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Alá Lys 305 310 315 320

Thr Lys Pro Arg Glu Glu Gin Phe Asn Ser Thr Phe Arg Val Val Ser 325 330 335

Val Leu Thr Val Val His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys 340 345 350

Cys Lys Val Ser Asn Lys Gly Leu Pro Alá Pro Ile Glu Lys Thr Ile 355 360 365

Ser Lys Thr Lys Gly Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro 370 375 380

Pro Ser Arg Glu Glu Met Thr Lys Asn Gin Val Ser Leu Thr Cys Leu 385 390 395 400

Val Lys Gly Phe Tyr Pro Ser Asp Ile Alá Val Glu Trp Glu Ser Asn 405 410 415

Gly Gin Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Met Leu Asp Ser 420 425 430

150

ΕΡ 2 379 594 Β1

Asp Gly Ser Phe Phe 435

Leu

Tyr Ser Lys Leu 440

Thr Val Asp 445

Lys

Ser Arg

Trp Gin Gin 450

Gly Asn Val

Phe

455

Ser Cys

Ser Val Met 460

His

Glu Alá Leu

His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu Ser Pro Gly Lys 465 470 475 <210> 36 <211> 475 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 36

151

EP 2 379 594 Β1

Met Asp Met Arg Val Pro Ala Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Ala Arg Cys Gin Val Gin Leu Val Gin Ser Gly Ala Glu 20 25 30

Val Lys Lys Pro Gly Ala Ser Val Lys Val Ser Cys Lys Ala Ser Gly 35 40 45

Tyr Thr Phe Thr Asp Tyr Tyr Met Tyr Trp Val Arg Gin Ala Pro Gly 50 55 60

Gin Gly Leu Glu Trp Met Gly Trp Ile Ser Pro Asn Ser Gly Gly Thr 65 70 75 80

Asn Tyr Ala Gin Lys Phe Gin Gly Arg Val Thr Met Thr Arg Asp Thr 85 90 95

Ser Ile Ser Thr Ala Tyr Met Glu Leu Ser Arg Leu Arg Ser Asp Asp 100 105 110

Thr Ala Val Tyr Tyr Cys Val Arg Gly Gly Tyr Ser Gly Tyr Ala Gly 115 120 125

Leu Tyr Ser His Tyr Tyr Gly Met Asp Val Trp Gly Gin Gly Thr Thr 130 135 140

Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu 145 150 155 160

Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly Cys

152

ΕΡ 2 379 594 Β1

165

170

175

Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser 180 185 190

Gly Alá Leu Thr Ser Gly Val His Thr Phe Pro Alá Val Leu Gin Ser 195 200 205

Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Asn 210 215 220

Phe Gly Thr Gin Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser Asn 225 230 235 240

Thr Lys Val Asp Lys Thr Val Glu Arg Lys Cys Cys Val Glu Cys Pro 245 250 255

Pro Cys Pro Alá Pro Pro Val Alá Gly Pro Ser Val Phe Leu Phe Pro 260 265 270

Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr 275 280 285

Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Gin Phe Asn 290 295 300

Trp Tyr Val Asp Gly Val Glu Val His Asn Alá Lys Thr Lys Pro Arg 305 310 315 320

Glu Glu Gin Phe Asn Ser Thr Phe Arg Val Val Ser Val Leu Thr Val 325 330 335

Val His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser 340 345 350

Asn Lys Gly Leu Pro Alá Pro Ile Glu Lys Thr Ile Ser Lys Thr Lys 355 360 365

Gly Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Glu 370 375 380

Glu Met Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe 385 390 395 400

Tyr Pro Ser Asp Ile Alá Val Glu Trp Glu Ser Asn Gly Gin Pro Glu 405 410 415

Asn Asn Tyr Lys Thr Thr Pro Pro Met Leu Asp Ser Asp Gly Ser Phe 420 425 430

153

ΕΡ 2 379 594 Β1

<td> Phe Leu</td><td> Tyr 435</td><td> Ser Lys</td><td> Leu Thr Val Asp Lys 440</td><td> Ser Arg Trp Gin Gin 445</td><td> Gly</td>

<td> Asn Val</td><td> Phe</td><td> Ser Cys</td><td> Ser Val Met His Glu</td><td> Alá Leu His Asn His</td><td> Tyr</td>

<td> 450</td><td></td><td></td><td> 455</td><td> 460</td><td></td>

<td> Thr Gin</td><td> Lys</td><td> Ser Leu</td><td> Ser Leu Ser Pro Gly</td><td> Lys</td><td></td>

<td> 465</td><td></td><td></td><td> 470</td><td> 475</td><td></td>

<td> &lt;210&gt; 37</td><td></td><td></td><td></td><td></td><td></td>

<td> &lt;211&gt; 479</td><td></td><td></td><td></td><td></td><td></td>

<td> &lt;212&gt; PRT</td><td></td><td></td><td></td><td></td><td></td>

<td colspan="2"> &lt;213&gt; Artificial Sequence</td><td></td><td></td><td></td><td></td>

<td> &lt;220&gt;</td><td></td><td></td><td></td><td></td><td></td>

<td> &lt;221 &gt; source</td><td></td><td></td><td></td><td></td><td></td>

<td colspan="5"> &lt;223&gt; /note=Description of Artificial Sequence: Synthetic polypeptide</td><td></td>

<td> &lt;400&gt; 37</td><td></td><td></td><td></td><td></td><td></td>

<td> Met Asp</td><td> Met</td><td> Arg Val</td><td> Pro Alá Gin Leu Leu</td><td> Gly Leu Leu Leu Leu</td><td> Trp</td>

<td> 1</td><td></td><td> 5</td><td> 10</td><td> 15</td><td></td>

<td> Leu Arg</td><td> Gly</td><td> Alá Arg</td><td> Cys Glu Val Gin Leu</td><td> Val Glu Ser Gly Gly</td><td> Gly</td>

<td></td><td></td><td> 20</td><td> 25</td><td> 30</td><td></td>

<td> Leu Val</td><td> Lys</td><td> Pro Gly</td><td> Gly Ser Leu Arg Leu</td><td> Ser Cys Alá Alá Ser</td><td> Gly</td>

<td></td><td> 35</td><td></td><td> 40</td><td> 45</td><td></td>

<td> Phe Thr</td><td> Phe</td><td> Gly Asn</td><td> Alá Trp Met Ser Trp</td><td> Val Arg Gin Alá Pro</td><td> Gly</td>

<td> 50</td><td></td><td></td><td> 55</td><td> 60</td><td></td>

<td> Lys Gly</td><td> Leu</td><td> Glu Trp</td><td> Val Gly Arg Ile Lys</td><td> Ser Lys Thr Asp Gly</td><td> Gly</td>

<td> 65</td><td></td><td></td><td> 70</td><td> 75</td><td> 80</td>

<td> Thr Thr</td><td> Asp</td><td> Tyr Alá</td><td> Alá Pro Val Lys Gly</td><td> Arg Phe Thr Ile Ser</td><td> Arg</td>

<td></td><td></td><td> 85</td><td> 90</td><td> 95</td><td></td>

<td> Asp Asp</td><td> Ser</td><td> Lys Asn</td><td> Thr Leu Tyr Leu Gin</td><td> Met Asn Ser Leu Lys</td><td> Thr</td>

<td></td><td></td><td> 100</td><td> 105</td><td> 110</td><td></td>

<td> Glu Asp</td><td> Thr</td><td> Alá Val</td><td> Tyr Phe Cys Thr Thr</td><td> Asp Arg Thr Gly Tyr</td><td> Ser</td>

<td></td><td> 115</td><td></td><td> 120</td><td> 125</td><td></td>

<td> Ile Ser</td><td> Trp</td><td> Ser Ser</td><td> Tyr Tyr Tyr Tyr Tyr</td><td> Gly Met Asp Val Trp</td><td> Gly</td>

<td> 130</td><td></td><td></td><td> 135</td><td> 140</td><td></td>

<td> Gin Gly</td><td> Thr</td><td> Thr Val</td><td> Thr Val Ser Ser Alá</td><td> Ser Thr Lys Gly Pro</td><td> Ser</td>

<td> 145</td><td></td><td></td><td> 150</td><td> 155</td><td> 160</td>

154

ΕΡ 2 379 594 Β1

Val Phe Pro Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala 165 170 175

Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val 180 185 190

Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala 195 200 205

Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val 210 215 220

Pro Ser Ser Asn Phe Gly Thr Gin Thr Tyr Thr Cys Asn Val Asp His 225 230 235 240

Lys Pro Ser Asn Thr Lys Val Asp Lys Thr Val Glu Arg Lys Cys Cys 245 250 255

Val Glu Cys Pro Pro Cys Pro Ala Pro Pro Val Ala Gly Pro Ser Val 260 265 270

Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr 275 280 285

Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu 290 295 300

Val Gin Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys 305 310 315 320

Thr Lys Pro Arg Glu Glu Gin Phe Asn Ser Thr Phe Arg Val Val Ser 325 330 335

Val Leu Thr Val Val His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys 340 345 350

Cys Lys Val Ser Asn Lys Gly Leu Pro Ala Pro Ile Glu Lys Thr Ile 355 360 365

Ser Lys Thr Lys Gly Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro 370 375 380

Pro Ser Arg Glu Glu Met Thr Lys Asn Gin Val Ser Leu Thr Cys Leu 385 390 395 400

Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn 405 410 415

Gly Gin Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Met Leu Asp Ser

155

EP 2 379 594 Β1

<td></td><td></td><td></td><td> 420</td><td></td><td></td><td></td><td></td><td> 425</td><td></td><td></td><td></td><td></td><td> 430</td><td></td><td></td>

<td> Asp</td><td> Gly</td><td> Ser 435</td><td> Phe</td><td> Phe</td><td> Leu</td><td> Tyr</td><td> Ser 440</td><td> Lys</td><td> Leu</td><td> Thr</td><td> Val</td><td> Asp 445</td><td> Lys</td><td> Ser</td><td> Arg</td>

<td> Trp</td><td> Gin 450</td><td> Gin</td><td> Gly</td><td> Asn</td><td> Val</td><td> Phe 455</td><td> Ser</td><td> Cys</td><td> Ser</td><td> Val</td><td> Met 460</td><td> His</td><td> Glu</td><td> Alá</td><td> Leu</td>

<td> His 465</td><td> Asn</td><td> His</td><td> Tyr</td><td> Thr</td><td> Gin 470</td><td> Lys</td><td> Ser</td><td> Leu</td><td> Ser</td><td> Leu 475</td><td> Ser</td><td> Pro</td><td> Gly</td><td> Lys</td><td></td>

<td colspan="3"> &lt;210&gt; 38 &lt;211&gt; 479 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 38

156

ΕΡ 2 379 594 Β1

Met Asp Met Arg Val Pro Alá Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Alá Arg Cys Glu Val Gin Leu Val Glu Ser Gly Gly Gly 20 25 30

Leu Val Lys Pro Gly Gly Ser Leu Arg Leu Ser Cys Alá Alá Ser Gly 35 40 45

Phe Thr Phe Gly Asn Alá Trp Met Ser Trp Val Arg Gin Alá Pro Gly 50 55 60

Lys Gly Leu Glu Trp Val Gly Arg Ile Lys Ser Lys Thr Asp Gly Gly 65 70 75 80

Thr Thr Asp Tyr Alá Alá Pro Val Lys Gly Arg Phe Thr Ile Ser Arg 85 90 95

Asp Asp Ser Lys Asn Thr Leu Tyr Leu Gin Met Asn Ser Leu Lys Thr 100 105 110

Glu Asp Thr Alá Val Tyr Tyr Cys Thr Thr Asp Arg Thr Gly Tyr Ser 115 120 125

Ile Ser Trp Ser Ser Tyr Tyr Tyr Tyr Tyr Gly Met Asp Val Trp Gly 130 135 140

Gin Gly Thr Thr Val Thr Val Ser Ser Alá Ser Thr Lys Gly Pro Ser 145 150 155 160

157

ΕΡ 2 379 594 Β1

Val Phe Pro Leu Alá Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Alá 165 170 175

Alá Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val 180 185 190

Ser Trp Asn Ser Gly Alá Leu Thr Ser Gly Val His Thr Phe Pro Alá 195 200 205

Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val 210 215 220

Pro Ser Ser Asn Phe Gly Thr Gin Thr Tyr Thr Cys Asn Val Asp His 225 230 235 240

Lys Pro Ser Asn Thr Lys Val Asp Lys Thr Val Glu Arg Lys Cys Cys 245 250 255

Val Glu Cys Pro Pro Cys Pro Alá Pro Pro Val Alá Gly Pro Ser Val 260 265 270

Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr 275 280 285

Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu 290 295 300

Val Gin Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Alá Lys 305 310 315 320

Thr Lys Pro Arg Glu Glu Gin Phe Asn Ser Thr Phe Arg Val Val Ser 325 330 335

Val Leu Thr Val Val His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys 340 345 350

Cys Lys Val Ser Asn Lys Gly Leu Pro Alá Pro Ile Glu Lys Thr Ile 355 360 365

Ser Lys Thr Lys Gly Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro 370 375 380

Pro Ser Arg Glu Glu Met Thr Lys Asn Gin Val Ser Leu Thr Cys Leu 385 390 395 400

Val Lys Gly Phe Tyr Pro Ser Asp Ile Alá Val Glu Trp Glu Ser Asn 405 410 415

158

ΕΡ 2 379 594 Β1

<td> Gly</td><td> Gin</td><td> Pro</td><td> Glu 420</td><td> Asn</td><td> Asn</td><td> Tyr</td><td> Lys</td><td> Thr 425</td><td> Thr</td><td> Pro</td><td> Pro</td><td> Met</td><td> Leu 430</td><td> Asp</td><td> Ser</td>

<td> Asp</td><td> Gly</td><td> Ser 435</td><td> Phe</td><td> Phe</td><td> Leu</td><td> Tyr</td><td> Ser 440</td><td> Lys</td><td> Leu</td><td> Thr</td><td> Val</td><td> Asp 445</td><td> Lys</td><td> Ser</td><td> Arg</td>

<td> Trp</td><td> Gin 450</td><td> Gin</td><td> Gly</td><td> Asn</td><td> Val</td><td> Phe 455</td><td> Ser</td><td> Cys</td><td> Ser</td><td> Val</td><td> Met 460</td><td> His</td><td> Glu</td><td> Alá</td><td> Leu</td>

<td> His 465</td><td> Asn</td><td> His</td><td> Tyr</td><td> Thr</td><td> Gin 470</td><td> Lys</td><td> Ser</td><td> Leu</td><td> Ser</td><td> Leu 475</td><td> Ser</td><td> Pro</td><td> Gly</td><td> Lys</td><td></td>

<td> &lt;210&gt; 39 &lt;211&gt; 478 &lt;212&gt; PRT</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<td colspan="3"> &lt;213&gt; Artificial Sequence</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 39

159

ΕΡ 2 379 594 Β1

Met Asp Met Arg Val Pro Alá Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Alá Arg Cys Gin Val Gin Leu Val Glu Ser Gly Gly Gly 20 25 30

Val Val Gin Pro Gly Arg Ser Leu Arg Leu Ser Cys Alá Alá Ser Gly 35 40 45

Phe Thr Phe Ser Ser Phe Gly Met His Trp Val Arg Gin Alá Pro Gly 50 55 60

Lys Gly Leu Glu Trp Val Alá Val Ile Ser Phe Asp Gly Ser Ile Lys 65 70 75 80

Tyr Ser Val Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn 85 90 95

Ser Lys Asn Thr Leu Phe Leu Gin Met Asn Ser Leu Arg Alá Glu Asp 100 105 110

Thr Alá Val Tyr Tyr Cys Alá Arg Asp Arg Leu Asn Tyr Tyr Asp Ser 115 120 125

Ser Gly Tyr Tyr His Tyr Lys Tyr Tyr Gly Leu Alá Val Trp Gly Gin 130 135 140

Gly Thr Thr Val Thr Val Ser Ser Alá Ser Thr Lys Gly Pro Ser Val 145 150 155 160

160

ΕΡ 2 379 594 Β1

Phe Pro Leu Alá Pro 165

Cys Ser Arg Ser Thr Ser 170

Glu Ser Thr Alá Alá 175

Leu Gly Cys Leu Val 180

Trp Asn Ser Gly Alá 195

Leu Gin Ser Ser Gly 210

Ser Ser Asn Phe Gly 225

Pro Ser Asn Thr Lys 245

Glu Cys Pro Pro Cys 260

Leu Phe Pro Pro Lys 275

Glu Val Thr Cys Val 290

Gin Phe Asn Trp Tyr 305

Lys Pro Arg Glu Glu 325

Leu Thr Val Val His 340

Lys Val Ser Asn Lys 355

Lys Thr Lys Gly Gin 370

Ser Arg Glu Glu Met 385

Lys

Gly Phe

Tyr Pro 405

Lys Asp Tyr Phe Pro Glu 185

Leu Thr Ser Gly Val His 200

Leu Tyr Ser Leu Ser Ser 215

Thr Gin Thr Tyr Thr Cys 230 235

Val Asp Lys Thr Val Glu 250

Pro Alá Pro Pro Val Alá 265

Pro Lys Asp Thr Leu Met 280

Val Val Asp Val Ser His 295

Val Asp Gly Val Glu Val 310 315

Gin Phe Asn Ser Thr Phe 330

Gin Asp Trp Leu Asn Gly 345

Gly Leu Pro Alá Pro Ile 360

Pro Arg Glu Pro Gin Val 375

Thr Lys Asn Gin Val Ser 390 395

Ser Asp

Ile Alá

Pro Val Thr Val Ser 190

Thr Phe Pro Alá Val 205

Val Val Thr Val Pro 220

Asn Val Asp His Lys 240

Arg Lys Cys Cys Val 255

Gly Pro Ser Val Phe 270

Ile Ser Arg Thr Pro 285

Glu Asp Pro Glu Val 300

His Asn Alá Lys Thr 320

Arg Val Val Ser Val 335

Lys Glu Tyr Lys Cys 350

Glu Lys Thr Ile Ser 365

Tyr Thr Leu Pro Pro 380

Leu Thr Cys Leu Val 400

Val Glu Trp Glu 410

Ser Asn Gly 415

161

ΕΡ 2 379 594 Β1

<td> Gin</td><td> Pro</td><td> Glu</td><td> Asn</td><td> Asn</td><td> Tyr</td><td> Lys</td><td> Thr</td><td> Thr</td><td> Pro</td><td> Pro</td><td> Met</td><td> Leu</td><td> Asp</td><td> Ser</td><td> Asp</td>

<td></td><td></td><td></td><td> 420</td><td></td><td></td><td></td><td></td><td> 425</td><td></td><td></td><td></td><td></td><td> 430</td><td></td><td></td>

<td> Gly</td><td> Ser</td><td> Phe</td><td> Phe</td><td> Leu</td><td> Tyr</td><td> Ser</td><td> Lys</td><td> Leu</td><td> Thr</td><td> Val</td><td> Asp</td><td> Lys</td><td> Ser</td><td> Arg</td><td> Trp</td>

<td></td><td></td><td> 435</td><td></td><td></td><td></td><td></td><td> 440</td><td></td><td></td><td></td><td></td><td> 445</td><td></td><td></td><td></td>

<td> Gin</td><td> Gin</td><td> Gly</td><td> Asn</td><td> Val</td><td> Phe</td><td> Ser</td><td> Cys</td><td> Ser</td><td> Val</td><td> Met</td><td> His</td><td> Glu</td><td> Alá</td><td> Leu</td><td> His</td>

<td></td><td> 450</td><td></td><td></td><td></td><td></td><td> 455</td><td></td><td></td><td></td><td></td><td> 460</td><td></td><td></td><td></td><td></td>

<td> Asn</td><td> His</td><td> Tyr</td><td> Thr</td><td> Gin</td><td> Lys</td><td> Ser</td><td> Leu</td><td> Ser</td><td> Leu</td><td> Ser</td><td> Pro</td><td> Gly</td><td> Lys</td><td></td><td></td>

<td> 465</td><td></td><td></td><td></td><td></td><td> 470</td><td></td><td></td><td></td><td></td><td> 475</td><td></td><td></td><td></td><td></td><td></td>

<td> &lt;210&gt; 40</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<td> &lt;211&gt; 479</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<td> &lt;212&gt; PRT</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<td colspan="3"> &lt;213&gt; Artificial Sequence</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 40

162

ΕΡ 2 379 594 Β1

Met Asp Met Arg Val Pro Alá Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Alá Arg Cys Glu Val Gin Leu Val Glu Ser Gly Gly Gly 20 25 30

Leu Val Lys Pro Gly Gly Ser Leu Arg Leu Ser Cys Alá Alá Ser Gly 35 40 45

Tyr Thr Phe Ser Thr Tyr Ser Met Asn Trp Val Arg Gin Alá Pro Gly 50 55 60

Lys Gly Leu Glu Trp Val Ser Ser Ile Ser Ser Ser Ser Ser Tyr Arg 65 70 75 80

Tyr Tyr Alá Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn 85 90 95

Alá Lys Asn Ser Leu Tyr Leu Gin Met Ser Ser Leu Arg Alá Glu Asp 100 105 110

Thr Alá Val Tyr Tyr Cys Alá Arg Glu Gly Val Ser Gly Ser Ser Pro 115 120 125

Tyr Ser Ile Ser Trp Tyr Asp Tyr Tyr Tyr Gly Met Asp Val Trp Gly 130 135 140

Gin Gly Thr Thr Val Thr Val Ser Ser Alá Ser Thr Lys Gly Pro Ser

163

ΕΡ 2 379 594 Β1

145

150

155

160

Val Phe Pro Leu Alá Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Alá 165 170 175

Alá Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val 180 185 190

Ser Trp Asn Ser Gly Alá Leu Thr Ser Gly Val His Thr Phe Pro Alá 195 200 205

Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val 210 215 220

Pro Ser Ser Asn Phe Gly Thr Gin Thr Tyr Thr Cys Asn Val Asp His 225 230 235 240

Lys Pro Ser Asn Thr Lys Val Asp Lys Thr Val Glu Arg Lys Cys Cys 245 250 255

Val Glu Cys Pro Pro Cys Pro Alá Pro Pro Val Alá Gly Pro Ser Val 260 265 270

Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr 275 280 285

Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu 290 295 300

Val Gin Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Alá Lys 305 310 315 320

Thr Lys Pro Arg Glu Glu Gin Phe Asn Ser Thr Phe Arg Val Val Ser 325 330 335

Val Leu Thr Val Val His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys 340 345 350

Cys Lys Val Ser Asn Lys Gly Leu Pro Alá Pro Ile Glu Lys Thr Ile 355 360 365

Ser Lys Thr Lys Gly Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro 370 375 380

Pro Ser Arg Glu Glu Met Thr Lys Asn Gin Val Ser Leu Thr Cys Leu 385 390 395 400

Val Lys Gly Phe Tyr Pro Ser Asp Ile Alá Val Glu Trp Glu Ser Asn 405 410 415

164

ΕΡ 2 379 594 Β1

<td colspan="2" rowspan="2"> Gly</td><td colspan="7"> Gin Pro Glu Asn Asn Tyr Lys</td><td colspan="3"> Thr Thr Pro</td><td colspan="2"> Pro Met</td><td colspan="2" rowspan="2"> Leu Asp 430</td><td rowspan="3"> Ser Arg</td>

<td colspan="4"> 420</td><td rowspan="2"> Leu</td><td rowspan="2"> Tyr</td><td rowspan="2"> Ser 440</td><td rowspan="2"> 425 Lys</td><td rowspan="2"> Leu</td><td rowspan="2"> Thr</td><td rowspan="2"> Val</td><td rowspan="2"> Asp 445</td>

<td> 5</td><td> Asp</td><td> Gly</td><td colspan="3"> Ser Phe Phe 435</td><td> Lys</td><td> Ser</td>

<td> 10</td><td> Trp</td><td> Gin 450</td><td> Gin</td><td> Gly</td><td> Asn</td><td> Val</td><td> Phe 455</td><td> Ser</td><td> Cys</td><td> Ser</td><td> Val</td><td> Met 460</td><td> His</td><td> Glu</td><td> Alá</td><td> Leu</td>

<td></td><td> His</td><td> Asn</td><td> His</td><td> Tyr</td><td> Thr</td><td> Gin</td><td> Lys</td><td> Ser</td><td> Leu</td><td> Ser</td><td> Leu</td><td> Ser</td><td> Pro</td><td> Gly</td><td> Lys</td><td></td>

465 470 475 <210> 41 <211 > 474 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic polypeptide <400> 41

Met Asp Met Arg Val Pro Alá Gin Leu Leu Gly Leu Leu Leu Leu Trp 15 10 15

Leu Arg Gly Alá Arg Cys Gin Val Gin Leu Val Glu Ser Gly Gly Gly 20 25 30

Val Val Gin Pro Gly Arg Ser Leu Arg Leu Ser Cys Alá Alá Ser Gly 35 40 45

Phe Thr Phe Ser Ser Tyr Gly Met His Trp Val Arg Gin Alá Pro Gly 50 55 60

Lys Gly Leu Glu Trp Val Alá Val Ile Trp Tyr Asp Gly Ser Asn Lys 65 70 75 80

Tyr Tyr Alá Asp Ser Val Lys Gly Arg Phe Ile Ile Ser Arg Asp Lys 85 90 95

Ser Lys Asn Thr Leu Tyr Leu Gin Met Asn Ser Leu Arg Alá Glu Asp 100 105 110

Thr Alá Val Tyr Tyr Cys Alá Arg Alá Gly Gly Ile Alá Alá Alá Gly 115 120 125

Leu Tyr Tyr Tyr Tyr Gly Met Asp Val Trp Gly Gin Gly Thr Thr Val 130 135 140

165

EP 2 379 594 Β1

Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala 145 150 155 160

Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly Cys Leu 165 170 175

Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly 180 185 190

Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser 195 200 205

Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Asn Phe 210 215 220

Gly Thr Gin Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser Asn Thr 225 230 235 240

Lys Val Asp Lys Thr Val Glu Arg Lys Cys Cys Val Glu Cys Pro Pro 245 250 255

Cys Pro Ala Pro Pro Val Ala Gly Pro Ser Val Phe Leu Phe Pro Pro 260 265 270

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys 275 280 285

Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Gin Phe Asn Trp 290 295 300

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 305 310 315 320

Glu Gin Phe Asn Ser Thr Phe Arg Val Val Ser Val Leu Thr Val Val 325 330 335

His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 340 345 350

Lys Gly Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Thr Lys Gly 355 360 365

Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu 370 375 380

Met Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 385 390 395 400

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn

166

ΕΡ 2 379 594 Β1

405 410 415

<td> Asn</td><td> Tyr</td><td> Lys</td><td> Thr 420</td><td> Thr</td><td> Pro</td><td> Pro</td><td> Met</td><td> Leu 425</td><td> Asp</td><td> Ser</td><td> Asp</td><td> Gly</td><td> Ser 430</td><td> Phe</td><td> Phe</td>

<td> Leu</td><td> Tyr</td><td> Ser 435</td><td> Lys</td><td> Leu</td><td> Thr</td><td> Val</td><td> Asp 440</td><td> Lys</td><td> Ser</td><td> Arg</td><td> Trp</td><td> Gin 445</td><td> Gin</td><td> Gly</td><td> Asn</td>

<td> Val</td><td> Phe 450</td><td> Ser</td><td> Cys</td><td> Ser</td><td> Val</td><td> Met 455</td><td> His</td><td> Glu</td><td> Alá</td><td> Leu</td><td> His 460</td><td> Asn</td><td> His</td><td> Tyr</td><td> Thr</td>

<td> Gin 465</td><td> Lys</td><td> Ser</td><td> Leu</td><td> Ser</td><td> Leu 470</td><td> Ser</td><td> Pro</td><td> Gly</td><td> Lys</td><td></td><td></td><td></td><td></td><td></td><td></td>

<td colspan="3"> &lt;210&gt; 42 &lt;211&gt; 13 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 42

Ser Gly Ser Ser Ser Asn Ile Gly Asn Asn Tyr Val Ser 15 10 <210> 43 <211>7 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 43

Asp Asn Asn Lys Arg Pro Ser 1 5 <210> 44 <211> 11 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 44

Gly Thr Trp Asp Ser Arg Leu Ser Alá Val Val 15 10

167

ΕΡ 2 379 594 Β1 <210> 45 <211> 13 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 45

Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn Tyr Val Tyr 15 10 <210> 46 <211>7 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 46

Arg Ser Asn Gin Arg Pro Ser 1 5 <210> 47 <211> 11 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 47

Alá Alá Trp Asp Asp Ser Leu Ser Gly Trp Val 15 10 <210> 48 <211> 11 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 48

Arg Alá Ser Gin Gly Ile Arg Asn Asp Leu Gly 15 10 <210> 49 <211>7

168

EP 2 379 594 B1 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 49

Alá Alá Ser Ser Leu Gin Ser 1 5 <210> 50 <211>9 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 50

Leu Gin Tyr Asn Ile Tyr Pro Trp Thr 1 5 <210> 51 <211> 11 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 51

Gin Gly Asp Ser Leu Arg Ser Phe Tyr Alá Ser 15 10 <210> 52 <211>7 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 52

Gly Lys Asn Asn Arg Pro Ser 1 5 <210> 53 <211> 11 <212> PRT

169

ΕΡ 2 379 594 Β1 <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 53

Asn Ser Arg Asp Ser Ser Val Tyr His Leu Val 15 10 <210> 54 <211> 16 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 54

Lys Ser Ser Gin Ser Leu Leu His Ser Alá Gly Lys Thr Tyr Leu Tyr 15 10 15 <210> 55 <211>7 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 55

Glu Val Ser Asn Arg Phe Ser 1 5 <210> 56 <211>9 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 56

Met Gin Ser Phe Pro Leu Pro Leu Thr 1 5 <210> 57 <211> 16 <212> PRT <213> Artificial Sequence

170

ΕΡ 2 379 594 Β1 <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 57

Arg Ser Ser Gin Ser Leu Leu His Ser Phe Gly Tyr Asn Tyr Leu Asp 15 10 15 <210> 58 <211>7 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 58

Leu Gly Ser Asn Arg Ala Ser 1 5 <210> 59 <211>9 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 59

Met Gin Ala Leu Gin Thr Pro Phe Thr 1 5 <210> 60 <211> 16 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 60

Lys Ser Ser Gin Ser Leu Leu His Ser Asp Gly Lys Thr Tyr Leu Tyr 15 10 15 <210> 61 <211>7 <212> PRT <213> Artificial Sequence <220>

<221 > source

171

ΕΡ 2 379 594 Β1 <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 61

Arg Asn Asn Gin Arg Pro Ser 1 5 <210> 62 <211> 13 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 62

Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn Thr Val Asn 15 10 <210> 63 <211>7 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 63

Thr Asn Asn Gin Arg Pro Ser 1 5 <210> 64 <211> 11 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 64

Alá Alá Arg Asp Glu Ser Leu Asn Gly Val Val 15 10 <210> 65 <211> 16 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide

172

ΕΡ 2 379 594 Β1 <400> 65

Lys Ser Ser Gin Ser Leu Leu His Ser Asp Gly Arg Asn Tyr Leu Tyr 15 10 15 <210> 66 <211> 11 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 66

Arg Alá Ser Gin Gly Ile Arg Lys Asp Leu Gly 15 10 <210> 67 <211>7 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 67

Gly Alá Ser Ser Leu Gin Ser 1 5 <210> 68 <211>9 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 68

Leu Gin Tyr Asn Ser Phe Pro Trp Thr 1 5 <210> 69 <211> 12 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 69

173

ΕΡ 2 379 594 Β1

Arg Alá Ser Gin Ser Val Ser Ser Gly Tyr Leu Thr 15 10 <210> 70 <211>7 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 70

Gly Alá Ser Ser Arg Alá Thr 1 5 <210> 71 <211>9 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 71

Gin Gin Tyr Gly Asn Ser Leu Cys Arg 1 5 <210> 72 <211>9 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 72

Gin Gin Tyr Gly Asn Ser Leu Ser Arg 1 5 <210> 73 <211>5 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 73

Ser Phe Gly Met His 1 5

174

ΕΡ 2 379 594 Β1 <210> 74 <211> 17 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 74

Val Ile Ser Phe Asp Gly Ser Ile Lys Tyr Ser Val Asp Ser Val Lys 15 10 15

Gly <210> 75 <211> 21 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 75

Asp Arg Leu Asn Tyr Tyr Asp Ser Ser Gly Tyr Tyr His Tyr Lys Tyr 15 10 15

Tyr Gly Met Alá Val 20 <210> 76 <211>5 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 76

Asn Alá Trp Met Ser 1 5 <210> 77 <211> 19 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide

175

ΕΡ 2 379 594 Β1 <400> 77

Arg Ile Lys Ser Thr Thr Asp Gly Gly Thr Thr Asp Tyr Alá Alá Pro 15 10 15

Val Lys Gly <210> 78 <211> 20 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 78

Asp Arg Thr Gly Tyr Ser Ile Ser Trp Ser Ser Tyr Tyr Tyr Tyr Tyr 15 10 15

Gly Met Asp Val 20 <210> 79 <211>5 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 79

Ser Tyr Alá Met Ser 1 5 <210> 80 <211> 17 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 80

Alá Ile Ser Gly Ser Gly Gly Arg Thr Tyr Tyr Alá Asp Ser Val Lys 15 10 15

Gly <210> 81 <211> 21

176

EP 2 379 594 B1 <212> PRT <213> Artificial Sequence <220>

<221 > source <223> /note=Description of Artificial Sequence: Synthetic peptide <400> 81

Asp Gin Arg Glu Val Gly Pro Tyr Ser Ser Gly Trp Tyr Asp Tyr Tyr 15 10 15