Anti-VEGF antibodies

p0001Anti-VEGF antibodies.

Background of the invention

Field of the Invention

p0002This invention relates, in general, to anti-VEGF antibodies and, in particular, to humanized anti-VEGF antibodies and variants of anti-VEGF antibodies.

Description of the related technique

p0003It is now well established that angiogenesis It is involved in the pathogenesis of a variety of disorders. These include solid tumors, neovascular syndromes intraocular such as proliferative retinopathies or degeneration age-related macular (AMD), rheumatoid arthritis, and psoriasis (Folkman <i>et al. J. Biol. Chem</i>. 267: 10931-10934 (1992); Klagsbrun <i>et al. Annu Rev. Physiol</i>. 53: 217-239 (1991); and Garner A,<i>Vascular diseases In: Pathobiology of ocular disease. A dynamic approach</i>. Garner A, Klintworth GK, Eds. 2nd Edition Marcel Dekker, NY, p. 1625-1710 (1994)). In the case of solid tumors, neovascularization allows cells tumors get an advantage of growth and autonomy proliferative compared to normal cells. By consequently, a correlation between the density of microvessels in tumor sections and in patients who survive breast cancer as well as several different tumors (Weidner <i>et to the. N Engl J Med</i> 324: 1-6 (1991); Horak<i>et to the. Lancet</i> 340: 1120-1124 (1992); and Macchiarini<i>et al. Lancet</i> 340:145-146 (1992)).

p0004The search for positive regulators of the Angiogenesis has produced many candidates, which include aFGF, bFGF, TGF-?, TGF-?, HGF, TNF-?, Angiogenin, IL-8, etc. (Folkman<i>et al</i>. and Klagsbrun<i>et to the</i>). The negative regulators identified so far include thrombospondin (Good <i>et al. Proc. Natl. Acad. Sci. USES</i>. 87: 6624-6628 (1990)), the fragment Prolactin 16 kilodalton N-terminal (Clapp<i>et al. Edocrinology</i>, 133: 1292-1299 (1993)), angiostatin (O'Reilly <i>et al. Cell</i>, 79: 315-328 (1994)) and endostatin (O'Reilly <i>et al. Cell</i>, 88:277-285 (1996)).

p0005The work done during the last years has established the key role the endothelial growth factor Vascular (VEGF) in the regulation of normal and abnormal angiogenesis (Ferrara <i>et al. Endocr Rev</i>. 18: 4-25 (1997)). The discovery that the loss of even a single allele of VEGF results in embryonic lethality, suggests that this factor plays an irreplaceable role in development and vascular system differentiation (Ferrara <i>et al</i>.). In addition, VEGF has been shown to be a key mediator in the neovascularization associated with tumors and intraocular disorders (Ferrara <i>et al</i>.). VEGF mRNA is overexpresses in most human tumors examined (Berkaman <i>et al. J Clin Invest</i>91: 153-159 (1993); Brown<i>et al. Human Pathol</i>. 26: 86-91 (1995); Brown<i>et al. Cancer Beef</i>. 53: 4727-4735 (1993); Mattern<i>et al. Brit. J. Cancer</i>. 73: 931-934 (1996); and Dvorak<i>et al. Am J. Pathol</i>. 146: 1029-1039 (1995)). In addition, the concentration of VEGF in eye fluids is highly correlated with the presence of active vessel proliferation blood in patients with diabetes or other retinopathies related to ischemia (Aiello <i>et al. N. Engl. J. Med</i>. 331: 1480-1487 (1994)). In addition, recent studies have demonstrated the location of VEGF in neovascular membranes Choroidal in patients affected by AMD (Lopez <i>et al. Invest. Ophtalmo Vis. Sci</i>. 37: 855-868 (1996)). The anti-VEGF neutralizing antibodies suppress the growth of a diversity of human tumor cell lines in nude mice (Kim <i>et al. Nature</i> 362: 841-844 (1993); Warren<i>et al. J. Clin. Invest</i>. 95: 1789-1797 (1995); Borgström<i>et to the. Cancer Res</i>. 56: 4032-4039 (1996); and Melnyk<i>et al. Cancer Res</i>. 56: 921-924 (1996)) and they also inhibit intraocular angiogenesis in disorder models ischemic retinals (Adamis <i>et al. Arch. Ophthalmol</i>. 114: 66-71 (1996)). Therefore, the antibodies monoclonal anti-VEGF or other inhibitors of VEGF action are promising candidates for the treatment of solid tumors and various neovascular disorders intraocular

Summary of the invention

p0006As defined in the claims, the invention provides anti-VEGF antibodies and anti-VEGF antibody variants with properties desirable from a therapeutic perspective, including strong binding affinity for VEGF; the ability to inhibit proliferation VEGF induced endothelial cells <i>in vitro</i>; and the ability to inhibit VEGF-induced angiogenesis <i>in alive</i>.

p0007Anti-VEGF antibody humanized or anti-VEGF antibody variant preferred in this document binds to human VEGF with a value K d of not more than about 1 x 10-8 M and preferably not more than about 5 x 10-9 M. In addition, the humanized anti-VEGF antibody or variant can have an ED50 value of no more than about 5 nM to inhibit VEGF induced proliferation of endothelial cells <i>in vitro</i>. Humanized anti-VEGF antibodies or variants of particular interest in this document are those that inhibit at least about 50% of tumor growth in an A673 tumor model <i>in vivo</i>, at a dose of antibody 5 mg / kg

p0008In one embodiment, the antibody anti-VEGF has a variable domain of the chain heavy and light, where the variable domain of the heavy chain comprises hypervariable regions with the following sequences of amino acids: CDRH1 (GYX1 FTX2 YGMN, where X1 is T or D and X2 is N or H; SEC.ID. No. 128), CDRH2 (WINTYTGEPTYAADFKR; SEQ ID No. 2) and CDRH3 (YPX_ {1} YYGX_ {2} SHWYFDV, where X_ {1} is Y or H and X2 is S or T; SEC.ID. No. 129). For example, the domain heavy chain variable can comprise the sequences of amino acids of CDRH1 (GYTFNYGMN; SEQ ID No. 1), CDRH2 (WINTYTGEPTYAADFKR; SEC.ID. No. 2) and CDRH3 (YPHYYGSSHWY FDV; SEC.ID. No. 3). Preferably, the three hypervariable regions of Heavy chain are provided in a human flanking region, for example, in the form of a contiguous sequence represented by the following formula: FR1-CDRH1-FR2-CDRH2-FR3-CDRH3-FR4.

p0009The invention also provides a domain. heavy chain variable of an antibody anti-VEGF comprising the sequence of amino acids:

p0010EVQLVESGGGLVQPGGSLRLSCAASGYX_ {1} FTX_ {2} YGMNWVRQAPGKGLEWVGWINTYTGEPTYAADFKR RFTFSLDTSKSTAYLQMNSLRAEDTAVYYCAKYPX_ {3} YYGX_ {V} SHYGX_ {V} SHYGX_ {V} SHYGX_DV (SEQ ID 125), where X 1 is T or D; X 2 is N or H; X_ {3} is Y or H and X_ {4} is S or T. A domain sequence particularly useful heavy chain variable is that of the humanized antibody F (ab) -12 of Example 1 and comprises the sequence of the heavy chain variable domain of SEQ ID No. 7. These sequences of the variable domain of the chain Preferred weights can be combined with the following sequences of the variable domain of the light chain or with other sequences of the variable domain of the light chain, preferred, provided that The antibody produced in this way binds to human VEGF.

p0011The antibody of the invention has the light chain variable domain sequences that are defined in the claims that can be combined with the sequences of the variable domain of the heavy chain identified above or with other sequences of the heavy chain variable domain, to condition that the antibody produced in this way maintains the ability to join human VEGF. For example, the variable domain of the light chain can comprise hypervariable regions with the following amino acid sequences: CDRL1 (SASQDISNYLN; SEC.ID.No. 4), CDRL2 (FTSSLHS; SEC.ID.No. 5) and CDRL3 (QQYSTVPWT; SEC.ID.No. 6). Preferably, the three hypervariable regions of the chain lightweight are provided in a human flanking region, by example, in the form of a contiguous sequence represented by the following formula: FR1-CDRL1-FR2-CDRL2-FR3-CDRL3-FR4.

p0012In one embodiment, the invention provides a variable domain of the light chain of an antibody humanized anti-VEGF comprising the sequences of amino acids:

p0013DIGX_ {1} TQSPSSLSASVGDRVTITCSASQDISNYLNWYQQKPGKAPKVLIYFTSSLHSGVPSRFSGSGSGTDF TLT ISSLQPEDFATYYCQQYSTVPWTFGQGTKVEIKR (SEC.ID.No. 124), where X_ {1} is M or L. A sequence of the light chain variable domain particularly useful is that of the humanized antibody F (ab) -12 of Example 1 and comprises the sequence of the light chain variable domain of SEQ ID NO. 8.

p0014An antibody of the invention, which is defined in the claims may be a variant of an antibody parental anti-VEGF (said parental antibody is preferably a humanized anti-VEGF antibody or human), where the variant joins human VEGF and comprises a amino acid substitution in a hypervariable region of the domain variable heavy and light antibody chain parental anti-VEGF. The variant preferably has one or more substitutions in one or more hypervariable regions of the anti-VEGF antibody. Preferably, the substitution or substitutions are in the variable domain of the heavy chain of the parental antibody. For example, the replacement or amino acid substitutions may be in the CDRH1 and / or CDRH3 of the variable domain of the heavy chain. Preferably, there are substitutions in these two hypervariable regions. In this document shows that such variants of "affinity matured "bind to human VEGF more strongly than the antibody parental anti-VEGF from which they have been generated, is that is, they have a K_ {d} value that is significantly less than that of the parental anti-VEGF antibody. Preferably, the variant has an ED50 value to inhibit the VEGF induced proliferation of endothelial cells <i>in vitro</i> which is at least about 10 times lower, preferably at least about 20 times lower, and more preferably at least about 50 times lower, than the of the parental anti-VEGF antibody. A variant Particularly preferred is variant Y0317 of Example 3, which It has a CDRH1 that comprises the amino acid sequence: GYDFTHYGMN (SEQ ID No. 126) and a CDRH3 comprising the sequence of amino acids: YPYYYGTSHWYFDV (SEC.ID.No. 127). These regions Hypervariables and CDRH2 are usually provided in a region human flanking, for example, resulting in a domain heavy chain variable comprising the sequence of amino acids of SEQ ID No. 116. These variable domain sequences of the heavy chain are optionally combined with a domain variable of the light chain comprising the sequence of amino acids of SEQ ID No. 124, and preferably the sequence of amino acids of the light chain variable domain of SEQ ID NO. 115

p0015Various forms of the antibody are contemplated in this document. For example, the anti-VEGF antibody it can be a full length antibody (for example, having an intact human Fc region) or an antibody fragment (for example, a Fab, Fab 'or F (ab') 2). In addition, the antibody can be marked with a detectable marker, immobilized in one phase solid and / or conjugate with a heterologous compound (such as a cytotoxic agent).

p0016Diagnostic and therapeutic uses are contemplated for the antibody. A diagnostic application concerns a method for determining the presence of VEGF protein comprising expose a sample that is believed to contain VEGF protein to anti-VEGF antibody and determine the binding of antibody to the sample. A kit comprising the antibody and instructions for using the antibody to detect the VEGF protein

p0017The invention also provides: an acid isolated nucleic encoding the antibody; a vector that comprises that nucleic acid, optionally operatively linked to control sequences recognized by a host cell transformed with the vector; a host cell that comprises that vector; a process to produce the antibody, which comprises culturing the host cell so that the acid is expressed nucleic and, optionally, recover the antibody from the culture of host cells (for example, from the culture medium of host cells). The invention also provides a composition comprising the anti-VEGF antibody and a pharmaceutically acceptable carrier or diluent. The composition For therapeutic use it is sterile and can be lyophilized. The invention it also provides a method to treat a mammal that suffers from a tumor or retinal disorder, which comprises administering an amount therapeutically effective anti-VEGF antibody to mammal.

Brief description of the drawings

p0018Figures 1A and 1B represent the sequences of amino acids of the heavy chain variable domain (SEC.ID. No. 9) and the light chain (SEC.ID. No. 10) of muMAbVEGF A.4.6.1, the heavy chain variable domain (SEQ ID No. 7) and light chain (SEQ ID No. 8) of F (ab) humanized (F (ab) -12) and flanking regions human consensus (hum III for subgroup III heavy chains (SEC.ID. No. 11); hum \ kappaI for subgroup I of light chains κ (SEQ ID No. 12)). Figure 1A aligns the sequences of the variable domain of the heavy chain and Figure 1B aligns the Light chain variable domain sequences. Asterisks indicate differences between F (ab) -12 humanized and the murine Mab or between F (ab) -12 and the human flanking region. The Determinant Regions of Complementarity (CDR) are underlined.

p0019Figure 2 is a tape diagram of the model of the VL and VH domains of F (ab) -12 humanized VL domain is shown in brown with color CDRs cinnamon. The side chain of the L46 residue is shown in yellow. He VH domain is shown in purple with CDR in pink. The chains sides of the remains of VH changed from human to murine are They show in yellow.

p0020Figure 3 represents the inhibition of VEGF-induced mitogenesis by F (ab) -12 Humanized anti-VEGF of Example 1. They were seeded capillary endothelial cells obtained from bovine adrenal cortex at a density of 6 x 10 3 cells / well in six plates wells, as described in Example 1. VEGF muMAb were added A.4.6.1 or rhuMAb VEGF (IgG1; F (ab) -12) at indicated concentrations. After 2-3 hours, it added rhVEGF165 to the final concentration of 3 ng / ml. After five or six days, the cells were treated with trypsin and they counted. The values ​​shown are means of determinations duplicates The variation of the average does not exceed 10%.

p0021Figure 4 shows growth inhibition of the tumor <i>in vivo</i> by F (ab) -12 Humanized anti-VEGF of Example 1. They were injected A673 rhabdomyosarcoma cells in nude mice BALB / ca a density of 2 x 10 6 per mouse. Starting 24 hours after the inoculation of the tumor cells, the animals were injected with a control MAb, muMAb VEGF A.4.6.1 or rhuVEGF MAb (IgG1; F (ab) -12) twice a week, by way intraperitoneal The dose of control MAb was 5 mg / kg; the MAb anti-VEGF were given at 0.5 or 5 mg / kg, as indicated (n = 10). Four weeks after the cell injection tumor, the animals underwent euthanasia and the tumors and weighed. *: significant difference when compared with the control group by ANOVA (p &lt;0.05).

p0022Figures 5A and 5B show respectively the amino acid sequences of the variable domains of the chain light and heavy antibody A.4.6.1 murine (SEQ ID No. 10 for VL and SEC.ID. No. 9 for the VH) and the hu2.0 variants (SEC.ID. No. 13 for the VL and SEC.ID. No. 14 for the VH) and hu2.10 (SEC.ID. No. 15 for the VL and SEC.ID. No. 16 for the VH) of humanized A.4.6.1 of the Example 2. Sequence numbering is in accordance with Kabat <i>et al., Sequences of Proteins of Immunological Interest</i>, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD. (1991) and mismatches are indicated by asterisks (A.4.6.1 murine vs hu2.0) or points (hu2.0 vs hu2.10). The hu2.0 variant contains only CDR (labeled) sequences of the grafted murine antibody in a consensus flanking region of subgroup I of light chains human κ (SEC.ID.12) and the consensus flanking region of the subgroup III of heavy chains (SEC.ID. No. 11). hu2.10 was the clone Humanized consensus obtained from classification experiments of phages described in this document.

p0023Figure 6 represents the flanking remains set as a target for randomization in Example 2.

p0024Figure 7 represents the construction of phagemid for superficial fusion presentation Fab-pIII in phages. The phagemid encodes a version humanized from the Fab fragment for antibody A.4.6.1 fused to a portion of the M13 gene III envelope protein. The fusion protein consists of the Fab bound to the carboxyl terminal of the heavy chain to a single glutamine residue (from the suppression of an amber codon in <i>E. coli supE</i>), after the C-terminal region of gene III protein (residues 249-406). The transformation in<i>E. coli</i>F +, followed by phage super-infection M13KO7 auxiliary, produces phagemid particles in which a small portion of these shows a single copy of the protein from fusion.

p0025Figures 8A-E represent the double stranded nucleotide sequence (SEQ ID No. 99) for the phMB4-19-1.6 antibody vector presented in phages in Example 3 and the two sequences of amino acids encoded in this way (SEQ ID No. 130 and No. 100).

p0026Figures 9A and 9B respectively represent an alignment of amino acid sequences for domains variables of the light and heavy chain of variants affinity affinity anti-VEGF in Example 3, in comparison with F (ab) -12 of Example 1 (SEC.ID. No. 8 and No. 7 for the variable domains of the light chain and heavy, respectively). The CDRs are underlined and named such as L, light chain, or H, heavy chain, and numbers 1-3. The remains are listed sequentially in the VL and VH domains, unlike the Kabat numbering scheme. The template molecule, MB1.6 (SEQ ID No. 101 and No. 102 for the variable domains of the light and heavy chain, respectively) together with the variants: H2305.6 (SEC.ID. No. 103 and No. 104 for variable domains of the light and heavy chain, respectively), Y0101 (SEC.ID. No. 105 and No. 106 for the variable domains of the light and heavy chain, respectively), and Y0192 (SEQ ID No. 107 and No. 108 for the variable domains of the light and heavy chain respectively). The differences with F (ab) -12 are shown in boxes shaded.

p0027Figures 10A and 10B represent an alignment of the amino acid sequences for the variable domains of the light and heavy chain respectively of variants matured affinity anti-VEGF of Example 3 in comparison with F (ab) -12 of Example 1 (SEC.ID. No. 8 and No. 7 for the variable domains of the light chain and heavy, respectively). The CDRs are underlined and named by L, light chain, or H, heavy chain, and numbers 1-3. The variants are called Y0243-1 (SEC.ID. No. 109 and No. 110 for domains light and heavy chain variables, respectively), Y0238-3 (SEC.ID. No. 111 and No. 112 for domains light and heavy chain variables, respectively), Y0313-1 (SEC.ID. No. 113 and No. 114 for domains variables of the light and heavy chain, respectively), and Y0317 (SEC.ID. No. 115 and No. 116 for the variable domains of the chain light and heavy, respectively). The differences with F (ab) -12 are shown in boxes shaded.

p0028Figure 11 represents the test results of HuVEC activity in Example 3 for variants Y0238-3, Y0192 and Y0313-1 as well as F (ab) -12 full length of Example 1.

p0029Figure 12 represents the inhibition of VEGF-induced mitogenesis by F (ab) -12 of full length of Example 1 (rhuMAb VEGF), a Fab fragment of F (ab) -12 of Example 1 (rhuFab VEGF), and a Fab fragment of the matured affinity variant Y0317 of Example 3 (rhuFab VEGF (matured affinity)).

Detailed description of the preferred embodiments

I.

Definitions

p0030The term "human VEGF" as used in this document refers to the cell growth factor of the Human vascular endothelium of 165 amino acids, and the factors of vascular endothelial cell growth of 121, 189 and 206 related amino acids, as described by Leung <i>et al., Science</i> 246: 1306 (1989), and Houck <i>et al., Mol. Endocrin</i>. 5: 1806 (1991) together with allelic and processed forms of origin natural of those growth factors.

p0031The present invention provides antibodies anti-VEGF antagonists that are able to inhibit one or more of VEGF activities, for example, its activity mitogenic or angiogenic. VEGF antagonists work interfering with the binding of VEGF to a cellular receptor, disabling or eliminating cells that have been activated by VEGF, or interfering with the activation of vascular endothelial cells after binding of VEGF to a cellular receptor. All these intervention points by a VEGF antagonist will be considered equivalents for the purposes of this invention.

p0032The term "VEGF receiver" or "VEGFr" as used herein refers to a cellular receptor for VEGF, usually a cell surface receptor found in vascular endothelial cells, as well as variants thereof maintain the ability to join hVEGF. An example of a receiver VEGF is the receiver <i>fms</i> tyrosine kinase type (<i>flt</i>), a transmembrane receptor of the tyrosine kinase family. DeVries<i>et al., Science</i> 255: 989 (1992); Shibuya<i>et al., Oncogene</i> 5: 519 (1990). The receptor <i>flt</i> comprises a extracellular domain, a transmembrane domain, and a domain intracellular with tyrosine kinase activity. Extracellular domain is involved in the union of VEGF while the domain Intracellular is involved in signal transduction. Other example of a VEGF receiver is the receiver<i>flk-1</i> (also mentioned as KDR). Matthews<i>et al., Proc. Nat. Acad. Sci</i>. 88: 9026 (1991); Terman<i>et al., Oncogene</i> 6: 1677 (1991); Terman <i>et al., Biochem. Biophys Res. Commun</i>. 187: 1579 (1992). The union of VEGF to receiver <i>flt</i> results in the formation of at least two high molecular weight complexes, which have molecular weight apparent of 205,000 and 300,000 Dalton. It is believed that the complex of 300,000 Dalton is a dimer that comprises two receptor molecules bound to a single VEGF molecule.

p0033The term "epitope A.4.6.1" when used in this document, unless otherwise indicated, refers to the region of human VEGF to which antibody A.4.6.1 binds described in Kim <i>et al., Growth Factors</i> 7:53 (1992) and Kim<i>et al. Nature</i> 362:841 (1993).

p0034"Treatment" refers to both treatment therapeutic as prophylactic or preventive measures. The individuals in need of treatment include those who already they have the disorder as well as those in which the disorder is to prevent it

p0035"Mammal" for treatment purposes is refers to any animal classified as a mammal, including humans, domestic and farm animals, and zoo animals, sports or companion, such as dogs, horses, cats, cows, etc. Preferably, the mammal is a human.

p0036"Antibodies" (Ab) and "immunoglobulins" (Ig) are glycoproteins that have the same characteristics structural. While antibodies show specificity binding to a specific antigen, immunoglobulins include both antibodies and other antibody-like molecules that lack of antigen specificity. The polypeptides of the last type are produce, for example, at low levels by the lymphoid system already levels increased by myelomas.

p0037"Native antibodies" and "immunoglobulins native "are usually heterotetrameric glycoproteins of approximately 150,000 Daltons of two identical light chains (L) and two identical heavy chains (H). Each light chain is attached to a heavy chain by a covalent disulfide bond, although the amount of disulfide bonds varies between heavy chains of different immunoglobulin isotypes. Each heavy and light chain it also has intracatenary disulfide bridges spaced so regular. Each heavy chain has at one end a variable domain (V_ {H}) followed by several constant domains. Each light chain it has a variable domain at one end (V_ {L}) and a domain constant at its other end; the constant domain of the chain lightweight is aligned with the first constant domain of the chain heavy, and the variable domain of the light chain is aligned with the variable domain of the heavy chain. It is believed that remains of particular amino acids form a contact surface between the variable domains of the light and heavy chain.

p0038The term "variable" refers to the fact of that certain portions of the variable domains differ greatly measured in the sequence between antibodies and used in binding and specificity of each particular antibody for its antigen particular. However, the variability is not distributed. evenly across all variable domains of antibodies. It is concentrated in three segments called hypervariable regions both in the variable domains of the light chain and the heavy chain The most highly conserved portions of Variable domains are called flanking regions (FR). The variable domains of native heavy and light chains Each comprises four FR (FR1, FR2, FR3 and FR4, respectively), which largely adopt a configuration lamina-?, connected by three regions hypervariables, which form connecting loops, and in some cases they are part of the lamina-β structure. The hypervariable regions in each chain are held together in close proximity to FR and, with hypervariable regions from another chain, contribute to the formation of the binding site to antibody antigen (see Kabat <i>et al., Sequences of Proteins of Immunological Interest</i>, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD. (1991), pages 647-669). Constant domains are not directly involved in the binding of the antibody to an antigen, but show various effector functions, such as participation of the antibody in cell-dependent toxicity of antibody.

p0039The term "hypervariable region" when used herein refers to the amino acid residues of a antibody that are responsible for antigen binding. The region Hypervariable comprises amino acid residues of a "region complementarity determinant "or" CDR "(that is, the residues 24-24 (L1), 50-56 (L2) and 89-97 (L3) in the variable domain of the chain light and 31-35 (H1), 50-65 (H2) and 95-102 (H3) in the variable domain of the chain heavy Kabat<i>et al., Sequences of Proteins of Immunological Interest</i>, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD. (1991)) and / or the remains of a "loop hypervariable "(that is, residues 26-32 (L1), 50-52 (L2) and 91-96 (L3) in the variable domain of the light chain and 26-32 (H1), 53-55 (H2) and 96-101 (H3) in the variable domain of the heavy chain; Chothia and Lesk <i>J. Mol. Biol</i>. 196: 901-917 (1987)). The remains "flanking" or "FR" are the remains of the domains variables other than the remnants of the hypervariable region such as It is defined in this document.

p0040Digestion of antibodies with papain produces two identical antigen binding fragments, called fragments "Fab", each with a single antigen binding site, and a residual "Fc" fragment, whose name reflects its ability to crystallize easily. Pepsin treatment produces a F (ab ') 2 fragment having two combination sites of antigen and is still capable of antigen cross-linking.

p0041"Fv" is the minimum antibody fragment which contains a complete site of recognition or union with antigen. This region consists of a dimer of a variable domain of the heavy chain and one of the light chain in strong association not covalent It is in the configuration in which the three regions hypervariables of each variable domain interact to define an antigen binding site on the dimer surface V_ {H} -V_ {L}. Collectively, the six regions hypervariables give the antibody specificity of binding to antigen. However, even a single variable domain (or means of an Fv comprising only three specific hypervariable regions for an antigen) has the ability to recognize and bind antigen, although at a lower affinity than the complete binding site.

p0042The Fab fragment also contains the domain light chain constant and the first constant domain (CH1) of heavy chain Fab 'fragments differ from Fab fragments by the addition of a few residues in the carboxyl terminal of CH1 domain of the heavy chain that includes one or more cysteines of the hinge region of the antibody. Fab'-SH is the denomination in this document for Fab 'in which the rest or cysteine ​​residues of the constant domains contain a group free thiol. F (ab ') 2 antibody fragments in a principle occurred as pairs of Fab 'fragments that have hinge cysteines between them. Others are also known chemical couplings of antibody fragments.

p0043The "light chains" of antibodies (immunoglobulins) of any vertebrate species can be assigned to two distinctly different types, called kappa (\ kappa) and lambda (λ), based on the amino acid sequences of their constant domains

p0044Depending on the amino acid sequence of the constant domain of their heavy chains, immunoglobulins are Assign to different classes. There are five main classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, and several of these they can be further divided into subclasses (isotypes), by example, IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. Domains heavy chain constants that correspond to different classes of immunoglobulins are called α, δ, ε, γ, and µ, respectively. The structures are well known of subunits and three-dimensional configurations of Different kinds of immunoglobulins.

p0045The term "antibody" is used in this document in the broadest sense and specifically covers monoclonal antibodies (including monoclonal antibodies of full length), polyclonal antibodies, antibodies multispecific (e.g. bispecific antibodies), and antibody fragments while showing activity biological desired.

p0046"Antibody fragments" comprise a portion of a full length antibody, generally the variable or antigen binding domain thereof. The examples of antibody fragments include Fab, Fab 'fragments, F (ab ') 2, and Fv; bivalent fragments; antibodies linear; single chain antibody molecules; and antibodies multi-species formed from fragments of antibodies

p0047The term "monoclonal antibody" as used herein refers to an antibody obtained from of a population of substantially homogeneous antibodies, that is, the individual antibodies that comprise the population are identical except for possible mutations of natural origin that They may be present in minor amounts. Antibodies monoclonal are highly specific, being directed against a unique antigenic site. In addition, in contrast to preparations of conventional (polyclonal) antibodies that typically include different antibodies directed against different determinants (epitopes), each monoclonal antibody is directed against a unique determinant in the antigen. The "monoclonal" modifier indicates the character of the antibody that is obtained from a population substantially homogeneous of antibodies, and it is not construed that it requires an antibody production by any particular method. For example, the monoclonal antibodies to be used according to the present invention can be made by the hybridoma method first described by Kohler <i>et al., Nature</i> 256: 495 (1975), or can be done by recombinant DNA methods (see, for example, U.S. Patent No. 4,816,567). "Antibodies monoclonal "can also be isolated from libraries of phage antibodies using the techniques described in Clackson<i>et al., Nature</i> 352: 624-628 (1991) and Marks<i>et al., J. Mol. Biol</i>. 222: 581-597 (1991), for example.

p0048Monoclonal antibodies include specifically in this document "chimeric" antibodies (immunoglobulins) in which a portion of the heavy chain and / or lightweight is identical or homologous to corresponding sequences in antibodies obtained from a particular species or belonging to a particular class or subclass of antibody, while the rest of the chain or chains is identical or homologous to sequences corresponding in antibodies obtained from another species or that belong to another class or subclass of antibody, as well as fragments of said antibodies, while showing activity Biological Desired (US Patent No. 4,816,567; and Morrison <i>et al., Proc. Natl. Acad. Sci. USA</i> 81:6851-6855 (1984)).

p0049The "humanized" forms of antibodies do not humans (e.g. murine) are chimeric antibodies that contain a minimal sequence obtained from a non-immunoglobulin human For the most part, humanized antibodies are human immunoglobulins (receptor antibody) in which the residues of the hypervariable region of the receptor are replaced by remnants of the hypervariable region of a non-human species (antibody donor) such as a mouse, rat, rabbit or non-human primate that It has the specificity, affinity, and capacity desired. In some cases, the remains of the flanking region (FR) of the Human immunoglobulin are replaced by non-human waste corresponding. In addition, humanized antibodies can comprise residues that are not found in the receptor antibody or in the donor antibody. These modifications are made to further improve the function of the antibody. Usually, the humanized antibody will comprise substantially all of the minus one, and typically two variable domains, in which all or substantially all of the hypervariable regions correspond to those of a non-human immunoglobulin and all or substantially all of the FRs are those of a sequence of a human immunoglobulin. The humanized antibody optionally will also comprise at least a portion of a constant region (Fc) of an immunoglobulin, typically that of a human immunoglobulin. For more details, see jones <i>et al., Nature</i> 321: 522-525 (1986); Reichmann<i>et al., Nature</i> 332: 323-329 (1988); and lend, <i>Curr. Op. Struct. Biol</i>. 2:593-596 (1992).

p0050Fv chain "antibody fragments simple "or" sFv "comprise the domains V_ {H} and V_ {L} of the antibody, where these domains are present in a single polypeptide chain. Generally, the Fv polypeptide also comprises a polypeptide linker between domains VH and V_ {L} that allows the sFv to form the desired structure for antigen binding. For an analysis of sFv see Pluckthun in<i>The Pharmacology of Monoclonal Antibodies</i>, vol. 113, Rosenburg and Moore eds. Springer-Verlag, New York, p. 269-315 (1994).

p0051The term "bivalent fragments" is refers to small antibody fragments with two binding sites to antigen, said fragments comprising a variable domain of the heavy chain (V_ {H}) connected to a variable domain of the light chain (V L) in the same polypeptide chain (V_ {H} -V_ {L}). Using a linker that is too short to allow pairing between two domains in the same chain, the domains are forced to match the complementary domains of another chain and create two binding sites to antigen. Bivalent fragments are described more fully in, for example, EP 404,097; WO 93/11161; and Hollinger <i>et al., Proc. Natl. Acad. Sci. USA</i> 90:6444-6448 (1993).

p0052The expression "linear antibodies" when used throughout this application refers to the antibodies described in Zapata <i>et al. Eng Protein</i>. 8 (10): 1057-1062 (1995). Briefly, these antibodies comprise a pair of tandem Fd segments (V_ {H} -C_ {H} 1-V_ {H} -C_ {1}) which form a pair of antigen binding regions. Antibodies linear can be bispecific or monospecific.

p0053A "variant" of an antibody anti-VEGF, refers in this document to a molecule that differs in the amino acid sequence of a sequence of amino acids of the parental anti-VEGF antibody due to the addition, deletion and / or substitution of one or more traces of amino acids in the sequence of the parental antibody. In the preferred embodiment, the variant comprises one or more substitutions of amino acids in one or more hypervariable regions of the antibody parental For example, the variant can comprise at least one, for example, from about one to about ten, and preferably from about two to about five, substitutions in one or more hypervariable regions of the antibody parental Generally, the variant will have a sequence of amino acids that have at least 75% sequence identity of amino acids with the sequences of the variable domain of the chain heavy or light parental antibody (for example, as in SEQ ID No. 7 or No. 8), more preferably at least 80%, more preferably at least 85%, more preferably at least the 90%, and much more preferably at least 95%. Identity u homology regarding this sequence is defined in this document as the percentage of amino acid residues in the sequence candidate who are identical to the remains of the parental antibody, after aligning the sequences and entering gaps, if it is necessary, to achieve the maximum percentage of identity of sequence. No extension, deletion, or insertion N-terminal, C-terminal, or internal in the antibody sequence will be interpreted to affect the sequence identity or homology. The variant maintains the ability to bind human VEGF and preferably has properties which are superior to those of the parental antibody. For example, the variant may have a stronger binding affinity, capacity enhanced to inhibit VEGF induced cell proliferation endothelial and / or increased ability to inhibit angiogenesis VEGF induced <i>in vivo</i>. To analyze these properties, a Fab form of the variant must be compared to a Fab form of the parental antibody or a full-length form of the variant to a full length form of the parental antibody, by example, although it has been found that the antibody format anti-VEGF affects its activity in the trials of Biological activity described in this document. The variant of a antibody of particular interest in this document is one that shows an enhancement of at least about 10 times, preferably at least about 20 times, and more preferably at least about 50 times, in the activity biological when compared to the parental antibody.

p0054The "parental" antibody in this document it is one that is encoded by an amino acid sequence used for the preparation of the variant. Preferably, the antibody parental has a human flanking region and, if present, has a constant region or regions of human antibody. By For example, the parental antibody can be a humanized antibody or human.

p0055An "isolated" antibody is one that has been identified or separated and / or recovered from a component of its medi or natural. The polluting components of its natural environment are materials that could interfere with diagnostic uses or therapeutic for the antibody, and may include enzymes, hormones, and other protein or non-protein solutes. In realizations preferred, the antibody will be purified (1) to more than 95% by weight of the antibody determined by the Lowry method, and more preferably to more than 99% by weight, (2) to a sufficient degree to obtain at least 15 amino acid sequence residues N-terminals or internal by the use of a rotating bowl sequencer, or (3) until homogeneous by SDS-PAGE under reducing or non-reducing conditions using Coomassie blue or, preferably, staining with silver. A isolated antibody includes antibody <i>on-site</i> in cells recombinants since at least one component of the natural environment of antibody will not be present. Generally, however, the isolated antibody will be prepared for at least one step of purification.

p0056The term "designated epitope" when used in this document it refers to the anti-VEGF antibody fused to an "epitope signal". The pointing polypeptide of epitope has enough remains to provide an epitope against which an antibody can be made, but it is sufficiently short so as not to interfere with antibody activity anti-VEGF The epitope pointer preferably is unique enough that the antibody against it does not react substantially crosswise with other epitopes. The appropriate pointing polypeptides generally have at least 6 amino acid residues and usually between about 8-50 amino acid residues (preferably between approximately 9-30 remains). Examples include the flu HA signaling polypeptide and its 12CA5 antibody (Field <i>et to the. Mol. Cell Biol</i>. 8: 2159-2165 (1988)); he c-myc signal and antibodies 8F9, 3C7, 6E10, G4, B7 and 9E10 in front of it (Evan <i>et al., Mol. Cell Biol</i>. 5 (12): 3610-3616 (1985)); and the flag of the Herpes Simplex virus glycoprotein D (gD) and its antibody (Paborsky <i>et al., Protein Engineering</i> 3 (6): 547-553 (1990)). In certain embodiments, the epitope signal is an "epitope binding to rescue receiver. "As used herein, the term "rescue receptor binding epitope" refers to a epitope of the Fc region of an IgG molecule (for example, IgG1, IgG2, IgG3, or IgG4) that is responsible for increase serum half-life <i>in vivo</i> of the molecule IgG

p0057The term "cytotoxic agent" as used in this document it refers to a substance that inhibits or prevents the cell function and / or cause cell destruction. The intention is to that the term includes radioactive isotopes (for example, I 131, I 125, Y 90 and Re 186), agents chemotherapeutics, and toxins such as enzymatically toxins active of bacterial, fungal, plant or animal origin, or fragments thereof.

p0058A "chemotherapeutic agent" is a Chemical compound useful in the treatment of cancer. The examples of Chemotherapeutic agents include Adriamycin, Doxorubicin, 5-Fluorouracil, Cytosine Arabinoside ("Ara-C"), Cyclophosphamide, Tiotepa, Taxotere (docetaxel), Busulfan, Cytoxin, Taxol, Methotrexate, Cisplatin, Melfalan, Vinblastine, Bleomycin, Etoposide, Ifosfamide, Mitomycin C, Mitoxantrone, Vincristine, Vinorelbine, Carboplatin, Teniposide, Daunomycin, Carminomycin, Aminopterin, Dactinomycin, Mitomycins, Spiramincins (see US Patent No. 4,675,187), Melfalan and other related nitrogen mustards.

p0059The term "prodrug" as used in this request refers to a precursor form or derived from a pharmaceutically active substance that is less cytotoxic for tumor cells compared to the parental drug and is capable of activating or becoming enzymatically in the parental form more active See, for example, Wilman, "Prodrugs in Cancer Chemotherapy " <i>Biochemical Society Transactions</i>, 14, p. 375-382, 615th Meeting Belfast (1986) and Stella <i>et to the</i>., "Prodrugs: A Chemical Approach to Targeted Drug Delivery ", <i>Directed Drug Delivery</i>Borchardt <i>et to the</i>., (ed.), p. 247-267, Humana Press (1985). The prodrugs of this invention include, but are not limited to, prodrugs containing phosphate, prodrugs containing thiophosphate, sulfate-containing prodrugs, prodrugs that contain peptides, prodrugs modified by D-amino acids, glycosylated prodrugs, prodrugs containing β-lactam, prodrugs containing optionally substituted phenoxyacetamide or prodrugs containing optionally substituted phenylacetamide, 5-fluorocytosine and other prodrugs of 5-fluorouridine that can become the drug Cytotoxic free more active. Examples of cytotoxic drugs which can be derivatized into a prodrug form for use in This invention includes, but is not limited to, agents. chemotherapeutic agents described above.

p0060The word "marker" when used in this document refers to a detectable compound or composition that is conjugates directly or indirectly to the antibody. The marker can be detectable by itself (for example, markers of radioisotopes or fluorescent markers) or, in the case of a Enzymatic marker, can catalyze the chemical alteration of a compound or substrate composition that is detectable.

p0061By "solid phase" is meant a matrix non-aqueous to which the antibody of the present can adhere invention. Examples of solid phases included in this document they include those formed partially or completely of glass (by for example, controlled pore glass), polysaccharides (for example, agarose), polyacrylamides, polystyrene, polyvinyl alcohol and silicones In certain embodiments, depending on the context, the phase solid may comprise the well of a test plate; in others it is a purification column (for example, a column of affinity chromatography). This term also includes a phase Discontinuous solid of discrete particles, such as those described in US Patent No. 4,275,149.

p0062A "liposome" is a small vesicle composed of various types of lipids, phospholipids and / or a surfactant that is useful for the delivery of a drug (such as the anti-VEGF antibodies described in this document and, optionally, a chemotherapeutic agent) to a mammal. The components of the liposome are usually placed in a bilayer formation, similar to the lipid placement of biological membranes An "isolated" nucleic acid molecule it is a nucleic acid molecule that is identified and separated of at least one pollutant nucleic acid molecule that is generally associated in the natural source of the nucleic acid of antibody. An isolated nucleic acid molecule is distinct in the form or means in which it is found in nature. Thus, the isolated nucleic acid molecules differ from the nucleic acid molecule that exists in natural cells. Without However, an isolated nucleic acid molecule includes a molecule. of nucleic acid contained in cells that generally express the antibody where, for example, the nucleic acid molecule is in a different chromosomal location from that of cells natural

p0063The expression "control sequences" is refers to DNA sequences necessary for the expression of a coding sequence operatively linked in an organism private host The control sequences that are appropriate for prokaryotes, for example, include a promoter, optionally an operating sequence, and a ribosome binding site. It's known that eukaryotic cells use promoters, signals from polyadenylation, and enhancers.

p0064A nucleic acid is "bound so operational "when placed in a functional relationship with another nucleic acid sequence. For example, the DNA for a Presence or secretion leader is operatively linked to a DNA for a polypeptide if it is expressed as a preprotein that participates in the secretion of the polypeptide; a promoter or enhancer is operatively linked to a coding sequence if influences sequence transcription; or a binding site to ribosome is operatively linked to a coding sequence if placed so that it facilitates translation. Usually, "operably linked" means that the DNA sequences that join are contiguous and, in the case of a secretory leader, contiguous in the reading phase. However, enhancers do not They have to be contiguous. The union is achieved by linking in suitable restriction sites. If such sites do not exist, it use synthetic oligonucleotide adapters or linkers of according to conventional practice.

p0065As used in this document, the expressions "cell", "cell line", and "cell culture" are used interchangeably and all these denominations include progeny. Therefore, the words "transformants" and "cells transformed "include the primary subject cell and cultures obtained from it without considering the amount of transfers. It is also understood that the entire progeny may not be exactly identical in DNA content, due to deliberate mutations or involuntary It includes the mutant progeny that has the same biological function or activity that is investigated in the cell transformed into a principle. When different are intended denominations, it will be clear from the context.

II.

Models for carrying out the invention

p0066Later examples describe the production of humanized anti-VEGF antibodies and variants with desirable properties from a therapeutic perspective that include: (a) strong binding affinity for the VEGF antigen; (b) ability to inhibit VEGF induced proliferation of endothelial cells <i>in vitro</i>; and (c) ability to inhibit VEGF induced angiogenesis <i>in vivo</i>.

p0067The affinities of the antibodies can determined as described in the subsequent examples. The Humanized antibodies or preferred variants are those that bind VEGF human with a K_ {d} value of no more than about 1 x 10-7 M; preferably not more than about 1 x 10-8 M; and more preferably no more than about 5 x 10-9 M.

p0068Apart from antibodies with strong affinity of human VEGF binding, it is also desirable to select antibodies humanized or variants that have other beneficial properties from a therapeutic perspective. For example, the antibody can be one that inhibits endothelial cell growth in response to VEGF In one embodiment, the antibody may be able to inhibit Bovine capillary endothelial cell proliferation in response to an almost maximum efficiency concentration of VEGF (3 ng / ml). Preferably, the antibody has a dose value 50 (ED50) effective of no more than about 5 nM, preferably no more than about 1 nM, and much more preferably no more than approximately 0.5 nM, to inhibit proliferation induced by VEGF endothelial cells in this "growth assay endothelial cell ", that is, at these concentrations the antibody is able to inhibit endothelial cell growth VEGF induced <i>in vitro</i> at 50% An "essay of "endothelial cell growth" preferred involves cultivating capillary endothelial cells obtained from adrenal cortex in presence of Eagle medium modified by Dulbecco low glucose (DEMEM) (GIBCO) supplemented with 10% calf serum, glutamine 2 mM, and antibiotics (growth medium), essentially as described in Example 1 below. These endothelial cells are seeded at a density of 6 x 10 3 cells per well, in 6-well plates in growth medium. Antibody parental anti-VEGF (control) antibody Humanized anti-VEGF or variant is added after at concentrations that vary between 1 and 5000 ng / ml. After 2-3 hours, purified VEGF is added to a final concentration of 3 ng / ml. For specificity control, each antibody can be added to endothelial cells to the 5000 ng / ml concentration, alone or in the presence of 2 ng / ml of bFGF. After five or six days, the cells dissociate by Trypsin exposure and are counted in a Coulter counter (Coulter Electronics, Hialeah, FL). The data can be analyzed by a Four parameter curve adjustment program (KaleidaGraph).

p0069Anti-VEGF antibody humanized or preferred variant can also be one that has a tumor suppression activity <i>in vivo</i>. For example, him antibody can suppress the growth of A673 cells from human rhabdomyosarcoma or cells MDA-MB-435 breast carcinoma in nude mice For tumor studies<i>in vivo</i>, he cultured human rhabdomyosarcoma A673 cells (ATCC; CRL1598) or MDA-MB-435 cells (available by ATCC) in DMEM / F12 supplemented with 10% fetal bovine serum, 2 mM glutamine and antibiotics as described in Example 1 later. BALB / c nude mice are injected subcutaneously 6-10 week old female with 2 x 10 6 tumor cells in the dorsal area in a volume of 200 µl. The animals are then treated with the humanized antibody or variant and a control antibody without activity in this assay. He Humanized anti-VEGF MAb or variant is administered to a dose of 0.5 and / or 5 mg / kg. Each MAb is administered twice per week intraperitoneally in a volume of 100 µl, starting 24 hours after cell inoculation Tumor Tumor size is determined at weekly intervals. Four weeks after inoculation of tumor cells, the Animals undergo euthanasia and tumors are removed and weighed. Statistical analysis can be performed by ANOVA. Preferably, the antibody in this "tumor assay <i>in vivo</i>"inhibits about 50-100%, preferably about 70-100% and more preferably approximately 80-100% of the growth of Human A673 tumor cells at a dose of 5 mg / kg.

p0070In the preferred embodiment, the antibody humanized or variant fails to elicit an immunogenic response on the administration of a therapeutically effective amount of antibody to a human patient. If a response is provoked immunogenic, the response will preferably be such that the antibody still provide a therapeutic benefit to the patient treated with it.

p0071The humanized antibody or variant is also preferably one that is capable of inhibiting induced angiogenesis by VEGF in a human, for example, to inhibit growth human tumor and / or inhibit intraocular angiogenesis in disorders retinals

p0072Preferred antibodies bind to the "epitope A.4.6.1 "as defined in this document. To search antibodies that bind to the epitope of a human VEGF bound by a antibody of interest (for example, those that block the binding of antibody A.4.6.1 to human VEGF), an assay of routine cross blocking such as described in <i>Antibodies, A Laboratory Manual</i>, Cold Spring Harbor Laboratory, Ed Harlow and David Lane (1988). As an alternative, a mapping of epitopes, for example, as described in Champe <i>et al., J. Biol. Chem</i>, 270: 1388-1394 (1995), for determine if the antibody binds to an epitope of interest.

p0073The antibodies of the preferred embodiment in this document have a heavy chain variable domain that It comprises an amino acid sequence represented by the formula: FR1-CDRH1-FR2-CDRH2-FR3-CDRH3-FR4, where "FR1-4" represent the four regions flanking and "CDRH1-3" represent the three hypervariable regions of a heavy chain variable domain of an anti-VEGF antibody. FR1-4 derive from a "consensus sequence" (ie amino acids most common of a class, subclass or subgroup of heavy chains or light human immunoglobulins) as in the examples later. Many sequences of flanking regions of human antibodies are collected in Kabat <i>et al., supra</i>, by example. The FR of the heavy chain variable domain is provides by a consensus sequence of a subgroup of human immunoglobulins compiled by Kabat <i>et al., supra</i>. The subgroup of human immunoglobulins is subgroup III of heavy human chains (for example, as in SEQ ID No. 11).

p0074The FR sequence of the variable domain of the chain heavy human has substitutions in it, for example, where the rest of human FR is replaced by a non-human rest corresponding (by "corresponding non-human rest" is it means the rest not human with the same positional numbering of Kabat than the rest of human interest when the human and non-human sequences), but replacement with the rest No human is not necessary. For example, a replacement of a FR rest other than the corresponding non-human rest can selected by phage display (see Example 2 later). The FR residues of the variable domain of the chain heavy that may contain substitutions include any one or more of the remains of FR with number: 37H, 49H, 67H, 69H, 71H, 73H, 75H, 76H, 78H, 94H (numbering of Kabat remains used here). Preferably at least two, or at least three, or At least four of these remains. A combination particularly Preferred FR substitutions is: 49H, 69H, 71H, 73H, 76H, 78H, and 94H.

p0075With respect to the hypervariable regions of the heavy chain, these have the following sequences of amino acids:

CDRH1

p0076GYX_ {1} X_ {2} X_ {3} X_ {4} YGX_ {5} N (SEQ ID No. 117), where X 1 is D, T or E, but preferably it is D or T; X 2 is F, W, or Y, but preferably it is F; X_ {3} is T, Q, G or S, but preferably it is T; X4 is H or N; and X_ {5} it is M or I, but preferably it is M.

CDRH2

p0077WINTX_ {1} TGEPTYAADFKR (SEQ ID No. 118), where X_ {1} is Y or W, but preferably it is Y.

CDRH3

p0078YPX_ {1} YX_ {2} X_ {3} X_ {4} X_ {5} HWYFDV (SEQ ID No. 119), where X 1 is H or Y; X_ {2} is Y, R, K, I, T, E, or W, but preferably it is Y; X_ {3} is G, N, A, D, Q, E, T, K, or S, but preferably is G; X_ {4} is S, T, K, Q, N, R, A, E, or G, but preferably it is S or T; and X_ {5} is S or G, but preferably it is S.

p0079The variable domain of the heavy chain optionally includes what has been called "CDR7" in this document in FR3 (that is, it is part of it) (see Figures 9B and 10B), where CDR7 can have the following sequence of amino acids:

CDR7

p0080X_ {1} SX_ {2} DX_ {3} X_ {4} X_ {5} X_ {6} TX_ {7} (SEC.ID. No. 120), where X_ {1} is F, I, V, L, or A, but preferably it is F; X 2 is A, L, V, or I, but preferably is L; X 3 is T, V or K, but preferably it is T; X_ {4} is S or W, but preferably it is S; X_ {5} is S or K, but preferably it is K; X 6 is N, or S, but preferably it is S; and X 7 is V, A, L or I, but preferably it is A.

p0081The antibodies of the preferred embodiment in this document have a variable domain of the light chain that It comprises an amino acid sequence represented by the formula: FR1-CDRL1-FR2-CDRL2-FR3-CDRL3-FR4, where "FR1-4" represent the four regions flanking and "CDRL1-3" represent the three hypervariable regions of a light chain variable domain of an anti-VEGF antibody. FR1-4 derive from a "consensus sequence" (ie amino acids most common of a class, subclass or subgroup of heavy chains or light human immunoglobulins) as in the examples later. The FR of the light chain variable domain is provides by a consensus sequence of a subgroup of human immunoglobulins compiled by Kabat <i>et al., supra</i>. The subgroup of human immunoglobulins is subgroup I of chains light kappa (for example, as in SEC.ID. No. 12).

p0082The FR sequence of the variable domain of the light chain has substitutions in it, for example, where the rest of human FR is replaced by a mouse rest corresponding, but the replacement with the non-human rest does not it is necessary. For example, a replacement of a different remainder of the corresponding non-human rest can be selected by the phage display (see Example 2 below). Preferably only 46L is replaced. In another embodiment it replace both 4L and 46L (enumeration of Kabat remains used here).

p0083With respect to the CDRs, these have the following amino acid sequences:

CDRL1

p0084X_ {1} AX_ {2} X_ {3} X_ {4} X_ {5} SNYLN (SEQ ID No. 121), where X 1 is R or S, but preferably is S; X 2 is S or N, but preferably it is S; X_ {3} is Q or E, but preferably it is Q; X 4 is Q or D, but preferably it is D; and X5 is I or L, but preferably it is I.

CDRL2

p0085FTSSLHS (SEC.ID. No. 122).

CDRL3

p0086QQYSX_ {1} X_ {2} PWT (SEC.ID.No. 123), where X1 is T, A or N, but preferably it is T; and X_ {2} is V or T, but preferably it is V.

p0087Anti-VEGF antibodies preferred humanized are those that have the sequences of the variable domains of the heavy and / or light chain of F (ab) -12 in Example 1 and variants of same as forms of matured affinity including variants Y0317, Y0313-1 and Y0238-3 in Example 3, Y0317 being the preferred variant. Methods to generate humanized anti-VEGF antibodies of interest in this document are elaborated in more detail to continuation.

A. Preparation of antibody

p0088The methods to humanize antibodies non-human anti-VEGF and generate variants of anti-VEGF antibodies are described in the examples later. To humanize an anti-VEGF antibody, The non-human antibody starting material is prepared. When a variant must be generated, the parental antibody is prepared. The exemplary techniques to generate said starting material of non-human antibodies and parental antibodies will be described on The following sections.

(i) Antigen preparation

p0089The VEGF antigen to be used for the production of antibodies can be, for example, intact VEGF or a fragment of VEGF (for example, a fragment of VEGF comprising the "epitope A.4.6.1 "). Other forms of VEGF useful for generating Antibodies will be apparent to those skilled in the art. He VEGF antigen used to generate the antibody is preferably Human VEGF, for example, as described in Leung <i>et al., Science</i> 246: 1306 (1989), and Houck <i>et al., Mol. Endocrin</i>. 5:1806 (1991).

(ii) Polyclonal antibodies

p0090Polyclonal antibodies are raised preferably in animals by multiple subcutaneous injections (sc) or intraperitoneal (ip) of the relevant antigen and a adjuvant It may be useful to conjugate the relevant antigen to a protein that is immunogenic in the species to be immunized, for example, California limpet hemocyanin, serum albumin, bovine thyroglobulin, or soybean trypsin inhibitor using a bifunctional or derivatizing agent, for example, maleimidobenzoyl sulfosuccinimide ester (conjugation through of cysteine ​​residues), N-hydroxysuccinimide (a through lysine residues), glutaraldehyde, succinic anhydride, SOCl_ {2}, or R 1 N = C = NR, where R and R 1 are alkyl groups different.

p0091Animals are immunized against the antigen, immunogenic conjugates, or derivatives combining, for example, 100 \ mug or 5 \ mug of the protein or conjugate (for rabbits or mice, respectively) with 3 volumes of complete adjuvant of Freund and inject the solution intradermally in multiple sites. One month later the animals are stimulated with 1/5 to 1/10 of the original amount of the peptide or conjugate in complete adjuvant of Freund by subcutaneous injection at multiple sites. From seven to 14 days later blood is drawn from the animals and the serum is assay for antibody titer. The animals are stimulated to the parking lot of the title. Preferably, the animal is stimulates with the conjugate of the same antigen, but conjugated to a different protein and / or through a crosslinking reagent different. Conjugates can also be made in culture of Recombinant cells as fusion proteins. In addition, they are used appropriately aggregating agents such as alum for Boost the immune response.

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(iii) Monoclonal antibodies

p0092Monoclonal antibodies can be made using the hybridoma method first described by Kohler<i>et al., Nature</i>, 256: 495 (1975), or can be done by methods of recombinant DNA (US Patent No. 4,816,567).

p0093In the hybridoma method, a mouse is immunized or another appropriate host animal, such as a hamster or macaque monkey, as described above to cause lymphocytes that produce or be able to produce antibodies that will bind specifically to the protein used for immunization. As alternatively, lymphocytes can be immunized <i>in vitro</i>. The lymphocytes are then fused to myeloma cells using a appropriate melting agent, such as polyethylene glycol, to form a hybridoma cell (Goding, <i>Monoclonal Antibodies: Principles and Practice</i>, P. 59-103 (Academy Press, 1986)).

p0094Hybridoma cells prepared in this way they are sown and grown in an appropriate culture medium that preferably contains one or more substances that inhibit the growth or survival of non-fused myeloma cells parental For example, if the parental myeloma cells they lack the enzyme hypoxanthine guanine phosphoribosyltransferase (HGPRT or HPRT), the culture medium for hybridomas typically will include hypoxanthine, aminopterin, and thymidine (HAT medium), substances that prevent the growth of deficient cells in HGPRT.

p0095Preferred myeloma cells are the one that merge effectively, support stable high level production of antibody by antibody producing cells selected, and are sensitive to a medium such as the HAT medium. Between these, the preferred myeloma cell lines are lines of murine myeloma, such as those obtained from mouse tumors MOP-21 and MC-11 available by the Salk Institute Cell Distribution Center, San Diego, California USA, and cells SP-2 or X63-Ag8-653 available by the American Type Culture Collection, Rockville, Maryland USA Myeloma cell lines have also been described human and mouse-human heteromyeloma for production of human monoclonal antibodies (Kozbor, <i>J. Immunol</i>., 133: 3001 (1984); Brodeur<i>et al., Monoclonal Antibody Production Techniques and Applications</i>, P. 51-63 (Marcel Dekker, Inc., New York, 1987)).

p0096The culture medium in which the hybridoma cells are tested for antibody production monoclonal directed against the antigen. Preferably, the binding specificity of the monoclonal antibodies produced Hybridoma cells are determined by immunoprecipitation or by a binding test <i>in vitro</i>, such as radioimmunoassay (RIA) or bound enzyme immunosorbent assay (ELISA).

p0097The binding affinity of the monoclonal antibody it can, for example, be determined by the Scatchard analysis of Munson <i>et al., Anal. Biochem</i>., 107:220 (1980).

p0098After identifying that the cells of hybridoma produce antibodies of specificity, affinity, and / or desired activity, clones can be subcloned by procedures of limiting dilution and growth by conventional methods (Goding, <i>Monoclonal Antibodies: Principles and Practice</i>, P. 59-103 (Academia Press, 1986)). The means of appropriate cultures for this purpose include, for example, medium D-MEM or RPMI-1640. In addition, the hybridoma cells can grow <i>in vivo</i> as tumors ascites in an animal.

p0099Monoclonal antibodies secreted by subclones properly separated from the culture medium, fluid ascites, or serum by purification procedures of conventional immunoglobulins such as, for example, protein A-Sepharose, hydroxylapatite chromatography, gel electrophoresis, dialysis, or chromatography of affinity.

p0100DNA encoding monoclonal antibodies it is easily isolated and sequenced using procedures conventional (for example, using oligonucleotide probes that are able to specifically bind to genes that encode the heavy and light chains of monoclonal antibodies). The hybridoma cells serve as the preferred source of said DNA. A Once isolated, the DNA can be placed in expression vectors, which They are then transferred into host cells such as<i>E. coli</i>, ape COS cells, hamster ovary cells Chinese (CHO), or myeloma cells that do not produce protein immunoglobulin otherwise, to obtain the synthesis of monoclonal antibodies in recombinant host cells. Recombinant antibody production will be described with more detail later.

(iv) Humanization and variants of the sequence of amino acids

p0101Examples 1-2 below describe procedures for the humanization of an antibody anti-VEGF In certain embodiments, it may be desirable to generate variants of the amino acid sequence of these humanized antibodies, particularly when they improve the binding affinity or other biological properties of the antibody humanized Example 3 describes methodologies to generate amino acid sequence variants of an antibody anti-VEGF with enhanced affinity in relation to the parental antibody

p0102The amino acid sequence variants of the anti-VEGF antibody are prepared by introducing appropriate nucleotide changes in antibody DNA anti-VEGF, or by peptide synthesis. These variants include, for example, deletions, and / or insertions and / or residue substitutions in the amino acid sequences of the anti-VEGF antibodies of the examples in this document. Any combination of deletion, insertion, and replacement is done to reach the final construction, on condition that the final construction has the desired characteristics. Amino acid changes can also alter processes post-translational antibody Humanized anti-VEGF or variant, such as changing the number or position of glycosylation sites.

p0103A useful method for the identification of certain residues or regions of the anti-VEGF antibody that are Preferred positions for mutagenesis is called "mutagenesis by Alanine scan ", as described by Cunningham and Wells<i>Science</i>, 244: 1081-1085 (1989). Here, it identify a remainder or group of target residues (e.g., residues loaded such as arg, asp, his, lys, and glu) and replaced by a negatively charged or neutral amino acid (more preferably alanine or polyalanin) to achieve the interaction of amino acids with the VEGF antigen. The amino acid positions that demonstrate functional sensitivity to substitutions are then perfected introducing additional or different variants to, or for, the replacement sites Therefore, although the site to introduce a variation of the amino acid sequence is predetermined, the nature of the mutation <i>per se</i> need not be default For example, to analyze the behavior of a mutation at a given site, mutagenesis is directed by exploration of wing or random to the codon or target region and variants of expressed anti-VEGF antibodies are screened for desired activity Alanine scan mutagenesis is described in Example 3.

p0104Insertions in the amino acid sequence include amino and / or carboxy-terminal fusions that they vary in length from a residue to polypeptides containing one hundred or more remains, as well as insertions within the sequence of a single rest or multiple amino acid residues. The examples of terminal inserts include an antibody anti-VEGF with a methionyl moiety N-terminal or antibody fused to a epitope signaling Other variants of molecule insertion Anti-VEGF antibody include fusion to N or C-terminal of anti-VEGF antibody of an enzyme or a polypeptide that increases serum half-life of the antibody (see below).

p0105Another type of variant is a variant of amino acid substitution. These variants have at least one remainder of amino acid removed in the antibody molecule anti-VEGF and a different remainder inserted in your place. The sites of greatest interest for replacement mutagenesis they include hypervariable regions, although they are also contemplated FR alterations. Conservative substitutions are shown in Table 1 with the title of "preferred substitutions". Yes such substitutions result in a change in activity biological, then more substantial changes can be introduced, called "exemplary substitutions" in Table 1, or as further described below in reference to classes of amino acids, and the products are explored.

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TABLE 1

<figref>1</figref>

p0106Substantial modifications in the Biological properties of the antibody are achieved by selecting substitutions that differ significantly in the effect on the maintenance of (a) the structure of the polypeptide central axis in the replacement area, for example, as a sheet conformation or helical, (b) the charge or hydrophobicity of the molecule in the target site, or (c) the volume of the side chain. The remains of Natural origin are divided into groups based on the properties Common side chain:

p0107(1) hydrophobic: norleucine, met, ala, val, leu, ile;

p0108(2) neutral hydrophilic: cys, ser, thr;

p0109(3) acids: asp, glu;

p0110(4) basic: asn, gln, his, lys, arg;

p0111(5) remains that influence the orientation of the chain: gly, pro; and

p0112(6) aromatic: trp, tyr, phe.

p0113Non-conservative substitutions will imply Exchange a member of these classes for another class.

p0114You can also replace any remaining cysteine ​​not involved in the maintenance of the conformation appropriate humanized anti-VEGF antibody or variant, usually by serine, to improve stability Oxidative molecule and avoid aberrant crosslinks. To conversely, a link or bonds of cysteine ​​to improve its stability (particularly when the antibody is an antibody fragment such as a fragment Fv).

p0115A particularly preferred type of variant of substitution involves replacing one or more remnants of the region hypervariable of a parental antibody (for example, an antibody humanized or human). Generally, the variant or variants results selected for further development will have improved biological properties in relation to the antibody parental from which they were generated. A suitable way to generate said substitution variants is affinity maturation using phage display (see Example 3 in this document). Briefly, several sites of the hypervariable region are mutated (by example, 6-7 sites) to generate all possible amino acid substitutions at each site. Variants of antibodies generated in this way are shown in a way monovalent filamentous phage particles as fusions to Gene III product of M13 packaged in each particle. The variants presented in phages are then explored for their activity biological (e.g. binding affinity) as described in this document. To identify sites in the hypervariable region candidates for modification, mutagenesis can be performed by Alanine scan (see Example 3) to remnants of the region hypervariable identified that contribute significantly to the antigen binding. Alternatively, or in addition, it can be beneficial. analyze a crystal structure of the complex antigen-antibody to identify points of contact between antibody and human VEGF. These remains of contact and adjacent remains are candidates for replacement of according to the techniques elaborated in this document. Once said variants are generated, the variants panel is subjected to scan as described herein and antibodies with superior properties in one or more relevant assays can be selected for further development.

p0116Another type of amino acid variant of antibody alters the original glycosylation pattern of antibody. By altering means deleting one or more remains of carbohydrates found in the antibody, and / or add one or more glycosylation sites that are not present in the antibody.

p0117Glycosylation of antibodies typically takes place with links N or links O. Link N refers to the binding of the rest of carbohydrate to the side chain of a remainder of asparagine The tripeptide sequences asparagine-X-serine and asparagine-X-threonine, where X is any amino acid except proline, are the sequences of recognition for the enzymatic binding of the rest of carbohydrate to the asparagine side chain. Therefore, the presence of any of these tripeptide sequences in a polypeptide creates a potential glycosylation site. Glycosylation by O-linkage refers to the union of one of the sugars N-acetylgalactosamine, galactose, or xylose at a hydroxyamino acid, more commonly serine or threonine, although 5-hydroxyproline can also be used or 5-hydroxylysine.

p0118The addition of glycosylation sites to antibody is achieved in a practical way by altering the sequence of amino acids so that it contains one or more of the sequences tripeptide described above (for glycosylation sites by link N). The alteration can also be made by adding, or substitution by one or more serine or threonine residues to the original antibody sequence (for glycosylation sites by link O).

p0119The nucleic acid molecules that encode variants of the amino acid sequence of the antibody Anti-VEGF are prepared by a variety of methods known in the art. These methods include, but without limitation, isolation from a natural source (in the case of naturally occurring amino acid sequence variants) or oligonucleotide-mediated (or directed) mutagenesis preparation site), PCR mutagenesis, or cassette mutagenesis of a previously prepared variant or a non-variant version of the anti-VEGF antibody.

(v) Human antibodies

p0120As an alternative to humanization, they can Generate human antibodies. For example, it is now possible produce transgenic animals (eg, mice) that are able, on immunization, to produce a complete repertoire of human antibodies in the absence of endogenous production of immunoglobulins For example, it has been described that the deletion homozygous for the gene of the binding region of the heavy chain of the antibody (JH) in chimeric or mutant mice in the line germinal results in complete production inhibition endogenous antibody. Transfer the gene series from human germline immunoglobulins in said mice germline mutants will result in the production of human antibodies on antigen stimulation. See for example, Jakobovits <i>et al., Proc. Natl. Acad. Sci. USA</i>, 90: 2551 (1993); Jakobovits<i>et al., Nature</i>, 362: 255-258 (1993); Bruggermann<i>et al., Year in Immuno</i>., 7:33 (1993); and U.S. Patent No. 5,591,669, No. 5,589,369 and No. 5,545,807. Antibodies can also be obtained. of libraries presented in phages (Hoogenboom <i>et al., J. Mol. Biol</i>., 227: 381 (1991); Marks<i>et al., J. Mol. Biol</i>., 222: 581-597 (1991); and U.S. Patent No. 5,565,332 and No. 5,573,905). As discussed above, the human antibodies can also be generated by B cells activated <i>in vitro</i> (see US Patent No. 5,567,610 and No. 5,229,275).

(vi) Antibody fragments

p0121In certain embodiments, the antibody Humanized anti-VEGF or variant is a fragment of antibody. Various production techniques have been developed of antibody fragments. Traditionally, these fragments are obtained by proteolytic digestion of intact antibodies (see, for example, Morimoto <i>et al., Journal of Biochemical and Biophysical Methods</i> 24: 107-117 (1992) and Brennan<i>et al., Science</i> 229: 81 (1985)). However, these fragments can now be produced directly by cells recombinant hosts. For example, the fragments Fab'-SH can recover directly from <i>AND. coli</i> and chemically couple to form fragments F (ab ') 2 (Carter <i>et al. Bio / Technology</i>10: 163-167 (1992)). In another embodiment, the F (ab ') 2 is formed using the GCN4 leucine zipper to promote the assembly of the F (ab ') 2 molecule. From according to another approach, the Fv, Fab or fragments F (ab ') 2 can be isolated directly from a culture of recombinant host cells. Other techniques for Production of antibody fragments will be apparent to those specialist doctors.

(vii) Multispecific antibodies

p0122In some embodiments, it may be desirable. generate multispecific anti-VEGF antibodies (by example, bispecific) humanized or variants that have binding specificities for at least two different epitopes. The exemplary bispecific antibodies can bind two epitopes different from VEGF protein. As an alternative, it can be combined an anti-VEGF arm with an arm that joins one signaling molecule in a leukocyte such as a molecule T cell receptor (for example, CD2 or CD3), or Fc receptors for IgG (FcγR), such as FcγRI (CD64), FcγRII (CD32) and FcγRIII (CD16) to concentrate the mechanisms of cell defense towards the cell that expresses VEGF. Antibodies Bispecific can also be used to locate agents cytotoxic for cells expressing VEGF. These antibodies they have a VEGF binding arm and an arm that binds to the agent cytotoxic (for example, saporin, anti-interferon-?, alkaloid of vinca, ricin A chain, methotrexate or isotope hapten radioactive). Bispecific antibodies can be prepared as full length antibodies or as antibody fragments (for example, bispecific antibodies F (ab ') 2).

p0123According to another approach to do bispecific antibodies, the contact surface can be designed between a pair of antibody molecules to maximize the percentage of heterodimers that recover from cell culture recombinant The preferred contact surface comprises the minus a part of domain C_ {H} 3 of a constant domain of a antibody. In this method, one or more chains are replaced small amino acid sides of the contact surface of the first antibody molecule with larger side chains (for example, tyrosine or tryptophan). "Cavities" of identical or similar size to the chain or large side chains in the contact surface of the second antibody molecule replacing large side chains of amino acids with others small (for example, alanine or threonine). This provides a mechanism to increase the production of the heterodimer over others unwanted end products such as homodimers. See the WO96 / 27011 published September 6, 1996.

p0124Bispecific antibodies include cross-linked or "heteroconjugate" antibodies. For example, one of the antibodies in the heteroconjugate can bind avidin, the other to biotin. Heteroconjugate antibodies can be made using any suitable crosslinking method. The agents of Appropriate crosslinking are well known in the art, and are described in US Patent No. 4,676,980, together with several crosslinking techniques.

p0125The techniques to generate antibodies bispecific from antibody fragments have also been described in the bibliography. For example, the antibodies Bispecific can be prepared using chemical bonds. Brennan<i>et al., Science</i> 229: 81 (1985) describes a procedure where intact antibodies are proteolytically cleaved to generate F (ab ') 2 fragments. These fragments are reduced by presence of dithiol complexing sodium arsenite agent for stabilize the vicinal dithiols and prevent the formation of intermolecular disulfide The Fab 'fragments generated afterwards are converted into thionitrobenzoate derivatives (TNB). One of the Fab'-TNB derivatives are then converted into the Fab'-thiol by reduction with mercaptoethylamine and mixing with an equimolecular amount of the other derivative Fab'-TNB to form the bispecific antibody. The bispecific antibodies produced can be used as agents for the selective immobilization of enzymes. In one embodiment additional more, the Fab'-SH fragments recovered directly from <i>E. coli</i> can chemically join <i>in vitro</i> to form bispecific antibodies. Shalaby<i>et al., J. Exp. Med</i>. 175:217-225 (1992).

p0126Various techniques have also been described for make and isolate bispecific antibody fragments directly of recombinant cell culture. For example, they have occurred bispecific antibodies using leucine zippers. Kostelny<i>et al., J. Immunol</i>. 148 (5): 1547-1553 (1992). Leucine zipper peptides of Fos proteins and Jun joined the Fab 'portions of two different antibodies by gene fusion. Antibody homodimers were reduced to hinge region to form monomers and then reoxidized to form antibody heterodimers. This method can also be used for the production of antibody homodimers. The "bivalent fragments" technology described by Hollinger<i>et al., Proc. Natl. Acad. Sci. USA</i>90: 6444-6448 (1993) has provided a mechanism alternative to make bispecific antibody fragments. The fragments comprise a heavy chain variable domain (V_ {H}) connected to a light chain variable domain (V_ {L}) by a linker that is too short to allow pairing between the two domains of the same chain. By consequently, the domains V_ {H} and V_ {L} of a fragment are force to match domains V_ {L} and V_ {H} complementary to another fragment, thereby forming two sites antigen binding. Another strategy has also been reported. to make bispecific antibody fragments by the use of single chain Fv dimers (sFv). See gruber<i>et al., J. Immunol</i>. 152: 5368 (1994). Alternatively, the antibody bispecific can be a "linear antibody" produced as It is described in Zapata <i>et al. Eng Protein</i>. 8(10):1057-1062 (1995).

p0127Antibodies with more than two are contemplated valences For example, antibodies can be prepared Triespecific. Tutt<i>et al., J. Immunol</i>. 147:60 (1991).

(viii) Other modifications

p0128Other antibody modifications are contemplated. Humanized anti-VEGF or variant. For example, you can it is desirable to modify the antibody of the invention with respect to the effector function, so that the effectiveness of the antibody in the treatment of cancer, for example. For example, a cysteine ​​residue or moieties may be introduced into the Fc region, thereby allowing the formation of a disulfide bond Intercatenary in this region. The homodimeric antibody generated in this way it can have an improved internalization capacity and / or complement-mediated cell elimination and cytotoxicity antibody-dependent cell (ADCC). See Caron<i>et al., J. Exp Med</i>. 176: 1191-1195 (1992) and Shopes, B.<i>J. Immunol</i>. 148: 2918-2922 (1992). They can also prepare homodimeric antibodies with antitumor activity enhanced using heterofunctional crosslinkers as described in Wolff <i>et al., Cancer Research</i> 53: 2560-2565 (1993). Alternatively, an antibody can be designed that has dual Fc regions and can thereby have lysis capabilities by complement and enhanced ADCC. See Stevenson <i>et al., Anti-Cancer Drug Design</i> 3:219-230 (1989).

p0129The invention also concerns immunoconjugates. which comprise the antibody described herein conjugated to a cytotoxic agent such as a chemotherapeutic agent, toxin (for example, an enzymatically active toxin of origin bacterial, fungal, plant or animal, or fragments thereof), or a radioactive isotope (i.e. a radioconjugate).

p0130Chemotherapeutic agents useful in generation of said immunoconjugates have been described above. Enzymatically active toxins and fragments thereof that can be used include diphtheria toxin A chain, non-binding fragments of diphtheria toxin, chain A of the exotoxin (from <i>Pseudomonas aeruginosa</i>), ricin A chain, string A, string A, modecin, alpha-sarcin proteins <i>Aleurites fordii</i>, diantin proteins, proteins <i>American Phytolacca</i>(PAPI, PAPII, and PAP-S), momordica inhibitor charantia, curcine, crotine, saponaria officinalis inhibitor, gelonin, mitogelin, restrictocin, phenycline, anomycin and trichothecenes A variety of radionuclides is available for anti-VEGF antibody production radioconjugates Examples include 212 Bi, 131 I, 131 In, 90 Y and 186 Re.

p0131The antibody and agent conjugates Cytotoxic are made using a variety of binding agents of bifunctional proteins such as N-succinimidyl-3- (2-pyridyldithiol) propionate (SPDP), iminothiolane (IT), bifunctional derivatives of imidoesters (such as dimethyl adipimidate HCL), active esters (such as disuccinimidyl suberate), aldehydes (such as glutaraldehyde), bis-azido compounds (such as bis- (p-azidobenzoyl) hexanediamine), derivatives bis-diazonium (such as bis (p-diazonium benzoyl) ethylenediamine), diisocyanates (such as 2,6-diisocyanate toluene), and active bis-fluorine compounds (such as 1,5-difluoro-2,4-dinitrobenzene). For example, a ricin immunotoxin can be prepared as described in Vitetta <i>et al., Science</i> 238: 1098 (1987). He acid 1-Isothiocyanatobenzyl-3-methyldiethylenetriaminopentaacetic acid (MX-DTPA) carbon-labeled 14 is an agent exemplary chelator for conjugation of radionuclides to antibody. See document WO94 / 11026.

p0132In another embodiment, the antibody can conjugate to a "receptor" (such as streptavidin) for use in tumor pre-marking where the conjugate antibody-receptor is administered to the patient, followed by withdrawal of unbound conjugate circulation using a removal agent and then administering a "ligand" (eg, avidin) that is conjugated to an agent cytotoxic (for example, a radionuclide).

p0133Anti-VEGF antibodies described in this document can also be formulated as immunoliposomes Liposomes containing the antibody will prepared by methods known in the art, such as described in Epstein <i>et al., Proc. Natl. Acad. Sci. USA</i>82: 3688 (1985); Hwang<i>et al. Proc Natl Acad. Sci. USA</i>77: 4030 (1980); and U.S. Patent No. 4,485,045 and No. 4,544,545. Liposomes with enhanced circulation time are described in U.S. Patent No. 5,013,556.

p0134Particularly useful liposomes can be generated by the reverse phase evaporation method with a composition lipid comprising phosphatidylcholine, cholesterol and PEG-derived phosphatidylethanolamine (PEG-PE). The Liposomes are extruded through filters of defined pore size to produce liposomes with the desired diameter. Fragments Fab 'of the antibody of the present invention can be conjugated to liposomes as described in Martin <i>et al., J. Biol. Chem</i>. 257: 286-288 (1982) by a reaction of disulfide exchange. Optionally the liposome contains a chemotherapeutic agent (such as Doxorubicin). See Gabizon<i>et al., J. National Cancer Inst</i>. 81(19):1484(1989).

p0135The antibody of the present invention also can be used in ADEPT by conjugating the antibody to an enzyme of prodrug activation that converts a prodrug (for example, a peptide chemotherapeutic agent, see document WO81 / 01145) to an active anti-cancer drug. See for example, WO 88/07378 and US Patent No. 4,975,278.

p0136The enzyme component of the useful immunoconjugate for ADEPT includes any enzyme capable of functioning in a prodrug so that it becomes its most cytotoxic form active

p0137Enzymes that are useful in the method of this invention include, but is not limited to, alkaline phosphatase, useful to convert phosphate-containing prodrugs into free drugs; arylsulfatase, useful for converting prodrugs containing sulfate in free drugs; cytosine deaminase, useful for convert non-toxic 5-fluorocytosine into the drug anti-cancer, 5-fluoroacyl; proteases, such as serratia protease, thermolysin, subtilisin, carboxypeptidases and cathepsins (such as cathepsins B and L), which are useful for converting prodrugs that contain peptides in free drugs; D-alanylcarboxypeptidases, useful for converting prodrugs containing substituents D-amino acids; carbohydrate cleavage enzymes such as β-galactosidase and neuraminidase, useful for converting glycosylated prodrugs into free drugs; β-lactamase, useful for converting drugs derivatives with β-lactams in free drugs; and penicillin amidases, such as penicillin V amidase or Penicillin G amidase, useful for converting derivatized drugs in their amine nitrogens with phenoxyacetyl or phenylacetyl groups, respectively, in free drugs. As an alternative, they can be used antibodies with enzymatic activity, also known in the technique like "abzimas", to convert the prodrugs of the invention in free active drugs (see, for example, Massey,<i>Nature</i> 328: 457-458 (1987)). The conjugates antibody-abzyme can be prepared as described in this document to supply the abzyme to the population of tumor cells.

p0138The enzymes of this invention can bind covalently to anti-VEGF antibodies by techniques well known in the art such as the use of heterobifunctional crosslinking reagents analyzed previously. Alternatively, proteins of fusion comprising at least the antigen binding region of an antibody of the invention bound to at least a portion functionally active of an enzyme of the invention using techniques of recombinant DNA well known in the art (see, for example, Neuberger <i>et al., Nature</i> 312:604-608 (1984)).

p0139In certain embodiments of the invention, it may it is desirable to use an antibody fragment, instead of a intact antibody, to increase tumor penetration, by example. In this case, it may be desirable to modify the fragment of antibody to increase its serum half-life. This can achieved, for example, by incorporating a binding epitope to rescue receptor in the antibody fragment (for example, by mutation of the appropriate region in the antibody fragment or incorporating the epitope into a peptide signaler that is then fuses the antibody fragment at the end or in the middle, by example, by synthesis of DNA or peptides). See the document WO96 / 32478 published October 17, 1996.

p0140The rescue receptor binding epitope generally constitutes a region where any one or more remains amino acids of one or two loops of an Fc domain are transferred to an analogous position of the antibody fragment. Even more preferably, three or more remains of one or two are transferred domain loops Fc. Even more preferred, the epitope is taken from the CH2 domain of the Fc region (eg, of an IgG) and is transfers to the region CH1, CH3, or VH, or more than one of said antibody regions. Alternatively, the epitope is takes the CH2 domain from the Fc region and transfers to the region C L or V L region, or both, of the antibody fragment.

p0141In preferred embodiments, the binding epitope The rescue receiver comprises the sequence: PKNSSMISNTP (SEC.ID.No. 17), and optionally also comprises a selected sequence among the group consisting of HQSLGTQ (SEC.ID. No. 18), HQNLSDGK (SEQ ID No. 19), HQNISDGK (SEQ ID No. 20), or VISSHLGQ (SEQ ID No. 21), particularly when the antibody fragment is a Fab or F (ab ') 2 or the rescue receptor binding epitope is a polypeptide containing the sequence or sequences: HQNLSDGK (SEQ ID No. 19), HQNISDGK (SEQ ID No. 20), or VISSHLGQ (SEQ ID No. 21) and the sequences: PKNSSMISNTP (SEC.ID. No. 17).

p0142Covalent antibody modifications Humanized anti-VEGF or variant are also included within the scope of this invention as defined in the claims. They can be done by chemical synthesis or by excision Enzymatic or chemical antibody, if applicable. They are introduced other types of covalent modifications of the antibody in the molecule reacting labeled amino acid residues of the antibody with an organic derivatizing agent that is capable of react with selected side chains or residues N or C-terminals Covalent modifications Exemplary polypeptides are described in US Patent No. 5,534,615. A preferred type of covalent modification of the antibody comprises binding of the antibody to one of a variety of non-protein polymers, for example, polyethylene glycol, propylene glycol, or polyoxyalkylenes, as described in the U.S. Patent No. 4,640,835; No. 4,496,689; No. 4,301,144; No. 4,670,417; No. 4,791,192 or No. 4,179,337.

B. Vectors, host cells and methods recombinant

p0143The invention also provides an acid. asylum nucleic encoding the antibody Humanized or variant anti-VEGF, vectors and cells hosts that comprise nucleic acid, and techniques recombinants for antibody production.

p0144For recombinant antibody production, the nucleic acid encoding it can be isolated and inserted into a replication vector for additional cloning (amplification of DNA) or for expression. In another embodiment, the antibody can be produced by homologous recombination, for example, as described in US Patent No. 5,204,244. The DNA that encodes the monoclonal antibody is isolated and easily sequenced using conventional procedures (for example, using probes oligonucleotides that are capable of specifically binding to genes encoding the heavy and light chains of the antibody). Is it so Many vectors available. The components of the vector generally include, but are not limited to, one or more of the following: signal sequence, an origin of replication, one or more genes markers, an enhancer element, a promoter, and a sequence of termination of transcription, for example, as described in the U.S. Patent No. 5,534,615 issued July 9, 1996.

p0145The appropriate host cells for cloning and expression of DNA in vectors in this document, are prokaryotic cells, yeasts, or higher eukaryotes described above. The appropriate prokaryotes for this purpose include eubacteria, such as organisms Gram-negative or Gram-positive, by example, Enterobacteriaceae such as <i>Escherichia</i>, by example, <i>E. coli, Enterobacter, Erwinia, Klebsiella, Proteus, Salmonella</i>, for example, <i>Salmonella typhimurium, Serratia</i>, for example <i>Serratia marcescans</i>, and<i>Shigella</i>, as well as <i>Bacilli</i> such as <i>B. subtilis and B. licheniformis</i> (for example, <i>B. licheniformis</i> 41P described in document DD 266,710 published on April 12, 1989), <i>Pseudomonas</i> such as <i>P. aeruginosa</i>, and<i>Streptomyces</i>. A cloning host of <i>E. coli</i>preferred is <i>E. coli</i> 294 (ATCC 31,446), although appropriate other strains such as <i>E. coli</i> B, <i>E. coli</i> X1776 (ATCC 31,537), and <i>E. coli</i> W3110 (ATCC 27,325). These examples are Illustrative rather than limiting.

p0146In addition to prokaryotes, microbes eukaryotes such as fungi or filamentous yeasts are appropriate cloning or expression hosts for vectors that encode anti-VEGF antibody. <i>Saccharomyces cerevisiae</i>, or common bread yeast, is the most usually used among host microorganisms lower eukaryotes However, they are usually available several other genera, species, and strains, and are useful in this document, such as <i>Schizosaccharomyces pombe</i>; hosts <i>Kluyveromyces</i> such as, for example, <i>K. lactis, K. fragilis</i> (ATCC 12.424), <i>K. bulgaricus</i> (ATCC 16.045),<i>K. wickeramii</i> (ATCC 24.178), <i>K. waltii</i> (ATCC 56,500),<i>K. drosophilarum</i> (ATCC 36.906), <i>K. thermotolerans</i>, and<i>K. marxiamis; yarrowia</i> (EP 402,226); <i>Pichia pastoris</i>(EP 183,070); <i>Candida; Trichoderma reesia</i> (EP 244,234); <i>Neurospora crassa; Schwanniomyces</i> such as<i>Schwanniomyces occidentalis</i>; and filamentous fungi such for example, <i>Neurospora, Penicillium, Tolypocladium</i>, and hosts <i>Aspergillus</i> such as <i>A. nidulans</i> and<i>A. niger</i>.

p0147The appropriate host cells for the Glycosylated anti-VEGF antibody expression is obtained from multicellular organisms. Examples of cells from Invertebrates include plant and insect cells. They have identified numerous strains and variants of baculovirus and cells corresponding permissive insect hosts of hosts such as <i>Spodoptera frugiperda</i> (caterpillar), <i>Aedes aegypti</i> (mosquito), <i>Aedes albopictus</i> (mosquito),<i>Drosophila melanogaster</i> (fruit fly), and <i>Bombyx I died</i>. A variety of strains are available to the public viral for transfection, for example, the variant L-1 of <i>Autographa californica</i> NPV and strain Bm-5 of <i>Bombyx mori</i> NPV, and such viruses may be used herein as viruses according to the present invention, particularly for cell transfection of<i>Spodoptera frugiperda</i>. They can also be used as host plant cell cultures of cotton, corn, Potato, soy, petunia, tomato, and tobacco.

p0148However, interest has been greater in cells of vertebrates, and the propagation of vertebrate cells in cultivation (tissue cultures) has become a procedure of routine. Examples of mammalian host cell lines are the CV1 monkey kidney line transformed by SV40 (COS-7, ATCC CRL 1651); kidney line human embryonic (293 or 293 cells subcloned for the growth in suspension culture, Graham <i>et al., J. Gen Virol</i>. 36:59 (1977)); hamster breeding kidney cells (BHK, ATCC CCL 10); Chinese hamster ovary cells / -DHFR (CHO, Urlaub<i>et al., Proc Natl. Acad. Sci. USA</i> 77: 4216 (1980)); cells of mouse sertoli (TM4. Mather, <i>Biol. Reprod</i>. 23: 243-251 (1980)); monkey kidney cells (CV1 ATCC CCL 70); African green monkey kidney cells (VERO-76. ATCC CRL-1587); cells of human cervical carcinoma (HELA, ATCC CCL 2); kidney cells canine (MDCK, ATCC CCL 34); buffalo rat liver cells (BRL 3A, ATCC CRL 1442); human lung cells (W138, ATCC CCL 75); human liver cells (Hep G2, HB 8065); mouse breast tumor (MMT 060562, ATCC CCL51); TR1 cells (Mather<i>et al., Annals NY Acad. Sci</i>. 383: 44-68 (1982)); MRC 5 cells; FS4 cells; and a human hepatoma line (Hep G2).

p0149Host cells are transformed with the expression or cloning vectors described above for production of anti-VEGF antibodies and grown in conventional nutrient medium modified in the appropriate manner for induce promoters, select transformants, or expand genes that encode the desired sequences.

p0150The host cells used to produce the anti-VEGF antibody of this invention can Cultivate in a variety of media. The means available in the market such as Ham's F10 (Sigma), Minimum Essential Medium ((MEM), Sigma)), RPM1I-1640 (Sigma), and Middle Eagle Modified by Dulbecco ((DMEM), Sigma) are suitable for cultivation host cells. In addition, any of the means described in Ham <i>et al., Meth. Enz</i>. 58:44 (1979), Barnes<i>et al., Anal. Biochem</i>. 102: 255 (1980), U.S. Pat. No. 4,767,704; No. 4,657,866; No. 4,927,762; No. 4,560,655; o Nº 5,122,469; WO 90/03430; WO 87/00195; or US Pat. Re. No. 30,985 can be used as culture media for cells hosts. Any of these means can be supplemented if it is necessary with hormones and / or other growth factors (such as insulin, transferrin, or epidermal growth factor), salts (such as sodium chloride, calcium, magnesium and phosphate), buffers (such as HEPES), nucleotides (such as adenosine and thymidine), antibiotics (such as the drug GENTAMYCIN ™), trace elements (defined as inorganic compounds usually present at final concentrations in the micromolar range), and glucose or An equivalent energy source. Other ones may also be included. any necessary supplements at appropriate concentrations that would be known to those skilled in the art. The culture conditions, such as temperature, pH, and the like, are those previously used with host cells selected for expression, and will be apparent to the specialist in the technique

p0151When recombinant techniques are used, the antibody can be produced intracellularly, in space periplasmic, or secreted directly in the middle. If he antibody is produced intracellularly, as a first stage, particulate debris, host cells or lysed fragments, they are removed, for example, by centrifugation or ultrafiltration. Sump <i>et al., Bio / Technology</i> 10: 163-167 (1992) describes a procedure to assimilate antibodies that they secrete the periplasmic space of <i>E. coli</i>. Briefly, I know thaw the cell paste in the presence of sodium acetate (pH 3.5), EDTA, and phenylmethylsulfonylfluoride (PMSF) for approximately 30 minutes. Cellular debris can be removed by centrifugation When the antibody is secreted in the medium, the supernatants of said expression systems generally first they are concentrated using a protein concentration filter commercially available, for example, a unit of Amicon or Millipore Pellicon ultrafiltration. It can include a protease inhibitor such as PMSF in any of the stages above to inhibit proteolysis and may be included antibiotics to prevent the growth of contaminants adventitious

p0152The antibody composition prepared from of these cells can be purified using, for example, hydroxylapatite chromatography, gel electrophoresis, dialysis, and affinity chromatography, being with affinity chromatography The preferred purification technique. How appropriate is the Protein A as affinity ligand depends on the species and isotype of any immunoglobulin Fc domain that is present in the antibody. Protein A can be used to purify antibodies which are based on heavy chains γ1, γ2, or γ4 (Lindmark <i>et al., J. Immunol. Meth</i>. 62: 1-12 (1983)). G protein is recommended for all mouse and human γ3 isotypes (Guss <i>et al., EMBO J</i>. 5: 1567-1575 (1986)). The matrix a the one that binds the affinity ligand is, in the most frequent way, agarose, but other matrices are available. The matrices mechanically stable such as controlled pore glass or poly (styrenedivinyl) benzene allow speeds of faster flow and shorter processing times than those They can be obtained with agarose. When the antibody comprises a domain C_ 3, Bakerbond ABX ™ resin (JT Baker, Phillipsburg, NJ) is useful for purification. They are also appropriate other techniques for protein purification such as fractionation in an ion exchange column, ethanol precipitation, reverse phase HPLC, chromatography on silica, SEPHAROSE ™ heparin chromatography, single chromatography anionic or cationic exchange resin (such as a column of polyaspartic acid), chromatoenfoque, SDS-PAGE, and precipitation on ammonium sulfate, depending on the antibody to recover.

p0153After any stage or stages of preliminary purification, the mixture comprising the antibody of interest and pollutants can undergo chromatography of low pH hydrophobic interaction using a pH elution buffer between about 2.5-4.5, preferably performed at low salt concentrations (for example, solution approximately 0-0.25 M saline).

C. Pharmaceutical formulations

p0154Therapeutic antibody formulations are prepare to store them by mixing the antibody that has the desired degree of purity with vehicles, excipients or stabilizers physiologically acceptable optimal (<i>Remington's Pharmaceutical Sciences</i> 16th edition, Osol, A. Ed. (1980)), in the form of lyophilized formulations or aqueous solutions. The vehicles, acceptable excipients, or stabilizers are non-toxic to recipients of the dosages and concentrations used, and include buffers such as phosphate, citrate, and other acids organic; antioxidants that include ascorbic acid and methionine; preservatives (such as octadecyldimethylbenzylammonium chloride; hexamethonium chloride; benzalkonium chloride benzethonium; phenolic, butyl or benzyl alcohol; parabens of alkyl such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol); low molecular weight polypeptides (less than about 10 remains); proteins, such as albumin serum, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates that include glucose, mannose, or dextrins; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose or sorbitol; salts that form counterions such as sodium; metal complexes (for example, complexes Zn-protein); and / or nonionic surfactants such such as TWEEN ™, PLURONICS ™ or polyethylene glycol (PEG).

p0155The formulation in this document can also contain more than one active compound, if necessary, for the particular condition being treated, preferably those of complementary activities that do not adversely influence between yes (see section F below). These molecules are present appropriately in combination in amounts that are effective for the intended purpose.

p0156The active ingredients can also be trapped in microcapsules prepared, for example, by techniques of coacervation or polymerization between surfaces, for example, hydroxymethylcellulose or gelatin microcapsule and microcapsule of poly- (methyl methacrylate), respectively, in release systems of colloidal drugs (for example, liposomes, microspheres of albumin, microemulsions, nanoparticles and nanocapsules) or in macroemulsions These techniques are described in<i>Remington's Pharmaceutical Sciences</i> 16th edition, Osol, A. Ed. (1980).

p0157The formulations to use for administration<i>in vivo</i> They must be sterile. This is easily achieved by filtration through sterile filtration membranes.

p0158Release preparations can be prepared sustained. Appropriate examples of release preparations Sustained include semipermeable matrices of hydrophobic polymers which contain the antibody, which are said matrices in the form of shaped articles, for example, films, or microcapsules. The Examples of sustained release matrices include polyesters, hydrogels (for example, poly (2-hydroxyethyl methacrylate), or poly (vinyl alcohol), polylactides (US Pat. No. 3,773,919), copolymers of L-glutamic acid and γ ethyl-L-glutamate, acetate non-degradable ethylene vinyl acid copolymers lactic-degradable glycolic acid such as Lupron Depot ™ (injectable microspheres composed of acid copolymer lactic-glycolic acid and leuprolide acetate), and acid poly-D - (-) - 3-hydroxybutyric. While polymers such as ethylene vinyl acetate and lactic acid-glycolic acid allow a release of molecules for more than a hundred days, certain hydrogels release proteins for shorter periods of time. When they remain antibodies encapsulated in the body for a long time, can be denatured or added as a result of exposure at humidity at 37 ° C, resulting in a loss of activity Biological and possible changes in immunogenicity. They can develop reasonable strategies for stabilization depending of the mechanism involved. For example, if it is discovered that the aggregation mechanism is link formation Intermolecular SS through exchange of thio-disulfide, stabilization can be achieved modifying the sulfhydryl moieties, lyophilizing from acid solutions, controlling moisture content, using appropriate additives, and developing matrix compositions specific polymeric.

D. Non-therapeutic uses of the antibody

p0159The antibodies of the invention can be used. as affinity purification agents. In this process, the antibodies are immobilized in a solid phase such as a resin Sephadex or filter paper, using methods well known in the technique. The immobilized antibody is contacted with a sample containing the VEGF protein (or fragment thereof) to purify, and then wash the support with an appropriate solvent which will remove substantially all the material in the sample except VEGF protein, which binds to the immobilized antibody. Finally, the support is washed with another appropriate solvent, such as buffer glycine, pH 5.0, which will release the VEGF protein from the antibody.

p0160Anti-VEGF antibodies also may be useful in diagnostic assays for VEGF protein, for example, by detecting its expression in cells, tissues, or serum specific. Such diagnostic methods may be useful in cancer diagnosis

p0161For diagnostic applications, the antibody It is typically marked with a detectable moiety. Are appropriate numerous markers that can generally be grouped into following categories:

p0162(a) Radioisotopes, such as 35 S, 14 C, 125 I, 3 H, e 135 I. The antibody can be labeled with the radioisotope using the techniques described in <i>Current Protocols in Immunology</i>, Volumes 1 and 2, Coligen <i>et to the</i>., Ed. Wiley-Interscience, New York, New York, Pubs. (1991), for example, and radioactivity can be measured using a scintillation account.

p0163(b) Fluorescent markers such as chelates rare earths (curopium chelates) or fluorescein or their derivatives, rhodamine and its derivatives, dansyl, lysamine, phycoerythrin and red Texas are appropriate. Fluorescent markers can be conjugated to the antibody using the techniques described in <i>Current Protocols in Immunology, supra</i>, for example. Fluorescence can quantify using a fluorimeter.

p0164(c) Various markers are available enzyme-substrate and U.S. Patent No. 4,275,149 It provides an analysis of some of them. The enzyme usually catalyzes a chemical alteration of the chromogenic substrate that can Measured using various techniques. For example, the enzyme can catalyze a color change in a substrate, which can be measured from a spectrophotometric mode Alternatively, the enzyme can alter the fluorescence or chemiluminescence of the substrate. The techniques to quantify a change in fluorescence have been described previously. The chemiluminescent substrate becomes excited electronically by a chemical reaction and then it can emit light that can be measured (using a chemiluminometer, for example) or donate energy to a fluorescent acceptor. Examples of bookmarks Enzymatic include luciferase (for example, luciferase from firefly and bacterial luciferase; U.S. Patent No. 4,737,456), Luciferin, 2,3-dihydroftalacininediones, malate dehydrogenase, urease, peroxidase such as horseradish peroxidase rusticano (HRPO), alkaline phosphatase, β-galactosidase, glucoamylase, lysozyme, saccharide oxidases (e.g. glucose oxidase, galactose oxidase, and glucose-6-phosphate dehydrogenase), heterocyclic oxidases (such as uricase and xanthine oxidase), lactoperoxidase, microperoxidase, and the like. The Techniques for conjugating enzymes to antibodies are described in O ' Sullivan <i>et al</i>., Methods for the Preparation of Enzyme-Antibody Conjugates for use in Enzyme Immunoassay, in <i>Methods in Enzym</i>. (ed J. Langone &amp; H. Van Vunakis), Academic press, New York, 73: 147-166 (1981).

p0165The examples of combinations substrate enzymes include, for example:

p0166(i) horseradish peroxidase (HRPO) with hydrogen peroxidase as a substrate, where hydrogen peroxidase oxidizes a precursor with color (for example, orthophenylenediamine (OPD) or 3,3 ', 5,5'-tetramethylbenzidine hydrochloride (TMB));

p0167(ii) alkaline phosphatase (AP) with of para-nitrophenyl as a chromogenic substrate; and

p0168(iii) β-D-galactosidase (β-D-Gal) with a substrate chromogenic (for example, p-nitrophenyl-? -D-galactosidase) or the fluorogenic substrate 4-methylumbelliferyl-? -D-galactosidase.

p0169Other numerous combinations enzyme-substrate are appropriate for Technical specialists. For a general analysis of these, see US Patents No. 4,275,149 and No. 4,318,980.

p0170Sometimes the marker is conjugated indirectly with the antibody. The specialist in the technique will be aware of various techniques to achieve this. For example, him antibody can be conjugated with biotin and any of the three main categories of bookmarks mentioned above can conjugate with avidin, or vice versa. Biotin binds in a way avidin selective and thus the marker can be conjugated with the antibody in this indirect mode. As an alternative, for achieve indirect conjugation of the label with the antibody, the antibody is conjugated with a small hapten (for example, digoxin) and one of the different types of markers mentioned previously conjugated with an antibody anti-hapten (for example, antibody anti-digoxin). In this way, the indirect conjugation of the marker with the antibody.

p0171In another embodiment of the invention, the anti-VEGF antibody does not need to be labeled, and the Its presence can be detected using a labeled antibody which binds to the anti-VEGF antibody.

p0172The antibodies of the present invention can be used in any known test method, such as tests of competitive union, direct or indirect sandwich tests, and immunoprecipitation assays. Zola,<i>Monoclonal Antibodies: A Manual of Techniques</i>, P. 147-158 (CRC Press, Inc. 1987).

p0173Competitive union trials are based on the ability of a marked pattern to compete with the analyte of the test sample to join with a limited amount of antibody. The amount of VEGF protein in the test sample is inversely proportional to the amount of pattern that reaches bind to antibodies. To facilitate the determination of the amount of pattern that comes to join, usually Insolubilize antibodies before or after the competition, from so that the pattern and the analyte that bind to the antibodies can be adequately separated from the standard and analyte that They remain without joining.

p0174Sandwich tests involve the use of two antibodies, each capable of binding to an immunogenic portion different, or epitope, of the protein to be detected. In a type essay sandwich, the analyte of the test sample is joined by a first antibody that is immobilized on a solid support, and subsequently a secondary antibody binds to the analyte, forming in this way an insoluble tripartite complex. See, for example, U.S. Patent No. 4,376,110. The secondary antibody can by itself be marked with a detectable remainder (type tests direct sandwich) or can be measured using an antibody anti-immunoglobulin that is marked with a residue detectable (indirect sandwich test). For example, a type of sandwich test is an ELISA test, in which case the rest detectable is an enzyme.

p0175For immunohistochemistry, the tumor sample can be fresh or frozen or can be embedded in paraffin and fixed with a preservative such as formalin, for example.

p0176Antibodies can also be used to diagnostic tests <i>in vivo</i>. Generally the antibody is labeled with a radionuclide (such as 111 In, 99 Tc, 14 C, 131 I, 125 I, 3 H, 32 P or 35 S) of so that the tumor can be located using immunocentelleography.

E. Diagnostic kits

p0177For convenience, the antibody of the The present invention can be provided in a kit, that is, a packaged combination of reagents in predetermined quantities with instructions to perform in diagnostic test. When he antibody is labeled with an enzyme, the kit will include substrates and cofactors required by the enzyme (for example, a substrate precursor that provides the detectable chromophore or fluorophore). In addition, other additives such as stabilizers may be included, buffers (for example, a blocking buffer or a lysis buffer) and Similar. The relative amounts of the various reagents can vary widely to provide solution concentrations of reagents that substantially optimize the sensitivity of the test. Particularly, the reagents can be provided as dry powders, usually lyophilized, including excipients which, in solution, will provide a reagent solution that It has the proper concentration.

F. Therapeutic uses for the antibody

p0178For therapeutic applications, the antibodies anti-VEGF of the invention are administered to a mammal, preferably a human, in a dosage form pharmaceutically acceptable, as discussed above, including those that can be administered to a human via intravenously as a bolus or by continuous infusion over a period of time, intramuscularly, intraperitoneally, intracerebroespinal, subcutaneous, intraarticular, intrasynovial, intrathecal, oral, topical, or by inhalation. Antibodies too are properly administered intratumorally, peritumorally, intralesional, or perilesional, to exert therapeutic effects local as well as systemic. The intraperitoneal route is expected be particularly useful, for example, in the treatment of tumors ovarian

p0179For the prevention or treatment of disease, the appropriate dosage of antibody will depend on the type of disease to be treated, as defined above, the severity and course of the disease, if the antibody is administered for preventive or therapeutic purposes, therapy previous, the patient's medical history and response to antibody, and the judgment of the doctor you are attending. He antibody is properly administered to the patient at one time or during a series of treatments.

p0180The anti-VEGF antibodies are useful in the treatment of various diseases and disorders Neoplastic and non-neoplastic. Neoplasms and conditions Related that are susceptible to treatment include carcinomas breast, lung carcinomas, gastric carcinomas, carcinomas esophageal, colorectal carcinomas, liver carcinomas, ovarian carcinomas, tecomas, arrenoblastomas, carcinomas cervical, endometrial carcinoma, endometrial hyperplasia, endometriosis, fibrosarcomas, choriocarcinoma, head cancer and neck, nasopharyngeal carcinoma, laryngeal carcinoma, hepatoblastoma, Kaposi's sarcoma, melanoma, skin carcinomas, hemangioma, cavernous hemangioma, hemangioblastoma, carcinomas of the pancreas, retinoblastoma, astrocytoma, glioblastoma, schwannoma, oligodendroglioma, medulloblastoma, neuroblastoma, rhabdomyosarcoma, osteogenic sarcoma, leiomyosarcomas, tract carcinomas urinary, thyroid carcinomas, Wilm's tumor, cell carcinoma renal, prostate carcinoma, abnormal vascular proliferation associated with phacomatosis, edema (such as that associated with tumors brain) and Meig syndrome.

p0181The non-neoplastic conditions that are susceptible to treatment include rheumatoid arthritis, psoriasis, arteriosclerosis, diabetic retinopathies and other retinopathies proliferatives that include retinopathy of prematurity, fibroplasia retrolental, neovascular glaucoma, related macular degeneration with age, thyroid hyperplasias (including disease of Severe), corneal transplantation and transplantation of other tissues, chronic inflammation, pulmonary inflammation, nephrotic syndrome, preeclampsia, ascites, pericardial effusion (such as associated with pericarditis), and pleural effusion.

p0182Macular degeneration associated with age (AMD) is a cause that leads to severe vision loss in elderly population The exudative form of AMD is characterized by Choroidal neovascularization and detachment of cells from the retinal pigment epithelium. As choroidal neovascularization is associated with a drastic worsening in prognosis, it expect the anti-VEGF antibodies of the present invention are especially useful in reducing the severity of AMD

p0183Depending on the type and severity of the disease, about 1 µg / kg to about 50 mg / kg (for example, 0.1-20 mg / kg) of antibody is a dosage initial candidate for patient administration, therefore, for for example, by one or more administrations separately, or by infusion keep going. A typical daily or weekly dosage may vary from about 1 µg / kg to about 20 mg / kg or more, depending on the factors mentioned above. For repeated administrations the treatment is repeated several days or more, depending on the condition, until suppression happens Desired symptoms of the disease. However, they can be Other dosage regimens are useful. The progress of this therapy It is easily controlled by conventional techniques and tests, including, for example, tumor radiographic images.

p0184The efficacy of the antibody in preventing and treating the disease can be improved by administering the antibody in a manner serial or in combination with another agent that is effective for those purposes, such as tumor necrosis factor (TNF), an antibody able to inhibit or neutralize the angiogenic activity of the factor of acid or basic fibroblast growth (FGF) or the factor of hepatocyte growth (HGF), an antibody capable of inhibiting or neutralize the clotting activities of the tissue factor, the protein C, or protein S (see Esmon <i>et al</i>., Publication of PCT Patent No. WO 91/01753, published February 21, 1991), an antibody capable of binding to the HER2 receptor (see Hudziak <i>et to the</i>., PCT Patent Publication No. WO 89/06692, published on 27 July 1989), or one or more conventional therapeutic agents such as, for example, alkylating agents, acid antagonists folic, nucleic acid metabolism antimetabolites, antibiotics, pyrimidine analogues, 5-fluoroacyl, cisplatin, purine nucleosides, amines, amino acids, nucleosides of thiazole, or corticosteroids. These other agents may be present in the composition being administered or may be administered separately In addition, the antibody is administered properly serially or in combination with treatments radiological, which involve both irradiation and administration of radioactive substances

p0185Vascularization of tumors can be fought in combination therapy The antibody and one or more are administered. anti-VEGF antagonists other than patients that have tumor at therapeutically effective doses as determines, for example, by observing the necrosis of the tumor or its metastatic foci, if any. This therapy is continued until no more beneficial effects are observed or clinical exams are not show tumor trace or any metastatic focus. Then administers TNF, alone or in combination with such an auxiliary agent as interferon alpha, beta or gamma, antibody anti-HER2, heregulin, antibody anti-heregulin, factor D, interleukin-1 (IL-1), interleukin-2 (IL-2), factor of stimulation of granulocyte-macrophage colonies (GM-CSF), or reagents that promote coagulation vascular in tumors, such as antibody anti-protein C, antibody anti-protein S, or C4b binding protein (see Esmon <i>et al</i>., PCT Patent Publication No. WO 91/01753, published on February 21, 1991), or heat or radiation.

p0186How auxiliary agents will vary in their efficacy, it is desirable to compare its impact on the tumor by Matrix scan in a conventional way. The administration of anti-VEGF antibody and TNF repeats until it is Get the desired clinical effect. Alternatively, the antibody anti-VEGF is administered together with TNF and, optionally, auxiliary agent or agents. Sometimes when find solid tumors in the extremities or in others locations susceptible to circulation isolation In general, the direct therapeutic agents in this document are administered to the tumor or isolated organ. It can be administered FGF antagonist or platelet-derived growth factor (PDGF), such as a neutralizing antibody anti-FGF or anti-PDGF, to the patient together with the anti-VEGF antibody. The treatment with anti-VEGF antibodies can be suspended from Optimal manner during periods of wound healing or desired neovascularization.

G. Manufacturing items

p0187A manufacturing article is contemplated that contains useful materials for the treatment of disorders described above. The article of manufacture comprises a container and a marker. Appropriate containers include, by example, boats, vials, syringes, and test tubes. The containers can be formed from a variety of materials such Like glass or plastic. The container contains a composition that It is effective for the treatment of the condition and may have a route sterile access (for example, the container can be a bag or a vial of intravenous solution that has a plug that can be pierced by a hypodermic injection needle). The active agent in the Composition is the anti-VEGF antibody. Marker in, or in association with, the container indicates that the composition is Use to treat the condition of choice. The article of manufacture it can additionally comprise a second container comprising a pharmaceutically acceptable buffer, such as a saline solution buffered with phosphate, Ringer's solution and dextrose solution. It can also include other desirable materials from a point of commercial and useful view, including other buffers, diluents, filters, needles, syringes, and leaflets with instructions for your use.

Example 1

p0188This example describes the production of humanized anti-VEGF antibodies with properties desirable from a therapeutic point of view.

Materials and methods

p0189<i>Cloning of MAb A.4.6.1 Murino and Construction of Chimeric Mouse-Human Fab</i>: MAb A.4.6.1 murine anti-VEGF has been previously described by Kim<i>et al., Growth Factors</i> 7:53 (1992) and Kim <i>et al., Nature</i> 362: 841 (1993). Total RNA from cells was isolated hybridoma that produce Mab A.4.6.1. anti-VEGF using RNAsol (TEL-TEST) and transcribed to the inverse to cDNA using the Oligo-dT primer and the SuperScript II system (GIBCO BRL, Gaithersburg), MD). The combinations of degenerate, based oligonucleotide primers in the N-terminal amino acid sequences of the light and heavy chains of the antibody, were synthesized and They used as direct primers. Inverse primers were based in the 4 flanking sequences obtained from the kV subgroup of light chains and subgroup II of murine heavy chains (Kabat<i>et al., Sequences of Proteins of Immunological Interest</i>. 5th ed Public Health Service, National Institutes of Health, Bethesda, MD. (1991)). After amplification by chain reaction of Polymerase (PCR) DNA fragments were ligated to a vector of TA cloning (Invitrogen, San Diego, CA). Eight were sequenced clones of each of the heavy and light chains. They subcloned a clone with a consensus sequence for the chain VL domain lightweight and one with a consensus sequence for the VH domain of the heavy chain, respectively, in vector pEMX1 containing the human CL and CH1 domains (Werther <i>et al., J. Immunol</i>. 157: 4986-4995 (1996)), generating a mouse chimera- human. This chimeric F (ab) consists of the VH domain A.4.6.1 complete murine fused to a human CH1 domain in the amino acid SerH113 and the VL domain A.4.6.1 murine complete fused to a human CL domain in amino acid LisL107. The expression and purification of the chimeric F (ab) were identical to those of the humanized F (ab). The chimeric F (ab) was used as pattern in binding assays.

p0190<i>Computer Graphics Models of F (ab) Humanized and Murine</i>: The sequences of the VL and VH domains (Figures 1A and 1B) were used to construct a graphic model Computer of the VL-VH domains A.4.6.1 murine. This model was used to determine which flanking remains should incorporated into the humanized antibody. Also built a Humanized F (ab) model to verify selection correct of the murine flanking remains. The construction of the Models were performed as previously described (Carter <i>et al., Proc. Natl. Acad. Sci. USA</i> 89: 4285-4289 (1992) and Eigenbrot <i>et al., J. Mol. Biol</i>. 229:969-995 (1993)).

p0191<i>Construction of humanized F (ab)</i>: Plasmid pEMX1 used for mutagenesis and expression of F (ab) in <i>E. coli</i> has been previously described (Werther <i>et al., supra</i>). Briefly, the plasmid contains a DNA fragment which encodes a consensus sequence of subgroup I of strings light human κ (VL \ kappa-CL) and a consensus sequence of subgroup III of human heavy chains (VHIII-CHI) and an alkaline phosphatase promoter. He Use of consensus sequences for VL and VH has been described previously (Carter <i>et al., supra</i>).

p0192To build the first variant of F (ab) of A.4.6.1. humanized, F (ab) -1, site directed mutagenesis was performed (Kunkel <i>et al., Proc. Natl. Acad. Sci. USA</i> 82: 488-492 (1985)) in a pEMX1 template containing deoxyuridine. The six CDRs agree with Kabat <i>et al., supra</i>, they changed to the sequence of A.4.6.1 murine. Therefore, F (ab) -1 consists of a complete human flanking region (VL? subgroup I and VH subgroup III) with the six complete murine CDR sequences. He they constructed plasmids for all other variants of F (ab) from the plasmid mold of F (ab) -1. Plasmids were transformed into the strain XL-1 Blue de <i>E. coli</i> (Stratagene, San Diego, CA) for the preparation of double stranded and stranded DNA simple. For each variant, the DNA that was completely sequenced encode the light and heavy chains using the method of dideoxynucleotides (Sequenase, US Biochemical Corp., Cleveland, OH). Plasmids were transformed into strain 16C9 of<i>AND. coli</i>, a derivative of MM294, were seeded in broth plates Luria containing 50 µg / ml carbenicillin, and selected A simple colony for protein expression. The simple colony was grown in 5 ml of Luria broth-100 mg / ml of carbenicillin for 5-8 hours at 37 ° C. The 5 ml of culture was added to 500 ml of AP5-50 µg / ml of carbenicillin and allowed to grow for 20 hours in a flask of agitated stirring of 4 liters at 30 ° C. The AP5 medium consists of 1.5 g of glucose, 11.0 g of Hycase SF, 0.6 g of yeast extract (certificate), 0.19 g of MgSO4 (anhydrous), 1.07 g of NH 4 Cl, 3.73 g of KCl, 1.2 g of NaCl, 120 ml of triethanolamine 1 M, pH 7.4, at 1 liter of water and then filtered in sterility to through a 0.1 mm Sealkeen filter. The cells were collected by centrifugation in a 1 liter centrifuge canister at 3000 xg and the supernatant was collected. After freezing for 1 hour, the sediment was resuspended in 25 ml of 10 mM Tris-EDTA 1 mM-20% sucrose, pH 8.0, cold. 250 were added ml of 0.1 M benzamidine (Sigma. St Louis, MO) to inhibit proteolysis After stirring gently on ice for 3 hours, The sample was centrifuged at 40,000 xg for 15 minutes. Later applied the supernatant to 1 protein column G-Sepharose CL-4B (Pharmacia, Uppsala, Sweden) (0.5 ml bed volume) balanced with Tris 10 mM-1 mM EDTA, pH 7.5. The column was washed with 10 ml. of 10 mM Tris-1 mM EDTA, pH 7.5, and eluted with 3 ml of 0.3 M glycine, pH 3.0, in 125 ml of 1 M Tris, pH 8.0. Later exchanged the F (ab) buffer for PBS using a Centricon 30 (Amicon, Beberly, MA) and concentrated to a final volume of 0.5 ml. SDS-PAGE gels were run for all F (ab) to determine the purity and molecular weight of each variant was verified by mass spectrometry by electrospray

p0193<i>Construction and Expression of Chimeric IgG and Humanized</i>: For the generation of human IgG variants of A.4.6.1 chimeric (chIgG1) and humanized (huIgG1), were subcloned the appropriate murine or humanized VL and VH domains (F (ab) -12, Table 2) in described pRK vectors previously, separately (Eaton <i>et al., Biochemistry</i>25: 8343-8347 (1986)). The AND that encodes the string light and heavy complete of each variant was verified by dideoxynucleotide sequencing.

p0194For the transient expression of the variants, heavy and light chain plasmids were co-infected in 293 human cells (Graham <i>et al., Gen. Virol</i>. 36: 59-74 (1977)), using a discharge procedure effectiveness (Gorman <i>et al., DNA Prot. Eng. Tech</i>. 2: 3-10 (1990)). The medium was changed to one free of serum and was collected daily for up to five days. The antibodies were purified from the combined supernatants using Protein- Sepharose CL-4B (Pharmacia). He exchanged the buffer of the eluted antibody for PBS using a Centricon-30 (Amicon), concentrated to 0.5 ml, was filtered under sterile conditions using a Millex-GV (Millipore, Bedford, MA) and stored at 4 ° C.

p0195For stable expression of the IgG1 variant Humanized final (rhuMAb VEGF), ovarian cells were transfected Chinese hamster (CHO) with dicistronic vectors designed to co-express both heavy and light chain (Lucas <i>et al., Nucleic Acid Res</i>. 24: 1774-79 (1996)). The Plasmids were introduced into DP12 cells, a proprietary derivative of the CHO-KI DUX B11 cell line developed by L. Chasin (Columbia University), by lipofection and selected for growth in GHT-free medium (Chisholm, V. High efficiency gene transfer in mammalian cells. In: Glover, DM, Hames, BD. <i>DNA Cloning 4. Mammalian systems</i>. Oxford Univ. Press, Oxford p. 1-41 (1996)). They were chosen randomly approximately 20 non-amplified clones and it reseeded in 96-well plates. Relative productivity specific to each clone was monitored using an ELISA to quantify the full length human IgG accumulated in each well after 3 days, and a fluorescent stain, Calcein AM, as a substitute marker for the amount of viable cells per well. Based on this data, several non-clones were chosen amplified for further amplification in the presence of increasing concentrations of methotrexate. Clones were chosen individuals who survived 10, 50 and 100 nM methotrexate and were transfected into 96-well plates for the exploration of productivity. A clone was expanded, which showed so reproducible high specific productivity, in T flasks and used to inoculate a rotating culture. After several passes, it they used suspension-adapted cells to inoculate cultures of production in serum-free media containing GHT, supplemented with various hormones and protein hydrolysates. He collected cell culture fluid, which contained rhuMAb VEGF, was purified using A-Sepharose protein CL-4B The purity after this stage was \ sim99%. Post homogeneity purification was performed using an ion exchange chromatography step. He Endotoxin content of the final purified antibody was &lt; 0.10 eu / mg

p0196<i>Quantification of F (ab) and IgG</i>: For quantify the F (ab) molecules, plates were coated ELISA with 2 µg / ml of goat F (ab) anti-human IgG (Organon Teknika, Durham, NC) in 50 mM carbonate buffer, pH 9.6, at 4 ° C overnight and blocked with 0.5% bovine serum albumin PBS (blocking buffer) at room temperature for 1 hour. The standards (0.78-50 ng / ml of human F (ab)) are acquired from Chemicon (Temecula, CA). Dilutions were incubated Serial samples in PBS-serum albumin 0.5% bovine-0.05% 20-fluororbate (assay buffer) in the palcas for 2 hours. F (ab) binding was detected using Goat anti-human IgG F (ab) labeled with horseradish peroxidase (Organon Teknika), followed by 3,3 ', 5,5'-tetramethylbenzidine (Kirkegaard &amp; Perry Laboratories, Gaithersburg, MD) as a substrate. The plates are They washed between the stages. The absorbance at 450 nm was read in a Vmax plate reader (Molecular Devices, Menlo Park, CA). The curve pattern was adjusted using a regression curve adjustment program Nonlinear four parameters. The point data that fell in the interval of the standard curve was used to calculate the F (ab) concentrations of the samples. Concentration of the full length antibody was determined using goat Fc anti-human IgG (Cappel, Westchester, PA) for the capture and Fc of goat anti-human IgG marked with horseradish peroxidase (Cappel) for detection. It was used Human IgG1 (Chemicon) as a standard.

p0197<i>Union test to VEGF</i>: To measure the VEGF binding activity of F (ab), plates were coated ELISA with 2 µg / ml of rabbit F (ab ') 2 against Fc of Human IgG (Jackson ImmunoResearch, West Grove, PA) and were blocked with blocking buffer (described above). The middle diluted conditioned containing 3 ng / ml KDR-IgG (Park <i>et al., J. Biol. Chem</i>. 296: 25646-25645 (1994)) in blocking buffer, it was incubated in the bale for 1 hour. Patterns were incubated (6.9-440 ng / ml of F (ab) chimeric) and serial twice dilutions of samples with 2 nM biotinylated VEGF for 1 hour in tubes. Later transferred the tube solutions to the ELISA plates and incubated for 1 hour. After washing, VEGF was detected biotinylated bound to KDR using streptavidin labeled with horseradish peroxidase (Zymed, South San Francisco, CA or Sigma, St Louis, MO) followed by 3,3 ', 5,5'-tetramethylbenzidine as substrate. The titration curves were adjusted with a curve fit program non-linear four-parameter regression (KaleidaGraph, Synergy Software, Reading PA). The concentrations of the variants of F (ab) corresponding to the midpoint of absorbance in the pattern titration curve, and after divided by the concentration of the pattern corresponding to the point absorbance medium of the standard titration curve. The essays for full length IgG were the same as for F (ab) except that the test buffer contained serum 10% human.

p0198<i>BIAcore ™ Biosensor Assay</i>: The union to VEGF of the humanized and chimeric F (ab) was compared using a BIAcore ™ biosensor (Karlsson <i>et al., Methods</i>: <i>TO Comparison to Methods in Enzymology</i> 6: 97-108 (1994)). F (ab) concentrations were determined by quantitative amino acid analysis. VEGF joined a chip CM-5 biosensor through the amine groups Primary according to the manufacturer's instructions (Pharmacia). The dissociation kinetics was measured by saturating the chip with F (ab) (35 µL of F (ab) 2 µM at a flow rate of 20 µl / min) and then switching to buffer (PBS-0.05% polysorbate 20). Punctual data from 0-4500 seconds were used for the analysis of Kinetics of dissociation. The dissociation rate constant (k_ {off}) was obtained from the slope of the graph of ln (R0 / R) versus time, where R0 is the signal at = 0 and R is the signal at each specific time.

p0199Association kinetics was measured using Serial dilutions of F (ab) twice (0.0625-2 mM). The slope, K_ {s}, was obtained from the graph of ln (-dR / dt) versus time for each concentration of F (ab) using the BIAcore ™ kinetic evaluation program described in the Pharmacia Biosensor manual. R is the signal at time t. Data between 80 and 168, 148, 128, 114, 102, and 92 seconds are used for F (ab) 0.0625, 0.125, 0.25, 0.5, 1, and 2 mM respectively. The association speed constant (k_ {on}) it was obtained from the slope of the graph of K_ {s} versus concentration of F (ab). At the end of each cycle, the F (ab) bound by injecting 5 µl of 50 mM HCl at a rate 20 µl / min flow to regenerate the chip.

p0200<i>Endothelial Cell Growth Assay</i>: Capillary endothelial cells obtained from bark were cultured bovine adrenal in the presence of Eagle's medium modified by Low glucose Dulbecco (DMEM) (GIBCO) supplemented with serum 10% veal, 2 mM glutamine, and antibiotics (medium of growth), essentially as previously described (Leung<i>et al., Science</i> 246: 1306-1309 (1989)). For mitogenic assays, endothelial cells were seeded at a density of 6 x 10 3 cells per well, in 6 plates wells, in growth medium. Then muMAb VEGF were added A.4.6.1 or rhuMAb VEGF at concentrations ranging between 1 and 5000 ng / ml After 2-3 hours, rhVEGF165 was added expressed by <i>E. coli</i> at a final concentration of 3 ng / ml. For specificity control, each antibody was added to endothelial cells at the concentration of 5000 ng / ml, alone or in the presence of 2 ng / ml of bFGF. After five or six days, the dissociated cells by trypsin exposure and counted in a Cuolter counter (Coulter Electronics, Hialeah, FL). The variation of the average did not exceed 10%. The data was analyzed by a four parameter curve adjustment program (KaleidaGraph).

p0201<i>Tumor Studies <u>Live</u></i>: He cultured human rhabdomyosarcoma A673 cells (ATCC; CRL 1598) as previously described, in DMEM / F12 supplemented with serum 10% fetal bovine, 2 mM glutamine and antibiotics (Kim <i>et al., Nature</i> 362: 841-844 (1993) and Borgström <i>et al., Cancer Res</i>. 56: 4032-4039 (1996)). Were injected subcutaneously nude mice BALB / c females of 6-10 weeks of age, with 2 x 10 6 cells tumor in the dorsal area in a volume of 200 µl. Then, the animals were treated with muMAb VEGF A.4.6.1, rhuMAb VEGF or a MAb control directed against gp120 protein (Kim <i>et al., Nature</i> 362: 841-844 (1993)). They were administered both anti-VEGF MAb at doses of 0.5 and 5 mg / kg; he Control MAb was given at the dose of 5 mg / kg. Each MAb was administered twice a week intraperitoneally in a volume of 100 µm, starting 24 hours after cell inoculation Tumor Each group consisted of 10 mice. Tumor size It was determined at weekly intervals. Four weeks after inoculation of tumor cells, the animals underwent Euthanasia and tumors were removed and weighed. The analysis Statistical was performed by ANOVA.

Results

p0202<i>Humanization</i>: The consensus sequence was used for subgroup III of human heavy chains and subgroup k I of human light chains as a flanking region for humanization (Kabat <i>et al., supra</i>) (Figures 1A and 1B). This flanking region has been used successfully in humanization of other murine antibodies (Werther <i>et al., supra</i>; Sump<i>et al., supra</i>; Lend<i>et al., J. Immunol</i>. 151: 2623-2632 (1993); and Eigenbrot<i>et al., Protein</i> 18: 49-62 (1994)). CDR-H1 included residues H26-H35. The other CDRs agreed with Kabat <i>et al., supra</i>. All humanized variants were made at first and explored for union as the F (ab) expressed in <i>AND. coli</i>. Typical yields of shake flasks of 500 ml was 0.1-0.4 mg of F (ab).

p0203F (ab) chimeric was used as a standard in the binding assays In the initial variant, F (ab) -1, the remains of CDR were transferred of the murine antibody to the human flanking region and, based on the methods of the murine and humanized F (ab), the rest in position H49 (Wing in human) was changed to murine Gly. In addition, the F (ab) consisting of the chimeric heavy chain / light chain of F (ab) -1 (F (ab) -2) and the heavy chain of F (ab) -1 / light chain chimeric (F (ab) -3) were generated and were rehearsed for binding. F (ab) -1 showed a affinity more than 1000 times reduced than that of chimeric F (ab) (Table 2). Comparing the binding affinities of F (ab) -2 and F (ab) -3 se suggests that the remnants of the flanking region in the VH domain of F (ab) -1 need to be altered to increase the Union.

TABLE 2

Union of variants of F (ab) Anti-VEGF Humanized to VEGF a

<figref>3</figref>

<pre listing-type="other">\ dotable {\ tabskip \ tabcolsep # \ hfil \ tabskip0ptplus1fil \ dddarstrut \ cr} { \ begin {minipage} {150mm} ^ {a} Variants of F (ab) anti-VEGF were incubated with biotinylated VEGF and then transferred to ELISA plates coated with KDR-IgG (Park et al., supra). \ end {minipage} \ cr \ begin {minipage} {150mm} b The murine remains are underlined; the rest numbers are according to Kabat et al., supra. \ end {minipage} \ cr \ begin {minipage} {150mm} ^ {c} The mean and standard deviation they are the average of the proportions calculated for each of the independent trials; the EC50 for chimeric F (ab) was 0.049 ± 0.013 mg / ml (1.0 nM). \ end {minipage} \ cr}</pre>

p0204Changing the human remains H71 and H72 to their murine equivalents in F (ab) -4 the union in 4 times (Table 2). The inspection of the models of F (ab) murine and humanized it is suggested that the rest L46, interned on the contact surface VL-VH and that interacts with CDR-H3 (Figure 2), it could also play a role in determining the conformation of CDR-H3 and / or affect the relationship of VL domains and VH. When the murine Val is exchanged for the human Leu in L46 (F (ab) -5), the binding affinity increases by almost 4 times (Table 2). Three other interned remains were evaluated in the flanking region based on molecular models: H49, H69 and H78. The H69 position may affect the conformation of CD-H2 while position H78 can affect the conformation of CDR-H1 (Figure 2). When each one was changed individually from the human equivalent to the murine, the union improved in 2 times in each case (F (ab) -6 and F (ab) -7, Table 2). When both changed simultaneously, the improvement in the union was 8 times (F (ab) -8, Table2). The rest H49 was included in a principle like the murine Gly; when he switched to the wing equivalent human consensus, the union was reduced by 15 times (F (ab) -9, Table 2).

p0205In F (ab) -10 and F (ab) -11 two remains were changed in the loop flanking 3, FR-3, to its murine equivalents: AsnH76 to be murine (F (ab) -10) and LysH75 to Ala murine (F (ab) -11). Both achieved an improvement relatively small at the junction (Table 2). Finally, in the H94 position the human and murine sequences have, in the most frequent, an Arg (Kabat <i>et al., supra</i>). In F (ab) -12, this Arg is replaced by the Lys uncommon found in the murine antibody (Figure 1A) and this gives as a result a union that was 2 times smaller than that of F (ab) chimeric (Table 2). The F (ab) -12 it was also compared to the chimeric F (ab) using the system BIAcore ™ (Pharmacia). Using this technique, the K_ {d} of the Humanized F (ab) -12 was 2 times weaker than that of chimeric F (ab) due to a slower K_ {on} and a Faster K_ {off} (Table 3).

TABLE 3

Union of variants of F (ab) Anti-VEGF to VEGF Using System ^ a BIAcore ™

<figref>5</figref>

<pre listing-type="other">\ dotable {\ tabskip \ tabcolsep # \ hfil \ tabskip0ptplus1fil \ dddarstrut \ cr} { \ begin {minipage} {150mm} ^ {a} The amount of F (ab) attached, in resonance units (RU), was measured using a system BIAcore ™ when 2 µg of F (ab) was injected into a chip containing 2480 RU of immobilized VEGF. The kinetics of dissociation (k off) was measured by saturating the chip with F (ab) and then controlling the dissociation after changing the buffer. The kinetic association (k on) was measured using dilutions Serials of F (ab) twice. The equilibrium constant of dissociation K d was calculated as k off / k on. \ end {minipage} \ cr \ begin {minipage} {150mm} ^ {b} quim-F (ab) is a chimeric F (ab) with murine VL and VH domains fused to CL and CH1 domains of the chain heavy human. \ end {minipage} \ cr}</pre>

p0206Full length mABs were built merging the VL and VH domains of the chimeric F (ab) and the variant F (ab) -12 to constant domains of the human k light chain and the human IgG1 heavy chain. He 12-length full length IgG1 (F (ab) -12 fused to human IgG1) showed a junction that was 1.7 times weaker than that of chimeric IgG1 (Table 4). Both 12-IgG1 and chimeric IgG1 bound slightly less well than the original murine MAb A.4.6.1 (Table 4).

TABLE 4

Union of variants of IgG Anti-VEGF a VEGF a

<figref>6</figref>

<pre listing-type="other">\ dotable {\ tabskip \ tabcolsep # \ hfil \ tabskip0ptplus1fil \ dddarstrut \ cr} { \ begin {minipage} {150mm} ^ {a} IgG variants anti-VEGF were incubated with biotinylated VEGF and then transferred to ELISA plates coated with KDR-IgG (Park et al., (1994), supra). \ end {minipage} \ cr \ begin {minipage} {150mm} <b> chIgG1 is a chimeric IgG1 with murine VL and VH domains fused to CL and human IgG1 heavy chains; the EC50 for chIgG1 was 0.113 ± 0.013 µg / ml (0.75 nM). \ end {minipage} \ cr \ begin {minipage} {150mm} ^ {c} murIgG1 is a muMabVEGF A.4.6.1 purified fluid ascitic. \ end {minipage} \ cr \ begin {minipage} {150mm} ^ {d} 12-IgG1 is the VL and VH domains of F (ab) -12 fused to CL and heavy chains of IgG1 human. \ end {minipage} \ cr}</pre>

p0207<i>Biological Studies</i>: rhuMAb VEGF and muMAb VEGF A.4.6.1 were compared for their ability to inhibit the bovine capillary endothelial cell proliferation in response at a maximum efficiency concentration of VEGF (3 ng / ml). How I know illustrated in Figure 3, the two MAb were essentially equivalent, both in potency and in effectiveness. ED50 values ​​were respectively 50 ± 5 ng / ml and 48 ± 8 ng / ml (~ 0.3 nM). In both cases a 90% inhibition was achieved at 500 ng / ml concentration (sim3 nM). Neither muMAb VEGF A.4.6.1 nor rhuMAb VEGF had no effect on basal proliferation or stimulated by bFGF of capillary endothelial cells, confirming that the inhibition is specific for VEGF.

p0208To determine whether such equivalence is also applies to a system <i>in vivo</i>, the two antibodies are compared for their ability to suppress cell growth A673 of human rhabdomyosarcoma in nude mice. The studies Previous have shown that muMAb VEGF A.4.6.1 has an effect drastic inhibitor in this tumor model (Kim <i>et al., Nature</i>362: 841-844 (1993) and Borgström <i>et al., Cancer Beef</i> 56: 4032-4039 (1996)). As shown in the Figure 4, at both doses tested (0.5 and 5 mg / kg), both antibodies markedly suppressed tumor growth as assessed by tumor weight measurements 4 weeks after cell inoculation. The decrease in the weight of tumor compared to the control group was respectively 85% and 93% at each dose in animals treated with VEGF muMAb A.4.6.1 versus 90% and 95% in those treated with rhuMAb VEGF. He they obtained similar results with the carcinoma cell line of MDA-MB 435 breast.

Example 2

Antecedent

p0209In this example, antibody A.4.6.1 murine anti-VEGF analyzed above was humanized by randomization of a small series of flanking remains and by monovalent presentation of the resulting library of molecules of antibody on the surface of filamentous phages to identify high affinity flanking sequences by selection affinity based.

Materials and methods

p0210<i>Construction of the Phagemid Vector Anti-VEGF, pMB4-19</i>: The mAb A.4.6.1 murine anti-VEGF has been analyzed earlier in Example 1. The first variant of F (ab) of humanized A.4.6.1, hu2.0, was constructed by directed mutagenesis site using a plasmid pAK2 template containing Deoxyuridine (Carter <i>et al., Proc. Natl. Acad. Sci. USA</i>. 89: 4285-4289 (1992)) encoding a light chain V_ {L} \ kappaI-C \ kappa_ {1} and an Fd fragment of the heavy chain V H III-C H I. The A.4.6.1 transplanted CDR sequences were chosen according to the Kabat sequence definition <i>et al., supra</i>, except for CDR-H1 that included residues 26-35. The Fab coding sequence subcloned into the phagemid vector phGHamg3 (Bass <i>et al., Proteins</i> 8: 309-314 (1990) and Lowman <i>et al., Biochemistry</i> 30: 10832-10838 (1991)). This construction, pMB4-19, encodes the Fab of humanized A.4.6.1 initial, hu2.0, with the C-terminal of the chain heavy precisely fused to the carboxyl portion of the M13 gene III envelope protein. pMB4-19 is similar in construction to pDH188, a plasmid previously described for the monovalent presentation of Fab fragments (Garrard <i>et al., Biotechnology</i> 9: 1373-1377 (1991)). The Notable differences between pMB4-19 and pDH188 include a shorter segment of gene III of M13 (codons 249-406) and the use of an amber stop codon immediately after the Fd fragment of the heavy chain of the antibody. This allows the expression of the secreted heavy chain or heavy chain-gene III fusions in strains suppressors <i>sup</i> from <i>E. coli</i>.

p0211<i>Expression and Purification of the Fab Fragment of A.4.6.1 humanized</i>: The 34B8 strain of <i>E. coli</i>, a strain no suppressor, was transformed with phagemid pMB4-19 or variants thereof. Simple colonies were grown during one night at 37 ° C in 2 ml of 2YT containing 50 µg / ml of carbenicillin These cultures were diluted in 200 ml of AP5 medium (Chang <i>et al. Gene</i> 55: 189-196 (1987)) that contained 20 µg / ml carbenicillin and incubated for 26 hours at 30 ° C. The cells were sedimented at 4000 xg and frozen at -20 ° C for at least 2 hours. Then the resuspended cell pellets in 5 ml of 10 mM Tris-HCl (pH 7.6) containing 1 mM EDTA, stirred at 4 ° C for 90 minutes and They were centrifuged at 10,000 xg for 15 minutes. The supernatant is applied to a protein G column of 1 ml streptococcus-Sepharose (Pharmacia) and se washed with 10 ml of 10 mM MES (pH 5.5). The bound Fab fragment is eluted with 2.5 ml of 100 mM acetic acid and neutralized immediately with 0.75 ml of 1M Tris-HCl, pH 8.0. The buffer of Fab preparations was exchanged for PBS and concentrated using Centricon-30 concentrators (Amicon) Typical Fab yields were \1 mg / l of culture, after protein G purification. Samples of Purified Fabs were characterized by mass spectrometry by electrospray, and concentrations were determined by amino acid analysis

p0212<i>Construction of the Fab Phagemid Library Anti-VEGF</i>: The phagemid library A.4.6.1 humanized was constructed by site-directed mutagenesis of according to the method of Kunkel <i>et al., Methods Enzymol</i>. 204: 125-139 (1991). A derivative of pMB4-19 containing TAA stop triplets in the codons of VH 24, 37, 67 and 93 to use as the mold of mutagenesis (all sequence numbering according to Kabat<i>et al., supra</i>). The modification was to avoid the subsequent background contamination by wild type sequences. The codons set as target for randomization were 4 and 71 (light chain) and 24, 37, 67, 69, 71, 73, 75, 76, 78, 93 and 94 (heavy chain).

p0213To randomize heavy chain codons 67, 69, 71, 73, 75, 76, 78, 93, and 94 with an oligonucleotide single mutagenic, first two were pre-assembled 126-serum oligonucleotides from fragments 60 and 66-serum by assisted enzymatic ligation by mold. Specifically, 1.5 nmol of the 5'phosphorylated oligonucleotide 503-1 (5'-GAT TTC AAA CGT CGT<u>NYT</u> ACT <u>WTT</u>TCT AGA GAC AAC TCC AAA AAC ACA <u>BYT</u> TAC CTG CAG ATG AAC-3 '(SEQ ID No. 22)) or 503-2 (5'-GAT TTC AAA CGT CGT <u>NYT</u> ACT <u>WTT</u>TCT <u>TTA</u> GAC <u>ACC</u> CBT <u>GCA</u><u>AGC</u> HERE<u>BYT</u> TAC CTG CAG ATG AAC-3 '(SEQ ID No. 23)) with 1.5 nmol of 503-3 (5'-AGC CTG CGC GCT GAG GAC ACT GCC GTC TAT TAC TGT <u>DYA</u><u>ARG</u> TAC CCC CAC TAT TAT GGG-3 '(SEQ ID No. 24)) (codons randomized underlines; N = A / G / T / C; W = A / T; B = G / T / C; D = G / A / T; R = A / G; Y = C / T). Then, 1.5 nmol of the template oligonucleotide was added (5'-CTC AGC GCG CAG GCT GTT CAT CTG CAG GTA-3 '(SEQ ID No. 25)), with the sequence complementary to the 5 'end of 503-1 / 2 and the end 3 'of 503-3, to hybridize to each end of the binding union. Was added<i>Taq</i> ligase (ligase New England Biolabs) thermostable and buffer, and the mixture of reaction was subjected to 40 rounds of thermal cycling, (95 ° C for 1.25 minutes; 50 ° C for 5 minutes) to cycle the oligonucleotide mold between the bound and unbound links. The product of 126-serum oligonucleotides were purified on a gel of 6% urea / TBE polyacrylamide and extracted from polyacrylamide in buffer. The two 126-meter products are combined in an equal proportion, precipitated in ethanol and finally they were solubilized in 10 mM Tris-HCl, EDTA 1mM The oligonucleotide product 126-mérico was 504-01 marked.

p0214Randomization of flanking codons selected (V L, ​​4, 71, V H, 24, 37, 67, 69, 71, 73, 75, 76, 93, 94) was achieved in two stages. First, the randomization of V_ {L} preparing three additional derivatives of Modified pMB4-19 mold. The flanking codons 4 and 71 of the light chain were replaced individually or by pairs using the two mutagenic oligonucleotides 5'-GCT GAT ATC CAG <u>TTG</u> ACC CAG TCC CCG-3 '(SEQ ID No. 26) and 5'-TCT GGG ACG GAT <u>TAC</u> ACT CTG ACC ATC-3 'SEC.ID.No. 27) .The mold containing deoxyuridine was prepared from each One of these new derivatives. Together with the original mold, you are four constructions encoded each of the four possible combinations of light chain flanking sequences (Table 5).

p0215Oligonucleotides were used 504-1, a mixture of two oligonucleotides 126-mérica (see above), and 5'-CGT TTG TCC TGT GCA <u>RYT</u> TCT GGC TAT ACC TTC ACC AAC TAT GGT ATG AAC TGG <u>RTC</u> CGT CAG GCC CCG GGT AAG-3 '(SEQ ID No. 28) to randomize codons flanks of the heavy chain using each of the molds just described. The four libraries were electroporated in XL-1 Blue cells from <i>E. coli</i> (Stratagene) and They combined. The total amount of independent transformants is estimated at> 1.2 x 10 8, approximately 1500 times greater than the maximum amount of DNA sequences in the library.

p0216A diversity of systems has been developed to functionally present antibody fragments in the surface of filamentous phages. Winter<i>et al., Ann. Rev. Immunol</i>. 12, 433 (1994). These include presenting the single chain Fab or Fv fragments (scFv) as fusions to cover proteins of gene III or bacteriophage gene VIII M13 The system selected in this document is similar to described by Garrard <i>et al., Biotechn</i>, 9, 1373 (1991) in the that a Fab fragment is presented monovalently as a fusion of gene III (Figure 7). This system has two remarkable features. In particular, unlike scFv, the Fab fragments have no tendency to form dimeric species, the presence of which can prevent the selection of Stronger binders due to the greed effects. Additionally, the monovalence of the protein presented eliminates a potential secondary source of greed effects that could be, otherwise, the result of the presence of multiple copies of a protein in each phagemid particle. Bass and Wells,<i>Protein</i> 8: 309 (1990) and Lowman <i>et al., Biochemistry</i> 30:10832 (1991).

p0217Phagemid particles that present the Fab fragments of humanized A.4.6.1 propagated in cells XL-1 Blue by <i>E. coli</i>. Briefly, the cells containing the randomized pMB4-19 construction They were grown overnight at 37 ° C in 25 ml of 2YT medium containing 50 µg / ml carbenicillin and approximately 10 10 auxiliary phages M13KO7 (Vieira &amp; Messing <i>Methods Enzymol</i>. 153: 3-11 (1987)). Stock solutions of phagemids were purified from the culture supernatants by precipitation with a solution of saline polyethylene glycol, and resuspended in 100 µl of PBS (sim10 14) phagemids / ml).

p0218<i>Selection of Fab Variants of A.4.6.1 Humanized</i>: The purified VEGF_ {121} (100 \ mul to 10 µg / ml in PBS) was coated in a plate well microtiter overnight at 4 ° C. The solution of coating was discarded and this well, in addition to a well not coated, they were blocked with 6% skim milk for 1 hour and they were washed with PBS containing 0.05% TWEEN 20 ™ (detergent). Then, 10 µl of phagemid stock solution was added, diluted to 100 µl with 20 mM Tris (pH 7.5) containing BSA at 0.1% and 0.05% TWEEN 20 ™ to each well. After 2 hours it the wells were washed and the bound phage eluted with 100 µl of 0.1 M glycine (pH 2.0), and neutralized with 25 µl of 1 M Tris pH 8.0. An aliquot of this was used to title the amount of eluted phage. The remaining phage eluted from the well coated with VEGF was propagated for use in the next selection cycle. He they made a total of 8 rounds of selection after what They selected 20 individual clones and sequenced (Sanger<i>et al., Proc. Natl. Acad. Sci. USA</i>. 74:5463-5467(1977)).

p0219<i>Determination of union affinities a VEGF</i>: Association speed constants (k_ {on}) and of dissociation (k_ {off}) for the union of Fab variants of A.4.6.1 humanized to VEGF_ 121 were measured by resonance of superficial plasmon (Karlsson <i>et al., J. Immun. Methods</i>145: 229-240 (1991)) in a Pharmacia instrument BIAcore VEGF_ 121 was immobilized covalently in the Biosensor chip by primary amino groups. The union of Fab variants of humanized A.4.6.1 were measured by solutions Fab fluids in 0.05% PBS / TWEEN 20 ™ on the chip at a flow rate of 20 µl / min. After each measure of binding, the residual Fab of the immobilized ligand was removed by washing with 5 µl of 50 mM aqueous HCl at 3 µl / min. The profiles of binding were analyzed by nonlinear regression using a model of single monovalent junction (BIAevaluation v2.0 program; Pharmacia).

Results

p0220<i>Construction of A.4.6.1 Humanized</i>: He constructed a Fab fragment of humanized A.4.6.1 (hu2.0, Figures 5A and 5B), in which the CDRs of A.4.6.1 are grafted into a region human flank V_ {L} \ kap-VH {III}. All other residues in hu2.0 were kept as the sequence human The binding of this variant to VEGF was so weak as to Be undetectable. Based on the relative affinity of other variants of A.4.6.1 humanized weak junction, the K_ for the binding of hu2.0 was estimated at> 7 µM. This contrasts with an affinity of 1.6 nM for a chimeric Fab construction consisting of the domains V_ {L} and V_ {H} intact as of A.4.6.1 murine and the constant human domains Therefore the union of hu2.0 to VEGF was at least 4000 times reduced in relation to the chimera.

p0221<i>Antibody Library Design</i>: He group of changes of the flanking region in the sequence human flanking in this document is shown in Table 5 and in Figure 6.

<pre listing-type="other">\ vskip1.000000 \ baselineskip</pre>

TABLE 5

Key Flanking Remains Important for the Union of Antigen and Marked for Randomization

<figref>7</figref>

<pre listing-type="other">\ dotable {\ tabskip \ tabcolsep # \ hfil \ tabskip0ptplus1fil \ dddarstrut \ cr} { a) Amino acid diversity in phagemid library. \ cr \ begin {minipage} {150mm} ^ b V H were randomized 71, 73, 75, 76 to produce the complete murine VH III tetrad (L71 / T73 / A75 / S76) or complete human (R71 / D73 / K75 / N76). \ End {minipage} \ cr}</pre>

p0222A concern in the design of the library of phagemids of humanized A.4.6.1 was that the remains fixed as objective for randomization were widely distributed along the sequences V_ {L} and V_ {H}. The limitations in length of synthetic oligonucleotides requires that the simultaneous randomization of each of these positions flanking can only be achieved through the use of multiple oligonucleotides. However, as the total amount of oligonucleotides increases, the effectiveness of mutagenesis decreases (that is, the proportion of mutants obtained that incorporate the sequence obtained from all mutagenic oligonucleotides). For avoid this problem, two elements were incorporated into the Library construction. The first was to prepare four molds of different mutagenesis that encode each of the possible flanking combinations of V L. This was simple to do given the limited diversity of the flanking region of the light chain (only 4 different sequences), but it was beneficial in that eliminated the need for two oligonucleotides from the strategy of mutagenesis The second was that they were pre-assembled two 126 base oligonucleotides from synthetic fragments More smalls. This made possible the randomization of codons 67, 69, 71, 73, 75, 76, 93 and 94 of VH with a single oligonucleotide long, instead of the two smaller ones. Therefore, the final randomization mutagenesis strategy employed only two oligonucleotides simultaneously in four molds different.

p0223<i>Selection of Fab from A.4.6.1 Humanized from Strong Union</i>: Fab phagemid library variants of A.4.6.1 humanized was selected based on VEGF binding. He enrichment of the functional phagemid measured by comparison of phage titres eluted from a microtiter plate well coated with VEGF versus eluted from a well of the plate uncoated microtiter, increased to the seventh round of affinity framing. After an additional round of classification, 20 clones were sequenced to identify the remains Preferred flanks selected in each randomized position. These results, summarized in Table 6, show a strong consensus among all selected clones. Ten of the twenty clones had the identical DNA sequence, called hu2.10. From the thirteen random flanking positions were selected eight substitutions in hu2.10 (V L, ​​71; V H, 37, 71, 73, 75, 76, 78 and 94). It is interesting to note that the remains V_ {H} 37 (Ile) and 78 (Val) were not selected in the sequence V_ {III} human or in the murine sequence A.4.6.1. This result suggests that some flanking positions can benefit from the increase of diversity beyond human flanking sequences and target parental murine.

TABLE 6

Selected Sequences of the Fab Library Humanized phagemids A.4.6.1

<figref>8</figref>

p0224The differences between hu2.0 and A.4.6.1 murine are underlined. The number of identical clones that identify each sequence selected by phage indicated in parenthesis. The stripes in the sequences of the selected clones per phage indicate the selection of the flanking sequence V_ {L} \ kappaI-V_ {H} III (ie, as in hu2.0).

p0225There were four unique amino acid sequences different among the ten remaining clones analyzed: hu2.1, hu2.2, hu2.6 and hu2.7. All these clones, in addition to hu2.10, contained identical substitutions in the flanking region in positions 37 (Ile), 78 (Val) and 94 (Lys) of V_ {H}, but kept the human VH III consensus sequence in positions 24 and 93. Four clones had lost the chain coding sequence lightweight and did not bind VEGF when tested in an ELISA test with phages (Cunningham <i>et al. EMBO J</i>. 13: 2508-251 (1994)). Such artifacts often can be minimized by reducing the number of classification cycles or propagating libraries in solid media.

p0226<i>Expression and Affinity of Variant Union of A.4.6.1 Humanized</i>: Variants selected by phage hu2.1, hu2.2, hu2.6, hu2.7 and hu2.10 were expressed in <i>E. coli</i> using shake flasks and Fab fragments were purified from the periplasmic extracts by protein affinity chromatography. The recovered Fab yields for these five clones varied from 0.2 (hu2.6) to 1.7 mg / l (hu2.1). The affinity of each of these Antigen variants (VEGF) was measured by plasmon resonance surface in a BIAcore instrument (Table 7). The analysis of this binding data shows that the consensus clone hu2.10 possessed the higher affinity for VEGF among the five variants tested. By therefore, the Fab phagemid library was selectively enriched for the strongest binding clone. The K_ {D} calculated for hu2.10 it was 55 nM, at least 125 times stronger than for hu2.0 that didn't It contains changes in the flanking region (K_ {D}> 7 µM). The four other selected variants all showed one more union weak to VEGF, varying below a K_ {D} of 360 nM for the weaker (hu2.7). It is interesting to note that the K_ {D} for hu2.6, 67 nM, was only slightly lower than that of hu2.10 and In addition, only one copy of this clone was found among the 20 clones sequenced. This may be due to the low level of expression and presentation, as was the case when the soluble Fab of This variant However, despite the proportion of expression lower, this variant is useful as a humanized antibody.

TABLE 7

VEGF binding affinity of Fab variants A.4.6.1 humanized

<figref>10</figref>

<tables><table><tgroup cols="1"><tbody><row><entry>* hu2.10V = hu2.10 with the mutation V_ {L} Leu \ rightarrow Val</entry></row><row><entry>The values Estimates in Biacore binding measurements are +/- 25%</entry></row><row><entry>** Too weak to measure; estimate lower link</entry></row></tbody></tgroup></table></tables>

p0227<i>Additional Improvement of the hu2.1 Variant Humanized</i>: Despite the great improvement in the affinity of antigen on the initial humanized variant, the union of hu2.10 to VEGF still it was 35 times weaker than a chimeric Fab fragment that contained the V_ {L} and V_ {H} domains of murine A.4.6.1. This difference considerable suggested that additional optimization of the region humanized flanking might be possible through mutations additional. Of the Vernier remains identified by Foote &amp; Winter <i>J. Mol. Biol</i>. 224: 487-499 (1992), only residues V L 46, V H 2 and V H 48 differ in the flanking region of A.4.6.11 versus the flanking region Human V L γ-V H III (Figures 5A and 5B) but they were not randomized in the phagemid library. A Molecular model of the Fv fragment of humanized A.4.6.1 showed that V_ {L} 46 is located on the contact surface V_ {L} -V_ {H} and could influence the conformation of CDR-H3. In addition, this amino acid is almost always leucine in most flanking regions V_ {L} \ kappa (Kabat <i>et al., supra</i>), but it is valine in A.4.6.1. By consequently, a Leu \ rightarrowVal substitution was made in this position in the background of hu2.10. The kinetic analysis of union for this new variant, hu2.10V, indicated an additional improvement 6 times in the K_D for VEGF binding, demonstrating the importance of valine at position VL 46 in the antibody A.4.6.1. The K_ {d} for hu2.10V (9.3 nM) was therefore 6 times the of the chimera. In contrast to V L 46, no improvement was observed in the binding affinity of hu2.10 for the replacement of V_ {2} or V_ {H} 48 with the corresponding remainder of murine A.4.6.1.

Example 3

p0228In this example, CDR randomization was used, affinity maturation by monovalent presentation in Fab phages, and cumulative combination of mutations to enhance the affinity of a humanized anti-VEGF antibody.

p0229<i>Construction of Antibody pY0101 Humanized</i>: The antibody vector presented by phage phMB4-19-1.6 (see figures 8A-E) was used as parental. In this construction, the anti-VEGF function is expressed as a Fab fragment with its heavy chain fused to the g3p N-terminal truncated. Both heavy and light chains are under the phoA promoter control with an upstream stII signal sequence for secretion in the periplasm. Punctual mutations were made outside the CDR regions by site-directed mutagenesis for improve affinity for VEGF with oligonucleotides HL-242, HL-243, HL-245, HL-246, HL-254, HL-256, and HL-257 as shown in Table 8 a continuation:

TABLE 8

Oligos for Directed Mutations

<figref>11</figref>

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p0230The resulting variant was called Y0101 (Figures 9A and 9B).

p0231<i>Construction of the First Generation of Antibody-Phage Libraries</i>: To avoid contamination by wild type sequence, molds were prepared with the TAA stop codon at the sites set for randomization and were used to build libraries by site-directed mutagenesis with oligonucleotides that use the degenerate NNS codon (where N is an equal mixture of A, G, C, and T while S is an equal mixture of G and C) for mutagenesis by saturation. VL1 and VH1 were chosen as potential candidates for affinity enhancement (Figures 9A and 9B). In the CDR, it they built two libraries from the pY0101 mold. VL1 was mutilated using mold stop oligonucleotides HL-248 and HL-249 (Table 9) and the HL-258 library oligonucleotides and HL-259 (Table 10). Similarly, it they built three libraries for VH3 using oligonucleotides of the stop mold HL-250, HL-251, and HL-252 (Table 9), and the oligonucleotides of library HL-260, HL-261, and HL-262 (Table 10). Library building is summarize in Tables 9 and 10 below.

TABLE 9

Mold oligos for mutagenesis

<figref>12</figref>

TABLE 10

Random Oligos for Construction of Libraries

<figref>13</figref>

p0232The products of mutagenesis reactions randomized electroporated in XL1-Blue cells of<i>E. coli</i> (Stratagene) and were amplified making them grow 15-16 hours with the auxiliary phage M13KO7. The complexity of each library, which varies from 2 x 10 7 to 1.5 x 10 8, was estimated based on the sowing of the transformation initial in carbenicillin plates.

p0233<i>Initial Affinity Selections</i>: For each selection round, they were explored approximately 10 9 -10 10 phage for plate binding (Nunc Maxisorp 96 wells) coated with 2 µg / ml of VEGF (recombinant; residue version 9-109) in 50 mM carbonate buffer, pH 9.6, and blocked with milk powder 5% in 50 mM carbonate buffer, pH 9.6. After a union of 1-2 hours at room temperature, in the presence of 0.5% bovine serum albumin and 0.05% TWEEN 20 ™ in PBS, He removed the phage solution, and the plate was washed ten times with PBS / TWEEN ™ (TWEEN 20 ™ 0.05% in PBS buffer). Typically for select enhanced affinity variants with speeds of slow dissociation, plates were incubated with PBS / TWEEN ™ buffer over a period of time that progressively lengthened for each selection round (from 0 minutes for the first round, to 3 hours for the ninth round of selection). After removing the buffer PBS / TWEEN ™, the remaining phages were eluted with 0.1 M HCl and they neutralized immediately with 1/3 volumes of 1M Tris, pH 8.0. Eluted phages spread by infecting cells XL1-Blue from <i>E. coli</i> (Stratagene) for the Next selection cycle.

p0234Sequencing data showed that both VL1 libraries, even after the eighth / ninth round of classification, remained varied, tolerating various types of remains at randomization sites. In contrast, VH3 libraries they kept only the wild type remains or had Very conservative substitutions. This suggested that VL1 was more exposed to the solvent and fell off the contact surface of Union. In contrast, VH3 showed no chain substitutions. drastically different sides and therefore could be more intimately involved in antigen binding.

p0235<i>ELISA Test of Affinity Phages of Union</i>: For each of these libraries, clones were tested representative (those represented by abundant sequences) to their affinities in relation to the parental clone pY0101 in one trial Phage ELISA. In that test, the phages are first diluted in series to determine a fractional saturation titer that after remained constant and was used to incubate with concentrations VEGF varied (starting at 200 nM at 0 nM) in solution. Then the mixture was transferred on a pre-coated plate with VEGF (2 µg / ml) and blocked with 5% milk powder, and allowed to equilibrate for 1 hour at room temperature. Subsequently, the phage solution was removed and the phages joined remaining were detected with a rabbit antibody solution anti-phage mixed with a goat conjugate anti-rabbit radish peroxidase rabbit. After one hour of incubation at room temperature, the plate was revealed with a chromogenic substrate, <i>or</i>-phenylenediamine (Sigma). The reaction was stopped with the addition of ½ volume of H 2 SO 4 2.5 M. The optical density was measured at 492 nm in a spectrophotometric plate reader.

p0236Although all the selected clones of these five libraries showed similar or weaker affinities than those of the wild type pY0101 in the phage ELISA test, a particular variant (pY0192) of the library HL-258 showed an apparent advantage (approximately 10 times) at the level of expression or presentation of the phage in relationship with pY0101. This clone contained mutations S24R, S26N, Q27E, D28Q, and I29L in the VL region (Figure 9A). In addition, it was discovered that This variant had a false mutation, M34I, in VH. This variant showed no significant differences in affinity for binding to VEGF compared to the pY0101 variant. To improve the level of Fab presentation in the phage, and the proportion signal-noise for phage ELISA assays, it incorporated the corresponding substitutions in pY0192 in VL1 in the antecedent mold to build both CDR Ala-mutants as the second generation of anti-VEGF libraries.

p0237<i>Wing of the CDR's Wing Anti-VEGF</i>: To determine the energy with the which contributes each of the amino acids in the CDR regions and by so much better select the target remains for randomization, the CDR regions were explored replacing alanine for each remainder. Each Ala mutant was constructed using directed mutagenesis of site with a synthetic oligonucleotide encoding the substitution of specific alanine. When Ala was the rest of the wild type, Ser was substituted to test the effect of a substitution of side chain The phage clones that had an Ala mutation single were purified and tested in phage ELISA as has been previously described. The results of the Ala scan showed that a replacement of Ala in various positions can have an effect, which varies from reductions of 2 to> 150 times, in antigen binding affinity compared to pY0192. Further, a previous observation was confirmed that VH3, but not VL1, is involved in antigen binding. The scan results CDR wing are summarized in Table 11 below.

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(Table goes to page following)

TABLE 11

Relative VEGF affinities of Fab variants of wing scan

<figref>16</figref>

<figref>17</figref>

p0238All variants are in the antecedent of pY0192 ("wt"; see Figures 9A-B). IC50 se determined in a competitive phage ELISA assay.

p0239The greatest effects of Ala substitutions they are seen in CDR H1, H2, and H3, including Y27A (reduction of 34 times affinity), N31A, Y32A, W50A, N52A, Y99A, S106A and W108A (each reduction of> 150 times); N35A (62-fold reduction), P100A (38-fold reduction) and F110A (25-fold reduction). In contrast, only one solution in VL had a big impact on the binding affinity, W96A (reduction of> 150 times). These results point to the three HRC CDRs as the determinants main energy sources of the union of Fab to VEGF, with some contribution of VL3.

p0240<i>Design of the CDR Mutation Libraries Second Generation</i>: Two additional libraries were designed which were randomized in the remains existing in version Y0192 anti-VEGF, based on the inspection of the structure crystalline In VH2, the remains were randomized 52-55 because they fall on the contact surface of union with VEGF. It was also set as a target for randomization an additional region of the Fab, called "CDR7" (see Figure 10B), because several remains in this loop, which do not contact VEGF, they have contacts with the VH loops of the antibody. These represent potential sites for affinity improvement through effects secondary in the remains of the contact surface. He Randomized remains L72, T74, and S77 in this library CDR7.

p0241Also based on the crystalline structure, it rebuilt one of the original CDR libraries to re-test the potential for maturation of affinity in the CDR of VH1. Remains 27, 28, and 30-32 using the new history of Y0192.

p0242<i>Second Generation Selections Anti-VEGF libraries</i>: based on the results of exploration of Ala as well as the crystalline structure of the antigen-antibody complex (F (ab) -12), a total of Seventeen libraries using the pY0192 template and oligonucleotides stop mold (encoding a stop codon at sites set as target for randomization) YC-80, YC-100, YC-102, HL-263, and HL-264 (Table 9 above). The corresponding randomization oligonucleotides (which employ NNS at the sites set as a target for randomization) were YC81, YC-101, YC-103, HL-265, and HL-266 (Table 10 previous). The resulting transformants produced libraries with complexities that vary from 6 x 10 7 to 5 x 10 suggesting that the libraries are complete in covering all possible variants. Phage libraries are classified for 7-8 rounds using conditions described in the Table 12 below.

TABLE 12

Conditions for Secondary Variant Selections from Fab

<figref>18</figref>

p0243ELISA buffer contained bovine serum albumin 0.5% and TWEEN 20 ™ 0.05% in PBS. VEGF was included in the buffer of incubation to minimize the re-union of VEGF phages coated on the plate surface. The classification of these libraries produced phage enrichments for 7 to 8 rounds of selection.

p0244<i>ELISA phage assays in Second Clones Generation</i>: After eight rounds of selection, they were isolated from ten to twenty clones of each plate library containing carbenicillin that harbored colonies of <i>E. coli</i>. (XL1) that they had been infected with a combination of eluted phages. The colonies were isolated and grown with auxiliary phage to get single stranded DNA to sequence it. Substitutions in CDR selected for the most favorable VEGF binding, deduced from the AND sequences of the phagemid clones. He shows a sampling of the clones selected in Table 13 a continuation.

TABLE 13

Protein sequences of variants Second Fab phage library anti-VEGF generation

<figref>20</figref>

<figref>21</figref>

<figref>22</figref>

p0245The sequence of the randomized region is only Sample deduced from DNA sequencing.

p0246When several clones were tested along with the parental clone pY10192 in phage ELISA assay, none showed a distinctive improvement over the parental clone. This could be explained by the time in scale in which the trial was performed (&lt;3 hours).

p0247To quantify the improvement in antigen binding on the parental clone, several DNAs of the variants were transformed anti-VEGF in strain 34B8 of <i>E. coli</i>, expressed as Fab, and purified by passing the download periplasmic through a G protein column (Pharmacia) as described in Example 2 above.

p0248<i>CDR Combination Variants</i>: For further enhance the affinity of VEGF binding, mutations were combined found in the phage presentation in different CDRs to create multiple CDR mutants. In particular, the mutations were combined identified in the phage variants of the VH1, VH2 libraries, and improved affinity VH3 (Table 14) to test the avidity of your contributions to union affinity.

TABLE 14

Combination Anti-VEGF variants CDR

<figref>23</figref>

p0249Mutations of parental vectors indicated were combined with those of the indicated oligonucleotides by site-directed mutagenesis to produce variants of combination listed.

p0250Version Y0317 is equivalent to Y0313-1 except that the background mutation in VL1 he retired and his sequence reverted to that of Y0101. The effects of mutate H101Y and S105T were tested by constructing a mutant of reversion from Y0238-3.

p0251<i>BIAcore analysis</i>: The affinities of union a VEGF of the Fab fragments were calculated from the association and dissociation rate constants measured using a surface plasmon resonance system BIAcore-2000 ™ (BIAcore, Inc., Piscataway, NJ). He activated a biosensor chip for VEGF covalent coupling using hydrochloride N-ethyl-N '- (3-dimethylaminopropyl) -carbodiimide (EDC) and N-hydroxysuccinimide (NHS) according to Provider instructions (BIAcore, Inc., Piscataway, NJ). He exchanged the VEGF buffer for 20 mM sodium acetate, pH 4.8, and diluted to approximately 50 µg / ml. An aliquot was injected (35 \ mul) at a flow rate of 2 \ mul / min to get approximately 700-1400 response units (RU) of coupled protein. Finally, 1 M ethanolamine was injected as blocking agent

p0252For kinetic measurements, they were injected Serb twice dilutions of Fab in PBS / TWEEN ™ buffer (TWEEN 0.05% 20 ™ in phosphate buffered saline) and 25 ° C at flow rate of 10 µl / min. Association speeds and dissociation were calculated using conventional protocols (Karlsson<i>et al. J. Immun. Methods</i> 145: 229-240 (1991)). Dissociation equilibrium constants, Kd, ​​a from surface plasmon resonance measurements (SPR) like koff / kon. The data is shown in Table 15 a continuation.

<figref>24</figref>

<pre listing-type="other">\ dotable {\ tabskip \ tabcolsep # \ hfil \ tabskip0ptplus1fil \ dddarstrut \ cr} { \ begin {minipage} {150mm} * The dissociation rate observed probably reflects an upper limit for the speed of actual dissociation in these experiments, although the speed of dissociation approaches the limit of detection by BIAcore. \ End {minipage} \ cr}</pre>

p0253The BIAcore ™ data in Table 15 shows that in Several variants have an improved affinity over Y0192. By example, a variant of CDRH1, Y0243-1, showed a affinity enhanced 4.1 times, which arises from T28D mutations and N31H The Y0238-3 variant showed at least one improvement of 14 times in the affinity of union on Y0192. Both mutations of CDRH3 contribute to the improved affinity of Y0238-3 because the reversion of T105 to S (variant Y0313-3) reduces the affinity of Y0238-3 from 0.15 nM to 0.65 nM (see Table 15). Enhancement of higher affinity in relation to Y0192 was seen for Y0313-1, which contained mutations in CDRH3 combined with mutations in CDRH1.

p0254<i>Inhibition Cell Based Assay VEGF</i>: Several versions of antibody A.4.6.1 were tested anti-VEGF for its ability to antagonize VEGF (recombinant; version 1-165) in the induction of HuVEC growth (endothelial cells of the umbilical vein human). 96-well plates were seeded with 1000 HuVEC per well and left in test medium (F12: DMEM 50:50 supplemented with 1.5% diafiltered fetal bovine serum) for 24 hours. The VEGF concentration used to induce cells was determined first titrating the amount of VEGF that can stimulate 80% of the maximum synthesis of DNA. Then new test medium was added containing fixed amounts of VEGF (final concentration 0.2 nM), and increasing concentrations of Fab or Mab anti-VEGF After 40 hours of incubation, it is measured DNA synthesis by incorporation of tritiated thymidine. Cells were pulsed with 0.5 µCi per well of [3H] -thymidine for 24 hours and were collected for count them, using a gamma counter TopCount.

p0255The results (Figure 11) show that the shape of full length IgG of F (ab) -12 was significantly more potent in VEGF inhibition activity than the Fab form (here, Y0192 was used). However, both variants Y0238-3 and Y0313-1 showed a VEGF inhibition activity even more potent than that of Fab's Y0192 or Mab of F (ab) -12. Comparing the Fab forms, the Y0313-1 variant seemed> 30 more powerful than the wild type Fab. It should be noted that the amount of VEGF (0.2 nM) used in this test is potentially limiting for the determination of an exact IC50 for the mutant. For example, if the binding affinity (Kd) of the mutant is in fact <0.2 nM, the IC50 in this experiment will be greater than in VEGF lower concentration conditions. Therefore the result maintains the conclusion that the affinity variant enhanced has at least a 30-fold improvement in affinity for VEGF, and that effectively blocks the activity of VEGF <i>in vitro</i>. How variant Y0317 differs from Y0313-1 only in the reversal of the VL1 sequence at wild type (Figure 10A), it is predicted that Y0317 will have a activity similar to Y0313-1.

p0256Variant Y0317 (Fab) and the Humanized F (ab) -12 variant of Example 1 (full length and Fab) for its ability to inhibit proliferation of capillary endothelial cells in response to a almost maximum efficiency concentration of VEGF using the assay described in Example 1. As illustrated in Figure 12, Y0317 was markedly more effective in inhibiting the proliferation of Bovine capillary endothelial cells that forms length complete and Fab of F (ab) -12 in this trial. He Y0317 mature affinity Fab showed an ED50 value in this trial which was at least about 20 times lower than the Fab of F (ab) -12.

(1) INFORMACIÓN GENERAL:

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<dl><dt>(i)</dt><dd>SOLICITANTE:</dd></dl>

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<dl><dt>(A)</dt><dd>NOMBRE: Genentech, Inc.</dd></dl>

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<dl><dt>(B)</dt><dd>CALLE: One DNA Way</dd></dl>

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<dl><dt>(C)</dt><dd>CIUDAD: South San Francisco</dd></dl>

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<dl><dt>(E)</dt><dd>PAÍS: EEUU</dd></dl>

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<dl><dt>(F)</dt><dd>CÓDIGO POSTAL (ZIP): CA 94080</dd></dl>

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<dl><dt>(ii)</dt><dd>TÍTULO DE LA INVENCIÓN: Anticuerpos Anti-VEGF</dd></dl>

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<dl><dt>(iii)</dt><dd>NÚMERO DE SECUENCIAS: 130</dd></dl>

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<dl><dt>(iv)</dt><dd>FORMA LEGIBLE POR ORDENADOR:</dd></dl>

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<dl><dt>(A)</dt><dd>TIPO DE MEDIO: Disco flexible</dd></dl>

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<dl><dt>(B)</dt><dd>ORDENADOR: PC IBM compatible</dd></dl>

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<dl><dt>(C)</dt><dd>SISTEMA OPERATIVO: PC-DOS/MS-DOS</dd></dl>

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<dl><dt>(E)</dt><dd>PROGRAMA: PatentIn Release Nº 1.0, Versión Nº 1.30</dd></dl>

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<dl><dt>(v)</dt><dd>DATOS DE LA SOLICITUD ACTUAL:</dd></dl>

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<dl><dt /><dd>NÚMERO DE SOLICITUD: EP 03004199.0</dd></dl>

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<dl><dt>(vi)</dt><dd>DATOS DE LA SOLICITUD ANTERIOR:</dd></dl>

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<dl><dt>(A)</dt><dd>NÚMERO DE SOLICITUD: PCT/US98/06604</dd></dl>

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<dl><dt>(B)</dt><dd>FECHA DE PRESENTACIÓN: 03-ABR-1998</dd></dl>

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<dl><dt>(vi)</dt><dd>DATOS DE LA SOLICITUD ANTERIOR:</dd></dl>

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<dl><dt>(A)</dt><dd>NÚMERO DE SOLICITUD: US 08/833,504</dd></dl>

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<dl><dt>(B)</dt><dd>FECHA DE PRESENTACIÓN: 07-ABR-1997</dd></dl>

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<dl><dt>(vi)</dt><dd>DATOS DE LA SOLICITUD ANTERIOR:</dd></dl>

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<dl><dt>(A)</dt><dd>NÚMERO DE SOLICITUD: US 08/908,469</dd></dl>

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<dl><dt>(B)</dt><dd>FECHA DE PRESENTACIÓN: 06-AGO-1997</dd></dl>

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(2) INFORMACIÓN PARA LA SEC ID Nº: 1:

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<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

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<dl><dt>(A)</dt><dd>LONGITUD: 10 aminoácidos</dd></dl>

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<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

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<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

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<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

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<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

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<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 1:</dd></dl>

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\sa{Gly Tyr Thr Phe Thr Asn Tyr Gly Met Asn}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 2:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 17 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 2:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Trp Ile Asn Thr Tyr Thr Gly Glu Pro Thr Tyr Ala Ala Asp Phe Lys}

\sac{Arg}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 3:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 14 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 3:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro His Tyr Tyr Gly Ser Ser His Trp Tyr Phe Asp Val}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 4:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 11 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 4:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Ser Ala Ser Gln Asp Ile Ser Asn Tyr Leu Asn}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 5:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 7 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 5:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Phe Thr Ser Ser Leu His Ser}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 6:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 9 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 6:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Gln Gln Tyr Ser Thr Val Pro Trp Thr}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 7:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 123 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 7:</dd></dl>

<figref>26</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 8:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 108 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 8:</dd></dl>

<figref>27</figref>

<figref>28</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 9:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 123 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 9:</dd></dl>

<figref>29</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 10:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 108 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 10:</dd></dl>

<figref>30</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 11:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 113 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 11:</dd></dl>

<figref>31</figref>

<pre listing-type="other">\newpage</pre>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 12:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 108 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 12:</dd></dl>

<figref>32</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 13:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 107 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 13:</dd></dl>

<figref>33</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 14:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 123 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 14:</dd></dl>

<figref>35</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 15:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 107 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 15:</dd></dl>

<figref>36</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 16:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 123 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 16:</dd></dl>

<figref>37</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 17:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 11 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 17:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Pro Lys Asn Ser Ser Met Ile Ser Asn Thr Pro}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 18:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 8 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 18:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{His Gln Ser Leu Thr Gly Thr Gln}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 19

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 8 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 19:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{His Gln Asn Leu Ser Asp Gly Lys}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 20:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 8 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 20:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{His Gln Asn Ile Ser Asp Gly Lys}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 21:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 8 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 21:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Val Ile Ser Ser His Leu Gly Gln}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 22:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 67 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 22:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GATTTCAAAC GTCGTNYTAC TWTTTCTAGA GACAACTCCA AAAACACABY TTACCTGCAG <pre listing-type="other">\hfill</pre> 60

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>ATGAAAC <pre listing-type="other">\hfill</pre> 67

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 23:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 66 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 23:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GATTTCAAAC GTCGTNYTAC TWTTTCTTTA GACACCTCCG CAAGCACABY TTACCTGCAG <pre listing-type="other">\hfill</pre> 60

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>ATGAAC <pre listing-type="other">\hfill</pre> 66

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 24:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 60 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 24:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>AGCCTGCGCG CTGAGGACAC TGCCGTCTAT TACTGTDYAA RGTACCCCCA CTATTATGGG <pre listing-type="other">\hfill</pre> 60

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 25:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 30 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 25:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>CTCAGCGCGC AGGCTGTTCA TCTGCAGGTA <pre listing-type="other">\hfill</pre> 30

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 26:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 27 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 26:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GCTGATATCC AGTTGACCCA GTCCCCG <pre listing-type="other">\hfill</pre> 27

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 27:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 27 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 27:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>TCTGGGACGG ATTACACTCT GACCATC <pre listing-type="other">\hfill</pre> 27

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 28:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 75 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 28:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>CGTTTGTCCT GTGCARYTTC TGGCTATACC TTCACCAACT ATGGTATGAA CTGGRTCCGT <pre listing-type="other">\hfill</pre> 60

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>CAGGCCCCGG GTAAG <pre listing-type="other">\hfill</pre> 75

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 29:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 24 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 29:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GATATCCAGT TGACCCAGTC CCCG <pre listing-type="other">\hfill</pre> 24

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 30:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 21 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 30:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GCTCCGAAAG TACTGATTTA C <pre listing-type="other">\hfill</pre> 21

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 31:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 54 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 31:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>CGTCGTTTCA CTTTTTCTGC AGACACCTCC AGCAACACAG TATACCTGCA GATG <pre listing-type="other">\hfill</pre> 54

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 32:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 25 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 32:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>CTATTACTGT GCAAAGTACC CCCAC <pre listing-type="other">\hfill</pre> 25

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 33:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 24 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 33:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GGGACGGATT TCACTCTGAC CATC <pre listing-type="other">\hfill</pre> 24

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 34:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 26 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 34:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GGTATGAACT GGGTCCGTCA GGCCCC <pre listing-type="other">\hfill</pre> 26

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 35:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 57 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 35:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>CGTCGTTTCA CTTTTTCTTT AGACACCTCC AAAAGCACAG CATACCTGCA GATGAAC <pre listing-type="other">\hfill</pre> 57

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 36:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 53 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 36:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GGGTCACCAT CACCTGCTAA GCATAATAAT AATAAAGCAA CTATTTAAAC TGG <pre listing-type="other">\hfill</pre> 53

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 37:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 52 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 37:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GCGCAAGTCA GGATATTTAA TAATAATAAT AATGGTATCA ACAGAAACCA GG <pre listing-type="other">\hfill</pre> 52

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 38:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 48 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 38:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GTCTATTACT GTGCAAAGTA ATAACACTAA TAAGGGAGCA GCCACTGG <pre listing-type="other">\hfill</pre> 48

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 39:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 49 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 39:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GGTACCCCCA CTATTATTAA TAATAATAAT GGTATTTCGA CGTCTGGGG <pre listing-type="other">\hfill</pre> 49

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 40:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 53 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 40:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>CACTATTATG GGAGCAGCCA CTAATAATAA TAAGTCTGGG TCAAGGAACC CTG <pre listing-type="other">\hfill</pre> 53

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 41:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 53 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 41:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>TCCTGTGCAG CTTCTGGCTA ATAATTCTAA TAATAAGGTA TGAACTGGGT CCG <pre listing-type="other">\hfill</pre> 53

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 42:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 52 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 42:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GAATGGGTTG GATGGATTA CTAATAATAA GGTTAACCGA CCTATCGTGC GG <pre listing-type="other">\hfill</pre> 52

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 43:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 49 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 43:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>CTGTGCAAAG TACCCGTAAT ATTAATAATA ATAACACTGG TATTTCGAC <pre listing-type="other">\hfill</pre> 49

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 44:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 48 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 44:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>CGTTTCACTT TTTCTTAGA CTAATCCAAA TAAACAGCAT ACCTGCAG <pre listing-type="other">\hfill</pre> 48

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 45:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 46 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 45:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GAATGGGTTG GATGGATTTA ATAATAATAA GGTGAACCGA CCTATG <pre listing-type="other">\hfill</pre> 46

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 46:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 53 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 46:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GGGTCACCAT CACCTGCNNS GCANNSNNSN NSNNSAGCAA CTATTTAAAC TGG <pre listing-type="other">\hfill</pre> 53

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 47:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 52 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 47:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GCGCAAGTCA GGATATTNNS NNSNNSNNSN NSTGGTATCA ACAGAAACCA GG <pre listing-type="other">\hfill</pre> 52

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 48:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 48 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 48:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GTCTATTACT GTGCAAAGNN SNNSCACNNS NNSGGGAGCA GCCACTGG <pre listing-type="other">\hfill</pre> 48

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 49:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 49 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 49:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GGTACCCCCA CTATTATNNS NNSNNSNNST GGTATTTCGA CGTCTGGGG <pre listing-type="other">\hfill</pre> 49

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 50:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 54 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 50:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>CACTATTATG GGAGCAGCCA CNNSNNSNNS NNSGTCTGGG GTCAAGGAAC CCTG <pre listing-type="other">\hfill</pre> 54

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 51:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 53 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 51:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>TCCTGTGCAG CTTCTGGCNN SNNSTTCNNS NNSNNSGGTA TGAACTGGGT CCG <pre listing-type="other">\hfill</pre> 53

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 52:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 52 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 52:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GAATGGGTTG GATGGATTAA CNNSNNSNNS GGTNNSCCGA CCTATGCTGC GG <pre listing-type="other">\hfill</pre> 52

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 53:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 49 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 53:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>CTGTGCAAAG TACCCGNNST ATNNSNNSNN SNNSCACTGG TATTTCGAC <pre listing-type="other">\hfill</pre> 49

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 54:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 48 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 54:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>CGTTTCACTT TTTCTNNSGA CNNSTCCAAA NNSACAGCAT ACCTGCAG <pre listing-type="other">\hfill</pre> 48

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 55:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 47 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 55:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GAATGGGTTG GATGGATTNN NSNNSNNSNN SGGTGAACCG ACCTATG <pre listing-type="other">\hfill</pre> 47

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 56:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 56:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Arg Gly Thr Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 57:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 57:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Ile Asn Lys Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 58:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 58:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Tyr Gly Thr Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 59:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 59:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Tyr Asn Gly Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 60:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 60:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Ile Ala Lys Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 61:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 61:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Arg Asp Asn Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 62:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 62:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Trp Gly Thr Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 63:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 63:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Arg Gln Asn Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 64:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 64:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Arg Gln Ser Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 65:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 65:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Arg Asn Thr Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 66:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 66:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Lys Asn Thr Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 67:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 67:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Ile Glu Arg Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 68:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 68:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Arg Asn Ala Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 69:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 69:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Thr Thr Arg Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 70:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 70:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Glu Gly Thr Ser Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 71:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 71:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Arg Gln Arg Gly His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 72:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 72:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Thr Gly Arg Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 73:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 73:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Thr Asn Thr Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 74:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 74:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Arg Lys Gly Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 75:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 75:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Thr Gly Ser Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 76:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 76:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Arg Ser Gly Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 77:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 77:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Thr Asn Arg Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 78:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 78:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Arg Asn Ser Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 79:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 79:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Lys Glu Ser Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 80:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 80:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Arg Asp Ala Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 81:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 81:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Arg Gln Lys Gly His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 82:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 82:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Lys Gly Gly Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 83:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 12 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 83:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Gly Ala Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 84:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 84:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Arg Gly Glu Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 85:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 13 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 85:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Tyr Pro Tyr Tyr Arg Ser Thr Ser His Trp Tyr Phe Asp}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 86:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 10 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 86:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Gly Tyr Asp Phe Thr His Tyr Gly Met Asn}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 87:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 10 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 87:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Gly Tyr Glu Phe Gln His Tyr Gly Met Asn}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 88:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 10 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 88:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Gly Tyr Glu Phe Thr His Tyr Gly Met Asn}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 89:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 10 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 89:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Gly Tyr Asp Phe Gly His Tyr Gly Met Asn}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 90:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 10 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 90:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Gly Tyr Asp Phe Ser His Tyr Gly Met Asn}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 91:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 10 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 91:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Gly Tyr Glu Phe Ser His Tyr Gly Met Asn}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 92:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 10 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 92:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Phe Ser Val Asp Val Ser Lys Ser Thr Ala}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 93:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 10 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 93:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Phe Ser Leu Asp Lys Ser Lys Ser Thr Ala}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 94:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 10 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 94:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Phe Ser Leu Asp Val Trp Lys Ser Thr Ala}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 95:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 10 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 95:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

\sa{Phe Ser Ile Asp Lys Ser Lys Ser Thr Ala}

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 96:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 42 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 96:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GCAAAGTACC CGTACTATTA TGGGACGAGC CACTGGTATT TC <pre listing-type="other">\hfill</pre> 42

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 97:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 48 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 97:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>GTCACCATCA CCTGCAGCGC AAGTCAGGAT ATTAGCAAGT ATTTAAAC <pre listing-type="other">\hfill</pre> 48

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 98:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 33 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 98:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<pre listing-type="other">\vskip0.400000\baselineskip</pre>

<pre listing-type="other">\hskip-.1em\dddseqskip</pre>CCGTACTATT ATGGGAGCAG CCACTGGTAT TTC <pre listing-type="other">\hfill</pre> 33

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 99:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 6072 pares de bases</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: ácido nucleico</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: ADN</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ix)</dt><dd>CARACTERÍSTICAS:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>NOMBRE/CLAVE: CDS</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>LOCALIZACIÓN: 390..1101</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ix)</dt><dd>CARACTERÍSTICAS:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>NOMBRE/CLAVE: mat_péptido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>LOCALIZACIÓN: 459</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ix)</dt><dd>CARACTERÍSTICAS:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>NOMBRE/CLAVE: CDS</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>LOCALIZACIÓN: 1185..2423</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ix)</dt><dd>CARACTERÍSTICAS:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>NOMBRE/CLAVE: mat_péptido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>LOCALIZACIÓN: 1254</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 99:</dd></dl>

<figref>38</figref>

<figref>39</figref>

<figref>40</figref>

<figref>41</figref>

<figref>42</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 100:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 413 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 100:</dd></dl>

<figref>43</figref>

<figref>44</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 101:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 110 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 101:</dd></dl>

<figref>45</figref>

<pre listing-type="other">\newpage</pre>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 102:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 119 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 102:</dd></dl>

<figref>46</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 103:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 110 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 103:</dd></dl>

<figref>47</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 104:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 118 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 104:</dd></dl>

<figref>48</figref>

<pre listing-type="other">\newpage</pre>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 105:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 110 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 105:</dd></dl>

<figref>49</figref>

<figref>50</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 106:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 118 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 106:</dd></dl>

<figref>51</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 107:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 110 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 107:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<figref>52</figref>

<pre listing-type="other">\newpage</pre>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 108:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 118 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 108:</dd></dl>

<figref>53</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 109:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 110 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 109:</dd></dl>

<figref>54</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 110:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 118 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 110:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<figref>55</figref>

<pre listing-type="other">\newpage</pre>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 111:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 110 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 111:</dd></dl>

<figref>56</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 112:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 118 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 112:</dd></dl>

<figref>57</figref>

<figref>58</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 113:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 110 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 113:</dd></dl>

<figref>59</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 114:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 118 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 114:</dd></dl>

<figref>60</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 115:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 110 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 115:</dd></dl>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

<figref>61</figref>

<pre listing-type="other">\newpage</pre>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 116:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 118 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(xi)</dt><dd>DESCRIPCIÓN DE LA SECUENCIA: SEC ID Nº: 116:</dd></dl>

<figref>62</figref>

<pre listing-type="other">\vskip1.000000\baselineskip</pre>

(2) INFORMACIÓN PARA LA SEC ID Nº: 117:

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(i)</dt><dd>CARACTERÍSTICAS DE LA SECUENCIA:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>LONGITUD: 10 aminoácidos</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>TIPO: aminoácido</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(C)</dt><dd>CADENA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>TOPOLOGÍA: desconocida</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ii)</dt><dd>TIPO DE MOLÉCULA: proteína</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ix)</dt><dd>CARACTERÍSTICAS:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>NOMBRE/CLAVE: Sitio modificado</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>LOCALIZACIÓN: 3</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(D)</dt><dd>OTRA INFORMACIÓN: /nota= "Donde la X en la posición 3 representa Ácido aspártico, Treonina o Ácido glutámico".</dd></dl>

<pre listing-type="other">\vskip0.800000\baselineskip</pre>

<dl><dt>(ix)</dt><dd>CARACTERÍSTICAS:</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(A)</dt><dd>NOMBRE/CLAVE: Sitio modificado</dd></dl>

<pre listing-type="other">\vskip0.500000\baselineskip</pre>

<dl><dt>(B)</dt><dd>LOCALIZACIÓN: 4</dd></dl>