Acylated glucagon analogues

Acylated glucagon analogs {Acylated glucagon analogues}

The present invention relates to acylated glucagon analog of, e.g., their medical use in the treatment of obesity and diabetes.

Obesity and diabetes is increasing worldwide health problem that also a variety of diseases, especially cardiovascular diseases (CVD), obstructive sleep apnea, stroke, peripheral arterial disease, vascular complications are associated with smiling and osteoarthritis.

The people of 246 million with diabetes worldwide, and by 2025 380 000 002 are estimated to have diabetes. People are higher / more has additional cardiovascular risk factors, including LDL, and triglycerides and low HDL.

Cardiovascular disease represents the death rate about 50% of people with diabetes, obesity and the morbidity and mortality of diabetes emphasizes the medical need for effective treatment options.

Pre pro glucagon (preproglucagon) is associated with a plurality of structural pro glucagon-derived peptides, for example, glucagon (Glu), glucagon-like peptide -1 (GLP-1), glucagon-like peptide -2 (GLP-2), and auxins Sat module Lin (OXM) 158 amino acid precursor that is processed differently from the tissue to form a polypeptide. These molecules are a wide variety of physical function, for example, glucose homeostasis (homeostasis), insulin secretion, gastric emptying (gastric emptying) and chapter growth (intestinal growth) is associated with the regulation of food intake as well.

Glucagon-free (pre) - amino acid of the glucagon sequence corresponding to 53 to 81 and also Having a 29-amino acid peptide. Lin sat dioxin module (OXM) is an octa-peptide carboxy-terminal extension of 37 amino acid peptide that contains a complete amino acid sequence of glucagon 29 having a (pre-pro-glucagon 82 to 89 amino acids, the sequence Lys-rg-Asn-Arg- an Asn-Asn-Ile-Ala has "peptide via 1" or IP-1 to bulrium; complete sequence of human dioxin Sat module according Lin Major, biologically active fragments of GLP-1 is a free-30-amino acid corresponding to amino acid 98 to 127 of the pro glucagon, it is produced as a C- terminal amidated peptides.

Glucagon helps maintain the blood glucose level was not binding in the glucagon receptor on the blood cell (hepatocyte). This will cause the liver to release glucose via the glucose is stored in the glucose decomposition Gen Gen form. As these storage water is exhausted, glucagon synthesize additional glucose by the glucose to stimulate new liver. The glucose is released into the blood, to prevent the development of hypoglycemia. In addition, glucagon has been demonstrated to increase the lipolysis is also to reduce the weight.

GLP-1 can reduce the blood sugar level elevated to improve the glucose insulin secretion promoting and also promote weight loss primarily by reducing food intake.

The dioxin Saturday modules Lin (oxyntomodulin) are released into the blood in proportion to the calorie content of meals and response to food intake. Sat mechanism of action of auxin module Lynn has not been well understood. In particular, exclusively through the glucagon receptor and GLP-1 receptor, or through at least one yet unidentified receptors, the effect of the hormone is unknown whether or not they are affected.

Other peptide coupling to activate the glucagon and GLP-1 receptor, and (Hjort, etc., Journal of Biological Chemistry, 269, 30121-30124,1994) also shown to inhibit food intake and reduce weight gain (International Application Publication WO 2006 / 134340; WO 2007/100535; WO 2008/101017, WO 2008/152403, WO 2009/155257 and WO 2009/155258).

Stabilization of the peptide has been shown to provide a better pharmacokinetic profile for a number of drugs. In particular, addition of one or more polyethylene glycol (PEG) or an acyl group has been shown to extend the half-life of GLP-1 and other peptides as with a short peptide such as plasma stability.

The International Patent Publication WO 00 / 55184A1 and WO 00/55119 a wide range of peptides, in particular the method is acylation of GLP-1 technology. Madsen, etc. (J. Med Chem.. 2007, 50, 6126-6132), the acylated GPL-1 (Liraglutide) at position 20 are described and can also provide the data for its stability.

Chemical stabilization of PEG OXM (PEGlation) and C- terminal acylated also International Patent Publication WO2007 / 100535, WO08 / 071972 and Druce, MR, etc., Endocrinology 2009, 150 (4), the analog selected from the pharmacokinetic 1712-1721 It was found to improve the profile.

Recently, PEG screen of glucose analogues have a significant effect on the pharmacokinetic profile of the test compounds (International Patent Publication WO2008 / 101017) also appeared to interfere with the effect of these compounds.

<p num="0015">The present invention provides a compound having the formula.</p><p num="0016">R^One-ZR²</p><p num="0017">Wherein</p><p num="0018">R^OneIs H, C_One_-4 Alkyl, acetyl, formyl, benzoyl and acetyl, or trifluoromethyl,</p><p num="0019">R²Is OH or NH₂and;</p><p num="0020">In addition, Z is a peptide having the formula (I):</p><p num="0021">His-X2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-X12-Tyr-Leu-Asp-X16-X17-Ala-Ala-X20-X21-Phe-Val-X24-Trp-Leu-X27-X28-Ala-X30; (I)</p><p num="0022">here, </p><p num="0023">X2 is selected from Ser or Aib;</p><p num="0024">X12 is selected from Lys, Arg, and Leu;</p><p num="0025">X16 is selected from Arg and X;</p><p num="0026">X17 is selected from Arg and X;</p><p num="0027">X20 is selected from Arg, His, and X;</p><p num="0028">X21 is selected from Asp and Glu;</p><p num="0029">X24 is selected from Ala and X;</p><p num="0030">X27 is selected from Leu and X;</p><p num="0031">X28 is selected from Arg and X;</p><p num="0032">X30 is not present, or X;</p><p num="0033">Wherein X16, X17, X20, X24, X27, X28, X30 and at least one of X;</p><p num="0034">Wherein each X moiety is independently also Glu, Lys, Ser, Cys, Dbu, is selected from the group consisting of Dpr and Orn;</p><p num="0035">The side chains of at least one residue X is the formula (i) or (ii) coupled to a lipophilic substituent, and having:</p><p num="0036">(i) Z^One, Where Z^OneDirectly bonded to the side chain of X is the lipophilic portion; or</p><p num="0037">(ii) Z^OneZ², Where Z^OneIs a lipophilic region, Z²And Z is a spacer^OneThe Z²Coupled via the side chain of X is; </p><p num="0038">Z is a single HSQGTFTSDYSKYLDS-K (hexadecanoyl -γ-Glu) is not a -AAHDFVEWLLRA.</p><p num="0039">X30 is absent or may not be present. In these embodiments, when X30 is present, the Lys may be desirable.</p><p num="0040">In certain embodiments, any of the X residues, and in particular any residues X coupled to a lipophilic substituent is independently selected from Lys, Glu or Cys. </p><p num="0041">In certain embodiments, </p><p num="0042">X16 is selected from Glu, Lys, and Ser;</p><p num="0043">X17 is selected from Lys and Cys;</p><p num="0044">X20 is selected from His, Lys, Arg and Cys;</p><p num="0045">X24 is selected from Lys, Glu, and Ala;</p><p num="0046">X27 is selected from Leu and Lys; And / or</p><p num="0047">X28 is selected from Ser, Arg and Lys.</p><p num="0048">Particular combinations of the residues that may be present in the peptide of formula (I) include: </p><p num="0049">X2 is Aib and X17 is also a Lys;</p><p num="0050">X2 is Aib and X17 is also Cys;</p><p num="0051">X2 is Aib and X20 is also Cys;</p><p num="0052">X2 is Aib and X28 is also a Lys;</p><p num="0053">X12 is Arg and is also X17 is Lys;</p><p num="0054">X12 is Leu and is also X17 is Lys;</p><p num="0055">X12 is Lys and is also X20 is Lys;</p><p num="0056">X12 is Lys and is also X17 is Lys;</p><p num="0057">X16 is Lys and is also X17 is Lys;</p><p num="0058">X16 is Ser and is also X17 is Lys;</p><p num="0059">X17 is Lys is also the X20 is Lys;</p><p num="0060">X17 is Lys and is also X21 Asp;</p><p num="0061">X17 is Lys and is also X24 is Glu;</p><p num="0062">X17 is Lys and is also X27 Leu;</p><p num="0063">X17 is Lys is also the X27 is Lys;</p><p num="0064">X17 is Lys and is also X28 Ser;</p><p num="0065">X17 is Lys is also the X28 is Arg;</p><p num="0066">X20 is Lys and is also X27 Leu;</p><p num="0067">X21 is Asp and is also X27 Leu;</p><p num="0068">The X2 is Aib, X12 is Lys and is also X16 is Ser;</p><p num="0069">The X12 is Lys, X17 is Lys and is also X16 Ser;</p><p num="0070">The X12 is Arg, X17 is Lys is also the X16 is Glu;</p><p num="0071">X16 is Glu, X17 is Lys is also the X20 is Lys;</p><p num="0072">X16 is Ser, X21 is Asp and is also X24 is Glu;</p><p num="0073">X17 is Lys, X24 is Glu is also the X28 is Arg;</p><p num="0074">X17 is Lys, X24 is Glu is also the X28 is Lys;</p><p num="0075">X17 is Lys, X27 is Leu and is also X28 Ser;</p><p num="0076">X17 is Lys, X27 is Leu is also the X28 is Arg;</p><p num="0077">The X20 is Lys, X24 is Glu and is also X27 Leu;</p><p num="0078">And X20 is Lys, X27 is Leu and is also X28 Ser;</p><p num="0079">And X20 is Lys, X27 is Leu, and also the X28 is Arg;</p><p num="0080">X16 is Ser, X20 is His, X24 is Glu and is also X27 Leu;</p><p num="0081">X17 is Lys, and X20 is His, X24 is Glu, and X28 is Ser and also;</p><p num="0082">X17 is Lys, X20 is Lys, X24 is Glu and is also X27 Leu; or</p><p num="0083">X17 is Cys, and X20 is Lys, Glu, and X24 is also X27 is Leu.</p><p num="0084">Peptide may be preferable to contain only a single amino acid of the type that can be derivatized by addition of a lipophilic substituent of the formula (I). For example, the peptide has only one Lys residue to the lipophilic substituent bound to the residue, and can only contain a Cys residues, or only one of the Glu residue.</p><p num="0085">The compounds of the invention may have one or more intramolecular cross-linking (intramolecular bridge) in the peptide sequence of formula (I). Each of these cross-linking is typically between two amino acid residues of the side chains (i.e., between amino acids A and A + 4) of the formula (I) are separated by three amino acids in the linear amino acid sequence may be formed.</p><p num="0086">More specifically, the crosslinking may be formed between the pair of residues 16 and 20, 17 and 21, 20 and 24, or 24 and 28 of the side chain. Two side chains may be bonded to each other by a covalent bond or through an ionic interaction. Therefore these residues may comprise a pair of reverse charged side chain to form the salt cross-linked by ionic interactions. For example, while one of the moieties, which can be Glu or Asp, the others may be a Lys or Arg. A pair of Lys and Glu and Lys and Asp may also react to form a lactam ring.</p><p num="0087">Examples of suitable pairs of residues located at the 16 and 20 include:</p><p num="0088">X16 is Glu is also the X20 is Lys;</p><p num="0089">X16 is Glu is also the X20 is Arg;</p><p num="0090">X16 X20 is Lys and also is Glu; And</p><p num="0091">X16 is Arg is also the X20 is Glu.</p><p num="0092">Examples of suitable pairs of residues located at the 17 and 21 include:</p><p num="0093">X17 is Arg is also the X21 is Glu;</p><p num="0094">X17 X21 is Lys and also is Glu;</p><p num="0095">X17 is Arg is also the X21 is Asp; And</p><p num="0096">X17 X21 is Lys and also is Asp.</p><p num="0097">Examples of suitable pairs of residues located at the 20 and 24 include: </p><p num="0098">In addition, X24 and X20 is Glu is Lys;</p><p num="0099">In addition, X24 and X20 is Glu is Arg;</p><p num="0100">In addition, X24 and X20 is Lys is Glu; And</p><p num="0101">In addition, X24 and X20 is Arg is Glu.</p><p num="0102">Examples of suitable pairs of residues located at the 24 and 28 include:</p><p num="0103">X24 is Glu is also the X28 is Lys;</p><p num="0104">X24 is Glu is also the X28 is Arg;</p><p num="0105">In addition, X28 and X24 is Lys is Glu; And</p><p num="0106">In addition, X28 and X24 is Arg is Glu.</p><p num="0107">For example, Lys and Glu of mating to form a lactam ring can be particularly preferred, especially between positions 24 and 28. </p><p num="0108">Moiety contained in the crosslinking molecule will be apparent that there may be derivatised with lipophilic substituent. So, if the cross-linking moiety X contained in the molecule, at least one other moiety X is coupled to a lipophilic substituent or substituents.</p><p num="0109">But want to be limited to any particular theory, this intramolecular cross-linking to stabilize the alpha helix structure of the molecule and thus GLP-1 receptor and possibly also believed to increase the efficacy and / or selectivity at the glucagon receptor. </p><p num="0110">The compound may have the following formula: </p><p num="0111">R^One-ZR²</p><p num="0112">Wherein</p><p num="0113">R^OneIs H, C_One_-4Alkyl, acetyl, formyl, benzoyl or trifluoroacetyl, and;</p><p num="0114">R²Is OH or NH₂and; Also</p><p num="0115">Z is a peptide having the following general formula (IIa), and:</p><p num="0116">His-Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-X12-Tyr-Leu-Asp-X16-X17-Ala-Ala-X20-X21-Phe-Val-X24-Trp-Leu-Leu-X28-Ala; (IIa)</p><p num="0117">here</p><p num="0118">X12 is selected from Lys, Arg, and Leu;</p><p num="0119">X16 is selected from Ser and X;</p><p num="0120">X17 is X;</p><p num="0121">X20 is selected from His and X;</p><p num="0122">X21 is selected from Asp and Glu;</p><p num="0123">X24 is selected from Ala and Glu;</p><p num="0124">X28 is selected from Ser, Lys and Arg; Also</p><p num="0125">Wherein each X moiety is independently selected from Glu, Lys, and is selected from the group consisting of Cys;</p><p num="0126">Wherein X is a side chain of at least one moiety is coupled to a lipophilic substituent having the formula (i) or (ii):</p><p num="0127">(i) Z^One, Where Z^OneDirectly bonded to the side chain of X is the lipophilic portion; or</p><p num="0128">(ii) Z^OneZ², Where Z^OneIs a lipophilic region, Z²A spacer, and Z^OneThe Z²It is coupled to the side chain of X through. </p><p num="0129">Alternatively, the compound can have the following formula:</p><p num="0130">R^One-ZR²</p><p num="0131">Wherein</p><p num="0132">R^OneIs H, C_One_-4 Alkyl, acetyl, formyl, benzoyl or trifluoroacetyl, and;</p><p num="0133">R²Is OH or NH₂and; Also</p><p num="0134">Z is a peptide having the following formula (IIb), and:</p><p num="0135">His-Ser-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-X12-Tyr-Leu-Asp-X16-X17-Ala-Ala-X20-X21-Phe-Val-X24-Trp-Leu-Leu-X28-Ala; (IIb)</p><p num="0136">here</p><p num="0137">X12 is selected from Lys, Arg, and Leu;</p><p num="0138">X16 is selected from Ser and X;</p><p num="0139">X17 is X;</p><p num="0140">X20 is selected from His and X;</p><p num="0141">X21 is selected from Asp and Glu;</p><p num="0142">X24 is selected from Ala and Glu;</p><p num="0143">X28 is selected from Ser, Lys and Arg; Also</p><p num="0144">Wherein each X is independently selected from Glu, Lys, and is selected from the group consisting of Cys;</p><p num="0145">Wherein X is a side chain of at least one moiety is coupled to a lipophilic substituent having the formula (i) or (ii):</p><p num="0146">(i) Z^One, Where Z^OneDirectly bonded to the side chain of X is the lipophilic portion; or</p><p num="0147">(ii) Z^OneZ², Where Z^OneIs the lipophilic part, Z²Is a spacer, and Z^OneThe Z²Coupled via the side chain of X is;</p><p num="0148">Z is a single HSQGTFTSDYSKYLDS-K (hexadecanoyl -γ-Glu)) - AAHDFVEWLLRA is not.</p><p num="0149">The compound may have the following formula: </p><p num="0150">R^One-ZR²</p><p num="0151">Wherein</p><p num="0152"> R^OneIs H, C_One_-4 Alkyl, acetyl, formyl, benzoyl or trifluoroacetyl, and;</p><p num="0153">R²Is OH or NH₂and; Also</p><p num="0154">Z is a peptide having the following formula (IIIa):</p><p num="0155">His-Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-X12-Tyr-Leu-Asp-Ser-X17-Ala-Ala-X20-X21-Phe-Val-X24-Trp-Leu-Leu-X28-Ala; (IIIa)</p><p num="0156">here </p><p num="0157">X12 is selected from Lys and Arg;</p><p num="0158">X17 is X;</p><p num="0159">X20 is selected from His and X;</p><p num="0160">X21 is selected from Asp and Glu;</p><p num="0161">X24 is selected from Ala and Glu;</p><p num="0162">X28 is selected from Ser, Lys and Arg; Also</p><p num="0163">Wherein each X moiety is independently selected from Glu, Lys, and Cys;</p><p num="0164">Wherein X is a side chain of at least one moiety is coupled to a lipophilic substituent having the following formula (i) or (ii): </p><p num="0165">(i) Z^One, Where Z^OneDirectly bonded to the side chain of X is the lipophilic portion; or</p><p num="0166">(ii) Z^OneZ², Where Z^OneIs a lipophilic region, Z²A spacer, and Z^OneThe Z²It is coupled to the side chain of X through.</p><p num="0167">Alternatively, the compound can have the following formula: </p><p num="0168">R^One-ZR²</p><p num="0169">Wherein</p><p num="0170"> R^OneIs H, C_One_-4 Alkyl, acetyl, formyl, benzoyl or trifluoroacetyl, and;</p><p num="0171">R²Is OH or NH₂and; Also</p><p num="0172">Z is the formula (IIIb) having a peptide:</p><p num="0173">His-Ser-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-X12-Tyr-Leu-Asp-Ser-X17-Ala-Ala-X20-X21-Phe-Val-X24-Trp-Leu-Leu-X28-Ala; (IIIb)</p><p num="0174">here </p><p num="0175">X12 is selected from Lys or Arg;</p><p num="0176">X17 is X;</p><p num="0177">X20 is selected from His and X;</p><p num="0178">X21 is selected from Asp and Glu;</p><p num="0179">X24 is selected from Ala and Glu;</p><p num="0180">X28 is selected from Ser, Lys and Arg; Also</p><p num="0181">Wherein each X moiety is independently selected from Glu, Lys, and Cys;</p><p num="0182">Wherein X is a side chain of at least one moiety is coupled to a lipophilic substituent having the following formula (i) or (ii):</p><p num="0183">(i) Z^One, Where Z^OneDirectly bonded to the side chain of X is the lipophilic portion; or</p><p num="0184">(ii) Z^OneZ², Where Z^OneIs a lipophilic region, Z²And Z is a spacer^OneThe Z²Coupled via the side chain of X is;</p><p num="0185">Z is a single HSQGTFTSDYSKYLDS-K (hexadecanoyl -γ-Glu)) - AAHDFVEWLLRA not.</p><p num="0186">The compound may have the following formula: </p><p num="0187">R^One-ZR²</p><p num="0188">Wherein</p><p num="0189">R^OneIs H, C_One_-4 Alkyl, acetyl, formyl, benzoyl or trifluoroacetyl, and;</p><p num="0190">R²Is OH or NH₂and; Also</p><p num="0191">Z is the formula (IVa) is a peptide having:</p><p num="0192">His-Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-X12-Tyr-Leu-Asp-Ser-X17-Ala-Ala-His-X21-Phe-Val-X24-Trp-Leu-Leu-X28-Ala; (IVa)</p><p num="0193">here</p><p num="0194">X12 is selected from Lys and Arg;</p><p num="0195">X17 is X;</p><p num="0196">X21 is selected from Asp and Glu;</p><p num="0197">X24 is selected from Ala and Glu;</p><p num="0198">X28 is selected from Ser, Lys and Arg;</p><p num="0199">Where X is selected from the group consisting of Glu, Lys, and Cys; Also</p><p num="0200">Where X is coupled to the side chain of a lipophilic substituent having the formula (i) or (ii):</p><p num="0201">(i) Z^One, Where Z^OneDirectly bonded to the side chain of X is the lipophilic portion; or</p><p num="0202">(ii) Z^OneZ², Where Z^OneIs a lipophilic region, Z²Is a spacer, and Z^OneThe Z²It is coupled to the side chain of X through.</p><p num="0203">Alternatively, the compound can have the following formula: </p><p num="0204">R^One-ZR²</p><p num="0205">Wherein</p><p num="0206">R^OneIs H, C_One_-4 Alkyl, acetyl, formyl, benzoyl or trifluoroacetyl, and;</p><p num="0207">R²Is OH or NH₂and; Also</p><p num="0208">Z is a peptide having the following general formula (IVb):</p><p num="0209">His-Ser-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-X12-Tyr-Leu-Asp-Ser-X17-Ala-Ala-His-X21-Phe-Val-X24-Trp-Leu-Leu-X28-Ala; (IVb)</p><p num="0210">here </p><p num="0211">X12 is selected from Lys and Arg;</p><p num="0212">X17 is X;</p><p num="0213">X21 is selected from Asp and Glu;</p><p num="0214">X24 is selected from Ala and Glu;</p><p num="0215">X28 is selected from Ser, Lys and Arg;</p><p num="0216">Where X is selected from the group consisting of Glu, Lys, and Cys; Also</p><p num="0217">Where X is coupled to the side chain of a lipophilic substituent having the following formula (i) or (ii): </p><p num="0218">(i) Z^One, Where Z^OneDirectly bonded to the side chain of X is the lipophilic portion; or</p><p num="0219">(ii) Z^OneZ², Where Z^OneIs a lipophilic region, Z²Is a spacer, and Z^OneThe Z²Coupled via the side chain of X is;</p><p num="0220">Z is a single HSQGTFTSDYSKYLDS-K (hexadecanoyl -γ-Glu)) - AAHDFVEWLLRA is not.</p><p num="0221">Alternatively, the compound can have the following formula:</p><p num="0222">R^One-ZR²</p><p num="0223">Wherein</p><p num="0224">R^OneIs H, C_One_-4 Alkyl, acetyl, formyl, benzoyl or trifluoroacetyl, and; </p><p num="0225">R²Is OH or NH₂and; Also</p><p num="0226">Z is a peptide having the following formula (V) is:</p><p num="0227">His-Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Tyr-Leu-Asp-Ser-Lys-Ala-Ala-His-Asp-Phe-Val-Glu-Trp-Leu-Leu-X28; (V)</p><p num="0228">here</p><p num="0229">X28 is Ser or absence and or;</p><p num="0230">X17 is X;</p><p num="0231">Where X is selected from the group consisting of Glu, Lys, and Cys; Also</p><p num="0232">Where X is coupled to the side chain of a lipophilic substituent having the following formula (i) or (ii):</p><p num="0233">(i) Z^One, Where Z^OneDirectly bonded to the side chain of X is the lipophilic portion; or</p><p num="0234">(ii) Z^OneZ², Where Z^OneIs a lipophilic region, Z²Is a spacer, and Z^OneThe Z²It is coupled to the side chain of X through.</p><p num="0235">In certain embodiments of the present invention, peptides of formula (I) may have the following sequence: </p><p num="0236">HSQGTFTSDYSKYLDSKAAHDFVEWLLRA;</p><p num="0237">HSQGTFTSDYSKYLDKKAAHDFVEWLLRA;</p><p num="0238">HSQGTFTSDYSKYLDSKAAKDFVEWLLRA;</p><p num="0239">HSQGTFTSDYSKYLDSKAAHDFVEWLKRA;</p><p num="0240">HSQGTFTSDYSKYLDSKAAHDFVEWLLKA;</p><p num="0241">HSQGTFTSDYSRYLDSKAAHDFVEWLLRA;</p><p num="0242">HSQGTFTSDYSLYLDSKAAHDFVEWLLRA;</p><p num="0243">HSQGTFTSDYSKYLDSKAAHDFVEWLLRAK;</p><p num="0244">HSQGTFTSDYSKYLDSKAAHDFVEWLLSAK;</p><p num="0245">HSQGTFTSDYSKYLDSKAAHDFVEWLKSA;</p><p num="0246">HSQGTFTSDYSKYLDSKAAHDFVKWLLRA;</p><p num="0247">HSQGTFTSDYSKYLDSCAAHDFVEWLLRA;</p><p num="0248">HSQGTFTSDYSKYLDSCAAHDFVEWLLSA;</p><p num="0249">HSQGTFTSDYSKYLDSKAACDFVEWLLRA;</p><p num="0250">HSQGTFTSDYSKYLDKSAAHDFVEWLLRA;</p><p num="0251">H-Aib-QGTFTSDYSKYLDSKAAHDFVEWLLSA;</p><p num="0252">H-Aib-QGTFTSDYSKYLDSKAAHDFVEWLLSAK;</p><p num="0253">H-Aib-QGTFTSDYSKYLDSKAARDFVAWLLRA;</p><p num="0254">H-Aib-QGTFTSDYSKYLDSKAAKDFVAWLLRA;</p><p num="0255">H-Aib-QGTFTSDYSKYLDSKAAHDFVEWLLRA; </p><p num="0256">H-Aib-QGTFTSDYSKYLDSKAAHDFVEWLLKA;</p><p num="0257">H-Aib-QGTFTSDYSKYLDSKAAKDFVAWLLSA;</p><p num="0258">H-Aib-QGTFTSDYSKYLDSKAAHDFVAWLLKA;</p><p num="0259">H-Aib-QGTFTSDYSKYLDKKAAHDFVAWLLRA;</p><p num="0260">H-Aib-QGTFTSDYSRYLDSKAAHDFVEWLLSA;</p><p num="0261">H-Aib-QGTFTSDYSKYLDSKAAHDFVKWLLSA;</p><p num="0262">H-Aib-QGTFTSDYSLYLDSKAAHDFVEWLLSA;</p><p num="0263">H-Aib-QGTFTSDYSKYLDSCAAHDFVEWLLSA; </p><p num="0264">H-Aib-QGTFTSDYSKYLDSKAACDFVEWLLRA; </p><p num="0265">H-Aib-QGTFTSDYSKYLDK () KAAE () DFVEWLLRA; </p><p num="0266">H-Aib-QGTFTSDYSKYLDSKAAHDFVE () WLLK () A;</p><p num="0267">H-Aib-QGTFTSDYSKYLDSKAAK () DFVE () WLLRA; </p><p num="0268">H-Aib-QGTFTSDYSKYLDSK () AAHE () FVEWLLKA; or</p><p num="0269">H-Aib-QGTFTSDYSKYLDSK () AAKE () FVEWLLRA.</p><p num="0270">In certain embodiments, these peptides may have a lipophilic substituent in the position marked "*" as follows: </p><p num="0271">HSQGTFTSDYSKYLDS-K * -AAHDFVEWLLRA;</p><p num="0272">HSQGTFTSDYSKYLD-K * -KAAHDFVEWLLRA;</p><p num="0273">HSQGTFTSDYSKYLDSKAA-K * -DFVEWLLRA;</p><p num="0274">HSQGTFTSDYSKYLDSKAAHDFVEWL-K * -RA;</p><p num="0275">HSQGTFTSDYSKYLDSKAAHDFVEWLL-K * -A;</p><p num="0276">HSQGTFTSDYSRYLDS-K * -AAHDFVEWLLRA;</p><p num="0277">HSQGTFTSDYSLYLDS-K * -AAHDFVEWLLRA;</p><p num="0278">HSQGTFTSDYSKYLDSKAAHDFVEWLLRA-K *;</p><p num="0279">HSQGTFTSDYSKYLDSKAAHDFVEWLLSA-K *;</p><p num="0280">HSQGTFTSDYSKYLDSKAAHDFVEWL-K * -SA;</p><p num="0281">HSQGTFTSDYSKYLDSKAAHDFV-K * -WLLRA;</p><p num="0282">HSQGTFTSDYSKYLDS-C * -AAHDFVEWLLRA;</p><p num="0283">HSQGTFTSDYSKYLDS-C * -AAHDFVEWLLSA;</p><p num="0284">HSQGTFTSDYSKYLDSKAA-C * -DFVEWLLRA;</p><p num="0285">HSQGTFTSDYSKYLD-K * -SAAHDFVEWLLRA;</p><p num="0286">H-Aib-QGTFTSDYSKYLDS-K * -AAHDFVEWLLSA;</p><p num="0287">H-Aib-QGTFTSDYSKYLDSKAAHDFVEWLLSA-K *; </p><p num="0288">H-Aib-QGTFTSDYSKYLDS-K * -AARDFVAWLLRA;</p><p num="0289">H-Aib-QGTFTSDYSKYLDSKAA-K * -DFVAWLLRA; </p><p num="0290">H-Aib-QGTFTSDYSKYLDSKAAHDFVEWLL-K * -A;</p><p num="0291">H-Aib-QGTFTSDYSKYLDS-K * -AAHDFVEWLLKA;</p><p num="0292">H-Aib-QGTFTSDYSKYLDS-K * -AAHDFVEWLLRA;</p><p num="0293">H-Aib-QGTFTSDYSKYLDSKAA-K * -DFVAWLLSA;</p><p num="0294">H-Aib-QGTFTSDYSKYLDSKAAHDFVAWLL-K * -A;</p><p num="0295">H-Aib-QGTFTSDYSKYLD-K * -KAAHDFVAWLLRA;</p><p num="0296">H-Aib-QGTFTSDYSRYLDS-K * -AAHDFVEWLLSA;</p><p num="0297">H-Aib-QGTFTSDYSKYLDSKAAHDFV-K * -WLLSA;</p><p num="0298">H-Aib-QGTFTSDYSLYLDS-K * -AAHDFVEWLLSA;</p><p num="0299">H-Aib-QGTFTSDYSKYLDS-C * -AAHDFVEWLLSA;</p><p num="0300">H-Aib-QGTFTSDYSKYLDSKAA-C * -DFVEWLLRA; </p><p num="0301">H-Aib-QGTFTSDYSKYLD-S * -KAAHDFVEWLLSA;</p><p num="0302">H-Aib-QGTFTSDYSKYLDK () K * AAE () DFVEWLLRA; </p><p num="0303">H-Aib-QGTFTSDYSKYLDSK * AAHDFVE () WLLK () A;</p><p num="0304">H-Aib-QGTFTSDYSKYLDSK * AAK () DFVE () WLLRA; </p><p num="0305">H-Aib-QGTFTSDYSKYLDSK () AAHE () FVEWLLK * A; or</p><p num="0306">H-Aib-QGTFTSDYSKYLDSK () AAK * E () FVEWLLRA.</p><p num="0307">Acid residue marked with "()" will be involved in the intramolecular bond as ring lactam. One or more side chains of the residues X (s) is coupled to a lipophilic substituent. For example, the side chains of residues X may be bonded to the lipophilic substituent. Alternatively, two of the moieties X, or even two or more side chains may be bonded to the lipophilic substituent.</p><p num="0308">For example, X16, X17, X20 and X28, at least one can be coupled to the lipophilic substituent. In this case, X30 can not be present. When X30 is present, typically it is coupled to a lipophilic substituent.</p><p num="0309">Thus the compounds are positions 16, 17, 20, 24, 27, 28 or 30, preferably at position 16, 17 or 20, in particular, have only one lipophilic substituent in position 17. </p><p num="0310">Alternatively, the compounds each of positions 16, 17, 20, 24, 27, 28 or precisely in a 30-can have two lipophilic substituents. Preferably only one or two lipophilic substituents are present in one position 16, 17 or 20.</p><p num="0311">Therefore, the compound is in position 16 and 17, 16 and 20, 16 and 24, 16 and 27, 16 and 28 or 16 and 30; 17 and 20, 17 and 24, 17 and 27, 17 and 28 or 17 and 30 in; 20 and 24, 20 and 27, 20 and 28 or 20 and 30; At 24 and 27, 24 and 28 or 24 and 30; 27 and 28 or 27 and 30; Or 28, and may have a lipophilic substituent in the 30.</p><p num="0312">In a further embodiment, the compound is position 16, 17, 20, 24, 27, 28 or more of the positions selected from 30, may have one or more lipophilic substituents (providing a total of 3 or more). However, up to two locations may preferably be derivatized in this way.</p><p num="0313">Z^OneMay have 10 to 24 carbon atoms, for example 10 to 22 carbon atoms, for example, hydrocarbons having 10 to 20 carbon atoms. At least 11 carbon atoms, and / or 18 carbon atoms may have. For example, the hydrocarbon chain of 12, 13, 14, 15, 16, 17 or 18 carbon atoms may have. Therefore Z^One Are dodecanoyl, 2-butyl octanoyl, tetra-decanoyl, hexadecanoyl, heptanoic decanoyl, may be octa decanoyl or eicosyl Sano one portion.</p><p num="0314">If independently, is present, Z²It may have to include one or more amino acids which may date mug. For example, Z²It may be a γ-Glu, Glu, β-Ala or ε-Lys moiety, or 4-amino nobuta Russo, 8-amino or 8-amino-3,6-dioxa-octanoyl octanoyl region.</p><p num="0315">Z^One And Z²Particular combinations of dodecanoyl -γ-Glu, hexadecanoyl -γ-Glu, -Glu hexadecanoyl, hexadecanoyl - [3-amino-propane alkanoyl], hexadecanoyl - [8-amino-oxazole alkanoyl], hexadecanoyl -ε-Lys, 2- butyl octanoyl -γ-Glu, Octavio decanoyl -γ-Glu and hexadecanoyl [4-amino nobuta Russo] can be.</p><p num="0316">In particular embodiments, Z can have the following formula:</p><p num="0317">HSQGTFTSDYSKYLD-K (hexadecanoyl -γ-Glu) -KAAHDFVEWLLRA;</p><p num="0318">HSQGTFTSDYSKYLDSKAAHDFVEWL-K (hexadecanoyl -γ-Glu) -RA;</p><p num="0319">HSQGTFTSDYSKYLDSKAA-K (hexadecanoyl -γ-Glu) -DFVEWLLRA;</p><p num="0320">HSQGTFTSDYSKYLDSKAAHDFVEWLL-K (hexadecanoyl -γ-Glu) -A;</p><p num="0321">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl -γ-Glu) -AAHDFVEWLLRA;</p><p num="0322">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl -γ-Glu) -AARDFVAWLLRA;</p><p num="0323">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl -γ-Glu) -AAHDFVEWLLSA;</p><p num="0324">H-Aib-QGTFTSDYSKYLDSKAAHDFVEWLL-K (hexadecanoyl -γ-Glu) -A;</p><p num="0325">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl -γ-Glu) -AAHDFVE () WLLK () A;</p><p num="0326">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl -γ-Glu) -AAHDFVEWLLKA;</p><p num="0327">HSQGTFTSDYSKYLDS-K (hexadecanoyl -γ-Glu) -AAHDFVEWLLRA;</p><p num="0328">H-Aib-QGTFTSDYSKYLDSKAA-K (hexadecanoyl -γ-Glu) -DFVAWLLRA;</p><p num="0329">H-Aib-QGTFTSDYSKYLDS-K (dodecanoyl -γ-Glu) -AAHDFVEWLLSA;</p><p num="0330">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl [3-amino-propanoyl]) - AAHDFVEWLLSA;</p><p num="0331">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl [8-amino-octanoyl]) - AAHDFVEWLLSA;</p><p num="0332">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl -ε-Lys) -AAHDFVEWLLSA;</p><p num="0333">HSQGTFTSDYSKYLDS-K (hexadecanoyl) -AAHDFVEWLLSA;</p><p num="0334">HSQGTFTSDYSKYLDS-K (octa-decanoyl -γ-Glu) -AAHDFVEWLLSA;</p><p num="0335">HSQGTFTSDYSKYLDS-K ([2- butyl octanoyl] -γ-Glu) -AAHDFVEWLLSA;</p><p num="0336">HSQGTFTSDYSKYLDS-K (hexadecanoyl [4-amino nobuta Russo]) - AAHDFVEWLLSA;</p><p num="0337">HSQGTFTSDYSKYLDS-K (octa-decanoyl -γ-Glu) -AAHDFVEWLLSA;</p><p num="0338">HSQGTFTSDYSKYLDS-K (hexadecanoyl -E) -AAHDFVEWLLSA;</p><p num="0339">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl) -AAHDFVEWLLSA;</p><p num="0340">H-Aib-QGTFTSDYSKYLDS-K (octa-decanoyl -γ-Glu) -AAHDFVEWLLSA;</p><p num="0341">H-Aib-QGTFTSDYSKYLDS-K ([2- butyl octanoyl] -γ-Glu) -AAHDFVEWLLSA;</p><p num="0342">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl [4-amino nobuta Russo]) - AAHDFVEWLLSA;</p><p num="0343">H-Aib-QGTFTSDYSKYLDS-K (octa-decanoyl -γ-Glu) -AAHDFVEWLLSA; or</p><p num="0344">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl -E) -AAHDFVEWLLSA.</p><p num="0345">Acid residue marked with "()" will be involved in the intramolecular bond as ring lactam.</p><p num="0346">In a further embodiment, Z has the formula:</p><p num="0347">H-Aib-QGTFTSDYS-K (hexadecanoyl-iso-Glu) -YLDSKAAHDFVEWLLSA;</p><p num="0348">H-Aib-QGTFTSDYSKYLD-K (hexadecanoyl-iso-Glu) -KAAHDFVEWLLSA;</p><p num="0349">H-Aib-QGTFTSDYSKYLDSKAA-K (hexadecanoyl-iso-Glu) -DFVEWLLSA;</p><p num="0350">H-Aib-QGTFTSDYSKYLDSKAAHDFV-K (hexadecanoyl-iso-Glu) -WLLSA;</p><p num="0351">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl-iso-Lys) -AARDFVAWLLRA;</p><p num="0352">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl-iso-Glu) -AAKDFVEWLLSA;</p><p num="0353">H-Aib-QGTFTSDYSKYLDE-K (hexadecanoyl-iso-Glu) -AAHDFVEWLLSA;</p><p num="0354">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl-iso-Glu) -AAHEFVEWLLSA;</p><p num="0355">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl-iso-Glu) -AAEDFVEWLLSA;</p><p num="0356">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl-iso-Glu) -AAHDFVEWLLEA.</p><p num="0357">In a further aspect, Z has the formula:</p><p num="0358">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl-iso-Glu) -AAHDFVEWLLS;</p><p num="0359">H-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl-iso-Glu) -AAHDFVEWLL;</p><p num="0360">In a further aspect, Z has the formula:</p><p num="0361">H-Aib-EGTFTSDYSKYLDS-K (hexadecanoyl-iso-Glu) -AAHDFVEWLLSA;</p><p num="0362">The present invention provides a compound having the formula: </p><p num="0363">R^One-ZR²</p><p num="0364">Wherein</p><p num="0365">R^OneIs H, C_One_-4 Alkyl, acetyl, formyl, benzoyl or trifluoroacetyl, and;</p><p num="0366">R²Is OH or NH₂and; Also</p><p num="0367">Z is the formula (I) is a peptide having:</p><p num="0368">His-X2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-X12-Tyr-Leu-Asp-X16-X17-Ala-Ala-X20-X21-Phe-Val-X24-Trp-Leu-X27-X28-Ala-X30, (I); </p><p num="0369">here</p><p num="0370">And X2 is Ser or Aib;</p><p num="0371">X12 is selected from Lys, Arg or Leu;</p><p num="0372">X16 is Arg or X;</p><p num="0373">X17 is Arg or X;</p><p num="0374">X20 is Arg, or His and X;</p><p num="0375">And X21 is Asp or Glu;</p><p num="0376">X24 is Ala or X;</p><p num="0377">X27 is Leu or X;</p><p num="0378">X28 is Arg or X;</p><p num="0379">The absence of either the X or X30; Also</p><p num="0380">Wherein each X moiety is independently selected from Glu, Lys, Ser, Cys, Dbu, is selected from the group consisting of Dpr and Orn;</p><p num="0381">The side chains of at least one residue X is the following formula (i) or (ii) is coupled to a lipophilic substituent having: </p><p num="0382">(i) Z^One, Where Z^OneDirectly bonded to the side chain of X is the lipophilic portion; or</p><p num="0383">(ii) Z^OneZ², Where Z^OneIs a lipophilic region, Z²Is a spacer, and Z^OneThe Z²Coupled via the side chain of X is; </p><p num="0384">Z is a single HSQGTFTSDYSKYLDS-K (hexadecanoyl -γ-Glu) is not a -AAHDFVEWLLRA.</p><p num="0385">X30 may be present or absent. In this case, X30 is present in these embodiments it may be desirable in the Lys.</p><p num="0386">In certain embodiments, any residue X, and in particular any residues X coupled to a lipophilic substituent is independently selected from Lys, Glu or Cys.</p><p num="0387">The compound may have the following formula:</p><p num="0388">R^One-ZR²</p><p num="0389">Wherein</p><p num="0390"> R^OneIs H, C_One_-4 Alkyl, acetyl, formyl, benzoyl or trifluoroacetyl, and;</p><p num="0391">R²Is OH or NH₂and; Also</p><p num="0392">Z is the formula (IIa) is a peptide having:</p><p num="0393">His-Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-X12-Tyr-Leu-Asp-X16-X17-Ala-Ala-X20-X21-Phe-Val-X24-Trp-Leu-Leu-X28-Ala, (IIa);</p><p num="0394">here </p><p num="0395">X12 is selected from Lys, Arg or Leu;</p><p num="0396">X16 is Ser or X;</p><p num="0397">X17 is X;</p><p num="0398">X20 is His or X;</p><p num="0399">And X21 is Asp or Glu;</p><p num="0400">The X24 is Ala or Glu;</p><p num="0401">X28 is Ser, Lys or Arg, and; Also</p><p num="0402">Wherein each X moiety is independently selected from Glu, Lys, or is selected from the group consisting of Cys;</p><p num="0403">The side chains of at least one residue X is the following formula (i) or (ii) is coupled to a lipophilic substituent having:</p><p num="0404">(i) Z^One, Where Z^OneDirectly bonded to the side chain of X is the lipophilic portion; or</p><p num="0405">ii) Z^OneZ², Where Z^OneIs a lipophilic region, Z²A spacer, and Z^OneThe Z²It is coupled to the side chain of X through.</p><p num="0406">Alternatively, the compound can have the following formula:</p><p num="0407">R^One-ZR²</p><p num="0408">Wherein</p><p num="0409">R^OneIs H, C_One_-4 Alkyl, acetyl, formyl, benzoyl or trifluoroacetyl, and;</p><p num="0410">R²Is OH or NH₂and; Also</p><p num="0411">Z is a peptide having the following formula (IIb), and:</p><p num="0412">His-Ser-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-X12-Tyr-Leu-Asp-X16-X17-Ala-Ala-X20-X21-Phe-Val-X24-Trp-Leu-Leu-X28-Ala, (IIb);</p><p num="0413">here </p><p num="0414">X12 is selected from Lys, Arg or Leu;</p><p num="0415">X16 is Ser or X;</p><p num="0416">X17 is X;</p><p num="0417">X20 is His or X;</p><p num="0418">And X21 is Asp or Glu;</p><p num="0419">The X24 is Ala or Glu;</p><p num="0420">X28 is Ser, Lys or Arg, and; Also</p><p num="0421">Wherein each X moiety is independently selected from Glu, Lys, or is selected from the group consisting of Cys;</p><p num="0422">Wherein X is a side chain of at least one moiety is coupled to a lipophilic substituent having the formula (i) or (ii):</p><p num="0423">(i) Z^One, Where Z^OneDirectly bonded to the side chain of X is the lipophilic portion; or</p><p num="0424">(ii) Z^OneZ², Where Z^OneIs a lipophilic region, Z²Is a spacer, and Z^One The Z²Coupled via the side chain of X is;</p><p num="0425">Z is a single HSQGTFTSDYSKYLDS-K (hexadecanoyl -γ-Glu)) - AAHDFVEWLLRA is not.</p><p num="0426">The compound may have the following formula: </p><p num="0427">R^One-ZR²</p><p num="0428">Wherein</p><p num="0429">R^OneIs H, C_One_-4 Alkyl, acetyl, formyl, benzoyl or trifluoroacetyl, and;</p><p num="0430">R²Is OH or NH₂and; Also</p><p num="0431">Z is a peptide having the following formula (IIIa):</p><p num="0432">His-Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-X12-Tyr-Leu-Asp-Ser-X17-Ala-Ala-X20-X21-Phe-Val-X24-Trp-Leu-Leu-X28-Ala, (IIIa);</p><p num="0433">here </p><p num="0434">X12 is selected from Lys or Arg;</p><p num="0435">X17 is X;</p><p num="0436">X20 is His or X;</p><p num="0437">The X21 is Asp or Glu;</p><p num="0438">The X24 is Ala or Glu;</p><p num="0439">X28 is Ser, Lys or Arg, and; Also</p><p num="0440">Wherein each X moiety is independently selected from Glu, Lys, or Cys;</p><p num="0441">The side chains of at least one residue X is the following formula (i) or (ii) is coupled to a lipophilic substituent having:</p><p num="0442">(i) Z^One, Where Z^OneDirectly bonded to the side chain of X is the lipophilic portion; or</p><p num="0443">(ii) Z^OneZ², Where Z^OneIs a lipophilic region, Z²Is a spacer, and Z^OneThe Z²It is coupled to the side chain of X through. </p><p num="0444">Alternatively, the compound can have the following formula: </p><p num="0445">R^One-ZR²</p><p num="0446">Wherein</p><p num="0447">R^OneIs H, C_One_-4 Alkyl, acetyl, formyl, benzoyl or trifluoroacetyl, and;</p><p num="0448">R²Is OH or NH₂and; Also</p><p num="0449">Z is the formula (IIIb) having a peptide:</p><p num="0450">His-Ser-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-X12-Tyr-Leu-Asp-Ser-X17-Ala-Ala-X20-X21-Phe-Val-X24-Trp-Leu-Leu-X28-Ala, (IIIb);</p><p num="0451">here </p><p num="0452">X12 is selected from Lys or Arg;</p><p num="0453">X17 is X;</p><p num="0454">X20 is His or X;</p><p num="0455">And X21 is Asp or Glu;</p><p num="0456">The X24 is Ala or Glu;</p><p num="0457">X28 is Ser, Lys or Arg, and; Also</p><p num="0458">Wherein each X moiety is independently selected from Glu, Lys, or Cys;</p><p num="0459">The side chains of at least one residue X is the following formula (i) or (ii) is coupled to a lipophilic substituent having:</p><p num="0460">(i) Z^One, Where Z^OneDirectly bonded to the side chain of X is the lipophilic portion; or</p><p num="0461">(ii) Z^OneZ², Where Z^OneIs a lipophilic region, Z²Is a spacer, and Z^OneThe Z²Coupled via the side chain of X is;</p><p num="0462">Z is a single HSQGTFTSDYSKYLDS-K (hexadecanoyl -γ-Glu)) - AAHDFVEWLLRA is not.</p><p num="0463">The compound may have the following formula: </p><p num="0464">R^One-ZR²</p><p num="0465">Wherein</p><p num="0466">R^OneIs H, C_One_-4 Alkyl, acetyl, formyl, benzoyl or trifluoroacetyl, and;</p><p num="0467">R²Is OH or NH₂and; Also</p><p num="0468">Z is a peptide having the following formula (IVa):</p><p num="0469">His-Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-X12-Tyr-Leu-Asp-Ser-X17-Ala-Ala-His-X21-Phe-Val-X24-Trp-Leu-Leu-X28-Ala, (IVa);</p><p num="0470">here </p><p num="0471">X12 is selected from Lys or Arg;</p><p num="0472">X17 is X;</p><p num="0473">And X21 is Asp or Glu;</p><p num="0474">The X24 is Ala or Glu;</p><p num="0475">X28 is Ser, Lys or Arg, and;</p><p num="0476">Where X is selected from the group consisting of Glu, Lys, or Cys; Also</p><p num="0477">Where X is the side chain of the following formula (i) or (ii) is coupled to a lipophilic substituent having:</p><p num="0478">(i) Z^One, Where Z^OneCoupled to the side chain of X is the lipophilic portion; or</p><p num="0479">(ii) Z^OneZ², Where Z^OneIs a lipophilic region, Z²Is a spacer, and Z^OneThe Z²It is coupled to the side chain of X through. </p><p num="0480">Alternatively, the compound can have the following formula:</p><p num="0481">R^One-ZR²</p><p num="0482">Wherein</p><p num="0483">R^OneIs H, C_One_-4 Alkyl, acetyl, formyl, benzoyl or trifluoroacetyl, and;</p><p num="0484">R²Is OH or NH₂and; Also</p><p num="0485">Z is the formula (IVb) with a peptide;</p><p num="0486">His-Ser-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-X12-Tyr-Leu-Asp-Ser-X17-Ala-Ala-His-X21-Phe-Val-X24-Trp-Leu-Leu-X28-Ala, (IVb);</p><p num="0487">here </p><p num="0488">X12 is selected from Lys or Arg;</p><p num="0489">X17 is X;</p><p num="0490">The X21 is Asp or Glu;</p><p num="0491">The X24 is Ala or Glu;</p><p num="0492">X28 is Ser, Lys or Arg, and;</p><p num="0493">Where X is selected from the group consisting of Glu, Lys, or Cys; Also</p><p num="0494">Where X is the side chain of the following formula (i) or (ii) is coupled to a lipophilic substituent having:</p><p num="0495">(i) Z^One, Where Z^OneDirectly bonded to the side chain of X is the lipophilic portion; or</p><p num="0496">(ii) Z^OneZ², Where Z^OneIs a lipophilic region, Z²Is a spacer, and Z^OneThe Z²Coupled via the side chain of X is;</p><p num="0497">Z is a single HSQGTFTSDYSKYLDS-K (hexadecanoyl -γ-Glu)) - AAHDFVEWLLRA is not.</p><p num="0498">In a further aspect, the present invention provides a composition comprising a compound, or a salt or derivative as defined herein with a carrier. In a preferred embodiment, the composition is a composition of a pharmaceutically acceptable carrier is also the carrier is pharmaceutically acceptable. The acid addition salts are pharmaceutically acceptable salts of the compounds, such as nitrate or chloride may be a salt.</p><p num="0499">The above-mentioned compounds show a use for preventing weight gain or promote weight loss. "Preventing" refers to inhibiting or decreasing weight gain in comparison with non-treated, and does not necessarily mean to include the complete stop of the increase in weight. Peptide is the cause of reduced and / or increased energy consumption of the food intake, may result in the observed effect on body weight. And their effect on the body weight, independently of the compounds of the invention may have a beneficial effect on circulating glucose levels, glucose tolerance, and / or circulating cholesterol levels, and this reduces the circulation LDL levels and increasing the HDL / LDL ratio It can be. Therefore, directly or indirectly, treatment of compounds of the present invention are any condition cause, or which is characterized them by excess weight, for example, obesity, morbid obesity, obesity-related inflammation, obesity-related gallbladder disease, obesity-induced sleep apnea It can be used for the treatment and / or prevention. These compounds are also diabetes jeonjeung, insulin resistance, glucose tolerance, type 2 diabetes, the I-type diabetes, high blood pressure or atherosclerosis lipid disorders metabolism (or their metabolic risk of two or more combinations of parameters), atherosclerosis, arteriosclerosis, coronary artery heart disease, can be used in the treatment of peripheral arterial disease, stroke, and micro-vascular disease. In these symptoms, the effect of these compounds can result in connection with their effects on the weight, or those with one or independently.</p><p num="0500">Therefore, the present invention provides the use of a compound of the invention in the treatment of symptoms as described above to a subject in need thereof. </p><p num="0501">The invention also provides the use of a compound of the invention for use in the treatment of symptoms such as those for, in particular for use in the medical treatment described above. </p><p num="0502">The invention also provides the use of a compound of the invention in the manufacture of a medicament for the treatment of the symptoms described above. </p><p num="0503">The compounds of the invention may be administered, diabetes, obesity, as part of a combination therapy with a medicament for the treatment of high blood pressure or more than the target lipid.</p><p num="0504">In this case, the two active agents may be provided together or separately, in which can be provided as the same pharmaceutical formulation or as a separate part of the formulation.</p><p num="0505">Thus compounds of the invention (or a salt thereof) is not limited, for anti-diabetic drugs, such as metformin, a sulfonylurea, Glynn arsenide (glinide), DPP-IV inhibitors, glitazones (glitazone), or insulin in combination with and it can be used. In a preferred embodiment, the compound or a salt thereof to achieve a suitable control blood sugar, insulin, DPP-IV inhibitors, sulfonylureas or metformin, in particular used in combination with a sulfonylurea or metformin. In a more preferred embodiment, the compound or its salts can be used in combination with metformin, a sulfonylurea, an insulin or insulin analogue to achieve adequate glycemic control. Examples of insulin analogues include, but are not limited Lanta's (Lantus), look no feed (Novorapid), Huma log (Humalog), Novo Mix (Novomix), evil Tra edition (Actraphane) HM, L'Mir (Levemir) and Bahia de la ( and a Apidra).</p><p num="0506">The compound or a salt thereof is not limited anti-obesity agents, for example, glucagon-like peptide receptor-1 agonists (agonist), peptide YY or an analogue, cannabinoid receptor 1 antagonist (antagonist), a lipase inhibitor, a melanocortin receptor The 4 agonists, or in combination with melanin concentrating hormone receptor antagonists can be used. </p><p num="0507">The compound or a salt thereof is not limited, for anti-hypertensive agent, e.g., angiotensin-converting enzyme inhibitors, angiotensin II receptor blocker, a diuretic, a beta-blocker, or calcium can be further used in combination with channel blockers. </p><p num="0508">The compound or a salt thereof, wherein the cholesterol, for example, but are not limited to, statins (statin), fibrate (fibrate), may be used in combination with the new age (niacin) or a cholesterol absorption inhibitor. </p>

Figure 1. 100 nmol / kg of the mouse at a dose subcutaneous (sc) pharmacokinetic profile of the compound 13 after administration. Figure 2. Effect of subcutaneous administration of the compound for 21 days for oral glucose tolerance in long-term intake of high-fat C57BL / 6J mice 11 (10 nmol / kg). Data represent the mean ± SEM. Figure 3. diabetic (db / db) mice treated vehicle or compound 7 (12.7 nmol / kg) was also 4 weekly in HbA1c was measured in whole blood sample collected from the treated mice (20 mu l) (Cobas® Application Notes : A1C-2). △ HbA1c (%) was subtracted and their HbA1c (%) in the process starting from the [Delta] HbA1c (%) for 4 weeks was calculated for each mouse. in db / db mice △ HbA1c (%) were treated 4 weeks with vehicle = 100%. * (P = 0.03, Student t- test). 4. The long-term intake of high-fat also C57BL / 6J 21 yl effects of subcutaneous administration of compound 11 on body weight in mice. Data are expressed as mean + SEM. Figure 5. Diet induced obese (DIO) mice were treated with vehicle or compound 4 weeks 7 (12.7 nmol / kg) was also prepared from the collected blood plasma sample. Total cholesterol were measured in each plasma sample (Cobas® Application Note CHOL2). *** (P &lt;0.0001, Student's t- test). Data represents the mean + SEM. Figure 6. Dietary-induced obese (DIO) mice were treated with vehicle or compound 7 (12.7 nmol / kg) was also prepared from the collected blood plasma sample. LDL and HDL cholesterol were measured in each plasma sample (Cobas® Application Note HDLC3 and LDL_C). *** (P &lt;0.0001, Student's t- test). Data represents the mean + SEM. Figure 7. Effect of high fat intake C57BL / 6J subcutaneous administration of GluGLP-1 agonist for the weight gain in mice. Data are mean ± SEM. Black line: vehicle (PBS), gray line: low dose (0.5 nmol / kg), a broken line: high dose (5nmol / kg). Figure 8 a high-fat diet C57BL / 6J mouse administered at 2, 4, 6, 8, 10 and 12 to avoid the effect of acute administration of Compound 7 on glucose tolerance, and after oral administration. Data are expressed as mean + SEM. 9. Children also ride local C57BL / 6J effect of compound 7 and subcutaneous administration of exendin-4 on food intake / body weight in mice. Data are mean + SEM. * = p &lt;0.05 vs. vehicle fat little ride. Data are expressed as mean + SEM. Figure 10. Older obese C57BL / 6J cumulative food intake / effects of subcutaneous administration of the compounds 7 and exendin-4 for the body weight in mice. Data are mean + SEM. * = p &lt;0.05 vs. vehicle obese elderly. Data are expressed as mean + SEM. Figure 11. aged obese C57BL / 6J excipient on plasma lipid levels in mice, and the effects of subcutaneous administration of exendin -4 (10 nmol / kg) and Compound 11 (10 nmol / kg). Data are mean + SEM. Figure 12. Mouse Compound 1 and Compound 11 (2 doses of 0.5 and 5 in nmol / kg) or vehicle was treated twice a day for two weeks. It was sacrificed on day bokgae the liver and weighed. Compound 1 "liver weight / body weight ratio" is significantly increased at higher doses. Compound 11 did not affect the "liver weight / body weight ratio" at two doses (0.5 and 5 nmol / kg). Compound 1 is not a double acylation GluGLP-1 agonists, and the compounds 11 is functional for a long time acylated double GluGLP-1 agonist (12). Figure 13. diabetic (db / db) mice treated vehicle, or compound 11 (12.7 nmol / kg) in four weeks, and, HbA1c was measured in a whole blood sample collected from the treated mice (20 mu l) (Cobas® Application Notes : A1C-2). △ HbA1c (%) was subtracted and their HbA1c (%) in the process starting from the HbA1c (%) 4 week was calculated for each mouse. in db / db mice △ HbA1c (%) were treated 4 weeks with vehicle = 100%. * (P = 0.03, Student t- test).

Through the present specification the first half, a naturally occurring conventional one letter and three letter code for amino acids is used as well, another amino acid, for example Aib (a-amino-isobutyric acid), Orn (ornithine), Dbu (2 , 4-diamino-butyric acid), and the three letter codes generally acceptable for Dpr (2, 3-diamino-propanoyl acid) is used.

The term "natural glucagon" (naive glucagon) is a sequence With natural human glucagon means.

Peptide sequences of the compounds of the invention are at least positions 18, 20, 24, 27, 28 and 29 differ from the sequences of natural glucagon. In addition, this may be different from that of natural glucagon in positions 12, 16 and 17 of one or more.

Natural glucagon have the Arg in position 18. The compounds of the invention on both the glucagon and GLP-1 receptor, but in particular have a small hydrophobic residues Ala at position 18 is believed to increase the effectiveness in the GLP-1 receptor.

Position 27 of native glucagon, 28 and 29 in the moiety is considered to provide a selectivity for the glucagon receptor hyeonjeon. Natural glucagon substituted in these positions with respect to the sequence, in particular, Ala 29 position, without potentially significant reduction in the effect on the glucagon receptor, can increase the potency and / or selectivity for the GLP-1 receptor. Additional compounds which may be included in the present invention examples of which include the Arg at position 28 and Leu at position 27. Furthermore, when in position 28 is Arg at position 24 with Glu crosslinking to form a molecule which may be particularly preferred since it is possible to increase its influence on the effect in the GLP-1 receptor.

Substitution also reduces the oxidation effect of the Met residue at position 27, a naturally occurring (that is Leu, Lys or Glu), thus increasing the chemical stability of the compounds.

Asn residue at position 28 of the naturally occurring (eg, Arg, with Ser) is substituted sikimyeo also reduce the effect talah folk acidic solution, thereby increasing the stability of the compounds.

Potentially effective and / or selective in the GLP-1 receptor of the glucagon receptor without significant loss effect is also standing on the C- terminal portion of the peptide alpha-may be increased by the introduction of residues that are likely to stabilize the helical structure. The helical portion of the molecule may be desirable but essential to the amphiphilic property is not considered. The introduction of residues such as Ala in Leu and / or position 24 in the position 22 may be secondary. Additionally or alternatively, charged residues are positions 16, 20, 24, and 28 can be introduced in one or more than one. Therefore, the charge may be all the positions 24 and 28 residues, positions 20, 24, and 28 can all be charged residues at either position 16 or 20, 24, and 28 can be both positively charged residues in the. For example, residues at position 20 may be Arg or His, in particular His. Residue in position 24 may be Glu, Lys or Arg, particularly Glu. Residue in position 28 may be Arg. The introduction of crosslinking in a molecule of the molecule as described above may also serve to stabilize the helix characteristic between positions 24 and 28, for example.

Substituted with one or two of the naturally occurring residue Gln at position 20 and 24 also reduces the effect on the talah folk of the acid solution, thereby increasing the stability of the compounds.

The substitution at position 12 of the native glucagon sequence (i.e. Arg, Leu) is to increase the selectivity in the effects and / or GLP-1 receptor in the two receptors.

C- terminal cleavage peptide (truncation) does not reduce the selectivity of the effect and / or GLP-1 of the two receptors. In particular, the cut or cutting in two positions 28 and 29 in the position 29 does not reduce the effect of any of the two receptors, receptor.

(If that is present the position 16, 17, 20, 24, 27 and 28, and / or 30) at least one moiety represented by X of the side chain is coupled to a lipophilic substituent. The combination of the lipophilic substituent is a side chain of a specific effect on certain advantages (e.g., reduced) can be provided in a non-bonded side chain it will be appreciated that the position could. The inventors have found that the compounds of the present invention provides a balance between the advantages and benefits of the specific substitutions to the native glucagon sequence acylated.

The composition of the present invention to further enhance the stability of the compound, increase bioavailability, increase solubility, decrease adverse effects, achieve the well-known time treatment (chronotherapy) to those skilled in the art, and also the patient's In order to receive state (compliance) or increase its specific combination, for example, share, through the hydrophobic and electrostatic interactions, a drug carrier, may be blended in the drug delivery system and advanced drug delivery system or in can be attached . Carrier, for example, drug delivery systems and advanced drug delivery systems include, but are not limited to, polymers, for example cellulose and derivatives, polysaccharides, for example dextran and derivatives, starch and derivatives, poly (vinyl alcohol), acrylate and methacrylate polymers, polylactic and polyglycolic acid and their block copolymers, polyethylene glycols, carrier proteins, for example albumin, gels, for example, thermal gelation system (thermogelling systems), for example the art the well-known block copolymeric systems, micelles (micelles) to, liposomes, microspheres (microspheres), nano-particles, liquid crystals (liquid crystals) and their dispersion, L2 phase (phase) and their dispersion, (these phase behavior in lipid-water systems well known in the field of (phase behavior)), the polymeric micelles, the multi-layer emulsion, self-emulsifying, self-micro-emulsifying, include cyclodextrin and derivatives thereof, and Den trimmer.

Other groups have attempted to extend the half-life of the double GluGLP-1 agonist compounds by PEG and derivatized (International Application Publication WO 2008/101017). However, derivatization is likely the most effective when applied to the C- terminus of the molecule than at the central core of the peptide backbone, and the effect of these compounds is reduced compared to the corresponding unmodified peptide.

On the other hand, the compounds of the invention while having a remarkably delayed pharmacokinetic profile than the non-modified peptide corresponding to maintain a high effect on the glucagon and GLP-1 receptor.

Natural glucagon has a Ser at position 16. Ala, Gly or Thr is substituted is shown to significantly reduce the climb between the active adenylate in the glucagon receptor (such Unson Proc. Natl. Acad. Sci. 1994, 91, 454-458). Thus, the derivatised with lipophilic substituent position 16 as shown surprisingly the compounds described herein, was not expected to produce a compound having an effect in the glucagon receptor. In International Application Publication WO 2008/101017, which is a negatively charged residue at position 16 has been found to be preferred in order to minimize the loss of effect.

In the presence of a basic amino acid at position 17 and 18 is generally considered to be necessary to enable the complete glucagon receptor (Unson like. J. Biol Chem.. 1998, 273, 10308-10312). The present inventors have found that, when the position 18 is alanine, at position 17 revealed that a compound of a higher effect by the hydrophobic amino acid substitution. Even amino acid is derivatized with a lipophilic substituent in position 17 as well as the compound to maintain nearly full effect in the glucagon and GLP-1 receptor, it shows a significantly delayed pharmacokinetic profile. This conversion to the basic amine side chain amide group in neutral if the derivatized lysine at position 17.

The inventors have also noted from the other, even in studies that the substitution at position 20 must be a basic amino acid having a side chain length of 4 to 6 atoms in order to enhance the GLP-1 receptor activity as compared to glucagon, the nearest 20-acylated found that compounds of highly active double agent (International Application Publication WO 2008/101017). Compound described herein shall have the GLP-1 and glucagon receptor activity when the 20 position is substituted with lysine is also acylated.

<u>Peptide</u><u> synthesis</u>

Peptides of the invention compounds can be prepared by standard synthetic methods, recombinant expression system, or any other suitable method. Thus, for example, the peptides can be synthesized in a number of ways, including the following methods:

(a) the synthesis of the peptide to the solid phase or the liquid phase method or by a short step by step assemble and separate and purify the end product peptide;

(b) expressing a nucleic acid encoding a peptide structure within a host cell and recovering the expression product from the host cell culture; or

and (c) performing the cell-free in vitro expression of a nucleic acid construct encoding the peptide, and then recovering the expression product.

Or the method (a), to obtain fragments of the peptides by any combination of (b) and (c), after the ligation of the fragments to obtain the peptide and recovering the peptide.

It may be desirable to synthesize the analogues of the present invention using the solid or liquid phase peptide synthesis. In this context, and International Application Publication WO 98/11125 is incorporated, and also, among many others, Fields, GB, etc., 2002, "Principles and practice of solid-phase peptide synthesis". Synthetic Peptides (2nd Edition), and embodiments are cited here.

<u>Lipophilic</u><u> Substituent</u>

One or more of the amino acid side chains in the compounds of the present invention, a lipophilic substituent Z^OneIt is coupled to. But want to be limited by theory, the lipophilic substituent is thought to block the compounds of the invention from the enzymatic degradation to enhance the half-life of the compound in combination with the albumin in the blood stream. Further, for example, it can control the effect of the compound to the glucagon receptor and / or GLP-1 receptor.

In certain embodiments only one amino acid side chain is coupled to a lipophilic substituent. In other embodiments the two amino acid side chains are respectively coupled to the lipophilic substituent. In another embodiment the three or more amino acid side chain is coupled to each of the lipophilic substituent. In this case, a compound containing two or more lipophilic substituents, which may be the same or different.

Lipophilic substituent Z^OneIt is or can be covalently attached to one element in the amino acid side chains, or alternatively the spacer Z²It may be bonded to the amino acid side chains by.

The term "the combination" (conjugated) is used herein to describe a structural relationship between a physical attachment site or a single identifiable chemical, and these areas. It is not to take to imply any particular synthesis method.

Spacer Z²If it is present, and is used to provide space between the compound and a lipophilic part.

Lipophilic substituent is attached to the amino acid side chain or an ester, sulfonyl ester, thioesters, can be attached to the spacer via an amide or sulfonamide. Therefore, preferably, to be understood as forming a part of the lipophilic substituent is an acyl group, the sulfonyl group, the N atom, O atom or S and contain atoms which ester, sulfonyl ester, thioester, amide, or sulfonamide will. Preferably, the acyl groups in the lipophilic substituent forms part of an amide or ester with the amino acid side chain or spacer.

Lipophilic substituent may contain from 10 to 24 C atoms, for example 10 to 22 C atoms, for example, a hydrocarbon chain having 10 to 20 C atoms. Preferably, it has an at least 11 C atoms, which have a 18 C atoms or less, also preferably used. For example, the hydrocarbon chain of 12, 13, 14, 15, 16, and may contain 17 or 18 carbon atoms. The hydrocarbon chain may be linear or branched may also be saturated or unsaturated. From the above description, a hydrocarbon chain is preferably to be understood that the optionally substituted amino acid side chain or spacer, for example, an acyl group, the sulfonyl group, the N atom, O atom or a region which forms part of the attachment to the S atom. Most preferably, the hydrocarbon chain is substituted with acyl, and thus is a hydrocarbon chain alkanoyl group, for example, dodecanoyl, 2-butyl octanoyl, tetra-decanoyl, hexadecanoyl, decanoyl heptadecyl, octadecyl or eicosyl decanoyl Sano may be part of a group.

As mentioned above, the lipophilic substituent Z^OneZ is a spacer²It may be bonded to the amino acid side chains by. When present, the spacer is attached to the lipophilic substituent and the amino acid side chain. The spacer is an ester, sulfonyl ester, thioester, amide or sulfonamide by independently may be attached to the amino acid side chain and the lipophilic substituent. Thus, acyl, sulfonyl, N atom, O atom or S atom selected from the two areas may comprise independently. Spacers are linear C_One_-10 More preferably a hydrocarbon chain or a linear C_One_-5 It may be formed of a hydrocarbon chain. In addition, the spacers are C_One_-6 Alkyl, C_One_-6 Alkylamine, C_One_-6 Alkyl, hydroxy and C_One_-6 It may be substituted with one or more substituents selected from alkyl carboxy.

The spacer may for example be any of the naturally occurring date of the glass or unnatural amino acid. For example, the spacer is Gly, Pro, Ala, Val, Leu, Ile, Met, Cys, Phe, Tyr, Trp, His, Lys, Arg, Gln, Asn, α-Glu, β-Glu, ε-Lys, Asp, Ser, Thr, Gaba, Aib, b-Ala (i.e. 3-amino-propanoyl), 4-amino nobuta Russo, 5 acetaminophen other Russo, 6-amino-hexanoyl, 7-amino-heptanoic Russo, 8-amino- octanoyl, 9-amino Nono nano-day, 10-amino-8-amino-3,6-decanoyl or may be a cup of de-octanoyl. In a specific embodiment, the spacer is Glu, γ-Glu, ε-Lys, β-Ala (i.e. 3-amino-propanoyl), 4-amino nobuta Russo, 8-amino or 8-amino -3,6-oxazol Russo - The dioxa octanoyl. In the present invention, γ-Glu Glu and isopropyl are used interchangeably.

Amino acid side chain is a lipophilic substituent combination is Glu, Lys, Ser, Cys, Dbu, Dpr or the side chain of the Orn residue. For example, this may be a side chain of Lys, Glu, or Cys residue. When two or more side-chain has a lipophilic substituent, these may be selected independently from the residue. Thus, amino acid side chains or may include a spacer for forming a lipid substituent parent ester, sulfonyl ester, thioester, amide, or sulfonamide, carboxyl, hydroxyl, thiol, an amide, or an amine group.

Lipophilic portion Z^One And a spacer Z²Examples of the lipophilic substituent is represented by the following general formula containing.

<img id="i0001" he="65" wi="124" file="pct00001.tif" img-format="tif" />

Wherein the Lys residue from the peptide of formula (I) is γ-Glu side chain spacer via an amide bond (Z²) To be covalently attached to. Hexadecanoyl group (Z^One) Are covalently attached to the spacer through the γ-Glu is an amide bond. If such a combination of a lipophilic part and a spacer coupled to the Lys residue, for example the specific compound represented by the following formula can be referred to as a short notation K (hexadecanoyl -γ-Glu). γ-Glu may also be referred to as iso-Glu also hexadecanoyl group can be mentioned as a palmitoyl group. Therefore, the short notation (hexadecanoyl -γ-Glu) is, for example, International Patent Publication PCT / GB2008 / 004121 abbreviated notation as used in ((iso Glu (Palm)) or (iso Glu (palmitoyl)) equal to It will be obvious that.

Skilled in the art will recognize the appropriate technique for the preparation of compounds of the present invention. Examples of suitable chemicals are published international application, WO 98/08871, WO 00/55184, WO 00/55119, Madsen, etc. (J. Med Chem.. 2007, 50, 6126-32), and the like Knudsen. 2000 (J. Med Chem. 43, 1664-1669) for the reference.

PEG screen and / or acylating a short half-life (T_One_/2) A, which causes a burst (burst) increase in GluGLP-1 agonist concentration. Therefore, glucagon receptor is exposed to receive a burst once daily throughout the treatment period (or double) glucagon receptor activating action (agonism).

But not limited to any theory, the burst is repeated on the glucagon receptor activation causes of action GluR exposure is chaos in lipids and free fatty acids transported between the liver and adipose tissue, and fat accumulation in the liver as a result.

GluR exposed to the constant action of glucagon receptor activation will block the accumulation of fat in the liver. Therefore, if you shorten the process of glucagon or double GluGLP-1 agonist as a repetitive action and glucagon was discovered that fat accumulation in the liver due to extensions (Chan, etc., 1984. Exp. Mol. Path. 40, 320-327).

But the cause when processing double GluGLP-1 agonist repeatedly acting long changes in liver size in normal weight and destination (zoom (enlarge) or reduced (shrunken)), to normalize liver lipid content (Day, etc., 2009; Nat. Chem.Biol. 5, 749-57).

<u>efficacy</u>

GPL-1 or glucagon (Glu) to when combining related compounds used as an indication of the agonist activity on the receptor, but, in general, to use a biological assay for measuring the delivery within the information cell, causing to combine the compounds to relevant receptor desirable. For example, activation of the glucagon receptor by glucagon agonist will promote cell cyclic (cyclic) AMP (cAMP) formation. Similarly, activation of the GLP-1 receptor on GPL-1 agonists will promote cell cAMP formation. Therefore, these two cAMP in appropriate cells expressing a receptor production is used to search the relevant receptor activation. If each type of the cells using an appropriate pair of expressing a receptor, but which do not express the others, the agonist activity of the two types of receptors can be used to determine.

The technician will know the types of analysis are also suitable embodiments are provided below. GLP-1 receptor and / or the glucagon receptor may have the sequence of the receptor, such as those described in the embodiments. For example, the method is the primary accession number GI: 4503947 (NP_000151.1) having a human glucagon receptor (glucagon - R) and / or the primary accession number GI: 166795283 (NP_002053.3) having a human glucagon-like peptide-1 receptor (GLP-1 R) and can be made using. (When referring to the sequence of the precursor protein and analyzed it will be appreciated that to take advantage of the mature protein without a signal sequence).

EC₅₀ Value may be used as a measurement value of an agonist effect from a given receptor. EC₅₀ Value is a measure of the concentration of compound required to achieve half of the maximum activity of the compound in a particular analysis. Thus, for example, EC of natural glucagon in a special analysis₅₀ [GLP-1R] lower than the EC₅₀ [GLP-1R] This compound may be regarded as having a having a higher effect than glucagon in GLP-1R.

The compounds described herein typically dual agonists Glu-GLP-1, that is, they may promote the cAMP formation in the glucagon receptor and GLP-1R both. And the promotion of the respective receptors can be determined by independent analysis, it may be compared to each other later.

EC of the glucagon receptor₅₀ Value (EC₅₀ [Glucagon - R]) of the GPL-1 receptor for a given compound EC₅₀ Value (EC₅₀ [GLP-1 R]) and by comparison, it can be measured relative glucagon selectivity (%) of the compound:

Glucagon -R relative selectivity [compound] = (1 / EC₅₀ [Glucagon -R]) x100% / (1 / EC₅₀ [Glucagon -R] + 1 / EC₅₀ [GLP-1 R])

Similarly, the relative selectivity nor GLP-1R can be measured:

GLP-1R relatively selective [compound] = (1 / EC₅₀ [GLP-1 R]) x 100% / (1 / EC₅₀ [Glucagon -R] + 1 / EC₅₀ [GLP-1 R])

The relative selectivity of the compounds makes it possible to compare their effects on GLP-1 or glucagon receptor directly and its effect on the other receptors. For example, if the relative selectivity of the GLP-1 compound higher, it is more effective for the compounds as compared to GLP-1 receptor and the glucagon receptor.

Using the analysis described below, we have found that GLP-1 has a relative selectivity of human glucagon is about 5%.

The compounds of the present invention therefore has a higher relative selectivity than human glucagon GLP-1R, in particular the level of glucagon -R agonist activity, the compound is in the GLP-1R agonist activity (ie GLP-1 receptor in a higher level than glucagon It will exhibit greater effect) a. Absolute effect of a particular compound in the glucagon and GLP-1 receptor, as long as the proper relative GLP-1R selectivity is achieved, either higher or lower than that of natural human glucagon, it will be understood that lower or substantially equal to this.

Nevertheless, the compounds of the present invention is lower than human glucagon EC₅₀ You may have a [GLP-1 R]. This compound is lower than glucagon EC₅₀ But you can have a [GLP-1R], more than 10 times the human glucagon, less than 10 times the human glucagon, less than five times higher than the human glucagon, EC 2-fold higher or lower than the human glucagon₅₀ You can keep the [glucagon -R].

Glucagon -R and of certain compounds for the GLP-1R EC₅₀It may be desirable that it be less than 1 nM.

The compounds of the present invention is less than twice that of human glucagon EC₅₀ It may have [glucagon -R]. The compound is less than twice that of human glucagon article also human Luca Gon of less than half, less than 5 of one of the human glucagon, or less than one tenth of the human glucagon EC₅₀ You may have a [GLP-1 R].

The relative selectivity of the GLP-1 compound may be less than 95% and more than 5%. For example, the compound is 5-20%, 10-30%, 20-50%, 30-70%, or the relative selectivity of 50 - 80%, or 30-50%, 40-60%, 50-70 It may have a relative selectivity% or 75-95%.

<u>Therapeutic</u><u> use</u>

The compounds of the present invention can provide an attractive treatment option for metabolic disorders, including obesity and diabetes (diabetes).

Diabetes is a metabolic disease characterized by including hyperglycemia resulting from insulin secretion, insulin action, or both of the deficiency. Acute symptoms of diabetes include excessive production of urine, thirst saenggim and compensatory increase in water intake, vision problems, unexplained weight loss, lethargy, and changes in energy metabolism. Chronic hyperglycemia of diabetes is long-term damage, dysfunction and failure of various organs, can significantly associated with the eyes, kidneys, nerves, heart and blood vessels. Diabetes is characterized by the Pathogenetic by default classified as Type 1 diabetes, type 2 diabetes and gestational diabetes.

Type 1 diabetes represents a 5 to 10% of all diabetes cases also caused by auto-immune destruction of insulin-secreting pancreatic β- cells.

Type 2 diabetes is a result of a metabolic disorder in the composite set indicates the addition of 90-95% of diabetes cases. Type 2 diabetes is a result of endogenous insulin production is insufficient to maintain the plasma glucose levels as a diagnostic threshold value or less.

Pregnancy diabetes refers to any degree of glucose intolerance in pregnancy identified.

Jeonjeung diabetes (pre-diabetes) is a fasting glucose and impaired glucose damaged including resistance and also shows what happens when the blood glucose level is elevated, but not more than the defined level for the clinical diagnosis of diabetes.

Type 2 diabetes, and most people with diabetes jeonjeung, for example the additional metabolic risk factors, central obesity (excessive fat tissue around the abdominal), atherogenic lipid disorders (blood lipid disorders such as high triglycerides, low HDL cholesterol and / or high LDL cholesterol, they raise the plaque accumulation in the arterial walls), blood pressure (hypertension), blood clot formation promoting conditions (eg, high blood fibrinogen or plasminogen activator inhibitor-1), and inflammation promoting state (e. g., C- reactive protein, elevated serum) due to the higher risk in the sign of a state of increased morbidity and mortality.

Conversely, obesity is jeonjeung diabetes, type 2 diabetes, as well as, for example, gives an increased risk of developing certain types of cancer, gallbladder disease and obstructive sleep apnea.

Lipid disorders (dyslipidaemia) is associated with an increased risk of cardiovascular disease. High-density lipoprotein (HDL) is the clinical importance is due to the presence of the inverse correlation between the (inverse correlation) plasma HDL levels and risk of atherosclerotic disease. Most of the cholesterol in the atherosclerotic plaque is stored derived from LDL and thus increase the concentration of low density lipoprotein (LDL) is closely related to atherosclerosis. The HDL / LDL ratio is particularly shown clinical risk of atherosclerosis and coronary artery atherosclerosis.

But want to be limited to any particular theory, it is believed that the compounds of the invention are dual acting GluGLP-1 agonist. Dual agonist (dual agonist) is having an effect on the lipid metabolism of GLP-1, for example, in the blood glucose levels and food intake for, it is possible to combine the effects of glucagon. Thus, it facilitates the removal of excess fatty tissue, and induce weight loss, and persistent, and may serve to promote hypercholesterolemia (glycaemic) per control. Double GluGLP-1 agonist may also act to reduce the cardiovascular risk factors such as high cholesterol and LDL- cholesterol.

Thus compounds of the invention prevent the increase in weight (for example, loss of appetite, feeding, food intake, caloric intake, and / or by regulation of energy consumption), promote weight loss, reduce excess weight to or treatment of obesity It can be used as a pharmaceutical preparation for. Here are morbidly obese, but obesity is of course not limited to such diseases and related health symptoms, such as, including obesity-related inflammation, gallbladder disease, obesity and obesity-induced sleep apnea. The compounds of the invention are also insulin resistance, glucose intolerance, diabetes jeonjeung, fasting plasma glucose increase, type 2 diabetes, hypertension, lipid disorders (or their metabolic risk factors, a combination of), atherosclerosis, arteriosclerosis, coronary heart disease, peripheral arterial It can be used for the treatment of diseases and stroke. These are all symptoms that can be associated with obesity. However, the effects of the compounds of the invention for these symptoms is or can be adjusted in whole or in part through the effect of the weight, or else can be independent.

<u>A pharmaceutical composition</u>

Compound, or a salt thereof of the present invention, in a pharmaceutically acceptable carrier, typically can be formulated as a compound or a pharmaceutical composition prepared for storage or administration that include a salt of the invention a therapeutically effective amount of .

The compounds of the present invention a therapeutically effective amount will vary depending upon the route of administration, the treatment type of the target mammal and the physical characteristics of the specific mammal being considered. On the determination of the relationship of these factors and such amounts it is well known to the skilled practitioner in the medical field. This amount and the method of administration can be adjusted to achieve optimal efficacy also well known to those skilled in the medical field, it will vary according to the body weight, diet, medication, and other factors in combination. The size of the dose for human use and administration plans to be guided by the results obtained are in accordance with the present invention can be also confirmed by properly designed clinical trials.

The effective dosage and treatment protocol from a low dose in laboratory animals and then navigate through the effect, by changing the mode of administration it may be determined systematically by a conventional method to increase the dose. Many factors may be considered by a clinician when determining an optimal dosage for a given object. These considerations are known to those skilled.

The term "pharmaceutically acceptable carrier" includes any of the standard pharmaceutical carriers. A pharmaceutically acceptable carrier for therapeutic use are well known in the pharmaceutical field, as well, for example, Remington's Pharmaceutical Sciences, Mack Publishing Co. (AR Gennaro edit. 1985) have been described in. For example, in a slightly acidic or physiological pH sterile saline and phosphate-buffered saline it can be used. pH buffer is phosphate, citrate, acetate, tris / hydroxymethyl) aminomethane (TRIS), N-tris (hydroxymethyl) methyl-3-aminopropane sulfonic acid (TAPS), ammonium bicarbonate, diethanolamine, histidine days and, a buffer may be preferred are ginil, lysine, or acetate or mixtures thereof. The term includes any drug listed in the United States Pharmacopeia for use in animals, including human in addition.

"Pharmaceutically acceptable salts" denotes salts of the compound term. Salts include, for example, pharmaceutically acceptable salts include acid addition salts and basic salts. Acid addition salts include hydrochloride salts, citrate salts and acetate salts. Basic salts such as alkaline earth metal is an alkali metal, an alkali metal cation such as calcium for example, the sodium, potassium, and ammonium ions⁺N (R³)₃(R⁴) Where R³ And R⁴ It is independently an optionally substituted C_One_-6-alkyl, Optionally substituted C₂_-6-alkenyl, Optionally substituted aryl, or optionally comprises a salt selected from those showing a substituted heteroaryl. Other pharmaceutically acceptable salts such as the "Remington's Pharmaceutical Sciences", 17th edition. Ed. Alfonso R. Gennaro (Ed.), Are described in Mark Publishing Company, Easton, PA, USA, 1985 and more recent editions, and pharmaceutical technology advance (Encyclopaedia of Pharmaceutical Technology).

"Treatment" is an approach for obtaining beneficial or desired clinical results. For the purposes of this invention, beneficial or desired clinical results include, but are not limited to, whether or not possible, whether detectable, relief of symptoms, decrease in disease extent, of disease stabilization (i.e., not worsening) state of disease progression, delay or slower, improved or palliation of the disease state, and a car (or any whole or in part). "Treatment" can also mean a longer life compared to expected survival if not treated. "Treatment" is an intervention performed with the intention of preventing the development of disease, the pathology of the disease or change. Thus, "treatment" refers to therapeutic treatment and prophylactic or room noted action. Those in need of treatment are those already with the disorder include those who try to prevent the disease as well. Treatment is not meant as compared pathology or condition (for example, weight gain, high blood sugar) is not a treatment means for reducing or inhibiting the growth of, and also necessarily imply a complete stop of the associated symptoms.

The pharmaceutical compositions may be in unit dosage form. In such form, the composition is divided into unit doses containing appropriate quantities of the active component. The unit dosage form is a package formulation, the package containing a formulation of the discrete quantities of, for example, can be a packaged tablets, capsules, and powders in vials or ampoules form. Unit dosage forms also a capsule, may be a cachet, or tablet itself, or may be any suitable number of these packaged form. This can provide a pen type single dosage injectable form, for example. The compositions may be formulated in any suitable administration route and means. The carrier or diluent is pharmaceutically acceptable, including oral, rectal, nasal, or parenteral (including subcutaneous, intramuscular, intravenous, intradermal, and transdermal including) those used in formulations suitable for administration. The formulations may be present in a convenient unit dosage form can also be prepared by any of the methods well known in the art of pharmacy.

Subcutaneous or transdermal administration patterns can be particularly suitable for the compounds described herein.

<u>Combination therapy</u>

The compounds of the invention may be administered, diabetes, obesity, lipid disorders, or as part of a combination therapy for the treatment of high blood pressure.

In this case, the two active agents can be provided as part of the same pharmaceutical formulation also together or separately also as a separate dosage form.

Therefore, the compound (or salt thereof) of the present invention is a therapeutic agent for diabetes, for example, but are not limited to, metformin, sulfonylureas, nateglinide (glinide), DPP-IV inhibitors, glitazones (glitazone), or insulin in combination with and it can be used. In a preferred embodiment, the compounds or salts thereof in order to achieve adequate glycemic control, insulin, DPP-IV inhibitors, sulfonylurea, or metformin, a sulfonylurea or metformin used in particular in combination. In a more preferred embodiment, the compound or a salt thereof is used in combination with insulin or insulin analogue to achieve adequate glycemic control. Examples of insulin analogues include, but are not limited to, Lantus (Lantus), look no feed (Novorapid), Huma log (Humalog), Novo Mix (Novomix), evil Trapani plate HM (Actraphane HM), L'Mir (Levemir) and Bahia including the driver (Apidra).

While the compound or the salt thereof is not limited for further anti-obesity agent, e.g., glucagon-like peptide receptor 1 agonists, peptide YY or an analogue, cannabinoid receptor antagonists, lipase inhibitors, melanocortin receptor 4 agonist, or a melamine It can be used in combination with concentrating hormone receptor antagonists.

The compound or a salt thereof is, for example anti-hypertensive, but are not limited to, angiotensin-converting enzyme inhibitors, angiotensin II receptor blockers, diuretics, beta-blockers may be used, or in combination with calcium channel blockers.

While the compound or the salt thereof is not limited to lipid disorder therapeutic agent, for example, statins (statin), fibrate (fibrate), New Age (niacin) and / or can be used in combination with a cholesterol absorption inhibitor.

<b>method</b>

<u>Acylated</u><u> In general synthesis of glucagon analogs</u>

Solid phase peptide synthesis, using standard Fmoc chemistry was carried out in a CEM Liberty peptide synthesizer. TentaGel S Ram resin (1g; 0.25 mmol / g) is swelled in and NMP (10 ml) prior to use, with the DCM and NMP was transferred between the tube and the reaction vessel.

Coupling:

NMP / DMF / DCM (1: 1: 1; 0.2 M; 5 ml) in the amino acid Fmoc- HATU / NMP (0.5 M; 2 ml) and DIPEA / NMP (2.0 M; 1 ml) and with a CEM Discover microwave unit with a resin It was added. Coupling a mixture of the saturated nitrogen through the mixture while heating period of 5 minutes at 75 &amp; lt; 0 &amp; gt; C. Then the resin was washed with NMP (4 x 10 ml).

Deprotection

Piperidine / NMP (20%; 10 ml) is added to the resin for the initial de-protected, and the mixture was microwave; was heated with (30 seconds 40 &amp; lt; 0 &amp; gt; C). The reaction vessel is drained and the piperidine / NMP in the second portion; were added and heated again (20% 10 ml) (75 &amp; lt; 0 &amp; gt; C; 3 minutes). Then the resin was washed with NMP (6 x 10 ml).

Chain acylated

Fmoc-Lys (ivDde) ​​-OH or an amino acid having a side chain protecting group or alternatively introduced at the position perpendicular to the acylation. N- terminal of the peptide backbone of the following Boc₂Using Boc-protected or O or alternatively at the end coupled with the Boc-protected Boc-protected amino acid. Although the peptide is attached to the resin, and side chain protecting group is orthogonal with hydrazine hydrate (2-4%), freshly prepared in NMP was cut for 2 x 15 selectively time. US-protected lysine side chain is first Fmoc-Glu-OtBu or another spacer amino acid and coupled to, deprotection with piperidine and using the peptide coupling method as described above was acylated with a lipophilic region.

Used abbreviations are:

ivDde: 1- (4,4- dimethyl-2,6-dioxo-cyclohexylidene) 3-methyl-butyl

Dde: 1- (4,4- dimethyl-2,6-dioxo-cyclohexylidene) ethyl

DCM: dichloromethane

DMF: <i>N, N</i>-Dimethylformamide

DIPEA: diisopropylethylamine

EtOH: ethanol

Et₂O: diethyl ether

HATU:　<i>N</i>- [(Dimethylamino) -1<i>H</i>-1,2,3-triazoles [4,5<i>b</i>] Pyridin-1-ylmethylene] -<i>N</i>Methyl methane aminium hexafluorophosphate <i>N</i>- Oxide

MeCN: acetonitrile

NMP:　 <i>N</i>- Methylpyrrolidone

TFA: trifluoroacetic acid

TIS: triisopropyl silane

cutting:

Resins EtOH (3 x 10 ml) and Et₂Washed with O (3 x 10 ml) and dried to constant weight at room temperature (RT). Crude (crude) peptides TFA / TIS / water (95 / 2.5 / 2.5; 40 ml of, 2 hours; room temperature) were cut from the resin by treatment with. Most of the TFA was removed under reduced pressure and the crude peptide is precipitated and washed three times with diethyl ether and dried to constant weight at room temperature.

HPLC purification of the crude peptides

Crude peptide C-18 column (5 cm; 10 mu m), and fraction collortor equipped purified PerSeptive Biosystems VISION Workstation at least 90% by preparative reverse phase HPLC using the buffer A (0.1% TFA, aqueous) and buffer B with a gradient (gradient) in (0.1% TFA, MeCN 90%, aqueous) 35 ml / min were performed. Fractions are analyzed by HPLC and MS analysis for, and the relevant fractions were mixed and lyophilized. The final product was characterized by HPLC and MS.

<u>Human glucagon - and </u><u>GLP</u><u>- Generation of a cell line expressing receptors</u>

Human glucagon receptor (glucagon -R) (primary accession number P47871) or human glucagon-like peptide-1 receptor (GLP-1R) cDNA encoding the (primary accession number P43220) is a cDNA clone BC104854 (MGC: 132514 / IMAGE: 8143857) or BC112126 (MGC: 138331 / IMAGE: 8327594) were each cloned from. The DNA coding for glucagon or GLP-1 R -R using primers coding for the terminal restriction sites for sub-cloning was amplified by PCR. 5'-end primer was encrypted by adding approximately kojak consensus sequence (near Kozak consensus sequence) to ensure efficient translation. Glucagon suitability of the DNA coding for the GLP-1R is -R and was confirmed by DNA sequencing. Glucagon -R or PCR product coding for the GLP-1R is neomycin (G418) were subcloned into mammalian expression vectors containing the resistance marker.

Glucagon or GLP-1 R -R mammalian expression vector encoding a trans faction was in HEK293 cells by standard calcium phosphate method trans faction. Trans faction seeded 48 hours the cells to limited dilution cloning and were selected with 1 mg / ml G418 in the culture medium. Three weeks after collecting the glucagon and GLP-1 R expressing cells -R 12 survival colon, proliferation and tested by glucagon and GLP-1 R -R efficacy analysis as described below. One of -R glucagon expression of GLP-1R expression clones and one clone was selected for compound profile.

<u>Glucagon receptor, and </u><u>GLP</u><u>1 receptor efficacy analysis</u>

Human glucagon - R, or HEK293 cells expressing the human GLP-1 R and is seeded with 40,000 cells per well of 0.01% in the trace amount of the 96-well plate coated with poly -L- lysine, and cultured in the growth medium one day 100㎕ It was grown. The analysis day, the growth medium was removed and the cells were washed once with 200㎕ tie rod buffer (Tyrode buffer). Increasing concentrations of test peptide, 100 mM IBMX, and 6 mM 100㎕ tie rod in the cell buffer containing glucose and incubated for 15 minutes at 37 &amp; lt; 0 &amp; gt; C. 25 mu l 0.5 M HCl was added to stop the reaction and incubated 60 minutes on ice. cAMP content of Perkin-Elmer was evaluated using FlashPlate® cAMP Kit from (Perkin-Elmer). Compared to the reference compound (glucagon and GLP-1) EC₅₀ And relative potency was assessed by computer-assisted adjustment curve.

<u>After subcutaneous administration in mouse plasma </u><u>Peptide</u><u>Glu</u><u>-</u><u>GLP1</u><u> Biochemical analysis of a screening method for the quantification of the agent</u>

Mice were subcutaneously (SC) administration at a dose of 100 nmol / kg. Sacrificing the mouse and the blood following points: 0.5, 2, 4, 6, 16, and collected at 24 hours. Plasma samples are analyzed using a protein precipitation, and then were analyzed by solid phase extraction (SPE) and liquid chromatography-mass spectrometry (LC-MS).

<u>High fat intake </u><u>C57Bl</u><u>/ 6J mice and </u><u>HbA1c</u><u>db</u><u>/</u><u>db</u><u> Oral glucose tolerance test in mice (</u><u>OGTT</u><u>), Blood lipids and body weight</u>

Male mice (long-term intake of high-fat C57Bl / 6J, short-term high-fat intake C57Bl / 6J and db / db) food and free access to the water was purified. These light, temperature and humidity controlled room (12-hour light: 12 hour dark cycle, 2000/0800 light on / off at a time; 24 &amp; lt; 0 &amp; gt; C; 50% relative humidity) and housed in groups of 5 to 6.

Animals were injected subcutaneously over a period of three days 100㎕ vehicle (once a day) to treat and purify the animal injections. Blood samples were taken from the tail end or from the eye. Animals were randomized prior to treatment.

Mice GluGLP-1 agonist or vehicle (injection volume = 2.5 ml / kg) were treated with daily subcutaneous twice. Through the overall analysis, the weight is recorded daily and used for administration of the peptides of the correction weight dose. Peptide solution was freshly prepared just prior to administration.

Oral glucose tolerance test (OGTT) was performed shortly after the animals fasted object. In order to prevent disturbing the food intake, the animals are fasted during the OGTT was maintained. After peptide administration, blood samples were taken early. It was then returned to the phosphate buffer (pH = 7.4) provided the glucose (1 g / kg) dissolved in the oral dose, and (5 ml / kg), animals their home cages (t = 0). All blood glucose (BG) is t = 15 minute, 30 minute t =, t = 60 minute, t = 90 minutes and was measured at t = 120 minutes.

BG concentration of blood (&lt;5 mu l; Elite Autoanalyser, Bayer, Denmark) according to the manufacturer's instructions in the drop was analyzed by the immobilized glucose oxidase method using.

<u>HbA1c</u><u> Measure</u>

It can access a long-term effect of the compound on the glucose level of the object by measuring the level of hemoglobin A1C (HbA1c). HbA1c is a glycated form of hemoglobin, and its level in the cell reflects the average glucose level of the cell is exposed during his lifetime. In the mouse, HbA1c is relevant biomarkers for the previous 4 weeks the average blood sugar level because the conversion is restricted to red blood cells HbA1c life of about 47 days (Abbrecht &amp; Littell, 1972;.. J. Appl Physiol 32, 443- 445).

HbA1c is measured by the HbA1c in the sample react with the soluble antigen, wherein -HbA1c - based on nephelometric inhibition immunoassay (TINIA) to form an antibody composite. The addition of poly-heptene is reacted with an excess of antibody, wherein -HbA1c insoluble antibody-complex to form a poly-heptane, which can be measured by Turbidimetry 2008. The free hemoglobin in a hemolyzed sample is converted into derivatives having a characteristic absorption spectrum, which is pre-determined in the incubation with dichromate. The final result is expressed as% HbA1c in total hemoglobin (Cobas® application notes A1C-2).

<u>Cholesterol level measurement</u>

The enzyme assay is a colorimetric method. In the presence of magnesium ions, dextran sulfate microns LDL, VLDLA and keys that are resistant to PEG- modified enzyme (chylomicron) and water-soluble complex it is selectively formed. HDL cholesterol is enzymatically measured by the cholesterol esterase and cholesterol oxidase is coupled with PEG at the amino group. The cholesterol esters are quantitatively destroyed in Colo free sterols and fatty acids. Call-LESS HDL cholesterol (choles) -4- en-3-one and H₂O₂Enzymatic oxidation and is also formed by H₂O₂It is measured by a colorimetric measurement (Cobas® application notes HDLC3).

Direct measurement of the LDL is used in selective micelles and lipid-soluble compounds and interaction of the glycoprotein of the LDL (VLDL and key azithromycin) by the non-ionic detergent. The combination of the sugar compound and the detergent is enable selective measurement of LDL in the blood plasma. Test principle is the same as that of cholesterol and HDL, but due to the sugar and detergent is only LDL- cholesterol ester to free cholesterol and fatty acids only destruction. Free cholesterol is then oxidized and formed H₂O₂Quantitative measures include colorimetric (Application Note LDL_C, Cobas®).

<b><u>High fat intake </u></b><b><u>C57BL</u></b><b><u>/ 6J mice from weight gain </u></b>

6-week old C57Bl / 6J male mice eat high fat diet (HFD) (D12492, Research Diet Inc., New Brunswick, USA), and free access to water, were purified by 4 weeks in their new environment. Animals by subcutaneous injection for a period of three days excipients 100㎕, before the start of the peptide treatment was fined for animal handling and scanning. Mouse exendin-4, compound 3, compound 6, compound 7, compound 8, compound 11 and compound 12 were treated twice daily with subcutaneous or excipient. Through the overall analysis, which was used to record weight every day, administration of the peptides of the correction weight dose. All animals were sacrificed on the same day by the neck vertebrae dislocation.

<b>High fat intake </b><b>C57BL</b><b>/ 6J mice at 2, 4, 6, 8, 10 and 12 hours after oral glucose administration</b><b>tolerance</b>

6-week old C57Bl / 6J male mice to a high fat diet (D12492, Research Diet Inc., New Brunswick, USA), and to access freely to water was purified in their new environment. Animals by subcutaneous injection for a period of three days as an excipient, was fined for animal handling and scanning. Taking a blood sample from the end of the tail was measured glucose. Blood glucose (mM) concentration of the drop according to the manufacturer's manual of blood (&lt;5 mu l; Contour Autoanalyser, Bayer, Denmark) was analyzed by using immobilized glucose oxidase method. After four weeks of high fat diet, and the basis weight of the animal, body weight it was used for administration of the peptides of the correction weight dose. Oral glucose tolerance test (OGTT) is the object 4 weeks fasted animals were then performed. Single peptide or vehicle administration. 2, 4, 6, 8, 10 and at 12 hours after, an initial blood sample was taken (t = 0 min). Immediately after, providing the glucose (1g / kg) of the oral dose and the animals were returned to their home cages (t = 0). BG level T = 15 minute, t = 30 minute, T = measured at 60 minutes and T = 90 minutes. Immediately after blood sampling, all animals is CO₂ After spinal anesthesia were sacrificed by neck dislocation.

<b>Children and the elderly Obesity dry </b><b>C57BL</b><b>/ 6J mice food intake in</b>

C57BL / 6J mice are treated with high-fat diet for 11 days to C57BL / 6J mice are treated for 52 weeks with High Fat Diet.

3 days before analysis, mice were transferred to the individual basis weight of the cage. Before analysis 4 days, mice were treated daily by purification and treatment by subcutaneous injection. The food was removed at 20:00 the day before the experiment. The basis weight of mice in the experiment day 0 and T = time (8:00) and t = 12 hours (20:00), exendin-4, compound 7, or were treated with subcutaneous administration of the vehicle. Immediately after the treatment (t = 0), In the mouse the basis weight of the food in advance, and the accumulated food intake is t = 1, 2, 4, 8, and basis weight of the remaining food was measured after 12 and 24 hours. After t = basis weight of food and animals in 24 hours, mice were sacrificed by dislocation of the neck vertebrae.

<b>Hepatocellular </b><b>cAMP</b><b> Formation</b>

<b>Experimental Procedure</b>

Walkersvill Lonza, Inc. The primary human hepatocytes are obtained from a carefully washed and incubated in 100 μM IBMX and 0.1% 15 minutes 37 &amp; lt; 0 &amp; gt; C to dissolve the peptide in the buffer TB in TB buffer supplemented with casein. Prior to addition to the cells, the peptide was warmed back to 37 &amp; lt; 0 &amp; gt; C diluted solution. The reaction was stopped by the addition of 25 mu l of ice-cold 0.5 M HCl, the cells were incubated for 60 minutes on ice. CAMP content in the well of a 96-well microtiter 'lashPlates' covered with glare and anti-antibody -cAMP, 75㎕ sodium acetate buffer, was determined by addition of an acid extract of 25㎕ from well to pH 6.2. 10μCi [of 100㎕ to each well¹²⁵I] cAMP, was added to the solution, incubated overnight at 4 &amp; lt; 0 &amp; gt; C and the plate, coupled to the flash plate (FlashPlate) [¹²⁵I] The amount of cAMP is a program on TopCount NXT "[¹²⁵I] cAMP flash plates were calculated using the 10 minutes "a.

Peptide was tested in a concentration range of 0.1 to 1000 nM.

<b>Data Analysis and Statistics</b>

The amount of cAMP produced by the cells was calculated by extrapolation to the cAMP standard curve.

EC₅₀ Value was evaluated by adjusting the cAMP data, according to the following equation using the Sigma Plot:

<img id="i0002" he="14" wi="133" file="pct00002.tif" img-format="tif" />

The invention is further illustrated by the following examples.

<b>C57BL</b><b>/ 6J mouse liver weight / body weight</b>

Mouse Compound 1 and Compound 11 (2 doses of 0.5 and 5 in nmol / kg), or 2 weeks was subcutaneously twice daily treatment with a vehicle. Through the overall analysis, body weight was recorded daily and used for administration of the peptides of the correction weight dose. It was sacrificed on day basis weight was taken out of the liver.

<b>Example</b>

<b>Example</b><b> 1: Compound Synthesis </b><b>Peptide</b><b> characteristic</b>

Synthesis example:

Compound 9 is TentaGel S Ram resin as described above (1,17 g; 0.23 mmol / g) was measured in CEM and free peptide synthesizer using the Fmoc- chemistry (Liberty Peptide Synthesizer). Fmoc-Lys (ivDde)-OH is at position 17 was similar proline using Fmoc-Phe-Thr (. Psi. Me, Me pro)-OH and Fmoc-Asp (OtBu)-Ser (Psi., Me, Me pro) -OH was used in the peptide backbone. After completion of the peptide backbone of the resinous phase, Fmoc- N-terminal Boc group is then cut by hand in DCM₂O (226 mg) and DIEA (54㎕) it was protected with the Boc-. Next ivDde- group is newly prepared hydrazine hydrate / NMP (4%; 2 x 15 min) was cut into. Two building blocks remaining in the glass again CEM peptide synthesizer, Fmoc-Glu-OtBu, and the acid was added to the non-protected hexadecafluorovanadyl lysine side chain.

Peptide was cleaved from the resin as described above, the purification buffer A, (0.1% TFA, aqueous) and buffer B (0.1% TFA, 90% MeCN, water-based) of a mixture of 35 ml / min flow with Gemini-NX column (5 cm, 10 mu m, Cl 8) was carried out on. The product was over 47 minutes from 25% to 65% buffer B and eluted with a linear gradient and fractions (9 ml) was collected in a fraction collector. Related fractions and analyzed by HPLC and MS analysis for, receiving a fraction having at least 95% pure, and was freeze-dried as a white powder. 72mg of the obtained water had a purity of 97% as determined by analytical HPLC was also mass 3697.05 Da as determined by MS (calculated 3696.97 Da).

<b>Example</b><b> 2</b>

<b>GPL</b><b>The efficacy of 1 and glucagon receptor</b>

GluGLP-1 efficacy of the agent is glucagon, and R h and exposed to cells expressing hGLP-1 R with increasing concentrations of the following compounds listed in the acylation was evaluated by measuring cAMP formed as described in the methods.

The results are given in Table 1:

Table 1

<img id="i0003" he="174" wi="165" file="pct00003.jpg" img-format="jpg" />

Table 1a]

<img id="i0004" he="224" wi="165" file="pct00004.jpg" img-format="jpg" />

For compound 28 HH-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl-E) -AAHDFVEWLLSA-NH 2 can also be written as HH-Aib-QGTFTSDYSKYLDS-K (hexadecanoyl -αGlu) -AAHDFVEWLLSA-NH2.

<b>Example</b><b> 3: Pharmacokinetic screening</b>

Pharmacokinetic profile was determined for a variety of acylated compounds. Calculated T_1/2It is shown in Table 2 as compared to (via a misfire) Compound 1.

Table 2

<img id="i0005" he="75" wi="128" file="pct00005.jpg" img-format="jpg" />

All acylated compounds improved compared to compound 1 T_One_/2He had.

Sample pharmacokinetic profile for the compound 13 is shown in Fig.

<b>Example</b><b> 4: </b><b>DIO</b><b> Oral glucose tolerance test in mice</b>

Effect of long-term administration of high fat intake subcutaneously 21 days C57BL / 6J oral glucose tolerance in mice for the compound 11 (10 nmol / kg). High fat mice fasting and fasting blood glucose levels were measured (t = 0) by taking the initial blood sample. Next provide glucose (5 ml / kg of 1g / kg) oral dose of the blood glucose level is t = 30 minute, 60 minute t =, t = 90 minutes and was measured at t = 120 minutes. Compound 11 was significantly improved glucose tolerance (two-way ANOVA). Data represent the mean ± SEM.

<b>Example</b><b> 5:28 days </b><b>db</b><b>/</b><b>db</b><b> In mice </b><b>HbA1c</b>

Diabetic (db / db) mice are treated with vehicle or compound 7 and 4 weeks, HbA1c was measured in whole blood sample (20 mu l) collected from the treated mouse (Cobas® Application Note: A1C-2). The result is shown in Fig. △ HbA1c (%) is from the beginning to process HbA1c (%) in 4 weeks by subtracting his HbA1c (%) was calculated for each mouse. Treatment with Compound 7 is reduced markedly △ HbA1c (%) as compared to vehicle (P = 0.03; Student's t- test).

<b>Example</b><b> 6: Weight Loss</b>

Effect of 21 days for the subcutaneous administration of the compounds 11 weight long-term intake of high-fat C57BL / 6J were measured in mice. C57BI / 6J male mice on high fat diet (HFD) was treated with compound 11 (10nmol / kg) or vehicle (Bid; subcutaneously). Body weight was recorded daily and used for administration of the peptides of the correction weight by the overall dose analysis. Data represent the mean ± SEM in FIG. Compound 11 was significantly reduced body weight (p &lt;0.05).

<b>Example</b><b> 7. Total cholesterol and </b><b>HDL</b><b>/</b><b>LDL</b><b> ratio</b>

Diet-induced obesity (DIO) mice were treated 4 weeks with vehicle or Compound 7, and collecting plasma from the collected blood sample. Total cholesterol, LDL and HDL are each plasma sample: measured in (Cobas® Application Note CHOL2, HDLC3 and LDL_C) and the results are shown in Figs. Compound 7 is treated by the total cholesterol concentration significantly (P &lt;0.0001, Student's t- test) reduction sikyeotgo (Figure 5) was also significantly increased the HDL / LDL-ratio (P &lt;0.0001, Student's t- test) ( 6).

<b>Example</b><b> 8. After the local administration and </b><b>C57BL</b><b>/ 6J mice from weight gain</b>

Exendin-4, Compound 8, Compound 3, Compound 7, Compound 11, Compound 12 and Compound 6 short-term intake of high-fat C57BL / 6J mice the effect of subcutaneously administered 10 yl. C57BI / 6J male mice on high fat diet (HFD) is treated with compound (0.5 and 5 nmol / kg) or vehicle (Bid; subcutaneously). Record weight every day, and was used to analyze the peptide administration over the first half of the weight correction dose. Data represent the mean ± SEM in FIG.

Control peptide (exendin-4), as well as the compound 8 was significantly decreased weight gain in the two doses (0.5 and 5 nmol / kg). Compound 3, Compound 7, Compound 11, and Compound 12 was markedly reduced the weight gain at high doses (5 nmol / kg), low dose (0.5 nmol / kg) was not the (7). Compound 6 is a low dose (0.5 nmol / kg) was reduced significantly in the weight gain.

<b>Example</b><b> 9: fat intake </b><b>C57BL</b><b>/ Dose in 6J 2, 4, 6, 8, 10 and 12 hours after the oral glucose tolerance</b>

Oral glucose tolerance test (OGTT) was performed after 4 hours fasting animal objects. Compound 7 or vehicle administered 2, 4, 6, 8, 10 and 12 hours later, blood samples were taken initially (t = -0 min). Immediately after, it provided the glucose (1 g / kg) oral doses. BG level T = 15 minute, t = 30 minute, T = measured at 60 minutes and T = 90 minutes. Immediately after blood sampling, all animals is CO₂ Following spinal anesthesia were sacrificed by neck dislocation.

The analysis of the compound 7 (10 nmol / kg) administered subcutaneously by a high fat intake C57BL / 6J 2 in the mouse, 4, 6, 8, 10 and 12 hours after the administration of glucose tolerance (oral glucose tolerance as measured by a reduced AUC during the test Do) to thereby significantly increase.

<b>Example</b><b> 10. skinny and obese older children </b><b>C57BL</b><b>/ 6J mouse food intake</b>

C57BL / 6J mice were treated with high-fat diet for 11 days, C57BL / 6J mice are treated for 52 weeks with High Fat Diet.

On the day of the experiment, the mouse, and the basis weight, t = 0 h (8:00), and at t = 12 h (20:00) exendin-4, compound 7, or were treated with a subcutaneous injection of vehicle. Treatment (t = 0) immediately after, In the mouse pre cumulative basis weight of the food and the food intake of t = 1, 2, 4, 8, and basis weight of the remaining food was measured after 12 and 24 hours.

In young mice, dry, compound 7, 0-4, 0-8, statistically significantly food intake 0-12 and 0-24 from the point (p &lt;0.05) reduced. Exendin-4 is 0-2, 0-4, 0-8, statistically significantly the food intake in the point 0-12 and 0-24 (p &lt;0.05) reduced.

In obese mice of advanced age, compound 7 0-2, 0-4, 0-8, statistically significantly the food intake in the point 0-12 and 0-24 (P &lt;0.05) it decreased. Exendin-4 had a statistically significant decrease in food intake at any time point (p &lt;0.05).

<b>Example</b><b> 11:30 Notes about dietary lipids in fat mice </b><b>GluGLP</b><b>-1 Agonist effects of compound 11 administered subcutaneously for 3 weeks</b>

Excipients twice daily for three weeks for the lipid (PBS), 10 nmol / kg exendin-4, or 10 nmol / kg treated with the compound 11 (subcutaneously) three weeks of the mice treated with 30 weeks high fat diet for 30 weeks before treatment effect (Fig. 11). This effect is LDL, HDL and triglycerides (CHO: total cholesterol; HDL: high-density cholesterol; LDL: low-density cholesterol; TRIG: triglycerides; HDL / LDL: HDL and the ratio between LDL).

Compound 11 is cholesterol, HDL, LDL (P &lt;0.001) and triglycerides (P &lt;0.05), and significantly lower the other hand, the ratio of HDL / LDL was increased significantly (P &lt;0.001) (Fig. 11). The ratio of HDL / LDL is considered as a dangerous indication of heart disease. This ratio is higher, the risk of heart attack or other cardiovascular problems in less.

<b>Example</b><b> 12. hepatocellular </b><b>cAMP</b><b> Effect of the compounds on the formation 11</b>

All the tested peptides had a role as full agonists, except for pure GLP-1 agonist exendin-4, and will glue Tide (liraglutide) and with respect to promoting GluR cAMP formation. Ranking of the effect of the compound from Table 1&gt; glucagon&gt; Compound 11&gt; dioxin te Doolin &gt;&gt;&gt; can be observed that exendin-4 and the glue will Tide (Table 9).

Finally, the down-regulation is in contrast to those observed in HEK293 cells hGluR, in a high density E_MAX The cAMP response was observed.

[Table 9]

<img id="i0006" he="73" wi="157" file="pct00006.jpg" img-format="jpg" />

<b>Example</b><b> Processed 13.2 weeks </b><b>C57</b><b> The weight of a healthy mouse liver contrast</b>

A long-acting acylated double GluGLP-1 repeat processing by the agent, for example, compound 11 is acylated via a double GluGLP-1 changes in liver size compared with the agonist compound 1 (close or contract) this does not occur (Fig. 12).

<b>Example</b><b> 14. After 28 days </b><b>db</b><b>/</b><b>db</b><b> In the mouse </b><b>HbA1c</b>

Diabetic (db / db) mice are treated with vehicle or compound 11 and 4 weeks, HbA1c was measured in whole blood sample (20 mu l) collected from the above treated mice (Cobas ® Application Note: A1C-2). The results are shown in Fig. △ HbA1c (%) was subtracted and their HbA1c (%) 4 week time to process from the HbA1c (%) was calculated for each mouse. Treatment with Compound 11 had significantly reduced the △ HbA1c (%) as compared to vehicle (P = 0.03; Student's t- test).

&lt;110&gt; ZEALAND PHARMA A/S &lt;120&gt; ACYLATED GLUCAGON ANALOGUES &lt;130&gt; IPA120063 &lt;150&gt; EP 09251780.4 &lt;151&gt; 2009-07-13 &lt;150&gt; US 61/225,080 &lt;151&gt; 2009-07-13 &lt;150&gt; EP 10157240.2 &lt;151&gt; 2010-03-22 &lt;150&gt; DK PA 2010 00412 &lt;151&gt; 2010-05-10 &lt;160&gt; 123 &lt;170&gt; KopatentIn 2.0 &lt;210&gt; 1 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 1 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr 20 25 &lt;210&gt; 2 &lt;211&gt; 8 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 2 Lys Arg Asn Arg Asn Asn Ile Ala 1 5 &lt;210&gt; 3 &lt;211&gt; 37 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 3 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Lys Arg Asn 20 25 30 Arg Asn Asn Ile Ala 35 &lt;210&gt; 4 &lt;211&gt; 30 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 4 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Xaa Tyr Leu Asp Xaa 1 5 10 15 Xaa Ala Ala Xaa Xaa Phe Val Xaa Trp Leu Xaa Xaa Ala Xaa 20 25 30 &lt;210&gt; 5 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 5 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 6 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 6 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Xaa Tyr Leu Asp Xaa 1 5 10 15 Xaa Ala Ala Xaa Xaa Phe Val Xaa Trp Leu Leu Xaa Ala 20 25 &lt;210&gt; 7 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 7 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Xaa Tyr Leu Asp Xaa 1 5 10 15 Xaa Ala Ala Xaa Xaa Phe Val Xaa Trp Leu Leu Xaa Ala 20 25 &lt;210&gt; 8 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 8 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Xaa Tyr Leu Asp Ser 1 5 10 15 Xaa Ala Ala Xaa Xaa Phe Val Xaa Trp Leu Leu Xaa Ala 20 25 &lt;210&gt; 9 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 9 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Xaa Tyr Leu Asp Ser 1 5 10 15 Xaa Ala Ala Xaa Xaa Phe Val Xaa Trp Leu Leu Xaa Ala 20 25 &lt;210&gt; 10 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 10 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Xaa Tyr Leu Asp Ser 1 5 10 15 Xaa Ala Ala His Xaa Phe Val Xaa Trp Leu Leu Xaa Ala 20 25 &lt;210&gt; 11 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 11 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Xaa Tyr Leu Asp Ser 1 5 10 15 Xaa Ala Ala His Xaa Phe Val Xaa Trp Leu Leu Xaa Ala 20 25 &lt;210&gt; 12 &lt;211&gt; 28 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 12 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Xaa 20 25 &lt;210&gt; 13 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 13 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 14 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 14 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Lys 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 15 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 15 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Lys Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 16 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 16 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Lys Arg Ala 20 25 &lt;210&gt; 17 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 17 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Lys Ala 20 25 &lt;210&gt; 18 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 18 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Arg Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 19 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 19 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Leu Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 20 &lt;211&gt; 30 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 20 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala Lys 20 25 30 &lt;210&gt; 21 &lt;211&gt; 30 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 21 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala Lys 20 25 30 &lt;210&gt; 22 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 22 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Lys Ser Ala 20 25 &lt;210&gt; 23 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 23 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Lys Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 24 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 24 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Cys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 25 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 25 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Cys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 26 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 26 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Cys Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 27 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 27 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Lys 1 5 10 15 Ser Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 28 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 28 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 29 &lt;211&gt; 30 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 29 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala Lys 20 25 30 &lt;210&gt; 30 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 30 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Arg Asp Phe Val Ala Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 31 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 31 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Lys Asp Phe Val Ala Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 32 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 32 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 33 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 33 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Lys Ala 20 25 &lt;210&gt; 34 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 34 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Lys Asp Phe Val Ala Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 35 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 35 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Ala Trp Leu Leu Lys Ala 20 25 &lt;210&gt; 36 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 36 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Lys 1 5 10 15 Lys Ala Ala His Asp Phe Val Ala Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 37 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 37 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Arg Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 38 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 38 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Lys Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 39 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 39 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Leu Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 40 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 40 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Cys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 41 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 41 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Cys Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 42 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 42 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Lys 1 5 10 15 Lys Ala Ala Glu Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 43 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 43 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Lys Ala 20 25 &lt;210&gt; 44 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 44 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Lys Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 45 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 45 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Glu Phe Val Glu Trp Leu Leu Lys Ala 20 25 &lt;210&gt; 46 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 46 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Lys Glu Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 47 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 47 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 48 &lt;211&gt; 28 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 48 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Lys 1 5 10 15 Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 49 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 49 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Lys Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 50 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 50 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Lys Arg Ala 20 25 &lt;210&gt; 51 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 51 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Lys Ala 20 25 &lt;210&gt; 52 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 52 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Arg Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 53 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 53 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Leu Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 54 &lt;211&gt; 30 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 54 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala Lys 20 25 30 &lt;210&gt; 55 &lt;211&gt; 30 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 55 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala Lys 20 25 30 &lt;210&gt; 56 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 56 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Lys Ser Ala 20 25 &lt;210&gt; 57 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 57 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Lys Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 58 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 58 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Cys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 59 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 59 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Cys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 60 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 60 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Cys Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 61 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 61 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Lys 1 5 10 15 Ser Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 62 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 62 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 63 &lt;211&gt; 30 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 63 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala Lys 20 25 30 &lt;210&gt; 64 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 64 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Arg Asp Phe Val Ala Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 65 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 65 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Lys Asp Phe Val Ala Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 66 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 66 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Lys Ala 20 25 &lt;210&gt; 67 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 67 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Lys Ala 20 25 &lt;210&gt; 68 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 68 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 69 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 69 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Lys Asp Phe Val Ala Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 70 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 70 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Ala Trp Leu Leu Lys Ala 20 25 &lt;210&gt; 71 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 71 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Lys 1 5 10 15 Lys Ala Ala His Asp Phe Val Ala Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 72 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 72 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Arg Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 73 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 73 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Lys Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 74 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 74 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Leu Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 75 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 75 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Cys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 76 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 76 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Cys Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 77 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 77 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 78 &lt;211&gt; 28 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 78 His Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Lys Lys 1 5 10 15 Ala Ala Glu Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 79 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 79 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Lys Ala 20 25 &lt;210&gt; 80 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 80 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Lys Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 81 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 81 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Glu Phe Val Glu Trp Leu Leu Lys Ala 20 25 &lt;210&gt; 82 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 82 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Lys Glu Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 83 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 83 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Lys 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 84 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 84 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Lys Arg Ala 20 25 &lt;210&gt; 85 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 85 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Lys Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 86 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 86 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Lys Ala 20 25 &lt;210&gt; 87 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 87 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 88 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 88 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Arg Asp Phe Val Ala Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 89 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 89 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 90 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 90 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Lys Ala 20 25 &lt;210&gt; 91 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 91 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Lys Ala 20 25 &lt;210&gt; 92 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 92 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Lys Ala 20 25 &lt;210&gt; 93 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 93 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 94 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 94 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Lys Asp Phe Val Ala Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 95 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 95 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 96 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 96 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 97 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 97 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 98 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 98 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 99 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 99 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 100 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 100 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 101 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 101 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 102 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 102 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 103 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 103 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 104 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 104 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 105 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 105 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 106 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 106 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 107 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 107 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 108 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 108 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 109 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 109 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 110 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 110 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 111 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 111 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 112 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 112 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Lys 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 113 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 113 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Lys Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 114 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 114 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Lys Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 115 &lt;211&gt; 28 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 115 His Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser Lys 1 5 10 15 Ala Ala Arg Asp Phe Val Ala Trp Leu Leu Arg Ala 20 25 &lt;210&gt; 116 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 116 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Lys Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 117 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 117 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Glu 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 118 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 118 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Glu Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 119 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 119 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala Glu Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25 &lt;210&gt; 120 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 120 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Glu Ala 20 25 &lt;210&gt; 121 &lt;211&gt; 28 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 121 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser 20 25 &lt;210&gt; 122 &lt;211&gt; 27 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 122 His Xaa Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu 20 25 &lt;210&gt; 123 &lt;211&gt; 29 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt; &lt;223&gt; Peptide &lt;400&gt; 123 His Xaa Glu Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Lys Ala Ala His Asp Phe Val Glu Trp Leu Leu Ser Ala 20 25