MXPA06008404A - Pituitary adenylate cyclase activating peptide (pacap) receptor (vpac2) agonists and their pharmacological methods of use - Google Patents

Abstract

This invention provides novel peptides that function in vivo as agonists of the VPAC2 receptor. These insulin secretagogue polypeptides are shown to lower blood glucose in vivo upon glucose challenge. The polypeptides of this invention are also stable in formulation and have long half-lives. The peptides of the present invention provide a therapy for patients with decreased endogenous insulin secretion, for example, type 2 diabetics. The invention is also directed to a method of treating a metabolic disease in a mammal comprising administering a therapeutically effective amount of the peptides to said mammal.

Description

C ?, CH, CN. CO, CR, CU, CZ, DE, DK, DM, DZ, EC, EE, - as to the applicant 's entitlement to claim. the priorily ofthe EG, ES, Fl, GB. GD, GE, GH, GM, HR, HU, ID, IL. N, IS, earlier application (Rule 4.17 (iii)) for all designations JP, KE, KG, KP, KR. KZ, LC, LK, LR, LS, LT, LU, LV, MA, - as to ihe applicant's entitlement to claim the priorily of the MD, MG, MK, MN, MW, MX. MZ NA, NI, NO, NZ, OM. earlier application (Rule 4? 7 (iii)) for all designations PG, PH, PL, PT, RO, RU, SC, SD, SE, SG, SK, SL, SM, SY, - of inventorship (Rule 4.17 (iv )) for US only TJ. TM, TN, TR, TT, TZ, UA, UG, UZ, VC, VN, YU. ZA, ZM, ZW, ARIPO patent (BW GH GM, KE, LS, MW, MZ, PublisÃedn: NA, SD, SL, SZ, TZ, UG, ZM, ZW), Eurasian patent (AM, - without intemational search report and to be republished AZ. BY, KG, KZ, MD, UK, TJ. TM), European patent (AT, upon receipt of that report BE, BG, CH, CY, CZ, DE, DK, EE, ES, Fl, FR, GB, GR, HU, IS, IT, LT, LU, MC, NL, PL, PT, RO, SE, YES, SK, For lwo-letler codes and other abbreviations, referred to the "Gtiid- TR) .OAPl patent (BF, BJ, CF, CG, Cl, CM, GA, GN, GQ, ance Notes on Codes andAbbreviations "appearing at ihe beginGW, ML, MR, NE, SN, TD, TG) no regular ofeach issue ofthe PCT Gazelle. _ AGRONISTS OF THE RECEPTOR (VPAC2) OF THE PEPTIDE ACTIVATOR OF THE HYPOPHYSAL CYLASAN ADENYLATE (PACAP) AND ITS METHODS OF PHARMACOLOGICAL USE The present application claims the benefit of Provisional Application No.

Series 60 / 539,550 from the USA UU., Filed on January 27, 2004 and Provisional Application Serial No. 60 / 566,499 of the US. UU., Filed on April 29, 2004, whose content is included in its entirety by reference.

SCOPE OF THE INVENTION This invention relates to newly identified polypeptides and to the use of said polypeptides for therapeutic purposes. More specifically, the polypeptides of the present invention are useful for stimulating the release of insulin from pancreatic β cells in a glucose-dependent manner, thus providing a treatment option for people who have metabolic disorders, such as diabetes or glucose intolerance, a prediethetic state.

BACKGROUND OF THE INVENTION Diabetes is characterized by alterations in the metabolism of glucose, manifested, among other things, by an elevated blood glucose level in the diabetic patient. The underlying defects lead to a classification of diabetes into two main groups: type 1 diabetes or insulin dependent diabetes mellitus (IDDM), which arises when patients lack insulin-producing β cells in their pancreatic islet of Langerhans; and type 2 diabetes or non-insulin dependent diabetes mellitus (NIDDM), which occurs in patients with impaired β-cell function and impaired insulin action.

Currently, patients with type 1 diabetes receive treatment with insulin, while the majority of patients with type 2 diabetes are treated with agents that stimulate the function of the cells ß or with agents that increase the sensitivity of the tissues of the patients. patients towards insulin. Over time, almost half of the subjects with type 2 diabetes lose their response to these agents and should begin to receive insulin therapy. Next, we describe the drugs that are currently used to treat type 2 diabetes.

Alpha glucosidase inhibitors (eg, Precose® (Bayer Pharmaceuticals Corporation), Voglibose ™ (Takeda Pharmaceuticals Company Limited) and Miglitol® (Bayer Pharmaceuticals Corporation)) reduce postprandial glucose fluctuations by slowing glucose uptake of the intestine. These drugs are safe and provide treatment for people with mild to moderate diabetes. However, gastrointestinal side effects have been reported in the literature. Insulin sensitizers are drugs that increase the body's response to insulin. Thiozolidinediones, such as Avandia® (GlaxoSmithKIine, rosiglitazone) and Actos ™ (Takeda Pharmaceuticals Company Limited, pioglitazone) activate the peroxisome proliferator-activated receptor (PPAR) receptor of the gamma subtype and modulate the activity of a set of genes that They have not been well described. Rezulin ™ (Warner-Lambert Company, troglitazone), the first drug of this class, was withdrawn from the market because it caused elevated levels of liver enzymes and pharmacological hepatotoxicity. These hepatic effects do not seem to be a significant problem in patients using Avandia® and Actos ™. Anyway, it is recommended to perform a liver enzyme analysis every 2 months during the first year of therapy and, from now on, periodically. Avandia® and Actos ™ appear to be associated with fluid retention and edema. Another possible side effect is weight gain.

Avandia® is not indicated for use with insulin because congestive heart failure can occur. Insulin secretagogues (eg, sulfonylureas (SFU) and other agents that act via the adenosine triphosphate-dependent K + channel (ATP)) are another type of drug currently used to treat type 2 diabetes. SFUs are the standard therapy for patients with type 2 diabetes who have mild to moderate fasting blood glucose. SFUs have limitations that include the possibility of causing hypoglycaemia, weight gain, and high rates of primary and secondary failure. Between 10 and 20% of patients who receive treatment initially do not show a significant treatment effect (primary failure). Secondary failure is demonstrated by an additional loss of treatment effect, between 20 and 30%, after six months of receiving treatment with an SFU. Insulin treatment is necessary in 50% of subjects who responded to SFU after 5 to 7 years of therapy (Scheen, et al., Diabetes Res. Clin. Pract. 6: 533-543, 1989). Glucophage® (Lipha Corporation, Metformin HCl) is a biguanide that lowers blood glucose levels by reducing hepatic glucose production and increasing peripheral glucose uptake and utilization. The drug is effective in lowering the blood glucose level in subjects of mild to moderately affected by the disease, and does not have the side effects of weight gain or the possibility of causing hypoglycemia. However, Glucophage® has a number of side effects that include gastrointestinal disorders and lactic acidosis. Glucophage® is contraindicated in diabetic subjects older than 70 years and in subjects with renal or hepatic dysfunction. Finally, the primary and secondary failure rates of Glucophage® are similar to those of SFUs. Insulin treatment is instituted once the blood glucose level has not been adequately controlled by diet, exercise and oral medications This treatment has as disadvantages that it is injectable, can cause hypoglycemia and causes weight gain. Due to the problems posed by current treatments, new therapies are needed to treat type 2 diabetes. In particular, new treatments are needed to maintain normal insulin (glucose dependent) secretion. These new drugs should have the following characteristics: dependence on glucose to promote insulin secretion (that is, produce insulin secretion only in the presence of an elevated blood glucose level); low rates of primary and secondary failure; and preservation of the function of islet cells. The strategy for developing the new therapy disclosed herein is based on the signaling mechanism of cyclic adenosine monophosphate (cAMP) and its effects on insulin secretion. Cyclic AMP is an important regulator of the insulin secretion process. The elevation of this signaling molecule promotes the closing of the K + channels after the activation of the protein kinase A pathway. The closure of the K + channels causes the depolarization of the cells and the subsequent opening of the Ca ++ channels, which in turn leads to the exocytosis of insulin granules. In the absence of low glucose concentrations, the effects that occur in the secretion of insulin are scarce or nonexistent (Weinhaus, et al., Diabetes 47: 1426-1435, 1998). Secretagogues, such as pituitary adenylate cyclase activating peptide (PACAP), vasoactive intestinal peptide (VIP), and glucagon-like peptide 1 (glucagon-like peptide 1, GLP-1) ) use the cAMP system to regulate insulin secretion in a glucose-dependent manner (Komatsu, et al., Diabetes 46: 1928-1938, 1997; Filipsson, et al., Diabetes 50: 1959-1969, 2001; Drucker, Endocrinology 142: 521-527, 2001). Insulin secretagogues that act through elevation of cAMP, such as GLP-1, VIP, and PACAP can also increase insulin synthesis, in addition to insulin release (Skoglund, et al., Diabetes 49: 1156-1164, 2000; Borboni, et al., Endocrinology 140: 5530-5537, 1999). GLP-1 is released from the L cells of the intestine after a meal and functions as an incretin hormone (that is, it potentiates the glucose-induced insulin release of the pancreatic β-cell). It is a 37 amino acid peptide that is differentially expressed by the glucagon gene, depending on the type of tissue. With GLP-1, clinical data have been compiled that support the beneficial effect of increasing cAMP levels in cells ß. Infusions of GLP-1 in subjects with poor control of type 2 diabetes normalized their fasting blood glucose levels (Gutniak, et al., New Eng. J. Med. 326: 1316-1322, 1992) and by administering more prolonged infusions, the function of cells ß is improved until reaching the levels of normal subjects (Rachman, et al., Diabetes 45: 1524-1530, 1996). A recent report has shown that GLP-1 improves the ability of ß cells to respond to glucose in subjects with glucose intolerance (Byrne, et al., Diabetes 47: 1259-1265, 1998). However, all these effects are short-lived due to the short half-life of the peptide. Amylin Pharmaceuticals has carried out clinical trials with Exendin-4 (AC2993), a 39 amino acid peptide originally identified in the Gila Monster. Amylin has reported that clinical studies demonstrated better glycemic control in patients with type 2 diabetes treated with Exendin-4. However, the incidence of nausea and vomiting was significant. PACAP is a potent stimulator of insulin-dependent glucose secretion from pancreatic β-cells. Three different types of PACAP receptors have been described (PAC1, VPAC1 and VPAC2) (Harmar, et al., Pharmacol., Reviews 50: 265-270, 1998; Vaudry, et al., Pharmacol., Reviews 52: 269-324, 2000). The PACAP does not present selectivity for the receptors, and the three receptors have comparable activities and powers. The PAC1 is located predominantly in the central nervous system, while the VPAC1 and the VPAC2 have a wider distribution. The VPAC1 is located in the system central nervous system, and in the liver, lungs and intestine. VPAC2 is located in the central nervous system, pancreas, skeletal muscle, heart, kidney, adipose tissue, testicle and stomach. Recent work states that VPAC2 is responsible for the secretion of insulin from ß cells (Inagaki, et al., Proc. Nati, Acad. Sci. USA 91: 2679-2683, 1994, Tsutsumi, et al., Diabetes 51 : 1453-1460, 2002). This insulinotropic action of PACAP is mediated by the guanosine triphosphate (G) guanosine triphosphate binding proteins. The accumulation of intracellular cAMP, in turn, activates the non-selective cation channels in the β-cells, increased [Ca ++] and promotes the exocytosis of secretory granules containing insulin.

PACAP is the newest member of the superfamily of metabolic peptide hormones, neuroendocrines and neurotransmitters that exert their action through the signal transduction pathway mediated by cAMP (Arimura, Regul Peptides 37: 287-303, 1992). The biologically active peptides are released from the biosynthetic precursor in two molecular forms: as a 38 amino acid peptide (PACAP-38) and / or as a 27 amino acid peptide (PACAP-27) with an amidated carboxyl terminal (Arimura, supra). The highest concentrations of the two forms of the peptide are found in the brain and testis (Arimura, supra). The shortest form of the peptide, PACAP-27, shows a structural homology of 68% with respect to the vasoactive intestinal polypeptide (vasoactive intestinal polypeptide, VIP). However, the distribution of PACAP and VIP in the central nervous system suggests that these structurally related peptides have different neurotransmitter functions (Koves, et al., Neuroendocrinology 54: 159-169, 1991). Recent studies have shown biological effects other than PACAP-38, ranging from a role in reproduction (McArdle, Endocrinology 135: 815-817, 1994) to the ability to stimulate insulin secretion (Yada, et al., J. Biol. Chem. 269: 1290-1293, 1994). In addition, PACAP appears to play a role in the hormonal regulation of lipid and carbohydrate metabolism (Gray, et al., Mol. Endrocrinol., 15: 1739-47, 2001).; in the function circadian (Harmar, et al., Cell 109: 497-508, 2002); in the function of the immune system, growth, energy homeostasis and male reproduction (Asnicar, et al., Endrocrinol, 143: 3994-4006, 2002); in the regulation of appetite (Tachibana, et al., Neurosci, Lett 339: 203-206, 2003); as well as in acute and chronic inflammatory diseases, septic shock and autoimmune diseases (eg, systemic lupus erythematosus) (Well, Trends Mol. Med. 9: 211-217, 2003). Vasoactive intestinal peptide (VIP) is a 28 amino acid peptide that was isolated for the first time from the upper portion of the pig's small intestine (Said and Mutt, Science 169: 1217-1218, 1970; US Patent No. 3,879,371). ). This peptide belongs to a family of structurally related small polypeptides that includes helodermin, secretin, somatostatins and glucagon. The biological effects of VIP are mediated by the activation of membrane-bound receptor proteins that are coupled to the signaling system of intracellular cAMP. These receptors were originally known as VIP-R1 and VIP-R2; however, it was later discovered that they were the same receptors as VPAC1 and VPAC2. The VIP shows activities and powers comparable to VPAC1 and VPAC2. To improve the stability of VIP in human lung fluid, Bolin, et al., (Biopolymers 37: 57-66, 1995) developed a series of VIP variants designed to increase the propensity for a helical structure of this peptide and reduce the Proteolytic degradation. The substitutions were concentrated at positions 8, 12, 17 and 25-28, on which it was inferred that they were not important for binding to receptors. In addition, the sequence "GGT" was stuck to the C-terminal of the VIP muteins, with the hope of adding the cap in the helix with greater effectiveness. Finally, to further stabilize the helix, various cyclic variants were synthesized (U.S. Patent No. 5,677,419). While these efforts were not directed toward receptor selectivity, they gave two analogs that have higher selectivity for VPAC2 (Gourlet, et al., Peptides 18: 403-408, 1997; Xia, et al., J. Pharmacol. Exp. Ther., 281: 629-633, 1997).

Better peptides having the insulin-dependent insulin secretagogue activity of PACAP, GLP-1 or Exendin-4 are needed, but with fewer side effects and, preferably, they are stable in the formulation and have long plasma half-lives in vivo. This better in vivo half-life is obtained from peptides with lower clearance and lower susceptibility to proteolysis. Also, stricter control of plasma glucose levels can prevent long-term diabetic complications. In this way, new drugs for diabetes should provide a better quality of life to patients.

COMPENDIUM OF THE INVENTION This invention provides novel polypeptides that function in vivo as agonists of the VPAC2 receptor (hereinafter, VPAC2) and that are effective in the treatment of diseases and conditions that can be improved by agents that have agonist activity on it VPAC2. Preferably, the polypeptides of this invention are selective agonists of VPAC2 and have higher potency in VPAC2 than in VPAC1 and PAC1. For example, a mere enunciative tvtulo, these polypeptides stimulate the synthesis and release of insulin from the pancreatic β cells in a glucose-dependent manner and the subsequent reduction of plasma glucose. It has been shown that these polypeptide secretagogues decrease the blood glucose level in vivo to a greater extent than vehicle control by performing a glucose tolerance test. Also, the polypeptides of this invention are stable in the formulation and have long plasma half-lives and a long duration of action in vivo when they are derivatized. The polypeptides of the present invention have a better stability to proteolysis by dipeptidyl peptidase IV (DPP4) and in plasma, compared to PACAP or VIP. Although it has been reported that both VIP and PACAP27 are resistant to cleavage by DPP4 (Zhu, et al., J. Biol.

Chem 278: 22418-22423, 2003), Figure 2a demonstrates that these peptides are oliveated at longer points in time, while the peptides of the present invention are resistant to cleavage at the points in time evaluated. The derivatives of the present invention demonstrate an extended duration of action in vivo, which supports a dosage range of less than once per day, and once a week or more when they are derivatized. The polypeptides of the present invention provide a therapy for patients having, for example, metabolic disorders such as those that occur as a consequence of the reduction of endogenous insulin secretion, such as patients with type 2 diabetes, or for patients with intolerance. to glucose, a prediabetic state that is characterized by a mild alteration of insulin secretion. In addition, the polypeptides of the present invention may be useful for preventing and / or treating gestational diabetes, juvenile hereditary diabetes type 2 (maturity-onset diabetes of the young, MODY), latent autoimmune diabetes of the adult (latent autoimmune diabetes adult). , LADA) and the dyslipidemia associated with diabetes, and other diabetic complications, as well as hyperglycemia, hyperinsulinemia, glucose intolerance, impaired fasting glucose, dyslipidemia, hypertriglyceridemia, X syndrome and resistance. to insulin.

The polypeptides of the present invention can be used to prevent and / or treat obesity (eg, regulation of appetite and food intake), atherosclerotic disease, hyperlipidemia, hypercholesterolemia, low HDL levels, hypertension , cardiovascular disease (including atherosclerosis, coronary heart disease, coronary artery disease and hypertension), cerebrovascular disease and peripheral vessel disease; and to prevent and / or treat lupus, polycystic ovarian syndrome, carcinogenesis and hyperplasia, asthma, male reproductive problems, ulcers, sleep disorders, disorders of lipid and carbohydrate metabolism, dysfunction circadian, growth disorders, disorders of energy homeostasis, diseases immune, including autoimmune diseases (eg, systemic lupus erythematosus), as well as acute and chronic inflammatory diseases, septic shock, and other conditions identified herein; or they may perform other functions, which are described later in this. One aspect of the invention is a polypeptide selected from the group comprising the IDs. of SEQUENCE No. 1 to 148, and the fragments, derivatives and variants thereof that demonstrate at least one biological function that is substantially equal to that of the polypeptides of the SEQUENCE ID numbers mentioned (collectively, the "polypeptides of this invention "), including the functional equivalents thereof. Another application of this invention is a polypeptide selected from the group comprising SEQUENCE ID Nos. 1 to 37, and SEQUENCE ID Nos. 112 to 148, and fragments, derivatives and variants thereof that demonstrate at least one biological function that is substantially equal to that of the polypeptides of the mentioned SEQUENCE ID numbers. Another application of this invention is a polypeptide selected from the group comprising SEQUENCE ID Nos. 1 to 5, and SEQUENCE ID Nos. 112 to 115, and fragments, derivatives and variants thereof that demonstrate at least one biological function that is substantially equal to that of the polypeptides of the mentioned SEQUENCE numbers of ID. Another application of this invention is a polypeptide selected from the group comprising SEQUENCE IDs No. 1 and 112, and fragments, derivatives and variants thereof that demonstrate at least one biological function that is substantially equal to that of the polypeptides. of the mentioned SEQUENCE ID numbers. Antibodies and antibody fragments that selectively bind to the polypeptides of this invention are also provided. Such antibodies are useful for detecting the polypeptides of this invention, and can be identified and manufactured by methods well known in the art. Have been generated a polyclonal N-terminal IgG antibody and a C-terminal monoclonal Fab antibody recognized by the polypeptides of this invention.

The invention is also directed to a method for treating diabetes, disorders related to diabetes, and / or other diseases or conditions affected by the polypeptides of this invention, for example, affected by the function of the polypeptides of this invention as VPAC2 agonists, in a mammal, which involves administering to said mammal a therapeutically effective amount of any of the polypeptides of the present invention or any active polypeptide in VPAC2, such as those in SEQUENCE IDs No. 1 to 148. the manufacturing methods of the polypeptides of this invention are disclosed.

BRIEF DESCRIPTION OF THE ILLUSTRATION Figures 1 a-1d depict the amino acid sequences of the polypeptides of SEQUENCE ID Nos. 1 to 148. SEQUENCE ID Nos. 112-148 refer to peptides that are PEGylated on the C-terminal cysteine by a maleimide bond. The polyethylene glycol (PEG) can be of any length, for example, a 22 kD linear PEG or a 43 kD or larger branched PEG. Figure 1e refers to the standards of the related peptides.

Figures 2a and 2b depict the stability of VPAC2 analogs against proteolytic cleavage by DPP4.

Figure 3 represents the response to cAMP of the cells treated with the VPAC peptides. Figures 4a and 4b represent an assay with enzyme-linked immunosorbent substances (Enzyme Linked-lmmuno-Sorbent Assay, ELISA) for the detection of peptides. Figures 5a, 5b and 5c demonstrate the pharmacokinetic properties of the VPAC peptides.

Figures 6a, 6b, 6c and 6d illustrate efficacy in vivo.

DETAILED DESCRIPTION OF THE INVENTION This invention provides novel polypeptides, and fragments, derivatives and variants thereof that demonstrate at least one biological function that is substantially equal to that of the polypeptides of Figures 1a-1d (collectively, the polypeptides of this invention). nvention). The polypeptides of this invention function as VPAC2 agonists and can be used to prevent and / or treat diseases or conditions such as diabetes, including type 2 diabetes, gestational diabetes, juvenile hereditary diabetes type 2 (MODY) ( Herman, et al., Diabetes 43:40, 1994); latent autoimmune diabetes of the adult (LADA) (Zimmet, et al., Diabetes Med. 11: 299, 1994); and dyslipidemia associated with diabetes and other diabetic complications, as well as hyperglycemia, hyperinsulinemia, glucose intolerance, impaired fasting glucose, dyslipidemia, hypertriglyceridemia, syndrome X, and insulin resistance. In addition, the polypeptides of the present invention can also be used to prevent and / or treat obesity (eg, regulation of appetite and food intake), atherosclerotic disease, hyperlipidemia, hypercholesterolemia, low levels of HDL, hypertension, cardiovascular disease (including atherosclerosis, coronary heart disease, coronary artery disease and hypertension), cerebrovascular disease and peripheral vessel disease; and to prevent and / or treat lupus, polycystic ovary syndrome, carcinogenesis and hyperplasia, asthma, male reproductive problems, including motility of human sperm, ulcers, sleep disorders and other conditions identified in the I presented; or they may perform other functions, which are described later in this. Also, the polypeptides of this invention stimulate the release of insulin from pancreatic cells in a glucose-dependent manner. The polypeptides of this invention are also stable in aqueous and non-aqueous formulations, and exhibit a plasma half-life of more than one hour, for example, of more than 6 hours. The polypeptides of this invention are VPAC2 agonists. They are selective agonists of VPAC2 that have at least, for example, a selectivity for VPAC2 equal to 10 times what they have for VPAC1 and / or PAC1. The polypeptides of this invention stimulate the release of insulin in the plasma in a glucose-dependent manner without causing stasis or increased plasma glucose level which is counterproductive for the treatment, for example, of type 2 diabetes. polypeptides of this invention are selective agonists of the VPAC2 receptor; in this way, for example, they cause an increase in the release of insulin in the plasma, while being selective with respect to other receptors that are responsible for unpleasant or dangerous side effects, such as water retention in the gastrointestinal tract and / or unwanted cardiovascular effects, such as increased heart rate or blood pressure.

The polypeptides of this invention are also stable in aqueous and non-aqueous formulations. For example, the polypeptides of this invention exhibit a degradation of less than 10% at 37-40 ° C for a period of one week, when they are dissolved in water (at a pH between 7-8) or in a non-aqueous organic solvent . Also, the compositions and formulations of the present invention may comprise the polypeptides of the present invention, and one or more of pharmaceutically acceptable carriers, pharmaceutically acceptable diluents and pharmaceutically acceptable solvents. The polypeptides of this invention provide a therapy for patients with a reduction in endogenous insulin secretion or glucose intolerance, for example, type 2 diabetes. That is, the polypeptides of the present invention are long-acting VPAC2 agonists. which can be used to maintain, improve and restore insulin secretion stimulated by glucose. Also, a selective peptide agonist of VPAC2 receptor increases glucose-dependent insulin secretion in the pancreas, without causing side effects associated with non-selective activation of the other PACAP receptors. Certain terms that are used throughout this specification are defined below and others will be defined as they appear. Below is the abbreviation of a single letter for a specific amino acid, its corresponding amino acid and the abbreviation of three letters: A, alanine (wing); C, cysteine (cis); D, aspartic acid (asp); E, glutamic acid (glu); F, phenylalanine (phe); G, glycine (gly); H, histidine (his); I, isoleucine (ile); K, lysine (lys); L, leucine (leu); M, methionine (met); N, asparagine (asn); P, proline (pro); Q, glutamine (gln); R, arginine (arg); S, serine (being); T, threonine (thr); V, valine (val); W, tryptophan (trp); and Y, tyrosine (tyr). The terms "functional equivalent" and "substantially the same biological function or activity" mean the degree of biological activity that is comprised between approximately 30% and 100% or more of the biological activity demonstrated by the polypeptide with which it is being making the comparison, when the biological activity of each polypeptide is determined by the same procedure. For example, a polypeptide that is functionally equivalent to a polypeptide of Figure 1 is one that, when evaluated by the proximity scintillation assay for the cyclic AMP (cAMP) of Example 9, demonstrates the accumulation of cAMP in the cell line of Chinese hamster ovary (Chínese Hamster Ovary, CHO) expressing the human VPAC2 receptor. The terms "fragment", "derivative" and "variant", referred to the polypeptides of Figure 1, mean the fragments, derivatives and variants of the polypeptides that retain substantially the same function or biological activity as said polypeptides, as described more in detail below.

An analog includes a pro-polypeptide that includes within it the amino acid sequence of the polypeptide of this invention. The active polypeptide of this invention can be oligoed from the additional amino acids that complete the pro-polypeptide molecule by natural in vivo processes, or by procedures well known in the art, such as enzymatic or chemical cleavage. For example, the native 28 amino acid VIP peptide is naturally expressed as a much larger polypeptide which is then processed in vivo to release the active mature peptide of 28 amino acids. A fragment is a portion of the polypeptide that retains substantially similar functional activity, as described in the in vivo models disclosed herein. A derivative includes all modifications to the polypeptide that substantially preserve the functions disclosed herein and include an additional related structure and function (e.g., PEGylated or acetylated polypeptides having a longer half-life), fusion polypeptides that confer a longer half-life, targeted to specificity or additional activity, such as a reduction in toxicity to a proposed purpose, as described in more detail below . The fragment, derivative or variant of the polypeptides of the present invention can be (i) that in which one or more of the amino acid residues are replaced by a conserved or non-conserved amino acid residue (preferably a conserved amino acid residue) and said substituted amino acid residue may or may not be encoded by the genetic code; (ii) that in which one or more of the amino acid residues includes a substituent group; (iii) that in which the N-terminal acetyl group is replaced by another substituent in one or more of the first three amino acids or that in which one or more of the first three amino acids is replaced by an amino acid residue conserved or not conserved (preferably a conserved amino acid residue) and said substituted amino acid residue may or may not be encoded by the genetic code, in order to confer resistance to proteolysis; (iv) that in which the mature polypeptide is fused with another compound, such as a compound to increase the half-life of the polypeptide (e.g., polyethylene glycol or fatty acid); (v) that in which the additional amino acids are fused with the mature polypeptide, as a leader or secretory sequence, or a sequence that is used for the purification of the mature polypeptide or a propolypeptide sequence, or (vi) that in which the The polypeptide sequence is fused with a larger polypeptide (eg, a human albumin, an antibody or Fc to increase the duration of the effect). Said fragments, derivatives, and variants and the like are considered to be included within the scope of those skilled in the art in accordance with what is described herein. The derivatives of the present invention may contain conservative amino acid substitutions (defined in more detail below) which are made in one or more predicted, preferably nonessential, amino acid residues. A "non-essential" amino acid residue is a residue that can be altered from the natural sequence of a protein without altering biological activity, while an "essential" amino acid residue is necessary for biological activity. A "substitution of a conservative amino acid" is one in which the amino acid residue is replaced by an amino acid residue having a similar side chain. Families of amino acid residues that have similar side chains have been defined in the field. These families include amino acids with basic side chains (eg, lysine, arginine, histidine), acid side chains (eg, aspartic acid, glutamic acid), polar uncharged side chains (eg, glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), non-polar side chains (eg, alanine, valine, leuoin, isoleucine, proline, phenylalanine, methionine, tryptophan), branched beta side chains (eg, threonine, valine, isoleucine) and aromatic side chains (eg, tyrosine, phenylalanine, tryptophan, histidine). No non-conservative substitutions would be made for conserved amino acid residues or for amino acid residues residing within the domain of a conserved protein, such as residues 19 and 27 that are essential for the activity of the protein, such as VPAC2 activity and / or the selectivity for VPAC2. Fragments or biologically active portions include fragments of polypeptides suitable for use as a medicament, for generating antibodies, as a reagent for research, and for similar uses. Fragments include peptides that comprise amino acid sequences sufficiently similar to the amino acid sequences of a polypeptide of this invention or derived therefrom, and which exhibit at least one activity of that polypeptide, but which include fewer amino acids than the full length polypeptides disclosed at the moment. Generally, biologically active portions comprise a domain or motif having at least one activity of the polypeptide. A biologically active portion of a polypeptide can, for example, be a peptide having a length of five or more amino acids.

- Said biologically active portions can be prepared synthetically or by recombinant techniques, and can be evaluated in relation to one or more of the functional activities of a polypeptide of this invention by means disclosed herein and / or well known in the art. Variants of the polypeptides of this invention include polypeptides having an amino acid sequence sufficiently similar to the amino acid sequence of the SEQUENCE ID numbers of Figure 1 (a-d) or a domain thereof. The term "sufficiently similar" means a first amino acid sequence that contains a sufficient or minimum amount of amino acid residues identical or equivalent to those of a second amino acid sequence, such that the first and second amino acid sequence have a structural domain common and / or a common functional activity. For exampleD. , amino acid sequences that contain a common structural domain that is at least about 45% identical, about 75% to 98% identical, or that are identical are defined herein as sufficiently similar. For example, the variants may be sufficiently similar to the amino acid sequence of the polypeptides of this invention. Variants include polypeptides that differ in amino acid sequence due to mutagenesis. Variants that function as VPAC2 agonists can be identified by screening libraries combinatorias of. mutants, for example, mutants by truncation, of the polypeptides of this invention in relation to the agonist activity on the VPAC2. In addition, derivatives of the present invention include mature polypeptides that have been fused to another compound, for example a compound to increase the half-life of the polypeptide and / or reduce the possible immunogenicity of the polypeptide (e.g., polyethylene glycol ["PEG" ] or fatty acid). In the case of PEGylation, the fusion of the polypeptide with the PEG can be achieved by any means known to a person skilled in the art. For example, PEGylation can be achieved by first introducing a mutation in the cysteine in the polypeptide to provide a bond to which PEG can bind, followed by site-specific derivatization with PEG-maleimide. As an example, cysteine can be added to the C-terminus of the peptides (see, eg, Tsutsumi, et al., Proc. Nati, Acad. Sci. USA 97 (15): 8548-53, 2000; Veronese , Biomaterials 22: 405-417, 2001; Goodsoon and Katre, Bio / Technology 8: 343-346, 1990). In addition to the maleimide, there are numerous reactive Cys groups that are known to those skilled in the art of protein crosslinking, such as the use of alkyl halides and vinyl sulfones (see, eg, TE Creighton, Proteins, 2 a Ed., 1993). In addition, PEG can be introduced by direct binding to the C-terminal carboxylate group, to the side chain of a C-terminal, to an internal amino acid, such as Cys, Lys, Asp or Glu, or to non-natural amino acids containing side chain fractions. similar reactive The link between the PEG and the peptide crosslinking group may vary. For example, the commercially available 40 kDa reagent for Cys (mPEG2-MAL; Nektar, San Carlos, CA) employs a maleimide group for conjugation with Cys, and the maleimide group binds to PEG via a linkage containing a Lys. As a second example, the 43 kDa reagent PEG for Cys (GL2-400MA; NOF, Tokyo, Japan) commercially available employs a maleimide group for conjugation with the Cys, and the maleimide group binds to the PEG via a disubstituted alkane linkage.

The present invention exemplifies, to a mere enunciative title, the use of Cys as a crosslinking site. It is well known that other fractions present in amino acids such as the N-terminal amino group, the C-terminal carboxylate and the side chains of amino acids such as Lys, Arg, Asp or Glu, provide reactive groups with fractions suitable for covalent modification and binding to PEG. There are numerous examples of suitable crosslinking agents that are known to those skilled in the art (see, eg, T. E. Creighton, Proteins, 2 a Ed., 1993). Such crosslinking agents can be linked to PEG as illustrated, by way of example, by commercially available PEG derivatives containing commercially available amines, aldehydes, acetals, maleimide, succinimides and thiols, for example, Nektar and NOF (e.g. ., Harris, et al., Clin. Pharmokinet., 40, 539-551, 2001). The invention also provides chimeric or fusion polypeptides. The polypeptides of this invention may be composed of amino acids linked together by peptide linkages or modified peptide linkages (ie, peptide isosteres) and may contain amino acids other than the 20 amino acids encoded by the genes. The polypeptides can be modified by natural processes, such as post-translational processing, or by chemical modification techniques that are well known in the art. These modifications are well described in basic texts and in more detailed monographs, as well as in the research literature. Modifications can occur in any part of a polypeptide, including the peptide backbone, the amino acid side chains and the amino or carboxyl terminus. It will be appreciated that the same type of modification may be present in equal or different degrees at various sites of a given polypeptide. In addition, a given polypeptide can contain many types of modifications. The polypeptides may be branched, for example, as a result of ubiquitination, and may be cyclic, with or without branching. Cyclic polypeptides, branched polypeptides and branched cyclic polypeptides can be the result of natural post-translational processes or be manufactured by synthetic methods. Modifications include acetylation, acylation, ADP-ribosylation, amidation, covalent binding of flavin, covalent attachment of a heme moiety, covalent attachment of a nucleotide or nucleotide derivative, covalent attachment of a lipid or lipid derivative, covalent attachment of phosphatidylinositol, crosslinking, cyclization, disulfide bond formation, demethylation, covalent crosslink formation, cysteine formation, pyroglutamate formation, formulation, gamma carboxylation, glycosylation, glycosyl phosphatidylinositol anchoring (Glycosyl Phosphatidylinositol, GPl), hydroxylation, iodination, methylation , myristoylation, oxidation, pegylation, proteolytic processing, phosphorylation, reward, racemization, selenoylation, sulfation, addition of amino acids to proteins mediated by transfer RNA, such as arginylation and ubiquitination (see, eg, Proteins, Structure and Molecular Properties. 2nd ed., T. E. Creighton, W. H. Freeman and Company, New York (1993); Posttranslational Covalent Modification of Proteins. B. C. Johnson, ed., Academic Press, New York, pgs. 1-12 (1983); Seifter, et al., Meth. Enzymol 182: 626-646, 1990; Rattan, et al., Ann. N.Y. Acad. Sci. 663: 48-62, 1992). The polypeptides of the present invention include the polypeptides of Figure 1a-d (SEQUENCE ID No. 1 to 148), as well as the sequences that have insubstantial variations with respect to these. An "unsubstantial variation" would include any addition, substitution or deletion of the sequence that substantially maintains, at least, a biological function of the polypeptides of this invention, for example, the agonist activity on VPAC2, the selective agonist activity on VPAC2 or the insulin secretion activity demonstrated in the present. These functional equivalents may include polypeptides having at least about 90% identity, the 95% or 97% with the polypeptides of Figure 1a-d, and also include portions of said polypeptides having substantially the same biological activity. However, any polypeptide that has an insubstantial variation in the amino acid sequence with respect to the polypeptides of Figure 1a-d and demonstrating a functional equivalence as described in more detail herein is included in the description of the present invention. As is known in the art, the "similarity" between two polypeptides can be determined by comparing the amino acid sequence and its conserved amino acid substitutes of a polypeptide with the sequence of a second polypeptide. Said conservative substitutions include those described above, and those described by Dayhoff (The Atlas of Protein Sequence and Structure 5, 1978) and by Argos (EMBO J. 8: 779-785, 1989). For example, the amino acids belonging to one of the following groups represent conservative changes: - ala, pro, gly, gln, asn, ser, thr; - cys, ser, tyr, thr; - val, ile, leu, met, ala, phe; - lys, arg, his; - phe, tyr, trp, his; and - asp, glu. The polypeptides of this invention may be a product of synthetic chemical processes and, as such, a polypeptide of this invention isolated or purified, or a biologically active part thereof, may be substantially free of chemical precursors or other chemical substances when chemically synthesized. . For example, a polypeptide of this invention isolated may have less than about 30% (by dry weight) of non-polypeptide or contaminant material. When a peptide of this invention is produced by chemical synthesis, the preparations may contain less than about 30%, by dry weight, of chemical precursors or chemicals not included in the invention.

The polypeptides of this invention can conveniently be isolated as described in the specific examples included below. A purified polypeptide preparation, for example, has a minimum purity of about 70%; or a purity of about 85% to 99%. The purity of the preparations can be evaluated by any means known in the art, such as polyacrylamide gel electrophoresis with sodium dodecyl sulfate (SDS) and mass spectroscopy / liquid chromatography. Peptides and related compounds (e.g., small molecule) that are understood by those skilled in the art, such as mimetic chemicals, mimetic organic molecules or mimetic peptides, are also provided. As used herein, the terms "mimetic," "mimetic peptide," "mimetic organic molecule," and "mimetic chemical" are intended to include derivatives of peptides, peptide analogs, and chemical compounds "that they have an arrangement of the atoms in a three-dimensional orientation that is equivalent to that of a peptide of the present invention. The phrase "equivalent to" is understood, as used herein, to comprise the peptides or compounds having one or more substitutions of certain atoms or chemical fractions in said peptide, which have binding lengths, binding angles and dispositions in the mimetic compound that produce an arrangement or orientation of said atoms and fractions equal or sufficiently similar to have the biological function of the Peptides of the Invention In the mimetic peptides of the invention, the three-dimensional arrangement of the chemical constituents is structurally and / or functionally equivalent to the three-dimensional arrangement of the peptide backbone and the side chains of amino acid components of the peptide; as a result, said mimetic peptides, organic mimetic molecules and mimetic chemicals of the peptides of the invention have substantial biological activity. These terms are used according to what is known in the art, as illustrated, for example, Fauchere, (Adv. Drug Res. 15:29, 1986); Veber and Freidinger, (TINS p.392, 1985); and Evans, et al., (J. Med. Chem. 30: 1229, 1987), which is included herein by reference. It is understood that there is a pharmacophore for the biological activity of each peptide of the invention. In this field it is understood that a pharmacophore comprises an idealized three-dimensional definition of the structural requirements for biological activity. The mimetic peptides, organic mimetic molecules and mimetic chemicals can be designed to fit each pharmacophore with the current computer modeling software (computer-aided drug design). Said mimetics can be produced by a structure-function analysis, based on the positional information of the substituent atoms in the peptides of the invention. The peptides as provided by the invention can be advantageously synthesized by any of the chemical synthesis techniques known in the art, particularly the solid phase synthesis techniques, for example, using commercially available automatic peptide synthesizers. The mimetics of the present invention can be synthesized by the solid phase or solution phase methods conventionally used for the synthesis of peptides (see, eg, Merrifield, J. Amer. Chem. Soc. 85: 2149- 54, 1963, Carpino, Acc. Chem. Res. 6: 191-98, 1973, Birr, Aspects of the Merrifield Peptide Synthesis, Springer-Verlag: Heidelberg, 1978, The Peptides: Analysis, Synthesis, Biology, Vol. 2, 3 and 5, (Gross and Meinhofer, eds.), Academic Press: New York, 1979, Stewart, et al., Solid Phase Peptide Synthesis, 2nd ed., Pierce Chem. Co .: Rockford, lll. , 1984, Kent, Ann., Rev. Biochem., 57: 957-89, 1988, and Gregg, et al., Int. J. Peptide Protein Res. 55: 161-214, 1990, which are included in full in the present by reference). For example, the solid phase methodology can be used. Briefly, an N-protected C-terminal amino acid residue is bound to an insoluble support, such as polystyrene crosslinked with divinylbenzene, polyacrylamide resin, Kieselguhr / polyamide (Pepsyn K), controlled pore glass, cellulose, membranes of polypropylene, polyethylene rods coated with acrylic acid or other similar. The deprotection, neutralization and coupling cycles of successive protected amino acid derivatives are used to bind the C-terminal amino acids, according to the amino acid sequence. For some synthetic peptides, an FMOC strategy can be used, using an acid sensitive resin. In this regard, the solid supports are polystyrene resins crosslinked with divinylbenzene, which are commercially available in various functionalized forms, which include the chloromethyl resin, the hydroxymethyl resin, the paraacetamidomethyl resin, the benzydrylamine resin (BHA), the 4-methylbenzydrylamine resin (MBHA), oxime resins, 4-alkoxybenzyl alcohol resin (Wang resin), 4- (2 ', 4, -dimethoxyphenyl aminomethyl) -phenoxymethyl resin, 2,4-alkynyl resin -dimethoxybenzydrylamine and the resin of 4- (2, 4'-dimethoxyphenyl-FMOC-amino-methyl) -phenoxyacetamidonorleucyl-MBHA (resin Rink amide MBHA). In addition, acid-sensitive resins also provide C-terminal acids, if desired. A protective group of alpha amino acids is 9-fluorenylmethoxycarbonyl (FMOC) labile bases. Suitable protecting groups of amino acid side chain functionalities chemically compatible with the BOC (t-butyloxycarbonyl) and FMOC groups are well known in the art. When using FMOC chemistry, the following protected amino acid derivatives are preferred: FMOC-Cys (Trit), FMOC-Ser (But), FMOC-Asn (Trit), FMOC-Leu, FMOC-Thr (Trit), FMOC- Val, FMOC-Gly, FMOC-Lys (Boc), FMOC-Gln (Trit), FMOC-Glu (OBut), FMOC-His (Trit), FMOC-Tyr (But), FMOC-Arg (PMC (2.2 , 5,7,8-pentamethylchroman-6-sulfonyl)), FMOC-Arg (BOC) 2, FMOC-Pro and FMOC-Trp (BOC). The amino acid residues can be coupled using various coupling agents and chemistries known in the field, as direct coupling with DIC (diisopropyl-carbodiimide), DCC (dicyclohexylcarbodiimide), BOP (benzotriazolyl-N-oxitrisdimethylaminophosphonium hexafluorophosphate), PyBOP (benzotriazole hexafluorophosphate) -1-yl-oxy-tris-pyrrolidinophosphonium), PyBrOP (bromo-tris-pyrrolidinophosphonium hexafluorophosphate); by preformed symmetrical anhydrides; through active esters, such as pentafluorophenyl esters; or by preformed active HOBt (1-hydroxybenzotriazole) esters or by using fluoride and amino acid chlorides with FMOC, or by using N-carboxy amino acid anhydrides with FMOC. Activation with HBTU (2- (1H-benzotriazol-1-yl), 1, 1,3,3-tetramethyluronium hexafluorophosphate) or HATU (2- (1H-7- aza-benzotriazole-1 hexafluorophosphate) is preferred. -yl), 1, 1, 3,3-tetramethyluronium) in the presence of HOBt or HOAt (7- azahydroxybenzotriazoi). The solid phase method can be carried out manually, although automatic synthesis can be used by a commercially available peptide synthesizer (eg, Applied Biosystems 431A or similar, Applied Biosystems, Foster City, CA). In a typical synthesis, the first amino acid (C-terminal) is loaded into the chlorotryril resin. Successive deprotection (with 20% piperidine / NMP (N-methylpyrrolidone)) and coupling cycles according to the ABI FastMoc protocols (Applied Biosystems) can be used to generate the peptide sequence. Double and triple coupling can also be used, with the addition of a cap with acetic anhydride. The synthetic mimetic peptide can be oiled from the resin and deprotected by treatment with trifluoroacetic acid (trifluoroacetic acid, TFA) containing the appropriate antioxidants. Many of these cleavage reagents can also be used, such as reagent K (0.75 g of crystalline phenol, 0.25 mL of ethanedithiol, 0.5 mL of thioanisole, 0.5 mL of deionized water and 10 mL of TFA). and other reagents. The peptide can be separated from the resin by filtration and isolated by precipitation with ether. Further purification can be achieved by conventional methods, such as gel filtration and high performance liquid chromatography (HPLC) in inverted phase. The synthetic mimetic peptides according to the present invention can take the form of pharmaceutically acceptable salts, especially salts with the addition of a base, including the salts of organic bases and inorganic bases. Salts with base addition of the acidic amino acid residues can be prepared by treating the peptide with the base or base suitable inorganic, according to procedures well known to those skilled in the art, or also, the desired salt can be obtained directly by lyophilization of the appropriate base. In general, those skilled in the art will recognize that peptides such as those described herein can be modified by various chemical techniques, in order to produce peptides that have essentially the same activity as the unmodified peptide, and that optionally have other desirable properties . For example, the carboxylic acid groups of the peptide may be provided in the form of a salt of a pharmaceutically acceptable cation. The amino groups of the peptide may be in the form of a salt with the addition of a pharmaceutically acceptable acid, such as HCl, HBr, and salts of acetic, benzoic, toluene sulfonic, maleic, tartaric, and other organic salts, or may be converted into a amide. Thiols can be protected by any of several well-recognized protecting groups, such as acetamide groups. Those skilled in the art will also recognize methods for introducing cyclic structures in the peptides of this invention, in order to come closer to the native binding configuration. For example, a carboxy terminal or amino terminal cysteine residue can be added to the peptide, so that upon being oxidized the peptide contains a disulfide bond, thereby generating a cyclic peptide. Other methods of peptide cyclization include the formation of thioethers and amides and carboxyl and amino terminal esters. Specifically, various techniques are available to construct peptide derivatives and analogs with a desirable biological activity equal or similar to that of the corresponding peptide, but with a more favorable activity than the peptide with respect to solubility, stability and susceptibility to hydrolysis and proteolysis. Said derivatives and analogs include peptides modified at the N-terminal amino group, the C-terminal carboxyl group and / or the change of one or more of the amido bonds in the peptide by a non-amido linkage. It is understood that two or more of said modifications can be coupled in the structure of a peptide mimetic (eg, modification in the C-terminal carboxyl group and inclusion of a carbamate-CH2- linkage between two amino acids of the peptide). Modifications at the amino termini include alkylation, acetylation, the addition of a carbobenzoyl group and the formation of a succinimide group. Specifically, the N-terminal amino group can be reacted to form an amide group of the formula RC (O) NH ~, where R is an alkyl, for example, lower alkyl, and is added by the reaction with a halide of acid, RC (O) CI or acid anhydride. In general, the reaction can be carried out by contacting approximately equimolar or excess amounts (eg, about 5 equivalents) of an acid halide with the peptide in an inert diluent (eg, dichloromethane) , which preferably contains an excess (eg, about 10 equivalents) of a tertiary amine, such as diisopropylethylamine, to collect the acid generated during the reaction. The other conditions in which the reaction occurs are conventional (eg, room temperature for 30 minutes). Alkylation of the amino terminus to provide an N-substitution of a lower alkyl followed by a reaction with an acid halide, as described above, will provide an N-alkyl amide group of the formula RC (O) NR- Alternatively , the amino terminal can be covalently linked to the succinimide group by the reaction with succinic anhydride. An approximately equimolar amount or an excess of succinic anhydride (e.g., about 5 equivalents) is used and the terminal amino group is converted to succinimide by methods well known in the art, including the use of an excess (e.g. eg, 10 equivalents) of a tertiary amine, such as diisopropylethylamine, in an appropriate inert solvent (eg, dichloromethane), as described in Wolienberg, et al., (U.S. Patent No. 4,612,132) , which is incorporated herein in its entirety by reference. It is also understood that the succinic group may be substituted, for example, by a C2- to Ce-alkyl or -SR substituents, which are prepared in a conventional manner to provide the substituted N-terminal succinimide of the peptide. Sayings alkyl substituents can be prepared by the reaction of a lower olefin (C2- to C6-alkyl) with maleic anhydride in the manner described by Wollenberg, et al., supra., and substituents -SR can be prepared by the reaction of RSH with anhydride maleic, where R is what was defined above. In another advantageous application, the amino terminus can be derivatized to form a benzyloxycarbonyl-NH- or benzyloxycarbonyl-NH-substituted group. This derivative can be produced by reaction with an approximately equivalent amount or an excess of benzyloxycarbonyl chloride (CBZ-Cl) or a substituted CBZ-CI in a suitable inert diluent (eg, dichloromethane) containing, for example, a tertiary amine to collect the acid generated during the reaction. In yet another derivative, the N-terminus comprises a sulfonamide group by reaction with an equivalent amount or an excess (eg, 5 equivalents) of RS (O) 2CI in an appropriate inert solvent (dichloromethane) to convert the amine terminal in a sulfonamide, wherein R is an alkyl and preferably, a lower alkyl. For example, the inert diluent may contain an excess of a tertiary amine (eg, 10 equivalents), such as diisopropylethylamine, to collect the acid generated during the reaction. The other conditions in which the reaction occurs are conventional (eg, room temperature for 30 minutes). Carbamate groups can be produced at the amino terminus by reaction with an equivalent amount or an excess (eg, 5 equivalents) of R-OC (O) CI or R-OC (O) OC6H4-p-? 2 in a suitable inert diluent (e.g., dichloromethane) to convert the terminal amine to carbamate, where R is an alkyl and preferably a lower alkyl. For example, the inert diluent may contain an excess (eg, about 10 equivalents) of a tertiary amine, such as diisopropylethylamine, to collect any acid generated during the reaction. The other conditions in which the reaction occurs are conventional (eg, room temperature for 30 minutes). Urea groups can be formed at the amino terminus by reaction with an equivalent amount or an excess (eg, 5 equivalents) of RN = C = O in an appropriate inert diluent (e.g., dichloromethane) to convert the amine terminal in a urea group (ie,.

RNHC (O) NH-), where R is what was previously defined. For example, the inert diluent may contain an excess (e.g., about 10 equivalents) of a tertiary amine, such as diisopropylethylamine. The other conditions under which the reaction occurs are conventional (eg, room temperature for about 30 minutes). In preparing mimetic peptides where the C-terminal carboxyl group can be replaced by an ester (eg, -C (O) OR, where R is an alkyl and preferably a lower alkyl), the resins used to prepare peptide acids, and the peptide protected by the side chain can be olive-based with a suitable base and alcohol (eg, methanol). The protecting groups of the side chain can be removed in the usual way by treatment with hydrogen fluoride, in order to obtain the desired ester. By preparing mimetic peptides where the C-terminal carboxyl group is replaced by the amide -C (O) NR3R. a benzydrylamine resin is used as a solid support for peptide synthesis. Upon completion of the synthesis, treatment with hydrogen fluoride to release the peptide from the support directly results in the free peptide amide (ie, the C-terminus is -C (O) NH2). Alternatively, the use of the chloromethylated resin during peptide synthesis together with the reaction with ammonia to clivate the peptide protected by the side chain of the support gives the free peptide amide, and the reaction with an alkylamine or dialkylamine gives an alkylamide or dialkylamide protected by the side chain (ie, the C-terminal is -C (O) NRR ?, where R and Ri are alkyls and preferably, lower alkyls). Then the protection of the side chain is removed in the usual way, by treatment with hydrogen fluoride, in order to obtain free amides, alkylamides or dialkylamides. In another alternative application, the cyclization of the C-terminal carboxyl group or of a C-terminal ester can be induced by displacement of the -OH or the ester (-OR) of the carboxyl group or ester, respectively, with the amino group N-terminal to form a cyclic peptide. For example, after synthesis and cleavage to give the peptide acid, the free acid becomes a solution in a activated ester by an appropriate activating carboxyl group, such as dicyclohexylcarbodiimide (DCC), for example, in methylene chloride (CH2Cl2), dimethyl formamide (DMF) or mixtures thereof. The cyclic peptide is then formed by displacement of the activated ester with the N-terminal amine. Cyclization, rather than polymerization, can be improved by the use of highly diluted solutions according to methods well known in the art.

The mimetic peptides, as understood in the art and as provided by the invention, are structurally similar to the peptide of the invention, but have one or more peptide bonds that can be replaced by a bond selected from the group consisting of: -CH2NH- , -CH2S-, - CH2CH2 ~, ~ CH = CH- (cis and trans-conformers), -COCH2-, -CH (OH) CH2 - and - CH2SO-, by methods known in the art and described in more detail in the following references: Spatola, Chemistry and Biochemistry of Amino Acids, Peptides, and Proteins, (Weinstein, ed.), Marcel Dekker: New York, p. 267, 1983; Spatola, Peptide Backbone Modifications 1: 3, 1983; Morley, Trends Pharm. Sci. Pgs. 463-468, 1980; Hudson, et al., Int. J. Pept. Prot. Res. 14: 177-185, 1979; Spatola, et al., Life Sci. 38: 1243-1249, 1986; Hann, J. Chem. Soc. Perkin Trans. I 307-314, 1982; Almquist, et al., J. Med. Chem. 23: 1392-1398, 1980; Jennings-White, et al., Tetrahedron Lett. 23: 2533, 1982; Szelke, et al., EP045665A; Holladay, et al., Tetrahedron Lett. 24: 4401-4404, 1983; and Hruby, Life Sci. 31: 189-199, 1982; each of which is included herein by reference. Said mimetic peptides have significant advantages over polypeptide applications, which include for example, more economical production cost, greater chemical stability or better pharmacological properties (such as half-life, absorption, potency, efficacy, etc.), reduced antigenicity, and other properties. The mimetic analogs of the peptides of the invention can also be obtained using the principles of conventional or rational drug design. (see, eg, Andrews, et al., Proc. Alfred Benzon Symp. 28: 145-165, 1990; McPherson, Eur. J. Biochem. 189: 1-24, 1990; Hol, et al., in Molecular Recognition: Chemical and Biochemical Problems, (Roberts, ed.); Royal Society of Chemistry; p. 84-93, 1989a; Hol, Arzneim-Forsch. 39: 1016-1018, 1989b; Hol, Agnew Chem. Int. Ed. Engl. 25: 767-778, 1986; whose disclosures are included herein by reference). According to conventional drug design methods, the desired mimetic molecules can be obtained by randomly evaluating molecules whose structures have an attribute in common with the structure of a "native" peptide. The quantitative contribution that is obtained by changing a specific group of a binding molecule can be determined by measuring the biological activity of the putative mimic substance as compared to the activity of the peptide. In a rational drug design application, the mimetic substance is designed to share an attribute of the more stable three-dimensional conformation of the peptide. Thus, for example, the mimetic substance may be designed to possess chemical groups that are oriented in such a way as to be sufficient to cause ionic, hydrophobic or van der Waals interactions similar to those exhibited by the peptides of the invention, such as it is disclosed in the present. A method for performing rational mimetic substance design employs a computerized system capable of forming a representation of the three-dimensional structure of the peptide, such as those illustrated by Hol, 1989a; Hol, 1989b; and Hol, 1986. The molecular structures of the mimetic peptides, the mimetic organic molecules and the mimetic chemicals of the peptides of the invention can be produced using computer-aided design programs commercially available in the field. Some examples of such programs include SYBYL 6.5®, HQSAR ™ and ALCHEMY 2000 ™ (Tripos); GALAXY ™ and AM2000 ™ (AM Technologies, Inc., San Antonio, TX); CATALYST ™ and CERIUS ™ (Molecular Simulations, Inc., San Diego, CA); CACHE PRODUCTS ™, TSAR ™, AMBER ™, and CHEM-X ™ (Oxford Molecular Products, Oxford, CA) and CHEMBUILDER3D ™ (Interactive Simulations, Inc., San Diego, CA).

The mimetic peptides, the mimetic organic molecules and the mimetic chemicals produced using the peptides disclosed herein, using, for example, molecular modeling programs recognized in the field, can be produced using conventional synthetic chemical techniques and designed in such a way as to allow ultrafast screening, including combinatorial chemistry methods. The combinatorial methods useful for producing the mimetic peptides, the mimetic organic molecules and the mimetic chemicals of the invention include solid phase synthesis and combinatorial chemistry series, according to, for example, SIDDCO (Tuscon, Arizona); Tripos, Inc .; Calbiochem / Novabiochem (San Diego, CA); Symyx Technologies, Inc. (Santa Clara, CA); Medichem Research, Inc. (Lemont, IL); Pharm-Eco Laboratories, Inc. (Bethlehem, PA); or N.V. Organon (Oss, The Netherlands). In the production by combinatorial chemistry of the mimetic peptides, the mimetic organic molecules and the mimetic chemicals of the invention, the methods known in the art may be used, including, but not limited to, the techniques disclosed in Terrett, (Combinatorial Chemistry, Oxford University Press, London, 1998); Gallop, et al., J. Med. Chem. 37: 1233-51, 1994; Gordon, et al., J. Med. Chem. 37: 1385-1401, 1994; Look, et al., Bioorg. Med. Chem. Lett. 6: 707-12, 1996; Ruhland, et al., J. Amer. Chem. Soc. 118: 253-4, 1996; Gordon, et al., Acc. Chem. Res. 29: 144-54, 1996; Thompson and Ellman, Chem. Rev. 96: 555-600, 1996; Fruchtel and Jung, Angew. Chem. Int. Ed. Engl. 35: 17-42, 1996; Pavia, "The Chemical Generation of Molecular Diversity", Network Science Center, www.netsci.org, 1995; Adnan, et al., "Solid Support Combined Chemistry in Lead Discovery and SAR Optimization," Id., 1995; Davies and Briant, "Combinatorial Chemistry Library Design using Pharmacophore Diversity", Id., 1995; Pavia, "Chemically Generated Screening Libraries: Present and Future", Id., 1996; and U.S. Patent Nos. 5,880,972, 5,463,564, 5,331,573 and 5,573,905. The newly synthesized polypeptides can be substantially purified by preparative high performance liquid chromatography (see, e.g., Creighton, Proteins: Structures And Molecular Principles, WH Freeman and Co., New York, N.Y., 1983). The composition of a synthetic polypeptide of the present invention can be confirmed by analysis or amino acid sequencing, for example, by the Edman degradation procedure (Creighton, supra). In addition, any portion of the amino acid sequence of the polypeptide can be altered during direct synthesis and / or combined using chemical methods with the sequences of other proteins, in order to produce a variant of the polypeptide or a fusion polypeptide. Also included in this invention are antibodies and antibody fragments that selectively bind the polypeptides of this invention. Any type of antibody known in the field can be generated using methods well known in the art. For example, an antibody can be generated to bind specifically to an epitope of a polypeptide of this invention. The term "antibody", as used herein, includes intact immunoglobulin molecules, as well as fragments of etas, such as Fab, F (ab ') 2 and Fv, which are capable of binding to an epitope of a polypeptide of this invention. Generally, at least 6, 8, 10 or 12 contiguous amino acids are required to form an epitope. However, epitopes that involve non-contiguous amino acids may require more amino acids, for example, at least 15, 25 or 50 amino acids. An antibody that specifically binds to an epitope of a polypeptide of this invention can be used for therapeutic purposes, as well as in immunochemical assays, eg, Western blot, ELISA assays, radioimmunoassays, immunohistochemical assays, immunoprecipitations or other immunochemical assays. known in the field. Various immunoassays can be used to identify antibodies having the desired specificity. Numerous protocols for competitive binding or immunoradiometric assays are well known in the art. Generally, said immunoassays involve measuring the formation of a complex between an immunogen and an antibody that specifically binds the immunogen.

In general, an antibody that specifically binds to a polypeptide of this invention provides a detection signal at least 5, 10 or 20 times higher than a detection signal provided with other proteins when used in an immunochemical assay. Preferably, antibodies that specifically bind to a polypeptide of this invention do not detect other proteins in immunochemical assays and can immunoprecipitate a polypeptide of this invention from the solution. The polypeptides of this invention, or fragments thereof, can be used to immunize a mammal, such as a mouse, a rat, a rabbit, a guinea pig, a monkey or a human being, to produce polyclonal antibodies. If desired, a polypeptide of this invention or a fragment thereof can be conjugated to a carrier protein, such as bovine serum albumin, thyroglobulin and keyhole limpet hemocyanin. Depending on the host species, various adjuvants can be used to increase the immune response. Such adjuvants include, but are not limited to, Freund's adjuvant, mineral gels (eg, aluminum hydroxide) and surface-active substances (eg, lysolecithin, Pluronic polyalcohols, polyanions, peptides, oily emulsions, limpet hemocyanin. Californian and dinitrophenol). Among the adjuvants used in humans, BCG (Bacillus Calmette-Guerin) and Corynebacterium parvum are especially useful. Monoclonal antibodies that specifically bind to a polypeptide of this invention or a fragment thereof can be prepared using any technique that allows the production of antibody molecules by continuous cell lines in culture. These techniques include, but are not limited to, the hybridoma technique, the human B-cell hybridoma technique and the hybridoma technique with the Epstein Barr virus (Epstein-Barr virus, EBV) (Kohier, et al., Nature 256 : 495-97, 1985; Kozbor, et al., J. Immunol, Methods 81: 3142, 1985; Cote, et al., Proc. Nati, Acad. Sci. 80: 2026-30, 1983; Cole, et al. ., Mol. Cell Biol. 62: 109-20, 1984).

In addition, techniques developed for the production of "chimeric antibodies", the splicing of mouse antibody genes to human antibody genes can be used to obtain a molecule with the appropriate antigenic specificity and biological activity (Morrison, et al. al., Proc. Nati, Acad. Sci. 81: 6851-55, 1984; Neuberger, et al., Nature 312: 604-08, 1984; Takeda, et al., Nature 314: 452-54, 1985). Monoclonal antibodies and other antibodies can also be "humanized" to prevent a patient from developing an immune response against the antibody when used for therapeutic purposes. These antibodies may be sufficiently similar to human antibodies, in terms of sequence, to be used directly in therapies or may require the alteration of some key residues. The differences with respect to the sequence between rodent and human antibodies can be minimized by replacing the residues that differ from those present in human sequences by mutagenesis of individual residues directed to the sites or by screening entire regions that determine complementarity. Alternatively, humanized antibodies can be produced using recombinant methods (see, e.g., GB2188638B). Antibodies that specifically bind to a polypeptide of this invention may contain antigen binding sites that are partially or fully humanized, as disclosed in U.S. Patent No. 5,565,332. Alternatively, the techniques described for the production of single chain antibodies can be adapted using methods known in the art to produce single chain antibodies that specifically bind to a polypeptide of this invention. Antibodies with related specificity, but of different idiotypic composition can be generated by the exchange of chains of random immunoglobulin combinatorial libraries (Burton, Proc Nati Acad Sci 88: 11120-23, 1991). Single chain antibodies can also be constructed using a DNA amplification method, such as the polymerase chain reaction (polymerase chain reaction, PCR), which uses cDNA of the hybridoma as a template (Thirion, et al., Eur. J. Cancer Prev. 5: 507-11, 1996). The single chain antibodies may be monospecific or bispecific, and may be bivalent or tetravalent. The preparation of tetravalent, bispecific single chain antibodies is described, for example, in Coloma and Morrison (Nat. Biotechnol.15: 159-63, 1997). The preparation of bivalent, bispecific single chain antibodies is described in Mallender and Voss (J. Biol. Chem. 269: 199-206, 1994).

A nucleotide sequence encoding a single chain antibody can be constructed using manual or automatic nucleotide synthesis, cloned into an expression construct using standard recombinant DNA methods and introduced into a cell to express the coding sequence, as describe below. Alternatively, single chain antibodies can be produced using directly, for example, filamentous phage technology (Verhaar, et al., Int. J. Cancer 61: 497-501, 1995; Nicholls, et al., J. Immunol., Meth. 165: 81-91, 1993). Antibodies that specifically bind to a polypeptide of this invention can also be produced by inducing in vivo production in the lymphocyte population or by screening immunoglobulin libraries or panels of highly specific binding reagents as disclosed in the literature (Orlandi, et al. , Proc. Nati, Acad. Sci. 86: 38333-37, 1989; Winter, et al., Nature 349: 293-99, 1991). Other types of antibodies can be made and used for therapeutic purposes in the methods of the invention. For example, chimeric antibodies can be made, as disclosed in WO 93/03151. Binding proteins that are derived from immunoglobulins and that are multivalent and multispecific, such as "bivalent fragments" can also be prepared (see, eg, WO 94/13804). Human antibodies that have the ability to bind to the polypeptides of this invention can also be identified from the library HuCAL® from MorphoSys, as indicated below. A polypeptide of this invention can be coated in a well plate and incubated with the phage library linked to HuCAL® Fab from MorphoSys. Those Fabs linked to phages that do not bind to the polypeptide of this invention can be removed from the plate by washing, leaving only the phage that binds tightly to the polypeptide of this invention. The bound phage can be eluted, for example, by a change in pH or by elution with E. coli, and amplified by infection of E. coli hosts. This screening process known as "panning" can be repeated once or twice to obtain a larger population of antibodies that bind tightly to the polypeptide of this invention. Then, the Fabs from the enriched pool are expressed, purified and screened in an ELISA assay. The antibodies according to the invention can be purified by methods well known in the art. For example, the antibodies can be purified by affinity by passing them through a column to which a polypeptide of this invention is attached. Then, the bound antibodies can be eluted from the column using a buffer solution with a high salt concentration. Various terms are defined below, as used herein. In presenting the elements of the present invention or the preferred application (s) thereof, the articles "a", "the (a)" and "said (a)" are intended. mean that there is one or more of the elements. It is intended that the terms "comprising", "including" and "having" be inclusive and mean that there may be additional elements other than the aforementioned elements. The term "subject" as used herein includes mammals (e.g., humans and animals). The term "treatment" includes any process, action, application, therapy or other similar procedure, in which a subject, including a human being, receives medical assistance in order to improve the condition of the subject, direct or indirectly, or slow down the progression of a condition or disorder in the subject. The term "combination therapy" or "co-therapy" means the administration of two or more therapeutic agents to treat a condition and / or disorder. Said administration includes the co-administration of two or more therapeutic agents in a substantially simultaneous manner, for example, in a single capsule having a fixed proportion of active ingredients or in multiple separate capsules for each inhibiting agent. In addition, such administration includes the use of each type of therapeutic agent in a sequential manner. The phrase "therapeutically effective (a)" means the amount of each agent administered that will achieve the objective of improving the severity of a diabetic condition or disorder, while avoiding or minimizing the side effects associated with a particular therapeutic treatment. The term "pharmaceutically acceptable" means that the element in question is suitable for use in a pharmaceutical product. Since the polypeptides of the present invention have the ability to stimulate insulin secretion from pancreatic islet cells in vitro and cause a decrease in the blood glucose level in vivo, they can be used in the treatment of diabetes, including diabetes. type 2 diabetes (non-insulin-dependent diabetes mellitus). Such treatment can also delay the onset of diabetes and diabetic complications. The polypeptides can be used to prevent subjects with glucose intolerance from developing type 2 diabetes. Other diseases and conditions that can be treated or prevented using the peptides of the invention in methods of the invention include: juvenile hereditary diabetes type 2 (MODY) (Herman, et al., Diabetes 43:40, 1994); latent autoimmune diabetes of the adult (LADA) (Zimmet, et al., Diabetes Med. 11: 299, 1994); glucose intolerance (IGT) (Expert Committee on Classification of Diabetes Mellitus, Diabetes Care 22 (Sup. 1): S5, 1999); the impaired fasting glucose (IFG) (Charles, et al., Diabetes 40: 796, 1991); gestational diabetes (Metzger, Diabetes, 40: 197, 1991); and Metabolic Syndrome X. The polypeptides of the present invention may also be effective in disorders such as obesity, and to treat atherosclerotic disease, hyperlipidemia, hypercholesterolemia, low HDL levels, hypertension, cardiovascular disease (including cardiovascular disease). atherosclerosis, coronary heart disease, coronary artery disease and hypertension), cerebrovascular disease and peripheral vessel disease; and to treat lupus, polycystic ovarian syndrome, carcinogenesis and hyperplasia, asthma, male reproductive problems, ulcers, sleep disorders, disorders of lipid and carbohydrate metabolism, circadian dysfunction, disorders of growth, disorders of energy homeostasis, immune diseases, including autoimmune diseases (eg, systemic lupus erythematosus), as well as acute and chronic inflammatory diseases, and septic shock. The polypeptides of the present invention may also be useful for treating physiological disorders related, for example, with cell differentiation to produce lipid accumulator cells, regulation of insulin sensitivity and blood glucose levels that are related, example, with the abnormal function of pancreatic ß cells, tumors that secrete insulin and / or autoimmune hypoglycaemia due to autoantibodies against insulin, autoantibodies against the insulin receptor or autoantibodies that stimulate pancreatic ß cells, differentiation of macrophages leading to the formation of atherosclerotic plaques, inflammatory response, carcinogenesis, hyperplasia, adipocyte gene expression, adipocyte differentiation, pancreatic ß-cell mass reduction, insulin secretion, tissue sensitivity to insulin, cell proliferation of liposarcoma, enfer polycystic ovary, chronic anovulation, hyperandrogenism, - progesterone production, steroidogenesis, oxidoreduction potential and oxidative aggression in cells, production of nitric oxide synthase (NOS), increase of gamma glutamyl transpeptidase, catalase, triglyceride levels and HDL cholesterol and LDL in plasma, and for similar uses . The polypeptides of the invention can also be used in methods of the invention to treat the secondary causes of diabetes (Expert Committee on Classification of Diabetes Mellitus, Diabetes Care 22 (Sup. 1): S5, 1999). These secondary causes include excess glucocorticoids, excess growth hormone, pheochromocytoma and pharmacogenic diabetes. Drugs that can cause diabetes include, but are not limited to, pyriminil, nicotinic acid, glucocorticoids, phenytoin, thyroid hormone, ß-adrenergic agents, interferon-a, and drugs used to treat HIV infection. In addition, the polypeptides of the invention can be used for the treatment of asthma (Bolin, et al., Biopolymer 37: 57-66, 1995; . 677,419; showing that the R3P0 polypeptide has activity to relax the smooth muscle of the guinea pig trachea); for the induction of hypotension (VIP causes hypotension, tachycardia and facial flushing in asthmatic patients (Morice, et al., Peptides 7: 279-280, 1986; Morice, et al., Lancet 2: 1225-1227, 1983) for male reproductive problems (Siow, et al., Arch. Androl. 43 (1): 67-71, 1999); as an antiapoptosis / neuroprotective agent (Brenneman, et al., Ann. N. Y.

Acad. Sci. 865: 207-12, 1998); for cardioprotection during ischemic events (Kalfin, et al., J. Pharmacol. Exp. Ther.1268 (2): 952-8, 1994; Das, et al., Ann. NY Acad. Sci. 865: 297-308 , 1998), for the manipulation of the circadian clock and related disorders (Hamar, et al., Cell 109: 497-508, 2002; Shen, et al., Proc. Nati Acad. Sci. 97: 11575-80, 2000) and finally, as an antiulcer agent (Tuncel, et al., Ann. N. Y. Acad. Sci. 865: 309-22, 1998). The polypeptides of the present invention can be used alone or in combination with additional therapies and / or compounds that are known to those skilled in the art in the treatment of diabetes and disorders. related Alternatively, the methods and polypeptides described herein may be used, partially or completely, in combination therapy. The polypeptides of the invention can also be administered in combination with other therapies known to treat diabetes, including PPAR agonists, sulfonylureas, non-sulfonylureas secretagogues, α-glucosidase inhibitors, insulin sensitizers, insulin secretagogues, compounds that decrease hepatic glucose production, insulin and anti-obesity drugs. Said therapies can be administered before the administration of the polypeptides of the invention, together with these or subsequent to them. Insulin includes short and long-acting forms, and insulin formulations. The PPAR agonist may include agonists from any of the sub-units of PPAR or combinations thereof. For example, the PPAR agonist may include the PPAR-a, PPAR- ?, PPAR-d agonists, or any combination of two or three of the PPAR subunits. PPAR agonists include, for example, rosiglitazone, troglitazone, and pioglitazone. Sulfonylureas include, for example, glyburide, glimepiride, chlorpropamide, tolbutamide and glipizide. A-glucosidase inhibitors that may be useful for treating diabetes upon administration with a polypeptide of the invention include Precose®, Miglitol® and Voglibose ™. Insulin sensitizers that may be useful for treating diabetes include PPAR-? Agonists such as glitazones (eg, troglitazone, pioglitazone, englitazone, MCC-555, rosiglitazone, and the like); the biguanides, such as metformin and phenformin; inhibitors of the protein tyrosine phosphatase-1B (PTP-1 B); inhibitors of dipeptidyl peptidase IV (DP-IV); and the thiazolidinediones and not thiazolidinediones. Compounds that decrease hepatic glucose production that may be useful in treating diabetes by administering them with a polypeptide of the invention include metformin, such as Glucophage® and Glucophage XR®. Insulin secretagogues that may be useful for treating diabetes upon administration with a polypeptide of the invention include the sulfonylureas and non-sulfonylureas: GLP-1, gastric inhibitory peptide (Gastric inhibitor peptide, GIP), secretin, nateglinide, meglitinide, repaglinide, glibenclamide, glimepiride, chlorpropamide and glipizide. GLP-1 includes GLP-1 derivatives that have longer half-lives than native GLP-1, such as derivatized fatty acid GLP-1 and exendin. In one application of the invention, the polypeptides of the invention are used in combination with insulin secretagogues to increase the sensitivity of pancreatic β cells to the insulin secretagogue. The polypeptides of the invention can also be used in methods of the invention in combination with anti-obesity drugs. Anti-obesity drugs include β-3 agonists; the CB-1 antagonists; the neuropeptide Y inhibitors; the anorectics, such as, for example, sibutramine (Meridia®, Abbott Laboratories); and lipase inhibitors, such as, for example, orlistat (Xenical®, Roche Pharmaceuticals). The polypeptides of the invention can also be used in methods of the invention in combination with drugs commonly used to treat lipid disorders in diabetic patients. Such drugs include, but are not limited to, HMG-CoA reductase inhibitors., nicotinic acid, lipid-lowering drugs (eg, stanol esters, sterol glycosides such as tiqueside and azetidinones such as ezetimibe), acetyl-CoA-C-acetyltransferase inhibitors (acetyl-CoA C-acetyItransferase, ACAT) (such as avasimiba), bile acid sequestrants, bile acid reuptake inhibitors, inhibitors of microsomal triglyceride transport and fibric acid derivatives. Inhibitors of HMG-CoA reductase include, for example, lovastatin, simvastatin, pravastatin, fluvastatin, atorvastatin, rivastatin, itavastatin, cerivastatin and ZD-4522. Fibric acid derivatives include, for example, clofibrate, fenofibrate, bezafibrate, ciprofibrate, beclofibrate, etofibrate and gemfibrozil. Kidnappers include, for example, cholestyramine, colestipol and the dialkylaminoalkyl derivatives of a crosslinked dextran.

The polypeptides of the invention can also be used in combination with antihypertensive drugs, such as, for example, β-blockers and angiotensin converting enzyme (angiotensin converting enzyme) inhibitors. Examples of additional antihypertensive agents for use in combination with the polypeptides of the present invention include calcium channel blockers (type L and type T, eg, diltiazem, verapamil, nifedipine, amlodipine and mibefradil), diuretics (p. eg chlorothiazide, hydrochlorothiazide, flumetiazide, hydroflumethiazide, bendroflumethiazide, methylchlorothiazide, trichloromethiazide, polythiazide, benzthiazide, ethacrynic acid ticrinaphene, chlorthalidone, furosemide, musolimine, bumetanide, triamterene, amiloride, spironolactone), renin inhibitors, inhibitors of ACE (eg, captopril, zofenopril, fosinopril, enalapril, ceranopril, cilazopril, delapril, pentopril, quinapril, ramipril, lisinopril), angiotensin 1 receptor (AT-1) antagonists (eg, losarían, irbesartan, valsarians), endoyelin receptor (ET) antagonisms (eg, sifaxenia, airasemia, neutral endopeptidase inhibitors (neutral endopeptides) e, NEP), vasopepidase inhibitors (dual inhibitors of NEP-ACE) (p. eg, omapairilat and gemopafrilat) and niíraíos. Said co-therapies may be administered in any combination of two or more drugs (eg, a polypeptide of the invention in combination with an insulin sensitizer and an anti-obesity drug). Said co-therapies can be administered in the form of pharmaceutical compositions, as described above. On the basis of well-known assays used to determine the efficiency for the processing of the conditions identified earlier in mammals and by comparing these results with the results of known drugs used to trawl these conditions, the effective dose of the polypeptides of this invention can be easily determined for the processing of each desired indication. The principle of a positive principle (eg, polypeptides) that will be administered in the context of one of these conditions may vary greatly, depending on considerations such as the specific polypeptide and the dosage unit used, the mode of administration, the period of time, the age and sex of the rape patient, and the nature and exigency of the rarity condition. In general, the viral content of the active principle to be administered may range from about 0.00001 mg / kg to 1 mg / kg, and preferably ranges from about 0.0001 mg / kg to 0.1 mg / kg of body weight per day. A dosage unit may confer between about 0.01 mg and 20 mg of active ingredient, and may be administered one or more times per week. The weekly dose for administration by injection, including intravenous, intramuscular, subcutaneous and parenteral injections, and the use of infusion techniques can range from about 0.0001 to 01 mg / kg. The daily rectal dosage regimen may range from 0.001 to 1 mg / kg of total body weight. The transdermal concentration may be that required to maintain a daily dose of between 0.001 and 1 mg / kg. Of course, the initial and continuous dosage regimen specific to each patient will vary depending on the nature and severity of the condition, as determined by the referring physician making the diagnosis, the activity of the specific polypeptide used, the age of the patient, the diet of the patient, the time of administration, the route of administration, the rate of drug excretion, drug combinations and other similar factors. The desired mode of treatment and the dose amount of a polypeptide of the present invention can be determined by those skilled in the art using conventional treatment tests. The polypeptides of this invention can be used to achieve the desired pharmacological effect by administering to a patient in need thereof a pharmaceutically formulated pharmaceutical composition. For the purposes of this invention, a mammal is included per patient, including humans, that needs treatment for a specific condition or disease. Therefore, the present invention includes pharmaceutical compositions that are composed of a pharmaceutically acceptable carrier and an amount effective ferapéuficamenie of a polypeptide. A pharmaceutically acceptable vehicle is any vehicle that is relatively non-toxic and harmless to a patient at concentrations comparable to the effective action of the active principle, so that any side effects attributable to the vehicle do not contaminate the beneficial effects of the active principle. A therapeutically effective amount of a polypeptide is the amount that produces a result or exerts an influence on the specific condition that is being brought about. The polypeptides disclosed herein can be administered with a pharmaceutically acceptable carrier using any effective conventional dosage unit form, including, for example, immediate and prolonged release preparations, oral, parenteral or topical administration, or otherwise. The polypeptides of the invention can be administered parenterally, ie, intravenously, intramuscularly, interperitoneally or preferably, subcutaneously, * as injectable doses of the polypeptide in a physiologically acceptable diluent with a pharmaceutic vehicle which can be a liquid or a mixture of sterile liquids, such as water, saline, aqueous dexory, and related sugar solutions; an alcohol, such as ethanol, isopropanol or hexadecyl alcohol; glycols such as propylene glycol or polyethylene glycol; glycerol ketals, such as 2,2-dimethyl-1,1-dioxolane-4-mephanol; eferes such as poly (ethylene glycol) 400; an aceife; a fatty acid; an ester or glyceride of fatty acid; or an oily fatty acid glyceride with or without the addition of a pharmaceutically acceptable surfactant, such as a soap or detergent, a suspending agent, such as pectin, carbomers, methylcellulose, hydroxypropylmethylcellulose or carboxymethylcellulose, or an emulsifying agent and other pharmaceutical adjuvants. Some examples of oils that can be used in the parenteral formulations of this invention are those based on petroleum, animal, vegetable or synthetic origin, for example, peanut, soybean, sesame, cotton, corn or olive oil, Vaseline and mineral oil. Suitable fatty acids include oleic acid, stearic acid and isostearic acid.

Examples of suitable fatty acids are, for example, eyl oleate and isopropyl myristate. Suitable soaps include those of fatty alkali metal, ammonium and triethylamine salts, and suitable detergents include cationic detergents, for example, dimethyl dialkyl ammonium halides, alkyl pyridinium halides and alkylamine achetalides; anionic detergents, for example, the alkyl, aryl and olefin sulfonates; the alkyl, olefin, ether and monoglyceride sulfates; and the sulfosuccinates; non-ionic detergents, for example, fatty amine oxides, fatty acid alkanolamides and polyoxyethylene polypropylene copolymers; and amphoteric detergents, for example, alkyl-beta-aminopropionates and 2-alkylimidazoline quaternary ammonium salts, as well as mixtures. In general, the parenteral compositions of this invention may contain about 0.5% to 25% based on the weight of the active ingredient in the solution. Preservatives and buffer solutions can also be used in a venious way. In order to minimize or eliminate the irriiation at the injection site, it is possible for said compositions to contain a nonionic surfactant having a hydrophilic-lipophilic balance (HLB) of between about 12 and about 17. The amount of surfactant present in said formulation ranges from about 5% to 15% by weight. The surfactant may be a single component having the aforementioned HLB or it may be a mixture of two or more components that contains the desired HLB. Some examples of the surfacides used in parenteral formulations are the class of fatty acid esters of polyethylene sorbitan, for example, sorbitan monooleate and the adducts of molecular weight of ethylene oxide with a hydrophobic base, formed by condensation. of propylene oxide with propylene glycol. The pharmaceutical compositions may take the form of sterile injectable aqueous suspensions. Said suspensions may be formulated according to known methods using dispersing or wetting agents and suitable suspension agents, such as, for example, sodium carboxymethyl cellulose, methyl cellulose, hydroxypropylmethyl cellulose, sodium alginate, polyvinyl pyrrolidone, gum arabic gum and gum arabic; dispersing agents or humectanis which may be a naturally occurring phosphatide, such as lecithin, a product of the condensation of an alkylene oxide with a fatty acid, for example, polyoxyethylene stearate, a product of the condensation of an ethylene oxide with an alcohol long-chain aliphatic, for example, heptadecaethylenexiceanol, a product of the condensation of an ethylene oxide with a partial ester derived from a fatty acid and a hexitol, such as polyoxyethylene sorbitan monooleate, or a product of the condensation of an oxide of ethylene with a partial ester derived from a fatty acid and a hexitol anhydride, for example, polyoxyethylene sorbitan monooleate. The sterile injectable preparation may also be a sterile injectable solution or suspension in a parenterally acceptable non-toxic diluent or solvent. The diluents and solvents that can be used are, for example, water, Ringer's solution and isotonic sodium chloride solution. In addition, sterile fixed oils are conventionally employed as solvents or suspending media. For this purpose, any soft fixed oil, including monoglycerides or synthetic diglycerides, may be employed. In addition, fatty acids such as oleic acid can be used in the preparation of injectable forms. A composition of the invention can also be administered in the form of suppositories of the drug for rectal administration. These compositions can be prepared by mixing the drug (eg, polypeptide) with an appropriate non-irritating excipient which is solid at usual temperatures, but liquid at the rectal temperature and which, consequently, will melt in the rectum to release the drug. Such materials include, for example, cocoa butter and polyethylene glycol. Another formulation employed in the methods of the present invention employs transdermal delivery devices ("patches"). Said patches Transdermal agents can be used to provide a continuous or discontinuous infusion of the polypeptides of the present invention in controlled amounts. The preparation and use of transdermal patches for the administration of pharmaceutical agents is well known in the art (see, eg, US Pat. No. 5,023,252, which is included in the present by reference). Said patches can be elaborated for the administration of pharmaceutical agents in a continuous, pulsatile or on demand form. It may be desirable or necessary to introduce the pharmaceutical composition into the patient through a mechanical delivery device. The development and use of mechanical delivery devices for the administration of pharmaceutical agents is well known in the art. For example, direct techniques for administering a drug directly to the brain generally involve the placement of a catheter for administration of the drug into the patient's ventricular system in order to bypass the blood-brain barrier. One such implantable delivery system that is used for the transport of agents to specific anatomical regions of the body is described in U.S. Patent No. 5,011,472, which is included herein by reference. The compositions of the invention may also contain other conventional pharmaceutically acceptable composition ingredients, which are generally referred to as carriers or diluents, as needed or required. Any of the compositions of this invention can be preserved by the addition of an antioxidant, such as ascorbic acid, or by other suitable preservatives. Conventional procedures can be used to prepare said compositions in appropriate galenic forms. Commonly used pharmaceutical ingredients whose use may be appropriate to formulate the composition for their intended administration route include: acidifying agents, for example, by way of example, acrylic acid, citric acid, fumaric acid, hydrochloric acid, nitric acid; and alkalizing agents, for example, to a mere enunciative title, ammonia solution, ammonium carbonate, diethanolamine, monoethanolamine, potassium hydroxide, sodium borate, sodium carbonate, sodium hydroxide, triethanolamine, trolamine.

Other pharmaceutical ingredients include, but are not limited to, adsorbents (e.g., powdered cellulose and activated charcoal); aerosol propellants (e.g., carbon dioxide, CCI2F2, F2CIC-CCIF2 and CCIF3); air displacement agents (eg, nitrogen and argon); antifungal preservatives (eg, benzoic acid, butylparaben, eilaparaben, methylparaben, propylparaben, sodium benzoate); antimicrobial preservatives (eg, benzalkonium chloride, benzethonium chloride, benzyl alcohol, cetylpyridinium chloride, chlorobutanol, phenol, phenylethyl alcohol, phenylmercury nitrate and thimerosal); antioxidants (eg, ascorbic acid, ascorbyl palmitate, butylated hydroxyanisole, butylated hydroxytoluene, hypophosphorous acid, monothioglycerol, propyl galaxido, sodium ascorbate, sodium bisulfite, sodium formaldehyde sulfoxylate, sodium metabisulfite); bonding materials (eg, block polymers, natural and synthetic rubber, polyacrylates, polyurethanes, silicones and styrene-butadiene copolymers); buffering agents (eg, potassium mephosphate, potassium monobasic phosphate, sodium acetate, sodium citrate anhydrous and dihydrate sodium citrate); vegetables (eg, acacia syrup, aromatic syrup, aromatic elixir, cherry syrup, cocoa syrup, orange syrup, syrup, corn oil, mineral oil, peanut oil, sesame oil, bacteriostatic chloride injection sodium and bacteriostatic water for injection); chelating agents (e.g., disodium edetate and edetic acid); Aeolian (e.g., FD &C red No. 3, FD &C red No. 20, FD &C yellow No. 6, FD &C blue No. 2, D &C green No. 5, D &C orange No. 5, D &C red No. 8, caramel and red ferric oxide); clarifying agents (eg, bentonia); emulsifying agents (by way of example, acacia, cetomacrogol, cetyl alcohol, glyceryl monostearate, lecithin, sorbitan monooleate, polyethylene stearate 50); encapsulating agents (e.g., gelatin and cellulose acetal phthalate); flavors (eg, anise oil, cinnamon oil, cocoa, menthol, orange oil, mint and vanilla oil); humectants (e.g., glycerin, propylene glycol and sorbitol); levigating agents (eg, mineral oil and glycerin); oils (eg, peanut oil, mineral oil, olive oil, peanut oil, sesame oil and vegetable oil); ointment bases (eg lanolin, hydrophilic ointment, polyethylene glycol ointment, petroleum jelly, hydrophilic vaseline, white ointment, yellow ointment and rose water ointment); elements that increase penetration (transdermal administration) (eg, monohydroxy or polyhydroxy alcohols, saturated or unsaturated fatty alcohols, saturated or unsaturated fatty esters, saturated or unsaturated dicarboxylic acids, essential oils, phosphatidyl derivatives, cephalins, terpenes, amides, ethers, ketones and ureas); plasticizers (eg, diethyl phthalate and glycerin); solvents (eg, alcohol, corn oil, cottonseed oil, glycerin, isopropyl alcohol, mineral oil, oleic acid, peanut oil, purified water for injection, sterile water for injection and sterile water for irrigation); hardening agents (eg, cetyl alcohol, cetyl ester wax, microcrystalline wax, paraffin, styrene alcohol, white wax and yellow wax); bases for suppositories (eg, cocoa butter and polyethylene glycols (mixtures)); surfactants (e.g., benzalkonium chloride, nonoxynol 10, oxtoxinol 9, polysorbate 80, sodium lauric sulfate and sorbitan monopalmitate); suspending agents (eg, agar, bentonite, carbomers, sodium carboxymethylcellulose, hydroxyethylcellulose, hydroxypropylcellulose, hydroxypropylmethylcellulose, kaolin, methylcellulose, tragacane and Veegum); sweeteners (eg, aspartame, dexadose, glycerin, mannitol, propylene glycol, sodium saccharin, sorbiol, and sucrose); antiadherents for tablets (eg, magnesium stearate and talc); binders for tablets (eg, acacia, alginic acid, sodium carboxymethylcellulose, compressible sugar, ethyl cellulose, gelatin, liquid glucose, methylcellulose, povidone and pregelatinized starch); diluents for tablets and capsules (eg, dibasic calcium phosphate, kaolin, lactose, mannitol, microcrystalline cellulose, powdered cellulose, precipitated calcium carbonate, sodium carbonate, sodium phosphate, sorbitol and starch); agents for coating tablets (eg, liquid glucose, hydroxyethylcellulose, hydroxypropylcellulose, hydroxypropylmethylcellulose, methylcellulose, ethylcellulose, cellulose acetate phthalate and shellac); excipients for the direct compression of tablets (eg, dibasic calcium phosphate); tablet disintegrants (e.g. eg, alginic acid, calcium carboxymethyl cellulose; microcrystalline cellulose, potassium polacriline, sodium alginate, sodium starch glycolate and starch); slides for tablets (eg, colloidal silica, corn starch and talc); lubricants for tablets (eg, calcium stearate, magnesium stearate, mineral oil, stearic acid and zinc stearate); opacifiers for tablets / capsules (e.g., titanium dioxide); agents for polishing tablets (eg, Carnauba wax and white wax); thickening agents (eg, beeswax, cetyl alcohol and paraffin); tonicity agents (e.g., dexadose and sodium chloride); agents that increase viscosity (eg, alginic acid, bentonite, carbomers, sodium carboxymethylcellulose, methylcellulose, povidone, sodium alginate and tragacanth); and wetting agents (e.g., hepyadecaethylene oxyethanol, lecithins, polyethylene sorbitol monooleate, polyoxyethylene sorbitol monooleate and polyoxyethylene stearate). The polypeptides described herein can be administered as single pharmaceutical agents or in combination with one or more other pharmaceutical agents, where the combination does not cause unacceptable side effects. For example, the polypeptides of this invention can be combined with anti-obesity agents, anti-diabetic agents or agents for other known indications, and similar agents, as well as with mixtures and combinations thereof. The polypeptides described herein may also be used, in free base form or in compositions, for research and diagnostic purposes, or as analytical reference standards, and for other similar uses. Therefore, the present invention includes compositions comprising an inert carrier and an effective amount of a polypeptide identified by the methods described herein, or a salt or ester thereof. An inert carrier is any material that does not interact with the polypeptide that will be transported and that provides a support, a transport medium, volume, a traceable material, enyre things, to the polypeptide that will be transported. An effective amount of Polypeptide is the amount that produces a result or exerts an influence on the specific procedure that is being performed. It is known that the polypeptides undergo hydrolysis, deamidation, oxidation, racemization and isomerization in aqueous and non-aqueous media. Degradation by hydrolysis, deamidation or oxidation, for example, can be easily detected by capillary electrophoresis, mass spectrometry or Edman degradation. Without detriment to enzymatic degradation, polypeptides having a prolonged plasma half-life or biological residence time must, at a minimum, be stable in aqueous solution. It is essential that the polypeptide exhibit a degradation of less than 10% over a period of one day at body temperature. It is even more preferable that the polypeptide exhibit degradation of less than 5% over a period of one day at body temperature. Because the chronic diabetic patient receives treatment throughout his life, preferably, it is even more convenient to administer these therapeutic agents infrequently, if they are administered parenterally. Stability (ie, a small percentage of degradation) over a period of weeks at body temperature will allow dose administration less frequently. Stability (ie, a small percentage of degradation) to degradation through proteolysis, relative to DPP4 or plasma over periods of days to weeks will further support dose administration less frequently. The stability in years at refrigeration temperature will allow the manufacturer to present a liquid formulation and avoid, in this way, the discomfort of the reconstiution. In addition, the stability in an organic solvent will allow the polypeptide to be formulated in new galenic forms, such as implants. Formulations suitable for subcutaneous, intravenous, intramuscular and other similar administration; the appropriate pharmaceutical vehicles; and techniques for formulation and administration can be prepared by any of the methods well known in the art (see, e.g., Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, Pa., 20th edition, 2000). It should be apparent to a person with ordinary skill in the art that changes and modifications to this invention can be made, without departing from the spirit or scope of the invention, as hereby espoused.

EXAMPLES In order that this invention may be better understood, the following examples are included. These examples are included for illustrative purposes only, and should not be construed as limiting the scope of the invention in any way. All publications mentioned herein are included in their entirety by reference. Example 1. Peptide synthesis methodology To synthesize some of the polypeptides of the invention, the following general procedures were followed: Peptide synthesis was performed by the FMOC / t-butyl strategy (Penningion and Dunn, Peptide Synthesis Protocols, Volume 35, 1994) under continuous flow conditions, using polyethylene glycol-polystyrene resins from Rapp-Polymere (Rapp-Polymere, Tubingen , Germany). Upon completion of the synthesis, the peptides were oiled from the resin and deprotected using TFA / DTT / H2O / triisopropyl silane (88/5/5/2). The peptides were precipitated from the cleavage fluid using cold diethyl ether. The precipitate was washed three times with the cold ether and then dissolved in 5% acetic acid before lyophilization. The peptides were verified by means of an inverted phase chroma-ography on a YMC-Pack ODS-AQ column (YMC, Inc., Wilmington, NC) on an ALLIANCE® system from Waters (Waters Corporation, Milford, MA) using water / acetonitrile with 3% TFA as gradient from 0% to 100% acetonitrile, and by matrix-assisted laser-desorption ionization desorption / ionization mass spectrometry (MALDI) in a mass spectrometer MALDI VOYAGER DE ™ (Model 5-2386-00, PerSeptive BioSystems, Framingham, MA). The peptide sample was added to the Matrix buffer solution (50/50 dH2O / acetonitrile with 3% TFA) in 1/1 ratio. The pepidae that did not meet the purity criteria of > 95% were purified by reverse phase chromatography on a Delta Prep 4000 HPLC System from Waters (Waters Corporation, Milford, MA).

Example 2. Peptide Acetylation The peptides were synthesized by standard methods well known in the art. The peptides were synthesized with an Applied Biosystems 430A peptide synthesizer using FMOC chemistry with activation with HBTU in Rink amide resin, and the N-terminus was acetylated with acetic anhydride. The peptide was cleaved with 84.6% TFA, 4.4% phenol, 4.4% water, 4.4% thioanisole and 2.2% ethanedithiol. The peptides were precipitated from the cleavage fluid using tert-butylmethyl ether. The precipitate was washed with the cold ether and dried with argon. The peptides were purified with inverted phase C18 chromatography with linear gradients of water / acetonitrile with 0.1% TFA. The identity of the peptides was confirmed with MALDI mass spectrometry and electrospray, and with amino acid analysis.

Example 3. Peptide PEGylation The half-life of a peptide in vivo can be increased through the binding of a polyethylene glycol (PEG) fraction to the peptide, whereby the clearance of the peptide by the kidney is reduced and degradation of the peptide by the protease is reduced. The use of a peptide agonist of the VPAC2 receptor is severely limited by its very short half-life in vivo; the binding of a PEG fraction to the peptide (PEGylation) prolonged the peptide half-life sufficiently as to allow an application applied between one time per day and one time per week. The PEGylation can be carried out by any method known to those skilled in the art. However, in this example, PEGylation was performed by introducing a single cysteine mutation in the peptide, followed by PEGylation of the cysteine via a stable thio ether bond between the sulfhydryl of the peptide and the maleimide group of the reagent. methoxy- PEG-maleimide (Nekíar (Inhale / Shearwaíer), San Carlos, CA; NOF, Tokyo, Japan). The single cysteine can be introduced at the C-terminus of the peptide to minimize the possible reduction of activity by PEGylation. As an example, a two-fold molar excess of a mPEG-mal reagent (molecular weight 22kD and 43kD) was added to 1 mg of peptide (e.g., SEQUENCE ID No. 1 with a mutation in the cysteine in the C-terminal peptide) and was dissolved in a reaction buffer at pH 6 (0.1 M sodium phosphate / 0.1 M NaCl / 0.1 M ethylenediamine-acetic acid [EDTA]). After 30 minutes at room temperature, the reaction was terminated with the addition of a two-fold molar excess of dithiothreitol (DTT) to mPEG-mal. The peptide-PEG-mal reactive mixture was applied to a cation exchange column to remove residual PEG reagents, followed by a gel filtration column to remove residual free peptide. Purity, mass and number of PEGylated sites were determined by polyacrylamide gel electrophoresis with sodium dodecyl sulfate (Sodium dodecyl sulphate-PoIyacrylamide gel elecrophoresis, SDS-PAGE) and matrix-assisted laser desorption / ionization mass spectrometry (matrix-assisted laser-desorption ionization, MALD!) - time of flight (time of flight, TOF). When a 22 kD PEG was attached to the peptides of the present invention, the potent activation of the VPAC2 receptor was maintained. Likewise, the activation selectivity of VPAC2 receptors was also maintained, in comparison with VPAC1 and PAC1. It is possible that PEGylation with a smaller PEG (eg, a 22 kD linear PEG) is less likely to reduce the activity of the peptide, whereas a PEG of Larger size (eg, a branched PEG of 43kD) will be more likely to reduce activity. However, the larger PEG will further increase the plasma half-life, so that one injection per week can be administered (Harris, et al., Clin.Pharmacokinet, 40: 539-551, 2001). Example 4. Pharmaceutical composition: formulations IV and SC A sterile IV injectable formulation is prepared with 4 mg of a SEQ ID No. 1 polypeptide, or a derivatized polypeptide having a content equivalent to 4 mg of polypeptide, and 1 L of sterile saline solution, using any manufacturing process well known in the field. For the subcutaneous (SC) formulation, higher concentrations of the polypeptide can be used. In the case of the polypeptide identified as SEQUENCE ID No. 1, or of a derivatized polypeptide, 4 mg are dissolved in 100 mL of saline or dimethylsulfoxide (DMSO), and sterile vials previously subjected to aseptic filtration are filled with the composition. . Example 5. Peptide analysis by mass spectrometry Aliquots of forty pmol / 2 μl of peptides up to 10 μl were diluted with water. The buffer solution of N-2-hydroxy-ethylpiperazine-N'-2-ene-sulphonic acid (N-2-Hydroxy-ethylpiperazine-N'-2-Efhanesulfonic Acid, HEPES) was removed by applying 50% of the sample (20 pmol / 5 μl) to a C18 ZipTip conditioning from Millipore, according to the manufacturer's instructions. The sample was eluted from the ZipTip with a matrix (10 mg / ml of alpha-cyanohydroxycinnamic acid in acetonitrile (ACN) 50%, 0.1% TFA) directly on the MALDI plate. The samples were analyzed in a Voyager DE-PRO MALDI system from Applied Biosystems operated in the ion reflector mode. The data was collected in the range of 500 to 4000 Da, and the resulting masses were compared with the theoretical masses by manual calculations. Example 6. Analysis of Edman peptides Peptide samples were supplied for the Edman degradation at 1 nmol / 10 μl in 10 mM HEPES, pH 7.4, 5% TFA. Before Edman's analysis, he removed the salt from the HEPES buffer using a ProSorb sample cartridge from Applied Biosystems, according to the manufacturer's instructions. Briefly, the sample was applied to a polyvinylidene difluoride membrane (Polyvinylidene difluoride, PVDF) and washed with 0.1% TFA; then the membrane was removed and inserted into the protein sequencer for Edman degradation. The Edman degradation was carried out in a Procise 494HT protein sequencing system from Applied Biosystems, using the pulsed liquid phase method, according to the manufacturer's instructions. Sequence determinations were performed manually. Example 7. Stability of the peptides. The formulations described in Example 4 were placed in a chamber - constant stability. The peptides were also analyzed with respect to their stability to degradation in DPP4 and plasma solutions.

Periodically, samples were removed for analysis by capillary electrophoresis, mass spectrometry, Edman degradation, ELISA and peptide activity assays, which are sensitive methods for detecting the degradation of the polypeptide. The area of several peaks and the area of the polypeptide peak were added - precursor is divided by the total peak area. The quotient is the percentage purity.

Since there are impurities present in the new polypeptide, the change in purity is normalized by dividing the purity at different points in time by the initial purity. To measure stability to DPP4 and plasma, the peptides at 20 pmol / μl were incubated at 37 ° C in the presence of 300 pM DPP-IV in 100 mM HEPES, pH 7.4 (Figure 2a). At various points in time, the reaction (2 μl aliquot) was terminated by the addition of the 1 μM DPP-IV inhibitor and freezing. For the analysis of mass spectrometry MALDl, the following points were evaluated in the time: T = 0 h, 1 h, 5 h and 24 h. The results were plotted as a percentage of peptides or derivatives of intact peptides, in comparison with the degradation products.

Example 8. Binding of the peptides to the PACAP1 and VPAC1 and VPAC2 receptors. CHO cells were grown to confluence with an overexpression of the PAC1, VPAC1 and VPAC2 receptors, then they were scraped from their flasks and pelleted by gentle centrifugation in 50 ml tubes. ml. The pellets were resuspended in an iris-based homogenization buffer and homogenized in a Dounce tissue breaker with 30-40 manual strokes on ice. The suspension was centrifuged in an ultracentrifuge, which produced the pelletization of the membranes. This pellef was resuspended in a small amount of homogenization buffer and a concentration of proteins was determined through the use of a Pierce BCA kit. A binding reaction containing a membrane protein of 10 μg, 0.1 nM 125 I-PACAP 27 and a dose curve of the compound to be evaluated was incubated in a 96-well plate at 37 ° C for 20 minutes. The reaction was stopped by placing the plate on ice for 20 minutes. The reaction was added to a filter plate pre-incubated with 0.1% polyethylenimine (polyethylenimine, PEI) to prevent non-specific binding, processed with a vacuum manifold and washed several times with an albumin-based wash solution. bovine serum (bovine serum albumin, BSA). The filter plate was dried, scintillator was added and read on a MicroBeta counter. The data was analyzed and presented in Prism charts. Example 9. Elevation of cAMP in response to peptides CHO cells expressing VPAC2 peptides were plated in 96-well plates at 8 x 10 4 cells / well and cultured at 37 ° C for 24 hours in aMEM, nucleosides , glutamine (Gibco / BRL, Rockville, MD), fetal bovine serum (fetal bovine serum, FBS) 5%, Pen / Strep 100 μg / mL, hygromycin 0.4 mg / mL and Generation 1.5 mg / mL ( Gibco / BRL). The media was removed and the plates were washed with phosphate buffered saline (PBS). The cells were incubated with a peptide (in mM HEPES, 150 mM NaC1, 5 mM KCl, 2.5 mM CaCl2, 1.2 mM KH2P04, 1.2 mM MgSO4, 25 mM NaHCO3 (pH 7.4) with 1% BSA and 100 μM IBMX) for 15 minutes at 37 ° C. The cyclic AMP of the cell extracts was quantified using the scintillation proximity assay (SPA) direct screened assay system of cAMP (Amersham Pharmacia Biotech Inc., Piscataway, NJ). The amount of cAMP present in the lysates was determined by following the instructions provided with this kit. The amount of cAMP (in pmol) produced at each concentration of the peptide was plotted and analyzed by non-linear regression using the Prizm software to determine the median effective concentration (EC50) for each peptide. Alternatively, the elevation of cAMP in response to receptor activation can be measured in an indicator cell line, such as CHO, which not only expresses the desired receptor, but also expresses an indicator, such as luciferase, bound to a response element to the cAMP (cAMP response element, CRE). Said cells are placed in 96-well plates at the rate of 10 4 cells per well and are cultured at 37 ° C for 48 hours in aMEM, nucleosides, glutamine (Gibco / BRL, Rockville, MD), 5% FBS, Pen / Strep 100 μg / mL, hygromycin 0.4 mg / mL and Geneticin (Gibco / BRL) 1.5 mg / mL. Then the cells are incubated with a peptide for 6 hours, the media are removed and the Bright-Glo reagent (Promega) is added. The signal is delectated using a scintillation counter. The polypeptides of this invention are designed on the basis of VIP, which has been shown to lack activity in PAC1 (Vaudry, et al., Pharmacol, Rev. 52: 269-324, 2000). Therefore, it is believed that the polypeptides of this invention do not possess considerable activity in PAC1. In Figure 3, the results of this assay are shown with representative polypeptides of this invention. The peptides identified as ID of SEQUENCE No. 1 and 112 are potent agonists of the VPAC2 receptor, and activate the receptor at 100% of the maximum level of receptor activation achieved by the endogenous peptide PACAP-27. Also, the peptides identified as ID of SEQUENCE No. 1, 112, 152 and 154 are selective agonists of the VPAC2 receptor and have a very weak agonist activity with respect to VPAC1. PACAP-27 is a potent agonist of VPAC1. Example 10. Insulin secretion from dispersed rat islet cells Insulin secretion from dispersed rat islets mediated by a series of peptides of the present invention was measured as described below. The islets of Langerhans, isolated from SD rats (200-250 g), were digested using collagenase. Scattered islet cells were treated with trypsin, seeded in 96-well V-bottom plates and pelleted. The cells were then cultured overnight in media with or without peptides of this invention. The media was aspirated, and the cells were preincubated with Krebs-Ringer-HEPES buffer containing 3 mM glucose for 30 minutes at 37 ° C. The preincubation buffer was removed, and the cells were incubated at 37 ° C with Krebs-Ringer-HEPES buffer containing the appropriate concentration of glucose (eg, 8 mM), with or without peptides, during a appropriate period. In some studies, an appropriate concentration of GLP-1 was also included. A portion of the supernatant was removed and its insulin content was measured by SPA. The results were expressed as "number of times greater than the control" (fold over control, FOC). In this assay, an increase in insulin secretion from islet cells of dispersed rats was defined as an increase of at least 1, 4 times. The VPAC2 receptor agonist component of the polypeptides of this invention produced an increase in insulin secretion from scattered islet cells of at least between 1, 4, and 1.7 times. Example 11. Generation of specific antibodies against the peptides and measurement of the peptides by ELISA Specific polyclonal antibodies against the polypeptides of the present invention were generated by the synthesis of a specific fragment of a polypeptide of this invention using an ABI 433A peptide synthesizer. Then, the peptide was oiled from the resin and purified in an analytical and preparative HPLC System Gold system from Beckman. A MALDl Perspective mass spectrophotometer system was used to identify the correct product. The peptide was dried using a lyophilizer. Then, the peptide (2 mg) was conjugated with keyhole limpet hemocyanin (KLH) through the free sulfhydryl group of Cys. New Zealand white rabbits were immunized on Day 0, 14, 35, 56 and 77. On Day 0, each rabbit was given a subcutaneous injection of 250 Og of the peptide and complete Freund's adjuvant. Subsequent immunizations used 125 Dg of the peptide per rabbit. The bleeding began on Day 21 and continued at 21-day intervals thereafter. The purification of the antibodies against the peptide was carried out by passing the crude serum through a peptide specific affinity purification column. The titering of antibodies was determined by ELISA. A 96-well Immulon 4HBX plate was coated with a Morphosys C-terminal F (ab) antibody, specific against the peptides of the present invention and allowed to incubate overnight at 4 ° C. Then the plate was blocked to prevent non-specific binding. Then, the peptide standards (2500 ng / mL-160 pg / mL) were diluted in 33% plasma, and the samples were diluted in a 1: 3 ratio in buffer, followed by incubation for 1.5 hours at room temperature. ambient. After washing, a specific polyclonal N-terminal antibody against the peptides of this invention was incubated on the plate for one hour. Next, the horseradish peroxidase conjugated donkey-anti-rabbit antibody (horseradish peroxidase, HRP) was added, and the samples and standards were incubated for an additional hour. Detection was evaluated after incubation with 3,3 ', 5,5'-tetramethylbenzidine (TMB) solution, and the plate was read at OD45o optical density (Figure 4a).

In an α-lactamase form, the 96-well Immulon 4HBX plate was coated with a polyclonal N-terminal antibody, specific against the peptides of the present invention, and allowed to incubate overnight at 4 ° C. Then the plate was blocked to prevent non-specific binding. Then, the peptide standards (2500 ng / mL-160 pg / mL) were diluted in 50% plasma, and the samples were diluted in a 1: 2 ratio in buffer, followed by incubation for 1.5 hours at room temperature. ambient. After washing, a monoclonal anti-PEG antibody specific to the peptides of this invention was incubated for one hour on the plate. Next, the anti-mouse antibody conjugated with horseradish peroxidase (HRP) was added, and the samples and standards were incubated for an additional hour. Detection was evaluated after incubation with 3.3 'solution, 5,5'-tetramethylbenzidine (TMB), and the plate was read at OD450 (Figure 4b).

Example 12. Pharmacokinetics of the peptides after administration of IV and subcutaneous doses The plasma samples are transferred to a microcentrifuge tube and an equivalent volume of acetonitrile is added to the sample (final concentration of 50%). The sample is vortexed for approximately 5 minutes and allowed to stand on ice for 10 minutes. The sample is vigorously vortexed again for approximately 1 minute, and then centrifuged for 30 minutes in a microcentrifuge tube (4 ° C) at the maximum speed (approximately 15,000 x g). After centrifugation, the aqueous phase is carefully transferred to a clean centrifuge tube, and the sample is centrifuged for 5 minutes in a microcentrifuge tube (4 ° C) at the maximum speed (approximately 15,000 x g). The extracted sample is dried under vacuum using a Speed Vac SC110 (Savant) with the intermediate temperature until it dries. The sample is resuspended in an appropriate volume of sterile water and maintained at 4 ° C. Then, the sample is homogenized with ultrasound in an ultrasonic bath for 10 minutes at room temperature before analysis. The peptide concentration is determined using the reporter assay as described in Example 9. Using this method, the pharmacokinetic properties of the SEQUENCE No. 112 were compared with those of SEQUENCE ID No. 154 (Figure 5a). The peptidic acetylation present in SEQUENCE ID No. 112 demonstrates better pharmacokinetic properties, and is decelerated at 24 and 48 hours, while the non-silylated peptide of SEQUENCE ID No. 154 is only deduced at 6 o'clock. . Subsequent studies comparing the two peptides demonstrated that the SEQUENCE ID No. 112 had a half-life of 10.9 hours in the rat, while that of the SEQ ID NO: 154 not acetylated was approximately 6 hours ( Figure 5b). Plasma peptide concentrations can also be measured using the ELISA assay, as described in Example 11. Using this methodology, the pharmacokinetics of SEQUENCE ID No. 112 (Figure 5c) was further characterized. In the dog, SEQUENCE ID No. 112 is still present at a detectable concentration for one week. Example 13. Effect of PEGylated Peptides on Intraperitoneal Glucose Tolerance in Rats The in vivo activity of the PEGylated pepids of this invention, administered subcutaneously, was examined in rats. Raias who fasted during the night received a subcutaneous injection of conirol or PEGylated p / pid (1-100 μg / kg). Three hours more farde, the basal blood glucose level was measured, and the rats received 2 g / kg of glucose intraperitoneally. The blood glucose level was measured again after 15, 30 and 60 minutes. A PEGylated peptide of this invention significantly reduced blood glucose levels relative to the vehicle after the intraperitoneal glucose tolerance test (Intraperitoneal Glucose Tolerance Tesi, IPGTT), with a reduction of 17% -28% in the area under the curve (area under fhe curve, AUC) of glucose (Figures 6a-6d). It shows that the PEGylated peel has a prolonged acivity of glucose decrease in vivo. In addition to the glucose depletion activity of the PEGylated peptides of the present invention, it also indicates that the peptide has a prolonged in vivo half-life. PACAP-27 has a very short half-life in vivo (<10 minutes). The ability of the PEGylated peptides of the invention to lower the blood glucose level after 24. (Figure 6b), 41 (Figure 6c), 65 (Figure 6c) and 72 (Figure 6d) hours after administration of the Peptide is a clear indication that the peptide is present in the circulation at this point in time and consequently, has a prolonged half-life in relation to PACAP-27 (Figure 6c). Likewise, the peptides of SEQUENCE ID No. 112 demonstrate greater potency and efficiency at longer time points, compared to the SEQ ID No. 154 and 155 peptides (Figures 6b and 6d). The demonstration of the activity of the polypeptides of the present invention can be achieved through in vitro, ex vivo and in vivo assays that are well known in the art. For example, the following assays can be used to demonstrate the efficacy of a pharmaceutical agent for the diabetic travail and related disorders, such as syndrome X, glucose intolerance, impaired fasting glucose and hyperinsulinemia; atherosclerotic disease and related disorders, such as hypertriglyceridemia and hypercholesterolemia; and obesity. Method for measuring blood glucose levels Blood is drawn (from an eye vein or tail) to db / db beads (obtained from Jackson Laboratories, Bar Harbor, ME), and the mice are grouped according to the levels of blood glucose equivalent means. An oral dose (by gavage in a pharmaceutically acceptable carrier) of the test polypeptide is administered once a day for 14 days. At that time, blood is drawn back from the animals, from a vein in the eye or tail, and blood glucose levels are determined. In each case, glucose levels are measured with an Elite XL glucometer (Bayer Corporation, Elkhart, IN).

Method for Measuring an Effect on Cardiovascular Parameters Cardiovascular parameters are also evaluated (eg, heart rate and blood pressure) An oral dose of the vehicle or the test polypeptide is administered to SHR rats once per day for 2 weeks. Blood pressure and heart rate are determined using a method that involves placing a cuff in the tail, as described in Grinsell, et al., (Am. J. Hypertens., 13: 370-375, 2000). blood pressure and heart rate are monitored, as described in Shen, et al., (J. Pharmacol.Exp Therap 278: 1435-1443, 1996.) Method for measuring triglyceride levels Blood is drawn ( from a vein of the eye or tail) to hApoAl mice (obtained from Jackson Laboratories, Bar Harbor, ME), and the mice are grouped according to the serum equivalent triglyceride levels, are given an oral dose (by gavage). in a vehicle pharmaceutically acceptable) of the test polypeptide once per day for 8 days. Blood is drawn back from the animals, from the vein of the eye or tail, and serum triglyceride levels are determined. In each case, triglyceride levels are measured using a Technicon Axon Analyzer (Bayer Corporation, Tarrytown, NY). Method for measuring HDL cholesterol levels To determine plasma levels of HDL cholesterol, blood is extracted from hApoAl mice and grouped according to plasma levels of HDL cholesterol equivalent. The mice are given an oral dose of the test vehicle or polypeptide once a day for 7 days, and then blood is drawn again on day 8. The plasma is analyzed for HDL cholesterol using the Synchron Clinical System ( CX4) (Beckman Coulter, Fullerton, CA). Method to measure the levels of total cholesterol, HDL cholesterol, triglycerides and glucose In another in vivo test, blood is extracted from obese monkeys, then an oral dose of the test vehicle or polypeptide is administered once a day for 4 weeks, and then blood is drawn again. The serum is analyzed to determine the levels of cholesterol, cholesterol, HDL, triglycerides and glucose, using the Synchronic Clinical System (CX4) (Beckman Coulter, Fullerton, CA). The subclass analysis of lipoprofeins is performed by nuclear magnetic resonance (NMR) spectroscopy as described in Oliver, et al., (Proc. Nati. Acad. Sci. USA 98: 5306-5311 , 2001). All publications and patents mentioned in the above specification are included herein by reference. Various modifications and variations of the compositions and methods of the invention described will be apparent to those skilled in the art, without departing from the scope and spirit of the invention. While the invention has been described in connection with specific preferred applications, it should be understood that the invention as claimed does not have to be unduly limited to said specific applications. Indeed, it is intended that various modifications of the modes described above to carry out the invention, which are obvious to those skilled in the field of biochemistry and other related fields, are included within the scope of the following claims. Those skilled in the art will recognize or be able to determine, by performing only routine experiments, many equivalents to the particular applications of the invention described in the present.

Claims (53)

1. We claim: 1. A polypeptide selected from the group comprising SEQUENCE ID Nos. 1 to 148, and the fragments, derivatives and variants of this functional equivalent.
2. 2. The polypeptide of claim 1, wherein said polypeptide is selected from the group comprising SEQUENCE ID Nos. 1, 2, 3, 4, 5, 112, 113, 114, 115 and 116.
3. 3. An antibody that specifically binds to the polypeptide of claim 1.
4. 4. The antibody of claim 3, wherein said antibody is a polyclonal antibody.
5. 5. The antibody of claim 3, wherein said antibody is a monoclonal antibody.
6. 6. An antibody that binds specifically to polyethylene glycol.
7. 7. The antibody of claim 6, wherein said antibody is a polyclonal antibody.
8. 8. The antibody of claim 6, wherein said antibody is a monoclonal antibody.
9. 9. A method for detecting a polypeptide selected from the group comprising SEQUENCE ID Nos. 1 to 148 in a sample that involves: to. putting the sample in contact with an antibody of claim 3 or claim 6, b. detect said antibody, and or to establish a correlation between the detection of the antibody and the amount of polypeptide present in the sample.
10. 10. A method for detecting a polypeptide selected from the group comprising SEQUENCE ID Nos. 1 to 148 in a sample that involves: to. putting the sample in contact with a first antibody of claim 3 or claim 6, b. putting the sample in contact with a labeled second antibody, where the second antibody binds to the first antibody, or detects the marker, and d. to draw a correlation between the deification of the marker and the amount of polypeptide present in the sample.
11. 11. A kit for detecting a polypeptide selected from the group comprising SEQUENCE IDs No. 1 to 148 in a sample involving: a first antibody of claim 3 or claim 6, and a second antibody, where the second antibody binds to the first anfibody.
12. 12. A pharmaceutical composition comprising a therapeutically effective amount of a polypeptide of claim 1 or fragments, derivatives and variants thereof functionally equivalent, in combination with a pharmaceutically acceptable vehicle.
13. 13. The pharmaceutical composition of claim 12, wherein said polypeptide is selected from the group comprising SEQUENCE ID Nos. 1, 2, 3, 4, 5, 112, 113, 114, 115 and 116.
14. 14. A pharmaceutical composition comprising a therapeutically effective amount of a polypeptide of claim 1, or fragments, derivatives and variants thereof functionally equivalent, in combination with a pharmaceutically acceptable carrier, and one or more pharmaceutical agents.
15. 15. The pharmaceutical composition of claim 14, wherein said pharmaceutical agent is selected from the group comprising the ligands of PPAR, insulin secretagogues, sulfonylureas, a-glucosidase inhibitors, insulin sensitizers, compounds that decrease the production of hepatic glucose, insulin and insulin derivatives, biguanides, inhibitors of protein tyrosine phosphatase-1B, dipeptidyl pepidase IV and 11 befa-HSD, anti-obesity drugs, HMG inhibitors- CoA reductase, nicotinic acid, drugs that decrease lipids, ACAT inhibitors, bile acid sequestrants, bile acid reuptake inhibitors, inhibitors of microsomal triglyceride transport, fibric acid derivatives, β-blockers, ACE inhibitors, blockers of calcium channels, diuretics, renin inhibitors na, AT-1 receptor antagonisms, ET anolygonias, ET inhibitors, neutral endopeptidase inhibitors, vasopeptidase inhibitors, and nifaranes.
16. 16. A composition comprising an effective amount of a polypeptide of claim 1, or fragments, derivatives and variants thereof functionally equivalent, in combination with an inert carrier.
17. 17. A method to eradicate diabetes, comprising the step of administering to a subject in need thereof a positive epileptic mass of a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
18. 18. The method of claim 17, wherein said diabetes is selected from the group comprising type 2 diabetes, juvenile hereditary diabetes type 2, adult latent autoimmune diabetes and gestational diabetes.
19. 19. A method for treating syndrome X, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
20. 20. A method for bringing about disorders related to diabetes, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
21. 21. The method of claim 20, wherein said disorder related to the diabei is selected from the group comprising hyperglycemia, hyperinsulinemia, glucose intolerance, fasting glucose allyration, dyslipidemia, hypertriglyceridemia and resistance to glucose. insulin.
22. 22. A method for bringing the diabei, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a polypeptide of claim 1, in combination with one or more pharmaceutical agents.
23. 23. The method of claim 20, wherein said pharmaceutical agent is selected from the group comprising the PPAR agonists, the sulfonylureas, the non-sulfonylureas secretagogues, the a-glucosidase inhibitors, the insulin sensitizers, the insulin secretagogues, the compounds that decrease the production of hepatic glucose, insulin and agents confers obesity.
24. 24. The method of claim 23, wherein said diabetes is selected from the group comprising type 2 diabetes, early maturity diabetes in the young, adult latent autoimmune diabetes and gestational diabetes.
25. 25. A method for bringing syndrome X, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a polypeptide of claim 1, in combination with one or more pharmaceutical agents.
26. 26. The method of claim 25, wherein said pharmaceutic agent is selected from the group comprising PPAR agonisias, sulfonylureas, non-sulfonylureas secretagogues, α-glucosidase inhibitors, insulin sensitizers, insulin secretagogues, compounds that decrease the production of hepatic glucose, insulin and agents confers obesity.
27. 27. A method for treating disorders related to diabetes, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a polypeptide of claim 1, in combination with one or more pharmaceutical agents.
28. 28. The method of claim 27, wherein said transpore related to the diabei is selected from the group comprising hyperglycemia, hyperinsulinemia, glucose intolerance, impaired fasting glucose, dyslipidemia, hypertriglyceridemia and resistance to glucose. insulin.
29. 29. The method of claim 28, wherein said pharmaceutical agent is selected from the group comprising the PPAR agonists, the sulfonylureas, the non-sulfonylureas secretagogues, the a-glucosidase inhibitors, the insulin sensitizers, the insulin secretagogues, the compounds that decrease the production of hepatic glucose, insulin and anti-obesity agents.
30. 30. A method for bringing diabeids, Syndrome X or the diabes related to the diabei, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a polypeptide of claim 1, in combination with one or more selected agents of the group comprising the HMG-CoA inhibitors reducíaa, nicoïínico acid, lipid-lowering drugs, ACAT inhibitors, bile acid sequestrants, inhibitors of bile acid reuptake, inhibitors of microsomal иiglicérides ransfer, fibric acid derivatives, ß -blockers, ACE inhibitors, calcium channel blockers, diuretics, renin inhibitors, antagonists of AT-1 receptor, ET receptor aniagonists, neutral endopeptidase inhibitors , inhibitors of vasopeptidase and nitrates.
31. 31. The method of claim 30, wherein said diabetes-related disorder is selected from the group comprising hyperglycemia, hyperinsulinemia, glucose intolerance, impaired fasting glucose, dyslipidemia, hypertriglyceridemia and resistance to glucose. insulin.
32. 32. The method of any of claims 22 to 31, wherein the polypeptide of claim 1 and one or more pharmaceutical agents are administered as a single galenic form.
33. 33. A method for bringing the diabeiids or preventing the secondary causes of said disease, comprising the step of administering to a subject in need thereof an epistemically effective amount of a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
34. 34. The method of claim 33, wherein said secondary cause is selected from the group comprising excess glucocorticoids, excess growth hormone, pheochromocytoma and pharmacogenic diabetes.
35. 35. A method for treating diabetes or preventing secondary causes of said disease, comprising the step of administering to a subject, in need thereof a therapeutically effective amount of a polypeptide of claim 1, in combination with one or more pharmaceutical agents.
36. 36. The method of claim 35, wherein said pharmaceutic agent is selected from the group comprising the PPAR agonists, the sulfonylureas, the non-sulfonylureas secreyagogues, the a-glucosidase inhibitors, the insulin sensitizers, the insulin secreyagogues, the compounds that decrease the production of hepatic glucose, insulin and anti-obesity agents.
37. 37. A method for bringing respiratory disease, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
38. 38. A method for eradicating obesity, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
39. 39. A method for bringing cardiovascular disease, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
40. 40. The method of claim 39, wherein said cardiovascular disease is selected from the group comprising aerosol, coronary heart disease, coronary artery disease and hypertension.
41. 41 A method for treating disorders of lipid and carbohydrate metabolism, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
42. 42. A method to bring sleep disorders, which includes the passage of administering to a subject in need thereof an epidurally effective effect of a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
43. 43. A method for treating male reproductive disorders, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
44. 44. A method to eradicate the growth phenomena or energy homeosis, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
45. 45. A method for bringing about immunological diseases, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
46. 46. A method for treating autoimmune diseases, comprising the step of administering to a subject in need thereof a therapeutically effective effect of a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
47. 47. A method for bringing acute and chronic inflammatory diseases, comprising the step of administering to a subject in need thereof an epilephically effective amount of a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
48. 48. A method to eradicate the sepficémic shock, which comprises the step of administering to a subject in need of an effective lepraeuically effective amount of a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
49. 49. A method for stimulating the release of insulin in a glucose dependent manner in a subject in need thereof, by administering to said subject a polypeptide of claim 1 or a pharmaceutical composition of claim 12.
50. 50. The polypeptides according to claim 1 for the treatment and / or prophylaxis of diabetes and disorders related to diabetes.
51. 51. A medicament containing at least one polypeptide according to claim 1, in combination with at least one pharmaceutically acceptable and pharmaceutically safe carrier or excipient.
52. 52. The use of the polypeptides according to claim 1 for manufacturing a medicament for the treatment and / or prophylaxis of diabetes and diabetes-related disorders.
53. 53. A medicament according to claim 51 for the treatment and / or prophylaxis of the diabei. SUMMARY OF THE INVENTION This invention provides novel peptides that function in vivo as VPAC2 receptor agonists. It has been shown that these insulin secretagogue polypeptides lower the blood glucose level in vivo by performing a glucose tolerance test. The polypeptides of this invention are also stable in the formulation and have long half-lives. The peptides of the present invention provide a therapy for patients with a reduction in endogenous insulin secretion, for example, diabei-type 2. The invention is also directed to a method for treating a metabolic disease in a mammal comprising administering to said mammal. mammal a therapeutically effective amount of the peptides. 1/17 FIG. 1a