N-terminus Conformationally Constrained Glp-1 Receptor Agonist Compounds

Abstract

The disclosure provides N-terminus conformationally constrained compounds, which may comprise peptide mimetics and/or amino acid substitutions, which may be used in peptides, such as GLP-1 receptor agonist compounds, to induce .beta.-turn secondary structure at the N-terminus. The N-terminus conformationally constrained compounds may be used for research purposes; to produce GLP-1 receptor agonist compounds having improved GLP-1 receptor binding activity, enzymatic stability, or in vivo glucose lowering activity; and to develop GLP-1 receptor agonist compounds which have fewer amino acid residues. The disclosure also provides GLP-1 receptor agonist compounds, such as exendins, exendin analogs, GLP-1(7-37), GLP-1(7-37) analogs, comprising the N-terminus conformationally constrained compounds. The compounds are useful for treating various diseases, such as diabetes and obesity. The disclosure also provides methods for chemically synthesizing the N-terminus conformationally constrained compounds.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application is a divisional of U.S. application Ser. No. 13/260,702, filed on Sep. 27, 2011, which is a U.S. national stage of International Application No. PCT/US2010/28883, filed Mar. 26, 2010, which claims priority to U.S. Application No. 61/165,604 filed Apr. 1, 2009.

FIELD

[0002] Provided herein are N-terminus conformationally constrained GLP-1 receptor agonist compounds and therapeutic methods for their use.

BACKGROUND

[0003] Peptides and proteins play critical roles in the regulation of biological processes. Peptides, for example, play a regulatory role as hormones and inhibitors, and are also involved in immunological recognition. The significant biological role of peptides makes it important to understand their interactions with the receptors to which they bind.

[0004] The determination of the receptor-bound conformation of a peptide is invaluable for the rational design of peptide analogs. Marshall et al, Ann. Rep. Med. Chem., 13:227-238 (1978) disclose that peptides are characteristically highly flexible molecules, the structures of which are strongly influenced by the environment in which they reside. Thus, peptides are not generally useful for determining their receptor-bound conformation.

[0005] As no approach is available to predict which new ligand-receptor interactions will lead to antagonists and which will lead to agonists of greater or less potency, it is necessary to perform classical structure-function studies in a systematic way to provide information about the specific amino acid residues and functional groups in a peptide that are important to biological activity. Studies of this nature can utilize conformationally constrained peptide mimetics. For example, Hruby, Trends Pharmacol. Sci., 8:336-339 (1987) suggests that conformational constraints can provide information about the different requirements that a receptor has for a ligand to be an agonist or antagonist.

[0006] Generally, peptide mimetics can be defined as structures which serve as appropriate substitutes for peptides in interactions with receptors and enzymes. The development of rational approaches for discovering peptide mimetics is a major goal of medicinal chemistry. Such development has been attempted both by empirical screening approaches and by specific synthetic design. Specific design of peptide mimetics has utilized both peptide backbone modifications and chemical mimics of peptide secondary structures. The beta-turn has been implicated as an important site for molecular recognition in many biologically active peptides. Consequently, peptides containing conformationally constrained mimetics of beta-turns are particularly desirable.

[0007] There is a need in the art for new GLP-1 receptor agonist compounds that have good stability, resistance to degradation, and good glucagon-like peptide-1 (GLP-1) receptor binding activity and in vivo glucose lowering activity. To solve these needs, the disclosure herein provides, among other things, novel N-terminus conformationally constrained compounds, novel N-terminus conformationally constrained GLP-1 receptor agonist compounds containing modifications, such as peptide mimetics and/or amino acid substitutions, that provide a conformationally constrained N-terminus that results in improved GLP-1 receptor binding and in vivo blood glucose lowering activity.

SUMMARY

[0008] It was previously believed that the N-terminus of exendin-4 and exendin analogs was a random coil. It has now been unexpectedly discovered that the N-terminus shows a high beta-turn characteristic in a specific site, and therefore mimics the receptor bound conformation of this region of the peptides. The disclosure herein is based on this discovery.

[0009] Provided herein are N-terminus conformationally constrained compounds having the formula: Xaa.sub.1Xaa.sub.2Xaa.sub.3-Z and Xaa.sub.1Xaa.sub.2Xaa.sub.3Xaa.sub.4-Z, where the substituents are defined herein. These N-terminus conformationally constrained compounds may induce a .beta.-turn conformational constraint at the N-terminus when they are used in GLP-1 receptor agonist compounds. The N-terminus conformationally constrained compounds may be used for therapeutic purposes (e.g., treat diabetes); for research purposes; and to produce GLP-1 receptor agonist compounds having improved GLP-1 receptor binding activity, enzymatic stability, and improved in vivo glucose lowering activity. The disclosure provides pharmaceutical compositions comprising therapeutically effective amounts of the N-terminus conformationally constrained compounds. The disclosure also provides methods for synthesizing the N-terminus conformationally constrained compounds.

[0010] Provided herein are GLP-1 receptor agonist compounds, such as exendins, exendin analogs, GLP-1(7-37) (SEQ ID NO: 84), and GLP-1(7-37) analogs, comprising an N-terminus conformationally constrained compound having the formula Xaa.sub.1Xaa.sub.2Xaa.sub.3- or Xaa.sub.1Xaa.sub.2Xaa.sub.3Xaa.sub.4-, where the substituents are defined herein. In one embodiment, the GLP-1 receptor agonist compounds comprise Xaa.sub.1Xaa.sub.2Xaa.sub.3Xaa.sub.4, where the substituents are defined herein, at positions 1-4 at the N-terminus. In one embodiment, the GLP-1 receptor agonist compounds comprise Xaa.sub.1Xaa.sub.2Xaa.sub.3, where the substituents are defined herein, at positions 1-3 at the N-terminus. The disclosure provides pharmaceutical compositions comprising therapeutically effective amounts of these N-terminus conformationally-constrained GLP-1 receptor agonist compounds.

[0011] Provided herein are exendins and exendin analogs having the formula:

TABLE-US-00001 (SEQ ID NO: 1) Xaa.sub.1Xaa.sub.2Xaa.sub.3Xaa.sub.4TFTSDLSKQXaa.sub.14EEEAVRLFIEXaa.sub.2- 5LKN-Z; (SEQ ID NO: 2) Xaa.sub.1Xaa.sub.2Xaa.sub.3Xaa.sub.4TFTSDLSKQXaa.sub.14EEEAVRLFIEXaa.sub.2- 5LK-R.sub.10-Z; (SEQ ID NO: 3) Xaa.sub.1Xaa.sub.2Xaa.sub.3Xaa.sub.4TFTSDLSKQXaa.sub.14EEEAVRLFIEXaa.sub.2- 5 LKNGGPSSGAPPPS-Z;

where Xaa.sub.14, Xaa.sub.25, R.sub.10, and Z are defined herein; and at least one of Xaa.sub.1, Xaa.sub.2, Xaa.sub.3, and Xaa.sub.4 are modifications, such as peptide mimetics and/or amino acid substitutions, that induce a conformational constraint at the N-terminus. The disclosure provides pharmaceutical compositions comprising therapeutically effective amounts of these exendin analogs.

[0012] Provided herein are GLP-1(7-37) (SEQ ID NO: 84) and GLP-1(7-37) analogs having the formula:

TABLE-US-00002 (SEQ ID NO: 4) Xaa.sub.1Xaa.sub.2Xaa.sub.3Xaa.sub.4TFTSDVSSYXaa.sub.14EGQAAKEFIAXaa.sub.- 25 LVXaa.sub.28GRXaa.sub.31-Z;

where Xaa.sub.14, Xaa.sub.25, Xaa.sub.28, Xaa.sub.31, and Z are as defined herein; and at least one of Xaa.sub.1, Xaa.sub.2, Xaa.sub.3, and Xaa.sub.4 are modifications, such as peptide mimetics and/or amino acid substitutions, that induce a conformational constraint at the N-terminus. The disclosure provides pharmaceutical compositions comprising therapeutically effective amounts of these GLP-1(7-37) analogs.

[0013] Provided herein are GLP-1 receptor agonist compounds, such as exendins, exendin analogs, GLP-1(7-37) (SEQ ID NO: 84), and GLP-1(7-37) analogs wherein position 1 comprises an imidazole ring (e.g., His) and position 3 is proline; where the GLP-1 receptor agonist compounds bind in a RIN cell membrane receptor binding assay with an affinity of less than 1 nM (or less than 0.1 nM).

[0014] The disclosure provides methods for treating diabetes; treating insulin resistance; treating postprandial hyperglycemia; lowering blood glucose levels; lowering HbA1c levels; stimulating insulin release; reducing gastric motility; delaying gastric emptying; reducing food intake; reducing appetite; reducing weight; treating overweight; and treating obesity in patients in need by administering therapeutically effective amounts of the N-terminus conformationally constrained compounds and/or the N-terminus conformationally constrained GLP-1 receptor agonist compounds described herein.

BRIEF DESCRIPTION OF THE FIGURES

[0015] FIG. 1: FIG. 1A is exendin-4 amide (SEQ ID NO: 86); FIG. 1B is dAla.sup.2,Pro.sup.3-exendin-4 amide, and FIG. 1C is dAla.sup.2,Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4 (1-28) amide. FIG. 1D is a graph showing the change in blood glucose in mice administered the compounds shown in FIGS. 1A-C based on the in vivo blood glucose assay described in Example 17. The compounds were injected IP at t=0 immediately following a baseline sample in 2-hour fasted NIH/Swiss mice. Blood glucose samples were taken at t=30, 60, 120, 180, and 240 minutes with a ONETOUCH.RTM. ULTRA.RTM. (LifeScan, Inc., Milpitas, Calif.).

[0016] FIG. 2: FIG. 2A is an exendin analog (SEQ ID NO: 87), described, e.g., in WO 2007/139941. FIGS. 2B-J show the exendin analog of FIG. 2A having a modification at Glu.sup.3. R.sub.1 is GTFTSDLSKQLEEEAVRLFIEWLKQGGPSKEIIS-NH.sub.2 (SEQ ID NO: 5). FIG. 2K shows the exendin analog (SEQ ID NO: 5) of FIG. 2A having modifications at Gly.sup.2Glu.sup.3. FIGS. 2L-2U show N-terminus conformationally constrained compounds. FIGS. 2B-J provide examples of exendin analogs containing the N-terminus conformationally constrained compounds shown in FIGS. 2L-2U.

[0017] FIGS. 3A-G are dAla.sup.2,Pro.sup.3-exendin-4 (FIG. 3A), Ala.sup.2,Pro.sup.3-exendin-4 (SEQ ID NO: 88) (FIG. 3B), Pro.sup.3,Ala.sup.4-exendin-4 (SEQ ID NO: 89) (FIG. 3C), Ala.sup.2,Pro.sup.3,Ala.sup.4-exendin-4 (SEQ ID NO: 90) (FIG. 3D), Pro.sup.3,dAla.sup.4-exendin-4 (FIG. 3E), Val.sup.2,Pro.sup.3-exendin-4 (SEQ ID NO: 91) (FIG. 3F), and NMeAla.sup.2,Pro.sup.3-exendin-4 (SEQ ID NO: 92) (FIG. 3G). The compound in FIGS. 1B and 3A are the same.

[0018] FIG. 4 is a graph showing the change in blood glucose in mice administered the compounds shown in FIGS. 3G, 2I, and 2B based on the in vivo blood glucose assay described in Example 17. The compounds were injected IP at t=0 immediately following a baseline sample in 2-hour fasted NIH/Swiss mice. Blood glucose samples were taken at t=30, 60, 120, 180, and 240 minutes with a ONETOUCH.RTM. ULTRA.RTM. (LifeScan, Inc., Milpitas, Calif.).

[0019] FIG. 5: FIG. 5A is Leu.sup.14,Phe.sup.25-exendin-4(1-28) (SEQ ID NO: 93), described in WO 2007/139941. Leu.sup.14,Phe.sup.25-exendin-4(1-28) (SEQ ID NO: 93) refers to amino acid residues 1-28 in exendin-4 (i.e., exendin-4(1-28) (SEQ ID NO: 99), where the amino acid residue at position 14 in exendin-4 is replaced with Leu (i.e., Leu.sup.14), and the amino acid residue at position 25 is replaced with Phe (i.e., Phe.sup.25). FIGS. 5B-G show the exendin analog of FIG. 5A having a modification at Glu.sup.3 or Gly.sup.2Glu.sup.3 at the N-terminus. R.sub.4 is GTFTSDLSKQLEEEAVRLFIEFLKN-NH.sub.2 (SEQ ID NO: 6).

[0020] FIGS. 6A-E show the exendin analog of FIG. 5A having a modification at Gly.sup.2Glu.sup.3 or Gly.sup.2Glu.sup.3Gly.sup.4 at the N-terminus. The C-terminal amino acid in each compound shown in FIGS. 6A-E is amidated. FIGS. 6A through 6D disclose SEQ ID NOS 94 and 94-96, respectively.

[0021] FIG. 7 is a graph showing the change in blood glucose in mice administered the compounds shown in FIGS. 6A, 8B, 6E, 1C, 5B, and 5A based on the in vivo blood glucose assay described in Example 17. The compounds were injected IP at t=0 immediately following a baseline sample in 2-hour fasted NIH/Swiss mice. Blood glucose samples were taken at t=30, 60, 120, 180, and 240 minutes with a ONETOUCH.RTM. ULTRA.RTM. (LifeScan, Inc., Milpitas, Calif.).

[0022] FIG. 8: FIGS. 8A-D show the exendin analog in FIG. 5A having modifications at Gly.sup.2Glu.sup.3Gly.sup.4 at the N-terminus, where the C-terminal amino acid is amidated. FIGS. 8E-H show the exendin analog in FIG. 2A having modifications at Gly.sup.2Glu.sup.3Gly.sup.4 at the N-terminus. FIGS. 8A and 8E disclose SEQ ID NOS 97-98, respectively.

[0023] FIG. 9: FIGS. 9A-H show the exendin analog in FIG. 5A having a modification at Gly.sup.2Glu.sup.3 or Gly.sup.2Glu.sup.3Gly.sup.4 at the N-terminus. Each compound in FIGS. 9A-H is amidated at the C-terminal amino acid. FIGS. 9A through 9H disclose SEQ ID NOS 6, 6, 6, 6, 100, 6, 6 and 6, respectively. FIG. 9I is a graph showing the change in blood glucose in mice administered the compounds shown in FIGS. 9B, 9C, 9D, 9F, 5A, and 1C based on the in vivo blood glucose assay described in Example 17. The compounds were injected IP at t=0 immediately following a baseline sample in 2-hour fasted NIH/Swiss mice. Blood glucose samples were taken at t=30, 60, 120, 180, and 240 minutes with a ONETOUCH.RTM. ULTRA.RTM. (LifeScan, Inc., Milpitas, Calif.).

[0024] FIGS. 10A-I show the exendin analog in FIG. 5A having a modification at His.sup.1 at the N-terminus. R.sub.4 is Xaa.sub.2Xaa.sub.3GTFTSDLSKQLEEEAVRLFIEFLKN-NH.sub.2 (SEQ ID NO: 7), where Xaa.sub.2 is Gly, dAla, or Aib; and Xaa.sub.3 is Glu or Pro. Alternatively, R.sub.4 is Xaa.sub.2Xaa.sub.3GTFTSDLSKQLEEEA VRLFIEWLKNGGPSSGAPPPS-NH.sub.2 (SEQ ID NO: 8), where Xaa.sub.2 is Gly, dAla, or Aib; and Xaa.sub.3 is Glu or Pro; provided that Xaa.sub.3 is not Glu when Xaa.sub.2 is Gly, or Xaa.sub.2 is not Gly when Xaa.sub.3 is Glu. Alternatively, R.sub.4 is Xaa.sub.2Xaa.sub.3GTFTSDLSKQLEEEAVRLFIEWLKQGGPSKEIIS-OH (SEQ ID NO: 9), where Xaa.sub.2 is Gly, dAla, or Aib; and Xaa.sub.3 is Glu or Pro.

[0025] FIGS. 11A-B show exendin-4 of FIG. 1A having a modification at His.sup.1 at the N-terminus. R.sub.2 is GEGTFTSDLSKQLEEEAVRLFIEWLKNGGPSSGAPPPS-NH.sub.2 (SEQ ID NO: 10).

[0026] FIG. 12 shows the compound of FIG. 10E with an amino acid substitution of Trp.sup.25. The compound in FIG. 12 is amidated at the C-terminal amino acid residue (SEQ ID NO: 101).

[0027] FIG. 13 shows an exendin analog having a modification at His.sup.1Gly.sup.2Glu.sup.3. The compound in FIG. 13 is amidated at the C-terminal amino acid residue (SEQ ID NO: 102).

[0028] FIG. 14: FIG. 14A is a generic structure (where Xaa.sub.2 is, e.g., Gly, dAla, or Aib; and the other substituents are defined herein) of an exendin analog of FIG. 1A, 2A, or 5A having modifications at His.sup.1Gly.sup.2Glu.sup.3 at the N-terminus, where R is a peptide, such as any one of the following: GTFTSDLSKQLEEEAVRLFIEFLKN-NH.sub.2 (SEQ ID NO: 6); GTFTSDLSKQLEEEAV-RLFIEWLKQGGPSKEIIS-OH (SEQ ID NO: 5); or GTFTSDLSKQLEEEAVRLFIEWLKNGGPSSGAPPPS-NH.sub.2 (SEQ ID NO: 11). Aib is .alpha.-methylalanine FIGS. 14B-C are examples of compounds from the structure in FIG. 14A that have modifications at His.sup.1 and Glu.sup.3. The compounds in FIGS. 14B-C are amidated at the C-terminal amino acid residue. FIGS. 14B through 14C disclose SEQ ID NOS 6 and 6, respectively. FIG. 14D provides the generic structure (where Xaa.sub.2 is Gly, dAla, or Aib; and the other substituents are defined herein) of an N-terminus conformationally constrained compound. FIGS. 14E-R are exendin analogs comprising an N-terminus conformationally constrained compound. FIG. 14Q discloses SEQ ID NO: 38. R.sub.1 is GTFTSDLSKQLEEEAVRLF IEWLKQGGPSKEIIS-OH (SEQ ID NO: 5). R.sub.4 is GTFTSDLSKQLEEEAVRLFIEFLKN-NH.sub.2 (SEQ ID NO: 6). R.sub.5 is PGTFTSDLSKQLEEEAVRLFIEWLKNGGPSSGAPPPS-NH.sub.2 (SEQ ID NO: 12).

[0029] FIGS. 15A-D are exendin analogs that comprise isomers of nipecotic acid as the N-terminus conformationally constrained compound. R.sub.1 is FTSDLSKQLEEEAVRLFI EWLKQGGPSKEIIS-NH.sub.2 (SEQ ID NO: 13).

[0030] FIG. 16: FIGS. 16A-H are exendin analogs containing a modification at the N-terminus. R.sub.1 is FTSDLSKQLEEEAVRLFIEWLKQGGPSKEIIS-OH (SEQ ID NO: 13). R.sub.2 is FTSDLSKQLE EEAVRLFIEWLKNGGPSSGAPPPS-NH.sub.2 (SEQ ID NO: 14). R.sub.3 is FTSDVSSYLEGQAAKEFIAWLVKGRG-NH.sub.2 (SEQ ID NO: 15). FIGS. 16I-L are exendin analogs containing a modification at the N-terminus. The modification is designed to mimic amino acid residues His.sup.1Gly.sup.2Glu.sup.3. R.sub.4 is GTFTSDLSKQLEE EAVRLFIEFLKN-NH.sub.2 (SEQ ID NO: 6).

[0031] FIG. 17: FIGS. 17A-F are exendin analogs containing a thiazolidine-proline peptide mimetics at Gly.sup.2Glu.sup.3. R.sub.4 is GTFTSDLSKQLEEEAVRLFIEFLKN-NH.sub.2 (SEQ ID NO: 6). FIGS. 17G-N are exendin analogs having a modification at His.sup.1 and containing a thiazolidine peptide mimetic at Gly.sup.2Glu.sup.3. R.sub.1, R.sub.2, and R.sub.3 are each independently hydrogen, methyl, or ethyl. In this embodiment, Xaa.sub.3 is Glu, Asp, Pro, or Gly. R.sub.4 is GTFTSDLSKQLEEEAVRLFIEFLKN-NH.sub.2 (SEQ ID NO: 6).

[0032] FIG. 18: FIG. 18A is a process for preparing (5,5)-Glu-Gly-OH, a dipeptide mimetic that can be used to induce a .beta.-turn conformational constraint, for example, at the N-terminus in a GLP-1 receptor agonist compound. FIG. 18B is a generic structure of the compound that can be produced by the process shown in FIG. 18A. The skilled artisan can choose compounds with different stereochemistries during the reaction process to provide for various stereochemistries in the final product. * represents a chiral carbon atom.

[0033] FIG. 19: FIG. 19A is a process for preparing .gamma.-lactam Glu-Gly-OH, a dipeptide mimetic that can be used to induce a .beta.-turn conformational constraint, for example, at the N-terminus in a GLP-1 receptor agonist compound. FIG. 19B is a generic structure of the compound that can be produced by the process shown in FIG. 19A. The skilled artisan can choose compounds with different stereochemistries during the reaction process to provide for various stereochemistries in the final product. * represents a chiral carbon atom.

[0034] FIG. 20: FIG. 20A is a process for preparing (6,5)-Asp-Gly-OH, a dipeptide mimetic that can be used to induce a .beta.-turn conformational constraint, for example, at the N-terminus in a GLP-1 receptor agonist compound. FIG. 20B is a generic structure of the compound that can be produced by the process shown in FIG. 20A. The skilled artisan can choose compounds with different stereochemistries during the reaction process to provide for various stereochemistries in the final product. * represents a chiral carbon atom.

[0035] FIG. 21: FIG. 21A is a process for preparing .delta.-lactam Asp-Gly-OH and Asp-Ala-OH, both of which are dipeptide mimetics that can be used to induce a .beta.-turn conformational constraint, for example, at the N-terminus in a GLP-1 receptor agonist compound. FIG. 21B is a generic structure of the compound that can be produced by the process shown in FIG. 21A. The skilled artisan can choose compounds with different stereochemistries during the reaction process to provide for various stereochemistries in the final product. * represents a chiral carbon atom.

[0036] FIG. 22 is a process for preparing a peptide mimetic which can be used to induce a .beta.-turn conformational constraint, for example, at the N-terminus in a GLP-1 receptor agonist compound. FIG. 22 discloses SEQ ID NOS 103-104, respectively, in order of appearance.

DETAILED DESCRIPTION

[0037] "GLP-1 receptor agonist compounds" refer to compounds that elicit a biological activity of an exendin reference peptide (e.g., exendin-4) or a GLP-1(7-37) (SEQ ID NO: 84) reference peptide when evaluated by art-known measures such as receptor binding studies or in vivo blood glucose assays as described, e.g., Examples 16 and 17, and by Hargrove et al, Regulatory Peptides, 141:113-119 (2007), the disclosure of which is incorporated by reference herein. GLP-1 receptor agonist compounds include, for example, native exendins, exendin analogs, native GLP-1, GLP-1 analogs, GLP-1(7-37) (SEQ ID NO: 84), and GLP-1(7-37) analogs.

[0038] The term "exendin" includes naturally occurring (or synthetic versions of naturally occurring) exendin peptides that are found in the salivary secretions of the Gila monster. Exendin-3 (HSDGTFTSDLSKQMEEEAVRLFIEWLKNGGPSSGAPPPS-NH.sub.2) (SEQ ID NO: 16) is present in the salivary secretions of Heloderma horridum and exendin-4 (FIG. 1A) is present in the salivary secretions of Heloderma suspectum. Exendins include the amidated forms, the acid form, the pharmaceutically acceptable salt form, and any other physiologically active form of the molecule. In one embodiment, the term exendin can be used interchangeably with the term "exendin agonist."

[0039] "Exendin analog" refers to peptides, peptides containing peptide mimetics, amino acid substitutions, and/or other modifications, peptides containing the N-terminus conformationally constrained compounds described herein, and/or other chemical moieties, or other compounds which elicit a biological activity similar to that of an exendin reference peptide (e.g., exendin-4), when evaluated by art-known measures such as receptor binding assays or in vivo blood glucose assays as described, e.g., Examples 16 and 17, and by Hargrove et al, Regulatory Peptides, 141:113-119 (2007), the disclosure of which is incorporated by reference herein. Preferably, the exendin analogs will bind in such receptor binding assays with an affinity of less than 1 .mu.M; an affinity of less than 5 nM; an affinity of less than 1 nM, or an affinity of less than 0.1 nM. In one embodiment, the term "exendin analog" refers to a peptide that has an amino acid sequence with 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions, insertions, deletions, or a combination of two or more thereof, when compared to the amino acid sequence of exendin-4 shown in FIG. 1A. In other embodiment, the term "exendin analog" refers to a peptide that has at least 85%, at least 88%, at least 90%, at least 93%, at least 95%, or at least 98% sequence identity to the amino acid sequence of exendin-4 shown in FIG. 1A. Exendin analogs include the amidated forms, the acid form, the pharmaceutically acceptable salt form, and any other physiologically active form of the molecule. In one embodiment, the term exendin analog can be used interchangeably with the term "exendin agonist analog."

[0040] "GLP-1(7-37) analogs" refers to peptides, peptides containing peptide mimetics and/or other modifications, peptides containing the N-terminus conformationally constrained compounds described herein, and/or other chemical moieties, or other compounds which elicit a biological activity similar to that of GLP-1(7-37) (SEQ ID NO: 84), when evaluated by art-known measures such as receptor binding assays or in vivo blood glucose assays as described, e.g., Examples 16 and 17, and by Hargrove et al, Regulatory Peptides, 141:113-119 (2007), the disclosure of which is incorporated by reference herein. In one embodiment, the term "GLP-1(7-37) analog" refers to a peptide that has an amino acid sequence with 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions, insertions, deletions, or a combination of two or more thereof, when compared to the amino acid sequence of GLP-1(7-37) (SEQ ID NO: 84). In one embodiment, the GLP-1(7-37) analog is GLP-1(7-36) (SEQ ID NO: 85). GLP-1(7-37) analogs include the amidated forms, the acid form, the pharmaceutically acceptable salt form, and any other physiologically active form of the molecule.

[0041] "N-Terminus conformationally constrained GLP-1 receptor agonist compounds" refers to compounds in which one, two, three, or four of the amino acid residues at positions 1-4 at the N-terminus of "GLP-1 receptor agonist compounds" (e.g., exendins, exendin analogs, GLP-1, GLP-1 analogs, GLP-1(7-37) (SEQ ID NO: 84), GLP-1(7-37) analogs) have been modified or substituted with amino acids (e.g., natural and/or non-natural amino acids), peptidomimetics, or beta-turn dipetidemimetics. This substitution(s) or modification(s) to the N-terminus of the parent GLP-1 receptor agonist compound changes the flexible random coil structure of this specific region into a more rigid secondary structure with beta-turn characteristics.

[0042] The glycine residues at positions 2 and 4 at the N-terminus of exendin-4 may indicate the presence of a .beta.-turn in this region. In order to produce a conformationally constrained N-terminus, exendin analogs having a mimetic or other structural modification that restricted the conformational flexibility of the His.sup.1 side chain were synthesized. Restricting the flexibility of the His.sup.1 side chain was hypothesized to provide structural information about the possible bioactive conformation of GLP-1 receptor agonist compounds and thus enhance GLP-1 receptor binding, in vivo blood glucose lowering activity, and enzymatic stability.

[0043] An Ala scan of exendin-4 showed that the Glu.sup.3 residue was important for biological activity. Additionally, Glu.sup.3 or Asp.sup.3 are present in many members of the super-family of glucagon-related peptides, which indicates the importance of an acidic side chain at that residue.

[0044] It was postulated that the negative charge of Glu.sup.3 or Asp.sup.3 interacted through an ionic bond with the positive charge of the His.sup.1 side chain to position the key imidazole ring of the His.sup.1 side chain in the right space for interaction with the GLP-1 receptor. It was thus proposed that a .beta.-turn would be formed by the sequence Gly.sup.2Glu.sup.3Gly.sup.4Thr.sup.5 (SEQ ID NO: 17) in the super-family of glucagon related peptides, such as exendin-4 and exendin analogs.

[0045] In order to maintain the negative charge of Glu.sup.3, thought to be essential for biological activity, 13-turn peptide mimetics were synthesized to mimic the amino acid residues Glu.sup.3Gly.sup.4. In one embodiment, the disclosure provides N-terminus conformationally constrained compounds of Formula (F):

Xaa.sub.1Xaa.sub.2Xaa.sub.3-Z;

wherein: Xaa.sub.1 is a compound of Formula (1), as described herein; Xaa.sub.2 is Gly, Ala, dAla, or Aib;

Xaa.sub.3 is:

[0046] ##STR00001## [0047] wherein * indicates a chiral carbon atom; and R.sub.2 is hydrogen or a C.sub.1-4 alkyl (e.g., methyl, ethyl); and

Z is:

[0047] [0048] (i) OH; [0049] (ii) NH.sub.2; [0050] (iii) TFTSDLSKQXaa.sub.14EEEAVRLFIEXaa.sub.25LKN-Z.sub.1 (SEQ ID NO: 18); [0051] (iv) TFTSDLSKQXaa.sub.14EEEAVRLFIEXaa.sub.25LKNGGPSSGAPPPS-Z.sub.1 (SEQ ID NO: 19); [0052] (v) TFTSDLSKQXaa.sub.14EEEAVRLFIEXaa.sub.25LK-R.sub.10-Z.sub.1 (SEQ ID NO: 20); or [0053] (vi) TFTSDVSSYXaa.sub.14EGQAAKEFIAXaa.sub.25LVXaa.sub.28GRXaa.sub.31-Z.sub.1 (SEQ ID NO: 21); [0054] wherein: [0055] Z.sub.1 is OH or NH.sub.2; [0056] Xaa.sub.14 is Leu or Met; [0057] Xaa.sub.25 is Phe or Trp; [0058] Xaa.sub.28 is Lys or Arg; [0059] Xaa.sub.31 is Gly or absent; and [0060] R.sub.10 is QGGPSKEIIS (SEQ ID NO: 22); QGGPSSGAPPPS (SEQ ID NO: 23); NG; NGG; NGGP (SEQ ID NO: 24); NGGPS (SEQ ID NO: 25); NGGPSS (SEQ ID NO: 26); NGGPSSG (SEQ ID NO: 27); NGGPSSGA (SEQ ID NO: 28); NGGPSSGAP (SEQ ID NO: 29); NGGPSSGAPP (SEQ ID NO: 30); NGGPSSGAPPP (SEQ ID NO: 31); NGGPSSGAPPS (SEQ ID NO: 32); NGGPSSGAPPSK (SEQ ID NO: 33); NGGPSSGAPPS(K).sub.2-6 (SEQ ID NO: 34); NGGPSSGAPPPSK (SEQ ID NO: 35); or NK.

[0061] When Z is OH or NH.sub.2, the compounds of Formula (F) are N-terminus conformationally constrained compounds. When Z is (iii), (iv), (v) or (vi), the compounds of Formula (F) are N-terminus conformationally constrained GLP-1 receptor agonist compounds.

[0062] Exemplary compounds of Formula (F) include the compounds in FIGS. 15A-D and 16A-H, each of which may be optionally amidated at the C-terminal amino acid residue. The reaction schemes for preparing the compounds are shown, e.g., in FIGS. 18-20.

[0063] Additional studies were undertaken to restrict the N-terminus conformation of GLP-1 receptor agonist compounds and it was unexpectedly discovered that the .beta.-turn in GLP-1 receptor agonist compounds, such as exendin and exendin analogs, was provided by His.sup.1Gly.sup.2Glu.sup.3Gly.sup.4 (SEQ ID NO: 36). Thus, GLP-1 receptor agonist compounds were produced to constrain or mimic a .beta.-turn defined by residues His.sup.1Gly.sup.2Glu.sup.3Gly.sup.4 (SEQ ID NO: 36) in exendin-4 and other GLP-1 receptor agonist compounds; or to constrain or mimic a .beta.-turn defined by residues His.sup.1Ala.sup.2Glu.sup.3Gly.sup.4 (SEQ ID NO: 37) in GLP-1, GLP-1 analogs, GLP-1(7-37) (SEQ ID NO: 84), or GLP-1(7-37) analogs.

[0064] Provided herein are N-terminus conformationally constrained compounds of Formula (A): Xaa.sub.1Xaa.sub.2Xaa.sub.3Xaa.sub.4-Z.

[0065] In one embodiment, the disclosure provides the compound of Formula (A):

Xaa.sub.1Xaa.sub.2Xaa.sub.3Xaa.sub.4-Z;

wherein: [0066] Xaa.sub.1 is a compound of Formula (1):

[0066] ##STR00002## [0067] wherein R.sub.20 and R.sub.21 are each independently a single bond or a carbon atom; R.sub.23, R.sub.24, R.sub.25 and R.sub.26 are each independently absent, hydrogen, hydroxyl, C.sub.1-4 alkyl, carboxyl, or C.sub.1-4 alkoxy; - - - - - - is a single bond or a double bond; and R.sub.21 is a chiral or achiral carbon atom; [0068] Xaa.sub.2 is Gly, dAla, Aib, Ala, Val, NMeAla, a compound of Formula (3), as described herein; or a compound of Formula (4) and described herein; and Xaa.sub.2 is absent when Xaa.sub.3 is:

[0068] ##STR00003## [0069] Xaa.sub.3 is Pro; a compound of Formula (2), as described herein; a compound of Formula (3), as described herein; a Compound of Formula (4), as described herein;

[0069] ##STR00004## [0070] Xaa.sub.4 is Gly, dAla, or Aib; and [0071] Z is OH or NH.sub.2. In other embodiments, Xaa.sub.3 is Pro. In other embodiments, Xaa.sub.2 is dAla, Aib, Ala, Val, NMeAla, a compound of Formula (3), or a compound of Formula (4).

[0072] In other embodiments, the disclosure provides the compound of Formula (A):

Xaa.sub.1Xaa.sub.2Xaa.sub.3Xaa.sub.4-Z;

wherein: [0073] Xaa.sub.1 is

[0073] ##STR00005## [0074] Xaa.sub.2 is Gly, dAla, Aib, Ala, Val, NMeAla, a compound of Formula (3), as described herein; or a compound of Formula (4), as described herein; and Xaa.sub.2 is absent when Xaa.sub.3 is:

[0074] ##STR00006## [0075] Xaa.sub.3 is Glu; Pro; a compound of Formula (2), as described herein; a compound of Formula (3), as described herein; a Compound of Formula (4), as described herein;

[0075] ##STR00007## [0076] Xaa.sub.4 is Gly, dAla, or Aib; and [0077] Z is OH or NH.sub.2.

[0078] Also provided herein are N-terminus conformationally constrained GLP-1 receptor agonist compounds of Formula (B)-(E):

TABLE-US-00003 (B) (SEQ ID NO: 1) Xaa.sub.1Xaa.sub.2Xaa.sub.3Xaa.sub.4TFTSDLSKQXaa.sub.14EEEAVRLFIEXaa.sub.2- 5LKN- Z; (C) (SEQ ID NO: 2) Xaa.sub.1Xaa.sub.2Xaa.sub.3Xaa.sub.4TFTSDLSKQXaa.sub.14EEEAVRLFIEXaa.sub.2- 5LK- R.sub.10-Z; (D) (SEQ ID NO: 3) Xaa.sub.1Xaa.sub.2Xaa.sub.3Xaa.sub.4TFTSDLSKQXaa.sub.14EEEAVRLFIEXaa.sub.2- 5LKNGG PSSGAPPPS-Z; (E) (SEQ ID NO: 4) Xaa.sub.1Xaa.sub.2Xaa.sub.3Xaa.sub.4TFTSDVSSYXaa.sub.14EGQAAKEFIAXaa.sub.2- 5LV Xaa.sub.28GRXaa.sub.31-Z.

The substituents for the compounds of Formula (A), (B), (C), (D), and (E) are as follows: [0079] Xaa.sub.1 is a compound of Formula (1), as described herein; [0080] Xaa.sub.2 is Gly; dAla; Aib; Ala; Val; NMeAla; a compound of Formula (3), as described herein; or a compound of Formula (4), as described herein; and Xaa.sub.2 is absent when Xaa.sub.3 is:

[0080] ##STR00008## [0081] Xaa.sub.3 is Pro; Glu; Asp; a compound of Formula (2), as described herein; a compound of Formula (3), as described herein; a Compound of Formula (4), as described herein;

[0081] ##STR00009## [0082] Xaa.sub.4 is Gly, dAla, or Aib; [0083] Xaa.sub.14 is Leu or Met; [0084] Xaa.sub.25 is Phe or Trp; [0085] Xaa.sub.28 is Lys or Arg; [0086] Xaa.sub.31 is Gly or absent; [0087] R.sub.10 is QGGPSKEIIS (SEQ ID NO: 22); QGGPSSGAPPPS (SEQ ID NO: 23); NG; NGG; NGGP (SEQ ID NO: 24); NGGPS (SEQ ID NO: 25); NGGPSS (SEQ ID NO: 26); NGGPSSG (SEQ ID NO: 27); NGGPSSGA (SEQ ID NO: 28); NGGPSSGAP (SEQ ID NO: 29); NGGPSSGAPP (SEQ ID NO: 30); NGGPSSGAPPP (SEQ ID NO: 31); NGGPSSGAPPS (SEQ ID NO: 32); NGGPSSGAPPSK (SEQ ID NO: 33); NGGPSSGAPPS(K).sub.2-6 (SEQ ID NO: 34); NGGPSSGAPPPSK (SEQ ID NO: 35); or NK; and [0088] Z is OH or NH.sub.2.

[0089] The compounds of Formula (A)-(E) may optionally be in the form of a pharmaceutically acceptable salt.

[0090] The compound of Formula (1) is:

##STR00010##

wherein R.sub.20 and R.sub.21 are each independently a single bond or a carbon atom; R.sub.23, R.sub.24, R.sub.25 and R.sub.26 are each independently absent, hydrogen, hydroxyl, C.sub.1-4 alkyl, carboxyl, amino, or C.sub.1-4 alkoxy; - - - - - - is a single bond or a double bond; and R.sub.21 is a chiral or achiral carbon atom.

[0091] In one embodiment for the compound of Formula (1), R.sub.20 and R.sub.21 are each independently a single bond or a carbon atom; R.sub.23 and R.sub.24 are each independently absent, hydrogen, hydroxy, a C.sub.1-4 alkyl, carboxyl, or a C.sub.1-4 alkoxy; R.sub.25 and R.sub.26 are each independently absent, hydrogen, hydroxy, a C.sub.1-4 alkyl, carboxyl, amino, or a C.sub.1-4 alkoxy; - - - - - - is a single bond or a double bond; and R.sub.21 is a chiral or achiral carbon atom.

[0092] In one embodiment for the compound of Formula (1), R.sub.20 and R.sub.21 are each independently a single bond or a carbon atom; R.sub.23 and R.sub.24 are each independently absent, hydrogen, hydroxy, methyl, ethyl, or carboxyl; R.sub.25 and R.sub.26 are each independently absent or hydrogen; - - - - - - is a single bond or a double bond; and R.sub.21 is a chiral or achiral carbon atom.

[0093] In one embodiment, the compound of Formula (1) is a compound of Formula (1a), (1b), (1c), (1d), (1e), (1f), (1g), (1h), (1j), (1k), (1m), or (1n):

##STR00011## ##STR00012##

[0094] In one embodiment, the compound of Formula (1) is a compound of Formula (1a), (1b), (1c), (1d), (1e), (1f), (1g), (1 h), (1j), (1k), or (1m).

[0095] In one embodiment, the compound of Formula (1) is a compound of Formula (1a), (1b), (1c), (1d), (1e), (1f), (1g), (1 h), or (1k).

[0096] In one embodiment, the compound of Formula (1) is a compound of Formula (1j) or (1m).

[0097] In one embodiment, the compound of Formula (1) is a compound of Formula (1n).

[0098] The compounds of Formula (2) and Formula (3) are:

##STR00013##

wherein Y.sub.1 and Z.sub.1 are each independently a single bond, a carbon, or a sulfur; and W.sub.1, W.sub.2 and W.sub.3 are each independently selected from hydrogen, C.sub.1-6 alkyl, C.sub.1-6 alkoxy, hydroxyl, and amino; and when one Y.sub.1 or Z.sub.1 is sulfur, the sulfur may be bonded to two oxygen atoms to form a sulfonyl group; and - - - - - - is or .

[0099] In one embodiment for the Compounds of Formula (2) and (3), Y.sub.1 and Z.sub.1 are each independently a single bond, a carbon, or a sulfur; and W.sub.1, W.sub.2 and W.sub.3 are each independently selected from hydrogen, C.sub.1-4 alkyl, C.sub.1-4 alkoxy, and amino; and when one Y.sub.1 or Z.sub.1 is sulfur, the sulfur may be bonded to two oxygen atoms to form a sulfonyl group; and - - - - - - is or .

[0100] In one embodiment for the Compounds of Formula (2) and (3), Y.sub.1 and Z.sub.1 are each independently a single bond, a carbon, or a sulfur; and W.sub.1, W.sub.2 and W.sub.3 are each independently selected from hydrogen, C.sub.1-4 alkyl, and C.sub.1-4 alkoxy; and when one Y.sub.1 or Z.sub.1 is sulfur, the sulfur may be bonded to two oxygen atoms to form a sulfonyl group; and - - - - - - is or .

[0101] In one embodiment for the Compounds of Formula (2) and (3), - - - - - - is .

[0102] In one embodiment for the Compounds of Formula (2), Y.sub.1 and Z.sub.1 are each independently a single bond, a carbon, or a sulfur; and W.sub.1, W.sub.2 and W.sub.3 are each independently selected from the group consisting of hydrogen, methyl, ethyl, and propyl; and - - - - - - is .

[0103] In one embodiment for the Compounds of Formula (3), Y.sub.1 and Z.sub.1 are each independently a single bond or carbon; and W.sub.1, W.sub.2 and W.sub.3 are each independently selected from the group consisting of hydrogen, methyl, ethyl, and propyl; and - - - - - - is .

[0104] In one embodiment, the compound of Formula (2) is a compound of Formula (2Z):

##STR00014##

wherein R.sub.40, R.sub.41, and R.sub.42 are each independently selected from the group consisting of hydrogen and C.sub.1-4 alkyl (preferably methyl or ethyl). Exemplary compounds of Formula (2Z) include compounds of Formula (2d), (2e), (2f), (2g), and (2h) described below.

[0105] In one embodiment, the compound of Formula (2) is a compound of Formula (2a), (2b), (2c), (2d), (2e), (2f), (2g), (2h), or (2j):

##STR00015##

[0106] In one embodiment, the compound of Formula (2) is a compound of Formula (2d), (2e), (20, (2g), or (2h).

[0107] In one embodiment, the compound of Formula (3) is a compound of Formula (3a), (3b), or (3c):

##STR00016##

[0108] The compound of Formula (4) is:

##STR00017##

wherein R.sub.30, R.sub.31, and R.sub.32 are each independently hydrogen or a C.sub.1-4 alkyl; or R.sub.30 and R.sub.31, together with the nitrogen.sup.1 and the carbon.sup.2, form a 5-membered or 6-membered heterocyclic ring; or R.sub.31 and R.sub.32, together with the carbon.sup.2, form a 3-, 4-, or 5-membered carbocyclic ring.

[0109] In one embodiment for the compound of Formula (4), R.sub.30, R.sub.31, and R.sub.32 are each independently hydrogen, methyl, or ethyl; or R.sub.30 and R.sub.31, together with the nitrogen.sup.1 and carbon.sup.2, form a 5-membered or 6-membered heterocyclic ring; or R.sub.31 and R.sub.32, together with the carbon.sup.2, form a 3-, or 4-membered carbocyclic ring;

[0110] In one embodiment, the compound of Formula (4) is a compound of Formula (4a), (4b), (4c), (4d), or (4e):

##STR00018##

[0111] In one embodiment for the Compounds of Formula (A)-(F), Xaa.sub.2 is dAla, Aib, Ala, Val, or NMeAla.

[0112] In one embodiment for the Compounds of Formula (A)-(F), Xaa.sub.2 is a compound of Formula (3).

[0113] In one embodiment for the Compounds of Formula (A)-(F), Xaa.sub.2 is a compound of Formula (4).

[0114] In one embodiment for the Compounds of Formula (A)-(F), Xaa.sub.2 is Gly.

[0115] In one embodiment for the Compounds of Formula (A)-(F), Xaa.sub.2 is dAla.

[0116] In one embodiment for the Compounds of Formula (A)-(F), Xaa.sub.2 is dAla or Aib.

[0117] In one embodiment for the Compounds of Formula (A)-(F), Xaa.sub.3 is Pro.

[0118] In one embodiment for the Compounds of Formula (A)-(F), Xaa.sub.3 is Glu.

[0119] In one embodiment for the Compounds of Formula (A)-(F), Xaa.sub.3 is a compound of Formula (2), as described herein.

[0120] In one embodiment for the Compounds of Formula (A)-(F), Xaa.sub.3 is a compound of Formula (3), as described herein.

[0121] In one embodiment for the Compounds of Formula (A)-(F), Xaa.sub.2 is absent and Xaa.sub.3 is

##STR00019##

[0122] In one embodiment for the Compounds of Formula (A)-(F), Xaa.sub.4 is Gly or dAla.

[0123] In one embodiment for the Compounds of Formula (A)-(F), Z is NH.sub.2.

[0124] For the Compounds of Formula (A)-(F): in one embodiment R.sub.10 is QGGPSKEIIS (SEQ ID NO: 22); in one embodiment R.sub.10 is NG; in one embodiment R.sub.10 is NGG; in one embodiment R.sub.10 is NGGP (SEQ ID NO: 24); in one embodiment R.sub.10 is NGGPS (SEQ ID NO: 25); in one embodiment R.sub.10 is NGGPSS (SEQ ID NO: 26); in one embodiment R.sub.10 is NGGPSSG (SEQ ID NO: 27); in one embodiment R.sub.10 is NGGPSSGA (SEQ ID NO: 28); in one embodiment R.sub.10 is NGGPSSGAP (SEQ ID NO: 29); in one embodiment R.sub.10 is NGGPSSGAPP (SEQ ID NO: 30); in one embodiment R.sub.10 is NGGPSSGAPPP (SEQ ID NO: 31); in one embodiment R.sub.10 is NGGPSSGAPPS (SEQ ID NO: 32); in one embodiment R.sub.10 is NGGPSSGAPPSK (SEQ ID NO: 33); in one embodiment R.sub.10 is NGGPSSGAPPS(K).sub.2-6 (SEQ ID NO: 34); in one embodiment R.sub.10 is NGGPSSGAPPPSK (SEQ ID NO: 35); and in one embodiment R.sub.10 is NK; and in one embodiment R.sub.10 is QGGPSSGAPPPS (SEQ ID NO: 23).

[0125] For the Compounds of Formula (A)-(F): in one embodiment Xaa.sub.14 is Met and Xaa.sub.25 is Trp; in one embodiment Xaa.sub.14 is Leu and Xaa.sub.25 is Phe; in one embodiment Xaa.sub.14 is Met and Xaa.sub.25 is Phe; and in one embodiment Xaa.sub.14 is Leu and Xaa.sub.25 is Trp.

[0126] With respect to the compounds of Formula (D), Xaa.sub.2, Xaa.sub.3, Xaa.sub.14, and Xaa.sub.25 cannot simultaneously be Gly, Glu, Met, and Trp, respectively, except when the compound of Formula (1) is a compound of Formula 1(j) or 1(m). Thus, when Xaa.sub.2, Xaa.sub.3, Xaa.sub.14, and Xaa.sub.25 are Gly, Glu, Met, and Trp, respectively, the compound of Formula (D) may be one of the following:

##STR00020##

[0127] In one embodiment, the N-terminus conformationally constrained GLP-1 receptor agonist compound may be Pro.sup.3-exendin-4 (SEQ ID NO: 40); Pro.sup.3,Leu.sup.14-exendin-4 (SEQ ID NO: 41); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4 (SEQ ID NO: 42); Pro.sup.3-exendin-4(1-28) (SEQ ID NO: 43); Pro.sup.3,Leu.sup.14-exendin-4(1-28) (SEQ ID NO: 44); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-28) (SEQ ID NO: 45); Pro.sup.3-exendin-4(1-36) (SEQ ID NO: 46); Pro.sup.3,Leu.sup.14-exendin-4(1-36) (SEQ ID NO: 47); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-36) (SEQ ID NO: 48); or HGPGTFTSDLSKQLEEEAVRLFIEWLKQGGPSKEIIS (SEQ ID NO: 49), each of which may optionally be amidated and which may optionally be in the form of a pharmaceutically acceptable salt.

[0128] In one embodiment, the N-terminus conformationally constrained GLP-1 receptor agonist compound may be Pro.sup.3-exendin-3 (SEQ ID NO: 50); Pro.sup.3-exendin-4(1-29) (SEQ ID NO: 51); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-29) (SEQ ID NO: 52); Pro.sup.3-exendin-4(1-30) (SEQ ID NO: 53); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-30) (SEQ ID NO: 54); Pro.sup.3-exendin-4(1-31) (SEQ ID NO: 55); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-31) (SEQ ID NO: 56); Pro.sup.3-exendin-4(1-32) (SEQ ID NO: 57); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-32) (SEQ ID NO: 58); Pro.sup.3-exendin-4(1-33) (SEQ ID NO: 59); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-33) (SEQ ID NO: 60); Pro.sup.3-exendin-4(1-34) (SEQ ID NO: 61); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-34) (SEQ ID NO: 62); Pro.sup.3-exendin-4(1-35) (SEQ ID NO: 63); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-35) (SEQ ID NO: 64); Pro.sup.3-exendin-4(1-37) (SEQ ID NO: 65); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-37) (SEQ ID NO: 66); Pro.sup.3-exendin-4(1-38) (SEQ ID NO: 67); or Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-38) (SEQ ID NO: 68), each of which may optionally be amidated and which may optionally be in the form of a pharmaceutically acceptable salt.

[0129] Examples of the compounds of Formula (A), (B), (C), (D), and (E) are shown in FIGS. 1B-C, 2A-U, 3A-G, 5B-G, 6A-E, 8A-H, 9A-H, 10A-I, 11A-B, 12, 13, 14A-Q, 15C-D, 16I-L, and 17A-N.

[0130] The N-terminus conformationally constrained compounds (e.g., compounds of Formula (A) and (F)) and the N-Terminus conformationally constrained GLP-1 receptor agonist compounds (e.g., compounds of Formula (B)-(F)) described herein (collectively referred to as "the compounds") can optionally be covalently linked to one or more polymers to provide beneficial biological properties to the compounds. Such beneficial properties may include conferring another therapeutic property to the compounds; increasing the in vivo half life of the compounds; decreasing the rate of clearance of the compounds by the kidney; decreasing the immunogenicity of the compounds; decreasing the proteolysis rate of the compounds; or increasing the stability of the compounds. Exemplary polymers that can be covalently linked to the compounds include peptides, polyethylene glycols, albumin, fatty acids, dextran, polyamino acids, alkyl chains, immunoglobulins, signaling moieties, gelatin, polyvinyl pyrrolidone, polyvinyl alcohol, N-(2-hydroxypropyl)-methacrylamide, and the like. In one embodiment, two or more polymers (e.g., peptides, polyethylene glycols, albumin, fatty acids, dextran, polyamino acids, alkyl chains, immunoglobulins, gelatin, polyvinyl pyrrolidone, polyvinyl alcohol, N-(2-hydroxypropyl)-methacrylamide) are covalently attached together and linked to the N-terminus conformation constrained compounds described herein. For example, the two more polymers that are linked together may be polyethylene glycol(s) and fatty acid(s) or a peptide, polyethylene glycol(s), and fatty acids.

[0131] In one embodiment, the compounds are linked to another peptide to provide additional therapeutic benefits. Such peptides include amylin, amylin agonist analogs (e.g., amylin analogs that function as amylin agonists), PYY, PYY analogs, GIP, GIP analogs, leptin, metraleptin, leptin analogs, metraleptin analogs, and the like. Hybrid peptides comprising the compounds and another therapeutic peptide are described, for example, in WO 2005/077072 and WO 2007/022123, the disclosures of which are incorporated by reference herein.

[0132] In one embodiment, compounds are linked to one, two, or three polyethylene glycols. In one embodiment, the compounds are linked to one polyethylene glycol. The polyethylene glycol can have a molecular weight from about 200 daltons to about 80,000 daltons; from about 5,000 from about 10,000 daltons to about 60,000 daltons; from about 10,000 daltons to about 50,000 daltons; or from about 15,000 daltons to about 40,000 daltons. The polyethylene glycol may be linear or branched.

[0133] In one embodiment, compounds are linked to one or two polyethylene glycols, where the polyethylene glycol is further linked to a lipophilic moiety. In one embodiment, the polyethylene glycol may have a molecular weight from about 200 to about 7,000 daltons or from about 500 to about 5,000 daltons. The lipophilic moiety may be an alkyl group (e.g., C.sub.1-20 alkyl group; C.sub.1-10 alkyl group; C.sub.1-6 alkyl group; C.sub.1-4 alkyl group), a fatty acid (e.g., C.sub.4-28 fatty acid; C.sub.4-20 fatty acid; C.sub.4-10 fatty acid), cholesteryl, adamantyl, and the like. The alkyl group may be linear or branched, preferably linear. In one embodiment, the fatty acid is an acetylated fatty acid or an esterified fatty acid. The -(polyethylene glycol)-(lipophilic moiety) may be linked to the compound at a C-terminal amino acid residue, an N-terminal amino acid residue, an internal amino acid residue (e.g., an internal Lys amino acid residue), or a combination thereof (e.g., the compound is linked at the N-terminal and C-terminal amino acid residues via a lysine residue). Examplary peptides linked to such groups are shown in Example 20.

[0134] In one embodiment, the compounds are linked to a polyamino acid. Exemplary polyamino acids include poly-lysine, poly-aspartic acid, poly-serine, poly-glutamic acid, and the like. The polyamino acid may be in the D or L form, preferably the L form. The polyamino acids may comprise from 1 to 12 amino acid residues; from 2 to 10 amino acid residues; or from 2 to 6 amino acid residues.

[0135] In one embodiment, compounds are linked to a fatty acid. The fatty acid may be a C.sub.4-C.sub.28 fatty acid chain, a C.sub.8-C.sub.24 fatty acid chain, or a C.sub.10-C.sub.20 fatty acid chain. In one embodiment, the fatty acid is an acetylated fatty acid. In one embodiment, the fatty acid is an esterified fatty acid.

[0136] In one embodiment, the compounds are linked to albumin. The albumin may be a recombinant albumin, serum albumin, or recombinant serum albumin. In another embodiment, the compounds are linked to an albumin-fatty acid (i.e., an albumin linked to a fatty acid).

[0137] In one embodiment, the compounds are linked to an immunoglobulin or an immunoglobulin Fc region. The immunoglobulin may be IgG, IgE, IgA, IgD, or IgM. In one embodiment, the compounds are linked to an IgG Fc region or an IgM Fc region. The immunoglobulin Fc region is (i) the heavy chain constant region 2(C.sub.H2) of an immunoglobulin; (ii) the heavy chain constant region 3(C.sub.H3) of an immunoglobulin; or (iii) both the heavy chain constant regions 2(C.sub.H2) and 3(C.sub.H3) of an immunoglobulin. The immunoglobulin Fc region may further comprise the hinge region at the heavy chain constant region. Other embodiments for the immunoglobulin Fc region that can be linked to exendin analog peptides are described in WO 2008/082274, the disclosure of which is incorporated by reference herein.

[0138] In one embodiment, the compounds are linked to one or more signalling moieties. Exemplary signalling moieties include, biotin, antigens, antibodies, receptors, enzymes, chemiluminescent groups, photoreactive groups, fluorescent groups, heavy metal-containing compounds (e.g., ferritin), and the like.

[0139] When the compounds described herein are covalently linked to one or more polymers, such as those described herein, any linking group known in the art can be used. The linking group may comprise any chemical group(s) suitable for linking the peptide to the polymer. Alternatively, compounds can be directly attached to the polymer without any linking group. Exemplary linking groups include amino acids, maleimido groups, dicarboxylic acid groups, succinimide groups, or a combination of two or more thereof. Methods for linking peptides to one or more polymers are known in the art and described, for example, in U.S. Pat. No. 6,329,336; U.S. Pat. No. 6,423,685; U.S. Pat. No. 6,924,264; WO 2005/077072, WO 2007/022123, WO 2007/053946; WO 2008/058461; and WO 2008/082274, the disclosures of which are incorporated by reference herein.

[0140] The compounds described herein may be prepared using biological, chemical, and/or recombinant DNA techniques that are known in the art. Exemplary methods are described in U.S. Pat. No. 6,872,700; WO 2007/139941; WO 2007/140284; WO 2008/082274; WO 2009/011544; and US Publication No. 2007/0238669, the disclosures of which are incorporated herein by reference. Other methods for preparing the compounds are set forth herein.

[0141] The compounds described herein may be prepared using standard solid-phase peptide synthesis techniques, such as an automated or semiautomated peptide synthesizer. Typically, using such techniques, an alpha-N-carbamoyl protected amino acid and an amino acid attached to the growing peptide chain on a resin are coupled at room temperature in an inert solvent (e.g., dimethylformamide, N-methylpyrrolidinone, methylene chloride, and the like) in the presence of coupling agents (e.g., dicyclohexylcarbodiimide, 1-hydroxybenzo-triazole, and the like) in the presence of a base (e.g., diisopropylethylamine, and the like). The alpha-N-carbamoyl protecting group is removed from the resulting peptide-resin using a reagent (e.g., trifluoroacetic acid, piperidine, and the like) and the coupling reaction repeated with the next desired N-protected amino acid to be added to the peptide chain. Suitable N-protecting groups are well known in the art, such as t-butyloxycarbonyl (tBoc) fluorenylmethoxycarbonyl (Fmoc), and the like. The solvents, amino acid derivatives and 4-methylbenzhydryl-amine resin used in the peptide synthesizer may be purchased from Applied Biosystems Inc. (Foster City, Calif.).

[0142] Solid phase peptide synthesis may be carried out with an automatic peptide synthesizer (Model 430A, Applied Biosystems Inc., Foster City, Calif.) using the NMP/HOBt (Option 1) system and tBoc or Fmoc chemistry (See Applied Biosystems User's Manual for the ABI 430A Peptide Synthesizer, Version 1.3B Jul. 1, 1988, section 6, pp. 49-70, Applied Biosystems, Inc., Foster City, Calif.) with capping. Boc-peptide-resins may be cleaved with HF (-5.degree. C. to 0.degree. C., 1 hour). The peptide may be extracted from the resin with alternating water and acetic acid, and the filtrates lyophilized. The Fmoc-peptide resins may be cleaved according to standard methods (e.g., Introduction to Cleavage Techniques, Applied Biosystems, Inc., 1990, pp. 6-12). Peptides may be also be assembled using an Advanced Chem Tech Synthesizer (Model MPS 350, Louisville, Ky.).

[0143] Peptides may be purified by RP-HPLC (preparative and analytical) using a Waters Delta Prep 3000 system. A C4, C8 or C18 preparative column (10.mu., 2.2.times.25 cm; Vydac, Hesperia, Calif.) may be used to isolate peptides, and purity may be determined using a C4, C8 or C18 analytical column (5.mu., 0.46.times.25 cm; Vydac). Solvents (A=0.1% TFA/water and B=0.1% TFA/CH.sub.3CN) may be delivered to the analytical column at a flow rate of 1.0 ml/min and to the preparative column at 15 ml/min. Amino acid analyses may be performed on the Waters Pico Tag system and processed using the Maxima program. Peptides may be hydrolyzed by vapor-phase acid hydrolysis (115.degree. C., 20-24 h). Hydrolysates may be derivatized and analyzed by standard methods (Cohen et al, The Pico Tag Method: A Manual of Advanced Techniques for Amino Acid Analysis, pp. 11-52, Millipore Corporation, Milford, Mass. (1989)). Fast atom bombardment analysis may be carried out by M-Scan, Incorporated (West Chester, Pa.). Mass calibration may be performed using cesium iodide or cesium iodide/glycerol. Plasma desorption ionization analysis using time of flight detection may be carried out on an Applied Biosystems Bio-Ion 20 mass spectrometer.

[0144] The compounds described herein may also be prepared using recombinant DNA techniques using methods known in the art, such as Sambrook et al., Molecular Cloning: A Laboratory Manual, 2d Ed., Cold Spring Harbor (1989). Non-peptide compounds may be prepared by art-known methods. For example, phosphate-containing amino acids and peptides containing such amino acids, may be prepared using methods known in the art, such as described in Bartlett et al, Biorg. Chem., 14:356-377 (1986).

[0145] The disclosure also provides pharmaceutical compositions comprising at least one of the N-terminus conformationally constrained compounds or the N-terminus conformationally constrained GLP-1 receptor agonist compounds described herein and a pharmaceutically acceptable carrier. The N-terminus conformationally constrained compounds or the N-terminus conformationally constrained GLP-1 receptor agonist compounds can be present in the pharmaceutical composition in a therapeutically effective amount and can be present in an amount to provide a minimum blood plasma level for therapeutic efficacy.

[0146] Pharmaceutical compositions containing the compounds described herein may be provided for peripheral administration, such as parenteral (e.g., subcutaneous, intravenous, intramuscular), topical, nasal, or oral administration. Suitable pharmaceutically acceptable carriers and their formulation are described in standard formulation treatises, such as Remington's Pharmaceutical Sciences by Martin; and Wang et al, Journal of Parenteral Science and Technology, Technical Report No. 10, Supp. 42:2 S (1988).

[0147] The compounds described herein can be provided in parenteral compositions for injection or infusion. They can, for example, be suspended in water; an inert oil, such as a vegetable oil (e.g., sesame, peanut, olive oil, and the like); or other pharmaceutically acceptable carrier. In one embodiment, the compounds are suspended in an aqueous carrier, for example, in an isotonic buffer solution at a pH of about 3.0 to 8.0, or about 3.0 to 5.0. The compositions may be sterilized by conventional sterilization techniques or may be sterile filtered. The compositions may contain pharmaceutically acceptable auxiliary substances as required to approximate physiological conditions, such as pH buffering agents. Useful buffers include for example, acetic acid buffers. A form of repository or "depot" slow release preparation may be used so that therapeutically effective amounts of the preparation are delivered into the bloodstream over many hours or days following subcutaneous injection, transdermal injection or other delivery method. The desired isotonicity may be accomplished using sodium chloride or other pharmaceutically acceptable agents such as dextrose, boric acid, sodium tartrate, propylene glycol, polyols (such as mannitol and sorbitol), or other inorganic or organic solutes. Sodium chloride is preferred particularly for buffers containing sodium ions.

[0148] The compounds can also be formulated as pharmaceutically acceptable salts (e.g., acid addition salts) and/or complexes thereof. Pharmaceutically acceptable salts are non-toxic salts at the concentration at which they are administered. Pharmaceutically acceptable salts include acid addition salts such as those containing sulfate, hydrochloride, phosphate, sulfamate, acetate, citrate, lactate, tartrate, methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonate, cyclohexylsulfamate and quinate. Pharmaceutically acceptable salts can be obtained from acids such as hydrochloric acid, sulfuric acid, phosphoric acid, sulfamic acid, acetic acid, citric acid, lactic acid, tartaric acid, malonic acid, methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, cyclohexylsulfamic acid, and quinic acid. Such salts may be prepared by, for example, reacting the free acid or base forms of the product with one or more equivalents of the appropriate base or acid in a solvent or medium in which the salt is insoluble, or in a solvent such as water which is then removed in vacuo or by freeze-drying or by exchanging the ions of an existing salt for another ion on a suitable ion exchange resin.

[0149] Carriers or excipients can also be used to facilitate administration of the compounds. Examples of carriers and excipients include calcium carbonate, calcium phosphate, various sugars such as lactose, glucose, or sucrose, or types of starch, cellulose derivatives, gelatin, vegetable oils, polyethylene glycols and physiologically compatible solvents.

[0150] If desired, solutions of the above compositions may be thickened with a thickening agent such as methyl cellulose. They may be prepared in emulsified form, either water in oil or oil in water. Any of a wide variety of pharmaceutically acceptable emulsifying agents may be employed including, for example, acacia powder, a non-ionic surfactant (such as a Tween), or an ionic surfactant (such as alkali polyether alcohol sulfates or sulfonates, e.g., a Triton).

[0151] Compositions may be prepared by mixing the ingredients following generally accepted procedures. For example, the selected components may be simply mixed in a blender or other standard device to produce a concentrated mixture which may then be adjusted to the final concentration and viscosity by the addition of water or thickening agent and possibly a buffer to control pH or an additional solute to control tonicity.

[0152] The therapeutically effective amount of the compounds described herein to treat the diseases described herein will typically be from about 0.01 .mu.g to about 5 mg; about 0.1 .mu.g to about 2.5 mg; about 1 .mu.g to about 1 mg; about 1 .mu.g to about 50 .mu.g; or about 1 .mu.g to about 25 .mu.g. Alternatively, the therapeutically effective amount of the GLP-1 receptor agonist compounds may be from about 0.001 .mu.g to about 100 .mu.g based on the weight of a 70 kg patient; or from about 0.01 .mu.g to about 50 .mu.g based on the weight of a 70 kg patient. These therapeutically effective doses may be administered once/day, twice/day, thrice/day, once/week, biweekly, or once/month, depending on the formulation. The exact dose to be administered is determined, for example, by the formulation, such as an immediate release formulation or an extended release formulation. For transdermal, nasal or oral dosage forms, the dosage may be increased from about 5-fold to about 10-fold.

[0153] The compounds described herein and pharmaceutical compositions comprising the compounds are useful for treating diabetes. The diabetes can be Type I diabetes, Type II diabetes, or gestational diabetes. The methods for treating diabetes provide administering to a patient in need thereof a therapeutically effective amount of one or more of the N-terminus conformationally constrained compounds or the N-terminus conformationally constrained GLP-1 receptor agonist compounds described herein to treat diabetes in the patient.

[0154] The compounds described herein and pharmaceutical compositions comprising the compounds are useful for treating insulin resistance and stimulating insulin release. The methods for treating insulin resistance provide administering to a patient in need thereof a therapeutically effective amount of one or more of the N-terminus conformationally constrained compounds or the N-terminus conformationally constrained GLP-1 receptor agonist compounds described herein to treat insulin resistance in the patient. The methods for treating stimulating insulin release provide administering to a patient in need thereof a therapeutically effective amount of one or more of the N-terminus conformationally constrained compounds or the N-terminus conformationally constrained GLP-1 receptor agonist compounds described herein to stimulate insulin release in the patient.

[0155] The compounds described herein and pharmaceutical compositions comprising the compounds are useful for treating postprandial hyperglycemia. The methods for treating postprandial hyperglycemia provide administering to a patient in need thereof a therapeutically effective amount of one or more of the N-terminus conformationally constrained compounds or the N-terminus conformationally constrained GLP-1 receptor agonist compounds described herein to treat postprandial hyperglycemia in the patient.

[0156] The compounds described herein and pharmaceutical compositions comprising the compounds are useful for lowering blood glucose levels and lowering HbA1c levels. The methods for lowering blood glucose levels provide administering to a patient in need thereof a therapeutically effective amount of one or more of the N-terminus conformationally constrained compounds or the N-terminus conformationally constrained GLP-1 receptor agonist compounds described herein to lower blood glucose levels in the patient. In one embodiment, the blood glucose levels can be fasting blood glucose levels. The methods for lowering HbA1c levels provide administering to a patient in need thereof a therapeutically effective amount of one or more of the N-terminus conformationally constrained compounds or the N-terminus conformationally constrained GLP-1 receptor agonist compounds described herein to lower HbA1c levels in the patient. HbA1c levels are generally a long-term measure of a patient's blood glucose levels.

[0157] The compounds described herein and pharmaceutical compositions comprising the compounds are useful for reducing gastric motility and delaying gastric emptying. The methods for reducing gastric motility provide administering to a patient in need thereof a therapeutically effective amount of one or more of the N-terminus conformationally constrained compounds or the N-terminus conformationally constrained GLP-1 receptor agonist compounds described herein to reduce gastric motility in the patient. The methods for delaying gastric emptying provide administering to a patient in need thereof a therapeutically effective amount of one or more of the N-terminus conformationally constrained compounds or the N-terminus conformationally constrained GLP-1 receptor agonist compounds described herein to delay gastric emptying in the patient.

[0158] The compounds described herein and pharmaceutical compositions comprising the compounds are useful for reducing food intake, reducing appetite, increasing satiety, and reducing weight. The methods for reducing food intake provide administering to a patient in need thereof a therapeutically effective amount of one or more of the N-terminus conformationally constrained compounds or the N-terminus conformationally constrained GLP-1 receptor agonist compounds described herein to reduce food intake in the patient. The methods for reducing appetite provide or increasing satiety administering to a patient in need thereof a therapeutically effective amount of one or more of the N-terminus conformationally constrained compounds or the N-terminus conformationally constrained GLP-1 receptor agonist compounds described herein to reduce appetite in the patient. The methods for treating reducing weight provide administering to a patient in need thereof a therapeutically effective amount of one or more of the N-terminus conformationally constrained compounds or the N-terminus conformationally constrained GLP-1 receptor agonist compounds described herein to reduce weight in the patient. In the methods described herein, the patient may be in need of a reduced intake in food, of a reduced appetite, or of reduced weight. In other methods described herein, the patient may be desirous of having a reduced intake in food, of having a reduced appetite, or of having a reduced weight. The patient may be of any weight, and can be overweight or obese.

[0159] The compounds described herein and pharmaceutical compositions comprising the compounds are useful for treating overweight and obesity. The methods for treating overweight provide administering to a patient in need thereof a therapeutically effective amount of one or more of the N-terminus conformationally constrained compounds or the N-terminus conformationally constrained GLP-1 receptor agonist compounds described herein to treat overweight in the patient. The methods for treating obesity provide administering to a patient in need thereof a therapeutically effective amount of one or more of the N-terminus conformationally constrained compounds or the N-terminus conformationally constrained GLP-1 receptor agonist compounds described herein to treat obesity in the patient.

[0160] The disclosure also provides drug delivery devices having at least one therapeutically effective dose of the compounds described herein or the pharmaceutical composition containing the compounds described herein. The drug delivery devices can be single or multiple-use vials, single or multiple-use pharmaceutical pens, single or multiple-use cartridges, and the like. In one embodiment, the drug delivery devices contain the compounds or pharmaceutical compositions described herein in amounts capable of providing a patient with from about 7 to about 40 doses or enough doses to last about one week or about one month.

EXAMPLES

[0161] The following examples are for purposes of illustration only and are not intended to limit the scope of the claims.

Example 1

Preparation of Compound in FIG. 16A

[0162] A calculated 100 .mu.mol of Fmoc-Ser(OtBu)-Wang resin was weighed into a reaction vessel in a Symphony peptide synthesizer, and the peptide elongation was carried out following standard Fmoc peptide synthesis protocol up to residue Thr.sup.5. The resulting peptide-resin intermediate (0.3 g, 0.0927 mmol) was swollen in dimethylformamide (DMF) and to the slurry was added compound 7 (0.110 g, 2.2 eq), followed by O-(benzotriazol-1-yl)-N,N,N',N' tetramethyluronium hexafluorophosphate (HBTU) (0.035 g, 2.2 eq), N-hydroxybenzotriazole (HOBt) (0.03 g, 2.2 eq) and methylmorpholine (NMM) (0.04 mL, 4.4 eq). After 3 hours, the resin was washed with DMF 6.times., treated with 20% piperidine in DMF 2.times.25 min and washed with DMF 6.times.. The above cycle was repeated with Fmoc-Ala-OH and Fmoc-His(Trt)-OH followed by cleavage of the peptide from the resin with 10 ml TFA/H.sub.2O/PhOH/TIPS (95:2:2:1) (TFA is trifluoroacetic acid; TIPS is triisopropylsilyl), precipitated by methyl-tert-Butyl ether and the obtained residue applied to a reverse-phase high performance liquid chromatography (HPLC) column (C.sub.18, 20-50% CH.sub.3CN in 0.1% TFA/H.sub.2O over 30 min gradient) to afford the titled compound as a white powder (16.4 mg, 6%): Retention time in reverse phase-high performance liquid chromatography (RP-HPLC) (C.sub.18, 5-75% CH.sub.3CN in 0.1% TFA/H.sub.2O over 15 min) is 9.78 min; Calculated mass for C.sub.189H.sub.294N.sub.48O.sub.60S (M+H).sup.+ 4231.84, found by liquid chromatography/mass spectrometry (LC-MS) 1411.3 (M+3H).sup.3+, 1059.6 (M+4H).sup.4+, 2117.2 (M+2H).sup.2+.

Example 2

Preparation of Compound in FIG. 16B

[0163] A calculated 100 .mu.mol of Fmoc-Ser(OtBu)-Wang resin was weighed into a reaction vessel in a Symphony peptide synthesizer, and the peptide elongation was carried out following standard Fmoc peptide synthesis protocol up to residue Thr.sup.5. The resulting peptide-resin intermediate (0.3 g, 0.0927 mmol) was swollen in DMF and to the slurry was added compound 24 (0.122 g, 2.2 eq), followed by HBTU (0.035 g, 2.2 eq), HOBt (0.03 g, 2.2 eq) and NMM (0.04 mL, 4.4 eq). After 3 hours, the resin was washed with DMF 6.times., treated with 20% piperidine in DMF 2.times.25 min and washed with DMF 6.times.. The above cycle was repeated with Fmoc-His(Trt)-OH followed by cleavage of the peptide from the resin with 10 ml TFA/H.sub.2O/PhOH/TIPS (95:2:2:1), precipitated by methyl-tert-Butyl ether. The obtained crude peptide was dissolved in 1.5 mL of MeOH:ACN (1:1) (ACN is acetonitrile), followed by addition of 2M LiOH (0.6 mL) and the reaction stirred at RT for 6 h. The crude was dissolved and applied to a reverse-phase HPLC column (C.sub.18, 20-50% CH.sub.3CN in 0.1% TFA/H.sub.2O over 30 min gradient) to afford the titled compound as a white powder (15 mg, 6%): Retention time in RP-HPLC (C.sub.18, 5-75% CH.sub.3CN in 0.1% TFA/H.sub.2O over 15 min) is 9.82 min; Calculated mass for C.sub.188H.sub.294N.sub.48O.sub.60 (M+H).sup.+ 4186.72, found by LC-MS 1396.7 (M+3H).sup.3+, 1048.6 (M+4H).sup.4+, 2114.2 (M+2H).sup.2+.

Example 3

Preparation of Compound in FIG. 16C

[0164] A calculated 100 .mu.mol of Fmoc-Ser(OtBu)-Wang resin was weighed into a reaction vessel in a Symphony peptide synthesizer, and the peptide elongation was carried out following standard Fmoc peptide synthesis protocol up to residue Thr.sup.5. The resulting peptide-resin intermediate (0.3 g, 0.0927 mmol) was swollen in DMF and to the slurry was added compound 18 (0.131 g, 2.2 eq), followed by HBTU (0.035 g, 2.2 eq), HOBt (0.03 g, 2.2 eq) and NMM (0.04 mL, 4.4 eq). After 3 hours, the resin was washed with DMF 6.times., treated with 20% piperidine in DMF 2.times.25 min and washed with DMF 6.times.. The above cycle was repeated with Fmoc-His(Trt)-OH followed by cleavage of the peptide from the resin with 10 ml TFA/H.sub.2O/PhOH/TIPS (95:2:2:1), precipitated by methyl-tert-Butyl ether. The obtained crude peptide was dissolved in 1.5 mL of MeOH:ACN (1:1), followed by addition of 2M LiOH (0.6 mL) and the reaction stirred at RT for 6 h. The crude was dissolved and applied to a reverse-phase HPLC column (C.sub.18, 20-50% CH.sub.3CN in 0.1% TFA/H.sub.2O over 30 min gradient) to afford the titled compound as a white powder (19.7 mg, 8%): Retention time in RP-HPLC (C.sub.18, 5-75% CH.sub.3CN in 0.1% TFA/H.sub.2O over 15 min) is 8.58 min; Calculated mass for C.sub.189H.sub.294N.sub.48O.sub.605 (M+H).sup.+ 4230.80, found by LC-MS 1411. (M+3H).sup.3+, 1059.6 (M+4H).sup.4+, 2117.2 (M+2H).sup.2+.

Example 4

Preparation of Compound in FIG. 16E

[0165] The synthesis of this compound was accomplished following the same experimental procedure as described for Example 3. The only difference was the sequence of the peptide-resin intermediate, starting with Rink-amide resin. The crude was dissolved and applied to a reverse-phase HPLC column (C.sub.18, 20-50% CH.sub.3CN in 0.1% TFA/H.sub.2O over 30 min gradient) to afford the titled compound as a white powder (15.7 mg, 7%): Retention time in RP-HPLC (C.sub.18, 5-75% CH.sub.3CN in 0.1% TFA/H.sub.2O over 15 min) is 8.25 min; Calculated mass for C.sub.188H.sub.286N.sub.48O.sub.60S (M+H).sup.+ 4238.74, found by LC-MS 1414.8 (M+3H).sup.3+, 1061.6 (M+4H).sup.4+, 2121.2 (M+2H).sup.2+.

Example 5

Preparation of Compound in FIG. 16D

[0166] A calculated 100 .mu.mol of Fmoc-Ser(OtBu)-Wang resin was weighed into a reaction vessel in a Symphony peptide synthesizer, and the peptide elongation was carried out following standard Fmoc peptide synthesis protocol up to residue Thr.sup.5. The resulting peptide-resin intermediate (0.3 g, 0.0927 mmol) was swollen in DMF and to the slurry was added compound 11 (0.100 g, 2.2 eq), followed by HBTU (0.035 g, 2.2 eq), HOBt (0.03 g, 2.2 eq) and NMM (0.04 mL, 4.4 eq). After 3 h, the resin was washed with DMF 6.times., treated with 20% piperidine in DMF 2.times.25 min and washed with DMF 6.times.. The above cycle was repeated with Fmoc-Ala-OH and Fmoc-His(Trt)-OH followed by cleavage of the peptide from the resin with 10 ml TFA/H.sub.2O/PhOH/TIPS (95:2:2:1), precipitated by methyl-tert-Butyl ether and the obtained residue applied to a reverse-phase HPLC column (C.sub.18, 20-50% CH.sub.3CN in 0.1% TFA/H.sub.2O over 30 min gradient) to afford the titled compound as a white powder (20.3 mg, 10%): Retention time in RP-HPLC (C.sub.18, 5-75% CH.sub.3CN in 0.1% TFA/H.sub.2O over 15 min) is 9.74 min; Calculated mass for C.sub.188H.sub.294N.sub.48O.sub.60 (M+H).sup.+ 4186.72, found by LC-MS 1396.7 (M+3H).sup.3+, 1048.6 (M+4H).sup.4+, 2114.2 (M+2H).sup.2+.

Example 6

Preparation of Compound in FIG. 16F

[0167] A calculated 100 .mu.mol of Fmoc-Ser(OtBu)-Wang resin was weighed into a reaction vessel in a Symphony peptide synthesizer, and the peptide elongation was carried out following standard Fmoc peptide synthesis protocol up to residue Thr.sup.5. The resulting peptide-resin intermediate (0.3 g, 0.0927 mmol) was swollen in DMF and to the slurry was added compound 21 (0.125 g, 2.2 eq), followed by HBTU (0.035 g, 2.2 eq), HOBt (0.03 g, 2.2 eq) and NMM (0.04 mL, 4.4 eq). After 3 h, the resin was washed with DMF 6.times., treated with 20% piperidine in DMF 2.times.25 min and washed with DMF 6.times.. The above cycle was repeated with Fmoc-His(Trt)-OH followed by cleavage of the peptide from the resin with 10 ml TFA/H.sub.2O/PhOH/TIPS (95:2:2:1), precipitated by methyl-tert-Butyl ether. The obtained crude peptide was dissolved in 1.5 mL of MeOH:ACN (1:1), followed by addition of 2M LiOH (0.6 mL) and the reaction stirred at RT for 6 h. The crude was dissolved and applied to a reverse-phase HPLC column (C.sub.18, 20-50% CH.sub.3CN in 0.1% TFA/H.sub.2O over 30 min gradient) to afford the titled compound as a white powder (8 mg, 4%): Retention time in RP-HPLC (C.sub.18, 5-75% CH.sub.3CN in 0.1% TFA/H.sub.2O over 15 min) is 7.93 min; Calculated mass for C.sub.189H.sub.296N.sub.48O.sub.60 (M+H).sup.+ 4200.75, found by LC-MS 1401.8 (M+3H).sup.3+, 1051.6 (M+4H).sup.4+, 2102.2 (M+2H).sup.2+.

Example 7

Preparation of Compound in FIG. 16G

[0168] The synthesis of this compound was accomplished following the same experimental procedure as described for Example 3. The only difference was the sequence of the peptide-resin intermediate. The crude was dissolved and applied to a reverse-phase HPLC column (C.sub.18, 20-50% CH.sub.3CN in 0.1% TFA/H.sub.2O over 30 min gradient) to afford the titled compound as a white powder (27.4 mg, 18%): Retention time in RP-HPLC (C.sub.18, 5-75% CH.sub.3CN in 0.1% TFA/H.sub.2O over 15 min) is 8.25 min; Calculated mass for C.sub.153H.sub.228N.sub.40O.sub.47S (M+H).sup.+ 3411.83, found by LC-MS 1138.6 (M+3H).sup.3+, 854.5 (M+4H).sup.4+, 1707.9 (M+2H).sup.2+.

Example 8

Preparation of (5,5)-Glu-Gly-OH as shown in FIG. 18

[0169] Preparation of Boc-Asp(OBn)-OMe: Boc-Asp(OBn)-OH 2.4 g (1 equiv, 7.45 mmol) was dissolved in 17 mL of dry DMF in a 100 mL round-bottom flask. Finely ground K.sub.2CO.sub.3 (1.5 g, 11 mmol) was added to the solution to form a suspension. The mixture was cooled to 0.degree. C. in an ice-bath over five minutes. MeI (1 mL, 15 mmol) was then added to the mixture over 20 seconds under a positive flow of nitrogen. A yellow color developed within 30 min. The resulting mixture was stirred for 3 hours. The ice-bath was removed and 25 mL of water were added and the mixture left standing at room temperature 2 hours. The mixture was then treated with 30 mL of water and extracted with 3.times.25 mL AcOEt. The organics were washed with saturated NaHCO.sub.3 1.times., brine 3.times., dried over Na.sub.2SO.sub.4 filtered and concentrated to yield a yellow oil. This material was passed through a short silica column (eluting w/ AcOEt), after concentration of the pure fractions 2.42 g of Boc-Asp(OBn)-OMe as a yellow oil was obtained (98% yield). .sup.1H NMR (DMSO d6, 500 mHz): .delta. 7.36 (m, 5H), 5.10 (s, 2H), 4.40 (q, 1H), 3.6 (s, 3H), 2.80 (dd, 1H), 2.72 (dd, 1H), 1.38 (s, 9H). LCMS (C.sub.18, 2-98% CH.sub.3CN in 0.1% TFA/H.sub.2O over 6 min); Calculated mass for C.sub.17H.sub.23NO.sub.6 (M+H).sup.+ 338.23, found by LC-MS 338.2.

[0170] Preparation of Compound 1 in FIG. 18: Boc-Asp(OBn)-OMe (26.4 g, 78.3 mmol) was dissolved in a 500 mL round-bottom flask with 110 mL of dry THF and the mixture cooled at negative 42.degree. C. To this mixture was added lithium bis(trimethylsilyl)amide (LiHMDS) (173 mL, 1 M solution, 2.2 eq). This solution was stirred under a gentle argon flow for 45 min followed by slow addition (via syringe) of tert-butyl bromo acetate (14.5 mL, 1.25 eq). Thin layer chromatography (TLC) (Hex:AcOEt, 8/2) showed after 4 h>95% of expected product. The reaction was quenched by addition of saturated NH.sub.4Cl (70 mL). The mixture was evaporated and the residue re-dissolved with 100 mL of dichloromethane (DCM). The emulsion formed was separated by standing overnight. The organics were collected and washed with saturated NH.sub.4Cl 2.times., brine 1.times., dried over Na.sub.2SO.sub.4, filtered and concentrated to give Compound 1 as a red orange oil. Flash chromatography (Hex:AcOEt, 7:3) gave 27.4 g of a yellow oil (78% yield). .sup.1H NMR (DMSO d6, 500 mHz): .delta. 7.35 (m, 5H), 5.08 (s, 2H), 4.57 (dd, 1H), 4.05 (m, 1H), 3.52 (s, 3H), 1.39 (s, 9H), 1.34 (s, 9H). LCMS (C.sub.18, 2-98% CH.sub.3CN in 0.1% TFA/H.sub.2O over 6 min); Calculated mass for C.sub.23H.sub.33NO.sub.8 (M+H).sup.+ 452.23, found by LC-MS 452.2.

[0171] Preparation of Compound 2 in FIG. 18. Compound 2 is tert-butyl-(1S,2R)-2-[(carboxy)-3-tert-butoxycarbonyl)-1-metoxhycarbonyl)- ]propylcarbamate. Compound 1 (5.8 g, 12.8 mmol) was dissolved in 30 mL of a 6:4 mixture of MeOH/THF. Argon was bubbled in the catalyst for 5 minutes, followed by incorporation of two hydrogen balloons attached via syringe to the reaction flask. After 4 hours, LCMS shows complete transformation. The crude mixture was filtrated through celite and concentrated to yield 5.25 g of Compound 2 as an orange oil which was used without further purification in the next step. Calculated mass for C.sub.16H.sub.27NO.sub.8 (M+H).sup.+ 362.23, found by LC-MS 362.2.

[0172] Preparation of Compound 3 in FIG. 18. Compound 3 is tert-butyl-(1S,2R)-2-[(benzylthio)-carbonyl)-3-tert-butoxycarbonyl)-1-met- oxhycarbonyl)]propylcarbamate. Compound 2 (2.1 g, 5.8 mmol) was dissolved with DCM (7 mL). To this clear mixture was added ethylthiol (0.4 g, 1.1 eq) and 4-dimethylaminopyridine (DMAP) (0.07 g, 0.1 eq). To the clear solution was added dicyclohexyl carbodiimide (DCC) (1.3 g, 1.1 eq) as a solid and the mixture was stirred at room temperature for 5 hours. The mixture was diluted with DCM (40 mL) and washed with 1N HCl (2.times.), dried over Na.sub.2SO.sub.4, filtered and concentrated to give 2.15 g of Compound 3 as a yellow oil, which was used without further purification in the next step. Calculated mass for C.sub.18H.sub.31NO.sub.7S (M+H).sup.+ 406.13, found by LC-MS 406.2.

[0173] Preparation of Compound 4 in FIG. 18. Compound 4 is tert-butyl-(1S,2R)-3-(tert-butoxcarbonyl)-1-(methoxycarbonyl)-2-formylpro- pylcarbamate. To Compound 3 (5.9 g, 14.5 mmol) and Pd--C 10% wt (0.35 g) were added acetone (36 mL) and the mixture cooled down to about 4-8.degree. C. To this mixture was added drop wise, under positive flow of Argon, triethylsilyl (TES) (11.5 mL, 5 eq). After addition of TES the mixture was kept at 10-15.degree. C. After 3.5 hours LCMS showed no more starting material. The mixture was filtrated through celite, and the solution concentrated to give 6.6 g of Compound 4 as a green oil which was used without further purification in the next step. Calculated mass for C.sub.16H.sub.27NO.sub.7 (M+H).sup.+ 346.13, found by LC-MS 346.2.

[0174] Preparation of Compound 5 in FIG. 18. Compound 5 is methyl-[3S,4R,8R]-1-Aza-3-tert-butoxycarbonyl-4-(tert-butoxycarbonyl)meth- yl)-2-oxo-6-thiabicyclic[3.3.0]-octane-8-carboxylate. To Compound 4 (1.3 g, 3.76 mmol), L-Cys-OH--HCl (0.98 g, 1.5 eq) and 4.degree. A molecular sieves (2g) was added dry pyridine (10 mL) and the mixture stirred, under argon, at room temperature for 4 hours in high pressure vessel. After this time, 3.5 mL more of pyridine were added and the reaction was stirred at 50.degree. C. for 5 days. After this time, the crude mixture was filtered through celite, the solution was re-dissolved in AcOEt and washed with 2N HCl 2.times., dried over Na.sub.2SO.sub.4, filtered and concentrated to yield 1.2 g of Compound 5 as a yellowish semisolid, which was used without further purification in the next step. Calculated mass for C.sub.19H.sub.30N.sub.2O.sub.7S (M+H).sup.+ 431.13, found by LC-MS 431.2.

[0175] Preparation of Compound 6 in FIG. 18. Compound 6 is [3S,4R,8R]-1-Aza-3-amino-4-(tert-butoxycarbonyl)methyl)-2-oxo-6-thiabicyc- lic[3.3.0]-octane-8-carboxylate. Crude Compound 5 was treated with 10 mL of a 50% TFA-DCM mixture at 0.degree. C. The mixture was let to warm up at 10.degree. C. and stirred for 2.5 hours. The mixture is then concentrated and the residue re-dissolved with 5 mL of a 2M LiOH solution and stirred at room temperature for 2.5 hours. LCMS analysis showed complete transformation to Compound 6. The reaction was concentrated and the residue passed through a short column packed with ion-exchange resin (H.sup.+, Dowex), eluting with 1:1 MeOH/H.sub.2O. The residue was concentrated and then lyophilized to give 1.8 g of crude Compound 6 as a yellow semisolid, which was used without further purification in the next step. Calculated mass for C.sub.13H.sub.20N.sub.2O.sub.5S (M+H).sup.+ 317.13, found by LC-MS 317.2.

[0176] Preparation of Compound 7 in FIG. 18. Compound 7 is the (5,5)-Glu-Gly bicylic dipeptide mimetic. Compound 7 is also referred to as [3S,4R,8R]-1-Aza-3-fluorenylmethyl-carbonyl-4-(tert-butoxycarbonyl)met- hyl)-2-oxo-6-thiabicyclic[3.3.0]-octane-8-carboxylate. Crude Compound 6 (1.2 g, 3.8 mmol) was dissolved in 17 mL of dry DCM. To this solution was added FmocOSu (1.8 g, 1.4 eq) followed by N,N-diisopropylethylamine (DIEA) (1.3 mL, 2 eq). The reaction was stirred at room temperature for 3 hours. To the mixture was added 50 mL of DCM and washed with 2M HCl 2.times., brine 1.times., dried over Na.sub.2SO.sub.4, filtered and concentrated to give 1.42 g of a yellow oil. Purification by flash chromatography using an increasing polarity solvent gradient (Hex:AcOEt 1:1 to DCM:MeOH 9:1) gave 0.150 g of pure Compound 7. Calculated mass for C.sub.28H.sub.30N.sub.2O.sub.7S (M+H).sup.+ 539.13, found by LC-MS 539.2.

Example 9

Preparation of Compound 11 in FIG. 19

[0177] Compound 9 is 1-Aza-3-amino-tert-butoxylcarbonyl-4-(tert-butoxycarbonyl)methyl)-2-oxo-1- -methoxyacetyl. Compound 4 was prepared as described above. A solution of Compound 4 (preparation described in Example 8) (4.1 g, 12 mmol) and the HCl salt of NH.sub.2-Gly-OMe (1.66 g, 1.1 eq) in 20 mL of DMF were stirred at room temperature. To this mixture was added NaBH(OAc).sub.3 (5.0 g, 2 eq) dissolved in 18 mL of DMF. After 1 hour LCMS shows complete transformation to the desired secondary amine 8. Calculated mass for C.sub.19H.sub.34N.sub.2O.sub.8 (M+H).sup.+ 419.1, found by LC-MS 419.2. To this crude mixture was added AcOEt (80 mL), washed with sat. NaHCO.sub.3 3.times., water 1.times., brine 1.times., dried over Na.sub.2SO.sub.4, filtered and concentrated to give 3.9 g of a crude yellow oil. LCMS of this material showed that the five-member ring lactam (Compound 9) was formed during work-up. The residue was concentrated to give 3.9 g of crude Compound 9 as a yellow oil, which was used without further purification in the next step. Calculated mass for C.sub.18H.sub.30N.sub.2O.sub.7 (M+H).sup.+ 387.1, found by LC-MS 387.2.

[0178] Preparation of Compound 10 in FIG. 19. Compound 10 is 1-amino-4-(tert-butoxy-carbonyl)methyl)-2-oxo-1-methoxyacetyl. Crude Compound 9 (3.8 g) was cooled at 10.degree. C. in an ice-water bath. To this stirred mixture was added in a drop wise manner 15 mL of a 50% solution of TFA in DCM. The mixture was stirred at that temperature for 2 h. LCMS showed a selective N-Boc deprotection. The mixture was concentrated to yield crude compound 10 as clear semisolid, which was used without further purification in the next step. Calculated mass for C.sub.13H.sub.22N.sub.2O.sub.5 (M+H).sup.+ 287.1, found by LC-MS 287.2.

[0179] Preparation of Compound 11 in FIG. 19. Compound 11 is a .gamma.-lactam-Glu-Gly bicyclic dipeptide mimetic which is 1-Aza-3-aminofluorenylmethylcarbonyl-4-(tert-butoxycarbonyl)methyl)-2-oxo- -1-acetylcarboxylate. Crude Compound 10 (2.8 g, 9.8 mmol) was dissolved in 8 mL of a 1:1 mixture of dioxane/MeOH at room temperature. To this mixture was added 15 mL of a 2M LiOH (2.8 eq) and the mixture stirred at room temperature for 3 hours. The mixture was then concentrated and passed trough a short column of Dowex H.sup.+ ion-exchange resin. The pooled fractions containing M+1=273 by LCMS were collected and lyophilized. This crude material was then dissolved in DCM (40 mL) followed by addition of DIEA (3.5 mL, 2 eq) and FmocOSu (3.9 g, 1.2 eq). The mixture was stirred at room temperature for 3 hours. LCMS analysis shows no more starting material, thus to the reaction was added AcOEt (60 mL), washed with sat. NaHCO.sub.3 3.times., water 1.times., brine 1.times., dried over Na.sub.2SO.sub.4, filtered and concentrated to give 2.5 g of a crude Compound 11. The mixture was purified by flash chromatography using AcOEt:Hex (1:1). Collection of the pure fractions identified by LCMS gave 100 mg of pure Compound 11. Calculated mass for C.sub.27H.sub.30N.sub.2O.sub.7 (M+H).sup.+ 495.1, found by LC-MS 495.2.

Example 10

Preparation of Compound 18 in FIG. 20

[0180] Compound 1 (i.e., tert-butyl-1(1S,2R)-2-[(benzyloxy)carbonyl)-3-tert-butoxycarbonyl)-1-meto- xhycarbonyl)]propylcarbamate) was prepared as described in Example 8 above.

[0181] Preparation of Compound 12 in FIG. 20. Compound 12 is tert-Butyl-(1S,2R)-2-[(benzylcarboxylate)-3-carboxy)-1-metoxhycarbonyl)]p- ropylcarbamate. To Compound 1 (13.9 g, 30.8 mmol) was added 45 mL of a 2M solution of HCl in diethyl ether. The clear yellow solution was stirred at room temperature for 18 hours. The mixture was triturated with cold ether which gave a yellow foam. Drying of this solid gave 8.32 g (84% yield) of the hydrochloride salt intermediate which was used without further purification in the next step. This HCl salt crude (8.3 g, 25 mmol) was dissolved in THF (80 mL). To this clear solution was added TEA (7.6 mL, 2.2 eq) followed by Boc.sub.2O (6.5 g, 1.2 eq). The mixture was stirred for 18 hours at room temperature. The mixture was then concentrated, re-dissolved in AcOEt (150 mL), washed with 1N HCl 2.times., sat. NaCl 1.times., dried over Na.sub.2SO.sub.4, filtered, concentrated and dried under high vacuum to yield Compound 12 as a brownish oil (11.1 g, 100% crude yield), which was used without further purification in the next step. Calculated mass for C.sub.19H.sub.25NO.sub.8 (M+H).sup.+ 396.1, found by LC-MS 396.2.

[0182] Preparation of Compound 13 in FIG. 20. Compound 13 is tert-Butyl-(1S,2R)-2-[(benzylcarboxylate)-3-ethylthiocarboxylate)-1-metox- hycarbonyl)]propylcarbamate. DCC (5.9 g, 1.1 eq) was added to a solution of crude Compound 12 (10.4 g, 26.2 mmol), EtSH (2.15 mL, 1.1 eq) and 4-dimethylaminopyridine (DMAP) (0.32 g, 0.1 eq) dissolved in DCM (28 mL). After 6 hours the mixture was concentrated, re-dissolved in AcOEt (150 mL) washed with 1N HCl 2.times., sat. NaCl 1.times., dried over Na.sub.2SO.sub.4, filtered, concentrated and dried under high vacuum to yield Compound 13 as a brown oil (7.6 g), which was used without further purification in the next step. Calculated mass for C.sub.21H.sub.29NO.sub.7S (M+H).sup.+ 440.1, found by LC-MS 440.2.

[0183] Preparation of Compound 14 in FIG. 20. Compound 14 is (1S,2R)-2-[(benzyl-carboxylate)-3-ethylthiocarboxylate)-1-metoxhycarbonyl- )]propylamine hydrochloride salt. Crude Compound 13 (7.6 g, 17.2 mmol) was dissolved in ethyl ether (8 mL), followed by addition of a 2N solution of HCl in diethyl ether (40 mL). The clear mixture was stirred at room temperature for 18 hours. The mixture was concentrated to give Compound 14 as an orange oil (7.5 g) which was used without further purification in the next step. Calculated mass for C.sub.16H.sub.21NO.sub.5S (M+H).sup.+ 338.1, found by LC-MS 338.2.

[0184] Preparation of Compound 15 in FIG. 20. Compound 15 is (1S,2R)-2-[(benzyl-carboxylate)-3-ethylthiocarboxylate)-1-metoxhycarbonyl- )]propyl-L-N-Boc-Ala. Crude Compound 14 (6.6 g, 17.7 mmol), Boc-L-Ala-OH (3.7 g, 1.1 eq) and 2-(7-aza-1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluoride (HATU) (12.1 g, 1.8 eq) were dissolved in ACN (150 mL). To this solution was added DIEA (6.1 mL, 2 eq) and the mixture was stirred at room temperature for 6 hours. The mixture was concentrated, re-dissolved in AcOEt (150 mL), washed with sat NH.sub.4Cl 2.times., sat. NaCl 1.times., dried over Na.sub.2SO.sub.4, filtered, concentrated and dried under high vacuum to yield compound 15 as a brown oil (16.5 g of crude). Purification by flash chromatography using Hex; AcOEt (6:4) gave pure compound 15 (3.55 g, 40% yield). Calculated mass for C.sub.24H.sub.34N.sub.2O.sub.8S (M+H).sup.+ 511.1, found by LC-MS 511.2.

[0185] Preparation of Compound 16 in FIG. 20. Compound 16 is (1S,2R)-2-[(benzyl-carboxylate)-3-formyl)-1-metoxhycarbonyl)]propyl-L-N-B- oc-Ala. Triethylsilane (5.6 mL, 5 eq) was slowly added, under argon, to a solution of Compound 15 (3.5 g, 6.9 mmol) and Pd--C (0.6 g) dissolved in acetone (16 mL) in an ice bath (.about.4.degree. C.). After addition of TES the reaction was stirred at 10-20.degree. C. After 4 hours, thin layer chromatography (TLC) showed complete transformation. The reaction was filtered and concentrated to give an off-green oil. Purification by flash chromatography using Hex; AcOEt (6:4) gave pure Compound 16 (2.9 g, 93% yield). Calculated mass for C.sub.22H.sub.30N.sub.2O.sub.8 (M+H).sup.+ 452.1, found by LC-MS 452.2.

[0186] Preparation of Compound 17 in FIG. 20. Compound 17 is (3R,6S,7R)-7-(benzyloxy-carbonyl)-6-amino-(L-Boc-Ala)-hexahydro-5-oxo-2H-- thiazolo[3,2-a]pyridine-3-carboxylic acid. In a high pressure vial crude Compound 16 (2 g, 4.55 mmol) was dissolved in dry pyridine (12 mL). To this mixture was added L-Cys-OH (0.88 g, 1.6 eq) followed by addition of 2.7 g of activated 4.degree. A molecular sieves (powder). The mixture was stirred vigorously at room temperature for 4 hours. Dry-pyridine (10 mL) was added to the mixture, the vial was capped and the mixture stirred at 50.degree. C. for 4 days. The mixture was then filtered through a bed of celite and washed with tetrahydrofuran (THF) and MeOH. The mixture was concentrated to give Compound 17 as yellow foam (1.6 g) which was used without further purification in the next step. Calculated mass for C.sub.24H.sub.31N.sub.3O.sub.8S (M+H).sup.+ 522.1, found by LC-MS 522.2.

[0187] Preparation of Compound 18 in FIG. 20. Compound 18 is (3R,6S,7R)-7-(benzyloxy-carbonyl)-6-amino-(L-Ala)-hexahydro-5-oxo-2H-thia- zolo[3,2-a]pyridine-3-carboxylic acid. In a round bottom flask crude Compound 17 (1.6 g, 3.1 mmol) was dissolved in dioxane (15 mL) followed by addition of 4M HCl. The mixture was stirred at room temperature and after 3 hours LCMS analysis showed complete conversion to the free amine. The mixture was concentrated, dissolved with DCM, MeOH, concentrated again and then dried under high vacuum to give the hydrochloride salt as a brownish semisolid (1.5 g) which was used without further purification in the next step. Calculated mass for C.sub.24H.sub.31N.sub.3O.sub.8S (M+H).sup.+ 522.1, found by LC-MS 522.2.

[0188] This HCl salt was dissolved in DCM (14 mL) followed by addition of DIEA (2 mL, 3.5 eq) and FmocOSu (1.2 g, 1.1 eq) in portions as solid. The mixture was stirred for 3 hours showing complete transformation by LCMS. The mixture was concentrated, dissolved in AcOEt (60 mL), washed with 2N HCl 2.times., brine, dried over Na.sub.2SO.sub.4, filtered and concentrated to give 2.1 g of a brownish oil. Purification by flash chromatography using Hex; AcOEt (5:95) followed by 100% MeOH gave pure compound 18 (0.821 g, 44% yield). Calculated mass for C.sub.34H.sub.33N.sub.3O.sub.8S (M+H).sup.+ 644.1, found by LC-MS 644.2.

Example 11

Preparation of Compounds in FIG. 21

[0189] Preparation of compound 16 has been described in Example 10.

[0190] Preparation of Compound 19 in FIG. 21. Compound 19 is (1S,2R)-[2-(benzylcarboxylate)-3-formyl-4-(tert-butoxy-N-2-methyl acetyl)-1-(methoxycarbonyl)]butyl-L-N-Boc-Ala. Crude compound 16 (1.01 g, 2.24 mmol) and L-Ala-OtBu. HCl (0.449 g, 1.1 eq) were dissolved in DMF (4.5 mL) at room temperature. NaBH(OAc).sub.3 (0.95 g, 2 eq) was dissolved separately in DMF (4.5 mL) at room temperature, the two solutions were mixed and stirred at room temperature for about 2 hours. The reaction was washed with saturated NaHCO.sub.3 solution 2.times., water 1.times., sat. NaCl 1.times., dried over Na.sub.2SO.sub.4, filtered, concentrated to yield 0.92 g of Compound 19 as a colorless oil, which was used without further purification in the next step. Calculated mass for C.sub.29H.sub.45N.sub.3O.sub.9 (M+H).sup.+ 579.68, found by LC-MS 579.6.

[0191] Preparation of Compound 20 in FIG. 21. Compound 20 is (1S,2R)-tert-butyl-[4-(benzylcarboxylate)-3-(N-L-Boc-Ala)-2-oxopiperidine- )-1-methyl-1-yl]acetate. Crude Compound 19 (0.550 g, 0.95 mmol) was dissolved in DMF (3 mL), DIEA (0.445 mL, 2 eq) was heated using microwaves at 140.degree. C. for 20 min. The reaction was checked and then washed with 0.5M HCl 2.times., water 1.times., sat. NaCl 1.times., dried over Na.sub.2SO.sub.4, filtered, concentrated to give 0.50 g of Compound 20 as a light brownish oil, which was used without further purification in the next step. Calculated mass for C.sub.28H.sub.41N.sub.3O.sub.8 (M+H).sup.+ 547.64, found by LC-MS 547.6.

[0192] Preparation of Compound 21 in FIG. 21. Compound 21 is (1S,2R)-[4-(benzyl-carboxylate)-3-(N-L-Fmoc-Ala)-2-oxopiperidine)-1-methy- l-1-yl]acetic acid. Crude Compound 20 (0.501 g, 0.91 mmol) was dissolved in 5 mL of 20% TFA in DCM and stirred for about an hour at room temperature. The reaction was concentrated and then dried under high vacuum to give the corresponding TFA salt as brownish oil (0.47 g) which was used without further purification in the next step. Calculated mass for C.sub.19H.sub.25N.sub.3O.sub.6 (M+H).sup.+ 505.64, found by LC-MS 505.6

[0193] This TFA salt (0.47 g, 0.91 mmol) was dissolved in aqueous 10% Na.sub.2CO.sub.3 solution (12 mL) and DMF (4 mL), and cooled down to 0.degree. C. A solution of FmocOSu (0.472 g, 1.5 eq) in DMF (8 mL) was added dropwise to the cold aqueous solution. After 5 minutes the ice bath was removed and the reaction was stirred overnight, quenched by adding 40 mL water followed by washings with AcOEt. The aqueous layer was acidified using 2N HCl (pH=2) and washed with AcOEt 3.times., NaCl 1.times., dried over Na.sub.2SO.sub.4, filtered and concentrated. Purification was done by washing the crude compound with Hex:AcOEt (6:4) 2.times. at room temperature to give Compound 21 as light colored oil (0.290 g, 52% yield). Calculated mass for C.sub.34H.sub.35N.sub.3O.sub.8 (M+H).sup.+ 613.24, found by LC-MS 613.2.

[0194] Preparation of Compound 22 in FIG. 21. Compound 22 is (1S,2R)-[2-(benzylcarboxylate)-3-(formyl)-4-(tert-butoxy-N-acetyl)-1-(met- hoxycarbonyl)]butyl-L-N-Boc-Ala. Crude compound 16 (1.01 g, 2.24 mmol) and AcOH. NH.sub.2-Gly-OtBu (0.478 g, 1.1 eq) were dissolved in dry DMF (4.5 mL) at room temperature. NaBH(OAc).sub.3 (0.95 g, 2 eq) was dissolved separately in dry DMF (4.5 mL) at room temperature, the two solutions were mixed and stirred at room temperature for about 2 hours. The reaction was washed with saturated NaHCO.sub.3 solution 2.times., water 1.times., sat. NaCl 1.times., dried over Na.sub.2SO.sub.4, filtered and concentrated to yield 1.09 g of Compound 22 as a colorless oil, which was used without further purification in the next step. Calculated mass for C.sub.28H.sub.43N.sub.3O.sub.9 (M+H).sup.+ 565.68, found by LC-MS 565.3.

[0195] Preparation of Compound 23 in FIG. 21. Compound 23 is (1S,2R)-tert-butyl-[4-(benzylcarboxylate)-3-(N-L-Boc-Ala)-2-oxopiperidine- )-1-yl]acetate. Crude compound 22 (0.550 g, 0.95 mmol) was dissolved in DMF (3 mL), DIEA (0.445 mL, 2 eq) and then heated using microwaves at 140.degree. C. for 20 min (Biotage.TM., Initiator8). The reaction was checked by LCMS. The reaction crude was washed with 0.5M HCl 2.times., water 1.times., sat. NaCl 1.times., dried over Na.sub.2SO.sub.4, filtered and concentrated to yield 0.41 g of Compound 23 as a light brownish oil, which was used without further purification in the next step. Calculated mass for C.sub.27H.sub.39N.sub.3O.sub.8 (M+H).sup.+ 533.64, found by LC-MS 533.6.

[0196] Preparation of Compound 24 in FIG. 21. Compound 24 is (1S,2R)-[4-(benzylcarboxylate)-3-(N-L-Fmoc-Ala)-2-oxopiperidine)-1-yl]ace- tic acid. Crude Compound 23 (0.41 g, 0.76 mmol) was dissolved in 5 mL of 20% TFA in DCM; reaction was stirred for about an hour at room temperature. The reaction was concentrated and then dried under high vacuum to give the corresponding TFA salt as a brownish oil (0.45 g) which was used without further purification in the next step. Calculated mass for C.sub.18H.sub.23N.sub.3O.sub.6 (M+H).sup.+ 491.3, found by LC-MS 491.4

[0197] This TFA salt (0.45 g, 0.92 mmol) was dissolved in aqueous 10% Na.sub.2CO.sub.3 solution (12 mL) and DMF (4 mL), and cooled down to 0.degree. C. A solution of FmocOSu (0.472 g, 1.5 eq) in DMF (8 mL) was added dropwise to the cold aqueous solution. After 5 min, the ice bath was removed and the reaction was stirred overnight, then quenched by adding about 40 mL water, followed by washings with AcOEt. The aqueous layer was acidified using 2N HCl (pH=2) and washed with AcOEt 3.times., NaCl 1.times., dried over Na.sub.2SO.sub.4, filtered, concentrated. Purification was done by washing the crude compound with Hex:AcOEt (6:4) 2.times. at room temperature giving Compound 24 as light colored oil (0.15 g, 27% yield). Calculated mass for C.sub.33H.sub.33N.sub.3O.sub.8 (M+H).sup.+ 599.6, found by LC-MS 599.4.

Example 12

Preparation of Compounds in FIGS. 1A, 2A, and 5A

[0198] The compound in FIG. 1A may be prepared following the methods described, e.g., in U.S. Pat. No. 6,872,700, the disclosure of which is incorporated by reference. The compounds in FIGS. 2A and 5A may be prepared following the methods described, e.g., in WO 2007/139941, the disclosure of which is incorporated by reference.

Example 13

Preparation of Compounds in FIGS. 1B-C, 2B-U, 3A-G, 5B-G, 6A-E, 10A-I, 11A-C, 12, 13A-B, 14B-C, 14E-R

[0199] For each compound in FIGS. 1B, 1C, 2B, 2C, 2D, 2E, 2F, 2G, 2H, 2I, 2J, 2K, 3A, 3B, 3C, 3D, 3E, 3F, 3G, 5B, 5C, 5D, 6A, 6B, 6C, 6D, and 6E, a calculated 100 .mu.mol of Fmoc-Ser(OtBu)-Wang resin or Fmoc Rink amide resin was weighed into a reaction vessel in a Symphony peptide synthesizer, and peptide elongation was carried out following standard Fmoc peptide synthesis protocol. If an unnatural amino acid was used at position 2 this was incorporated manually in a polypropylene syringe. Cleavage of the peptide from the resin was done with 10 mL TFA/H.sub.2O/PhOH/TIPS (95:2:2:1), then precipitated by methyl-tert-Butyl ether. The crude was dissolved and applied to a reverse-phase HPLC column (C.sub.18, 20-50% CH.sub.3CN in 0.1% TFA/H.sub.2O over 30 min gradient) to afford the titled compounds as white powders. All characterized by LC-MS. The compounds in FIGS. 2L-U, 5E-G, 10A-I, 13B, 14B-C and 14E-K will be prepared following the methods described in this example.

[0200] With respect to the compounds in FIGS. 14L-R, a calculated 100 .mu.mol of Fmoc-Rink-amide resin was weighed into a reaction vessel in a Symphony peptide synthesizer, and peptide elongation was carried out following standard Fmoc peptide synthesis protocol. If an unnatural amino acid was used at position 2 this was incorporated manually in a polypropylene syringe. Only for the compounds in FIGS. 14N-O, this procedure was carried out under the following microwave-assisted conditions: A microwave vial was loaded with the corresponding peptide-resin intermediate (0.3 g, 0.14 mmol), Fmoc-AA-OH (4 eq), PyBrop (0.32 g, 4.8 eq), 2,6-lutidine (0.25 mL, 15 eq), dichloroethane (DCE) (2-3 mL) and DMF (about 0.3 mL). The vial was capped and heated with microwaves (Biotage.TM., Initiator8) at 100.degree. C. for 11 min. The resin was filtered, washed with DCE and MeOH. Cleavage of the peptide from the resin was done with 10 mL TFA/H.sub.2O/PhOH/TIPS (95:2:2:1), then precipitated by methyl-tert-Butyl ether. The crude was dissolved and applied to a reverse-phase HPLC column (C.sub.18, 20-50% CH.sub.3CN in 0.1% TFA/H.sub.2O over 30 min gradient) to afford the titled compounds as white powders. All characterized by LC-MS.

Example 14

Preparation of Compounds in FIGS. 8A-H

[0201] For the compounds in FIGS. 8B, 8C, 8D, 8E, 8F, 8G, and 8H, a calculated 100 mol of Fmoc-Ser(OtBu)-Wang or Fmoc-Rink-amide resin was weighed into a reaction vessel in a Symphony peptide synthesizer, and the peptide elongation was carried out following standard Fmoc peptide synthesis protocol. The corresponding L- or D-Cys(Trt)-OH was introduced at positions 2 and 4. Cleavage of the peptide from the resin with 10 mL TFA/H.sub.2O/PhOH/TIPS (95:2:2:1) then precipitated by methyl-tert-Butyl ether. The crude peptides were dried overnight under high vacuum.

[0202] Disulfide cylclization: Clear-OX resin (0.39 g, 3.times. molar excess) was swollen on DCM for 45 min at room temperature, then washed with DCM 2.times., DMF 3.times., MeOH 3.times., deionized water 3.times. and finally H.sub.2O:ACN (1:1) 3.times.. The corresponding crude peptide (0.1 g) was dissolved in degassed 1:1 v/v solution of 0.1M NH.sub.4OAc buffer (pH=6.5)/ACN. The peptide solution was then added to the pre-swollen Clear-OX resin and the slurry was shacked at room temperature. After 2-3 hours the cyclization was complete. The resin was washed with a small amount of ACN/H.sub.2O (1:1) solution, the filtrate concentrated to remove volatiles and then lyophilized. The crude was dissolved and applied to a reverse-phase HPLC column (C.sub.18, 20-50% CH.sub.3CN in 0.1% TFA/H.sub.2O over 30 min gradient) to afford the titled compounds as white powders. All characterized by LC-MS. The compound in FIG. 8A will be prepared following the methods described in this example.

Example 15

Preparation of Compounds in FIGS. 9A-H

[0203] For each compound in FIGS. 9A, 9B, 9C, 9D, 9E, 9G, and 9H, a calculated 100 .mu.mol of Fmoc-Rink amide resin was weighed into a reaction vessel in a Symphony peptide synthesizer, and the peptide elongation was carried out following standard Fmoc peptide synthesis protocol. The amino acid at position 2 of all these compounds was introduced by special coupling conditions. A microwave vial was loaded with the corresponding peptide-resin intermediate (0.3 g, 0.14 mmol), Fmoc-AA-OH (4 eq), PyBrop (0.32 g, 4.8 eq), 2,6-lutidine (0.25 mL, 15 eq), dichloroethane (DCE) (2-3 mL) and DMF (-0.3 mL). The vial was capped and heated with microwaves (Biotage.TM., Initiator8) at 100.degree. C. for 11 min. The resin was filtered, washed with DCE and MeOH. For some compounds the above process was repeated twice. The resin is then treated in a cycle of deprotection, coupling with Fmoc-His(Trt)-OH, HBTU and HOBt, deprotection. Cleavage of the peptide from the resin was done with 10 mL TFA/H.sub.2O/PhOH/TIPS (95:2:2:1), then precipitated by methyl-tert-Butyl ether. The crude was dissolved and applied to a reverse-phase HPLC column (C.sub.18, 20-50% CH.sub.3CN in 0.1% TFA/H.sub.2O over 30 min gradient) to afford the titled compounds as white powders. All characterized by LC-MS. The compound in FIG. 9F will be prepared following the methods described in this example.

Example 16

In Vitro Assay

[0204] The compounds shown in Table 1 below were analyzed in an in vitro functional assay at the GLP-1 receptor. The assay was conducted as follows: Membrane fractions were prepared from confluent cultures of RIN m5f cells. Compounds were serially diluted with an assay buffer, and then added to a 96-well assay plate containing RIN m5f cell membranes in an ATP/GTP mixture. Cyclase activities were determined by measuring the production of cAMP induced through GLP-1 receptor activation. Quantification of cAMP production was achieved through a competitive chemiluminescence assay with a biotinylated-cAMP probe using Perkin Elmer Fusion.TM.-Alpha Microplate Analyzer (AlphaScreen.TM. technology). The compound EC.sub.50 values were obtained through fitting the concentration-response curves to a four-parameter logistic equation within GraphPad PRISM.RTM. software. The results of the assay are presented in Table 1 below.

Example 17

In Vivo Assay

[0205] Some compounds shown in Table 1 below were analyzed in an in vivo basal glucose lowering assay using the following procedure: A subcutaneous injection of either 200 .mu.l phosphate-buffered saline (PBS) vehicle or test article was given immediately following baseline glucose (t=0) to NIH/Swiss female mice. Tail blood glucose samples were measured at t=2 and 4 hours post dose using a OneTouch.RTM. Ultra.RTM. (LifeScan, Inc., a Johnson & Johnson Company, Milpitas, Calif.). Body weight was measured daily. Significant test sample effects were identified by ANOVA (p<0.05), followed by Dunnett's post test using GraphPad Prism version 4.00 for Windows (GraphPad Software, San Diego Calif.). The results are presented in Table 1 below.

TABLE-US-00004 TABLE 1 Compound in Cyclase GLP-1 Receptor In Vivo FIG. EC.sub.50 (nM) Glucose-Lowering Assay 1A 0.01 not tested 1B 0.004 not tested 1C 0.029 not tested 2A 0.008 greater than 3 hours 2B 0.0089 more than 4 hours 2C 6 not tested 2D 0.04 not tested 2E 2.15 not tested 2F 0.027 more than 4 hours 2G 0.89 not tested 2H 0.124 not tested 2I 0.054 more than 4 hours 2J 7.2 not tested 2K 0.29 similar to GLP-1 3A 0.007 more than 4 hours 3B 0.007 more than 2 hours 3C 0.022 not tested 3D 1.03 not tested 3E 0.06 more than 2 hours 3F 1.089 not tested 3G 0.065 similar to GLP-1 5A 1.83 more than 4 hours 5B 0.53 more than 4 hours 5C 1.056 not tested 5D 10.5 not tested 6A 0.144 similar to GLP-1 6B 0.52 not tested 6C 151.9 not tested 6D 0.282 not tested 6E 0.60 more than 4 hours 8B 1.26 similar to GLP-1 8C 968.6 not tested 8D 106.2 not tested 9A 0.21 not tested 9B 0.03 more than 4 hours 9C 0.05 more than 4 hours 9D 0.01 more than 4 hours 9E 133 not tested 9F 0.07 more than 4 hours 9G 1.9 not tested 9H 0.1 not tested 11A 0.155 not tested 11B 0.007 not tested 11C 0.011 not tested 12 0.028 not tested 13 0.274 not tested 14J 0.054 not tested 14K 3.5 not tested 14L 0.05 not tested 14M 0.2 (partial agonist) not tested 14N 0.004 not tested 14O 0.24 not tested 14P 0.25 not tested 14Q 0.02 not tested 14R 0.41 (partial agonist) not tested 15A 0.57 inactive 15B 1000 not tested 15C 2.46 inactive 15D 1.7 inactive 16A 0.52 inactive 16B 0.18 similar to GLP-1 16C 1.1 inactive 16D 1 inactive 16E 1 inactive 16F 0.61 about 30 minutes 16G 56 not tested 16H 1000 not tested 17A 105 not tested 17B 668 not tested 17C 10,000 not tested

Example 18

Preparation of Compounds in FIGS. 17A-F

[0206] A calculated 100 .mu.mol of Rink amide resin was weighed into a reaction vessel in a Symphony peptide synthesizer, and the peptide elongation was carried out following standard Fmoc peptide synthesis protocol up to residue Pro.sup.3. The resulting peptide-resin intermediate (0.3 g, 0.0927 mmol) was swollen in DCM with around 5% DMF and to the slurry was added Fmoc-Cys(Trt)-OH (0.351 g, 4 eq), followed by Bromo-tris-pyrrolidino-phosphonium hexafluorophosphate (PyBrop) (0.34 g, 4.8 eq) and 2,6-lutidine (0.27 mL, 15 eq). The vial was capped and heated in a microwave apparatus (Biotage Initiator8) at 100.degree. C. for 11 min. The resin was filtered and washed with DMF 6.times., treated with 20% piperidine in DMF 2.times.25 min and washed with DMF 6.times., DCM 4.times. followed by treatment with 2% TFA/1.5% TIS in DCM 4.times.10 min. then washed with DCM 4.times., DMF 2.times. and TMOF 3.times.. In a polypropylene syringe the resin is swollen in TMOF, followed by addition of compound 25 (0.2 g, 3 eq) and dry pyridine (0.06 mL, 5 eq). The resin was shaken at room temperature for 16 hours, followed by washings with TMOF 3.times., DCM 3.times., MeOH 3.times. and dried under high vacuum. Cleavage of the peptide from the resin was carried out with 10 ml TFA/H.sub.2O/PhOH/TIPS (95:2:2:1) (TFA is trifluoroacetic acid; TIPS is triisopropylsilyl), precipitated by methyl-tert-Butyl ether. The residue was dissolved in 0.8 ml of MeOH and 0.8 ml of ACN. To this solution 0.1 ml of DEA were added and the mixture stirred at room temperature overnight. The resulting residue was applied to a reverse-phase high performance liquid chromatography (HPLC) column (C.sub.18, 20-50% CH.sub.3CN in 0.1% TFA/H.sub.2O over 30 min gradient) to afford compound 17A as a white powder (5.3 mg, 3%): Retention time in reverse phase-high performance liquid chromatography (RP-HPLC) (C.sub.18, 5-75% CH.sub.3CN in 0.1% TFA/H.sub.2O over 15 min) is 8.71 min; Calculated mass for C.sub.146H.sub.228N.sub.38O.sub.43S (M+H).sup.+ 3235.74, found by liquid chromatography/mass spectrometry (LC-MS) 1079.6 (M+3H).sup.3+, 1619.9 (M+2H).sup.2+.

[0207] The synthesis of compounds 17B and 17C was performed similar as reported above for compound 17A, with the difference of using Fmoc-D-Cys(Trt)-OH and Fmoc-Pen-OH, respectively. After purification compound 17B was obtained as a white powder (8 mg, 7%): Retention time in reverse phase-high performance liquid chromatography (RP-HPLC) (C.sub.18, 5-75% CH.sub.3CN in 0.1% TFA/H.sub.2O over 15 min) is 8.55 min; Calculated mass for C.sub.146H.sub.228N.sub.38O.sub.43S (M+H).sup.+ 3235.74, found by liquid chromatography/mass spectrometry (LC-MS) 1079.6 (M+3H).sup.3.alpha., 1619.9 (M+2H).sup.2+. After purification compound 17C was obtained as a white powder (1.5 mg, 3%): Retention time in reverse phase-high performance liquid chromatography (RP-HPLC) (C.sub.18, 5-75% CH.sub.3CN in 0.1% TFA/H.sub.2O over 15 min) is 8.55 min; Calculated mass for C.sub.148H.sub.232N.sub.38O.sub.43S (M+H).sup.+ 3263.79, found by liquid chromatography/mass spectrometry (LC-MS) 1089.6 (M+3H).sup.3+, 1632.9 (M+2H).sup.2+.

Example 19

Preparation of Compound 25

[0208] In a round bottom flask Fmoc-His(Boc)-OH (4 g, 1 eq) was dissolved in DCM (50 mL). To this solution was added DCC (1.9 g, 1.1 eq), EtSH (0.7 mL, 1.1 eq) and DMAP (0.102 g, 0.1 eq). The mixture was stirred at room temperature for 6 h. The reaction mixture was filtered and the solution concentrated to yield 4.9 g of an off-white solid. The crude product was purified by flash chromatography using Hex:AcOEt (1:1). After concentration of the pure fractions the corresponding thioester was obtained as a clear oil (3.6 g, 84%). The thioester (0.69 g, 1 eq) was dissolved in THF (11 mL) and stirred under Argon atmosphere for few minutes. To this solution was added Pd--C (0.180 g) and the mixture stirred under Argon for 10 min followed by dropwise addition of TES (0.75 mL, 3.5 eq) and the mixture stirred at room temperature. After 4 h one more equivalent of TES was added, one hour later the crude mixture was filtered through a bed of silica/celite and washed with THF. The filtrate was concentrated to give a dark brown oil, which was purified by flash chromatography in a short (20 mL) silica gel column using Hex:AcOEt (2:8). After concentration of the pure fractions compound 25 was obtained as a clear semisolid (0.26 g, 50%).

Example 20

Modified Exendin Peptides

[0209] N-Terminus conformationally constrained GLP-1 receptor agonist compounds described herein were covalently linked to one or more polyethylene glycol and/or fatty acids, as described herein. In particular, the following twelve compounds 23A-L were prepared:

[0210] Compound 23A: 4-imidazopropionyl-dAla-PGTFTSDLSK.sup.12QMEEEAVRLFIE-WLKNGGPSSGAPPPS-NH.- sub.2, wherein K.sup.12 was modified with --C(.dbd.O)--CH.sub.2(OCH.sub.2CH.sub.2).sub.2NH--C(.dbd.O)CH.sub.2--(OCH- .sub.2CH.sub.2).sub.2NH--C(.dbd.O)--(CH.sub.2).sub.6--CH.sub.3 (SEQ ID NO:105).

[0211] Compound 23B: 4-imidazopropionyl-dAla-PGTFTSDLSK.sup.12QMEEEAVRLFIE-WLKNGGPSSGAPPPS-NH.- sub.2, wherein K.sup.12 was modified with --C(.dbd.O)--CH.sub.2(OCH.sub.2CH.sub.2).sub.2NH--C(.dbd.O)CH.sub.2(OCH.s- ub.2CH.sub.2).sub.2NH--C(.dbd.O)--CH(NH.sub.2)--(CH.sub.2).sub.7--CH.sub.3- (SEQ ID NO:106).

[0212] Compound 23C: 4-imidazopropionyl-dAla-PGTFTSDLSK.sup.12QMEEEAVRLFIE-WLKNGGPSSGAPPPS-NH.- sub.2, wherein K.sup.12 was modified with --C(.dbd.O)--CH.sub.2(OCH.sub.2CH.sub.2).sub.2NH--C(.dbd.O)CH.sub.2(OCH.s- ub.2CH.sub.2).sub.2NH--C(.dbd.O)--CH[NH--C(.dbd.O)(CH.sub.2).sub.6CH.sub.3- ]-(CH.sub.2).sub.7--CH.sub.3(SEQ ID NO:107).

[0213] Compound 23D: 4-imidazopropionyl-dAla-PGTFTSDLSK.sup.12QMEEEAVRLFIE-WLKNGGPSSGAPPPS-NH.- sub.2, wherein K.sup.12 was modified with --C(.dbd.O)--CH.sub.2(OCH.sub.2CH.sub.2).sub.2NH--C(.dbd.O)CH.sub.2(OCH.s- ub.2CH.sub.2).sub.2NH--C(.dbd.O)--CH.sub.2CH.sub.2--CH[C(OH)(.dbd.O)]--NH-- -C(.dbd.O)--(CH.sub.2).sub.16-[C(OH)(.dbd.O)] (SEQ ID NO:108).

[0214] Compound 23E: 4-imidazopropionyl-dAla-PGTFTSDLSKQMEEEAVRLFIEW-LK.sup.27NGGPSSGAPPPS-NH.- sub.2, wherein K.sup.27 was modified with --C(.dbd.O)--CH.sub.2(OCH.sub.2CH.sub.2).sub.2NH--C(.dbd.O)CH.sub.2(OCH.s- ub.2CH.sub.2).sub.2NH--C(.dbd.O)--(CH.sub.2).sub.6--CH.sub.3 (SEQ ID NO:109).

[0215] Compound 23F: 4-imidazopropionyl-dAla-PGTFTSDLSKQMEEEAVRLFIEW-LK.sup.27NGGPSSGAPPPS-NH.- sub.2, wherein K.sup.27 was modified with --C(.dbd.O)--CH.sub.2(OCH.sub.2CH.sub.2).sub.2NH--C(.dbd.O)CH.sub.2(OCH.s- ub.2CH.sub.2).sub.2NH--C(.dbd.O)--CH(NH.sub.2)--(CH.sub.2).sub.7--CH.sub.3 (SEQ ID NO:110).

[0216] Compound 23G: 4-imidazopropionyl-dAla-PGTFTSDLSKQMEEEAVRLFIEW-LK.sup.27NGGPSSGAPPPS-NH.- sub.2, wherein K.sup.27 was modified with --C(.dbd.O)--CH.sub.2(OCH.sub.2CH.sub.2).sub.2NH--C(.dbd.O)CH.sub.2(OCH.s- ub.2CH.sub.2).sub.2NH--C(.dbd.O)--CH[NH--C(.dbd.O)(CH.sub.2).sub.6CH.sub.3- ]-(CH.sub.2).sub.7--CH.sub.3 (SEQ ID NO:111).

[0217] Compound 23H: 4-imidazopropionyl-dAla-PGTFTSDLSKQMEEEAVRLFIEW-LK.sup.27NGGPSSGAPPPS-NH.- sub.2, wherein K.sup.27 was modified with --C(.dbd.O)--CH.sub.2(OCH.sub.2CH.sub.2).sub.2NH--C(.dbd.O)CH.sub.2(OCH.s- ub.2CH.sub.2).sub.2NH--C(.dbd.O)--CH.sub.2CH.sub.2--CH[C(OH)(.dbd.O)]--NH-- -C(.dbd.O)--(CH.sub.2).sub.16-[C(OH)(.dbd.O)](SEQ ID NO:112).

[0218] Compound 23I: 4-imidazopropionyl-dAla-PGTFTSDLSKQMEEEAVRLFIEWLKN-GGPSSGAPPPS.sup.39, wherein S.sup.39 was modified with -Lys(NH.sub.2)--C(.dbd.O)--CH.sub.2(OCH.sub.2CH.sub.2).sub.2NH--C(.dbd.O)- CH.sub.2(OCH.sub.2CH.sub.2).sub.2NH--C(.dbd.O)--(CH.sub.2).sub.6--CH.sub.3 (SEQ ID NO:113).

[0219] Compound 23J: 4-imidazopropionyl-dAla-PGTFTSDLSKQMEEEAVRLFIEWLKN-GGPSSGAPPPS.sup.39, wherein S.sup.39 was modified with -Lys(NH.sub.2)--C(.dbd.O)--CH.sub.2(OCH.sub.2CH.sub.2).sub.2NH--C(.dbd.O)- CH.sub.2(OCH.sub.2CH.sub.2).sub.2NH--C(.dbd.O)--CH(NH.sub.2)--(CH.sub.2).s- ub.7--CH.sub.3 (SEQ ID NO:114).

[0220] Compound 23K: 4-imidazopropionyl-dAla-PGTFTSDLSKQMEEEAVRLFIEWLKN-GGPSSGAPPPS.sup.39, wherein S.sup.39 was modified with -Lys(NH.sub.2)--C(.dbd.O)--CH.sub.2(OCH.sub.2CH.sub.2).sub.2NH--C(.dbd.O)- CH.sub.2(OCH.sub.2CH.sub.2).sub.2NH--C(.dbd.O)--CH[NH--C(.dbd.O)(CH.sub.2)- .sub.6CH.sub.3]-(CH.sub.2).sub.7--CH.sub.3 (SEQ ID NO:115).

[0221] Compound 23L: 4-imidazopropionyl-dAla-PGTFTSDLSKQMEEEAVRLFIEWLKN-GGPSSGAPPPS.sup.39, wherein S.sup.39 was modified with --C(.dbd.O)--CH.sub.2(OCH.sub.2CH.sub.2).sub.2NH--C(.dbd.O)CH.sub.2(OCH.s- ub.2CH.sub.2).sub.2NH--C(.dbd.O)--CH.sub.2CH.sub.2--CH[C(OH)(.dbd.O)]--NH-- -C(.dbd.O)--(CH.sub.2).sub.16-[C(OH)(.dbd.O)] (SEQ ID NO:116).

[0222] For each of Compound Nos. 23A-L above, a calculated 100 .mu.mol of Fmoc-Rink-amide resin was weighed into a reaction vessel in a Symphony peptide synthesizer, and peptide elongation was carried out following standard Fmoc peptide synthesis protocol. The dAla at position 2 was incorporated manually in a polypropylene syringe. The orthogonal protection (alloc group) of the side chain group (Lys.sup.12, Lys.sup.27 or Lys.sup.40) was performed as follows: The peptide-resin was swollen in DCM and dimethylamino borane-complex (6 eq) followed after about 3 minutes by tetrakis(triphenylphosphine)palladium(0) (0.1 eq). The resin was shaken for 15 minutes, washed with DCM 3.times. and the process repeated. Then, the resin was washed with DCM 3.times., 10% DIEA in DCM 2.times., DCM 3.times. and MeOH 2.times.. At this point, the peptide-resin gave a positive chloranil test. The resulting peptide-resin intermediate (0.3 g, 0.0927 mmol) was swollen in DMF and to the slurry was added the corresponding polyethylene glycol (2.2 eq), followed by HBTU (0.035 g, 2.2 eq), HOBt (0.03 g, 2.2 eq) and NMM (0.04 mL, 4.4 eq). After 3 hours, the resin was washed with DMF 6.times., treated with 20% piperidine in DMF 2.times.25 min and washed with DMF 6.times.. The above cycle was repeated with a second Fmoc-(polyethylene glycol)-OH in most cases. The coupling of the corresponding fatty acid chain was done using two different microwave-assisted methods: the corresponding acyl chloride (5 eq) and NMM (7 eq) were added to the resin swollen in a 1:1 DMF:DCM mixture, and then heated using microwaves at 75.degree. C. for 20 min (Biotage.TM., Initiator8); or the corresponding carboxylic or dicarboxylic acid (5 eq), HOAt (5 eq) and DIC (5 eq) were added to the slurry resin on DMF, and then heated using microwaves at 75.degree. C. for 15 min (Biotage.TM., Initiator8). Cleavage of the peptide from the resin was done with 10 mL TFA/H.sub.2O/PhOH/TIPS (95:2:2:1), then precipitated by methyl-tert-Butyl ether. The crude was dissolved and applied to a reverse-phase HPLC column (C.sub.18, 20-50% CH.sub.3CN in 0.1% TFA/H.sub.2O over 30 min gradient) to afford the titled compounds as white powders. All were characterized by LC-MS.

[0223] Each of these compounds was analyzed in an in vitro functional assay at the GLP-1 receptor following the methods described in Example 16. The results are in Table 2:

TABLE-US-00005 TABLE 2 Cyclase GLP-1 Receptor Compound EC.sub.50 (nM) 23A 0.027 23B 0.027 23C 0.06 23D 0.22 23E 0.3 23F 0.1 23G 0.7 23H 0.08 23I 0.01 23J 0.01 23K 0.02 23L 0.185

Example 21

Modified Exendin Peptides

[0224] N-Terminus conformationally constrained GLP-1 receptor agonist compounds described herein were covalently linked to one or more biotin, as described herein. In particular, the following compounds 24A-L were prepared:

TABLE-US-00006 Compound 24A: (SEQ ID NO: 1117) His-dAla-PGTFTSDLSKQMEEEAVRLFIEWL-Lys(biotin)- NGGPSSGAPPS-Lys[(NH.sub.2)(biotin)]. Compound 24B: (SEQ ID NO: 69) 4-imidazopropionyl-GEGTFTSDLSKQMEEEAVRLFIEWL- Lys(biotin)-NGGPSSGAPPS-Lys[(NH.sub.2)(biotin)]. Compound 24C: (SEQ ID NO: 70) 4-imidazopropionyl-GEGTFTSDLSKQMEEEAVRLFIEWLKN- Lys[(NH.sub.2)(biotin)]. Compound 24D: (SEQ ID NO: 71) 4-imidazopropionyl-APGTFTSDLSKQMEEEAVRLFIEWLKN- Lys[(NH.sub.2)(biotin)]. Compound 24E: (SEQ ID NO: 118) His-dAla-PGTFTSDLSKQMEEEAVRLFIEWLKNGGPSSGAPPPS- Lys(NH2)-(O-CH.sub.2-CH.sub.2).sub.2-KAKAEAEAKAKAEAEA-biotin. Compound 24F: (SEQ ID NO: 119) His-dAla-PGTFTSDLSKQMEEEAVRLFIEWLE[-(O-CH.sub.2-CH.sub.2).sub.2- (biotin)]-NGGPSSGAPPPS-NH.sub.2. Compound 24G: (SEQ ID NO: 120) His-dAla-PGTFTSDLSEK[-(O-CH.sub.2-CH.sub.2).sub.2-(biotin)]- QMEEEAVRLFIEWLKNGGPSSGAPPPS-NH.sub.2.

[0225] For each of Compound Nos. 24A-G above, a calculated 100 .mu.mol of Fmoc-Rink-amide resin was weighed into a reaction vessel in a Symphony peptide synthesizer, and peptide elongation was carried out following standard Fmoc peptide synthesis protocol. If an unnatural amino acid was used at position 2 this was incorporated manually in a polypropylene syringe. The orthogonal protection (alloc group) of the side chain group (Lys.sup.27, Lys.sup.28 or Lys.sup.40) was performed as follows: The peptide-resin was swollen in DCM and dimethylamino borane-complex (6 eq) followed after about 3 minutes by tetrakis(triphenylphosphine)palladium(0) (0.1 eq). The resin was shaken for 15 min, washed with DCM 3.times. and the process repeated. Then, the resin was washed with DCM 3.times., 10% DIEA in DCM 2.times., DCM 3.times. and MeOH 2.times.. At this point, the peptide-resin gave a positive chloranil test. The biotin moiety (one or two) was coupled to the free amino group using the standard solid-phase coupling conditions described above (HBTU, HOBt, NMM). Cleavage of the peptide from the resin was done with 10 mL TFA/H.sub.2O/PhOH/TIPS (95:2:2:1), then precipitated by methyl-tert-Butyl ether. The crude was dissolved and applied to a reverse-phase HPLC column (C.sub.18, 20-50% CH.sub.3CN in 0.1% TFA/H.sub.2O over 30 min gradient) to afford the titled compounds as white powders. All were characterized by LC-MS.

[0226] Each of these compounds was analyzed in an in vitro functional assay at the GLP-1 receptor following the methods described in Example 16. The results are in Table 3:

TABLE-US-00007 TABLE 3 Cyclase GLP-1 Receptor Compound EC.sub.50 (nM) 24A 0.04 24B 0.03 24C 0.07 24D 0.77 24E 0.06 24F 0.06 24G 0.01

Example 22

Compounds of FIG. 10

[0227] With respect to FIG. 10, the following compounds 10J-10N were made:

TABLE-US-00008 10J: (SEQ ID NO: 72) 4-imidazopropionyl-GPGTFTSDLSKQLEEEAVRLFIEWLKNGGPSSGAPPPS-NH.sub.2 10K: (SEQ ID NO: 121) 4-imidazopropionyl-dAla-PGTFTSDLSKQLEEEAVRLFIEWLKNGGPSSGAPPPS-NH.sub.2. 10L: (SEQ ID NO: 73) 4-imidazopropionyl-Aib-PGTFTSDLSKQLEEEAVRLFIEWLKNGGPSSGAPPPS-NH.sub.2 10M: (SEQ ID NO: 74) 4-imidazopropionyl-GEGTFTSDLSKQLEEEAVRLFIEWLKQGGPSKEIIS-OH 10N: (SEQ ID NO: 122) 4-imidazopropionyl-dAla-PGTFTSDLSKQLEEEAVRLFIEWLKQGGPSKEIIS-OH.

[0228] For each of Compounds 10J-N, a calculated 100 .mu.mol of Fmoc-Rink-amide resin was weighed into a reaction vessel in a Symphony peptide synthesizer, and peptide elongation was carried out following standard Fmoc peptide synthesis protocol. If an unnatural amino acid was used at position 2 this was incorporated manually in a polypropylene syringe. Only for the compound in FIG. 10L, this procedure was carried out under the following microwave-assisted conditions: a microwave vial was loaded with the corresponding peptide-resin intermediate (0.3 g, 0.14 mmol), Fmoc-AA-OH (4 eq), PyBrop (0.32 g, 4.8 eq), 2,6-lutidine (0.25 mL, 15 eq), dichloroethane (DCE) (2-3 mL) and DMF (.about.0.3 mL). The vial was capped and heated with microwaves (Biotage.TM., Initiator8) at 100.degree. C. for 11 min. The resin was filtered, washed with DCE and MeOH. Cleavage of the peptides from the resins was done with 10 mL TFA/H.sub.2O/PhOH/TIPS (95:2:2:1), then precipitated by methyl-tert-Butyl ether. The crude was dissolved and applied to a reverse-phase HPLC column (C.sub.18, 20-50% CH.sub.3CN in 0.1% TFA/H.sub.2O over 30 min gradient) to afford the titled compounds as white powders. All characterized by LC-MS.

[0229] Compounds 10J-L were analyzed in an in vitro functional assay at the GLP-1 receptor following the methods described in Example 16. The results are in Table 4:

TABLE-US-00009 TABLE 4 Cyclase GLP-1 Receptor Compound EC.sub.50 (nM) 10J 0.1 10K 0.03 10L 0.005 10M 0.007 10N 0.01

[0230] All publications and patent applications are incorporated herein by reference. Although the foregoing has been described in detail for purposes of clarity of understanding, it will be apparent to one of ordinary skill in the art that changes and modifications may be made without departing from the spirit or scope of the disclosure or appended claims.

Sequence CWU 1

1

122128PRTArtificial SequenceFormula (B); Description of Artificial Sequence Synthetic polypeptide 1Xaa Xaa Xaa Xaa Thr Phe Thr Ser Asp Leu Ser Lys Gln Xaa Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Xaa Leu Lys Asn 20 25 228PRTArtificial SequenceFormula (C); Description of Artificial Sequence Synthetic polypeptide 2Xaa Xaa Xaa Xaa Thr Phe Thr Ser Asp Leu Ser Lys Gln Xaa Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Xaa Leu Lys Xaa 20 25 339PRTArtificial SequenceFormula (D); Description of Artificial Sequence Synthetic polypeptide 3Xaa Xaa Xaa Xaa Thr Phe Thr Ser Asp Leu Ser Lys Gln Xaa Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Xaa Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 431PRTArtificial SequenceFormula (E); Description of Artificial Sequence Synthetic polypeptide 4Xaa Xaa Xaa Xaa Thr Phe Thr Ser Asp Val Ser Ser Tyr Xaa Glu Gly 1 5 10 15 Gln Ala Ala Lys Glu Phe Ile Ala Xaa Leu Val Xaa Gly Arg Xaa 20 25 30 534PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 5Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu Ala Val 1 5 10 15 Arg Leu Phe Ile Glu Trp Leu Lys Gln Gly Gly Pro Ser Lys Glu Ile 20 25 30 Ile Ser 625PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 6Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu Ala Val 1 5 10 15 Arg Leu Phe Ile Glu Phe Leu Lys Asn 20 25 727PRTArtificial SequenceR4; Description of Artificial Sequence Synthetic polypeptide; see specification as filed for detailed description of substitutions and preferred embodiments 7Xaa Xaa Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu 1 5 10 15 Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn 20 25 838PRTArtificial SequenceR4; Description of Artificial Sequence Synthetic polypeptide; see specification as filed for detailed description of substitutions and preferred embodiments 8Xaa Xaa Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu 1 5 10 15 Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser Ser 20 25 30 Gly Ala Pro Pro Pro Ser 35 936PRTArtificial SequenceR4; Description of Artificial Sequence Synthetic polypeptide; see specification as filed for detailed description of substitutions and preferred embodiments 9Xaa Xaa Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu 1 5 10 15 Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Gln Gly Gly Pro Ser Lys 20 25 30 Glu Ile Ile Ser 35 1038PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 10Gly Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu 1 5 10 15 Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser Ser 20 25 30 Gly Ala Pro Pro Pro Ser 35 1136PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 11Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu Ala Val 1 5 10 15 Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser Ser Gly Ala 20 25 30 Pro Pro Pro Ser 35 1237PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 12Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu Ala 1 5 10 15 Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser Ser Gly 20 25 30 Ala Pro Pro Pro Ser 35 1332PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 13Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu Ala Val Arg Leu 1 5 10 15 Phe Ile Glu Trp Leu Lys Gln Gly Gly Pro Ser Lys Glu Ile Ile Ser 20 25 30 1434PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 14Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu Ala Val Arg Leu 1 5 10 15 Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser Ser Gly Ala Pro Pro 20 25 30 Pro Ser 1526PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 15Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly Gln Ala Ala Lys Glu 1 5 10 15 Phe Ile Ala Trp Leu Val Lys Gly Arg Gly 20 25 1639PRTHeloderma horridum 16His Ser Asp Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 174PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 17Gly Glu Gly Thr 1 1824PRTArtificial SequenceZ; Description of Artificial Sequence Synthetic peptide; see specification as filed for detailed description and preferred embodiments 18Thr Phe Thr Ser Asp Leu Ser Lys Gln Xaa Glu Glu Glu Ala Val Arg 1 5 10 15 Leu Phe Ile Glu Xaa Leu Lys Asn 20 1935PRTArtificial SequenceZ; Description of Artificial Sequence Synthetic peptide; see specification as filed for detailed description and preferred embodiments 19Thr Phe Thr Ser Asp Leu Ser Lys Gln Xaa Glu Glu Glu Ala Val Arg 1 5 10 15 Leu Phe Ile Glu Xaa Leu Lys Asn Gly Gly Pro Ser Ser Gly Ala Pro 20 25 30 Pro Pro Ser 35 2024PRTArtificial SequenceZ; Description of Artificial Sequence Synthetic peptide; see specification as filed for detailed description and preferred embodiments 20Thr Phe Thr Ser Asp Leu Ser Lys Gln Xaa Glu Glu Glu Ala Val Arg 1 5 10 15 Leu Phe Ile Glu Xaa Leu Lys Xaa 20 2127PRTArtificial SequenceZ; Description of Artificial Sequence Synthetic peptide; see specification as filed for detailed description and preferred embodiments 21Thr Phe Thr Ser Asp Val Ser Ser Tyr Xaa Glu Gly Gln Ala Ala Lys 1 5 10 15 Glu Phe Ile Ala Xaa Leu Val Xaa Gly Arg Xaa 20 25 2210PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 22Gln Gly Gly Pro Ser Lys Glu Ile Ile Ser 1 5 10 2312PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 23Gln Gly Gly Pro Ser Ser Gly Ala Pro Pro Pro Ser 1 5 10 244PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 24Asn Gly Gly Pro 1 255PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 25Asn Gly Gly Pro Ser 1 5 266PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 26Asn Gly Gly Pro Ser Ser 1 5 277PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 27Asn Gly Gly Pro Ser Ser Gly 1 5 288PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 28Asn Gly Gly Pro Ser Ser Gly Ala 1 5 299PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 29Asn Gly Gly Pro Ser Ser Gly Ala Pro 1 5 3010PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 30Asn Gly Gly Pro Ser Ser Gly Ala Pro Pro 1 5 10 3111PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 31Asn Gly Gly Pro Ser Ser Gly Ala Pro Pro Pro 1 5 10 3211PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 32Asn Gly Gly Pro Ser Ser Gly Ala Pro Pro Ser 1 5 10 3312PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 33Asn Gly Gly Pro Ser Ser Gly Ala Pro Pro Ser Lys 1 5 10 3417PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 34Asn Gly Gly Pro Ser Ser Gly Ala Pro Pro Ser Lys Lys Lys Lys Lys 1 5 10 15 Lys 3513PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 35Asn Gly Gly Pro Ser Ser Gly Ala Pro Pro Pro Ser Lys 1 5 10 364PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 36His Gly Glu Gly 1 374PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 37His Ala Glu Gly 1 3839PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 38His Ala Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 3938PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 39Gly Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu Glu 1 5 10 15 Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser Ser 20 25 30 Gly Ala Pro Pro Pro Ser 35 4039PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 40His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 4139PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 41His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 4239PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 42His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 4328PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 43His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn 20 25 4428PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 44His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn 20 25 4528PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 45His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn 20 25 4636PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 46His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro 35 4736PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 47His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro 35 4836PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 48His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro 35 4937PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 49His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Gln Gly Gly Pro Ser 20 25 30 Lys Glu Ile Ile Ser 35 5039PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 50His Ser Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 5129PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 51His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly 20 25 5229PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 52His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn Gly 20 25 5330PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 53His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly 20 25 30 5430PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 54His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn Gly Gly 20 25 30 5531PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 55His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro 20 25 30 5631PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 56His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn Gly Gly Pro 20 25 30 5732PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 57His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 5832PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 58His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn Gly Gly Pro Ser 20 25 30 5933PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 59His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser

6033PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 60His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser 6134PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 61His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly 6234PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 62His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly 6335PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 63His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala 35 6435PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 64His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala 35 6537PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 65His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro 35 6637PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 66His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro 35 6738PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 67His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro 35 6838PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 68His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro 35 6939PRTArtificial SequenceCompound 24B; Description of Artificial Sequence Synthetic polypeptide 69Xaa Gly Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Ser Lys 35 7029PRTArtificial SequenceCompound 24C; Description of Artificial Sequence Synthetic peptide 70Xaa Gly Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Lys 20 25 7129PRTArtificial SequenceCompound 24D; Description of Artificial Sequence Synthetic peptide 71Xaa Ala Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Lys 20 25 7239PRTArtificial SequenceCompound 10J; Description of Artificial Sequence Synthetic polypeptide 72Xaa Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 7339PRTArtificial SequenceCompound 10L; Description of Artificial Sequence Synthetic polypeptide 73Xaa Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 7437PRTArtificial SequenceCompound 10M; Description of Artificial Sequence Synthetic polypeptide 74Xaa Gly Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Gln Gly Gly Pro Ser 20 25 30 Lys Glu Ile Ile Ser 35 7529PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 75His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly 20 25 7630PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 76His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly 20 25 30 7731PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 77His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro 20 25 30 7832PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 78His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 7933PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 79His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser 8034PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 80His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly 8135PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 81His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala 35 8237PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 82His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro 35 8338PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 83His Gly Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro 35 8431PRTHomo sapiens 84His Ala Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly 1 5 10 15 Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Arg Gly 20 25 30 8530PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 85His Ala Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly 1 5 10 15 Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Arg 20 25 30 8639PRTHeloderma suspectum 86His Gly Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 8737PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 87His Gly Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Gln Gly Gly Pro Ser 20 25 30 Lys Glu Ile Ile Ser 35 8839PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 88His Ala Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 8939PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 89His Gly Pro Ala Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 9039PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 90His Ala Pro Ala Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 9139PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 91His Val Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 9241PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 92His Ala Leu Ala Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met 1 5 10 15 Glu Glu Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly 20 25 30 Pro Ser Ser Gly Ala Pro Pro Pro Ser 35 40 9328PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 93His Gly Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn 20 25 9426PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 94Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu Ala 1 5 10 15 Val Arg Leu Phe Ile Glu Phe Leu Lys Asn 20 25 9528PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 95His Val Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn 20 25 9628PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 96His Ala Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn 20 25 9728PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 97His Cys Pro Cys Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn 20 25 9837PRTArtificial SequenceDescription of Artificial Sequence Synthetic polypeptide 98His Cys Pro Cys Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Gln Gly Gly Pro Ser 20 25 30 Lys Glu Ile Ile Ser 35 9928PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 99His Gly Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn 20 25 10024PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 100Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu Ala Val Arg 1 5 10 15 Leu Phe Ile Glu Phe Leu Lys Asn 20 10127PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 101Gly Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu 1 5 10 15 Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn 20 25 10225PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 102Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu Ala Val 1 5 10 15 Arg Leu Phe Ile Glu Trp Leu Lys Asn 20 25 1038PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 103Pro Gly Thr Phe Thr Ser Asp Leu 1 5 10422PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 104Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu Ala Val Arg Leu Phe 1 5 10 15 Ile Glu Phe Leu Lys Asn 20 10539PRTArtificial SequenceCompound 23A 105Xaa Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 10639PRTArtificial SequenceCompound 23B 106Xaa Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 10739PRTArtificial SequenceCompound 23C 107Xaa Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 10839PRTArtificial SequenceCompound 23D 108Xaa Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 10939PRTArtificial SequenceCompound 23E 109Xaa Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 11039PRTArtificial SequenceCompound 23F 110Xaa Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 11139PRTArtificial SequenceCompound 23G 111Xaa Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5

10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 11239PRTArtificial SequenceCompound 23G 112Xaa Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 11339PRTArtificial SequenceCompound 23I 113Xaa Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 11439PRTArtificial SequenceCompound 23J 114Xaa Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 11539PRTArtificial SequenceCompound 23K 115Xaa Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 11639PRTArtificial SequenceCompound 23L 116Xaa Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 11739PRTArtificial SequenceCompound 24A 117His Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Ser Lys 35 11839PRTArtificial SequenceCompound 24E 118His Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Ser Lys 35 11927PRTArtificial SequenceCompound 24F 119His Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Glu 20 25 12013PRTArtificial SequenceCompound 24G 120His Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Glu Lys 1 5 10 12139PRTArtificial SequenceCompound 10K 121Xaa Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser 20 25 30 Ser Gly Ala Pro Pro Pro Ser 35 12237PRTArtificial SequenceCompound 10N 122Xaa Xaa Pro Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu 1 5 10 15 Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Gln Gly Gly Pro Ser 20 25 30 Lys Glu Ile Ile Ser 35

Claims

1-130. (canceled)

131. A polypeptide comprising the amino acid sequence as set forth in Formula (C): TABLE-US-00010 (SEQ ID NO: 2) Xaa.sub.1Xaa.sub.2Xaa.sub.3Xaa.sub.4TFTSDLSKQXaa.sub.14EEEAVRLFIEXaa.sub.2- 5LK-R.sub.10-Z;

wherein: Xaa.sub.1 is His; or a compound of Formula (1); Xaa.sub.2 is Gly, dAla, Aib, Ala, Val, NMeAla, a compound of Formula (3), or a compound of Formula (4); and Xaa.sub.2 is absent when Xaa.sub.3 is: ##STR00021## Xaa.sub.3 is Pro; a compound of Formula (2); a compound of Formula (3); a Compound of Formula (4); ##STR00022## Xaa.sub.4 is Gly, dAla, or Aib; Xaa.sub.14 is Leu or Met; Xaa.sub.25 is Phe or Trp; R.sub.10 is QGGPSKEIIS (SEQ ID NO:22); NG; NGG; NGGP (SEQ ID NO:24); NGGPS (SEQ ID NO:25); NGGPSS (SEQ ID NO:26); NGGPSSG (SEQ ID NO:27); NGGPSSGA (SEQ ID NO:28); NGGPSSGAP (SEQ ID NO:29); NGGPSSGAPP (SEQ ID NO:30); NGGPSSGAPPP (SEQ ID NO:31); QGGPSSGAPPPS (SEQ ID NO:32); NGGPSSGAPPS (SEQ ID NO:33); NGGPSSGAPPSK (SEQ ID NO:34); NGGPSSGAPPS(K).sub.2-5 (SEQ ID NO:35); NGGPSSGAPPPSK (SEQ ID NO:36); or NK; and Z is OH or NH.sub.2; wherein the compound of Formula (1) is: ##STR00023## wherein R.sub.20 and R.sub.21 are each independently a single bond or a carbon atom; R.sub.23, R.sub.24, R.sub.25 and R.sub.26 are each independently absent, hydrogen, hydroxyl, C.sub.1-6 alkyl, carboxyl, amino, or C.sub.1-6 alkoxy; - - - - - - is a single bond or a double bond; and R.sub.21 is a chiral or achiral carbon atom; wherein the compound of Formula (2) is: ##STR00024## wherein Y.sub.1 and Z.sub.1 are each independently a single bond, a carbon, or a sulfur; and W.sub.1, W.sub.2 and W.sub.3 are each independently selected from hydrogen, C.sub.1-6 alkyl, C.sub.1-6 alkoxy, hydroxyl, and amino; and when one Y.sub.1 or Z.sub.1 is sulfur, the sulfur may be bonded to two oxygen atoms to form a sulfonyl group; and - - - - - - is or ; wherein the compound of Formula (3) is: ##STR00025## wherein Y.sub.1 and Z.sub.1 are each independently a single bond, a carbon, or a sulfur; and W.sub.1, W.sub.2 and W.sub.3 are each independently selected from hydrogen, C.sub.1-6 alkyl, C.sub.1-6 alkoxy, hydroxyl, and amino; and when one Y.sub.1 or Z.sub.1 is sulfur, the sulfur may be bonded to two oxygen atoms to form a sulfonyl group; and - - - - - - is or ; and wherein the compound of Formula (4) is: ##STR00026## wherein R.sub.30, R.sub.31, and R.sub.32 are each independently hydrogen or a C.sub.1-6 alkyl; or R.sub.30 and R.sub.31, together with the nitrogen.sup.1 and the carbon.sup.2, form a 5-membered or 6-membered heterocyclic ring; or R.sub.31 and R.sub.32, together with the carbon.sup.2, form a 3-, 4-, or 5-membered carbocyclic ring.

132. The polypeptide of claim 131, wherein Xaa.sub.3 is Pro.

133. The polypeptide of claim 132, wherein the polypeptide comprises the sequence set forth as: Pro.sup.3-exendin-4 (SEQ ID NO:40); Pro.sup.3,Leu.sup.14-exendin-4 (SEQ ID NO:41); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4 (SEQ ID NO:42); Pro.sup.3-exendin-4(1-28) (SEQ ID NO:43); Pro.sup.3,Leu.sup.14-exendin-4(1-28) (SEQ ID NO:44); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-28) (SEQ ID NO:45); Pro.sup.3-exendin-4(1-29) (SEQ ID NO:51); Pro.sup.3,Leu.sup.14-exendin-4(1-29) (SEQ ID NO:75); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-29) (SEQ ID NO:52); Pro.sup.3-exendin-4(1-30) (SEQ ID NO:53); Pro.sup.3,Leu.sup.14-exendin-4(1-30) (SEQ ID NO:76); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-30) (SEQ ID NO:54); Pro.sup.3-exendin-4(1-31) (SEQ ID NO:55); Pro.sup.3,Leu.sup.14-exendin-4(1-31) (SEQ ID NO:77); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-31) (SEQ ID NO:56); Pro.sup.3-exendin-4(1-32) (SEQ ID NO:57); Pro.sup.3,Leu.sup.14-exendin-4(1-32) (SEQ ID NO:78); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-32) (SEQ ID NO:58); Pro.sup.3-exendin-4(1-33) (SEQ ID NO:59); Pro.sup.3,Leu.sup.14-exendin-4(1-33) (SEQ ID NO:79); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-33) (SEQ ID NO:60); Pro.sup.3-exendin-4(1-34) (SEQ ID NO:61); Pro.sup.3,Leu.sup.14-exendin-4(1-34) (SEQ ID NO:80); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-34) (SEQ ID NO:62); Pro.sup.3-exendin-4(1-35) (SEQ ID NO:63); Pro.sup.3,Leu.sup.14-exendin-4(1-35) (SEQ ID NO:81); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-35) (SEQ ID NO:64); Pro.sup.3-exendin-4(1-36) (SEQ ID NO:46); Pro.sup.3,Leu.sup.14-exendin-4(1-36) (SEQ ID NO:47); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-36) (SEQ ID NO:48); Pro.sup.3-exendin-4(1-37) (SEQ ID NO:65); Pro.sup.3,Leu.sup.14-exendin-4(1-37) (SEQ ID NO:82); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-37) (SEQ ID NO:66); Pro.sup.3-exendin-4(1-38) (SEQ ID NO:67); Pro.sup.3,Leu.sup.14-exendin-4(1-38) (SEQ ID NO:83); Pro.sup.3,Leu.sup.14,Phe.sup.25-exendin-4(1-38) (SEQ ID NO:68); Pro.sup.3-exendin-3 (SEQ ID NO:50); or HGPGTFTSDLSKQLEEEAVRLFIEWLKQGGPSKEIIS (SEQ ID NO:49).

134. A method for treating diabetes, treating insulin resistance, treating postprandial hyperglycemia, lowering blood glucose levels, lowering HbA1c levels, stimulating insulin release, reducing gastric motility, delaying gastric emptying, reducing food intake, reducing appetite, reducing weight, treating overweight, or treating obesity in a patient in need thereof, the method comprising: administering to the patient a therapeutically effective amount of the peptide of claim 131 to treat diabetes, treat insulin resistance, treat postprandial hyperglycemia, lower blood glucose levels, lower HbA1c levels, stimulate insulin release, reduce gastric motility, delay gastric emptying, reduce food intake, reduce appetite, reduce weight, treat overweight, or treat obesity in the patient.