Methods for treating inflammatory bowel diseases with α4β7 integrin antagonists

Abstract

To provide a method for treating inflammatory bowel disease (IBD).SOLUTION: The present invention relates to methods of treating inflammatory bowel diseases, including with engineered peptides (e.g. peptide monomers and dimers comprising disulfide or thioether intramolecular bonds) that bind α4β7 integrin. In one aspect, the disclosure provides a method of treating an inflammatory bowel disease (IBD) in a subject in need thereof, comprising administering to the subject an α4β7 integrin antagonist, wherein the antagonist is administered to the patient orally at a dose of about 100 mg to about 500 mg, once or twice daily, wherein the antagonist is a peptide dimer compound comprising two peptides, or a pharmaceutically acceptable salt thereof.SELECTED DRAWING: Figure 22

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to U.S. Provisional Application No. 62 / 959,854, filed January 10, 2020, which is incorporated herein by reference in its entirety.

[0002] Sequence Listing This application was filed electronically via EFS-Web and includes an electronically submitted Sequence Listing in .txt format. The .txt file contains a Sequence Listing entitled "PRTH_052_01WO_ST25.txt," created on January 8, 2021, and having a size of approximately 7 kilobytes. The Sequence Listing contained in this .txt file is a part of the present specification and is incorporated herein by reference in its entirety.

[0003] FIELD OF THE INVENTION The present disclosure relates to methods of treating inflammatory bowel disease with engineered peptides (e.g., peptide monomers and dimers containing disulfide or thioether intramolecular bonds) that bind to α4β7 integrin. [Background technology]

[0004] Background of the Invention Integrins are non-covalently associated α / β heterodimeric cell surface receptors involved in numerous cellular processes ranging from cell adhesion and migration to gene regulation (Dubree, et al., Selective α4β7 Integrin Antagonist and Their Potential as Anti-inflammatory Agents, J. Med. Chem. 2002, 45, 3451-3457). Differential expression of integrins can regulate the adhesive properties of cells and recruit different leukocyte populations to specific organs in response to different inflammatory signals. If left untreated, integrin-mediated adhesion processes can lead to chronic inflammation and autoimmune diseases.

[0005] The α4 integrins, α4β1 and α4β7, play important roles in lymphocyte migration throughout the gastrointestinal tract. They are expressed on most leukocytes, including B and T lymphocytes, and mediate cell adhesion through binding to their respective primary ligands, vascular cell adhesion molecule (VCAM) and mucosal addressin cell adhesion molecule 1 (MAdCAM1), respectively. The proteins differ in binding specificity: VCAM binds to both α4β1 and, to a lesser extent, α4β7, while MAdCAM1 is highly specific for α4β7. In addition to pairing with the α4 subunit, the β7 subunit also forms a heterodimeric complex with the αE subunit to form α4β7, which is primarily expressed on intraepithelial lymphocytes (IELs) in the intestine, lung, and genitourinary tract. α4β7 is also expressed on dendritic cells in the intestine. The α4β7 heterodimer binds to E-cadherin on epithelial cells. IEL cells are thought to provide a mechanism for immune surveillance within the epithelial compartment. Therefore, blocking both α4β7 and α4β7 may be a useful method for treating intestinal inflammatory conditions.

[0006] Inhibitors of specific integrin-ligand interactions have been shown to be effective as anti-inflammatory agents for the treatment of various autoimmune diseases. For example, monoclonal antibodies exhibiting high binding affinity for α4β7 have shown therapeutic benefit in gastrointestinal autoinflammatory / autoimmune diseases (IDs), such as Crohn's disease and ulcerative colitis. However, these therapies interfere with the α4β1 integrin-ligand interaction, thereby resulting in dangerous side effects for patients. Therapies utilizing small molecule antagonists have shown similar side effects in animal models, hindering further development of these technologies. Recently, engineered peptides exhibiting high potency and stability, as well as high specificity for the α4β7 integrin, have been shown to be effective in the treatment of various immune disorders, including inflammatory bowel disease.

[0007] However, there is a need in the art for additional methods for using α4β7 antagonists and other agents to treat inflammatory disorders. Such methods are disclosed herein. [Prior art documents] [Non-patent literature]

[0008] [Non-Patent Document 1] Dubree, et al., Selective α4β7 Integrin Antagonist and Their Potential as Anti-inflammatory Agents, J.Med.Chem.2002,45,3451-3457 Summary of the Invention

[0009] The present disclosure provides compositions and methods for treating various diseases and conditions associated with α4β7 integrin signaling.

[0010] In one aspect, the disclosure provides a method of treating inflammatory bowel disease (IBD) in a subject in need thereof, comprising administering to the subject an α4β7 integrin antagonist, wherein the antagonist is orally administered to the patient once or twice daily at a dose of about 100 mg to about 500 mg, wherein the antagonist is a peptide dimeric compound comprising two peptides, or a pharmaceutically acceptable salt thereof, each of the two peptides having the sequence (optionally with an N-terminal Ac): 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH2 (SEQ ID NO: 5), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 6), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH2 (SEQ ID NO: 6), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 7), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-NH2 (SEQ ID NO: 7), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 8), or Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-NH2(SEQ ID NO: 8) wherein each of the two peptides comprises a thioether bond between 2-methylbenzoyl and Pen or a disulfide bond between the two Pens, and the two peptides are linked by a linker moiety attached to D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG).

[0011] In one embodiment, each of the two peptides has the sequence: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0012] In one embodiment, each of the two peptides has the sequence: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0013] In one embodiment, each of the two peptides has the sequence: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0014] In one embodiment, each of the two peptides has the sequence: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0015] In one embodiment, each of the two peptides has the sequence: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH (SEQ ID NO: 5), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0016] In one embodiment, each of the two peptides has the sequence: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0017] In one embodiment, the peptide dimer compound or a pharmaceutically acceptable salt thereof is TIFF2025186365000002.tif70132 or a pharmaceutically acceptable salt thereof.

[0018] In one embodiment, the peptide dimer compound or a pharmaceutically acceptable salt thereof is TIFF2025186365000003.tif81152 or a pharmaceutically acceptable salt thereof.

[0019] In one embodiment, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0020] In one embodiment, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0021] In one embodiment, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0022] In one embodiment, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0023] In one embodiment, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH (SEQ ID NO: 5); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0024] In one embodiment, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0025] Any of the peptides disclosed herein can include an N-terminal Ac.

[0026] In one embodiment, the peptide dimer compound or a pharmaceutically acceptable salt thereof is TIFF2025186365000004.tif70132 or a pharmaceutically acceptable salt thereof.

[0027] In one embodiment, the peptide dimer compound or a pharmaceutically acceptable salt thereof is TIFF2025186365000005.tif82153 or a pharmaceutically acceptable salt thereof.

[0028] In certain embodiments of the methods disclosed herein, the peptide dimeric compound or a pharmaceutically acceptable salt thereof is administered to a subject at a dose of about 100.0, 112.5, 125.0, 137.5, 150.0, 162.5, 175, 187.5, 200.0, 212.5, 225.0, 237.5, 250.0, 262.5, 275, 287.5, 300.0, 312.5, 325.0, 337.5, 350.0, 362.5, 375, 387.5, 400.0, 412.5, 425.0, 437.5, 450.0, 462.5, 475, 487.5, or 500.0 mg. In one embodiment, the peptide dimeric compound or its pharmaceutically acceptable salt is administered to a subject at a dose of about 150 mg or about 450 mg. In certain embodiments, the dose is administered to a subject twice a day.

[0029] In certain embodiments, the pharmaceutically acceptable salt of the peptide dimer compound is an acetate salt.

[0030] In certain embodiments of the methods disclosed herein, the administered dosage optionally results in an unsaturated blood receptor occupancy (RO%) as measured at peak blood or serum levels of the antagonist. In some embodiments, the administered dosage optionally results in an RO of less than 90%, less than 80%, less than 70%, less than 60%, or less than 50% as measured at peak blood or serum levels of the antagonist.

[0031] In certain embodiments of the methods disclosed herein, the methods inhibit MadCAM1-mediated T cell proliferation in the gastrointestinal tract.

[0032] In certain embodiments of the methods disclosed herein, the methods reduce cell surface expression of β7 on CD4+ T cells in the gastrointestinal tract.

[0033] In certain embodiments of the methods disclosed herein, the method comprises: i) induces internalization of α4β7 integrin on CD4+ T memory cells; ii) causes a reduction in adhesion of CD4+ T memory cells to MAdCAM1 in the gastrointestinal tract; and / or iii) inhibiting homing of T cells to the gastrointestinal tract, optionally to the ileal lamina propria, Peyer's patches, mesenteric lymph nodes, small intestine, and / or colon.

[0034] In certain embodiments of the methods disclosed herein, the IBD is ulcerative colitis.

[0035] In certain embodiments of the methods disclosed herein, the IBD is Crohn's disease.

[0036] In certain embodiments of the methods disclosed herein, the methods result in one or more of the following pharmacokinetic parameters in the plasma of the subject: Cmax (ng / mL) of 1 to 25; Tmax (hours) from 1 to 5 AUC between 10 and 250 t (ng.hour / mL) AUC between 10 and 300 inf (ng.h / mL), 3~10t 1 / 2 (time), AUC of 30 to 130 tau (ng.h / mL), Ctrough (ng / mL) of 1 to 5; an accumulated Cmax (ng.mL) of 0.5 to 2.5, and Accumulated AUC between 0.5 and 3.0 t (ng.h / mL).

[0037] In certain embodiments of the methods disclosed herein, the methods result in one or more of the following pharmacodynamic parameters in the plasma of the subject: ROmax (%) of 50 to 100, -20 to -60 receptor expression max Change in (%), Mean change (%) in receptor expression from -10 to -55; Steady-state ROmax (%) of 80-100; Average RO of 50-95 0-24 (h%), Average RO of 80-95 0-12 (h%), and Average RO of 70-90 12-24 (h%).

[0038] In another aspect, the present disclosure provides a method of treating an inflammatory disease or disorder in a subject in need thereof, comprising administering to the subject an α4β7 integrin antagonist, optionally at a dosage that results in unsaturated blood receptor occupancy (RO%) as measured at peak blood or serum levels of the antagonist. In certain embodiments, the antagonist is optionally administered at a dosage that results in a blood RO of less than 90%, less than 80%, less than 70%, less than 60%, or less than 50% as measured at peak blood or serum levels of the antagonist. In certain embodiments, the antagonist is present in a pharmaceutical composition formulated for a route of administration selected from oral, parenteral, subcutaneous, buccal, nasal, inhalation, topical, and rectal administration. In certain embodiments, the antagonist is administered orally or rectally.

[0039] In certain embodiments of any of the methods of the present disclosure, the inflammatory disease or disorder is selected from the group consisting of inflammatory bowel disease (IBD), adult IBD, pediatric IBD, adolescent IBD, ulcerative colitis, Crohn's disease, celiac disease (non-tropical sprue), seronegative arthropathy-related enteropathy, microscopic colitis, collagenous colitis, eosinophilic gastroenteritis, radiation therapy, chemotherapy, pouchitis occurring after proctocolectomy and ileoanal anastomosis, gastrointestinal cancer, pancreatitis, insulin-dependent diabetes mellitus, mastitis, cholecystitis, cholangitis, pericholecititis, chronic bronchitis, chronic sinusitis, asthma, primary sclerosing cholangitis, human immunodeficiency virus (HIV) infection in the GI tract, eosinophilic asthma, eosinophilic esophagitis, gastritis, colitis, microscopic colitis, and graft-versus-host disease (GVDH). In certain embodiments, the disease or disorder is an IBD, such as ulcerative colitis or Crohn's disease.

[0040] In certain embodiments, the antagonist is a peptide dimeric compound comprising two peptides, or a pharmaceutically acceptable salt thereof; Each of the two peptides has the sequence (optionally including an N-terminal Ac): 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH2 (SEQ ID NO: 5), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 6), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH2 (SEQ ID NO: 6), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 7), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-NH2 (SEQ ID NO: 7), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 8), or Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-NH2(SEQ ID NO: 8) wherein each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen or a disulfide bond between the two Pens, and the two peptides are linked by a linker moiety bonded to D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG).

[0041] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0042] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0043] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0044] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0045] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH (SEQ ID NO: 5); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0046] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0047] In certain embodiments, the peptide dimeric compound or a pharmaceutically acceptable salt thereof comprises: TIFF2025186365000006.tif70132 or a pharmaceutically acceptable salt thereof.

[0048] In certain embodiments, the peptide dimeric compound or a pharmaceutically acceptable salt thereof comprises: TIFF2025186365000007.tif81152 or a pharmaceutically acceptable salt thereof.

[0049] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0050] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0051] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0052] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0053] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH (SEQ ID NO: 5); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0054] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0055] In certain embodiments, the peptide dimeric compound or a pharmaceutically acceptable salt thereof comprises: TIFF2025186365000008.tif70132 or a pharmaceutically acceptable salt thereof.

[0056] In certain embodiments, the peptide dimeric compound or a pharmaceutically acceptable salt thereof comprises: TIFF2025186365000009.tif83156 or a pharmaceutically acceptable salt thereof.

[0057] In certain embodiments, the peptide dimeric compound or a pharmaceutically acceptable salt thereof has a molecular weight of about 5, 6, 7, 8, 9, 10, 12.5, 25.0, 37.5, 50.0, 62.5, 75, 87.5, 100.0, 112.5, 125.0, 137.5, 150.0, 162.5, 175, 187.5, 200.0, 212.5, 225 In certain embodiments, the subject is administered a dose of 0.0, 237.5, 250.0, 262.5, 275, 287.5, 300.0, 312.5, 325.0, 337.5, 350.0, 362.5, 375, 387.5, 400.0, 412.5, 425.0, 437.5, 450.0, 462.5, 475, 487.5, or 500.0 mg. In certain embodiments, the dose is administered to the subject once daily or twice daily.

[0058] In certain embodiments, the pharmaceutically acceptable salt of the peptide dimer compound is an acetate salt.

[0059] In a related aspect, the present disclosure provides a pharmaceutical composition comprising the peptide dimer compound disclosed in any one of claims 39 to 58 or a pharmaceutically acceptable salt thereof. In certain embodiments, the composition is formulated for oral delivery, and optionally, the composition comprises an enteric coating. In certain embodiments, the method comprises administering to a subject a pharmaceutical composition disclosed herein.

[0060] In certain embodiments of the methods and compositions disclosed herein, the antagonist or a pharmaceutically acceptable salt thereof inhibits binding of α4β7 integrin to MAdCAM1.

[0061] In certain embodiments of the methods and compositions disclosed herein, the antagonist or a pharmaceutically acceptable salt thereof or pharmaceutical composition is provided to a subject in need thereof at intervals sufficient to improve or ameliorate the condition. In certain embodiments, the intervals are selected from the group consisting of 24 consecutive hours, hourly, every 4 hours, once a day, twice a day, three times a day, four times a day, every other day, weekly, biweekly, and monthly. In certain embodiments, the antagonist or a pharmaceutically acceptable salt thereof or pharmaceutical composition is provided as an initial dose, followed by one or more subsequent doses, with the minimum interval between any two doses being less than one day, and each dose containing an effective amount of the antagonist. In some embodiments, the effective amount of the antagonist or a pharmaceutically acceptable salt thereof or pharmaceutical composition is sufficient to achieve at least one of the following: a) saturation of approximately 50% or more of the MAdCAM1 binding sites on the α4β7 integrin molecule; b) inhibiting α4β7 integrin expression on the cell surface by about 50% or more; and c) saturation of MAdCAM1 binding sites on the α4β7 molecule by about 50% or more and inhibition of α4β7 integrin expression on the cell surface by about 50% or more, where i) the saturation is maintained for a time period consistent with a dosing frequency of no more than twice daily, ii) the inhibition is maintained for a time period consistent with a dosing frequency of no more than twice daily, or iii) saturation and inhibition, each of which is maintained for a time period consistent with a dosing frequency of no more than twice daily. [The present invention 1001] 1. A method of treating inflammatory bowel disease (IBD) in a subject in need thereof, comprising administering to the subject an α4β7 integrin antagonist, wherein the antagonist is orally administered to the patient once or twice daily at a dose of about 100 mg to about 500 mg, wherein the antagonist is a peptide dimeric compound comprising two peptides, or a pharmaceutically acceptable salt thereof, each of the two peptides having the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH2 (SEQ ID NO: 5), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 6), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH2 (SEQ ID NO: 6), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 7), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-NH2 (SEQ ID NO: 7), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 8), or Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-NH2(SEQ ID NO: 8) wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen or a disulfide bond between the two Pens, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1002] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1) wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1003] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2) wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1004] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3) wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1005] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4) wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1006] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH2 (SEQ ID NO: 5) wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1007] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5) wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1008] The peptide dimer compound or a pharmaceutically acceptable salt thereof is TIFF2025186365000010.tif70132 or a pharmaceutically acceptable salt thereof. [The present invention 1009] The peptide dimer compound or a pharmaceutically acceptable salt thereof is TIFF2025186365000011.tif88164 or a pharmaceutically acceptable salt thereof. [The present invention 1010] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1) wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1011] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2) wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1020] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3) 1009. The method of claim 1009, wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1012] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4) wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1013] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH2 (SEQ ID NO: 5) wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1014] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5) wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1015] The peptide dimer compound or a pharmaceutically acceptable salt thereof is TIFF2025186365000012.tif70132 or a pharmaceutically acceptable salt thereof. [The present invention 1016] The peptide dimer compound or a pharmaceutically acceptable salt thereof is TIFF2025186365000013.tif88164 or a pharmaceutically acceptable salt thereof. [The present invention 1017] 1016. The method of any of claims 1001 to 1016, wherein the peptide dimer compound or a pharmaceutically acceptable salt thereof is administered to the subject at a dose of about 100.0, 112.5, 125.0, 137.5, 150.0, 162.5, 175, 187.5, 200.0, 212.5, 225.0, 237.5, 250.0, 262.5, 275, 287.5, 300.0, 312.5, 325.0, 337.5, 350.0, 362.5, 375, 387.5, 400.0, 412.5, 425.0, 437.5, 450.0, 462.5, 475, 487.5, or 500.0 mg. [The present invention 1018] 1017. The method of claim 1017, wherein said peptide dimer compound or a pharmaceutically acceptable salt thereof is administered to said subject at a dose of about 150 mg. [The present invention 1019] 1017. The method of claim 1017, wherein said peptide dimer compound or a pharmaceutically acceptable salt thereof is administered to said subject at a dose of about 450 mg. [The present invention 1020] 1019. The method of claim 1018 or 1019, wherein said dose is administered to said subject twice daily. [The present invention 1021] The method according to any one of claims 1001 to 1020, wherein said pharmaceutically acceptable salt of said peptide dimer compound is an acetate salt. [The present invention 1022] Any of the methods of claims 1001-1021, wherein the administered dosage optionally results in unsaturated blood receptor occupancy (RO%) as measured by peak blood or serum levels of said antagonist. [The present invention 1023] The method of claim 1022, wherein said administered dosage optionally results in an RO of less than 90%, less than 80%, less than 70%, less than 60%, or less than 50% as measured by peak blood or serum levels of said antagonist. [The present invention 1024] The method of any of claims 1001 to 1021, wherein the method inhibits MadCAM1-mediated T cell proliferation in the gastrointestinal tract. [The present invention 1025] The method of any of claims 1001 to 1021, wherein the cell surface expression of β7 on CD4+ T cells in the gastrointestinal tract is reduced. [The present invention 1026] i) induces internalization of α4β7 integrin on CD4+ T memory cells; ii) causes a reduction in adhesion of CD4+ T memory cells to MAdCAM1 in the gastrointestinal tract; and / or iii) inhibiting homing of T cells to the gastrointestinal tract, optionally the ileal lamina propria, Peyer's patches, mesenteric lymph nodes, small intestine, and / or colon; Any of the methods of 1001 to 1021 of the present invention. [The present invention 1027] The method according to any one of claims 1001 to 1026, wherein said IBD is ulcerative colitis. [The present invention 1028] The method according to any one of claims 1001 to 1026, wherein said IBD is Crohn's disease. [The present invention 1029] The following pharmacokinetic parameters in the plasma of said subject: Cmax (ng / mL) of 1 to 25; Tmax (hours) from 1 to 5 AUC between 10 and 250 t (ng.hour / mL) AUC between 10 and 300 inf (ng.h / mL), 3~10t 1 / 2 (time), AUC of 30 to 130 tau (ng.h / mL), Ctrough (ng / mL) of 1 to 5; an accumulated Cmax (ng.mL) of 0.5 to 2.5, and Accumulated AUC between 0.5 and 3.0 t (ng.hour / mL) resulting in one or more of the following: Any method of the present invention. [The present invention 1030] The following pharmacodynamic parameters in the plasma of said subject: ROmax (%) of 50 to 100, -20 to -60 receptor expression max Change in (%), Mean change (%) in receptor expression from -10 to -55; Steady-state ROmax (%) of 80-100; Average RO of 50-95 0-24 (h%), Average RO of 80-95 0-12 (h%), and Average RO of 70-90 12-24 (h%) Any of the methods of the present invention resulting in one or more of: [The present invention 1031] A method for treating an inflammatory disease or disorder in a subject in need thereof, comprising administering to the subject an α4β7 integrin antagonist, optionally administered at a dosage that results in unsaturated blood receptor occupancy (RO%) as measured by peak blood or serum levels of the antagonist. [The present invention 1032] The method of claim 1011, wherein the antagonist is optionally administered at a dosage that results in a blood RO of less than 90%, less than 80%, less than 70%, less than 60%, or less than 50%, as measured at peak blood or serum levels of the antagonist. [The present invention 1033] The method of any one of claims 1031 to 1032, wherein the antagonist is present in a pharmaceutical composition formulated for a route of administration selected from oral, parenteral, subcutaneous, buccal, nasal, inhalation, topical, and rectal administration. [The present invention 1034] The method of any of claims 1031 to 1033, wherein said antagonist is administered orally or rectally. [This invention 1035] 1036. The method of any of claims 1031 to 1034, wherein the disease or disorder is selected from the group consisting of inflammatory bowel disease (IBD), adult IBD, pediatric IBD, adolescent IBD, ulcerative colitis, Crohn's disease, celiac disease (non-tropical sprue), enteropathy associated with seronegative arthropathy, microscopic colitis, collagenous colitis, eosinophilic gastroenteritis, pouchitis occurring after radiation therapy, chemotherapy, proctocolectomy and ileoanal anastomosis, gastrointestinal cancer, pancreatitis, insulin-dependent diabetes mellitus, mastitis, cholecystitis, cholangitis, pericholangitis, chronic bronchitis, chronic sinusitis, asthma, primary sclerosing cholangitis, human immunodeficiency virus (HIV) infection of the GI tract, eosinophilic asthma, eosinophilic esophagitis, gastritis, colitis, microscopic colitis, and graft-versus-host disease (GVDH). [The present invention 1036] The method of any one of claims 1031 to 1035, wherein said disease or disorder is IBD. [This invention 1037] 1036. The method of claim 10, wherein said IBD is ulcerative colitis. [The present invention 1038] 1036. The method of claim 1036, wherein said IBD is Crohn's disease. [This invention 1039] the antagonist is a peptide dimer compound comprising two peptides, or a pharmaceutically acceptable salt thereof; Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH2 (SEQ ID NO: 5), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 6), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH2 (SEQ ID NO: 6), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 7), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-NH2 (SEQ ID NO: 7), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 8), or Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-NH2(SEQ ID NO: 8) comprising or consisting of any of Any of the methods of claims 1031 to 1038, wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, or a disulfide bond between the two Pens, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1040] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1) 1039. The method of claim 1039, wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [This invention 1041] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2) 1039. The method of claim 1039, wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1042] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3) 1039. The method of claim 1039, wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [This invention 1043] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4) 1039. The method of claim 1039, wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [This invention 1044] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH2 (SEQ ID NO: 5) 1039. The method of claim 1039, wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [This invention 1045] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5) 1039. The method of claim 1039, wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1046] The peptide dimer compound or a pharmaceutically acceptable salt thereof is TIFF2025186365000014.tif70132 or a pharmaceutically acceptable salt thereof. [This invention 1047] The peptide dimer compound or a pharmaceutically acceptable salt thereof is TIFF2025186365000015.tif88164 or a pharmaceutically acceptable salt thereof. [This invention 1048] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1) 1039. The method of claim 1039, wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [This invention 1049] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2) 1039. The method of claim 1039, wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [The present invention 1050] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3) 1039. The method of claim 1039, wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [This invention 1051] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4) 1039. The method of claim 1039, wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [This invention 1052] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH2 (SEQ ID NO: 5) 1039. The method of claim 1039, wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [This invention 1053] Each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5) 1039. The method of claim 1039, wherein each of the two peptides comprises a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety attached to the D-Lys amino acids of the two peptides, and the linker moiety is diglycolic acid (DIG). [This invention 1054] The peptide dimer compound or a pharmaceutically acceptable salt thereof is TIFF2025186365000016.tif70132 or a pharmaceutically acceptable salt thereof. [This invention 1055] The peptide dimer compound or a pharmaceutically acceptable salt thereof is TIFF2025186365000017.tif88164 or a pharmaceutically acceptable salt thereof. [This invention 1056] The peptide dimer compound or a pharmaceutically acceptable salt thereof is about 5, 6, 7, 8, 9, 10, 12.5, 25.0, 37.5, 50.0, 62.5, 75, 87.5, 100.0, 112.5, 125.0, 137.5, 150.0, 162.5, 175, 187.5, 200.0, 212.5, 225.0, 237.5, 250.0, 260.0, 270.0, 280.0, 290.0, 300.0, 310.0, 320.0, 330.0, 340.0, 350.0, 360.0, 370.0, 380.0, 390.0, 400.0, 410.0, 420.0, 430.0, 440.0, 450.0, 460.0, 470.0, 480.0, 490.0, 510.0, 520.0, 530.0, 540.0, 550.0, 560.0, 570.0, 580.0, 590.0, 600.0, 610.0, 620.0, 630.0, 640.0, 650.0, 660.0, 670.0, 680.0, 690.0, 700.0, 710.0, 720.0, 730.0, 740.0, 750.0, 760.0 62.5, 275, 287.5, 300.0, 312.5, 325.0, 337.5, 350.0, 362.5, 375, 387.5, 400.0, 412.5, 425.0, 437.5, 450.0, 462.5, 475, 487.5, or 500.0 mg of the compound of formula (I) administered to the subject. [This invention 1057] 1056. The method of claim 1056, wherein said dose is administered to said subject once daily or twice daily. [This invention 1058] The method according to any one of claims 1039 to 1057, wherein the pharmaceutically acceptable salt of the peptide dimer compound is an acetate salt. [This invention 1059] A pharmaceutical composition comprising a peptide dimer compound or a pharmaceutically acceptable salt thereof disclosed in any one of the present inventions 1039 to 1058. [The present invention 1060] 1059. The pharmaceutical composition of claim 1059, wherein said composition is formulated for oral delivery, and optionally said composition comprises an enteric coating. [This invention 1061] The method of any one of claims 1039 to 1058, comprising administering to the subject the pharmaceutical composition of any one of claims 1032 to 1034. [This invention 1062] The method of any of claims 1031 to 1058 or 1061, wherein said antagonist or a pharmaceutically acceptable salt thereof inhibits binding of α4β7 integrin to MAdCAM1. [This invention 1063] The method of any of claims 1031 to 1058 or 1061 to 1062, wherein the antagonist or a pharmaceutically acceptable salt thereof or the pharmaceutical composition is provided to the subject in need thereof at intervals sufficient to improve or ameliorate the disease state. [This invention 1064] 1063. The method of claim 1063, wherein the intervals are selected from the group consisting of 24 consecutive hours, hourly, every 4 hours, once a day, twice a day, three times a day, four times a day, every other day, weekly, biweekly, and monthly. [This invention 1065] The method of invention 1063 or 1064, wherein the antagonist or a pharmaceutically acceptable salt thereof or pharmaceutical composition is provided as an initial dose followed by one or more subsequent doses, the minimum interval between any two doses being less than one day, and each of the doses comprising an effective amount of the antagonist. [The present invention 1066] The effective amount of the antagonist or a pharmaceutically acceptable salt thereof or the pharmaceutical composition is selected from the group consisting of: a) saturation of approximately 50% or more of the MAdCAM1 binding sites on the α4β7 integrin molecule; b) inhibiting α4β7 integrin expression on the surface of said cells by about 50% or more; and c) about 50% or greater saturation of MAdCAM1 binding sites on the α4β7 molecule and about 50% or greater inhibition of α4β7 integrin expression on the surface of said cells, wherein (i) said saturation is maintained for a time period consistent with a dosing frequency of no more than twice daily, (ii) said inhibition is maintained for a time period consistent with a dosing frequency of no more than twice daily, or (iii) said saturation and said inhibition are each maintained for a time period consistent with a dosing frequency of no more than twice daily. The method of the present invention 1065 is sufficient to achieve at least one of the following. [Brief explanation of the drawings]

[0062] [Figure 1] 1 is a table showing T cell proliferation in response to indiacted treatment and inhibition of T cell proliferation by Compound A or vedolizumab. [Figure 2] 1 is a table showing CD45RO − naive and CD45RO + memory T cells in response to treatment with ati-CD3 or anti-CD3 + MAdaCAM. [Figure 3] 3A-B provide a table showing the increase in β7 expression during successive cycles of proliferation (FIG. 3A) and the decrease in surface expression of β7 in undivided CD4+ T cells in the presence of Compound A (FIG. 3B). [Figure 4] 1 is a graph showing reduction in surface β7 expression in five donors upon treatment with Compound A. [Figure 5] Figures 5A-C are graphs showing cytokine release after treatment with anti-CD3+MAdCAM1 and inhibition by Compound A of the following cytokines: IFNγ (Figure 5A), IL-23 (Figure 5B), and GM-CSF (Figure 5C). [Figure 6] Figures 6A-C are graphs showing cytokine release after treatment with anti-CD3+MAdCAM1 and inhibition by Compound A of the following cytokines: IL-10 (Figure 6A), IL-5 (Figure 6B), and TNFα (Figure 6C). [Figure 7] 1 is a graph showing % receptor occupancy (RO) in whole blood and Peyer's patches following administration of the indicated doses of Compound A. The table below the graph provides the % RO. [Figure 8] RO% in whole blood and Peyer's patches (top panel) is provided for six individual animals treated with the indicated amount of Compound A. RO% on day 14 is provided in the bottom panel. [Figure 9] Graphs are provided showing the concentration of Compound A in plasma and Peyer's patches after administration of the indicated doses of Compound A (left panel), and the RO% of Compound A in whole blood and Peyer's patches after administration of the indicated doses of Compound A (right panel). [Figure 10] 1 provides a graph showing the concentrations of Compound A detected in plasma and the indicated tissues at various time points after treatment. The bottom panel represents the data from the top panel plotted on an expanded scale. [Figure 11]1 is a graph summarizing various pharmacokinetic parameters of Compound A following administration of a single PO dose of 30 mg / kg in mice. [Figure 12] Graph showing the percentage of cultured cells bearing the indicated surface markers after the indicated treatments. For each cell type, the four bars from left to right correspond to the treatments indicated from top to bottom left. [Figure 13] Graph showing the percentage of cultured cells bearing the indicated surface markers after the indicated treatments. For each cell type, the four bars from left to right correspond to the treatments indicated from top to bottom left. [Figure 14] FIG. 1 is a graph showing α4β7 cell surface expression on PBMCs treated with Compound C or Compound D. FMO is used as a staining control. [Figure 15] FIG. 1 is a graph showing time-dependent α4β7 cell surface expression on CD4+ T memory cells treated with Compound A at 0 hours. [Figure 16] FIG. 1 is a graph showing concentration-dependent α4β7 cell surface expression on CD4+ T memory cells treated with Compound A at the indicated concentrations. [Figure 17] FIG. 1 is a graph showing concentration-dependent α4β7 cell surface expression on CD4+ T memory cells treated with Compound A at the indicated concentrations. [Figure 18] Graph showing concentration-dependent reduction in adhesion to MAdCAM1 on CD4+ T memory cells treated with Compound A at the indicated concentrations. [Figure 19] Graph showing the correlation between % reduction in adhesion to MAdCAM1 and % reduction in α4β7 expression. [Figure 20] 1 is a graph showing downregulation of α4β7 expression after treatment with Compound A, followed by recovery of α4β7 expression after treatment has ceased. [Figure 21] 1 is a graph showing the mean plasma concentrations over time following a single dose of Compound A in the amounts indicated. [Figure 22] 22A-B are graphs showing % receptor occupancy (FIG. 22A) and % receptor expression (FIG. 22B) over time following a single dose of Compound A in the amounts indicated. [Figure 23] Figures 23A-B are graphs showing the mean steady-state plasma concentrations of Compound A over time (Figure 23A) and % receptor occupancy (Figure 23B) following administration of 450 mg of Compound A as a liquid solution or immediate-release tablet. [Figure 24] 1 is a graph showing the correlation between plasma concentration of Compound A and receptor occupancy (%) after administration of Compound A. [Figure 25] 1 is a graph showing the mean receptor occupancy (%) in whole blood and Peyer's patches following administration of the indicated doses of Compound A. Relevant values ​​are provided in the table. [Figure 26] 1 provides graphs showing dose-dependent concentrations of Compound A in plasma (left panel) and Peyer's patches (right panel) following administration of the indicated doses of Compound A. [Figure 27] 1 is a table showing receptor occupancy in individual animals following treatment with Compound A at the indicated doses. DETAILED DESCRIPTION OF THE INVENTION

[0063] Detailed Description of the Invention Ulcerative colitis (UC) is a chronic inflammatory bowel disease (IBD) with a remitting and relapsing course, characterized by bloody diarrhea, abdominal cramps, and fatigue. The etiology is thought to result from an inappropriate immune response to gastrointestinal antigens and environmental triggers in genetically susceptible individuals. It is reported to be most prevalent in Europe and North America. UC has a significant negative impact on patients' quality of life and represents a high economic burden to health systems.

[0064] Inflammatory bowel diseases, such as ulcerative colitis, are managed with corticosteroids, 5-aminosalicylates, immunosuppressants, and, more recently, biologic agents targeted against specific mediators of inflammation. Treatment options for long-term treatment of ulcerative colitis are limited. 5-aminosalicylates, such as sulfasalazine, olsalazine, balsalazide, and various forms of mesalamine (e.g., Asacol, Pentasa, Lialda, Canasa), are effective only in mild to moderate disease, while patients with severe disease may be started on biologics. Several monoclonal antibodies against TNF-α (e.g., infliximab, adalimumab, golimumab, and certolizumab) are currently available. Drugs targeted against other cytokines involved in the inflammatory response, such as ustekinumab against IL-12 / IL-23, and the pan-JAK inhibitor tofacitinib, are some of the current treatment options available for inflammatory bowel disease, and several IL-23 and S1P1 inhibitors are also currently in clinical trials.

[0065] Despite a wide range of treatment options, treatment for inflammatory bowel disease remains limited, and available agents are not without risks. TNF-α inhibitors are ineffective in approximately one-fifth to one-third of patients, and 10–15% of treated patients who show initial benefit may lose response each year. Skin reactions are also the most common adverse reaction with anti-TNF therapy. These include immune-mediated complications such as injection site reactions, skin infections, psoriasis, and lupus-like syndromes, as well as rare skin cancers. Tofacitinib may increase the risk of infection and may increase the risk of thrombosis or thromboembolic events. There is growing recognition that alleviating the local inflammatory response may hold promise. Orally administered budesonide and 5-ASA are effective topically, and various other locally acting agents, including AMT-101, an oral biologic fusion protein that acts locally on interleukin-10, and TD-1473, a JAK inhibitor, have shown promise or are in clinical trials. Local delivery via oral administration may allow higher doses of drugs to be delivered to the target site without increasing systemic side effects.

[0066] Integrins are heterodimers that function as cell adhesion molecules. The α4 integrins, α4β1 and α4β7, are known to play an important role in lymphocyte migration throughout the gastrointestinal tract. They are expressed on most leukocytes, including B and T lymphocytes, monocytes, and dendritic cells, and mediate cell adhesion through binding to their respective primary ligands, vascular cell adhesion molecule (VCAM) and mucosal addressin cell adhesion molecule 1 (MAdCAM1), respectively. VCAM and MAdCAM1 differ in their binding specificity, in that VCAM binds to both α4β1 and α4β7, while MAdCAM1 is highly specific for α4β7.

[0067] The α4β7 integrin, which is primarily involved in the recruitment of leukocytes to the gastrointestinal (GI) tract, is present on the cell surface of a minority population of circulating T and B lymphocytes. Its primary ligand, MAdCAM1, is selectively expressed on the endothelium of the intestinal vasculature and is present at increased concentrations in inflamed tissues.

[0068] The present disclosure provides methods for treating IBD by inhibiting α4β7 integrin, for example, using a peptide dimer antagonist of α4β7 integrin, including, but not limited to, any of those disclosed herein. In particular, the present disclosure provides oral dosages of α4β7 integrin antagonists effective for treating IBD, including ulcerative colitis. In addition, the present disclosure provides pharmacokinetic and pharmacodynamic parameters of α4β7 integrin antagonists related to the antagonist's biological activities, such as inhibition of MAdCAM1-mediated T cell proliferation, reduction of T cell expression of β7 (and α4β7 integrin), internalization of α4β7 integrin on T cells, reduction of T cell homing to gastrointestinal tissues, reduction of cytokine release by T cells, reduction of T cell adhesion to MAdCAM1, and reduction of gastrointestinal inflammation. In certain embodiments, the T cells are CD4+ T memory cells.

[0069] Furthermore, it was previously thought that the mechanism underlying the use of α4β7 integrin antagonists to treat IBD involves the binding of the antagonist to α4β7 expressed on circulating T cells, which prevents T cells from binding to MAdCAM1 expressed on GI endothelial cells and thus prevents T cell extravasation into the inflamed gastrointestinal mucosa in IBD patients. Therefore, the goal was to achieve a maximum blood receptor occupancy (%RO), e.g., greater than 80%, greater than 90%, or even close to 100%, to block T cell binding and migration into the inflamed gastrointestinal mucosa.

[0070] In contrast, the present inventors have identified an alternative mechanism by which α4β7 integrin antagonists inhibit inflammation in inflamed tissues, such as inflamed gastrointestinal mucosa, by exerting a local effect. As disclosed in the accompanying Examples, when present in inflamed tissues, α4β7 integrin antagonists can inhibit MAdCAM1-mediated CD4+ T cell proliferation and cytokine production, which occurs through direct binding and stimulation of α4β7 integrin. This local effect does not require saturating blood receptor occupancy; instead, it has been demonstrated herein that oral administration of a subsaturating dose of the antagonist is sufficient to achieve a therapeutic effect, e.g., endoscopic or histological improvement. Accordingly, the present disclosure provides, inter alia, a method for treating IBD, comprising orally providing to a subject a subsaturating blood receptor occupancy dose of an α4β7 integrin antagonist, including, but not limited to, the peptide dimer compounds disclosed herein.

[0071] In certain aspects, the present disclosure provides methods for using α4β7 antagonist thioether peptide monomers and dimers as anti-inflammatory and / or immunosuppressive agents, e.g., for use in treating pathologies associated with the biological function of α4β7 or cells or tissues expressing MAdCAM1.

[0072] Aspects of the present invention relate to cyclized disulfide or thioether peptidic compounds that exhibit integrin antagonist activity, i.e., high specificity for α4β7 integrin. In certain embodiments, each peptide of the present invention comprises a downstream natural or unnatural amino acid and an upstream modified amino acid or aromatic group that can be bridged to form a cyclized structure via a disulfide or thioether bond. The peptides of the present invention demonstrate increased stability when orally administered as therapeutic agents.

[0073] In further related embodiments, the present invention provides methods for treating or preventing diseases or conditions associated with the biological function of integrin α4β7, comprising providing an effective amount of a peptide molecule of the present invention or a pharmaceutical composition of the present invention to a subject in need thereof. In certain embodiments, the disease or condition is inflammatory bowel disease. In certain embodiments, the inflammatory bowel disease is ulcerative colitis or Crohn's disease. In certain embodiments, the peptide molecule inhibits α4β7 binding to MAdCAM1. In certain embodiments, the peptide molecule or pharmaceutical composition is provided to a subject in need thereof at intervals sufficient to ameliorate the condition. In certain embodiments, the intervals are selected from the group consisting of 24 consecutive hours, hourly, every 4 hours, once daily, twice daily, three times daily, four times daily, every other day, weekly, biweekly, and monthly. In certain embodiments, the peptide molecule or pharmaceutical composition is provided as an initial dose followed by one or more subsequent doses, the minimum interval between any two doses being less than one day, and each dose comprising an effective amount of the peptide molecule. In certain embodiments, an effective amount of the peptide molecule or pharmaceutical composition is sufficient to achieve at least one of the following: a) about 50% or more saturation of MAdCAM1 binding sites on α4β7 integrin molecules, b) about 50% or more inhibition of α4β7 integrin expression on the cell surface, and c) about 50% or more saturation of MAdCAM1 binding sites on α4β7 molecules and about 50% or more inhibition of α4β7 integrin expression on the cell surface, wherein i) saturation is maintained for a period consistent with a dosing frequency of no more than twice daily, ii) inhibition is maintained for a period consistent with a dosing frequency of no more than twice daily, or iii) saturation and inhibition, each maintained for a period consistent with a dosing frequency of no more than twice daily. In certain embodiments, the peptide molecule is administered orally, parenterally, or topically.

[0074] definition As used herein, the singular forms "a," "and," and "the" include plural references unless the context clearly dictates otherwise.

[0075] Where the term "comprising" is used herein, it is understood that the invention also includes the same embodiments in which the term "comprising" is replaced with "consisting essentially of" or "consisting of."

[0076] As used herein, the following terms have the indicated meanings.

[0077] The term "peptide," as used herein, broadly refers to a structure comprising a sequence of two or more amino acids joined together by peptide bonds. In certain embodiments, it refers to a sequence of two or more amino acids joined together by peptide bonds. It should be understood that the term does not imply a polymer of amino acids of a specific length, nor is it intended to imply or distinguish whether the polypeptide is produced using recombinant technology, chemical synthesis, enzymatic synthesis, or naturally occurring. The term "peptide," as generally used herein, includes both peptide monomers and peptide dimers.

[0078] As used herein, the term "monomer" may also be referred to as a "peptide monomer," a "peptide monomer molecule," or a "monomeric peptide." The term "monomer" refers to a single sequence of two or more amino acids joined together by peptide bonds.

[0079] The term "dimer," as used herein, broadly refers to a peptide comprising two monomeric peptide subunits (e.g., thioether monomeric peptides) linked at their respective C- or N-termini. Dimers of the present invention may include homodimers and heterodimers that function as integrin antagonists. The term "dimer" may also be referred to herein as a "peptide dimer," a "peptide dimeric molecule," a "dimeric peptide," or a "dimeric compound." The term "monomeric peptide subunit" may also be referred to herein as a "monomer subunit," a "peptide monomer subunit," a "peptide subunit," a "peptide dimeric subunit," a "dimeric subunit," a "monomeric subunit," or a "subunit of a peptide dimer."

[0080] The term "thioether," as used herein, refers to a cyclizing covalent bond, i.e., a C-S bond, formed between an upstream amino acid or aromatic acid group and a downstream sulfur-containing amino acid, or isotope thereof.

[0081] The term "linker," as used herein, broadly refers to a chemical structure capable of linking two thioether monomer subunits together to form a dimer.

[0082] The term "L-amino acid," as used herein, refers to the "L" isomeric form of a peptide; conversely, the term "D-amino acid" refers to the "D" isomeric form of a peptide. While the amino acid residues described herein are preferably in the "L" isomeric form, residues in the "D" isomeric form can be substituted for any L-amino acid residue so long as the desired functionality is retained by the peptide.

[0083] Unless otherwise indicated, the term "NH2" as used herein refers to the free amino group present at the amino terminus of a polypeptide. The term "OH" as used herein refers to the free carboxy group present at the carboxy terminus of a peptide. Furthermore, the term "Ac" as used herein refers to acetyl protection via acylation of the N-terminus of a polypeptide. When indicated, "NH2" refers to the free amino group side chain of an amino acid. When indicated, the term "Ac" as used herein refers to the acylation of an amino acid with an NH2 group.

[0084] The term "carboxy" as used herein refers to -CO2H.

[0085] The terms "isostere" or "isosteric substitution," as used herein, refer to any amino acid or other analog moiety that has similar chemical and / or structural properties as a particular amino acid. In certain embodiments, an "isostere" or "suitable isostere" of an amino acid is another amino acid of the same type, and amino acids belong to the following classes based on the tendency of their side chains to contact polar solvents such as water: hydrophobic (low tendency to contact water), polar, or charged (energetically favorable contact with water). Charged amino acid residues include lysine (+), arginine (+), aspartate (-), and glutamate (-). Polar amino acids include serine, threonine, asparagine, glutamine, histidine, and tyrosine. Hydrophobic amino acids include alanine, valine, leucine, isoleucine, proline, phenylalanine, tryptophan, cysteine, and methionine. The amino acid glycine does not have a side chain and is difficult to assign to one of the above classes. However, glycine is often found on the surface of proteins, often within loops, providing greater flexibility to these regions, and isosteres may have similar characteristics. Proline has the opposite effect, providing rigidity to the protein structure by imposing specific torsion angles on segments of the polypeptide chain.

[0086] The term "cyclization," as used herein, refers to a reaction in which one portion of a polypeptide molecule is linked to another portion of a polypeptide molecule to form a closed ring, such as by the formation of a disulfide or thioether bond. In certain embodiments, the monomeric subunits of the peptide monomers and peptide dimers of the invention are cyclized via an intramolecular disulfide or thioether bond.

[0087] The term "receptor," as used herein, refers to a chemical group of a molecule on the surface of a cell or within the interior of a cell that has affinity for a specific chemical group or molecule. Binding between a peptide molecule and a targeting integrin can provide a useful diagnostic tool.

[0088] The term "integrin-associated disease," as used herein, refers to indications that manifest as a result of integrin binding and that can be treated through the administration of integrin antagonists.

[0089] The term "pharmaceutically acceptable salt," as used herein, refers to a salt or zwitterionic form of a compound of the present invention that is water- or oil-soluble or dispersible, suitable for the treatment of disease without undue toxicity, irritation, or allergic reaction, commensurate with a reasonable benefit / risk ratio, and effective for its intended use. Salts can be prepared during the final isolation and purification of the compound or separately by reacting an amino group with a suitable acid. Representative acid addition salts include acetate, adipate, alginate, citrate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, camphorate, camphorsulfonate, digluconate, glycerophosphate, hemisulfate, heptanoate, hexanoate, formate, fumarate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate (isethionate), lactate, maleate, mesitylene, and the like. The salts of the amino groups in the compounds of the present invention include sulfonate, methanesulfonate, naphthylenesulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, pamoate, pectinate, persulfate, 3-phenylproprionate, picrate, pivalate, propionate, succinate, tartrate, trichloroacetate, trifluoroacetate, phosphate, glutamate, bicarbonate, para-toluenesulfonate, and undecanoate. The amino groups in the compounds of the present invention can also be quaternized with methyl, ethyl, propyl, and butyl chlorides, bromides, and iodides; dimethyl sulfate, diethyl sulfate, dibutyl sulfate, and diamyl sulfate; decyl, lauryl, myristyl, and steryl chlorides, bromides, and iodides; and benzyl and phenethyl bromides. Examples of acids which can be used to form therapeutically acceptable addition salts include inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, and phosphoric acid, and organic acids such as oxalic acid, maleic acid, succinic acid, and citric acid.

[0090] The term "N(alpha) methylation," as used herein, describes the methylation of the alpha amine of an amino acid, also commonly referred to as N-methylation.

[0091] The term "acylating organic compound," as used herein, refers to various compounds having a carboxylic acid functionality that can be used to acylate the C-terminus and / or N-terminus of a peptide molecule. Non-limiting examples of acylating organic compounds include cyclopropylacetic acid, 4-fluorobenzoic acid, 4-fluorophenylacetic acid, 3-phenylpropionic acid, succinic acid, glutaric acid, cyclopentanecarboxylic acid, glutaric acid, succinic acid, 3,3,3-trifluoropropeonic acid, and 3-fluoromethylbutyric acid.

[0092] All peptide sequences are written according to the generally accepted convention with the α-N-terminal amino acid residue on the left and the α-C-terminus on the right. As used herein, the term "α-N-terminus" refers to the free α-amino group of an amino acid in a peptide, and the term "α-C-terminus" refers to the free α-carboxylic acid terminus of an amino acid in a peptide.

[0093] As used herein, the term "amino acid" or "any amino acid" refers to any and all amino acids, including naturally occurring amino acids (e.g., α-amino acids), unnatural amino acids, modified amino acids, and non-natural amino acids. This includes both D- and L-amino acids. Natural amino acids include those found in nature, such as the 23 amino acids that combine into peptide chains to form the building blocks of a wide variety of proteins. These are primarily L-stereoisomers, although several D-amino acids occur in bacterial envelopes and some antibiotics. "Non-standard" naturally occurring amino acids are pyrrolysine (found in methanogens and other eukaryotes), selenocysteine ​​(present in most eukaryotes as well as many non-eukaryotes), and N-formylmethionine (encoded by the start codon AUG in bacteria, mitochondria, and chloroplasts). "Unnatural" or "non-natural" amino acids are non-proteinogenic amino acids (i.e., amino acids that are not naturally encoded or found in the genetic code), either occurring in nature or chemically synthesized. Over 140 naturally occurring amino acids are known, with thousands more combinations possible. Examples of "unnatural" amino acids include β-amino acids (β 3 and β 2 ), homoamino acids, proline and pyruvate derivatives, tri-substituted alanine derivatives, glycine derivatives, ring-substituted phenylalanine and tyrosine derivatives, linear core amino acids, diamino acids, D-amino acids, alpha-methyl amino acids, and N-methyl amino acids. Unnatural or non-natural amino acids also include modified amino acids. "Modified" amino acids include amino acids (e.g., natural amino acids) that have been chemically modified to include a group, groups, or chemical moiety that is not naturally present on the amino acid.

[0094] Generally, the names of naturally occurring and non-naturally occurring aminoacyl residues used herein follow the naming conventions proposed by the IUPAC Commission on the Nomenclature of Organic Chemistry and the IUPAC-IUB Commission on Biochemical Nomenclature, as presented in "Nomenclature of α-Amino Acids (Recommendations, 1974)" Biochemistry, 14(2), (1975). To the extent that the names and abbreviations of amino acids and aminoacyl residues used in this specification and the appended claims differ from these suggestions, they will be made clear to the reader. Some abbreviations useful in describing this invention are defined below in Table 1, below. (Table 1) Abbreviations TIFF2025186365000018.tif40160TIFF2025186365000019.tif244160TIFF2025186365000020.tif246160TIFF20251863650 00021.tif237160TIFF2025186365000022.tif226160TIFF2025186365000023.tif241160TIFF2025186365000024.tif99160

[0095] Peptide antagonists The present invention generally relates to cyclic peptides, e.g., disulfide and thioether peptides, that have been shown to have integrin antagonist activity. In particular, the present invention relates to various peptides that form cyclized structures via intramolecular bonds, e.g., disulfide or thioether bonds, e.g., intramolecular disulfide or thioether bonds. While the disclosure provided herein generally relates to peptides with disulfide or thioether intramolecular bonds, it is understood that other cyclic peptide antagonists of the a4b7 integrin, including those containing intramolecular bonds of different nature, and cyclic peptide antagonists of the α4β7 integrin containing bonds between two peptide monomer subunits, can also be used to practice the methods disclosed herein. Certain embodiments relate to disulfide or thioether peptide monomers with integrin antagonist activity. Some embodiments relate to disulfide or thioether peptide dimers with integrin antagonist activity comprising hetero- or homo-monomeric thioether peptide subunits, where the disulfide or thioether peptide subunits are linked at either their C- or N-termini. Cyclization of peptides, peptide monomers, or peptide subunits has been shown to increase the potency, selectivity, and stability of peptide molecules, as described below. In some embodiments, dimerizing peptide monomers increases potency, selectivity, and / or stability compared to non-dimerized peptides. Exemplary peptides and genera that can be used in accordance with the methods disclosed herein are provided in the following patent application publications, each of which is incorporated by reference in its entirety: PCT Application Publication Nos. 2014 / 059213, 2014 / 165448, 2014 / 165449, 2015 / 176035, 2016 / 054411, and 2016 / 054445.

[0096] In some examples, the monomeric peptide further comprises a C-terminus and / or an N-terminus (or both a C-terminus and an N-terminus) containing a free amine. Similarly, a peptide dimer may comprise one or more C-terminus or N-terminus containing a free amine. Thus, users may modify either terminus to include a modifying group, such as PEGylation, e.g., small PEGylations (e.g., PEG4 to PEG13). Users may further modify either terminus through acylation. For example, in some examples, at least one of the N-terminus and C-terminus of a peptide molecule is acylated with an acylating organic compound selected from the group consisting of 2-Me-trifluorobutyl, trifluoropentyl, acetyl, octonyl, butyl, pentyl, hexyl, palmityl, trifluoromethylbutyric acid, cyclopentanecarboxylic acid, cyclopropylacetic acid, 4-fluorobenzoic acid, 4-fluorophenylacetic acid, and 3-phenylpropionic acid. In some examples, peptide molecules of the present invention comprise both a free carboxyl terminus and a free amino terminus, allowing users to selectively modify the peptide to achieve a desired modification. It is further understood that the C-terminal residue of the thioether peptides, e.g., thioether monomers, disclosed herein is an amide or an acid unless otherwise indicated. Accordingly, one of skill in the art will understand that the thioether peptides of the present invention can be selectively modified.

[0097] With respect to peptide dimers, it is understood that monomer subunits are dimerized to form peptide dimer molecules, e.g., the monomer subunits are joined or dimerized by a suitable linker moiety as defined herein. Some of the monomer subunits are shown to have C- and N-termini, both of which contain free amines. Thus, users can modify either terminal end of the monomer subunit to eliminate the free amine at either the C- or N-terminus, thereby allowing dimerization at the remaining free amine. Thus, some of the monomer subunits contain both a free carboxy or amide and a free amino terminus at the C-terminus, allowing users to selectively modify the subunit to achieve dimerization at the desired terminus. Thus, those skilled in the art will understand that the monomer subunits of the present invention can be selectively modified to achieve a single specific amine for desired dimerization.

[0098] It is further understood that the C-terminal residue of the monomer subunits disclosed herein contains -OH or -NH unless otherwise indicated. It is further understood that dimerization at the C-terminus can be facilitated by using a suitable amino acid having a side chain with an amine functionality, as is generally understood in the art. In certain embodiments, a linker is attached to the functional amine group of the C-terminal amino acid of each of the peptide monomer subunits to form a dimer. With respect to the N-terminal residue, it is generally understood that dimerization can be achieved via the free amine of the terminal residue or by using a suitable amino acid side chain with a free amine, as is generally understood in the art.

[0099] The peptide monomers and dimers of the present invention, or peptide subunits thereof, can further comprise one or more terminal modification groups. In at least one embodiment, the terminal end of the peptide is modified to include a terminal modification group selected from the non-limiting group consisting of DIG, PEG4, PEG13, PEG25, PEG1K, PEG2K, PEG4K, PEG5K, polyethylene glycol having a molecular weight of 400 Da to 40,000 Da, PEG having a molecular weight of 40,000 Da to 80,000 Da, IDA, ADA, glutaric acid, succinic acid, isophthalic acid, 1,3-phenylenediacetic acid, 1,4-phenylenediacetic acid, 1,2-phenylenediacetic acid, AADA, and suitable aliphatic, aromatic, and heteroaromatic compounds.

[0100] In some embodiments of the peptide dimers, peptide dimer subunits, or peptide monomers described herein, the N-terminus further comprises a suitable linker moiety or other modifying group. In some embodiments of the peptide monomers described herein, the N-terminus may further be acylated.

[0101] Non-limiting examples of terminal modification groups are provided in Table 2. Table 2: Exemplary terminal modification groups TIFF2025186365000025.tif74165TIFF2025186365000026.tif233165TIFF2025186365000027.tif129165

[0102] Linker moieties of the present invention can comprise any structure, length, and / or size compatible with the teachings herein. In at least one embodiment, the linker moiety is selected from the non-limiting group consisting of DIG, PEG4, PEG4-biotin, PEG13, PEG25, PEG1K, PEG2K, PEG3.4K, PEG4K, PEG5K, IDA, ADA, Boc-IDA, glutaric acid, isophthalic acid, 1,3-phenylenediacetic acid, 1,4-phenylenediacetic acid, 1,2-phenylenediacetic acid, triazine, Boc-triazine, IDA-biotin, PEG4-biotin, AADA, suitable aliphatic compounds, aromatic compounds, heteroaromatic compounds, and polyethylene glycol-based linkers having a molecular weight of approximately 400 Da to approximately 40,000 Da, or approximately 40,000 Da to approximately 80,000 Da.

[0103] If the linker is IDA, ADA, or any linker with a free amine, it can be acylated with an acylating organic compound selected from the group consisting of 2-me-trifluorobutyl, trifluoropentyl, acetyl, octonyl, butyl, pentyl, hexyl, palmityl, lauryl, oleoyl, trifluoromethylbutyric acid, cyclopentanecarboxylic acid, cyclopropylacetic acid, 4-fluorobenzoic acid, 4-fluorophenylacetic acid, 3-phenylpropionic acid, tetrahedro-2H-pyran-4carboxylic acid, succinic acid, and glutaric acid, straight-chain aliphatic acids having 10 to 20 carbon units, cholic acid, and other bile acids. In some instances, small PEGs (PEG4 to PEG13), Glu, or Asp are used as spacers before acylation.

[0104] In certain embodiments, a linker connects two monomeric subunits by connecting two sulfur-containing C-terminal amino acids or N-terminal amino acids. In some embodiments, the two sulfur-containing amino acids are connected by a linker comprising a dihalide, an aliphatic chain, or PEG. In certain embodiments, a linker connects two monomeric subunits by connecting the sulfur-containing C-terminal amino acids at the C-terminus of each monomeric subunit. In some embodiments, two sulfur-containing amino acids are connected by a linker comprising a homobifunctional maleimide crosslinker, a dihalide, 1,2-bis(bromomomethyl)benzene, 1,2-bis(chloromomethyl)benzene, 1,3-bis(bromomomethyl)benzene, 1,3-bis(chloromomethyl)benzene, 1,4-bis(bromomomethyl)benzene, 1,4-bis(chloromomethyl)benzene, 3,3'-bis-bromomethyl-biphenyl, or 2,2'-bis-bromomethyl-biphenyl. Certain haloacetyl crosslinkers contain iodoacetyl or bromoacetyl groups. These homobifunctional linkers may contain a spacer comprising PEG or an aliphatic chain.

[0105] Non-limiting examples of suitable linker moieties are provided in Table 3. Table 3: Exemplary linker moieties TIFF2025186365000028.tif108168TIFF2025186365000029.tif225168TIFF2025186365000030.tif21216 8TIFF2025186365000031.tif220168TIFF2025186365000032.tif218168TIFF2025186365000033.tif88168

[0106] Those skilled in the art will understand that certain amino acids and other chemical moieties are modified when they are attached to another molecule. For example, an amino acid side chain can be modified when it forms an intramolecular bridge with another amino acid side chain. In addition, when Homo-Ser-Cl is attached to an amino acid such as Cys or Pen via a thioether bond, the Cl moiety is released. Thus, as used herein, an amino acid such as Homo-Ser-Cl or a modified amino acid (e.g., Xaa) present in the peptide dimer of the present invention is referred to as a "modified" amino acid. 4 Position or Xaa 10 Reference to a nucleotide (position) is meant to include the form of that amino acid or modified amino acid present in the peptide both before and after formation of the intramolecular bond.

[0107] In certain embodiments, the methods disclosed herein are practiced using any of the following peptide antagonists of α4β7 integrin, although it is understood that the methods disclosed herein may also be practiced using other peptide antagonists, including those disclosed in the PCT applications incorporated herein by reference.

[0108] In some embodiments, the peptide antagonist is a peptide dimeric compound comprising two peptides, or a pharmaceutically acceptable salt thereof, wherein each of the two peptides has the sequence: 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4), 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5) or 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH2 (SEQ ID NO: 5), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 6), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH2 (SEQ ID NO: 6), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 7), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-NH2 (SEQ ID NO: 7), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 8), or Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-NH2(SEQ ID NO: 8) Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, or a disulfide bond between the two Pens, and the two peptides are linked by a linker moiety bonded to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG). The peptide may also contain an N-terminal Ac.

[0109] In certain embodiments of any of the peptide antagonists or pharmaceutically acceptable salts thereof, the pharmaceutically acceptable salt of the peptide dimeric compound is an acetate salt.

[0110] In certain embodiments, each of the two peptides has the sequence: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0111] In certain embodiments, each of the two peptides has the sequence: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0112] In certain embodiments, each of the two peptides has the sequence: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0113] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0114] In certain embodiments, each of the two peptides comprises or consists of the sequence. 2-Methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH2 (SEQ ID NO: 5), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0115] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0116] In certain embodiments, the peptide dimeric compound or a pharmaceutically acceptable salt thereof comprises: TIFF2025186365000034.tif70132 or a pharmaceutically acceptable salt thereof.

[0117] In certain embodiments, the peptide dimeric compound or a pharmaceutically acceptable salt thereof comprises: TIFF2025186365000035.tif81152 or a pharmaceutically acceptable salt thereof.

[0118] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0119] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0120] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3), Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0121] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0122] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH (SEQ ID NO: 5); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0123] In certain embodiments, each of the two peptides has the sequence: comprising or consisting of 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5); Each of the two peptides contains a thioether bond between 2-methylbenzoyl and Pen, and the two peptides are connected by a linker moiety attached to the D-Lys amino acids of the two peptides, the linker moiety being diglycolic acid (DIG).

[0124] In certain embodiments, the peptide dimeric compound or a pharmaceutically acceptable salt thereof comprises: TIFF2025186365000036.tif70132 or a pharmaceutically acceptable salt thereof.

[0125] In certain embodiments, the peptide dimeric compound or a pharmaceutically acceptable salt thereof comprises: TIFF2025186365000037.tif80150 or a pharmaceutically acceptable salt thereof.

[0126] In certain embodiments, the peptide dimer compound is Compound A or Compound B, as described in the accompanying Examples.

[0127] Peptide Biological Activity In certain embodiments, the peptide molecules disclosed herein have increased α4β7 binding affinity, increased selectivity for α4β1, and increased stability in simulated intestinal fluid (SIF) and the gastric environment under reducing conditions. These novel antagonist molecules demonstrate high binding affinity to α4β7, thereby preventing the binding between α4β7 and the MAdCAM1 ligand. Accordingly, these peptide molecules have been shown to be effective in eliminating and / or reducing inflammatory processes in various experiments.

[0128] Peptide monomer and dimer molecules bind to or associate with α4β7 integrin, disrupting or blocking the binding between α4β7 and MAdCAM1 ligand. In certain embodiments, peptide dimer and monomer molecules of the present invention inhibit or reduce the binding between α4β7 and MAdCAM1 ligand. In certain embodiments, peptides of the present invention reduce the binding between α4β7 and MAdCAM1 ligand by at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% compared to a negative control peptide. Methods for determining binding are known in the art and described herein, including, for example, ELISA assays.

[0129] In certain embodiments, peptide monomer or dimer molecules have an IC50 of <500 nM, <250 nM, <100 nM, <50 nM, <25 nM, or <10 nM. Methods for determining activity are known in the art and include any of those described in the accompanying examples.

[0130] In some embodiments, the peptide monomer or dimer molecules have a half-life of greater than 180 minutes when exposed to simulated intestinal fluid (SIF). Some implementations further provide peptide monomer or dimer molecules with half-lives of approximately 1 minute to approximately 180 minutes. Similarly, these peptides are stable to the gastric environment under reducing conditions, with half-lives of >120 minutes when tested in a DTT (dithiothreitol) assay.

[0131] In certain embodiments, the peptide monomer or dimer molecule has increased stability, increased gastrointestinal stability, and / or increased stability in simulated intestinal fluid (SIF) compared to a control peptide. In certain embodiments, the control peptide is a peptide that has the same or a highly related amino acid sequence (e.g., >90% sequence identity) as the peptide monomer or dimer, but does not form a cyclized structure via a thioether bond. In some embodiments of dimeric molecules, the control peptide is not dimerized. In certain embodiments, the only difference between the peptide monomer or dimer and the control peptide is that the peptide contains one or more amino acid substitutions that introduce one or more amino acid residues into the peptide, and the introduced residue forms a thioether bond with another residue in the peptide.

[0132] Methods for determining peptide stability are known in the art. In certain embodiments, the stability of a peptide (e.g., a peptide monomer or dimer described herein) is determined using a SIF assay, for example, as described in the accompanying Examples. In certain embodiments, a peptide monomer or dimer molecule of the present invention has a half-life under a given set of conditions (e.g., temperature) when exposed to SIF of more than 1 minute, more than 10 minutes, more than 20 minutes, more than 30 minutes, more than 60 minutes, more than 90 minutes, more than 120 minutes, more than 3 hours, or more than 4 hours. In certain embodiments, the temperature is about 25°C, about 4°C, or about 37°C, and the pH is physiological pH, i.e., about pH 7.4.

[0133] In some embodiments, half-life is measured in vitro using any suitable method known in the art, for example, in some embodiments, the stability of peptide monomer or dimer molecules of the invention is determined by incubating the peptide with pre-warmed human serum (Sigma) at 37° C. Samples are typically taken at various time points up to 24 hours, and the stability of the samples is analyzed by dissociating the peptide monomer or dimer from serum proteins and then analyzing for the presence of the peptide monomer or dimer of interest using LC-MS.

[0134] In certain embodiments, the peptide dimer or monomer molecule inhibits or reduces α4β7-mediated inflammation. In related embodiments, the peptide monomer or dimer of the present invention inhibits or reduces α4β7-mediated secretion or release of one or more cytokines (including any disclosed herein) by T cells, e.g., T cells in the GI mucosa in response to MAdCAM1. Methods for determining inhibition of cytokine secretion and inhibition of signaling molecules are known in the art.

[0135] In certain embodiments, the peptide monomer or dimer molecules demonstrate increased binding selectivity, in particular instances, the peptide monomer or dimer binds to α4β7 with at least 2-fold, 3-fold, 5-fold, or 10-fold greater affinity than the monomer or dimer binds to α4β1.

[0136] In some embodiments, peptide monomer or dimeric molecules demonstrate increased potency as a result of replacing various native aminoacyl residues with N-methylated analog residues. In certain embodiments, potency is measured as the IC50 for binding to α4β7, e.g., determined as described herein, although in some embodiments, potency indicates functional activity, e.g., according to a cell adhesion assay.

[0137] In certain embodiments, any of these superior properties of the peptides of the invention are measured relative to a control peptide.

[0138] Manufacturing method Peptides of the invention (e.g., peptide monomers or peptide dimers) can be synthesized by techniques known to those of skill in the art, for example, as disclosed in PCT Publication Nos. 2014 / 059213, 2014 / 165448, 2014 / 165449, 2015 / 176035, 2016 / 054411, or 2016 / 054445. Such techniques include the use of commercially available robotic protein synthesizers (e.g., the Symphony multiplex peptide synthesizer from Protein Technologies). In some embodiments, novel peptide monomer or dimer subunits are synthesized and purified using the techniques described herein.

[0139] Therapeutic Methods and Pharmaceutical Compositions In some embodiments, the present invention provides methods for treating an individual or subject suffering from, for example, a condition or symptom characterized by α4β7 integrin binding to MAdCAM1, the method comprising providing or administering to the individual or subject an integrin antagonist, e.g., a peptide molecule, described herein. In certain embodiments, the subject or individual is a mammal, e.g., a human or a non-human mammal such as a dog, cat, or horse. It is understood that the integrin antagonist can be present in a pharmaceutical composition, e.g., any of those disclosed herein. It is further understood that other agents that disrupt α4β7 integrin or MAdCAM1 signaling, or, for example, inhibit α4β7 integrin binding to MAdCAM1, can be used as alternatives to the antagonists disclosed herein.

[0140] In certain embodiments of the disclosed methods, the methods reduce cell surface expression of β7 on CD4+ T cells in the gastrointestinal tract.

[0141] In certain embodiments of the disclosed methods, the methods inhibit MAdCAM1-mediated T cell proliferation in the gastrointestinal tract.

[0142] In certain embodiments of the disclosed methods, the methods reduce cell surface expression of β7 on CD4+ T cells in the gastrointestinal tract.

[0143] In certain embodiments of the disclosed methods, the methods induce internalization of α4β7 integrin on CD4+ T memory cells.

[0144] In certain embodiments of the disclosed methods, the methods cause a reduction in adhesion of CD4+ T memory cells to MAdCAM1 in the gastrointestinal tract.

[0145] In certain embodiments of the disclosed methods, the methods inhibit homing of T cells to the gastrointestinal tract, optionally to the ileal lamina propria and / or Peyer's patches.

[0146] In certain embodiments of the methods of the present disclosure, the methods are used to treat IBD, optionally wherein the IBD is ulcerative colitis or Crohn's disease.

[0147] In certain embodiments of the disclosed methods, the methods result in one or more of the following pharmacokinetic parameters in the plasma of the subject: Cmax (ng / mL) of 1 to 25, optionally 4 to 12; Tmax (hours) from 1 to 5, optionally from 2 to 4; AUC of 10 to 250, optionally 50 to 150 t (ng.h / mL), AUC of 10–300, optionally 30–250 inf (ng.h / mL), 3~10, optionally 4~10 t 1 / 2 (time), AUC of 30 to 130 tau (ng.h / mL), Ctrough (ng / mL) of 1 to 5; an accumulated Cmax (ng.mL) of 0.5 to 2.5, optionally 2 to 3, and Accumulated AUC between 0.5 and 3.0 t (ng.h / mL).

[0148] In certain embodiments of these methods, the methods comprise orally providing an antagonist disclosed herein, optionally Compound A or Compound A, at a dose of about 150 mg twice daily or about 450 mg twice daily.

[0149] In certain embodiments of the disclosed methods, the methods result in one or more of the following pharmacodynamic parameters in the plasma of the subject: 50~100, optionally, 90~100 ROmax(%); 50-95, optionally 65-95 average RO(%); -20 to -60, optionally -35 to -60 receptor expression max Change in (%), mean change (%) in receptor expression from -10 to -55, optionally from -25 to -55; Steady-state ROmax (%) of 80-100; 75~90 or 50~95, optional, average RO of 65~95 0-24 (h%), Average RO of 80-95 0-12 (h%), and Average RO of 70-90 12-24 (h%).

[0150] In certain embodiments of these methods, the methods comprise orally providing an antagonist disclosed herein, optionally Compound A or Compound A, at a dose of about 150 mg twice daily or about 450 mg twice daily.

[0151] In certain embodiments of the methods disclosed herein, a subject is provided with a dose or amount of an α4β7 integrin antagonist (or other agent) that does not saturate blood receptors on circulating T cells, e.g., α4β7 integrin receptors. Thus, the dose or amount is one that results in a subsaturated blood receptor occupancy (RO%). In certain embodiments, the dose results in an RO% of less than 90%, less than 80%, less than 70%, less than 60%, less than 50%, less than 40%, less than 30%, less than 20%, or less than 10%. In certain embodiments, the RO% is less than 50% or less than 40%. The RO% can be measured at drug levels or maximum RO%. In certain embodiments, maximum RO is measured about 4 hours after the dose, while trough levels occur about 24 hours after the dose. In certain embodiments, the method is performed using a peptide dimer compound disclosed herein, e.g., Compound A or Compound B. In certain embodiments, the dose is provided orally or topically, e.g., rectally. In certain embodiments, subjects are provided with this dose once or twice daily.

[0152] In certain embodiments of the methods disclosed herein, a subject is provided with a dose or amount of an α4β7 integrin antagonist (or other agent) that achieves high antagonist levels and / or occupancy of T cell α4β7 in gastrointestinal tissue. In certain embodiments, the dose results in a T cell α4β7 occupancy of the GI mucosa of at least 95%, at least 90%, at least 80%, at least 70%, at least 60%, at least 50%, at least 40%, or at least 30%. In certain embodiments, the method is carried out using a peptide dimer compound disclosed herein, e.g., Compound A or Compound B. In certain embodiments, the dose is provided orally or topically, e.g., rectally. In certain embodiments, the subject is provided with this dose once or twice daily.

[0153] In certain embodiments of the methods disclosed herein, a subject is provided with an α4β7 integrin antagonist (or other agent) at a dose or amount that achieves a ratio of RO% in blood / RO% in Peyer's patches (or other GI tissue) of less than 1.0, less than 0.9, less than 0.8, less than 0.7, less than 0.6, or less than 0.5.

[0154] In certain embodiments, the subject has a blood glucose level of about 5, 6, 7, 8, 9, 10, 12.5, 25.0, 37.5, 50.0, 62.5, 75, 87.5, 100.0, 112.5, 125.0, 137.5, 150.0, 162.5, 175, 187.5, 200.0, 212.5, 225.0, 237.5, 250.0, 260.0, 270.0, 280.0, 290.0, 300.0, 310.0, 320.0, 330.0, 340.0, 350.0, 360.0, 370.0, 380.0, 390.0, 400.0, 410.0, 420.0, 430.0, 440.0, 450.0, 460.0, 470.0, 480.0, 490.0, 510.0, 520.0, 530.0, 540.0, 550.0, 560.0, 570.0, 580.0, 590.0, 600.0, 610.0, 620.0, 630.0, 640.0, 650.0, 660.0, 670.0, 680.0, 690.0, 700.0, 710.0, 720.0, 730.0, 740.0, 750.0, 760.0, It may be provided in any of the following doses or amounts: 62.5, 275, 287.5, 300.0, 312.5, 325.0, 337.5, 350.0, 362.5, 375, 387.5, 400.0, 412.5, 425.0, 437.5, 450.0, 462.5, 475, 487.5, or 500.0 mg. In some embodiments, the subject is provided with a dose of about 12.5, 25.0, 37.5, 50.0, 62.5, 75, 87.5, 100.0, 112.5, 125.0, 137.5, 150.0, 162.5, 175, 187.5, 200.0, 212.5, 225.0, 237.5, 250.0, 262.5, 275, 287.5, 300.0, 350.0, 400.0, 450.0, or 500.0 mg. In some embodiments, the subject is provided with a dose of about 70, 75, 80, 85, 90, 95, 100, 105, 110, 115, 120, 125, or 130 mg. In some embodiments, subjects are provided with a dose of about 85, 90, 95, 100, 105, 110, or 115 mg. In some embodiments, subjects are provided with a dose of about 95, 100, or 105 mg. In some embodiments, subjects are provided with a dose of about 100 mg. In some embodiments, subjects are provided with a dose ranging from about 100 mg to about 500 mg, optionally once daily or twice daily. In some embodiments, subjects are provided with a dose ranging from about 200 mg to about 1000 mg, optionally taken as a single dose or in divided doses twice daily (e.g., half the amount). In some embodiments, subjects are provided with a dose ranging from about 100 mg to about 1500 mg per day, optionally taken as a single dose or in divided doses twice daily (e.g., half the amount). In some embodiments, subjects are provided with a dose ranging from about 100 mg to about 1500 mg once daily or twice daily.In some embodiments, subjects are provided with a dose of about 100, 150, 200, 250, 300, 250, 400, 450, or 500 mg once or twice daily. In some embodiments, subjects are provided with a dose of about 200, 300, 400, 500, 600, 700, 800, 900, or 1000 mg per day, optionally taken as a single dose or as a divided dose twice daily (e.g., half the amount). In some embodiments, about 450 mg or about 150 mg is provided, optionally twice daily. In certain embodiments, subjects are provided with any of these doses twice daily, optionally orally. In certain embodiments, subjects are provided with this dose once or twice daily. In some embodiments, this dose is divided, and half is administered twice daily. In certain embodiments, the dose comprises a peptide dimer compound disclosed herein, e.g., Compound A or Compound B. In certain embodiments, the dose is optionally provided orally or topically, e.g., rectally, to treat IBD, such as ulcerative colitis.

[0155] In certain embodiments of any of the methods disclosed herein, a subject is provided with any of about 5, 6, 7, 8, 9, 10, 12.5, 25.0, 37.5, 50.0, 62.5, 75, 87.5, or 100.0 mg doses or amounts, optionally twice daily. In some embodiments, a subject is provided with a dose of about 6, 7, 8, 9, 10, 12.5, 25.0, or 37.5 mg. In some embodiments, a subject is provided with a dose ranging from about 5 mg to about 130 mg. In some embodiments, a subject is provided with a dose ranging from about 5 mg to about 50 mg. In some embodiments, a subject is provided with a dose ranging from about 5 mg to about 12.5 mg. In some embodiments, a subject is provided with a dose of about 8 mg. In some embodiments, a subject is provided with a dose of about 150 mg twice daily or a dose of about 450 mg twice daily. In certain embodiments, subjects are provided with any of these doses twice a day, optionally orally. In certain embodiments, subjects are provided with any of these doses once or twice a day. In certain embodiments, it is provided twice a day. In some embodiments, subjects are provided with a dose of about 8 mg. In some embodiments, subjects are provided with a dose of about 150 mg twice a day or a dose of about 450 mg twice a day. In certain embodiments, the dose comprises a peptide dimer compound disclosed herein, such as Compound A or Compound B. In certain embodiments, the dose is provided orally or topically, for example, rectally, for example, by suppository. In some embodiments, subjects are optionally provided with Compound A or Compound B twice a day at a dose of about 150 mg twice a day or a dose of about 450 mg twice a day, orally, for treating IBD, such as ulcerative colitis.

[0156] In certain embodiments of the methods disclosed herein, the method is for treating an individual or subject suffering from an inflammatory disease or disorder. In certain embodiments, the condition is an inflammatory condition of the digestive system. In certain embodiments, the subject is administered or provided with an α4β7 integrin antagonist at a dose or amount that results in a subsaturated blood receptor occupancy rate (RO%). In certain embodiments, the method is carried out using a peptide dimer compound disclosed herein, such as Compound A or Compound B. In certain embodiments, the dose is provided orally or topically, for example, rectally. In certain embodiments, the subject is provided with this dose once or twice daily.

[0157] In certain embodiments, the disease or disorder is selected from the group consisting of inflammatory bowel disease (IBD), adult IBD, pediatric IBD, adolescent IBD, ulcerative colitis, Crohn's disease, celiac disease (non-tropical sprue), seronegative arthropathy-associated enteropathy, microscopic colitis, collagenous colitis, eosinophilic gastroenteritis, radiation therapy, chemotherapy, pouchitis occurring after proctocolectomy and ileoanal anastomosis, gastrointestinal cancer, pancreatitis, insulin-dependent diabetes mellitus, mastitis, cholecystitis, cholangitis, pericholecititis, chronic bronchitis, chronic sinusitis, asthma, primary sclerosing cholangitis, human immunodeficiency virus (HIV) infection in the GI tract, eosinophilic asthma, eosinophilic esophagitis, gastritis, colitis, microscopic colitis, and graft-versus-host disease (GVDH). In certain embodiments, the disease or disorder is IBD. In some embodiments, the IBD is ulcerative colitis. In some embodiments, the IBD is Crohn's disease. In some embodiments, the subject is provided with Compound A or Compound B orally to treat ulcerative colitis or Crohn's disease.

[0158] In certain embodiments, the present disclosure provides a method for treating IBD in a subject in need thereof, comprising orally administering to the subject a peptide dimer compound disclosed herein, e.g., Compound A or Compound B, wherein the compound is administered at a dosage that results in a subsaturated blood receptor occupancy, e.g., an RO of less than 50%. In certain embodiments, the IBD is ulcerative colitis or Crohn's disease. In certain embodiments, the subject is treated with a steroid hormone (SHR) of about 5, 6, 7, 8, 9, 10, 12.5, 25.0, 37.5, 50.0, 62.5, 75, 87.5, 100.0, 112.5, 125.0, 137.5, 150.0, 162.5, 175, 187.5, 200.0, 212.5, 225.0, 237.5, 250.0, 260.0, 270.0, 280.0, 290.0, 310.0, 320.0, 330.0, 340.0, 350.0, 360.0, 370.0, 380.0, 390.0, 410.0, 420.0, 430.0, 440.0, 450.0, 460.0, 470.0, 480.0, 490.0, 510.0, 520.0, 530.0, 540.0, 550.0, 560.0, 570.0, 580.0, 590.0, 610.0, 620.0, 750.0 It may be provided in any of the following doses or amounts: 62.5, 275, 287.5, 300.0, 312.5, 325.0, 337.5, 350.0, 362.5, 375, 387.5, 400.0, 412.5, 425.0, 437.5, 450.0, 462.5, 475, 487.5, or 500.0 mg. In some embodiments, the subject is provided with a dose of about 12.5, 25.0, 37.5, 50.0, 62.5, 75, 87.5, 100.0, 112.5, 125.0, 137.5, 150.0, 162.5, 175, 187.5, 200.0, 212.5, 225.0, 237.5, 250.0, 262.5, 275, 287.5, 300.0, 350.0, 400.0, 450.0, or 500.0 mg. In some embodiments, the subject is provided with a dose of about 70, 75, 80, 85, 90, 95, 100, 105, 110, 115, 120, 125, or 130 mg. In some embodiments, subjects are provided with a dose of about 85, 90, 95, 100, 105, 110, or 115 mg. In some embodiments, subjects are provided with a dose of about 95, 100, or 105 mg. In some embodiments, subjects are provided with a dose of about 100 mg. In some embodiments, subjects are provided with a dose ranging from about 100 mg to about 500 mg, optionally once daily or twice daily.In some embodiments, subjects are provided with a dose ranging from about 200 mg to about 1000 mg, optionally taken as a single dose or in divided doses twice daily (e.g., half the amount). In some embodiments, subjects are provided with a dose ranging from about 100 mg to about 1500 mg per day, optionally taken as a single dose or in divided doses twice daily (e.g., half the amount). In some embodiments, subjects are provided with a dose ranging from about 100 mg to about 1500 mg once or twice daily. In some embodiments, subjects are provided with a dose of any of about 100, 150, 200, 250, 300, 250, 400, 450, or 500 mg once or twice daily. In some embodiments, the subject is provided with a dose of about 200, 300, 400, 500, 600, 700, 800, 900, or 1000 mg per day, optionally taken as a single dose or twice daily in divided doses (e.g., half the amount). In some embodiments, about 450 mg or about 150 mg is provided, optionally twice daily. In some embodiments, the subject is provided with an oral dose of about 150 mg twice daily or about 450 mg twice daily of Compound A or Compound B to treat ulcerative colitis (UC) or Crohn's disease. In certain embodiments, the method is used to treat a subject for ulcerative colitis. In certain embodiments, the subject has moderate to severe active UC. In certain embodiments, the subject has a biopsy-confirmed diagnosis of UC. In certain embodiments, a subject meets one or more (or all) of the inclusion criteria disclosed in the examples and does not meet one or more (or any) of the exclusion criteria disclosed in the examples.

[0159] In certain embodiments, the present disclosure provides a method of treating IBD (e.g., ulcerative colitis or Crohn's disease) in a subject in need thereof, comprising orally administering to the subject a peptide dimer compound disclosed herein, e.g., Compound A or Compound B, wherein the compound is administered at a dosage that results in one or more of the following pharmacokinetic parameters being satisfied in the subject's plasma: Cmax (ng / mL) of 1 to 25, optionally 4 to 12; Tmax (hours) from 1 to 5, optionally from 2 to 4; AUC of 10 to 250, optionally 50 to 150 t (ng.h / mL), AUC of 10–300, optionally 30–250 inf (ng.h / mL), 3~10, optionally 4~10 t 1 / 2 (time), AUC of 30 to 130 tau (ng.h / mL), Ctrough (ng / mL) of 1 to 5; an accumulated Cmax (ng.mL) of 0.5 to 2.5, optionally 2 to 3, and Accumulated AUC between 0.5 and 3.0 t (ng.h / mL).

[0160] In certain embodiments of these methods, IBD is ulcerative colitis or Crohn's disease.In certain embodiments, pharmacokinetic parameters are met within 1 hour, 2 hours, 3 hours, 4 hours, 6 hours, 8 hours, or 12 hours after administration.In certain embodiments, pharmacokinetic parameters are maintained for at least 1 hour, at least 2 hours, at least 3 hours, at least 4 hours, at least 6 hours, at least 8 hours, or at least 12 hours after administration.

[0161] In certain embodiments, the present disclosure provides a method of treating IBD in a subject in need thereof, comprising orally administering to the subject a peptide dimer compound disclosed herein, e.g., Compound A or Compound B, wherein the compound is administered at a dosage that results in one or more of the following pharmacodynamic parameters in the subject's plasma: 50~100, optionally, 90~100 ROmax(%); 50-95, optionally 65-95 average RO(%); -20 to -60, optionally -35 to -60 receptor expression max Change in (%), mean change (%) in receptor expression from -10 to -55, optionally from -25 to -55; Steady-state ROmax (%) of 80-100; Average RO of 75-90 0-24 (h%), Average RO of 80-95 0-12 (h%), and Average RO of 70-90 12-24 (h%).

[0162] In certain embodiments of these methods, IBD is ulcerative colitis or Crohn's disease.In certain embodiments, pharmacodynamic parameters are met within 1 hour, 2 hours, 3 hours, 4 hours, 6 hours, 8 hours, or 12 hours after administration.In certain embodiments, pharmacodynamic parameters are maintained for at least 1 hour, at least 2 hours, at least 3 hours, at least 4 hours, at least 6 hours, at least 8 hours, or at least 12 hours after administration.

[0163] In certain embodiments, the methods disclosed herein reduce (partially or completely) the activity of α4β7 in a subject. In certain embodiments, the methods reduce the proliferation of T cells containing α4β7 integrin, e.g., T cells present in the subject's gastrointestinal tissue, e.g., the gastrointestinal mucosa. In further embodiments, the methods inhibit the production or release of cytokines by T cells in the subject, e.g., T cells in the subject's gastrointestinal tissue, e.g., β7+ T cells. In certain embodiments, the methods reduce the production or release of any of the cytokines disclosed in the accompanying figures, e.g., IFN-gamma, interleukin-6 (IL-6), IL-8, IL-12 / 23p40, IL-15, IL-16, IL-13, vascular endothelial growth factor (VEGF), granulocyte-macrophage colony-stimulating factor (GM-CSF), tumor necrosis factor alpha (TNFα), or tumor necrosis factor β (TNFβ). In certain embodiments, the methods disclosed herein inhibit the production or release of cytokines by T cells, the release of which is promoted by binding to mucosal vascular addressin cell adhesion molecule 1 (MAdCAM1) in a gastrointestinal tissue, such as the gastrointestinal mucosa, in a subject. In certain embodiments, the T cells are CD45RO- naive or CD45RO+ memory T cells. In certain embodiments, the T cells are β7 + is.

[0164] In further related embodiments, the present invention includes methods for treating a subject, e.g., a mammal or human, suffering from a condition associated with the biological function α4β7, comprising providing or administering to the subject a peptide molecule described herein in an amount sufficient to inhibit (partially or completely) the biological function of α4β7 in tissues that express MAdCAM1, e.g., gastrointestinal tissues such as the gastrointestinal mucosa. In certain embodiments, the subject is provided with an effective amount of a peptide monomer or peptide dimer sufficient to at least partially inhibit the biological function of α4β7 in tissues that express MAdCAM1. In certain embodiments, the condition is inflammatory bowel disease.

[0165] In additional embodiments, the present invention includes a method of treating or preventing a disease or condition in a subject in need thereof, comprising providing or administering to the subject, e.g., a mammal, an effective amount of a peptide dimer or peptide monomer as described herein, wherein the disease or condition is inflammatory bowel disease (IBD) (including adult IBD, pediatric IBD, and adolescent IBD), ulcerative colitis, Crohn's disease, celiac disease (non-tropical sprue), seronegative arthropathy-associated enteropathy, microscopic colonic ulcers, or ulcerative colitis. In certain embodiments of the methods of treatment described herein, the subject is diagnosed with or is at risk of developing one of these diseases or conditions.

[0166] In certain embodiments of any of the methods of treatment described herein, the peptide molecule (or pharmaceutical composition comprising the peptide molecule) is administered to the individual by a mode of administration selected from the group consisting of oral, intravenous, peritoneal, intradermal, subcutaneous, intramuscular, intrathecal, inhalation, vaporization, spray, sublingual, buccal, parenteral, rectal, vaginal, and topical.

[0167] In certain embodiments, the present disclosure provides a steroid hormone therapy comprising a steroid hormone therapy comprising at least about 5, 6, 7, 8, 9, 10, 12.5, 25.0, 37.5, 50.0, 62.5, 75, 87.5, 100.0, 112.5, 125.0, 137.5, 150.0, 162.5, 175, 187.5, 200.0, 212.5, 225.0, 237.5, 250.0, 262.5, 275, 28 Provided are unit dose forms of the peptide dimer compounds disclosed herein comprising any of 7.5, 300.0, 312.5, 325.0, 337.5, 350.0, 362.5, 375, 387.5, 400.0, 412.5, 425.0, 437.5, 450.0, 462.5, 475, 487.5, or 500.0 mg. In some embodiments, the unit dose form contains about any of 12.5, 25.0, 37.5, 50.0, 62.5, 75, 87.5, 100.0, 112.5, 125.0, 137.5, 150.0, 162.5, 175, 187.5, 200.0, 212.5, 225.0, 237.5, 250.0, 262.5, 275, 287.5, 300.0, 350.0, 400.0, 450.0, or 500.0 mg. In some embodiments, the unit dose form contains about any of 70, 75, 80, 85, 90, 95, 100, 105, 110, 115, 120, 125, or 130 mg. In some embodiments, the unit dose form comprises about 85, 90, 95, 100, 105, 110, or 115 mg. In some embodiments, the unit dose form comprises about 95, 100, or 105 mg. In some embodiments, the unit dose form comprises about 100 mg. In some embodiments, the unit dose form comprises about 100-500 mg. In some embodiments, the unit dose form comprises about 100, 150, 200, 250, 300, 250, 400, 450, or 500 mg. In some embodiments, the unit dose form comprises about 450 mg or about 150 mg. In certain embodiments, the unit dose form comprises a pharmaceutical composition comprising a peptide dimer compound, e.g., any of those disclosed herein. In certain embodiments, it is formulated for oral administration, e.g., as a tablet.In certain embodiments, it is formulated for rectal administration, for example, as a suppository. In some embodiments, the unit dose form comprises about 450 mg or about 150 mg of Compound A or Copound B (or a pharmaceutically acceptable salt thereof). In certain embodiments, the unit dose form comprises a pharmaceutical composition comprising a peptide dimer compound, for example, any of those disclosed herein.

[0168] In certain embodiments, the peptide molecules of the present invention are present in pharmaceutical compositions further comprising one or more pharmaceutically acceptable diluents, carriers, or excipients. In certain embodiments, they are formulated as liquids or solids. In certain embodiments, they are formulated as tablets or capsules, or as liquid suspensions. Some embodiments of the present invention further provide a method for treating an individual with the α4β7 integrin antagonist peptide molecules of the present invention suspended in a sustained-release matrix. As used herein, a sustained-release matrix is ​​a matrix made of a material (usually a polymer) that can be degraded by enzymatic hydrolysis or acid-base hydrolysis, or by dissolution. When inserted into the body, enzymes and body fluids act on this matrix. The sustained-release matrix is ​​desirably selected from biocompatible materials such as liposomes, polylactide (polylactic acid), polyglycolide (polymer of glycolic acid), polylactide-co-glycolide (copolymer of lactic acid and glycolic acid), polyanhydrides, poly(ortho)esters, polypeptides, hyaluronic acid, collagen, chondroitin sulfate, carboxylic acids, fatty acids, phospholipids, polysaccharides, nucleic acids, polyamino acids, amino acids such as phenylalanine, tyrosine, isoleucine, polynucleotides, polyvinylpropylene, polyvinylpyrrolidone, and silicones. In particular, the biodegradable matrix is ​​a matrix of either polylactide, polyglycolide, or polylactide-co-glycolide (copolymer of lactic acid and glycolic acid).

[0169] In some aspects, the present invention provides pharmaceutical compositions for oral delivery. Various embodiments and peptide molecule compositions of the present invention can be prepared for oral administration according to any of the methods, techniques, and / or delivery vehicles described herein. Furthermore, those skilled in the art will understand that the peptide molecule compositions of the present invention can be modified or integrated into systems or delivery vehicles not disclosed herein but known in the art and adapted for use in oral delivery of small peptide molecules.

[0170] Oral dosage forms or unit doses suitable for use with the peptides of the present invention may include a mixture of peptide active drug components and non-drug components or excipients, as well as other non-recyclable materials that may be considered either components or packaging. Oral compositions may include at least one of liquid, solid, and semi-solid dosage forms. In some embodiments, oral dosage forms are provided that include an effective amount of the peptide molecules described herein, and the dosage form includes at least one of pills, tablets, capsules, gels, pastes, beverages, and syrups. In some examples, oral dosage forms are provided that are designed and configured to achieve delayed release of the thioether peptide molecules in the small intestine of a subject.

[0171] In one embodiment, oral pharmaceutical compositions comprising the peptides of the present invention comprise an enteric coating designed to delay release of the peptide molecules in the small intestine. In some instances, it is preferred that the pharmaceutical compositions of the present invention comprise an enteric coating that is soluble in gastric juice at a pH of about 5.0 or higher. In at least one embodiment, pharmaceutical compositions are provided that comprise an enteric coating comprising a polymer with a dissociable carboxyl group, such as derivatives of cellulose, including hydroxypropylmethylcellulose phthalate, cellulose acetate phthalate, and cellulose acetate trimellitate, and similar derivatives of cellulose and other carbohydrate polymers.

[0172] In one embodiment, a pharmaceutical composition comprising a peptide molecule described herein is provided within an enteric coating, which is designed to protect and release the pharmaceutical composition in a controlled manner within the subject's lower gastrointestinal system and to avoid systemic side effects. In addition to enteric coatings, the peptide molecules of the present invention can be encapsulated, coated, engaged, or otherwise associated with any suitable oral drug delivery system or component. For example, in some embodiments, the peptide molecules of the present invention are provided within a lipid carrier system, including at least one of polymer hydrogels, nanoparticles, microparticles, micelles, and other lipid systems.

[0173] To overcome peptide degradation in the small intestine, some implementations of the present invention include a hydrogel polymer carrier system in which the peptide molecules of the present invention are housed, whereby the hydrogel polymer protects the peptide from proteolytic degradation in the small intestine. The peptide molecules of the present invention can be further formulated for use with carrier systems designed to increase the dissolution kinetics of the peptide and enhance intestinal absorption. These methods include the use of liposomes, micelles, and nanoparticles to increase peptide penetration into the GI tract.

[0174] Various bioreactive systems may also be combined with one or more thioether peptide molecules of the present invention to provide pharmaceutical agents for oral delivery. In some embodiments, the peptide molecules of the present invention are used in combination with bioreactive systems, such as hydrogels, mucoadhesive polymers with hydrogen-bonding groups (e.g., PEG, poly(methacrylic) acid [PMAA], cellulose, Eudragit®, chitosan, alginate), and the like, to provide therapeutic agents for oral administration. Other embodiments include methods for optimizing or extending the drug residence time of the peptide molecules disclosed herein, in which the surface of the peptide molecule is modified to contain mucoadhesive properties through hydrogen bonding, linked mucin-bearing polymers, or / and hydrophobic interactions. These modified peptide molecules may demonstrate increased drug residence time within a subject, in accordance with a desired feature of the present invention. Furthermore, targeted mucoadhesive systems may specifically bind to receptors on the surface of enterocytes and M-cells, thereby further enhancing the uptake of particles containing the peptide molecules.

[0175] Other embodiments include methods for oral delivery of peptide molecules described herein, in which the peptide molecules are used in combination with a penetration enhancer that promotes transport of the peptide across the intestinal mucosa by increasing paracellular or transcellular permeation. For example, in one embodiment, a penetration enhancer is combined with a peptide molecule described herein, the penetration enhancer comprising at least one of a long-chain fatty acid, a bile salt, an amphiphilic surfactant, and a chelating agent. In one embodiment, a penetration enhancer comprising sodium N-[(hydroxybenzoyl)amino]caprylate is used to form a weak non-covalent association with the peptide molecule of the present invention, which facilitates membrane transport and further dissociation upon reaching the blood circulation. In another embodiment, the peptide molecule of the present invention is conjugated to oligoarginine, thereby increasing the cellular penetration of the peptide into various cell types. Furthermore, in at least one embodiment, a non-covalent bond is provided between the peptide molecule described herein and a penetration enhancer selected from the group consisting of cyclodextrins (CDs) and dendrimers, where the penetration enhancer reduces peptide aggregation and increases the stability and solubility of the peptide molecule.

[0176] When used in at least one of the treatments or delivery systems described herein, a therapeutically effective amount of the peptide molecule of the present invention can be used in pure form, or, if such a form exists, in the form of a pharmaceutically acceptable salt. As used herein, a "therapeutically effective amount" of a compound of the present invention is intended to describe a sufficient amount of the peptide molecule to treat an integrin-related disease (e.g., reduce inflammation associated with IBD) at a desired benefit / risk ratio applicable to any medical treatment. However, it will be understood that the total daily usage of the compounds and compositions of the present invention will be determined by the attending physician within the scope of sound medical judgment. The specific therapeutically effective dose level for any particular patient will depend on various factors, including: a) the disorder being treated and the severity of the disorder; b) the activity of the specific compound used; c) the specific composition used; the patient's age, weight, general health, sex, and diet; d) the time of administration, route of administration, and excretion rate of the specific compound used; e) the duration of treatment; f) drugs used in combination with or concurrently with the specific compound used, and similar factors known in the medical art.

[0177] Alternatively, the compound of the present invention can be administered as a pharmaceutical composition containing the peptide molecule of interest in combination with one or more pharmaceutically acceptable excipients. Pharmaceutically acceptable carriers or excipients refer to any type of non-toxic solid, semi-solid, or liquid filler, diluent, encapsulating material, or formulation aid. The composition can be administered parenterally, intracisternally, intravaginally, intraperitoneally, intrarectally, topically (by powder, ointment, drops, suppository, or transdermal patch, etc.), rectally, or orally. As used herein, the term "parenteral" refers to modes of administration including intravenous, intramuscular, intraperitoneal, intrasternal, subcutaneous, intradermal, and intraarticular injection and infusion.

[0178] Compositions for rectal or vaginal administration are preferably suppositories which can be prepared by mixing a compound of the present invention with a suitable non-irritating excipient or carrier such as cocoa butter, polyethylene glycol, or a suppository wax which is solid at room temperature but liquid at body temperature and therefore melts in the rectum or vaginal cavity and releases the active compound.

[0179] The total daily dose of the compositions of this invention to be administered to a human or other mammalian host in single or divided doses may be in the range of, for example, 0.0001 to 300 mg / kg, and more usually 1 to 300 mg / kg, of body weight per day. [Example]

[0180] Example 1 Compound A blocks MAdCAM1-mediated CD4+ T cell proliferation. Compound A, an oral gastrointestinal (GI)-restricted peptide antagonist of the α4β7 integrin, is being developed for the treatment of inflammatory bowel disease (IBD). Blockade of α4β7 binding to the mucosal addressin cell adhesion molecule-1 (MAdCAM1) is thought to treat IBD by preventing vascular T cell extravasation into the inflamed GI mucosa. To further explore the mechanism by which Compound A reduces GI inflammation, the following experiments were performed. Specifically, MAdCAM1-mediated CD4 + The potential local GI function of α4β7 was assessed by evaluating the ability of Compound A to inhibit T cell proliferation and cytokine production.

[0181] Compound A: TIFF2025186365000038.tif68128 ((2-benzyl)-(N-Me-R)-Ser-Asp-Thr-Leu-Pen-(Phe(4-tBu))-(β-homo-Glu)-(D-Lys)-OH)2 (SEQ ID NO: 5) and linker-DIG diglycolic acid.

[0182] PBMCs were purified from healthy human donors and enriched for CD4+ T cells. +T cells were fluorescently labeled and incubated for 3 days with plate-bound anti-CD3 alone or MAdCAM1 with or without inhibitors (or negative controls): Compound A (1 μM), an inactive analog (1 μM) as a negative control, or vedolizumab (500 ng / mL). Analysis of phenotype, distribution of T helper (Th) subsets, and RO% was performed by flow cytometry of freshly stained raw samples.

[0183] MAdCAM1 in combination with anti-CD3 significantly reduced CD4 T cell proliferation compared to anti-CD3 alone after 3 days of incubation. + Compound A significantly enhanced T cell proliferation (n=7, 12-87%) (Figure 1). Compound A completely abolished MAdCAM1-mediated proliferation (Figure 1). The level of inhibition was similar to that by vedolizumab (Figure 1). Blockade was not observed with an inactive analog (negative control, NEG), indicating dependence on Compound A binding to α4β7. Inhibition by Compound A was dependent on the activity of Compound A. Inhibition by Compound A was concentration-dependent. Average IC from four independent human donors 50 was 4.4 nM (Table 4).

[0184] Table 4: IC50 from four donors TIFF2025186365000039.tif36128

[0185] Immunophenotyping was performed using CD45RO - Naive and CD45RO + We found that proliferation of both naive and memory T cells occurred, shifting naive T cells to a memory cell phenotype (Figure 2). + The β7 surface expression was restricted to the undivided CD4 population, and successive cycles of proliferation showed an increase in β7 expression (Figure 3A). + T cells exhibited reduced internalization with Compound A (Figure 3B and Figure 4, tested in five donors). Among expanded memory T cells, the proportion of IFNγ-producing Th1 subsets was higher than the IL-17A-producing Th17 and IL-4-producing Th2 subsets (Table 5).

[0186] Table 5. Characteristics of expanded CD4+ T cells TIFF2025186365000040.tif36160

[0187] The α4β7-MAdCAM1 interaction is + CD4 + It promotes T cell proliferation and cytokine release, which may contribute to the chronic inflammatory response that occurs in the diseased gut of IBD patients, independently of T cell trafficking. Compound A's inhibition of MAdCAM1-mediated signaling through α4β7 supports the potential therapeutic benefit of an oral GI restriction approach, whereby Compound A is delivered locally to directly block α4β7 function within the GI tract.

[0188] Example 2 Compound A blocks MAdCAM1-mediated cytokine production. To further explore the mechanism by which Compound A reduces GI inflammation, the following experiments were performed. In particular, cytokine profiling was performed on T cells isolated from normal, healthy donors.

[0189] PBMCs were purified from three healthy human donors (donors 7, 10, and 11) and enriched for CD4+ T cells. + T cells were fluorescently labeled and incubated with plate-bound anti-CD3 alone, plate-bound anti-CD3 with MAdCAM1, or plate-bound anti-CD3 with MAdCAM1 and varying amounts of Compound A. Supernatant cytokine levels were quantified by MSD or Luminex platform multiplex assays for anti-CD3 alone and anti-CD3 + MAdCAM1 in the presence of varying concentrations of Compound A.

[0190] Multiplex profiling identified several cytokines, including IFNγ, IL-5, IL-6, IL-10, IL-13, GM-CSF, and TNFα, whose release was promoted by MAdCAM1 (Figures 5A-C and 6A-C). MAdCAM1-mediated cytokine production was inhibited by Compound A in a concentration-dependent manner. Concentration-dependent and complete inhibition of MAdCAM1-mediated production of specific cytokines by Compound A is shown in Figures 5A-C and 6A-C. α4β7-MAdCAM1 interaction promotes β7 + CD4 + It promotes T cell proliferation and cytokine release, which may contribute to the chronic inflammatory response that occurs in the diseased gut of IBD patients, independently of T cell trafficking. Compound A's inhibition of MAdCAM1-mediated signaling through α4β7 supports the therapeutic benefit of an oral GI restriction approach, whereby Compound A is delivered locally to directly block α4β7 function within the GI tract.

[0191] Example 3 Receptor occupancy in mice The next experiment was performed to examine receptor occupancy in whole blood and Peyer's patches in mice dosed with Compound B, an analog of Compound A.

[0192] Compound B: TIFF2025186365000041.tif71133 Linker = (Ac-Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-(Phe(4-tBu))-(β-homo-Glu)-(D-Lys)-NH2)2 (SEQ ID NO: 6) with DIG (diglycolic acid)

[0193] Three groups of female C57BL / 6 mice (N = 6 per group) were orally treated with either vehicle (Group 1), Compound B (3 mg / kg PO QD, Group 2), or Compound B (30 mg / kg PO QD, Group 3). One hour after dosing, mice were euthanized, and whole blood / plasma and Peyer's patches were collected. Peyer's patches were dispersed in 1 mL of RPMI medium containing 2% FBS without a washing step. Single-cell suspensions of whole blood and Peyer's patches (100 μL of the total 1 mL) were submitted for flow cytometry to determine α4β7 receptor occupancy. RO% = (1 - (% positive test samples / median % positive vehicle controls) * 100). Plasma and dispersed Peyer's patches in the single-cell suspension (500 μL of the total 1 mL) were also analyzed for drug exposure.

[0194] In both dose groups, receptor occupancy in Peyer's patches was significantly higher compared to whole blood (Figure 7). Receptor occupancy levels in whole blood or Peyer's patches were comparable between dose groups. At both the 3 mg / kg and 30 mg / kg doses, there was competitive (100%) receptor occupancy in Peyer's patches in some animals (Figure 8, top). RO% for various doses of Compound B is shown in Figure 8, bottom. PK and PD data are shown in Figure 9, where the doses of Compound B are indicated.

[0195] Similar experiments were performed using Compound A. Receptor occupancy was significantly higher in Peyer's patches compared to whole blood in both dose groups. Compared to the 3 mg / kg dose, the 30 mg / kg dose resulted in significantly higher receptor occupancy in whole blood (P<0.01) and Peyer's patches (P<0.001) (Figure 25). The effect was similar to that observed with Compound B, although quantitatively less significant. The same dose was used, potentially due to Compound A having greater activity than Compound B. There was also a significant dose-dependent increase in Compound A concentrations in both plasma and Peyer's patches (Figure 26). Compound A concentrations were significantly higher in Peyer's patches than in plasma at both dose levels. As shown in Figure 27, there was a dose-dependent increase in receptor occupancy in whole blood and Peyer's patches in animals dosed with Compound A, with complete (100%) receptor occupancy in Peyer's patches in animals at the 30 mg / kg dose.

[0196] Example 4 Tissue exposure of Compound A in mice This study was conducted to determine plasma, Peyer's patches (PP), and mesenteric lymph nodes (MLN), small intestine, and colon tissue exposure of Compound A following PO administration in healthy C57BL / 6 female mice.

[0197] Twelve treatment-naive C57BL / 6 female mice were assigned to the study. Animals were fasted overnight and administered a single dose of 30 mg / kg Compound A by oral gavage (PO) at a dose volume of 10 mL / kg. At 1, 3, and 6 hours (h) post-dose, four mice per time point were subjected to terminal bleeding, euthanized, and Peyer's patches (PPs), mesenteric lymph nodes (MLNs), small intestine, and colon were collected from each animal. Blood was processed to plasma, and plasma and tissue samples were submitted for pharmacokinetic (PK) analysis of Compound A levels using a qualified liquid chromatography-tandem mass spectrometry (LC-MS / MS) method.

[0198] Compound A concentrations in plasma and tissues were analyzed using a qualified liquid chromatography-tandem mass spectrometry (LC-MS / MS) method. Processed plasma and tissue samples were analyzed on an AB / MDS Sciex API4000 mass spectrometer. Positive ions were monitored in multiple reaction monitoring (MRM) mode. Quantification was performed by peak area ratio.

[0199] PK data analysis was performed using non-compartmental analysis (NCA) in Phoenix WinNonlin 8.1 (Certara USA Inc.). All concentration values ​​below the lower limit of quantification were treated as zero in the pharmacokinetic analysis. The maximum concentration (C max ) and C max Apparent time to (t max ) were obtained by observation. Area under the concentration versus time curve (AUC) was obtained by the linear trapezoidal method. All concentration data and PK parameters were reported to a maximum of three significant figures if values ​​were greater than 1 and to a maximum of three decimal places if values ​​were less than 1. Time parameters were reported to a maximum of two decimal places. Concentration data were plotted using Excel (Microsoft).

[0200] The mean plasma and tissue concentrations of Compound A in each animal are plotted in Figure 10. The resulting PK parameters are shown in Figure 11. After a single PO dose of 30 mg / kg, peak Compound A exposure was observed at 1 hour in the MLN, PP, and small intestine, at 3 hours in plasma, and at 6 hours in the colon. The mean Cmax value was greatest in the small intestine (13,300 ng / g), with mean values ​​approximately half that observed in the PP and colon. These gastrointestinal levels were much higher (more than 100-fold) than those in plasma (19.0 ng / mL) and MLN (56.8 ng / g). Similarly, the mean AUC values ​​in the small intestine, PP, and colon (48,600, 37,900, and 15,700 ng / g, respectively) were much higher (more than 60-fold) than those in plasma (95.4 ng / mL) and MLN (226 ng / g). These results demonstrated that Compound A had limited plasma and lymph node exposure when administered PO to otherwise healthy female mice. Dose analysis (data not shown) demonstrated that the dosing solution was 97.6% of the nominal concentration.

[0201] Example 5 Compound A inhibits gut homing of cultured T cells. T cells cultured in the presence of all-trans retinoic acid (ATRA) upregulate the gut-homing receptors CCR9, integrin α4 (α4), and integrin β7 (β7), and preferentially home to intestinal tissues (ileal lamina propria and Peyer's patches). The aim of this study was to analyze the gut homing of T cells cultured in the presence of compound A.

[0202] Purified CD3+ cells were isolated from B6.SJL (CD45.1+) donor mice and cultured in the presence of anti-CD3 / anti-CD28 beads and IL-2 to induce T cell activation and proliferation. In some culture conditions, Compound A and / or ATRA were added. To track cells in vivo, ATRA- and ATRA+ cells were labeled with CMFDA and CTFR, respectively. These labeled cells were then co-injected into C57BL / 6 (CD45.2+) recipient mice. There were four groups of recipient mice in this study. Vehicle (negative control) Anti-VLA-4 (in vivo treatment with anti-VLA-4 as a positive control) Compound A, 100 nM (test, in culture) Compound A, 1000 nM (testing, in culture)

[0203] The percentages of ATRA- and ATRA+ cells in the spleen, Peyer's patches (PP), and ileal lamina propria (LP) of recipient mice were measured by flow cytometry to assess cell homing.

[0204] As expected, cells cultured in the presence of ATRA+ / DMSO ("ATRA+ / DMSO cells") had a higher percentage of cells expressing the gut-homing receptor CCR9 and integrins α4 and β7 than cells cultured in the absence of ATRA ("ATRA- cells"). ATRA+ / Compound A cells had a lower percentage of integrin β7+ cells than ATRA+ / DMSO cells. The spleens of vehicle-treated mice contained a higher percentage of ATRA- than ATRA+ cells, as expected for this group, and the LPs of these mice contained a higher percentage of ATRA+ than ATRA- cells, confirming that ATRA+ cells preferentially homed to the intestine. Compared with vehicle-treated mice, anti-VLA-4-treated mice had approximately 10-fold lower percentages of ATRA+ cells in the LP and approximately 2-fold lower percentages of ATRA+ cells in the PP. These results confirmed that anti-VLA-4 treatment reduced the gut homing of ATRA+ cells, as expected for this positive control. The Compound A 1000 nM group had significantly lower percentages of CD45.1+ cells in the spleen and LP than the vehicle group. In addition, both Compound A groups had lower percentages of ATRA+ cells in the LP compared to vehicle, and the reduction approached statistical significance for the 1000 nM group.

[0205] method: Eighteen B6.SJL (CD45.1+) donor mice were acclimated for 3-9 weeks prior to the start of the study and were 11-16 weeks old at the time of culture setup (day 0). On day 0, spleen and lymph node cells were isolated from donor mice and pooled. CD3+ cells were enriched using STEMCELL Technologies kit catalog number 19851. The purity of the enriched cells was confirmed by flow cytometry.

[0206] Approximately 44% of the cells were then cultured at 1.5 × 10 cells / well in the presence of anti-CD3 / CD28 beads (Dynabeads, ThermoFisher 11453D) at a 1:1 cell-to-bead ratio. 6 The remaining cells were cultured at a cell concentration of 2 × 10 6The cells were cultured under the same conditions, except that either Compound A or DMSO was added to the cultures at a concentration of 0.1 μM. All-trans retinoic acid (ATRA) was then added to these cultures at a concentration of 0.1 μM. Table 6 below summarizes the culture conditions.

[0207] (Table 6) Culture conditions TIFF2025186365000042.tif77163

[0208] On day 1, IL-2 was added to all cultures to reach a concentration of 30 U / mL.

[0209] From day 2 to day 4, cultures were grown as needed by adding fresh medium while maintaining the following concentrations: 1000 mg / kg of sucrose; 30 U / mL IL-2 for all cultures For ATRA+ cultures, 0.1 μM ATRA 0.1% DMSO in ATRA+ cultures Compound A listed in Table 4

[0210] On day 5, cells from each culture were stained and analyzed by flow cytometry using the reagents listed in Table 7.

[0211] Table 7. Flow cytometry panel for assessment of gut-homing receptor expression TIFF2025186365000043.tif41128

[0212] Thirty-eight C57BL / 6 (CD45.2+) recipient mice were acclimated for 9 weeks before the start of the study (day 0) and were 16 weeks old at the time of cell transfer (day 5). On day 4, recipient mice were assigned to groups in a balanced manner to achieve similar mean body weights across groups.

[0213] On day 5, after removing CD3 / CD28 beads from the cell culture with a magnet, ATRA+ cells were labeled with CFTR, and ATRA- cells were labeled with CMFDA. Then, cells from each culture condition were counted. For each group, ATRA- cells and cells from one of the ATRA+ culture conditions were mixed at a 1:1 ratio according to Table 8 below, and then transferred into recipient mice. Approximately 13 million cells of each type (total of 26 million) were intravenously injected into each mouse.

[0214] Table 8: Treatment regimens Mice in group 2 were dosed once with anti-VLA-4 on day 5 prior to cell transfer. Anti-VLA-4 (PS / 2) antibody was purchased from BioXCell and kept at -80°C until needed. The antibody was diluted in sterile PBS to a final concentration of 1 mg / mL and dosed intraperitoneally at 10 mg / kg. No in vivo treatment was administered to other groups.

[0215] 20-22 hours after cell transfer, all mice were euthanized, and their blood, spleen, Peyer's patches, and small intestine were collected. Approximately 50 µL of plasma was isolated from each mouse's blood and stored on a dry incubator until further analysis.

[0216] For flow cytometry analysis, cells from the following tissues were isolated from each mouse: Spleen Peyer's patches Ileal lamina propria The isolated cells were counted and stained with anti-CD45.1 antibody and live / dead stain, and then acquired for flow cytometry analysis to determine the percentage of CD45.1+, ATRA+, and ATRA- cells in each tissue.

[0217] At the end of the culture period, flow cytometry analysis showed that a much smaller proportion of ATRA+ / DMSO cells than ATRA− cells expressed the gut-homing receptor CCR9 and integrins α4 and β7 (Figures 12 and 13). These findings confirmed that, as expected, cells cultured in the presence of ATRA upregulated gut-homing receptors. ATRA+ / Compound A cultures contained a smaller proportion of integrin β7+ cells than ATRA+ / DMSO cultures (Figure 12). Expression of integrin α4 was lower in ATRA+ / DMSO cultures compared with ATRA+ / Compound A cultures, whereas CCR9 expression appeared unaffected (Figure 13). These results indicated that Compound A inhibited upregulation, downregulated expression, or interfered with the detection of integrins β7 and α4. The results of tissue homing analysis are shown in Tables 9-13.

[0218] Table 9. Total number of isolated cells (x10 3 ) TIFF2025186365000045.tif50149*p<0.05 vs. vehicle

[0219] (Table 10) CD45.1+ cells / 10 3 living cells TIFF2025186365000046.tif50149*p<0.05 vs. vehicle

[0220] Table 11. ATRA+ and ATRA- cells / 10 in the spleen 3 living cells TIFF2025186365000047.tif30158*p<0.05 vs. vehicle

[0221] Table 12. ATRA+ and ATRA- cells / 10 in Peyer's patches 3 living cells TIFF2025186365000048.tif30158*p<0.05 vs. vehicle

[0222] Table 13. ATRA+ and ATRA- cells / 10 in ileal LP 3 living cells TIFF2025186365000049.tif30158*p<0.05 vs. vehicle **p<0.10

[0223] The number of cells isolated from the spleen, Peyer's patches, and ileal lamina propria (LP) in the vehicle group was as expected (Table 9), and the percentage of CD45.1+ cells isolated from these tissues was as expected for this model (Table 10).

[0224] Spleens from the vehicle group contained a greater proportion of ATRA- than ATRA+ cells (Table 11), whereas the lamina propria (LP) contained a greater proportion of ATRA+ than ATRA- cells (Table 13), confirming that ATRA+ cells preferentially homed to the intestine, as expected for this group.

[0225] The anti-VLA-4 group had approximately 1 / 10 the rate of ATRA+ cells in the LP and approximately 1 / 2 the rate of ATRA+ cells in the PP compared to vehicle mice (Tables 13 and 12), confirming that treatment reduced gut homing of ATRA+ cells, as expected for this positive control.

[0226] The anti-VLA-4 group had significantly fewer cells isolated from Peyer's patches and ileal lamina propria than the vehicle group (Table 9), which is typically observed in anti-VLA-4 treated mice, particularly for Peyer's patches.

[0227] The percentage of ATRA cells in the spleen of this group was significantly higher than that of the vehicle group, which is often observed in anti-VLA-4 treated mice and may be due to ATRA cells being blocked from homing to the intestine and consequently accumulating in the spleen.

[0228] The percentage of ATRA+ cells in the LP of mice from the Compound A group was found to be smaller than that of the vehicle group. This reduction was dose-dependent and approached statistical significance for cells treated with 1000 nM Compound A.

[0229] The Compound A, 1000 nM group had significantly fewer cells isolated from the spleen than the vehicle group, while both the 100 nM and 1000 nM groups had significantly fewer cells isolated from Peyer's patches (Table 9).

[0230] The Compound A, 1000 nM group also had a significantly lower percentage of CD45.1+ cells in the spleen and LP than the vehicle group (Table 10).

[0231] Overall, these results suggest that Compound A-treated cells have impaired homing to intestinal tissue.

[0232] Example 6 Compound A inhibits the upregulation of integrin B7. A flow cytometry-based in vitro assay was used to evaluate the internalization activity of peptide Compound A. This study showed that Compound A specifically triggered the internalization of α4β7 in human primary cells in a time- and dose-dependent manner. Compound A also triggered a reduction in α4β7 expression, resulting in CD4 + This led to a decrease in adhesion to MAdCAM1 by T memory cells, with an average decrease of up to 39% observed in α4β7 expression and an average decrease of up to 37% in adhesion to MAdCAM1. Furthermore, Compound A expression recovered to control levels after 5 days of additional incubation following removal of Compound A.

[0233] method Blood samples from human donors were obtained from the Stanford Blood Center (Stanford, CA) under an IRB-approved research protocol. Blood was drawn into BD Vacutainer sodium heparin blood collection tubes (BD Biosciences, catalog number 362753). Peripheral blood mononuclear cells (PBMCs) were isolated from the blood using SepMate-50 tubes and LymphoPrep according to the manufacturer's protocol. After PBMC isolation, CD4 + T memory cells were enriched.

[0234] To determine specificity, human PBMCs were incubated with either 100 nM Compound C (an analog of Compound A), Compound D (an inactive triple mutant peptide analog of Compound A), or no peptide in complete culture medium for 24 hours at 37° C. After incubation, an aliquot of cells from each reaction was stained for α4β7 expression.

[0235] To determine time and dose-dependent responses, purified human CD4 + T memory cells were incubated with either 10 nM Compound A for different time ranges (0, 1, 2, 4, 6, 24, 28, 30, and 48 hours) or various concentrations (0, 0.01, 0.1, 1, and 10 nM) of Compound A in complete culture medium for 24 hours at 37° C. After incubation, an aliquot of cells from each reaction was stained for α4β7 expression.

[0236] Purified human CD4 was used to determine its effect on α4β7 expression and MAdCAM1 adhesion. +T memory cells were incubated with various concentrations of Compound A (0, 0.01, 0.1, 1, and 10 nM) in complete culture medium for 2 hours at 37°C. After incubation, cells were washed extensively to remove excess peptide. For each reaction, an aliquot of cells was stained for α4β7 expression, while a separate aliquot was tested for adhesion to MAdCAM1.

[0237] To determine recovery after washout, human PBMCs were incubated with 10 nM Compound A in complete culture medium (without MnCl2) for 24 hours at 37°C. Aliquots of cells were collected before and 24 hours after peptide addition and stained for α4β7 expression. Cells were then thoroughly washed to remove excess peptide and incubated in fresh complete culture medium (without MnCl2) for an additional 7 days. After peptide washout, aliquots were stained for α4β7 expression on days 1, 2, 4, 5, and 7.

[0238] After peptide incubation, an aliquot of each reaction was stained for surface expression of α4β7 in preparation for flow cytometry. Cells were stained for 30 min at 4°C, washed twice in DPBS containing 0.5% BSA (PBS / BSA), incubated with streptavidin BV421 (1:1000 dilution) for 30 min at 4°C, washed twice in PBS / BSA, and then resuspended in PBS / BSA for analysis. Where relevant, "fluorescence minus one" (FMO) samples were used as staining controls.

[0239] Samples were analyzed by flow cytometry on a BD (Franklin Lakes, NJ) FACS Verse flow cytometer equipped with the following lasers: 405 nm (violet), 488 nm (blue), 561 nm (yellow-green), and 640 nm (red). + T memory cells are CD4 + , CD45RA - , CD197 + Identified as lymphocytes. CD4 +α4β7 expression in T memory cells was identified based on staining with the vedolizumab-BV421 conjugate, and staining was analyzed using BD FACSuite software, version 1.0.5. Where relevant, values ​​were normalized to peptide-free controls and expressed as percentages to allow evaluation of changes in the indicated parameters. Data were plotted and analyzed using Prism software (version 7, GraphPad, La Jolla, CA).

[0240] result Human PBMCs were incubated with 100 nM Compound B, Compound C, or no peptide and stained for α4β7 expression. Incubation with Compound B, but not Compound C or the no-peptide control, resulted in internalization of α4β7 (FIG. 14). These data indicate that internalization is dependent on binding of the peptide to α4β7.

[0241] Purified human CD4 + T memory cells were incubated with 10 nM Compound A for a range of times (0-48 h) or Compound A concentrations (0-10 nM) for 24 h and stained for α4β7 expression. The results show that Compound A-induced α4β7 internalization is time (Figure 15) and concentration (Figure 16) dependent, respectively. Similar results were obtained using PBMCs (data not shown).

[0242] Purified human CD4 +T memory cells were incubated with various concentrations of Compound A and then washed to remove excess peptide. Separate aliquots from each reaction were stained for α4β7 expression and tested for MAdCAM1 adhesion, with values ​​normalized to the respective "no peptide" control for each assay. Data revealed that Compound A reduced α4β7 expression (a range of 5.4-24.7% reduction normalized to the no peptide control) and adhesion to MAdCAM1 (a range of 18.4-36.4% reduction relative to the no peptide control) (Figures 17 and 18, respectively). These effects were strongly correlated, exhibiting an R-squared value of 0.968 (Figure 19). Purified human CD4 from a second donor was also stained for α4β7 expression and MAdCAM1 adhesion, with values ​​normalized to the respective "no peptide" control for each assay. + When the assay was repeated using T memory cells, a similar correlation was obtained (R-squared 0.940, complete data not shown). Data from two donors resulted in an average maximum 39% reduction in α4β7 expression and an average maximum 37% reduction in adhesion to MAdCAM1, as shown in Table 1.

[0243] Table 14. Maximum reduction in α4β7 expression and adhesion to MAdCAM1 TIFF2025186365000050.tif29160*Normalized to the percentage of the peptide-free control.

[0244] Human PBMCs were incubated with 10 nM Compound A or without peptide in MnCl2-free medium for 24 hours, then washed to remove excess peptide and resuspended in fresh medium without MnCl2. At 24 hours, α4β7 expression, as measured by MFI (5090 MFI), was only 20.6% different compared to the FMO control (4219 MFI). After removing Compound A, incubation was continued, and aliquots were removed and stained for α4β7 expression on days 1, 2, 4, 5, and 7. Results showed that after 4–5 days of additional incubation, downregulation of α4β7 expression in the presence of peptide was nearly restored to control levels (Figure 20).

[0245] conclusion This study demonstrated that Compound A specifically induces the internalization of α4β7 in human primary cells in a time- and dose-dependent manner. The reduction of α4β7 expression by Compound A was associated with the CD4 + This was highly correlated with reduced adhesion to MAdCAM1 by T memory cells. Compound A-induced internalization of α4β7 on cells required an additional 4-5 days of incubation to fully restore control expression levels.

[0246] Example 7 A Randomized, Double-Blind, Placebo-Controlled Study of Single and Multiple Ascending Doses of Compound A in Normal Healthy Volunteers Ulcerative colitis is a chronic inflammatory bowel disease with a remitting and relapsing course, characterized by bloody diarrhea, abdominal cramps, and fatigue. The etiology is thought to result from an inappropriate immune response to gastrointestinal antigens and environmental triggers in genetically susceptible individuals.

[0247] The α4β7 integrin, present on the cell surface of circulating memory T and B lymphocytes, is primarily responsible for leukocyte recruitment to the gastrointestinal mucosa and associated lymphoid tissues. The major ligand for α4β7, mucosal addressin cell adhesion molecule 1 (MAdCAM1), is selectively expressed on the endothelium of the gastrointestinal vasculature and is present at increased concentrations in inflamed tissues.

[0248] Vedolizumab is an intravenously administered humanized IgG monoclonal antibody directed against α4β7. It is approved for the treatment of moderate to severe ulcerative colitis and Crohn's disease in adult patients who have not responded to one or more conventional therapies, such as steroids, immunosuppressants, or tumor necrosis factor (TNF) inhibitors. Due to the inconvenience and potential systemic risks of injectable treatments, an oral GI-restrictive therapeutic agent that selectively targets the α4β7 integrin may offer significant benefit to ulcerative colitis patients. Compound A is an orally stable peptide that specifically binds to the α4β7 integrin on leukocytes and shows minimal systemic absorption (<1%) in animal studies. This study investigated the safety, tolerability, pharmacokinetics, and pharmacodynamics of oral Compound A in healthy male subjects.

[0249] Two pharmacokinetic / pharmacodynamic studies were conducted in healthy volunteers. Study 1 was the first in-human study in 40 men receiving a single dose of Compound A, 100-1400 mg, or placebo, and 57 men receiving multiple doses of Compound A, 100-1000 mg, or placebo. Study 2 was a randomized crossover study comparing multiple doses of Compound A, 450 mg twice daily, as a liquid solution and as an immediate-release tablet in 10 subjects.

[0250] No subjects discontinued due to treatment-emergent adverse events. Consistent with the gastrointestinal-restricted nature of the peptide, systemic exposure was minimal, with an approximately dose-proportional increase in AUC. With once-daily dosing, there was minimal accumulation and no time-dependent changes in pharmacokinetics. Administration of Compound A after a high-fat meal reduced peak plasma concentrations and AUC. Urinary excretion of intact drug was minimal (<0.1%), and there was a dose-related increase in fecal excretion of intact Compound A. A dose-dependent increase in blood receptor occupancy and a reduction in blood receptor expression were observed, supporting target engagement. Twice-daily dosing resulted in sustained receptor occupancy (143%) with low plasma fluctuations.

[0251] Compound A was generally well tolerated after single and multiple oral doses with low systemic exposure. Twice-daily dosing resulted in sustained pharmacokinetics and pharmacodynamics, supporting further investigation in efficacy studies.

[0252] method research design Two studies were conducted at a single clinical center.

[0253] Study 1 was a three-part, first-in-human study in healthy male volunteers to evaluate the safety, tolerability, pharmacokinetics, and pharmacodynamics of a liquid solution formulation of Compound A.

[0254] Part 1 was a randomized, placebo-controlled, double-blind study of single ascending doses of Compound A in 40 men divided into four equal cohorts. Dose escalation progressed from 100 mg, 300 mg, 1000 mg, and 1400 mg. Subjects in the 300 mg dose cohort were treated in a crossover fashion, first fasted and second after a high-fat meal. The high-fat meal consisted of two fried eggs in butter, two slices of bacon, two slices of buttered toast, 4 ounces of hash brown potatoes, and 240 ml of whole milk. During Part 1, subjects abstained from food and beverages other than water 10 hours before and 4 hours after dosing, except for subjects in the 300 mg dose cohort during contact treatment.

[0255] Part 2 was a randomized, placebo-controlled, double-blind, multiple-ascending-dose study in 50 male subjects, equally divided into five cohorts. Subjects received a once-daily dose of Compound A or placebo for 14 days. Doses evaluated in Part 2 included 100 mg, 300 mg, and 1000 mg. During Part 2, subjects in two cohorts (100 mg and 300 mg) received a meal approximately 30 minutes before each dose, while subjects in two cohorts (300 mg and 100 mg) abstained from food 10 hours before and 1 hour after dosing. An additional cohort of nine subjects in Part 2 received 300 mg of Compound A in a crossover fashion to evaluate the effect of meal timing on the pharmacokinetics and pharmacodynamics of Compound A. Subjects in this cohort received a meal 30, 60, or 90 minutes after dosing with Compound A.

[0256] Part 3 was an open-label, randomized, crossover, multiple-dose comparison of Compound A at 900 mg once daily and 450 mg twice daily as a liquid solution for 5 days. Part 3 subjects abstained from food 10 hours before and 1 hour after Compound A dosing.

[0257] The second study was a 5-day, multiple-dose pharmacokinetic and pharmacodynamic study comparing liquid and tablet formulations of Compound A administered twice daily at 450 mg in healthy men and women. Subjects were instructed to withhold food 10 hours before and 1 hour after the daily morning dose, and 1 hour before and 1 hour after the evening dose.

[0258] The study protocol, subject information, and informed consent form were reviewed and approved by an independent human research ethics committee. The study was conducted in accordance with the Declaration of Helsinki and the International Conference on Harmonization Good Clinical Practice guidelines for biomedical research involving human subjects, and all study procedures were performed by scientifically and medically qualified personnel. Written informed consent, explaining the nature, objectives, and potential risks and benefits of the study, was provided by the subjects prior to any study-related activities.

[0259] Research Subjects Both studies used similar screening and enrollment procedures. Subjects were screened within 21 days of enrollment. Eligible subjects were 18 to 55 years of age and had a body mass index (BMI) of 18 to 30 kg / m. 2 The subjects were in good general health, with no significant medical history or clinically significant abnormalities on physical examination. The first human study (Study 1) enrolled only men, and the study evaluating the tablet formulation (Study 2) enrolled both men and women who agreed to use highly effective contraceptive methods in accordance with the Clinical Trials Facilitation and Coordination Group guidelines for the duration of the study and for 90 days after the last dose.

[0260] Subjects were excluded if they had a history of clinically significant endocrine, gastrointestinal, cardiovascular, hematological, hepatic, immunological, renal, respiratory, or urinary abnormalities or diseases, or clinically significant laboratory abnormalities including loss of renal function (serum creatinine >106 umol / L or estimated creatinine clearance <80 mL / min), or alanine aminotransferase or aspartate aminotransferase values ​​>1.2 times normal.

[0261] procedure Study 1: The single and multiple ascending dose phases of the study consisted of sequential dose escalation in 10 subjects per dose cohort. Participants were randomized to receive Compound A or matching placebo as a 60 mL oral solution in an 8:2 ratio. Dose solutions were formulated in 50 mM phosphate buffer, pH 7.4, and prepared weekly by qualified pharmacists. Dosing solutions spanning the expected concentration range were demonstrated to be stable for 3 months when stored at 2-8°C.

[0262] Blood samples for pharmacokinetics were collected pre-dose and 48 hours post-dose. During the multiple ascending dose phase, blood samples were obtained on days 1-3 and 14-16. On day 8, samples were obtained 4 hours and 12 hours pre-dose. On day 10 of the MAD, subjects were required to collect all urine samples from 0-6, 6-12, 12-18, and 18-24 hours post-dose, and on day 11, subjects were required to collect a fecal sample.

[0263] The decision to proceed to the next dose level was made by the investigator and safety monitoring committee based on acceptable safety and tolerability of the lower dose.

[0264] Study 2: This was a randomized, open-label, two-treatment, two-period, multiple-dose study to determine the safety, tolerability, pharmacokinetics, and pharmacodynamics of Compound A immediate-release (IR) tablets and liquid solution. This study allowed for a comparison of the liquid and solid dose formulations investigated in the first human study. Subjects were randomized to receive 450 mg of Compound A twice daily (BID) for 5 days as IR tablets in one 300 mg and one 150 mg dose strength administered every 12 hours, and 450 mg of Compound A BID for 5 days as a liquid solution administered every 12 hours.

[0265] dosage The initial human starting dose in single- and multiple-dose studies was based on the no-observed-effect level (NOEL) from 28-day toxicity studies in rats and cynomolgus monkeys and consideration of the receptor occupancy demonstrated in cynomolgus monkeys. The NOEL determined in rats and monkeys was converted to a human-equivalent dose of approximately 145 mg using standard allometric scaling and a 10-fold safety margin. A starting dose of 100 mg was selected, with an initial escalation of approximately 3-fold.

[0266] The dose selected for Study 2, which compared the tablet and oral solution formulations, was based on the pharmacokinetic and pharmacodynamic profiles from Part 3 of Study 1 and the expected dose planned for the efficacy study in patients with moderate to severe ulcerative colitis.

[0267] Analysis method A validated high-performance liquid chromatography-tandem mass spectrometry (LC / MS / MS) method was used to assay compound A concentrations in plasma, urine, and fecal samples from Study 1 and in plasma and urine samples from Study 2. Drug and internal standards were extracted from the matrix by a protein precipitation procedure. The limits of quantification were 0.2 ng / mL, 20 ng / mL, and 100 ng / mL for plasma, urine, and feces, respectively. Sample stability was demonstrated for at least 100 days and over four freeze-thaw cycles for all matrices. The coefficients of determination for the calibration curves were at least 0.99 for all matrices. Inter-assay accuracy (% bias) ranged from -2.2% to 1.0% for plasma, -3.8% to 9.0% for urine, and -5.0% to 5.2% for feces. Inter-assay precision (% CV) ranged from 3.7% to 7.7% for plasma, 2.8% to 7.0% for urine, and 1.2% to 5.2% for feces. Reanalysis of the generated samples showed that >88% of the samples with valid reanalysis met the acceptance criteria, making the analytical method acceptable.

[0268] Study endpoints The primary endpoint of this first human study was safety and tolerability assessment after single and multiple dosing with Compound A. Secondary objectives were to characterize pharmacokinetics and pharmacodynamics, evaluate the effect of a high-fat meal on the pharmacokinetics of Compound A, and compare twice-daily and once-daily dosing. Safety assessments, adverse events, and laboratory evaluations are summarized narratively for placebo and each Compound A dose.

[0269] The endpoints of the second study comparing oral solution and tablet formulations were pharmacokinetics and pharmacodynamics.

[0270] Pharmacokinetic analysis Pharmacokinetic parameters were estimated by non-compartmental methods using Phoenix WinNonlin (Certara, Princeton NJ). Peak plasma concentrations (C max ) and time to peak plasma concentration (T max ) was the observed value. Elimination rates were estimated from the slope of the least squares regression on the terminal log-linear phase. The area under t ) was estimated by the linear trapezoidal method and the area under the plasma concentration-time curve up to infinity (AUC) by dividing the last quantifiable concentration by the elimination rate. ∞ The steady-state plasma concentration fluctuations were calculated as Calculated as TIFF2025186365000051.tif8128.

[0271] Pharmacodynamic assays for α4β7 receptor occupancy and receptor expression Translational biomarkers such as receptor occupancy have been validated as pharmacodynamic markers through their use in preclinical studies and clinical trials with vedolizumab 27~28In this study, a flow cytometry-based assay was designed to quantify the amount of α4β7 integrin on the cell surface occupied by Compound A, or the amount of α4β7 expression on the cell surface of circulating lymphocytes in response to engagement by Compound A. Briefly, in this assay, each heparinized whole blood sample was first treated with a saturating amount of unlabeled competitor peptide, which served as a "blocking" control for 100% receptor occupancy, or no peptide, which served as a "non-blocking" sample to measure the level of blockade by orally administered Compound A. After incubation, the blood was stained with a subsaturating concentration of Alexa647-labeled peptide, followed by staining with a cell surface marker panel (CD45, CD3, CD4, CD45RA, CD19, IgD, and the anti-α4β7 antibody vedolizumab). After staining was complete, the sample was treated with red blood cell lysis and fixation buffer, washed, and acquired on a flow cytometer. To quantify receptor occupancy on α4β7-expressing memory CD4+ T cells, we used the medium fluorescence intensity (MFI) of Alexa647-labeled peptide in vedolizumab + memory CD4+ T cells. Receptor occupancy was calculated according to the following formula: [RO percent] = (1 - ([unblocked] - [blocked]) / ([unblocked baseline] - [blocked baseline])) x 100.

[0272] α4β7 expression is defined by the MFI of vedolizumab in memory CD4+ T cells from unblocked samples. Receptor expression (RE) was calculated as the percent change in MFI from baseline of vedolizumab staining.

[0273] statistical analysis No formal sample size estimation was performed. In the single and multiple ascending dose studies, eight subjects received oral Compound A and two subjects received placebo in each dose cohort. Ten subjects were enrolled in a second study comparing the immediate-release tablet formulation with an oral solution. Enrollment in each study was deemed adequate to assess the tolerability and safety of Compound A and to allow for characterization of the pharmacokinetics and pharmacodynamics of Compound A.

[0274] result Subject characteristics and trends A total of 97 healthy male subjects were enrolled in Study 1, with 40 subjects enrolled in the single-dose phase and 57 subjects enrolled in the multiple-dose phase. 95 subjects completed Compound A or placebo dosing as scheduled. Two subjects withdrew consent for personal reasons unrelated to safety: one subject did not want to remain in the clinical unit, and one subject experienced discomfort with the intravenous cannula. The mean age was 28.7 years in the single-dose phase and 30.9 years in the multiple-dose phase.

[0275] Ten subjects were enrolled in Study 2, and nine subjects completed both treatments. One subject discontinued the study on Day 1 after oral solution treatment due to an adverse event of acute tonsillitis that was deemed unrelated to study medication.

[0276] Safety and Tolerability A total of 23 TEAEs were reported by 14 subjects during the single ascending dose phase. Of the 13 subjects who experienced TEAEs, 12 received Compound A (21 events) and 2 received placebo (2 events). All TEAEs were mild or moderate, except for a severe headache in a subject treated with 100 mg of Compound A, which was not considered treatment-related. All subjects recovered from the AEs, and no subjects withdrew due to AEs. No clinically relevant changes were observed in respiratory rate or vital signs, clinical laboratory parameters (hematology, coagulation, serum chemistry, or urinalysis), or electrocardiogram or QTc interval interpretation.

[0277] Safety and Tolerability Thirty subjects in the group receiving multiple doses of Compound A reported a total of 68 AEs. All but two were mild in severity. One report of upper respiratory tract infection was characterized as moderate, and one report of influenza, which occurred after release from the clinical unit, was characterized as severe and considered a serious adverse event. Four subjects receiving placebo reported a total of six mild TEAEs, primarily gastrointestinal disorders. Treatment-emergent adverse events reported in two or more subjects during the multiple ascending dose phase included abdominal discomfort, flatulence, upper respiratory tract infection, back pain, dizziness, and headache. Nervous system disorders, particularly headache, were the most commonly reported TEAE. No clinically relevant changes were observed in respiratory rate, vital signs, laboratory parameters, or electrocardiograms.

[0278] Safety and Tolerability Of the 10 subjects enrolled in Study 2, comparing immediate-release tablet and oral solution dosing, 9 subjects completed both treatments. One subject experienced a moderate adverse event of tonsillitis unrelated to treatment and discontinued the study. The incidence of treatment-emergent adverse events was similar across both treatments. The most common adverse event was headache; all other adverse events were reported by only one subject.

[0279] Safety and Tolerability Pharmacokinetics The mean plasma concentration-time profiles after a single dose of Compound A are shown in Figure 21. The single-dose pharmacokinetics of Compound A are summarized in Table 15.

[0280] Table 15. Single-dose pharmacokinetics of Compound A (mean ± SD) TIFF2025186365000052.tif69160 a Median (min, max) b N=4 c Not reported due to insufficient data d N=7

[0281] The median time to peak plasma concentration was 2 to 4 hours. The mean peak Compound A plasma concentration (C max ) increased from 2.11 mg / mL to 23.5 ng / mL, and AUC inf AUC increased from 16.5 ng.hr / mL to 260 ng.hr / mL. Across the dose range of 100 mg to 1400 mg of Compound A, inf Dose-proportional increase in and C max There was a slightly less than dose-proportional increase in the mean elimination half-life at the low doses (100 and 300 mg) and at the high doses (1000 and 1400 mg), the mean elimination half-life was 3.1 to 4.0 hours and 5.3 to 5.7 hours, respectively.

[0282] Safety and Tolerability The pharmacokinetics of Compound A after multiple doses are summarized in Table 16.

[0283] Table 16. Multiple-dose pharmacokinetics of Compound A (mean ± SD) TIFF2025186365000053.tif75160 a Median (min, max) b N=1 c Not reported due to insufficient data d N=7

[0284] C between the 100 mg and 300 mg dose groups in the fed state on day 14 and between the 300 mg and 1000 mg dose groups in the fasted state on day 14 max and AUC inf There was an approximately dose-proportional increase in C. The median time to peak plasma concentration ranged from 2 to 4 hours. The mean elimination half-life ranged from 5.2 to 7.7 hours. Consistent with the half-life, individual subject C at 300 mg and 1000 mg on Days 1 and 14 max and AUC tComparison of the values ​​suggested minimal accumulation (≦30%) with once-daily dosing. Comparison of the AUC values ​​on Day 1 and the AUC values ​​on Day 14 indicated no time-dependent changes in the pharmacokinetics of Compound A.

[0285] Twenty-four-hour urine and fecal collections were performed in the 300 mg and 1000 mg dose groups during the multiple ascending dose phase. Only a small portion of Compound A was recovered intact in urine over 24 hours, with recoveries of 0.028%, 0.056%, and 0.056% in the 300 mg fasting, 300 mg fed, and 1000 mg dose groups, respectively. There was a dose-related increase in 24-hour fecal recovery of Compound A in the 300 mg fasting, 300 mg fed, and 1000 mg dose groups, with 0.73%, 1.78%, and 16.8% of Compound A recovered intact, respectively.

[0286] The influence of food The effect of a high-fat meal on the pharmacokinetics of Compound A was evaluated in a crossover fashion at 300 mg during the single ascending dose portion of Study 1.

[0287] Administration of Compound A within 30 minutes of ingesting a high-fat meal reduced peak concentrations and exposure compared to the fasted state (Table 13). Mean Compound A peak plasma concentrations were 6.55 ng / mL in the fasted state and 1.58 ng / mL in the fed state. The median time to peak concentration was delayed by 1 hour after the high-fat meal.

[0288] The effect of the interval between Compound A dosing and food intake was investigated in Study 1. Subjects received a meal 30, 60, or 90 minutes after a single dose of 300 mg Compound A. The median time to peak Compound A plasma concentrations was 1, 2, and 4 hours for the 30, 60, and 90 minute treatment groups. C was significantly higher when food was delayed 60 or 90 minutes compared to 30 minutes after Compound A. max and AUC tValues ​​increased slightly, with only a small difference noted between the 60- and 90-minute delays. Based on the more favorable Cmax and AUCt values ​​noted for the 60-minute delay of food compared to the 30-minute delay, dosing for additional cohorts in multiple ascending doses incorporated a 1-hour fasting interval before and after dosing with Compound A.

[0289] Table 16 shows a comparison of the pharmacokinetics of 300 mg of Compound A after an overnight fast compared to abstaining from food within 1 hour of dosing with Compound A as part of the multiple ascending dose phase of Study 1. The median time to peak concentration was 4 hours after an overnight fast, compared to 2 hours when food was taken 1 hour after Compound A. Peak plasma concentrations were lower when Compound A was administered after an overnight fast compared to when food was administered 1 hour after dosing (Cmax on Day 1 was 7.23 ng / mL and 2.32 ng / mL for fasting and for eating 1 hour after the dose, respectively).

[0290] Pharmacokinetics of once-daily and twice-daily dosing The effects of the dosing regimen were evaluated in part 3 of study 1 in a randomized crossover fashion after 900 mg once daily for 5 days and 450 mg twice daily for 5 days.

[0291] Figure 25 shows the mean plasma concentration-time profiles following dosing of 900 mg once daily and 450 mg twice daily of Compound A. A pharmacokinetic summary comparing once-daily and twice-daily dosing is shown in Table 17.

[0292] Table 17. Pharmacokinetics of Compound A after once-daily and twice-daily dosing (mean ± SD) TIFF2025186365000054.tif83166 a Median (min, max) b Not reported due to insufficient data

[0293] Peak concentrations were observed at a median of 2 hours for both dosing regimens on Days 1 and 5. Steady-state peak concentrations were 14.2 ng / mL for once-daily dosing and 9.96 ng / mL for twice-daily dosing. Dose-adjusted areas under the curve for the dosing interval were comparable for the two treatment regimens. Consistent with the half-life of Compound A, there was minimal accumulation with once-daily dosing and approximately 1.6- to 1.7-fold accumulation with twice-daily dosing. Twice-daily dosing of 450 mg Compound A as a liquid solution resulted in sustained plasma concentrations compared with 900 mg once-daily, as reflected by lower peak-to-trough fluctuations (143% vs. 245%) and higher trough concentrations (3.25 ng / mL vs. 1.78 ng / mL) (Table 18).

[0294] Pharmacokinetics of Compound A liquid solution and immediate-release tablet formulations The steady-state pharmacokinetics of Compound A immediate release tablets administered as 450 mg twice daily for 5 days compared to the liquid solution used in the first human study are summarized in Table 18.

[0295] Table 18. Steady-state pharmacokinetics of Compound A after oral dosing of 450 mg twice daily as a liquid solution and as an IR tablet (mean ± SD) TIFF2025186365000055.tif76128 a Median (min, max)

[0296] Figure 23A shows the mean steady-state plasma concentration-time profiles of the two formulations. Both formulations had similar median time to peak concentration (2 hours), while the peak concentration was approximately 20% lower for the IR tablet compared to the liquid solution. The IR tablet formulation had approximately 85% bioavailability relative to the liquid solution. Twice-daily dosing of the tablet formulation resulted in a C max The IR tablet and liquid solution concentrations were comparable (1.86 ng / mL and 1.98 ng / mL, respectively), resulting in approximately 2-fold accumulation based on β-blockers and 1.6-fold accumulation based on AUC.

[0297] Pharmacodynamics α4β7 as measured by mean percent receptor occupancy and mean receptor expression after a single dose of Compound A + Memory CD4 + The mean pharmacodynamics of T cells are summarized in Table 19.

[0298] Table 19: Pharmacodynamics of Compound A after a single dose (mean ± SD). TIFF2025186365000056.tif67166RO max Maximum receptor occupancy, RE max Maximum receptor expression

[0299] The mean percent receptor occupancy and mean receptor expression time course after a single dose are shown in Figures 22A-B. Peak α4β7 memory CD4 + The mean time to T cell occupancy was approximately 4 hours. Mean peak receptor occupancy increased dose-related, ranging from 61.8% at 100 mg to 94.8% at 1400 mg. Peak receptor occupancy for the 1000 mg and 1400 mg dose cohorts was similar, indicating that receptor occupancy saturation was achieved by a single dose of approximately 1000 mg of Compound A. Mean change in receptor expression (RE) max ) increased with dose, ranging from −28.2% in the 100 mg Compound A dose group to −49.0% in the 1400 mg dose group (Table 19).

[0300] α4β7 after multiple doses of Compound A + Memory CD4 + The percent receptor occupancy of T cells is shown in Table 20.

[0301] Table 20. Multiple dose pharmacodynamics of Compound A (mean ± SD) TIFF2025186365000057.tif23964RO max Maximum receptor occupancy, RE max Maximum receptor expression

[0302] The mean peak memory T cell receptor occupancy rate after multiple doses of Compound A reached a peak at approximately 4 hours. The mean receptor occupancy rate percentage on day 1 for the multiple dose cohort was comparable to that of the single dose cohort. On day 1, the mean peak receptor occupancy rates after 300 mg and 1000 mg were 77.8% and 91.3%, respectively. With continued daily dosing for 14 days, the peak receptor occupancy rate percentage increased slightly. On day 14, the mean peak receptor occupancy rates after 300 mg and 1000 mg were 79.7% and 95.6%, respectively.

[0303] Administration of Compound A within 30 minutes of ingesting a high-fat meal reduced the pharmacodynamic effects, consistent with the effect of a high-fat meal on pharmacokinetics. Peak receptor occupancy after 300 mg of Compound A in the fasted state was 83.4%, compared to 61.4% when Compound A was administered within 30 minutes after a high-fat meal. Delaying food for 60 minutes after administration of Compound A improved the pharmacodynamic profile compared to dosing after a high-fat meal or consuming food within 30 minutes after dosing with Compound A. There was relatively little difference in steady-state (day 14) pharmacodynamic effects when Compound A was administered in the fasted state or when food was consumed 60 minutes after dosing with Compound A (Table 15).

[0304] The pharmacodynamic effects of 900 mg once daily and 450 mg twice daily were investigated as part of the multiple ascending dose phase of Study 1. A summary of the pharmacodynamic effects on receptor occupancy by dosing regimen is shown in Table 21.

[0305] Table 21: Pharmacodynamic summary of once-daily and twice-daily receptor occupancy (mean ± SD). TIFF2025186365000058.tif69166 a N=7

[0306] On Day 1, the 900 mg once-daily regimen achieved a mean peak receptor occupancy of 94.5%, while the 450 mg twice-daily regimen achieved a mean peak receptor occupancy of 86.5%. Although both treatment regimens resulted in similar peak receptor occupancy on Day 5 (94.9% and 91.9% for 900 mg QD and 450 mg BID, respectively), the twice-daily regimen provided a more sustained pharmacodynamic effect. Notably, the AUEC on Day 5 was higher with the twice-daily regimen compared with the once-daily regimen. The mean receptor occupancy based on the 24-hour area under the curve (AUEC) on Day 5 was 85.3% with the twice-daily regimen and 79.2% with the once-daily regimen. The BID regimen also provided a sustained effect, as indicated by the minimal difference in peak and trough receptor occupancy. In addition, the intersubject variability in receptor occupancy at trough for the 450 mg BID treatment was 11.3%–15.2% on day 5 compared with 26.3%–33.6% for the 900 mg QD treatment, suggesting a more consistent effect of the BID regimen.

[0307] Steady-state CD4 after twice-daily Compound A as IR tablets or liquid solution + The pharmacodynamics of percent receptor occupancy of α4β7 memory T cells are shown in Figure 23B. The pharmacodynamics of steady-state receptor occupancy are summarized in Table 22.

[0308] Table 22. Steady-state pharmacodynamics of Compound A after oral dosing of 450 mg twice daily as a liquid solution and as an IR tablet (mean ± SD) TIFF2025186365000059.tif51128RO max Maximum receptor occupancy, RE max Maximum receptor expression

[0309] Peak receptor occupancy was observed at 4 hours for both formulations. The mean steady-state peak receptor occupancy for the IR tablet was 91.9%, with a mean 24-hour receptor occupancy of 83.6%, compared to a peak receptor occupancy of 93.8% and a mean 24-hour receptor occupancy of 85.8% for the liquid solution.

[0310] Pharmacokinetic-pharmacodynamic correlation The in vivo Compound A plasma concentration-receptor occupancy relationship was characterized using the sigmoidal Emax (Hill) model (Figure 24). Estimated IC for receptor occupancy 50 and IC 80 were 0.69 ng / mL and 5.9 ng / mL, respectively.

[0311] Consideration Compound A is an oral, gut-restricted peptide that specifically binds to the α4β7 integrin on leukocytes and is being developed in Phase 2 studies as a potential oral treatment for patients with ulcerative colitis. The peptide's GI-restricted nature and enhanced gastrointestinal stability have the potential to enhance efficacy, allowing for localized effects while minimizing the potential for adverse events associated with systemic exposure.

[0312] The primary objective of these studies was to evaluate the safety / tolerability of Compound A after single and multiple dosing. Secondary objectives were to assess the pharmacokinetic and pharmacodynamic profile of Compound A after single and multiple ascending oral doses, to evaluate the effect of food on the pharmacokinetics and pharmacodynamics, to compare once-daily and twice-daily dosing, and to describe the pharmacokinetics and pharmacodynamics of an immediate-release formulation of Compound A.

[0313] In the first human study, Compound A was well tolerated after single doses of up to 1400 mg and multiple doses of up to 1400 mg once daily for 14 days. TEAEs were all mild, except for one report of severe headache after a single dose of the lowest dose of Compound A (100 mg) and one report of flu after 900 mg once daily. None of the TEAEs led to subject withdrawal from the study. Treatment-emergent adverse events noted in two or more subjects after repeated dosing included abdominal discomfort, flatulence, upper respiratory tract infection, back pain, dizziness, and headache, with headache being the most frequently reported TEAE. Treatment with Compound A did not result in any safety findings with respect to clinically significant changes in vital signs or laboratory values, and no evidence of QTc prolongation was observed. There were no differences in the treatment-emergent adverse event profile after twice-daily dosing of Compound A as an IR tablet or liquid solution.

[0314] After a single oral dose, Compound A had a moderate absorption rate with maximum plasma concentrations occurring at approximately 4 hours. The increase in AUC of Compound A was approximately dose-proportional, but max The increase in AUC was slightly less than dose-proportional. Compound A demonstrated low systemic exposure after single and multiple dosing. Terminal half-lives ranged from 3.1 to 5.7 hours in the fasted state and 5.2 to 7.7 hours in the fed state. Consistent with the terminal half-life, Compound A accumulation was approximately 0.9- and 1.6-fold when administered once daily and twice daily, respectively. Similar day 1 AUC inf and AUC on day 14 t There was no time-dependent pharmacokinetics, as evidenced by (Supplementary Table 4).

[0315] There was a dose-dependent increase in α4β7 receptor occupancy after Compound A administration, reaching a mean peak receptor occupancy of over 90% at the 900 mg dose. Trough receptor occupancy after once-daily dosing of 100 mg and 1000 mg Compound A was approximately 25.4% and 78.6%, respectively, and after twice-daily dosing of 450 mg Compound A was 79.2%. These receptor occupancy data indicate that Compound A concentrations are maintained at levels sufficient to allow for once- or twice-daily dosing. PK / PD correlation demonstrated an estimated IC of 0.69 ng / mL. 50 and an IC of 5.9 ng / mL. 80 showed concentration-dependent receptor occupancy with an asymptote at full receptor occupancy. The estimated IC for receptor occupancy shown in humans 50 (0.69 ng / mL) compares very favorably with the potency of Compound A (0.73 ng / mL) against recombinant MAdCAM1-expressing memory CD4+ T cells isolated from human peripheral blood mononuclear cells.

[0316] Systemic concentrations of Compound A after oral administration were generally low, consistent with the gut-restricted nature of the drug and the very low oral bioavailability (<1%) demonstrated in mice and cynomolgus monkeys. There was a dose-dependent increase in fecal recovery of Compound A after oral administration, ranging from approximately 1-2% at 300 mg to 16.8% at 1000 mg. Compound A is a small, disulfide-containing, cyclic peptide. Orally administered peptides encounter a harsh environment along the gastrointestinal tract, including pH conditions ranging from pH <2 in the stomach to pH 8 in the duodenum, as well as proteolytic enzymes such as gastric hydrolases (pepsin), pancreatic hydrolases (trypsin, chymotrypsin, elastase, aminopeptidase, and carboxypeptidase A and B), and intestinal brush border membrane-associated enzymes (carboxypeptidases, endopeptidases, and aminopeptidases). 29The highly acidic environment in the stomach leads to the degradation of peptide drugs through destabilization of their three-dimensional structure. The stability of peptides and proteins in the gastrointestinal tract is an inherent problem associated with oral administration, whether for local action or systemic delivery. Numerous studies have shown that various factors, such as amino acid sequence, molecular size, pH, and exposure to the gastrointestinal environment, including enzymatic action, play important roles in determining peptide stability and the likelihood of oral absorption. Cyclization via sulfide bond linkage and N-methylation may provide some resistance to enzymatic degradation and also improve the oral absorption of Compound A.

[0317] Low detectable intact concentrations in plasma after oral administration indicate that Compound A can penetrate the gastrointestinal barrier. Approximately 0.03% to 0.06% of the drug was detected intact in urine. Orally administered peptides typically have low oral bioavailability. Although systemic concentrations of Compound A were low, they were sufficient to achieve and maintain >80% receptor occupancy at the trough after once-daily or twice-daily dosing.

[0318] Administration of Compound A within 30 minutes of a high-fat meal reduced the oral absorption of Compound A. Although there was no direct correlation between systemic exposure and fecal recovery, the data directly indicated that there was a corresponding increase in fecal recovery following a high-fat meal in absorption reduction.

[0319] The steady-state pharmacokinetic and pharmacodynamic profiles of the immediate-release tablet formulation of Compound A were generally similar to those of the liquid formulation used in the first human study. Twice-daily dosing of 450 mg Compound A as an IR tablet resulted in sustained pharmacokinetics and a mean receptor occupancy of approximately 84%.

[0320] conclusion Ninety-seven healthy male volunteers were dosed with Compound A. In Part 1, a single ascending dose of Compound A up to a daily dose of 1400 mg was studied, along with the effect of food. In Part 2, multiple ascending doses up to 1000 mg were tested, administered once daily for up to 14 days. Additionally, 900 mg of Compound A once daily for 5 days was compared with 450 mg of Compound A twice daily for 5 days. The study drug was well tolerated, and no dose-limiting toxicities were observed. With one exception, all adverse events were mild to moderate in severity. One serious adverse event, characterized as severe, of influenza was reported approximately 36 hours after receiving Compound A and was possibly related to the study drug. The diagnosis was influenza A, confirmed by influenza swab testing. The subject recovered uneventfully.

[0321] The maximum tolerated dose for both single and multiple doses was 1400 mg for single doses and 1000 mg for multiple doses, the highest dose tested. Minimal plasma exposure was observed for both single and multiple doses, confirming that the drug was largely GI-restricted. A dose-dependent increase in blood receptor occupancy and a decrease in receptor expression were observed, thus supporting target engagement and pharmacological activity of Compound A in healthy volunteers.

[0322] To support the use of the tablet formulation, a multiple-dose crossover pharmacokinetic and pharmacodynamic study was conducted in 10 healthy subjects after dosing with 450 mg as an oral solution administered twice daily for 5 days or as an immediate-release tablet administered twice daily for 5 days. On average, the IR tablet had slightly lower peak Compound A plasma concentrations and AUC values ​​(approximately 15-18%) than the solution on Day 5, and this difference is not considered clinically significant. Mean steady-state peak receptor occupancy was >90% for both formulations, and mean receptor occupancy based on the 24-hour area under the effect curve (AUEC) on Day 5 was comparable for the two formulations.

[0323] Following single and multiple ascending doses, Compound A was safe and well tolerated when administered orally to healthy subjects across a wide dose range. Consistent with a GI-restricted peptide, Compound A had low systemic exposure with a pharmacokinetic profile supporting once- or twice-daily dosing. Twice-daily dosing of Compound A resulted in sustained receptor occupancy. The safety, tolerability, and PK / PD profile of Compound A in healthy subjects support the continued clinical evaluation of this novel, GI-restricted, targeted treatment for inflammatory bowel disease.

[0324] Example 8 A Randomized, Double-Blind, Placebo-Controlled Study Evaluating the Safety and Efficacy of Oral Compound A in Subjects With Moderately to Severely Active Ulcerative Colitis A Phase 2 randomized, double-blind, placebo-controlled clinical study in human patients with moderate to severe ulcerative colitis will be conducted to demonstrate the safety, tolerability, and efficacy of treatment with oral Compound A. The study will also evaluate the pharmacokinetics (PK) and pharmacodynamics (PD), as well as biomarker responses to treatment with oral Compound A.

[0325] research design This is a two-part study. Part 1 is a randomized, double-blind, placebo-controlled, parallel-design 12-week lead-in treatment period in patients with moderately to severely active UC, and Part 2 is a 40-week extension treatment period including subjects who successfully completed Part 1. Subjects who completed the 12-week visit in Part 1 are eligible to enter Part 2.

[0326] Part 1: Induction Treatment Period (ITP): Part 1 is a 12-week, randomized, double-blind, placebo-controlled, parallel-design study in adult subjects with moderate to severe active UC. Eligible subjects will be randomized 1:1:1 to receive Compound A 450 mg twice daily (BID), Compound A 150 mg BID, or placebo BID. Subjects must have a biopsy-confirmed diagnosis of UC. To meet the inclusion criteria, eligible subjects must have previously had an inadequate initial response, loss of response, or intolerance to older conventional therapies for UC (i.e., corticosteroids, aminosalicylates, or immunomodulators), or a previous inadequate initial response, loss of response, or intolerance to newer biologic therapies (i.e., TNFα antagonists or IL-12 / 23 antagonists). Subjects with a history of prior vedolizumab treatment will be excluded. Randomization will be stratified by prior failure of a TNFα antagonist or IL-12 / 23 antagonist.

[0327] Eligible subjects will meet the following inclusion criteria: Male and female subjects aged 18 (or country-specific minimum age of consent if >18) to 75 years old; • Subjects who understand the study procedures and agree to participate in the study by giving written informed consent; • A diagnosis of UC supported by adequate documentation of biopsy results consistent with UC; Moderate to severe active UC, and • Demonstrated inadequate response, loss of response, or intolerance to at least one oral aminosalicylate (5-ASA), corticosteroid, immunomodulator, or biologic agent (excluding vedolizumab); AND eligible subjects do not meet the following exclusion criteria: • Subjects with a current diagnosis of Crohn's disease (CD), indeterminate colitis (IC), microscopic colitis, ischemic colitis, or radiation colitis; • History of colonic dysplasia other than a completely removed low-grade dysplastic lesion; • History of active bacterial, viral, fungal, or mycobacterial infection requiring hospitalization or IV antibiotic / anti-infective treatment within 4 weeks of screening, or oral antibiotic / anti-infective within 2 weeks of screening; prior treatment with vedolizumab, natalizumab, or any agent targeting α4β7 or β1 integrins planned during the study; Positive stool test for C. difficile Chronic recurrent or severe infections, known primary or secondary immunodeficiency, Women who are pregnant or breastfeeding, or who are considering pregnancy during the study or within 30 days after the last dose of study drug, and • History of any major neurological disorder.

[0328] Eligible subjects will be randomized 1:1:1 to Compound A 450 mg twice daily (BID), Compound A 450 150 mg BID, or placebo BID.

[0329] Part 2: Extended Treatment Period (ETP): Subjects who complete the Week 12 visit in Part 1, including the Adapted Mayo Score component, are eligible to enter Part 2. All completers of Part 1 will be eligible to enter the extension treatment period in Part 2 at the investigator's discretion. Subjects will be assigned to the appropriate extension treatment group in a blinded manner. All subjects proceeding to Part 2 will receive Compound A.

[0330] Test Sample, Dosage and Mode of Administration: Compound A (300 mg and 150 mg) and corresponding placebo tablets are administered orally. Both Compound A strengths and placebo have the same appearance.

[0331] Outcome analysis: The primary outcome measure includes the proportion of subjects achieving clinical remission compared to placebo at week 12. Clinical remission is determined using the Adapted Mayo Score (the sum of three subscores from the Mayo Score). Stool Frequency Subscore (SFS) Rectal bleeding subscore (RBS) ●Endoscopic subscore (ESS)

[0332] Secondary outcome measures include individual comparisons between high and low doses of Compound A versus placebo. • Proportion of subjects with endoscopic improvement. • Proportion of subjects achieving endoscopic remission. • Percentage of subjects with histological enhancement. • Proportion of subjects achieving histologic remission. • Proportion of subjects with mucosal healing.

[0333] Other outcome measures include the proportion of subjects achieving clinical remission at week 52. Clinical remission is determined using the Adapted Mayo Score (the sum of three subscores from the Mayo Score). Stool Frequency Subscore (SFS) Rectal bleeding subscore (RBS) ●Endoscopic subscore (ESS).

[0334] Efficacy assessment Efficacy will be assessed, at least in part, based on the Mayo score, which includes four components: stool frequency subscore (SFS), rectal bleeding subscore (RBS), endoscopy subscore (ESS), and physician global assessment (PGA). Each individual score ranges from 0 to 3, with higher numbers indicating greater severity. The Complete Mayo score is the sum of all four subscores (SFS, RBS, ESS, and PGA) and ranges from 0 to 12 points. The Adapted Mayo score is the sum of the three subscores (SFS, RBS, and ESS). The Adapted Mayo score ranges from 0 to 9 points. • The Partial Mayo score is the sum of three subscores (SFS, RBS, and PGA) ranging from 0 to 9 points. • Endoscopic subscore (ESS) ≤ 1 (corrected to not include fragility in a score of 1).

[0335] result Treatment with any of the doses of Compound A is expected to be safe, and treatment with either 450 mg BID or 150 mg BID is expected to show statistically significant improvements in Complete Mayo score, Adapted Mayo score, and / or Partial Mayo score compared to treatment with placebo, thus demonstrating the efficacy of these doses of Compound A for treating ulcerative colitis.

[0336] All of the above U.S. patents, U.S. patent application publications, U.S. patent applications, foreign patents, foreign patent applications and / or non-patent publications referred to in this application and / or listed in this application data sheet are hereby incorporated by reference in their entirety.

[0337] The present invention may be embodied in other specific forms without departing from its structure, method, or other significant characteristics as broadly described herein and as claimed below. The described embodiments are to be considered in all respects as illustrative and not restrictive. The scope of the invention is, therefore, indicated by the appended claims rather than by the foregoing description. All changes that come within the meaning and range of equivalency of the claims are intended to be embraced within their scope.

[0338] Sequence information SEQUENCE LISTING <110> Protagonist Therapeutics, Inc. <120> METHODS FOR TREATING INFLAMMATORY BOWEL DISEASES WITH alpha4beta7 INTEGRIN ANTAGONISTS <150> US 62 / 959,854 <151> 2020-01-10 <160> 8 <170> PatentIn version 3.5 <210> 1 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Made in lab - peptide antagonist <220> <221> MOD_RES <222> (1)..(1) <223> Xaa is N-Methyl-Arginine <220> <221> MOD_RES <222> (1)..(1) <223> N-Methyl-Arginine modified with 2-methylbenzoyl <220> <221> MOD_RES <222> (6)..(6) <223> Xaa is penicillamine <220> <221> MOD_RES <222> (7)..(7) <223> Xaa is (S)-2-amino-3-(4-tert-butyl-phenyl)propionic acid <220> <221> MOD_RES <222> (8)..(8) <223> Xaa is beta-homoglutamic acid <220> <221> MOD_RES <222> (9)..(9) <223> D form Glu <220> <221> MOD_RES <222> (10)..(10) <223> D form Lys <400> 1 Xaa Ser Asp Thr Leu Xaa Xaa Xaa Glu Lys 1 5 10 <210> 2 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Made in lab - peptide antagonist <220> <221> MOD_RES <222> (1)..(1) <223> Xaa is N-Methyl-Arginine <220> <221> MOD_RES <222> (1)..(1) <223> N-Methyl-Arginine modified with 2-methylbenzoyl <220> <221> MOD_RES <222> (6)..(6) <223> Xaa is penicillamine <220> <221> MOD_RES <222> (7)..(7) <223> Xaa is (S)-2-amino-3-(4-tert-butyl-phenyl)propionic acid <220> <221> MOD_RES <222> (8)..(8) <223> Xaa is beta-homoglutamic acid <220> <221> MOD_RES <222> (10)..(10) <223> D form Lys <400> 2 Xaa Ser Asp Thr Leu Xaa Xaa Xaa Gly Lys 1 5 10 <210> 3 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Made in lab - peptide antagonist <220> <221> MOD_RES <222> (1)..(1) <223> Xaa is N-Methyl-Arginine <220> <221> MOD_RES <222> (1)..(1) <223> N-Methyl-Arginine modified with 2-methylbenzoyl <220> <221> MOD_RES <222> (6)..(6) <223> Xaa is penicillamine <220> <221> MOD_RES <222> (7)..(7) <223> Xaa is (S)-2-amino-3-(4-tert-butyl-phenyl)propionic acid <220> <221> MOD_RES <222> (8)..(8) <223> Xaa is beta-homoglutamic acid <220> <221> MOD_RES <222> (10)..(10) <223> D form Lys <400> 3 Xaa Ser Asp Thr Leu Xaa Xaa Xaa Pro Lys 1 5 10 <210> 4 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Made in lab - peptide antagonist <220> <221> MOD_RES <222> (1)..(1) <223> Xaa is N-Methyl-Arginine <220> <221> MOD_RES <222> (1)..(1) <223> N-Methyl-Arginine modified with 2-methylbenzoyl <220> <221> MOD_RES <222> (6)..(6) <223> Xaa is penicillamine <220> <221> MOD_RES <222> (7)..(7) <223> Xaa is (S)-2-amino-3-(4-tert-butyl-phenyl)propionic acid <220> <221> MOD_RES <222> (8)..(8) <223> Xaa is beta-homoglutamic acid <220> <221> MOD_RES <222> (9)..(9) <223> D form Pro <220> <221> MOD_RES <222> (10)..(10) <223> D form Lys <400> 4 Xaa Ser Asp Thr Leu Xaa Xaa Xaa Pro Lys 1 5 10 <210> 5 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Made in lab - peptide antagonist <220> <221> MOD_RES <222> (1)..(1) <223> Xaa is N-Methyl-Arginine <220> <221> MOD_RES <222> (1)..(1) <223> N-Methyl-Arginine modified with 2-methylbenzoyl <220> <221> MOD_RES <222> (6)..(6) <223> Xaa is penicillamine <220> <221> MOD_RES <222> (7)..(7) <223> Xaa is (S)-2-amino-3-(4-tert-butyl-phenyl)propionic acid <220> <221> MOD_RES <222> (8)…(8) <223> The stone is beta-homoglutamic acid. <220> <221> MOD_RES <222> (9)..(9) <223> D form Lys <400> 5 Thanks Ser Asp Thr Leu Thanks Thanks Lys 1 5 <210> 6 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Lab made - antagonist peptide <220> <221> MOD_RES <222> (1)..(1) <223> The stone is penicillamine. <220> <221> MOD_RES <222> (2)…(2) <223> The stone is N-Methyl-Arginine. <220> <221> MOD_RES <222> (7)…(7) <223> The stone is penicillamine. <220> <221> MOD_RES <222> (8)…(8) <223> The stone is (S)-2-amino-3-(4-tert-butyl-phenyl)propionic acid <220> <221> MOD_RES <222> (9)..(9) <223> The stone is beta-homoglutamic acid. <220> <221> MOD_RES <222> (10)..(10) <223> D form Lys <400> 6 Thank You Ser Asp Thr Leu Thank You Lys 1 5 10 <210> 7 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Lab made - antagonist peptide <220> <221> MOD_RES <222> (1)..(1) <223> The stone is penicillamine. <220> <221> MOD_RES <222> (2)…(2) <223> The stone is N-Methyl-Arginine. <220> <221> MOD_RES <222> (7)…(7) <223> The stone is penicillamine. <220> <221> MOD_RES <222> (8)…(8) <223> The stone is (S)-2-amino-3-(4-tert-butyl-phenyl)propionic acid <220> <221> MOD_RES <222> (9)..(9) <223> The stone is beta-homoglutamic acid. <220> <221> MOD_RES <222> (11)..(11) <223> D form Lys <400> 7 Thank You Ser Asp Thr Leu Thank You Pro Lys 1 5 10 <210> 8 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Lab made - antagonist peptide <220> <221> MOD_RES <222> (1)..(1) <223> The stone is penicillamine. <220> <221> MOD_RES <222> (2)…(2) <223> The stone is N-Methyl-Arginine. <220> <221> MOD_RES <222> (7)…(7) <223> The stone is penicillamine. <220> <221> MOD_RES <222> (8)…(8) <223> The stone is (S)-2-amino-3-(4-tert-butyl-phenyl)propionic acid <220> <221> MOD_RES <222> (9)..(9) <223> The stone is beta-homoglutamic acid. <220> <221> MOD_RES <222> (10)..(10) <223> D form Pro <220> <221> MOD_RES <222> (11)..(11) <223> D form Lys <400> 8 Thank You Ser Asp Thr Leu Thank You Pro Lys 1 5 10

Claims

1. A pharmaceutical composition comprising an α4β7 integrin antagonist for the treatment of inflammatory bowel disease (IBD) in a subject, The aforementioned antagonist is administered orally to the subject twice a day at a dose of approximately 100 mg to approximately 200 mg. The antagonist is a peptide dimer compound comprising two peptides, or a pharmaceutically acceptable salt thereof, wherein each of the two peptides has the following sequence: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1) 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2) 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3) 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4) 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5) 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH ２ (Sequence No. 5) Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 6), Pen-(-Me-｡rgg-3er-sppThr-eumm n-Phe(-tttuu--(β-ホモ-Glu)--mysss)-- ２ (Fashion 66)、 Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 7), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-NH ２ (SEQ ID NO:7), Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 8), or Pen-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-NH ２ (SEQ ID NO: 8) It includes or consists of any of the following: Each of the two peptides contains a thioether bond between the 2-methylbenzoyl and the Pen, or a disulfide bond between the two Pens. The two peptides are linked by a linker moiety attached to the D-Lys amino acid of the two peptides. The linker portion is diglycolic acid (DIG). Pharmaceutical composition.

2. Each of the two peptides mentioned above is (a) The aforementioned sequence: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Glu)-(D-Lys)-OH (SEQ ID NO: 1) It consists of, each of the two peptides contains a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety bonded to the D-Lys amino acid of the two peptides, and the linker moiety is diglycolic acid (DIG), (b) The aforementioned sequence: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Gly-(D-Lys)-OH (SEQ ID NO: 2) It consists of, each of the two peptides contains a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety bonded to the D-Lys amino acid of the two peptides, and the linker moiety is diglycolic acid (DIG), (c) Said array: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-Pro-(D-Lys)-OH (SEQ ID NO: 3) It consists of, each of the two peptides contains a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety bonded to the D-Lys amino acid of the two peptides, and the linker moiety is diglycolic acid (DIG), (d) Each of the two peptides has the sequence: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Pro)-(D-Lys)-OH (SEQ ID NO: 4) It consists of, each of the two peptides contains a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety bonded to the D-Lys amino acid of the two peptides, and the linker moiety is diglycolic acid (DIG), (e) The aforementioned sequence: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-NH ２ (Sequence No. 5) It consists of, each of the two peptides includes a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety bonded to the D-Lys amino acid of the two peptides, wherein the linker moiety is diglycolic acid (DIG), or (f) The aforementioned sequence: 2-methylbenzoyl-(N-Me-Arg)-Ser-Asp-Thr-Leu-Pen-Phe(4-tBu)-(β-homo-Glu)-(D-Lys)-OH (SEQ ID NO: 5) It consists of, each of the two peptides contains a thioether bond between the 2-methylbenzoyl and the Pen, and the two peptides are linked by a linker moiety bonded to the D-Lys amino acid of the two peptides, wherein the linker moiety is diglycolic acid (DIG). The pharmaceutical composition according to claim 1.

3. The peptide dimer compound or a pharmaceutically acceptable salt thereof The pharmaceutical composition according to claim 1, or a pharmaceutically acceptable salt thereof.

4. The peptide dimer compound or a pharmaceutically acceptable salt thereof The pharmaceutical composition according to claim 1, or a pharmaceutically acceptable salt thereof.

5. The pharmaceutical composition according to any one of claims 1 to 4, wherein the peptide dimer compound or a pharmaceutically acceptable salt thereof is administered to the subject twice daily in doses of about 100.0, 112.5, 125.0, 137.5, 150.0, 162.5, 175, 187.5, or 200.0 mg.

6. The pharmaceutical composition according to claim 5, wherein the peptide dimer compound or a pharmaceutically acceptable salt thereof is administered to the subject twice daily at a dose of about 150 mg.

7. The pharmaceutical composition according to any one of claims 1 to 6, wherein the pharmaceutically acceptable salt of the peptide dimer compound is an acetate salt.

8. The pharmaceutical composition according to any one of claims 1 to 7, wherein the IBD is ulcerative colitis or Crohn's disease.

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