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CN-121986111-A - Stable GLP-1 peptides and dual agonist peptides directed against GLP-1R and GIPR and methods of use thereof

CN121986111ACN 121986111 ACN121986111 ACN 121986111ACN-121986111-A

Abstract

The present application provides protease-resistant glucagon-like peptide 1 (eGLP-1) peptides, as well as protease-resistant dual agonist peptides directed to GLP-1R and GIPR, methods of making these peptides, as well as compositions comprising protease-resistant peptides and methods of treatment using these peptides.

Inventors

  • SCHACHT ARNE
  • A. KUMAR
  • D. M. gangaia
  • K. Vasudewan
  • A. Kamala

Assignees

  • 生物编辑股份有限公司

Dates

Publication Date
20260505
Application Date
20240726
Priority Date
20230727

Claims (20)

  1. 1. An engineered dual agonist polypeptide comprising an amino acid sequence of the formula: Formula II (SEQ ID NO: 111): Y X 2 E G T X 6 X 7 S D X 10 S I X 13 X 14 D X 16 I A Q X 20 A X 22 V Q X 25 X 26 I A G G P S S G A P P（I） Wherein the method comprises the steps of X 2 =v or K; [ telpoiside=a ] X 6 = F, P or S; [ telpoiside=f ] X 7 = T, C or E; [ telpoiside=t ] X 10 = Y, C or E; [ telpofungin=y ] X 13 = A, S, Y, N, I, L, R, V or K; [ telpoiside=a ] X 14 = L, K, H or I; [ telpopeptid=l ] X 16 = K, R, H or V; [ telpoiside=k ] X 20 = K, R, H, N; [ telpoiside = K ] X 22 = F, A, P; [ telpoiside = F ] X 25 = W, P, K, H or I, [ telpofungin = W ] X 26 = L or V, Or formula IV (SEQ ID NO: 383): Y X 2 E G T X 6 X 7 S D X 10 S I X 13 X 14 D X 16 I A X 19 X 20 A X 22 V Q X 25 X 26 I A G G P S S G A P P（IV）, Wherein X 2 = V or K, X 6 = F, P or S, X 7 = T, C or E, X 10 = Y, C or E, X 13 = A, S, Y, N, I, L, R, V or K, X 14 = L, K, H or I, X 16 = K, R, H or V, X 19 = Q or V, X 20 = K、R、H、N;X 22 = F、A、P;X 25 = W, P, K, H or I, and X 26 = L or V.
  2. 2. The engineered dual agonist polypeptide of claim 1, wherein X 2 = V;X 6 =p or S, X 7 =c or E, X 10 =c or E, X 13 = S;X 14 = L, H or I, X 16 =h or V, X 20 =h or N, X 22 =a or P, X 25 =p or I, and X 26 =l or V.
  3. 3. The engineered dual agonist polypeptide of claim 1, comprising the amino acid sequence of any one of SEQ ID NOs 112 to 147 or SEQ ID NO 381.
  4. 4. The engineered dual agonist polypeptide of claim 1, comprising the amino acid sequence of any one of SEQ ID NOs 113 to 116.
  5. 5. An engineered glucagon-like peptide-1 (eGLP-1) polypeptide comprising an amino acid sequence of the formula: Formula I: H G E G T S E S D V S X 12 X 13 X 14 E G Q A A Q E X 22 X 23 A X 25 X 26 V D G X 30 （I） Wherein the method comprises the steps of X 12 = S or Q; X 12 = S, Q or Y; X 14 = I or L; x 22 = V, I or F; X 23 = V or I; X 25 = V, I or W; x 26 = I or V, and X 30 = R or S, Or formula III (SEQ ID NO: 382): X 1 G E G T S E S D V S X 12 X 13 X 14 E X 16 Q A X 19 X 20 E X 22 X 23 A X 25 X 26 V D G X 30 （III）, Wherein X 1 = H or Y, X 12 = S, Q, A or M, X 13 = S, Q, Y or M, X 14 = I or L, X 16 = G or M, X 19 = a or T, X 20 = Q, V or D, X 22 = V, I or F, X 23 = V or I, X 25 = V, I or W, X 26 = I or V, and X 30 = R or S.
  6. 6. The engineered polypeptide of claim 5 comprising the amino acid sequence of any one of SEQ ID No. 2 to SEQ ID No. 14 or any one of SEQ ID No. 372 to SEQ ID No. 380.
  7. 7. The engineered polypeptide of claim 1 or claim 5, wherein the engineered polypeptide is substantially resistant to proteolytic degradation.
  8. 8. The engineered polypeptide of claim 1 or claim 5, wherein the engineered polypeptide retains at least substantially the same receptor potency as a corresponding wild-type unmodified polypeptide.
  9. 9. The engineered polypeptide of claim 1 or claim 5, wherein the engineered polypeptide is substantially resistant to degradation by DPP-IV, enkephalinase, alpha-chymotrypsin, trypsin, elastase or pepsin.
  10. 10. An isolated polynucleotide encoding: an engineered dual agonist polypeptide comprising an amino acid sequence of formula II (SEQ ID NO: 111) or formula IV (SEQ ID NO: 383), and/or An engineered glucagon-like peptide-1 (eGLP-1) polypeptide comprising the amino acid sequence of formula I (SEQ ID NO: 1) or formula III (SEQ ID NO: 382).
  11. 11. A pharmaceutical composition comprising: an engineered dual agonist polypeptide comprising an amino acid sequence of formula II (SEQ ID NO: 111) or formula IV (SEQ ID NO: 383), and/or An engineered glucagon-like peptide-1 (eGLP-1) polypeptide comprising the amino acid sequence of formula I (SEQ ID NO: 1) or formula III (SEQ ID NO: 382); And a carrier.
  12. 12. A host cell comprising the polynucleotide of claim 10.
  13. 13. The host cell of claim 12, wherein the cell is a bacterial cell, a plant cell, a yeast cell, or an algal cell.
  14. 14. A kit comprising the pharmaceutical composition of claim 11 or the host cell of claim 12, and a carrier.
  15. 15. A method of treating or preventing a disease or disorder caused by or characterized by impaired hypoglycemia or insulin release, comprising administering to a subject in need of treatment an effective amount of the engineered dual agonist polypeptide of claim 1 and/or the engineered glucagon-like peptide-1 (eGLP-1) of claim 5, the pharmaceutical composition of claim 11, or the host cell of claim 12.
  16. 16. A method of treating or preventing a condition caused by or characterized by overweight and treating obesity, morbid obesity, obesity-related inflammation, obesity-related gallbladder disease, obesity-induced sleep apnea, metabolic syndrome, pre-diabetes, insulin resistance, glucose intolerance, type 2 diabetes, type 1 diabetes, hypertension, atherogenic dyslipidemia, atherosclerosis, arteriosclerosis, coronary heart disease, peripheral arterial disease, stroke or microvascular disease in a subject, comprising administering to a subject in need of treatment an effective amount of the engineered dual agonist polypeptide of claim 1 and/or the engineered glucagon-like peptide-1 (eGLP-1) of claim 5, the pharmaceutical composition of claim 11 or the host cell of claim 12.
  17. 17. The method of claim 15 or 16, wherein the disease or disorder is diabetes or obesity.
  18. 18. The method of claim 15 or 16, wherein the dual agonist polypeptide, the eGLP-1 polypeptide, or the pharmaceutical composition is administered orally, by injection, or transdermally.
  19. 19. The method of claim 15 or 16, wherein the subject is a human, cat, dog, or non-human primate.
  20. 20. A pharmaceutical composition comprising an engineered glucagon-like peptide-1 (eGLP-1) polypeptide comprising an amino acid sequence selected from any one of SEQ ID No. 112 to SEQ ID No. 147 or SEQ ID No. 381.

Description

Stable GLP-1 peptides and dual agonist peptides directed against GLP-1R and GIPR and methods of use thereof Cross Reference to Related Applications The present application claims priority from U.S. provisional patent application 63/529,271 filed on day 27 of 7 in 2023 and U.S. provisional patent application 63/540,427 filed on day 26 of 9 in 2023. Incorporation of the sequence Listing A sequence listing in st.26 XML format titled 2950-17_pct_st26 was created at 7.26, 2024, containing 575,627 bytes, prepared according to 37 CFR 1.822 to 1.824, and filed with the present application at 7.26, 2024, which is incorporated herein by reference in its entirety. Technical Field The present invention relates to variants and stable forms of glucagon-like peptide 1 (GLP-1), methods of use and methods of production thereof, as well as dual agonists against GLP-1R and GIPR and their delivery by in vivo delivery platforms such as genetically modified bacteria to deliver therapeutic activity directly to the mucosa of an animal in need thereof. Background Type 2 diabetes is a disease caused by hyperglycemia resulting from insulin resistance and relative lack of insulin. Type 2 diabetes mellitus (T2 DM) is most common and is characterized by a number of interrelated metabolic disorders. Although a variety of therapeutic approaches such as telipopeptide (tirzepatide), semaglutin (semaglutide), liraglutide (liraglutide), GLP-1, GIP (7dty_p), medi7219, exenatide (Exenatide), to name a few, have been available for T2DM, treatments for domestic cats (Felis catus) and other species still need to be further improved. Peptide therapeutics are increasingly being used in the treatment of T2 DM. Oral administration represents a significant medical advance but faces the challenges of gastrointestinal instability and inefficient absorption into the circulation. Peptides of exenatide, telipopeptide, semaglutin and liraglutide were administered subcutaneously, whereas Medi7219 and J229 were administered orally. Drugs designed for T2DM may function as single agonists or dual/triple agonists. See Pechenov et al ,"Development of an orally delivered GLP-1 receptor agonist through peptide engineering and drug delivery to treat chronic disease,"Scientific Reports11(1):22521 (2021). GLP-1, a glucagon derived peptide produced by intestinal L cells, and GIP, produced by K cells, are both used to treat type 2 diabetes. Both GLP-1 and GIP stimulate insulin secretion, GLP-1 can delay gastric emptying while GIP does not. Exenatide is a GLP-1 analogue, originally found in Ji Ladu exendin (Gila monster) saliva, （Bond, "Exenatide (Byetta) as a novel treatment option for type 2 diabetes mellitus,"Proceedings (Baylor University. Medical Center)19(3):281–284 (2006)）. liraglutide (4APD_A) with 53% amino acid identity to GLP-1 shares 97% sequence identity with GLP-1, and its addition of C16 fatty acid side chain promotes drug binding to circulating serum albumin （Garber, "Long-acting glucagon-like peptide 1 receptor agonists：a review of their efficacy and tolerability,"Diabetes Care34( journal 2) S279-84 (2011). Furthermore, as employed in exenatide, substitution of alanine at position 8 for glycine (GLP-1-Gly 8) significantly enhanced insulinotropic action (Lin et al ,"Oral Delivery of Pentameric Glucagon-Like Peptide-1 by Recombinant Lactobacillus in Diabetic Rats,"PloS One11(9):e0162733 (2016)）. GIPR and GLP-1R receptors each comprise an N-terminal extracellular domain (ECD), a central domain consisting of seven transmembrane alpha helices, and a C-terminal cytoplasmic domain that mediates intracellular signaling by physical binding to G proteins. Tenipotent agonists against GIPR and GLP-1 have similar peptide-receptor binding interfaces, but structural changes are observed in the extracellular loop (ECL 1, ECL 3) and transmembrane (TM 1, TM 3) regions. The N-terminus of telpofungin has good overlap in both receptors and interacts with the central transmembrane domain, resulting in conformational rearrangement to activate the receptor. The Tyr-1 to Met-14 residues of the telipopeptide agonist are not in contact with the extracellular domain of the receptor. It is believed that hydrophobic interactions are involved in the binding process. The binding interface of GIP spans the alpha-helical C-terminal region of the peptide, which contains Asp-15 to Lys-30 residues that interact with the receptor extracellular domain, as shown in figure 2. As shown in FIG. 2, the C-terminus of the GLP-1 agonist interacts with the extracellular domain, while the N-terminus of GLP-1 interacts with the transmembrane domain of the GLP-1R receptor. The figures originate from Cong et al ,"Molecular insights into ago-allosteric modulation of the human glucagon-like peptide-1 receptor,"Nature Communications,12(1):3763 (2021). The helical peptide exposes hydrophobic residues to ECD, indicating that its binding is predominantly dominated by hydrophobic interactions. Structural studies