Gipr inhibitors and methods of use

Abstract

The present disclosure provides variant GIP peptides and polypeptides capable of inhibiting GIPR and methods and uses thereof.

Description

[0001] Cross-reference to related applications

[0002] This application claims priority to U.S. Provisional Application 63 / 561,960, filed March 6, 2024, the contents of which are incorporated herein by reference. Technical Field

[0003] This invention relates to polypeptides with GIPR inhibitory activity. Background Technology

[0004] Glucose-dependent insulinotropic peptide (GIP) is an incretin hormone that coordinates nutrient uptake and systemic metabolism. It is secreted primarily by endocrine K cells located in the proximal small intestine. GIP is a stimulatory ligand of the GIP receptor (GIPR). Upon nutrient stimulation, GIP is secreted into the bloodstream and activates GIPR, leading to insulin secretion, lipid deposition in adipose tissue, decreased bone resorption, increased bone formation, and increased gastrointestinal blood flow. GIPR is expressed in pancreatic β cells, adipose tissue, the central nervous system, and leukocyte subsets, playing a major role in maintaining glucose homeostasis and controlling metabolism. Therefore, GIPR is of significant importance as a target for metabolic disorders such as type 2 diabetes and obesity, although its presence in certain endocrine tumors makes it a potential target for cancer therapy. In addition to the full-length GIP (1–42), truncated endogenous variants of wild-type GIP with inhibitory activity, namely GIP(1–30)NH2, GIP(3–42), and GIP(3–30)NH2, have been identified. However, new GIPR inhibitors with improved properties, such as increased inhibitory activity, are still needed. Summary of the Invention

[0005] This invention relates to polypeptides with GIPR inhibitory activity.

[0006] In one embodiment, a polypeptide is provided comprising an N-terminal portion and a C-terminal portion, wherein the N-terminal portion comprises an amino acid sequence having substituted 0, 1, 2, 3, 4, 5, or 6 amino acids relative to any one of SEQ ID NO: 9-16, 25-48, and 73-114, and wherein the C-terminal portion comprises an amino acid sequence that is at least 70% identical to any one of SEQ ID NO: 2 and 166-168.

[0007] In one embodiment, a peptide is provided comprising an amino acid sequence having 0, 1, 2, 3, 4, 5, or 6 amino acid substitutions relative to any one of SEQ ID NO: 9-16, 25-48, and 73-114.

[0008] In one embodiment, a nucleic acid molecule encoding the polypeptide or peptide described herein is provided.

[0009] In one embodiment, a vector comprising the nucleic acid molecules described herein is provided.

[0010] In one embodiment, a host cell comprising the nucleic acid molecule or vector described herein is provided.

[0011] In one embodiment, a pharmaceutical composition is provided comprising the polypeptide, peptide, nucleic acid molecule or carrier described herein, and pharmaceutically acceptable loaders, excipients and / or stabilizers.

[0012] In one embodiment, a polypeptide, peptide, nucleic acid molecule, carrier, or pharmaceutical composition as described herein is provided for inhibiting GIPR signaling in cells.

[0013] In one embodiment, a method for inhibiting GIPR signaling in cells is provided, comprising contacting the cells with a polypeptide, peptide, nucleic acid molecule, carrier, or pharmaceutical composition described herein.

[0014] In one embodiment, a method is provided for treating or preventing a disease or condition related to GIPR signaling in a subject, comprising administering to the subject a polypeptide, peptide, nucleic acid molecule, carrier, or pharmaceutical composition described herein.

[0015] In one embodiment, the use of polypeptides, peptides, nucleic acid molecules, carriers, or pharmaceutical compositions as described herein for inhibiting GIPR signaling in cells is provided.

[0016] In one embodiment, the use of a polypeptide, peptide, nucleic acid molecule, carrier, or pharmaceutical composition as described herein is provided for the treatment or prevention of a disease or condition in a subject that is associated with GIPR signaling. Detailed Implementation

[0017] The inventors have discovered peptides that can be used to generate N-terminal variants of GIP with GIPR inhibitory activity. These peptides can be used to replace the N-terminal portion of GIP or GIP analogs to generate GIP variant polypeptides with GIPR inhibitory activity. As exemplified herein, the GIP variant polypeptides are generated via peptide synthesis. As shown in Example 1, the GIP variant polypeptides are approximately 100 times more potent than previously reported truncated GIP variants.

[0018] In this disclosure, sequences, compositions, and methods for carrying out the invention are presented by way of examples and embodiments. However, the invention is not limited to the described examples and embodiments, and those skilled in the art will understand that many other embodiments of the invention are possible without departing from the basic concept of the invention, and any such modifications will also fall within the scope of the invention. Other styles and configurations of the invention are contemplated to be readily incorporated into the teachings of the invention, and these configurations will be shown and described for purposes of clarity and disclosure, rather than for limitation.

[0019] The molecules of the present invention

[0020] It provides peptides that offer GIP inhibitory activity and can be incorporated into peptides, such as those in GIP variants.

[0021] In some embodiments, a polypeptide comprising an N-terminal portion and a C-terminal portion is provided, wherein the polypeptide is a GIP variant and a GIPR inhibitor. As used herein, “GIP variant,” “GIP variant polypeptide,” “GIP derivative,” or “GIP derivative polypeptide” according to the invention refers to a polypeptide derived from GIP, such as full-length human GIP (SEQ ID NO: 1) or truncated human GIP of 1-30, wherein the N-terminal portion comprises the peptide of the invention as described herein, and the C-terminal portion comprises at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 99%, at least 99.9%, or 100% of the same amino acid sequence as any one of SEQ ID NO: 2 and 166-168.

[0022] The GIP variants of this invention are modified GIP polypeptides containing one or more structural differences compared to wild-type GIP polypeptides. As used herein, the term "wild-type GIP" refers to a naturally occurring GIP structure found in an organism. As used herein, "wild-type GIP" refers to a GIP polypeptide isolated from an organism or a synthetically produced GIP peptide that has the same structure and / or function as GIP found in an organism. The terms "naturally occurring GIP," "native GIP," and "wild-type GIP" are used interchangeably.

[0023] Compared to wild-type GIP, GIP variants contain mutations, substitutions, additions, deletions, truncations, or other structural variations. In some embodiments, GIP variants contain mutations or other structural variations in the receptor-binding region of wild-type GIP. In some embodiments, GIP variants contain a truncation of the C-terminal portion of wild-type GIP.

[0024] As used herein, the term "ligand" refers to a molecule that specifically binds to one or more receptors. As used herein, "antagonist" is a polypeptide, peptide, small molecule, or other compound (ligand) that inhibits one or more biological activities induced by receptor activation, activation, or signaling. As used herein, "agonist" is a polypeptide, peptide, small molecule, or other compound (ligand) that induces one or more biological activities induced by receptor activation, activation, or signaling. As used herein, "antagonistic effect" refers to the activity of the antagonist, and "agonistic effect" refers to the activity of the agonist. As used herein, the terms "GIPR inhibitor" and "GIPR antagonist" are used interchangeably to refer to a polypeptide, peptide, small molecule, or other compound that inhibits one or more biological activities induced by GIPR activation, activation, or signaling. As used herein, the term "GIPR agonist" refers to a polypeptide, peptide, small molecule, or other compound that induces one or more biological activities induced by GIPR activation, activation, or signaling.

[0025] GIPR inhibitors can prevent ligand binding by partially or completely occupying one or more sites on GIPR required for its interaction (ortho-articulate inhibitors). For example, GIPR inhibitors can block the binding of natural GIP ligands to GIPR, preventing normal GIPR activation, or block the binding of synthetic GIPR agonists to GIPR. Alternatively, GIPR inhibitors can prevent ligand binding by binding to sites on GIPR and inducing GIPR to adopt one or more conformations that the ligand cannot recognize (allosteric inhibitors). GIPR inhibitors may inhibit all or only part of the GIPR intracellular signaling pathway.

[0026] The GIP variants of the present invention are GIPR antagonists. In some embodiments, the GIP variants may exhibit at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 50, or 100 times higher GIPR antagonism compared to truncated wild-type GIPs (e.g., GIP(1-30)NH2, GIP(3-42), GIP(3-30)NH2). In some embodiments, the GIP variants are antagonists that inhibit one or more biological activities of GIPR, as described herein. The amount of agonistic effect provided by a ligand is typically measured using its EC50 value. The EC50 of a given agonist ligand can be measured by determining the concentration of the agonist ligand required to elicit half of the maximum biological response. A smaller EC50 value measured at agonist ligand concentration indicates increased agonistic effect because a lower concentration of agonist ligand is required to elicit the maximum biological response. In contrast, the activity of the antagonist can be measured using the IC50 value obtained from an inhibitory potency assay. IC50 is generally defined as the antagonist concentration that inhibits 50% of the signal induced by a reference agonist via the receptor. IC50 is sometimes reported as pIC50, which is the negative logarithm of the IC50 value in moles per liter (molar concentration or M). In some embodiments, the GIP variants of the present invention inhibit GIPR with pIC50 values ​​greater than 7.00, 7.25, 7.50, 7.75, 8.00, 8.25, or 8.50. pIC50 can be measured using techniques known in the art, such as by inhibiting calcium flux induced by GIP agonistic activity of GIPR expressed on HEK cells.

[0027] When a receptor is activated by an agonist ligand, it may exhibit more than one biological response. The magnitude of the agonist or antagonist effect of the ligand can be measured relative to one or more biological responses exhibited by the activated receptor. Agonism can be measured by one or more of the following: increased receptor phosphorylation upon ligand binding; increased G protein signaling through the receptor upon ligand binding; increased recruitment of repressor proteins on the receptor upon ligand binding; increased intracellular calcium ion flux induction upon ligand binding; increased intracellular cyclic AMP production upon ligand binding; and / or increased receptor internalization upon ligand binding. Antagonism can be measured by inhibiting one or more of the above-mentioned receptor responses. The measurement of agonist or antagonist effects can be determined using conventional methods known in the art.

[0028] Calcium flux assays measure intracellular calcium flux induced by receptor agonism when a cell receptor binds to an agonist, such as GIPR being activated by GIP. Calcium flux assays are known in the art and can be disclosed as in WO 2008 / 012689 or, as in Gaertner et al., 2008. PNASThe procedure was performed as disclosed in , 105(46):17706-17711. In the exemplary calcium flux assay, Chinese hamster ovary (CHO) cells or human embryonic kidney (HEK) cells engineered to express GIPR were seeded at 20,000 cells / well in the wells of a 384-well plate with a black-walled, clear-bottomed plate. The test sample of the variant ligand was diluted in PBS supplemented with 1% BSA and 25 mM HEPES to generate a dilution series for the dose-response assay: a 12-point dose-response starting at 688 nM, with each treatment at a 2.5-fold dilution interval. The cells were loaded with the calcium-sensitive fluorescent dye (Screen Quest™ Fluo-8 Wash-Free Calcium Assay Kit, AAT Bioquest) according to the manufacturer’s instructions, followed by the addition of the test sample diluent or a separate medium. After 5 minutes, the cells were stimulated with 100 nM of the test or reference sample, and the fluorescence signal (excitation 490 nm, emission 525 nm) was recorded.

[0029] The isolation assay measures the internalization and / or downregulation of surface receptors on cells in response to cell exposure to receptor ligands. Isolation assays are known in the art and can be disclosed as in WO 2008 / 012689 or, as in Gaertner et al., 2008. PNAS The assay was performed as disclosed in , 105(46):17706-17711. In the exemplary isolation assay, CHO cells expressing the receptor of interest were seeded at a density of 80,000 cells / well. After overnight incubation, the medium was removed and replaced with medium containing the ligand, and the cells were incubated at 37°C for 1 hour. The medium was then removed, and the cells were fixed with 4% paraformaldehyde and washed twice with PBS. The cells were then labeled with fluorescently labeled anti-ligand antibodies in PBS supplemented with 1% BSA (PBS-1% BSA) on ice for 1 hour. The plate was washed three times with PBS-1% BSA, and the fluorescence value of each well was measured using a FLEXstation fluorometer (Molecular Devices). The results are expressed as a percentage control level of the surface receptor: 100 (mean fluorescence [with added ligand, anti-ligand] - mean negative control fluorescence [anti-ligand]) / (mean positive control fluorescence [without added ligand, anti-ligand] - mean fluorescence [anti-ligand]).

[0030] In one embodiment, the polypeptide of the present invention comprises an amino acid sequence having 0, 1, 2, 3, 4, 5, or 6 amino acid substitutions relative to any one of SEQ ID NO: 9-16, 25-48, and 73-114. In some embodiments, the amino acid sequence is located near the N-terminus of the polypeptide. In some embodiments, the position of the amino acid sequence is such that the starting position of the amino acid sequence is located within 15 residues of the N-terminus of the polypeptide, for example, within 15, 12, 10, 8, 6, 5, 4, 3, 2, 1 residues of the N-terminus, or is formed by the N-terminus of the polypeptide.

[0031] In one embodiment, the polypeptide of the present invention comprises an amino acid sequence having 0, 1, 2, 3, 4, 5, or 6 amino acid substitutions relative to any one of SEQ ID NO: 17-24, 49-72, and 115-165. In one embodiment, the polypeptide of the present invention comprises or consists of the amino acid sequence of any one of SEQ ID NO: 17-24, 49-72, and 115-165.

[0032] In some embodiments, the polypeptide according to the invention comprises an N-terminal portion and a C-terminal portion, wherein the N-terminal portion comprises an amino acid sequence having 0, 1, 2, 3, 4, 5, or 6 amino acid substitutions relative to any one of SEQ ID NOs: 9-16, 25-48, and 73-114, and the C-terminal portion comprises an amino acid sequence that is at least 70% identical to any one of SEQ ID NOs: 2 and 166-168. In some embodiments, the C-terminal portion is truncated relative to wild-type GIP, for example, truncated by 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or more C-terminal amino acids relative to wild-type GIP. In some embodiments, the truncated C-terminal portion of the GIP is SEQ ID NO: 166 or 168, which comprises a truncation of 13 C-terminal amino acids relative to human GIP.

[0033] In some embodiments, the polypeptide or peptide according to the invention comprises the amino acid sequence of any one of SEQ ID NO: 9-165 or a functional fragment thereof that retains GIPR inhibitory activity.

[0034] In some embodiments, the N-terminal portion comprises the amino acid sequence XXXXXXXISXY (SEQ ID NO: 3), where X is any amino acid. In some embodiments, the N-terminal portion comprises the amino acid sequence [D or W or S or I][S or G or W or L][F or W or G or N or C][F or H or W or L][R or E or I or L or A or P or G][N or R or P or C or T][W or F]IS[Y or D]Y (SEQ ID NO: 4). In some embodiments, the N-terminal portion comprises the amino acid sequence XXXXXXFISDY (SEQ ID NO: 5), where X is any amino acid. In some embodiments, the N-terminal portion comprises the amino acid sequence [D or W or S or I][S or G or W or L][F or W or G or N or C][F or H or W or L][R or E or I or L or A or P or G][N or R or P or C or T]FISDY (SEQ ID NO: 6). In some embodiments, the N-terminal portion comprises the amino acid sequence [D or I][L or G][C or W][F or W][E or G][T or R]FISDY (SEQ ID NO: 7). In some embodiments, the N-terminal portion comprises the amino acid sequence DGW[F or W][E or G][T or R]FISDY (SEQ ID NO: 8). In such embodiments, the C-terminal portion comprises an amino acid sequence that is at least 70% identical to any one of SEQ ID NO: 2 and 166-168.

[0035] In some embodiments, the N-terminal portion consists of no more than 11 amino acids, for example, no more than 11 or 10 amino acids. In some embodiments, the N-terminal portion consists of 11 amino acids or 10 amino acids. In one embodiment, the N-terminus of the C-terminal portion is directly linked to the C-terminus of the N-terminal portion, i.e., the N-terminal portion and the C-terminal portion are directly linked. In some embodiments, the N-terminal portion is linked to the C-terminal portion via a peptide linker. In some embodiments, the N-terminal portion is located at the extreme N-terminus of the polypeptide.

[0036] In some embodiments, a peptide is provided comprising an amino acid sequence having 0, 1, 2, 3, 4, 5, or 6 amino acids substituted relative to any one of SEQ ID NO: 9-16, 25-48, and 73-114. In some embodiments, the amino acid sequence is XXXXXXXISXY (SEQ ID NO: 3), where X is any amino acid. In some embodiments, the amino acid sequence is [D or W or S or I][S or G or W or L][F or W or G or N or C][F or H or W or L][R or E or I or L or A or P or G][N or R or P or C or T][W or F]IS[Y or D]Y (SEQ ID NO: 4). In some embodiments, the amino acid sequence is XXXXXXFISDY (SEQ ID NO: 5), where X is any amino acid. In some embodiments, the amino acid sequence [D or W or S or I][S or G or W or L][F or W or G or N or C][F or H or W or L][R or E or I or L or A or P or G][N or R or P or C or T]FISDY (SEQ ID NO: 6). In some embodiments, the peptide comprises the amino acid sequence [D or I][L or G][C or W][F or W][E or G][T or R]FISDY (SEQ ID NO: 7). In some embodiments, the peptide comprises the amino acid sequence DGW[F or W][E or G][T or R]FISDY (SEQ ID NO: 8).

[0037] Polypeptides and peptides are polymers comprising amino acids linked by peptide bonds. As used herein, the term "amino acid" is used to describe any amino acid that can be incorporated into a polypeptide or peptide, whether native or otherwise. An amino acid is a small molecule comprising an amine (-NH₂) group, a carboxyl group (-COOH), and a variable side chain (R group) characteristic of each amino acid. Amino acids are covalently linked by peptide bonds between the amine group of one amino acid and the carboxyl group of another amino acid to form a polypeptide. The amino acids within a polypeptide are generally referred to in the art as "residues".

[0038] Methionine residues may be conservatively substituted with non-oxidizable amino acid analogs or amino acid derivatives to reduce complications caused by methionine oxidation during synthesis. In some embodiments, one or more methionine residues in the polypeptides or peptides of the present invention, including SEQ ID NO: 17-24, 49-72, and 115-165, are conservatively substituted with amino acid analogs or amino acid derivatives, such as, but not limited to, ortholeucine (Nle). C-terminal residues of the polypeptide may be amidated. In some embodiments, the polypeptide of any one of SEQ ID NO: 2, 17-24, 49-72, and 115-168 comprises C-terminal amidation. It is known in the art that N-terminal glutamine or glutamate can spontaneously convert to pyroglutamic acid under various in vitro and in vivo conditions (Cao et al., 2022). J. Pharm. Sci. (111:335-344). In some embodiments, the N-terminal residues designated herein as glutamine (Q) are present in the form of pyroglutamate.

[0039] Peptides and peptides, including the GIP variants of the present invention, may contain modifications such as phosphorylation, glycosylation, ubiquitination, nitrosation, methylation, acetylation, esterification, acylation, isopreneation, alkylation, oxidation, or other modifications known in the art. In some embodiments, the GIP variants described herein contain one or more modifications selected from the group consisting of phosphorylation, glycosylation, ubiquitination, nitrosation, methylation, acetylation, esterification, acylation, isopreneation, alkylation, oxidation, or other modifications known in the art at their N-terminus and / or C-terminus.

[0040] Polypeptides and peptides, including the GIP variants of the present invention, may comprise non-proteinogenic amino acids and / or amino acid analogs, including artificial, synthetic, modified, or non-natural amino acids other than the 20 genetically encoded amino acids known in the art. Examples of non-proteinogenic amino acids or amino acid analogs that may be incorporated into the polypeptides or peptides of the present invention include, but are not limited to: ortholeucine, β-amino acids, homoamino acids, synthetic proline and pyruvate derivatives, 3-substituted alanine derivatives, glycine derivatives, cyclically substituted phenylalanine and tyrosine derivatives, substituted leucine derivatives, ornithine, linear core amino acids, N-methyl amino acids, N-acetyl amino acids, and amino acids with synthetic R-groups. Polypeptides and peptides may also comprise amino acid derivatives. As used herein, the term "amino acid derivative" describes an amino acid derived from a modification of one of the 20 genetically encoded amino acids. Amino acid derivatives may be synthetic, for example, through chemical reactions, or they may be naturally occurring in a living organism, such as in vivo metabolites. An example of an amino acid derivative is pyroglutamate / pyroglutamic acid, a glutamine cyclized derivative in which the free amino group of glutamate is cyclized to form a lactam. In some implementations, the GIP variants described herein comprise substitutions for one or more non-proteinogenic amino acids and / or amino acid analogs.

[0041] As used herein, the term "sequence identity" refers to the percentage of sequence identity between two nucleic acid (polynucleotide) or two amino acid (polypeptide) sequences. To determine the percentage of identity between two amino acid sequences or two nucleic acid sequences, the sequences are aligned for optimal comparison (e.g., a gap may be introduced into the first amino acid or nucleic acid sequence to optimize alignment with the second amino acid or nucleotide sequence). The amino acid residues or nucleotides at the corresponding amino acid or nucleotide positions are then compared. When a position in the first sequence is occupied by the same amino acid residue or nucleotide as the corresponding position in the second sequence, the molecules are identical at that position. The percentage of identity between two sequences is a function of the number of identical positions shared by the sequences (i.e., identity % = number of identical overlapping positions / total number of positions multiplied by 100%). Mathematical algorithms can also be used to determine the percentage of identity between two sequences. A non-limiting example of a mathematical algorithm for comparing two sequences is Karlin, S. and Altschul, S., 1990. PNAS The algorithm proposed in , 87(6):2264-2268, which was first proposed by Karlin, S. and Altschul, S., 1993, PNASModifications were made in 90(12):5873-5877. This type of algorithm was incorporated into the BLAST procedure. BLAST nucleotide searches can be performed using the NBLAST nucleotide procedure parameter set, for example, score=100, word length=12, to obtain nucleotide sequences homologous to a given nucleic acid molecule. BLAST protein searches can be performed using the XBLAST procedure parameter set, for example, score=50, word length=3, to obtain amino acid sequences homologous to a given polypeptide. For obtaining vacancy alignments for comparative purposes, methods such as Altschul et al., 1997, can be used. Nucleic Acids Res. The method described in 25(17):3389-3402 uses Gapped BLAST. Alternatively, PSI-BLAST can be used to perform iterative searches to detect distance relationships between molecules. When using BLAST, Gapped BLAST, and PSI BLAST procedures, the default parameters of the corresponding procedures (e.g., XBLAST and NBLAST) can be used (see, for example, the NCBI website). Another non-limiting example of a mathematical algorithm for sequence comparison is Myers, E. and Miller, W., 1988. Bioinformatics The algorithm described is 4(1):11-17. This algorithm is incorporated into the ALIGN program, which is part of the GCG sequence alignment software package. When comparing amino acid sequences using the ALIGN program, the PAM120 weighted residue table, a 12-fold vacancy length penalty, and a 4-fold vacancy penalty can be used. The percentage of identity between two sequences can be determined using techniques similar to those described above, regardless of the presence of vacancy. When calculating the percentage of identity, only perfect matches are typically counted.

[0042] Conservative substitution is a substitution in a peptide or polypeptide sequence that does not result in a significant loss of function or only a slight loss of function. If the loss of function relative to the function of a polypeptide having an unsubstituted sequence is less than 20%, less than 15%, less than 10%, less than 6%, or less than 4%, then such loss of function due to one or more conservative substitutions is considered insignificant. Conservative substitution is generally a substitution in which an amino acid side chain is replaced by an amino acid side chain that is related to or has similar physicochemical properties to the substituted residue. For example, such conservative substitution can be performed using one of the 20 natural amino acids according to Table 1, wherein amino acids in the same block of the middle column and preferably amino acids in the same row of the right column can substitute for each other. Conservative substitution can also be performed using amino acid analogs or amino acid derivatives, such as replacing methionine with ortholeucine.

[0043]

[0044] The polypeptides and peptides of the present invention can be prepared in a variety of ways, for example, using known protein chemistry techniques (e.g., chemical peptide synthesis) or molecular biology techniques (i.e., genetic engineering and fermentation - commonly referred to as biotechnology).

[0045] The polypeptides or peptides of the present invention can be prepared using known protein chemistry techniques, such as those described in Gaertner et al., 2008. PNAS , 105(46):17706-17711 or Akondi et al., 2021, Chimia As described in , 75(6):489-494.

[0046] The method for preparing the polypeptides or peptides of the present invention involves in vitro chemical synthesis. The polypeptides or peptides can be synthesized partially or entirely using chemical means. For example, solid-phase peptide synthesis can be used, such as methods based on tBoc or Fmoc chemistry. Enzymatic synthesis can also be used partially or entirely.

[0047] Furthermore, the polypeptides or peptides of the present invention can be prepared using genetic engineering. The polypeptides or peptides of the present invention can be produced by culturing host cells containing nucleic acid molecules expressing the polypeptides or peptides of the present invention under conditions that induce polypeptide or peptide expression. In some embodiments, the host cells are bacterial cells (e.g., *Escherichia coli*), yeast cells (e.g., *Saccharomyces cerevisiae*), or mammalian cells (e.g., human cells, mouse cells, CHO cells, HEK cells, HeLa cells).

[0048] Biosynthesis other than expression in host cells can be used; for example, the polypeptides or peptides of the present invention can be produced by in vitro RNA translation. For example, the polypeptides or peptides of the present invention can also be prepared by digesting longer polypeptides with proteases.

[0049] Biological methods, including genetic engineering, fermentation, and expression, are generally limited to the production of L-amino acid-based peptides, but manipulation of in vivo or in vitro translational mechanisms (e.g., aminoacyl-tRNA molecules) can be used to introduce D-amino acids (or other non-natural amino acids, such as iodotyrosine or methylphenylalanine, azidohoalanine, etc.). However, in cases involving D-amino acids, chemical synthesis is preferred. The peptides or polypeptides of the present invention may have covalent modifications at the C-terminus and / or N-terminus.

[0050] This invention provides polypeptides and peptides as described above, and also provides nucleic acid molecules encoding said polypeptides and peptides. In some embodiments, the nucleic acid molecules encoding the polypeptides and peptides of this invention are RNA or DNA. Those skilled in the art can design or identify nucleic acid molecules encoding said polypeptides or peptides of this invention using methods known in the art. In some embodiments, the nucleic acid molecules encoding the polypeptides or peptides of this invention are incorporated into a vector, such as a plasmid, a free organism, an artificial chromosome, a virus, or a viral vector. In some embodiments, the nucleic acid molecules encoding the polypeptides or peptides of this invention, or the vector containing said nucleic acid molecules, are contained within a host cell to enable the expression of the polypeptides or peptides of this invention. In some embodiments, the host cell is a bacterial cell, yeast cell, vertebrate cell, mammalian cell, human cell, or cell of an immortalized cell line, such as CHO cells, HEK cells, or HeLa cells.

[0051] Pharmaceutical Composition

[0052] The peptides or polypeptides described herein may be incorporated into pharmaceutical compositions for administration to subjects in need. The pharmaceutical compositions may contain pharmaceutically acceptable carriers, excipients, and / or stabilizers. The pharmaceutical compositions may be provided for use as medicines. The preparation of pharmaceutical compositions is well known to those skilled in the art.

[0053] The pharmaceutical compositions according to the invention can be administered to a subject in a therapeutically effective amount. As used herein, "therapeutically effective amount" refers to the amount of the pharmaceutical composition, polypeptide, or peptide therein that effectively provides a therapeutic, preventive, or diagnostic benefit to the subject. In some embodiments, the therapeutically effective amount of the pharmaceutical composition is the amount that induces a clinical response in a subject when treating a particular disease or condition. The determination of the therapeutically effective amount of the pharmaceutical composition is entirely within the capabilities of those skilled in the art. The therapeutically effective amount can vary depending on a variety of factors, such as the subject's condition, weight, sex, and age.

[0054] The pharmaceutical compositions described herein can be prepared in various pharmaceutical dosage forms, such as immediate-release, controlled-release, sustained-release, or targeted drug delivery systems. Commonly used dosage forms include, for example, solutions and suspensions, (micro)emulsions, ointments, gels, creams, pastes, foams, suppositories, vaginal suppositories, implants, patches, liposomes, tablets, sugar-coated pills, sugar tablets, soft-shell or hard-shell capsules, amorphous or crystalline powders, effervescent powders or tablets, aerosols, and lyophilized formulations. Depending on the route of administration used, special devices may be required to apply or administer a dosage form, such as syringes and needles, inhalers, pumps, injection pens, applicators, special flasks, or other administration devices, which may also be implanted in the body. The pharmaceutical dosage forms described herein can be manufactured by any method known in the art, such as conventional mixing, sieving, dissolving, melting, granulation, sugar pilling, tableting, suspension, extrusion, spray drying, grinding, emulsification, (nano / micro) encapsulation, embedding, or lyophilization processes.

[0055] In one embodiment, the pharmaceutical composition of the present invention may comprise more than one polypeptide or peptide, or a nucleic acid or carrier encoding said polypeptide or peptide of the present invention. In one embodiment, the pharmaceutical composition of the present invention may comprise (a) at least one polypeptide or peptide, or a nucleic acid or carrier encoding said polypeptide or peptide of the present invention; and (b) at least one additional agent or therapeutic agent. In one embodiment, the additional agent or therapeutic agent may be, for example, an anti-inflammatory drug, an immunosuppressant, an antibiotic, an antiviral agent, a small molecule drug, a cytotoxic agent, or an antibody. In some embodiments, the additional agent or therapeutic agent is formulated in combination with at least one polypeptide or peptide or a nucleic acid or carrier encoding said polypeptide or peptide, or formulated as a separate pharmaceutical composition, for example, for simultaneous or sequential administration. In some embodiments, the additional agent or therapeutic agent is bound to the polypeptide or peptide of the present invention.

[0056] The pharmaceutical compositions provided herein may also contain pharmaceutically acceptable carriers, excipients, and / or stabilizers known in the art. Acceptable carriers, excipients, or stabilizers are non-toxic to the recipient at the dose and concentration, and may include, for example, but not limited to, buffers such as phosphates, citrates, and other organic acids; antioxidants (including ascorbic acid and methionine); preservatives (e.g., octadecyl dimethyl benzyl ammonium chloride; hexamethyl diammonium chloride; benzalkonium chloride; benzyl chloride; phenol, butyl or benzyl alcohol; alkyl esters of p-hydroxybenzoate such as methylparaben or propylparaben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol); low molecular weight (less than about 10 residues) polyphenols. Peptides; proteins such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates (including glucose, mannose, or dextran); chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose, or sorbitol; salt-forming counterions such as sodium ions; metal complexes (such as Zn-protein complexes); and / or nonionic surfactants such as TWEEN™, PLURONICS™, or polyethylene glycol (PEG).

[0057] Methods and uses

[0058] In some embodiments, the polypeptides or peptides of the present invention are used to inhibit GIPR signaling in cells. Inhibition of GIPR signaling includes GIPR antagonism as described herein. In some embodiments, the cells are, but are not limited to, pancreatic β cells, pancreatic α cells, pancreatic υ cells, adipocytes, pericytes, mesothelial cells, endothelial cells, osteocytes, osteoblasts, osteoclasts, cardiomyocytes, neurons, oligodendrocytes, gastric mucus-secreting cells, ciliated cells, T cells, myeloid cells, immortalized cells expressing GIPR, or cancer cells expressing GIPR. In some embodiments, the cells are primary cells or transformed cells that naturally express GIPR or are genetically modified to express GIPR. In some embodiments, the cells are contacted with the polypeptides or peptides of the present invention in vitro. In some embodiments, the cells are contacted with the polypeptides or peptides of the present invention in vivo or in vitro.

[0059] In some embodiments, the polypeptides or peptides of the present invention are used to treat or prevent a subject from having a disease or condition associated with GIPR signaling. As used herein, "a disease or condition associated with GIPR signaling" means a disease or condition in which inhibition (e.g., antagonism) of GIPR signaling in the subject results in complete or partial treatment or prevention. In some embodiments, a disease or condition associated with GIPR signaling is a metabolic disorder, GIPR-mediated cancer, or a cancer expressing GIPR. In some embodiments, a disease or condition associated with GIPR signaling is obesity, hypertension, cardiovascular disease, non-alcoholic fatty liver disease, Cushing's syndrome, Alzheimer's disease, Parkinson's disease, metabolic syndrome, or diabetes (e.g., type I or type II). In some embodiments, the subject is a human.

[0060] As used herein, “treating” or “preventing” or “prevention” refers to methods for achieving beneficial or desired outcomes. Beneficial or desired outcomes may include, but are not limited to, alleviating or improving one or more symptoms or conditions, reducing the extent of disease, stabilizing the disease state, preventing disease progression, preventing disease spread, delaying or slowing disease progression, suppressing disease, delaying or slowing disease onset, and improving or alleviating the disease state. “Treatment” or “prevention” may also mean prolonging a patient’s survival beyond the expected survival without treatment, and may also mean temporarily suppressing disease progression or preventing disease onset. The distinction between “treatment” and “prevention” is that “treatment” typically occurs in subjects who already have the disease or condition, while “prevention” typically occurs in subjects who do not have the disease or condition. It should be understood that there may be overlap between treatment and prevention. For example, it is possible to “treat” a subject’s disease while simultaneously “preventing” the symptoms or progression of the disease.

[0061] The specific implementation schemes disclosed herein include, but are not limited to, the following: 1. A polypeptide comprising an N-terminal portion and a C-terminal portion, wherein the N-terminal portion comprises an amino acid sequence having substituted 0, 1, 2, 3, 4, 5, or 6 amino acids relative to any one of SEQ ID NO: 9-16, 25-48, and 73-114, and wherein the C-terminal portion comprises an amino acid sequence that is at least 70% identical to any one of SEQ ID NO: 2 and 166-168.

[0062] 2. The polypeptide as described in embodiment 1, wherein the amino acid substitution is a conservative substitution.

[0063] 3. The polypeptide of embodiment 1, wherein the N-terminal portion comprises an amino acid sequence of any one of SEQ ID NO: 9-16, 25-48, and 73-114.

[0064] 4. The polypeptide of embodiment 1, wherein the N-terminal portion comprises the amino acid sequence XXXXXXXISXY (SEQ ID NO: 3), wherein X is any amino acid.

[0065] 5. The polypeptide of embodiment 4, wherein the N-terminal portion comprises the amino acid sequence [D or W or S or I][S or G or W or L][F or W or G or N or C][F or H or W or L][R or E or I or L or A or P or G][N or R or P or C or T][W or F]IS[Y or D]Y (SEQ ID NO: 4).

[0066] 6. The polypeptide of embodiment 1, wherein the N-terminal portion comprises the amino acid sequence XXXXXXFISDY (SEQ ID NO: 5), wherein X is any amino acid.

[0067] 7. The polypeptide of embodiment 6, wherein the N-terminal portion comprises the amino acid sequence [D or W or S or I][S or G or W or L][F or W or G or N or C][F or H or W or L][R or E or I or L or A or P or G][N or R or P or C or T]FISDY (SEQ ID NO 6).

[0068] 8. The polypeptide of embodiment 6, wherein the N-terminal portion comprises the amino acid sequence [D or I][L or G][C or W][F or W][E or G][T or R]FISDY (SEQ ID NO: 7).

[0069] 9. The polypeptide of embodiment 6, wherein the N-terminal portion comprises the amino acid sequence DGW[F or W][E or G][T or R]FISDY (SEQ ID NO: 8).

[0070] 10. The polypeptide according to any one of embodiments 1-9, wherein the N-terminal portion consists of 10-11 amino acids.

[0071] 11. The polypeptide of any one of embodiments 1-10, wherein the C-terminal portion comprises the amino acid sequence of any one of SEQ ID NO:2 and 166-168.

[0072] 12. The polypeptide of any one of embodiments 1-11, wherein the polypeptide inhibits GIPR with a pIC50 greater than 7.00, greater than 7.25, greater than 7.50, greater than 7.75, greater than 8.00, greater than 8.25 or greater than 8.50.

[0073] 13. A peptide comprising an amino acid sequence having 0, 1, 2, 3, 4, 5 or 6 amino acid substitutions relative to any one of SEQ ID NO: 9-16, 25-48 and 73-114.

[0074] 14. The peptide as described in embodiment 13, wherein the amino acid substitution is a conservative substitution.

[0075] 15. The peptide as described in Embodiment 13, comprising the amino acid sequence of any one of SEQ ID NO: 9-16, 25-48, and 73-114.

[0076] 16. The peptide as described in embodiment 13, comprising the amino acid sequence XXXXXXX ISXY (SEQ ID NO: 3), wherein X is any amino acid.

[0077] 17. The peptide as described in embodiment 16, comprising the amino acid sequence [D or W or S or I][S or G or W or L][F or W or G or N or C][F or H or W or L][R or E or I or L or A or P or G][N or R or P or C or T][W or F]IS[Y or D]Y (SEQ ID NO: 4).

[0078] 18. The peptide as described in embodiment 13, comprising the amino acid sequence XXXXXXFISDY (SEQ ID NO: 5), wherein X is any amino acid.

[0079] 19. The peptide as described in embodiment 18, comprising the amino acid sequence [D or W or S or I][S or G or W or L][F or W or G or N or C][F or H or W or L][R or E or I or L or A or P or G][N or R or P or C or T]FISDY (SEQ ID NO6).

[0080] 20. The peptide as described in embodiment 18, comprising the amino acid sequence [D or I][L or G][C or W][F or W][E or G][T or R]FISDY (SEQ ID NO: 7).

[0081] 21. The peptide as described in embodiment 18, comprising the amino acid sequence DGW[F or W][E or G][T or R]FISDY (SEQ ID NO: 8).

[0082] 22. A nucleic acid molecule encoding a polypeptide as described in any one of embodiments 1-12 or a peptide as described in any one of embodiments 13-21.

[0083] 23. A carrier comprising a nucleic acid molecule as described in embodiment 22.

[0084] 24. A host cell comprising a nucleic acid molecule as described in embodiment 22 or a vector as described in embodiment 23.

[0085] 25. A pharmaceutical composition comprising a polypeptide as described in any one of embodiments 1-12, a peptide as described in any one of embodiments 13-21, a nucleic acid molecule as described in embodiment 22, or a carrier as described in embodiment 23, and a pharmaceutically acceptable carrier, excipient, and / or stabilizer.

[0086] 26. The pharmaceutical composition as described in embodiment 25, further comprising a therapeutic agent.

[0087] 27. The polypeptide as described in any one of embodiments 1-12, the peptide as described in any one of embodiments 13-21, the nucleic acid molecule as described in embodiment 22, the carrier as described in embodiment 23, or the pharmaceutical composition as described in embodiment 25 or 26, for inhibiting GIPR signaling in cells.

[0088] 28. The polypeptide, peptide, nucleic acid molecule, carrier, or pharmaceutical composition as described in embodiment 27, wherein the cell is a pancreatic β cell, pancreatic α cell, pancreatic υ cell, adipocyte, pericyte, mesothelial cell, endothelial cell, osteocyte, osteoblast, osteoclast, cardiomyocyte, neuron, oligodendrocyte, gastric mucus secretory cell, ciliated cell, T cell, myeloid cell, immortalized cell expressing GIPR, or cancer cell expressing GIPR.

[0089] 29. A polypeptide, peptide, nucleic acid molecule, carrier, or pharmaceutical composition as described in embodiment 27 or 28, wherein the cells are in vitro.

[0090] 30. A polypeptide, peptide, nucleic acid molecule, carrier, or pharmaceutical composition as described in embodiment 27 or 28, wherein the cells are in the body of a subject.

[0091] 31. The polypeptide as described in any one of embodiments 1-12, the peptide as described in any one of embodiments 13-21, the nucleic acid molecule as described in embodiment 22, the carrier as described in embodiment 23, or the pharmaceutical composition as described in embodiment 25 or 26, for the treatment or prevention of a disease or condition in a subject related to GIPR signaling.

[0092] 32. The polypeptide, peptide, nucleic acid molecule, carrier, or pharmaceutical composition as described in embodiment 31, wherein the disease or condition is obesity, hypertension, cardiovascular disease, non-alcoholic fatty liver disease, Cushing's syndrome, Alzheimer's disease, Parkinson's disease, metabolic syndrome, or diabetes.

[0093] 33. A method for inhibiting GIPR signaling in cells, comprising contacting the cells with a polypeptide as described in any one of embodiments 1-12, a peptide as described in any one of embodiments 13-21, a nucleic acid molecule as described in embodiment 22, a carrier as described in embodiment 23, or a pharmaceutical composition as described in embodiment 25 or 26.

[0094] 34. The method as described in embodiment 33, wherein the cells are pancreatic β cells, pancreatic α cells, pancreatic υ cells, adipocytes, pericytes, mesothelial cells, endothelial cells, osteocytes, osteoblasts, osteoclasts, cardiomyocytes, neurons, oligodendrocytes, gastric mucus-secreting cells, ciliated cells, T cells, myeloid cells, immortalized cells expressing GIPR or cancer cells expressing GIPR, immortalized cells expressing GIPR or cancer cells expressing GIPR.

[0095] 35. The method as described in embodiment 33 or 34, wherein the cells are in vitro.

[0096] 36. The method as described in embodiment 33 or 34, wherein the cells are in the subject.

[0097] 37. A method for treating or preventing a disease or condition related to GIPR signaling in a subject, comprising administering to the subject a polypeptide as described in any one of embodiments 1-12, a peptide as described in any one of embodiments 13-21, a nucleic acid molecule as described in embodiment 22, a carrier as described in embodiment 23, or a pharmaceutical composition as described in embodiment 25 or 26.

[0098] 38. The method as described in embodiment 37, wherein the disease or condition is obesity, hypertension, cardiovascular disease, non-alcoholic fatty liver disease, Cushing's syndrome, Alzheimer's disease, Parkinson's disease, metabolic syndrome, or diabetes.

[0099] 39. Use of a polypeptide as described in any one of embodiments 1-12, a peptide as described in any one of embodiments 13-21, a nucleic acid molecule as described in embodiment 22, a carrier as described in embodiment 23, or a pharmaceutical composition as described in embodiment 25 or 26, for inhibiting GIPR signaling in cells.

[0100] 40. The use as described in embodiment 39, wherein the cells are pancreatic β cells, pancreatic α cells, pancreatic υ cells, adipocytes, pericytes, mesothelial cells, endothelial cells, osteocytes, osteoblasts, osteoclasts, cardiomyocytes, neurons, oligodendrocytes, gastric mucus-secreting cells, ciliated cells, T cells, myeloid cells, immortalized cells expressing GIPR, or cancer cells expressing GIPR.

[0101] 41. The use as described in embodiment 39 or 40, wherein the cells are in vitro.

[0102] 42. The method as described in embodiment 39 or 40, wherein the cells are in the subject.

[0103] 43. Use of a polypeptide as described in any one of embodiments 1-12, a peptide as described in any one of embodiments 13-21, a nucleic acid molecule as described in embodiment 22, a carrier as described in embodiment 23, or a pharmaceutical composition as described in embodiment 25 or 26, for the treatment or prevention of a disease or condition in a subject related to GIPR signaling.

[0104] 44. The method as described in embodiment 43, wherein the disease or condition is obesity, hypertension, cardiovascular disease, non-alcoholic fatty liver disease, Cushing's syndrome, Alzheimer's disease, Parkinson's disease, metabolic syndrome, or diabetes.

[0105] In understanding the scope of this disclosure, the term "comprising" and its derivatives as used herein are intended as open-ended terms to indicate the presence of said features, elements, components, groups, integers, and / or steps, but do not exclude the presence of other unstated features, elements, components, groups, integers, and / or steps. The foregoing also applies to words with similar meanings, such as the terms "comprising," "having," and their derivatives. The term "consisting of" and its derivatives are intended as closed-ended terms to indicate the presence of said features, elements, components, groups, integers, and / or steps, but exclude the presence of other unstated features, elements, components, groups, integers, and / or steps. The term "mainly composed of" as used herein is intended to indicate the presence of said features, elements, components, groups, integers, and / or steps, as well as those that do not materially affect the essential and novel features of the features, elements, components, groups, integers, and / or steps.

[0106] As used herein, the singular forms “a(a)”, “an”, and “the” include a plural of indicators unless the content explicitly indicates otherwise. In embodiments that include an “additional” or “second” component, the second component, as used herein, is different from the other components or the first component. A “third” component is different from the other components, the first component, and the second component, and is also different from the further listed components or “additional” components.

[0107] All publications and patents cited herein are incorporated by reference in their entirety, as if each individual publication or patent were specifically and individually cited. In the event of any conflict between the definitions of terms in this disclosure and those in the cited publications or patents, the definitions provided herein shall prevail in describing the invention.

[0108] The invention will now be described by way of non-limiting examples with reference to the accompanying drawings.

[0109] Example 1

[0110] Basically, it uses Gaertner et al. (2008). PNAS ,105(46):17706-17711) and Akondi et al. (2021, Chimia The techniques described in [Journal Name], 75(6):489-494, were used to prepare GIP variants having a variant N-terminal portion (residues 0-10) and a conserved GIP C-terminal portion (residues 11-42). The GIP variants were chemically synthesized, and their pharmacological activity was tested on CHO cells expressing GIPR.

[0111] Table 2 summarizes the determined activities of the GIP variants, showing their sequence and inhibitory activities. The GIP variants were synthesized using protein modification sites (residues 0 to 10), as indicated (encoded by single-letter amino acids). The antagonistic potency of the analogs was assessed using CHO-GIPR cells in a calcium flux assay. "Av ANTAG pIC50" shows the mean measured pIC50 of the inhibition of GIP-mediated GIPR agonist-induced calcium flux, averaged from 2–3 independent replicates. As shown in Table 2, the GIP variant peptides are approximately 100-fold more potent than previously reported truncated GIP variants.

[0112] Table 2

[0113] Example 2

[0114] Other GIP variants based on hGIP-1P25 (SEQ ID NO: 24) were generated, and their inhibitory activity against GIPR was tested using HEK cells expressing GIPR. pIC50 was determined as described in Example 1.

[0115] In Group 1 experiments (Table 3), a single-substituted variant of hGIP-1P25 was generated, and its inhibitory effect on GIPR was tested. The Group 1 variants were compared with those of Yang B et al. (…). Mol Metab.The reference antagonist GIP (2022 Dec;66:101638. doi: 10.1016 / j.molmet.2022.101638) is disclosed. (5-31) Palmitoylated analogues ([N α -Ac, L14, R18,E21] hGIP (5-31) -K11 (γE-C16)) was compared. The results are disclosed in Table 3. “OAc” refers to N-acetylation of the N-terminal residue. All positions and substitutions are indicated relative to hGIP-1P25 (SEQ ID NO: 24).

[0116] Table 3

[0117] In the second set of experiments (Table 4), a bisubstituted variant of hGIP-1P25 was generated, and its inhibitory effect on GIPR was tested. The second set of variants was compared with the reference antagonist GIP. (5-31) Palmitoylated analogues ([N α -Ac, L14, R18, E21]hGIP (5-31) -K11 (γE-C16)) was compared. The results are disclosed in Table 4. “0Ac” refers to N-acetylation of the N-terminal residue. All positions and substitutions are indicated relative to hGIP-1P25 (SEQ ID NO: 24).

[0118] Table 4

[0119] In Group 3 experiments (Table 5), truncated variants of hGIP-1P25 were generated, and their inhibitory effect on GIPR was tested. The Group 3 variants were truncated to include positions 1-30 of hGIP-1P25 and included further substitutions or modifications. The Group 3 variants were compared with the reference antagonist GIP. (5-31) Palmitoylated analogues ([N α -Ac, L14, R18, E21] hGIP (5-31) -K11 (γE-C16)) was compared. The results are disclosed in Table 5. “0Ac” refers to N-acetylation of the N-terminal residue. All positions and substitutions are indicated relative to hGIP-1P25 (SEQ ID NO: 24).

[0120] Table 5

[0121] For purposes of illustration and description, the foregoing description of specific embodiments of the invention has been provided. These descriptions are not intended to be exhaustive or to limit the invention to the precise forms disclosed. Many modifications and variations are possible based on the foregoing teachings. The described embodiments were chosen and described in order to best explain the principles of the invention and its practical application, thereby enabling others skilled in the art to best utilize the invention and its various embodiments, making various modifications to suit a particular intended use. It should be understood that, as appropriate, various omissions or substitutions with equivalents may be considered, but are intended to cover the application or implementation without departing from the spirit or scope of the claims of the invention.

Claims

1. A polypeptide comprising an N-terminal portion and a C-terminal portion, wherein the N-terminal portion comprises an amino acid sequence having substituted 0, 1, 2, 3, 4, 5, or 6 amino acids relative to any one of SEQ ID NO: 16, 9-15, 25-48, and 73-114, and wherein the C-terminal portion comprises an amino acid sequence that is at least 70% identical to any one of SEQ ID NO: 2 and 166-168.

2. The polypeptide of claim 1, wherein the N-terminal portion comprises the amino acid sequence of any one of SEQ ID NO: 16, 9-15, 25-48, and 73-114.

3. The polypeptide of claim 1, wherein the N-terminal portion comprises the amino acid sequence XXXXXXFISDY (SEQ ID NO: 5), wherein X is any amino acid.

4. The polypeptide of claim 3, wherein the N-terminal portion comprises the amino acid sequence DGW[F or W][E or G][T or R]FISDY (SEQ ID NO: 8).

5. The polypeptide of any one of claims 1 to 4, wherein the C-terminal portion comprises the amino acid sequence of any one of SEQ ID NO: 2 and 166-168.

6. A peptide comprising an amino acid sequence having 0, 1, 2, 3, 4, 5 or 6 amino acid substitutions relative to any one of SEQ ID NO: 16, 9-15, 25-48 and 73-114.

7. The peptide of claim 6, comprising the amino acid sequence of any one of SEQ ID NO: 16, 9-15, 25-48, and 73-114.

8. The peptide of claim 6, comprising the amino acid sequence XXXXXXFISDY (SEQ ID NO: 5), wherein X is any amino acid.

9. The peptide of claim 8, comprising the amino acid sequence DGW[F or W][E or G][T or R]FISDY (SEQ ID NO: 8).

10. A nucleic acid molecule encoding a polypeptide as described in any one of claims 1 to 5 or a peptide as described in any one of claims 6 to 9.

11. A vector comprising the nucleic acid molecule as described in claim 10.

12. A host cell comprising the nucleic acid molecule of claim 10 or the vector of claim 11.

13. A pharmaceutical composition comprising a polypeptide as described in any one of claims 1 to 5, a peptide as described in any one of claims 6 to 9, a nucleic acid molecule as described in claim 10, or a carrier as described in claim 11, and a pharmaceutically acceptable carrier, excipient, and / or stabilizer.

14. The pharmaceutical composition of claim 13, further comprising a therapeutic agent.

15. The polypeptide of any one of claims 1 to 5, the peptide of any one of claims 6 to 9, the nucleic acid molecule of claim 10, the carrier of claim 11, or the pharmaceutical composition of claim 13 or 14, for inhibiting GIPR signaling in cells.

16. The polypeptide, peptide, nucleic acid molecule, carrier, or pharmaceutical composition of claim 15, wherein the cell is a pancreatic β cell, pancreatic α cell, pancreatic υ cell, adipocyte, pericyte, mesothelial cell, endothelial cell, osteocyte, osteoblast, osteoclast, cardiomyocyte, neuron, oligodendrocyte, gastric mucus-secreting cell, ciliated cell, T cell, myeloid cell, immortalized cell expressing GIPR, or cancer cell expressing GIPR.

17. The polypeptide, peptide, nucleic acid molecule, carrier, or pharmaceutical composition of claim 15 or 16, wherein the cells are in vitro.

18. The polypeptide, peptide, nucleic acid molecule, carrier, or pharmaceutical composition of claim 15 or 16, wherein the cells are in the body of the subject.

19. The polypeptide of any one of claims 1 to 5, the peptide of any one of claims 6 to 9, the nucleic acid molecule of claim 10, the carrier of claim 11, or the pharmaceutical composition of claim 13 or 14, for the treatment or prevention of diseases or conditions related to GIPR signaling in a subject.

20. The polypeptide, peptide, nucleic acid molecule, carrier, or pharmaceutical composition of claim 19, wherein the disease or condition is obesity, hypertension, cardiovascular disease, non-alcoholic fatty liver disease, Cushing's syndrome, Alzheimer's disease, Parkinson's disease, metabolic syndrome, or diabetes.

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