WO2022206587A1 - 一种多肽化合物及其应用 - Google Patents

一种多肽化合物及其应用 Download PDF

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WO2022206587A1
WO2022206587A1 PCT/CN2022/082973 CN2022082973W WO2022206587A1 WO 2022206587 A1 WO2022206587 A1 WO 2022206587A1 CN 2022082973 W CN2022082973 W CN 2022082973W WO 2022206587 A1 WO2022206587 A1 WO 2022206587A1
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substituted
unsubstituted
group
hydrogen
independently selected
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PCT/CN2022/082973
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English (en)
French (fr)
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孟广鹏
付晓平
李四军
马浩宇
张银
刘爽
徐苗
高剑
李元波
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成都诺和晟泰生物科技有限公司
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Priority to JP2023550657A priority Critical patent/JP2024507884A/ja
Priority to KR1020237028176A priority patent/KR20230133889A/ko
Priority to EP22778770.2A priority patent/EP4321525A1/en
Publication of WO2022206587A1 publication Critical patent/WO2022206587A1/zh

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/04—Linear peptides containing only normal peptide links
    • C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/17—Amino acids, peptides or proteins
    • A23L33/18—Peptides; Protein hydrolysates
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P5/00—Drugs for disorders of the endocrine system
    • A61P5/06—Drugs for disorders of the endocrine system of the anterior pituitary hormones, e.g. TSH, ACTH, FSH, LH, PRL, GH
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/08—Tripeptides
    • C07K5/0802—Tripeptides with the first amino acid being neutral
    • C07K5/0804—Tripeptides with the first amino acid being neutral and aliphatic
    • C07K5/081—Tripeptides with the first amino acid being neutral and aliphatic the side chain containing O or S as heteroatoms, e.g. Cys, Ser
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/10—Tetrapeptides
    • C07K5/1002—Tetrapeptides with the first amino acid being neutral
    • C07K5/1005—Tetrapeptides with the first amino acid being neutral and aliphatic
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/10—Tetrapeptides
    • C07K5/1024—Tetrapeptides with the first amino acid being heterocyclic
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
    • Y02P20/00—Technologies relating to chemical industry
    • Y02P20/50—Improvements relating to the production of bulk chemicals
    • Y02P20/55—Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups

Definitions

  • the present invention relates to the technical field of biomedicine, in particular to a polypeptide compound and its application.
  • ghrelin Growth hormone releasing peptide
  • GHSR growth hormone secretagogue receptor
  • ghrelin was discovered by Kojima et al. in mouse and human gastric endocrine cells and arcuate nucleus of hypothalamus. It is the only natural ligand of GHSR discovered so far. It contains 28 amino acid residues and has a molecular weight of 3.3kDa.
  • the ghrelin precursor protein of human and rat is composed of 117 amino acids, and the first 23 peptides of the N-terminal are characteristic of secretion signal peptide; amino acid-arginine) as its recognition site.
  • Human and rat ghrelin differ in only 2 amino acid residues, and the coding gene sequences share 82.9% homology.
  • the third serine at the N-terminal of ghrelin is the substantial part of its biological function.
  • des-acyl ghrelin (des-acyl ghrelin) had no biological activity, but the latest study found that both des-octyl ghrelin and ghrelin can promote the proliferation of notochord neuroepithelial; It has endocrine function, can promote cell proliferation, and has anti-apoptotic effect.
  • GHSR is an orphan G protein-coupled receptor, which is widely distributed in peripheral tissues, brain, intestine, kidney, pancreas, heart, adipose tissue, etc. .
  • the extensive distribution of GHSR plays an important role in various biological functions of ghrelin and its receptors.
  • the GHSR structure-encoding genome is highly conserved in different species, and its amino acid sequence shares 52% homology with the G protein-coupled protein receptor of motilin gene-related peptide.
  • GHSR is divided into 1a type and 1b type according to different exon coding, among which GHSR-1a is a functional receptor of ghrelin, which activates phospholipase C (PLC), inositol triphosphate (IP3) after binding to ghrelin , protein kinase C (PKC) and other biological effects.
  • PLC phospholipase C
  • IP3 inositol triphosphate
  • PLC protein kinase C
  • the non-functional receptor GHSR-1b has no biological activity.
  • the pulsatile release of growth hormone (GH) in the pituitary is mainly regulated by three factors, growth hormone releasing hormone (GHRH), somatostatin (SS) and ghrelin in the hypothalamus.
  • GHRH growth hormone releasing hormone
  • SS somatostatin
  • ghrelin can cooperate with GHRH to play a promoting role, and the three form a local neuroendocrine regulation feedback loop in the hypothalamus.
  • GHRH binds to its receptors and increases the level of intracellular cyclic adenosine monophosphate
  • ghrelin binds to its receptors, depolarizes and inhibits K + channels, causes an increase in intracellular IP3 concentration, and intracellular
  • the increase of Ca 2+ concentration eventually stimulates the secretion of GH.
  • the release of GH from pituitary somatotropin cells can also be controlled by growth hormone-releasing polypeptide (GHRP).
  • GHRP growth hormone-releasing polypeptide
  • GHRP- 6 H-His-D-Trp-Ala-Trp-D-Phe-Lys-CONH2
  • Such peptides and peptoid compounds can combine with GHSR-1a to generate agonistic activity, cause signal transduction, and thus regulate GH secretion, but they all have certain limitations in clinical development. Therefore, the development of structures with high activity, small doses, and low toxic side effects is a research goal of GHSR-1a receptor agonists.
  • the technical problem to be solved by the present invention is to provide a polypeptide compound and its application.
  • the prepared polypeptide compound has high activity as a GHSR-1a receptor agonist.
  • the present invention provides a polypeptide compound having the structure shown in formula I, or a stereoisomer, mixture or pharmaceutically acceptable salt thereof:
  • R 1 is selected from -NR 2 R 3 , -OR 2 or -SR 2 ;
  • R 1 is not a D- or L-amino acid
  • R2 and R3 are independently selected from hydrogen , deuterium, polymers derived from polyethylene glycol, acyclic substituted or unsubstituted aliphatic groups, substituted or unsubstituted alicyclic groups, substituted substituted or unsubstituted heterocyclyl, substituted or unsubstituted heteroarylalkyl, substituted or unsubstituted aryl, substituted or unsubstituted aralkyl.
  • W is selected from a single bond, a D-amino acid or an L-amino acid
  • U 1 is selected from any of the following structures:
  • R4 , R7 and R8 are independently selected from hydrogen, deuterium, amino, protecting groups, polymers derived from polyethylene glycol, acyclic substituted or unsubstituted aliphatic groups, substituted or unsubstituted substituted alicyclic, substituted or unsubstituted heterocyclyl, substituted or unsubstituted heteroarylalkyl, substituted or unsubstituted aryl, substituted or unsubstituted aralkane base or R 9 CO-;
  • Y is selected from halogen, amino, nitro, hydroxyl or cyano
  • R 5 is selected from -NR 2 R 3 , -OR 2 or -SR 2 ;
  • R 6 is selected from hydrogen, deuterium, acyclic substituted or unsubstituted aliphatic group, substituted or unsubstituted alicyclic group, substituted or unsubstituted heterocyclic group, substituted or unsubstituted substituted heteroarylalkyl, substituted or unsubstituted aryl, substituted or unsubstituted aralkyl;
  • n 1 and m 2 are independently selected from 0, 1, 2 or 3; in particular, when X is N, m 1 is 2, and m 2 is independently selected from 0, 1, 3; m 2 is 2, m 1 is independently selected from 0, 1, 3;
  • n3 and m4 are independently selected from 0, 1, 2 or 3;
  • n 1 , n 2 , n 3 and n 4 are independently selected from 0, 1, 2 or 3;
  • p 0, 1, 2, 3, 4 or 5;
  • U 2 is a single bond, or is selected from any of the following structures, and the carbonyl end of U 2 is connected to W:
  • R 9 is selected from hydrogen, acyclic substituted or unsubstituted aliphatic group, substituted or unsubstituted alicyclic group, substituted or unsubstituted heterocyclic group, substituted or unsubstituted Heteroarylalkyl, substituted or unsubstituted aryl, substituted or unsubstituted aralkyl.
  • the R 1 is selected from -NR 2 R 3 , -OR 2 or SR 2 .
  • R and R are independently selected from hydrogen , deuterium, polymers derived from polyethylene glycol, acyclic substituted or unsubstituted aliphatic groups, substituted or unsubstituted alicyclic groups , substituted or unsubstituted heterocyclyl, substituted or unsubstituted heteroarylalkyl, substituted or unsubstituted aryl, substituted or unsubstituted aralkyl.
  • the R 1 is selected from -NR 2 R 3 or -OR 2 , wherein R 2 and R 3 are independently selected from hydrogen, methyl, ethyl, hexyl, dodecyl or hexadecyl .
  • the R 1 is not a D-type or L-type amino acid.
  • amino acids refer to amino acid residues, specifically, the residues after amino acids react with amino groups or carboxyl groups to form polypeptides.
  • the W is independently selected from a single bond, a D-type amino acid or an L-type amino acid. More preferably, the W is selected from single bond, alanine, arginine, asparagine, cysteine, glutamine, aspartic acid, glutamic acid, glycine, histidine, isoleucine One or more of amino acid, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, valine residues kind.
  • the amino acid removes a molecule of water, and forms an amide bond with the adjacent group.
  • the residue refers to to form an amide bond by removing a molecule of water and adjacent groups, thereby forming a polypeptide compound.
  • the U 1 is selected from any of the following structures:
  • R4 , R7 and R8 are independently selected from hydrogen, deuterium, amino, protecting groups, polymers derived from polyethylene glycol, acyclic substituted or unsubstituted aliphatic groups, substituted or unsubstituted substituted alicyclic, substituted or unsubstituted heterocyclyl, substituted or unsubstituted heteroarylalkyl, substituted or unsubstituted aryl, substituted or unsubstituted aralkane or R 9 CO-; more preferably hydrogen, deuterium, amino, polymers derived from polyethylene glycol, acyclic substituted or unsubstituted C 1-10 aliphatic groups, substituted or unsubstituted Substituted C 3-10 alicyclic, substituted or unsubstituted C 2-10 heterocyclyl, substituted or unsubstituted C 2-20 heteroarylalkyl, substituted or unsubstituted C 6-12 aryl, substituted or unsub
  • R 9 is preferably hydrogen, acyclic substituted or unsubstituted aliphatic group, substituted or unsubstituted alicyclic group, substituted or unsubstituted heterocyclic group, substituted or unsubstituted Substituted heteroarylalkyl, substituted or unsubstituted aryl, substituted or unsubstituted aralkyl; more preferably hydrogen, acyclic substituted or unsubstituted C 1-10 aliphatic group, substituted or unsubstituted C3-10 alicyclic group, substituted or unsubstituted C2-10 heterocyclyl group, substituted or unsubstituted C2-20 heteroarylalkyl group, Substituted or unsubstituted C 6-12 aryl, substituted or unsubstituted C 6-12 aralkyl. It is further preferably hydrogen or C 1-6 alkyl; in some specific embodiments of the present invention, the R 9 is specifically
  • the Y is selected from halogen, amino, nitro, hydroxyl or cyano. More preferred is F, Cl, Br or amino.
  • the R 5 is independently selected from -NR 2 R 3 , -OR 2 or -SR 2 ; more preferably -NR 2 R 3 .
  • R 2 and R 3 are independently selected from hydrogen, methyl, ethyl or hexyl.
  • the R 6 is independently selected from hydrogen, deuterium, acyclic substituted or unsubstituted aliphatic group, substituted or unsubstituted alicyclic group, substituted or unsubstituted heterocyclic group Cyclic, substituted or unsubstituted heteroarylalkyl, substituted or unsubstituted aryl, substituted or unsubstituted aralkyl; more preferably hydrogen, deuterium, acyclic substituted or Unsubstituted C 1-10 aliphatic group, substituted or unsubstituted C 3-10 alicyclic group, substituted or unsubstituted C 2-10 heterocyclic group, substituted or unsubstituted C 2-20 heteroarylalkyl, substituted or unsubstituted C 6-20 aryl, substituted or unsubstituted C 6-12 aralkyl. Further preferred is hydrogen, C 1-6 alkyl, C 6-14 aryl or C 3-8
  • the U 1 is selected from the following structures:
  • R 6 is selected from hydrogen, substituted or unsubstituted C 1-6 alkyl, substituted or unsubstituted C 6-14 aryl, substituted or unsubstituted C 3-8 cycloalkyl; the substituted group
  • the group is preferably halogen, amino, nitro, hydroxy, acyl-substituted amino, ureido or guanidino.
  • the R 6 is selected from hydrogen, substituted or unsubstituted methyl, ethyl, propyl, isopropyl, butyl, isobutyl or tert-butyl.
  • the substituted group is selected from halogen, amino, nitro, hydroxyl, carboxamido, acetamido, propionamido, butanamido, ureido or guanidino.
  • R 7 and R 8 are independently preferably hydrogen, C 1-6 alkyl, C 6-14 aryl, C 3-8 cycloalkyl, C 2-10 acyl; more preferably hydrogen, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl, formyl, acetyl, propionyl or butyryl.
  • the U 1 is selected from any of the following structures:
  • R 10 and R 11 are independently preferably hydrogen, amino, nitro, hydroxyl, halogen, cyano, aminomethyl, aminoethyl, aminopropyl or aminobutyl.
  • the U 2 is a single bond, or is selected from any of the following structures (ie 4-amino-4-piperidinecarboxylic acid (Apc) or lysine (Lys), ornithine (Orn), sperm amino acid (Arg) residue), and the carbonyl terminus of U is attached to W:
  • m 1 and m 2 are independently selected from 0, 1, 2 or 3; in particular, when X is N, m 1 is 2, and m 2 is independently selected from 0, 1, 3; m 2 is 2, and m 1 is independently selected from 0, 1, and 3; that is, the structures containing m 1 and m 2 are not selected from pyridine groups.
  • n 3 and m 4 are independently selected from 0, 1, 2 or 3.
  • n 1 , n 2 , n 3 and n 4 are independently selected from 0, 1, 2 or 3.
  • p is 0, 1, 2, 3, 4 or 5.
  • the polypeptide compound has any of the following structures, or a stereoisomer, mixture or pharmaceutically acceptable salt thereof:
  • polypeptide compounds as described herein, their stereoisomers, mixtures thereof, their pharmaceutically acceptable salts can be carried out according to any conventional method known in the art, such as Using solid-phase peptide synthesis methods [Stewart J.M.y Young J.D., “Solid Phase Peptide Synthesis, 2nd edition", (1984), Pierce Chemical Company, Rockford, Illinois; Bodanzsky M.y Bodanzsky A., "The practice of Peptide Synthesis", (1994 ), Springer Verlag, Berlin; Lloyd Williams P.
  • methods of obtaining the polypeptide compounds of the present invention, their stereoisomers and mixtures thereof may comprise the following stages:
  • the C-terminus is bound to a solid support and the process is carried out in solid phase, thus comprising combining N-terminal protected and C-terminal free amino acids with N-terminal free and C-terminally bound to the polymer support Amino acid coupling; elimination of the group protecting the N-terminus; and repeating this procedure as many times as necessary to obtain compounds of the desired length, ultimately followed by cleavage of synthetic compounds from the original polymeric support.
  • solid phase synthesis can be performed with a convergent strategy: coupling the peptide to a polymeric support or to a peptide or amino acid pre-bound to the polymeric support.
  • Convergent synthesis strategies are widely known to those skilled in the art and are described in Lloyd-Williams P. et al., "Convergent Solid-Phase Peptide Synthesis", (1993), Tetrahedron, 49(48), 11065-11133.
  • the process of the present invention can include additional stages of C-terminal deprotection and/or cleavage of the peptide from the polymeric carrier in an indiscriminate sequence; after these terminal stages Functional groups can be modified.
  • Optional modification of the C-terminus can be performed when the polypeptide compound of formula (I) is immobilized to the polymer support or once the polypeptide compound has been separated from the polymer support.
  • the R residue can be introduced in a suitable solvent and base such as N,N-diisopropylethylamine (DIEA) or triethylamine or additives such as 1-hydroxybenzotriazole Compound HR 1 , wherein R 1 is -OR 2 , -NR2R3 or -SR2 , reacted with a complementary fragment corresponding to a compound of formula (I), wherein R1 is -NH2 ; or by first reacting the complementary fragment corresponding to a compound of formula (I) with, for example, sulfite The acid chloride preforms the acid halide, which is then reacted with HR 1 , thereby obtaining a peptide according to the invention of formula (I), wherein the fragment has functional groups that are not involved in the formation of the NC bond and are suitably protected with temporary or permanent protective groups ; or alternatively other R1 residues can be introduced by a process of simultaneous cleavage of the incorporated peptide from the polymeric
  • DIEA N
  • the present invention provides a composition comprising the above-mentioned polypeptide compound, and an acceptable adjuvant.
  • the above-mentioned composition may be a pharmaceutical composition or a health care product composition.
  • the adjuvants include, but are not limited to, carriers, diluents, excipients or adjuvants that are well known to those skilled in the art.
  • the carrier includes, but is not limited to, sterile water, saline, buffer, phosphate buffered saline, buffered sodium chloride, vegetable salts, minimal essential medium (MEM), MEM with HEPES, and the like.
  • the polypeptide compounds may exist alone, or in a mixture of two or more, or be more closely associated by complexation, crystallization, or ionic or covalent bonding.
  • the size of the rigid structure or flexible structure in the amino acid residue at the C-terminus of the polypeptide compound provided by the present invention is very important to maintain the configuration of the peptide bond in the sequence. Furthermore, the present invention introduces different structural types of functional groups at the C-terminus of the sequence, so that the The polypeptide compound can effectively bind to GHSR-1a and is suitable for treating, preventing, alleviating or diagnosing related diseases caused by disorders mediated by GHSR-1a.
  • the present invention provides the application of the above-mentioned polypeptide compound, or the above-mentioned preparation method of the polypeptide compound, or the above-mentioned composition, as an agonist of growth hormone secretagogue receptor, or in preparation for treatment, prevention, alleviation and / or medicines for diagnosing related diseases caused by disorders mediated by growth hormone secretagogue receptors, or application in health care products for promoting growth and development.
  • the growth hormone secretagogue receptor may also be referred to as a ghrelin receptor, a ghrelin receptor or a GHSR-1a receptor.
  • the related disease caused by the disorder mediated by the growth hormone secretagogue receptor is growth hormone deficiency.
  • the present invention provides the use of the above-mentioned polypeptide compound, or the polypeptide compound prepared by the above-mentioned preparation method, or the above-mentioned composition as a GHSR-1a agonist.
  • the present invention provides the use of the above-mentioned polypeptide compound, or the polypeptide compound prepared by the above-mentioned preparation method, or the above-mentioned composition in the preparation of a GHSR-1a agonist.
  • polypeptide compounds, compositions, or agonists of growth hormone secretagogue receptors can be administered in a variety of ways, depending on whether topical or systemic administration is desired and on the area to be treated.
  • the polypeptide compound, or composition thereof, or a GHSR-1a agonist thereof can be administered to a patient orally or rectally, or transmucosally, or enterally, or intramuscularly, or subcutaneously, or medullarily Intra, or intrathecal, or directly intraventricular, or intravenous, or intravitreal, or intraperitoneal, or intranasal, or intraocular.
  • protecting group refers to a group that blocks an organic functional group and can be removed under controlled conditions.
  • Protecting groups, their relative reactivity and the conditions under which they remain inert are known to those skilled in the art.
  • Examples of representative protective groups for amino groups are especially amidoacetate, amidobenzoic acid, amidopivalate; carbamates such as benzyloxycarbonyl (Cbz or Z), 2-chlorobenzyl ( CIZ), p-nitrobenzyloxycarbonyl (pNZ), tert-butoxycarbonyl (Boc), 2,2,2-trichloroethoxycarbonyl (Troc), 2-(trimethylsilyl)ethyl yloxycarbonyl (Teoc), 9-fluorenylmethyloxycarbonyl (Fmoc) or allyloxycarbonyl (Alloc), trityl (Trt), methoxytrityl (Mtt), 2,4-Dinitrophenyl (Dnp), N-1-(4,4-dimethyl-2,6-dioxocyclohex-1-ylidene)ethyl (Dde), 1-( 4,4-Dimethyl-2,6-dioxo-
  • esters such as tert-butyl ester (tBu), allyl ester (All), triphenylmethyl ester (Trt ester), cyclohexyl ester (cHx), benzyl Base ester (Bzl), o-nitrobenzyl ester, p-nitrobenzyl ester, p-methoxybenzyl ester, trimethylsilylethyl ester, 2-phenylisopropyl ester, fluorenyl methyl ester base ester (Fm), 4-(N-[1-(4,4-dimethyl-2,6-dioxo-cyclohexylene)-3-methylbutyl]amino)benzyl ester (Dmab ), preferably All, tBu, cHx, Bzl and Trt esters.
  • esters such as tert-butyl ester (tBu), allyl ester (All), triphenylmethyl ester (Trt ester),
  • the side chains of trifunctional amino acids can be protected with temporary or permanent protecting groups orthogonal to the N-terminal and C-terminal protecting groups during the synthetic process.
  • the indole group of the tryptophan side chain can be protected by a formyl group (For), Boc, Mts or can be used unprotected.
  • the piperidine group of the 4-amino-4-piperidinecarboxylic acid side chain is protected by Boc or Fmoc.
  • Arginine side chains can be protected by the following protecting groups: Tos, 4-methoxy-2,3,6-trimethylbenzenesulfonyl (Mtr), Alloc, Nitro, 2,2,4,6 , 7-Pentamethyldihydrobenzofuran-5-sulfonyl (Pbf) and 2,2,5,7,8-pentamethylchroman-6-sulfonyl (Pmc).
  • Amides such as acetate amide, benzoic acid amide, pivalic acid amide can be used to protect the amino groups of the lysine and ornithine side chains; carbamates such as Cbz or Z, CIZ, pNZ, Boc, Troc, Teoc , Fmoc or Alloc, Trt, Mtt, Dnp, Dde, ivDde, Adpoc, etc.
  • the protecting group strategy used is the following strategy: amino group is protected by Boc, carboxyl group is protected by Bzl, cHx or All, arginine side chain is protected by Tos, 4-amino-4 -
  • the piperidine group of the pipecolic acid side chain is protected by Fmoc
  • the tryptophan side chain is protected by For or Mts
  • the Apc lysine and ornithine side chains are protected by CIZ, Fmoc or Alloc.
  • the protecting group strategy used is the following strategy: amino group is protected by Fmoc, carboxyl group is protected by tBu, All or Trt ester, arginine side chain is protected by Pmc or Pbf , the piperidine group of the 4-amino-4-piperidinecarboxylic acid side chain is protected by Boc, the tryptophan side chain is protected or used unprotected by Boc, and the lysine and ornithine side chains are protected by Boc, Trt or Alloc protection.
  • protective group also includes polymeric supports in solid phase synthesis.
  • solid supports for use in the process of the present invention involve polystyrene supports, polyethylene glycol grafted to polystyrene, etc., such as and not limited to p-methyl Diphenylmethyl eucalyptus (MBHA) [Matsueda GR et al., "A p-methyl benzhydrylamine resin for improved solid-phase synthesis of peptide amides", (1981), Peptides, 2, 4550], 2-chlorotrityl Base resins [Barlos K.
  • MBHA p-methyl Diphenylmethyl eucalyptus
  • Trp Tryptophan
  • polypeptide As defined herein, the terms “polypeptide”, “peptide” and “amino acid sequence” are used interchangeably herein to refer to a polymer of amino acid residues of any length.
  • the polymer may be linear or branched, it may contain modified amino acids or amino acid analogs, and it may be interrupted by chemical moieties other than amino acids.
  • the term also includes amino acid polymerisations that have been modified naturally or artificially (eg disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation or any other manipulation or modification, eg conjugation to a label or biologically active component) thing.
  • peptide includes two or more naturally occurring or synthetic amino acids linked by covalent bonds (eg, amide bonds).
  • amino acid is defined as having at least one primary, secondary, tertiary or quaternary amino group and at least one acid group, wherein the acid group may be a carboxylic acid, sulfonic acid or phosphoric acid or a mixture thereof.
  • Amino groups can be "alpha”, “beta”, “gamma” to “omega” relative to the acid group.
  • Suitable amino acids include, but are not limited to, the 20 common naturally occurring amino acids found in peptides (eg, alanine, arginine, asparagine, cysteine, glutamine, aspartic acid, glutamic acid) , glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, valine D- and L-isomers of acid) and naturally occurring and non-naturally occurring amino acids prepared by organic synthesis or other metabolic pathways.
  • peptides eg, alanine, arginine, asparagine, cysteine, glutamine, aspartic acid, glutamic acid
  • glycine histidine
  • isoleucine leucine
  • lysine methionine
  • phenylalanine proline
  • serine threonine
  • tryptophan ty
  • the "backbone of an amino acid” may be substituted with one or more groups selected from halogen, hydroxy, guanidino, heterocyclic groups.
  • amino acid also includes within its scope glycine, alanine, valine, leucine, isoleucine, norleucine, methionine, proline, phenylalanine, tryptophan Amino acid, serine, threonine, cysteine, tyrosine, asparagine, glutamine, aspartic acid, glutamic acid, lysine, histidine, homocysteine, bovine Sulfonic acid, betaine, N-methylalanine, etc. Amino acids in (L) and (D) forms are included.
  • amino acid side chain refers to the moiety attached to the alpha-carbon of an amino acid.
  • amino acid side chain of alanine is methyl
  • amino acid side chain of phenylalanine is phenylmethyl
  • amino acid side chain of cysteine is thiomethyl
  • amino acid side chain of aspartic acid is Carboxymethyl
  • amino acid side chain of tyrosine is 4-hydroxybenzyl, and so on.
  • other non-naturally occurring amino acid side chains such as naturally occurring (eg, amino acid metabolites) or synthetically produced (eg, alpha-substituted amino acids).
  • acyclic aliphatic group encompasses linear or branched alkyl, alkenyl and alkynyl groups.
  • alkyl refers to a linear or branched saturated group having 1 to 24, preferably 1 to 16, more preferably 1 to 14, even more preferably 1 to 12, still more preferably 1, 2, 3, 4 , 5 or 6 carbon atoms and by a simple bond to the rest of the molecule including, for example and without limitation, methyl, ethyl, isopropyl, isobutyl, tert-butyl, heptyl, octyl, decyl, Dodecyl, lauryl, hexadecyl, octadecyl, pentyl, 2-ethylhexyl, 2-methylbutyl, 5-methylhexyl, etc.
  • alkynyl group refers to a linear or branched group having 2 to 24, preferably 2 to 16, more preferably 2 to 14, even more preferably 2 to 12, still more preferably 2, 3, 4, 5 or 6 carbon atoms, with one or more carbon-carbon triple bonds, preferably with 1, 2 or 3 carbon-carbon triple bonds, the bonds are conjugated or unconjugated, which are bonded to the rest of the molecule by simple bonding moieties, including, for example and without limitation, ethynyl groups, 1-propynyl, 2-propynyl, 1-butyl, 2-butyl, 3-butyl, pentyl, such as 1-pentyl, and the like.
  • Alkynyl groups can also contain one or more carbon-carbon double bonds, including, for example and without limitation, the groups but-1-en-3-ynyl, pent-4-en-1-ynyl, and the like.
  • alicyclic group is used in the present invention to encompass groups such as and not limited to cycloalkyl or cycloalkenyl or cycloalkynyl groups.
  • cycloalkyl refers to a saturated monocyclic or polycyclic aliphatic group having 3 to 24, preferably 3 to 16, more preferably 3 to 14, even more preferably 3 to 12, still more preferably 3, 4 , 5 or 6 carbon atoms and is bound to the rest of the molecule by a simple bond including, for example and without limitation, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, methylcyclohexyl, dimethyl Cyclohexyl, octahydroindene, decalin, dodecahydrophenarene, etc.
  • cycloalkenyl refers to a non-aromatic monocyclic or polycyclic aliphatic group having 5 to 24, preferably 5 to 16, more preferably 5 to 14, even more preferably 5 to 12, still more preferably 5 to 6 carbon atoms, with one or more carbon-carbon double bonds, preferably 1, 2 or 3 carbon-carbon double bonds, conjugated or unconjugated, by simple bonding to the rest of the molecule, including For example and without limitation, the cyclopent-1-en-1-yl group and the like.
  • cycloalkynyl refers to a non-aromatic monocyclic or polycyclic aliphatic group having 8 to 24, preferably 8 to 16, more preferably 8 to 14, even more preferably 8 to 12, still more preferably 8 or 9 carbon atoms with one or more carbon-carbon triple bonds, preferably 1, 2 or 3 carbon-carbon triple bonds, conjugated or unconjugated, which are bonded to the rest of the molecule by simple bonding, including For example and without limitation, the cyclooct-2-yn-1-yl group and the like. Cycloalkynyl groups can also contain one or more carbon-carbon double bonds, including, for example and without limitation, cyclooct-4-en-2-ynyl groups and the like.
  • aryl group refers to an aromatic group having 6 to 30, preferably 6 to 18, more preferably 6 to 10, still more preferably 6 or 10 carbon atoms, comprising 1, 2, 3 or 4 carbon atoms Aromatic rings, bonded or fused by carbon-carbon bonds, including, for example and without limitation, phenyl, naphthyl, diphenyl, indenyl, phenanthryl, or anthracenyl, etc.; or aralkyl groups.
  • aralkyl refers to an alkyl group substituted with an aromatic group, having 7 to 24 carbon atoms and including, for example and without limitation, -( CH2 ) 1-6 -phenyl, -( CH2 ) 1- 6- (1-naphthyl), -( CH2 ) 1-6- (2-naphthyl), -( CH2 ) 1-6 -CH(phenyl) 2 and the like.
  • heterocyclyl group refers to a 3-10 membered alkylated ring wherein one or more, preferably 1, 2 or 3 of the atoms in the ring are elements other than carbon such as nitrogen, oxygen or sulfur, and can be saturated or unsaturated.
  • heterocycles can be monocyclic, bicyclic, or tricyclic systems, which may include fused ring systems; and nitrogen, carbon, or sulfur atoms in residue heterocycles may be optionally oxidized; nitrogen atoms may be optionally quaternized; and the residue heterocyclyl may be partially or fully saturated or aromatic.
  • the term heterocyclyl most preferably refers to a 5 or 6 membered ring.
  • saturated heterocyclyl groups are dioxane, piperidine, piperazine, pyrrolidine, morpholine and thiomorpholine.
  • aromatic heterocyclyl groups also called heteroaromatic groups, are pyridine, pyrrole, furan, thiophene, benzofuran, imidazoline, hydroquinone, quinoline and naphthyridine.
  • heteroarylalkyl group refers to an alkyl group substituted with a substituted or unsubstituted aromatic heterocyclyl group, the alkyl group having 1 to 6 carbon atoms and the aromatic heterocyclyl group having 2 to 24 carbon atoms and 1 to 3 non-carbon atoms, and include, for example and without limitation, -( CH2 ) 1-6 -imidazolyl, -( CH2 ) 1-6 -triazolyl, -(CH 2 ) 1-6 -thienyl, -(CH 2 ) 1-6 -furanyl, -(CH 2 ) 1-6 -pyrrolidinyl and the like.
  • halogen or variations such as “halide” or “halo” refers to fluorine, chlorine, bromine and iodine.
  • heteroatom or variants such as “hetero-” as used herein refers to O, N, NH and S.
  • alkoxy refers to a straight or branched chain alkoxy group. Examples include methoxy, ethoxy, n-propoxy, isopropoxy, tert-butoxy, and the like.
  • amino refers to a group of the form -NR a R b , wherein R a and R b are independently selected from groups including but not limited to hydrogen, optionally substituted alkyl, optionally substituted alkenyl, The group of optionally substituted alkynyl and optionally substituted aryl.
  • substituents include, but are not limited to, the following groups that result in the formation of stabilizing moieties: aliphatic, alkyl, alkenyl, alkynyl, heteroaliphatic, heterocyclyl, aryl, heteroaryl, acyl, oxo , imino, thiocarbonyl, cyano, isocyano, amino, azido, nitro, hydroxyl, thiol and halo and any combination thereof, including but not limited to the following groups: aliphatic amino, hetero Aliphatic amino, alkylamino, heteroalkylamino, arylamino, heteroarylamino, alkylaryl, arylalkyl, aliphaticoxy, heteroaliphaticoxy, alkoxy, heteroalkoxy group, aryloxy, heteroaryloxy, aliphatic thio, heteroaliphatic thio, alkylthio, heteroalkylthio, arylthio, heteroarylthio,
  • a heteroatom such as nitrogen may have a hydrogen substituent and/or any suitable substituent as described herein that satisfies the valence of the heteroatom and results in the formation of a stabilizing moiety.
  • Compounds may contain one or more asymmetric centers and thus exist as racemates and racemic mixtures, single enantiomers, individual diastereomers and mixtures of diastereomers. All such isomeric forms of these compounds are expressly included herein.
  • Compounds may also be represented in a variety of tautomeric forms, in which case all tautomeric forms of the compounds described herein are expressly included herein (eg, alkylation of ring systems can result in multiple sites alkylation, all such reaction products are expressly included herein). All such isomeric forms of this compound are expressly included herein. All crystalline forms of the compounds described herein are expressly included herein.
  • the compounds of the present invention can exist as stereoisomers or mixtures of stereoisomers; for example, the amino acids that constitute them can independently of one another have the configuration L-, D- or racemic.
  • isomeric mixtures as well as racemic or diastereomeric mixtures, or pure diastereomers or enantiomers, depending on the number of asymmetric carbons and the isomer or isomeric mixture the presence of asymmetric carbons.
  • U2 can be -Lys-
  • U2 is selected from -L - Lys-, -D-Lys- or a mixture of both, racemic or non-racemic.
  • the preparation procedures described in this document allow those skilled in the art to obtain the respective stereoisomers of the compounds of the invention by selecting the amino acid of the correct configuration.
  • Pharmaceutically acceptable salts of the peptides of the present invention are also within the field of the present invention.
  • pharmaceutically acceptable salts means salts whose use in animals and more particularly in humans is recognized, and includes salts used to form base addition salts, whether they are inorganic or organic, Inorganic salts such as and not limited to lithium, sodium, potassium, calcium, magnesium, manganese, copper, zinc or aluminum, etc., or organic salts such as and not limited to ethylamine, diethylamine, ethanolamine, diethanolamine, arginine, lysine acid, histidine or piperazine, etc.; or acid addition salts, whether they are organic or inorganic, organic salts such as but not limited to acetate, citrate, lactate, malonate, maleate acid salt, tartrate, fumarate, benzoic acid, aspartate, glutamate, succinate, oleate, trifluoroacetate, oxalate, pamoate or gluconate Sugar salt
  • salt is not critical, provided that it is cosmetically or pharmaceutically acceptable.
  • Pharmaceutically acceptable salts of the peptides of the present invention can be obtained by conventional methods well known in the art (Berge S.M. et al., "Pharmaceutical Salts", (1977), J. Pharm. Sci., 66, 119, which is incorporated by reference in its entirety into this article).
  • the present invention provides a polypeptide compound having the structure shown in formula I, or a stereoisomer, mixture or pharmaceutically acceptable salt thereof.
  • the experimental results show that the polypeptide compounds provided by the present invention can effectively exhibit high agonistic activity to GHSR-1a.
  • Peptide synthesis was performed using standard Fmoc solid phase methods. Rink Amide resin is used, and the peptide chain is extended from the C-terminus to the N-terminus.
  • Protected amino acids include: Fmoc-Apc(Boc)-OH, Fmoc-D-Lys(Boc)-OH, Fmoc-D-Orn(Boc)-OH, Fmoc-Phe-OH, Fmoc-D-Trp(Boc)- OH, Fmoc-D-Bal-OH, Fmoc-D-Cit-OH, Fmoc-D-Arg(Pbf)-OH, Boc-D-Aba-OH, Fmoc-Lys(Boc)-OH, Fmoc-Lys( Alloc)-OH, Fmoc-Gly-OH, Fmoc-D-Ala-OH, Fmoc-D-Val-OH, Fmoc-D
  • the condensing agent is HBTU/HOBt/DIEA.
  • the deprotection reagent was piperidine/DMF solution. Crude peptides were dissolved in water and lyophilized for storage. It is separated and purified by medium pressure liquid chromatography or high performance liquid chromatography (HPLC), and the content of pure peptide is more than 90%.
  • Matrix-assisted laser desorption time-of-flight mass spectrometry MALDI-TOF-MS was used to determine the molecular weight of peptide sequences.
  • Deprotection reagent 20% v/v piperidine in DMF.
  • Deprotection Weigh 0.23 g (0.1 mmol) of Rink Amide resin and place it in a peptide synthesis reactor, then prepare the deprotection reagent according to the above concentration, add it to the resin, react at room temperature, drain it, and then again Piperidine/DMF was added, and after reaction at room temperature, it was drained and washed with DMF until the test was qualified.
  • Condensation reaction Add amino acid and condensing agent to DMF for activation under ice bath conditions, then add activation base to react to obtain activation solution, and finally add activation solution to resin. After reaction at room temperature, use 5% ninhydrin.
  • the color-developing reagent makes the resin develop color, the resin discolors, and the solvent is drained and washed with DMF. After the test is qualified, the solvent is drained, and the condensation reaction is complete at this time.
  • TFA needs to be cooled in an ice bath for 30 minutes or stored in a refrigerator before use; add the prepared lysis solution to the peptide resin under ice bath conditions, stir electromagnetically, the resin turns black, react under ice bath conditions for 30 minutes, and then remove Ice bath, continue to stir the reaction for 180 min at room temperature, the reaction is complete, add 200 ml of glacial ether under vigorous stirring, white crystals are precipitated, and continue to stir for 30 min; filter out the precipitate with a G4 sand core funnel, repeatedly wash with cold ether for 3 times, and air dry. Add 50 ml of double-distilled water and 5 ml of acetonitrile to fully dissolve the solid, suction filtration, and freeze-dried the filtrate to obtain 1.04 g of crude peptide.
  • the crude peptides were purified by medium pressure or high performance liquid chromatography.
  • the chromatographic column is a C18 column, and the eluent is acetonitrile, water and a small amount of acetic acid.
  • the column was pre-equilibrated with 200 ml of 15% acetonitrile/water/0.1% glacial acetic acid solution.
  • Example 2 This example is based on Example 1. The difference from Example 1 is that after the deprotection of the last amino acid at the N-terminus of the peptide chain is completed, 2 ml of acetic acid and 2 ml of DIEA are added, and the reaction is carried out for 30 min. .
  • GHSR-active compounds Screening of GHSR-active compounds is accomplished by recombinantly expressing the receptor.
  • the use of recombinant expression of GHSR offers several advantages, such as the ability to express the receptor in a defined cellular system so that the response of a compound to GHSR can be more easily distinguished from that of other receptors.
  • GHSR can be expressed in cell lines such as HEK293, COS7, and CHO that normally express GHSR without an expression vector, and the same cell line without an expression vector can be used as a control.
  • the activity of GHSR-1a can be measured using different techniques, eg, by detecting changes in the intracellular conformation of GHSR, changes in G-protein coupling activity, and/or changes in intracellular messengers, among others.
  • GHSR-1a activity is preferably measured using techniques such as the determination of intracellular Ca2 + .
  • techniques known in the art that can be used to measure Ca include the use of Calcium ion detection kit, etc.
  • the calcium detection kit uses calcium-sensitive indicators and blocking dyes to enable researchers to perform highly sensitive fluorescent screening for G protein-coupled receptors, ion channels, and other calcium-sensitive targets. In this experiment, FLIPR calcium 6 detection kit and FLIPR calcium 6-QF detection kit were used.
  • Cell seeding medium F12K + 10% fetal bovine serum.
  • a component take the test buffer and A component to room temperature (RT), add 10 ml of buffer to A component, vortex for 1-2 min, and store at -20°C;
  • X log value of compound concentration
  • Y Activation% or Inhibition%
  • Cytochrome P450-containing human liver microsomes (0.253 mg/mL protein) were mixed with test compounds (0.05-50 ⁇ M), CYPs substrates (10 ⁇ M acetaminophen, 5 ⁇ M diclofenac, 30 ⁇ M mephentoin, 5 ⁇ M dexamethasone Methorphan, 2 ⁇ M midazolam), 1.0 mM NADP were incubated at 37°C for 10 minutes. Naphthoflavone, sulfaphenazole, N-3-benzyl nivan, quinidine and ketoconazole were used as reference inhibitors. The results are shown in Table 3.

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Abstract

一种多肽化合物，具有式Ⅰ所示结构，或其立体异构体、混合物、药学上可接受的盐。实验结果表明，该多肽化合物能够有效地对GHSR-1a表现出较高的激动活性。

Description

一种多肽化合物及其应用
本申请要求于2021年03月30日提交中国专利局、申请号为202110343062.2、发明名称为“一种多肽化合物及其应用”的中国专利申请的优先权，其全部内容通过引用结合在本申请中。
技术领域
本发明涉及生物医药技术领域，尤其涉及一种多肽化合物及其应用。
背景技术
生长激素释放肽(ghrelin)是生长激素促分泌素受体(growth hormone secretagogue receptor，GHSR)的内源性配体。ghrelin是Kojima等在小鼠和人胃内分泌细胞及下丘脑弓状核中发现的，是目前为止发现的唯一的GHSR天然配体，它含有28个氨基酸残基，分子量为3.3kDa。人和大鼠的ghrelin前体蛋白由117个氨基酸组成，N端前23肽呈现分泌信号肽的特征；ghrelin的N端前4个氨基酸片段为其最小的活性中心，C末端的P-R结构(脯氨酸－精氨酸)为其识别部位。人和大鼠的ghrelin只有2个氨基酸残基不同，编码基因序列有82.9％的同源性。ghrelin在体内有两种分泌形式：一种是ghrelin的N端第3位丝氨酸发生了辛酰基化，另一种是该部位没有发生辛酰基化。ghrelin的N端第3位丝氨酸是其发挥生物学功能的实质部位。最初的研究认为去辛酰基化ghrelin(des-acyl ghrelin)不具备生物学活性，而最新研究发现，去辛酰基化和辛酰基化的ghrelin均能够促进脊索神经上皮的增殖；去辛酰基化ghrelin具有内分泌功能，能促进细胞增殖，具有抗凋亡作用。
GHSR是一种孤核的G蛋白耦联受体，除主要存在于啮齿动物和人的垂体、胃部外，还广泛分布于外周组织、脑、肠、肾脏、胰脏、心脏、脂肪组织等。GHSR分布的广泛性对ghrelin及其受体的多种生物学功能起重要作用。GHSR结构编码基因组在不同物种中高度保守，其氨基酸排列顺序与胃动素基因相关肽的G蛋白偶联蛋白受体有52％同源性。GHSR按不同的外显子编码分为1a型和1b型，其中GHSR-1a是ghrelin的功能性受体，该受体与ghrelin结合后激活磷脂酶C(PLC)、三磷酸肌醇(IP3)、蛋白激酶C(PKC)等发挥生物学效应。而非功能性受体GHSR-1b无生物学活性。
生长激素(growth hormone，GH)在垂体的脉冲式释放主要受下丘脑的生长激素释放激素(growth hormone releasing hormone，GHRH)、生长抑素 (Somatostatin，SS)和ghrelin三个因子调节。对GH的分泌，GHRH起促进作用，SS起抑制作用，ghrelin可与GHRH协同起到促进作用，三者在下丘脑形成局部神经内分泌调节反馈环。在调节GH分泌的系统中，GHRH与其受体结合，增加细胞内环磷酸腺苷的水平；ghrelin与其受体结合，K +通道的去极化和抑制，引起细胞内IP3浓度升高，细胞内的Ca 2+浓度升高，最终刺激GH的分泌。GH从垂体促生长激素细胞的释放也可受到生长激素释放多肽(GHRP)的控制。已经发现有一种六肽，H-His-D-Trp-Ala-Trp-D-Phe-Lys-CONH 2(GHRP-6)在包括人的几种物种中以剂量依赖性方式调节促生长激素细胞中释放生长激素(Bowers et.al.,Endocrinology 1984,114,1537-1545)。通过对GHRP-6的结构进行分析，研究人员又发现了一些GHRP类似物。
该类多肽以及类肽等化合物可以与GHSR-1a结合，产生激动活性，引起信号转导，从而调节GH的分泌，但在临床开发上均具有一定的局限性。因此，开发具有活性高、剂量小以及低毒副作用的结构是GHSR-1a受体激动剂的研究目标。
在保留或提高多肽的生理活性的同时，改善多肽的稳定性，是行之有效的药物开发策略之一。
发明内容
有鉴于此，本发明要解决的技术问题在于提供一种多肽化合物及其应用，制备的多肽化合物作为GHSR-1a受体激动剂具有较高的活性。
为达到上述目的，本发明提供了一种多肽化合物，具有式Ⅰ所示结构，或其立体异构体、混合物、药学上可接受的盐：
Figure PCTCN2022082973-appb-000001
其中，
R 1选自-NR 2R 3，-OR 2或-SR 2；
并且，R 1不为D型或L型氨基酸；
R 2和R 3独立地选自氢，氘，衍生自聚乙二醇的聚合物，非环状取代的或未经取代的脂族基团，取代的或未经取代的脂环基，取代的或未经取代的杂环基，取代的或未经取代的杂芳基烷基，取代的或未经取代的芳基，取代的或未经取代的芳烷基。
W选自单键、D型氨基酸或L型氨基酸；
U 1选自以下任一结构：
Figure PCTCN2022082973-appb-000002
其中，X和Z独立地选自CH-R 4，N-R 4，O，S，Se，S＝O或O＝S＝O；
R 4，R 7和R 8独立地选自氢，氘，氨基、保护基，衍生自聚乙二醇的聚合物，非环状取代的或未经取代的脂族基团，取代的或未经取代的脂环基，取代的或未经取代的杂环基，取代的或未经取代的杂芳基烷基，取代的或未经取代的芳基，取代的或未经取代的芳烷基或R 9CO-；
Y选自卤素、氨基、硝基、羟基或氰基；
R 5选自-NR 2R 3，-OR 2或-SR 2；
R 6选自氢，氘，非环状取代的或未经取代的脂族基团，取代的或未经取代的脂环基，取代的或未经取代的杂环基，取代的或未经取代的杂芳基烷基，取代的或未经取代的芳基，取代的或未经取代的芳烷基；
m 1和m 2独立地选自0，1，2或3；特别地，当X为N时，m 1为2，m 2则独立地选自0，1，3；m 2为2，m 1则独立地选自0，1，3；
m 3和m 4独立地选自0，1，2或3；
n 1，n 2，n 3和n 4独立地选自0，1，2或3；
p为0，1，2，3，4或5；
U 2为单键，或选自以下任一结构，且U 2的羰基端与W连接：
Figure PCTCN2022082973-appb-000003
R 9选自氢，非环状取代的或未经取代的脂族基团，取代的或未经取代的脂环基，取代的或未经取代的杂环基，取代的或未经取代的杂芳基烷基，取代的或未经取代的芳基，取代的或未经取代的芳烷基。
本发明中，所述R 1选自-NR 2R 3，-OR 2或SR 2。
其中，R 2和R 3独立地选自氢，氘，衍生自聚乙二醇的聚合物，非环状取代的或未经取代的脂族基团，取代的或未经取代的脂环基，取代的或未经取代的杂环基，取代的或未经取代的杂芳基烷基，取代的或未经取代的芳基，取代的或未经取代的芳烷基。更优选为氢，氘，衍生自聚乙二醇的聚合物，非环状取代的或未经取代的C 1-10脂族基团，取代的或未经取代的C 3-10脂环基，取代的或未经取代的C 2-10杂环基，取代的或未经取代的C 2-20杂芳基烷基，取代的或未经取代的C 6-12芳基或取代的或未经取代的C 6-12芳烷基。
本发明优选的，所述R 1选自-NR 2R 3或-OR 2，其中R 2和R 3独立地选自氢，甲基，乙基，己基，十二烷基或十六烷基。
本发明中，所述R 1不为D型或L型氨基酸。
本发明中，上述氨基酸指氨基酸残基，具体的，为氨基酸通过氨基或羧基反应，形成多肽后的残基。
本发明中，所述W独立地选自单键、D型氨基酸或L型氨基酸。更优选的，所述W选自单键、丙氨酸，精氨酸，天冬酰胺，半胱氨酸，谷氨酰胺，天冬氨酸，谷氨酸，甘氨酸，组氨酸，异亮氨酸，亮氨酸，赖氨酸，甲硫氨酸，苯丙氨酸，脯氨酸，丝氨酸，苏氨酸，色氨酸，酪氨酸，缬氨酸残基中的一种或多种。
本发明中，当所述W选自单键时，即U 2和R 1直接相连接。
当所述W选自D型氨基酸或L型氨基酸时，所述氨基酸脱去一分子水，和相邻基团形成酰胺键。
所述残基是指W通过脱去一分子水和相邻基团形成酰胺键，进而形成多 肽化合物。
本发明中，所述U 1选自以下任一结构：
Figure PCTCN2022082973-appb-000004
其中，X和Z独立地选自CH-R 4，N-R 4，O，S，Se，S＝O或O＝S＝O；更优选为N-R 4或O。
R 4，R 7和R 8独立地选自氢，氘，氨基、保护基，衍生自聚乙二醇的聚合物，非环状取代的或未经取代的脂族基团，取代的或未经取代的脂环基，取代的或未经取代的杂环基，取代的或未经取代的杂芳基烷基，取代的或未经取代的芳基，取代的或未经取代的芳烷基或R 9CO-；更优选为氢，氘，氨基、衍生自聚乙二醇的聚合物，非环状取代的或未经取代的C 1-10脂族基团，取代的或未经取代的C 3-10脂环基，取代的或未经取代的C 2-10杂环基，取代的或未经取代的C 2-20杂芳基烷基，取代的或未经取代的C 6-12芳基，取代的或未经取代的C 6-12芳烷基或R 9CO-。进一步优选为氢，氨基、C 1-6烷基，C 6-14芳基，C 3-8环烷基或C 2-10酰基。
所述R 9优选为氢，非环状取代的或未经取代的脂族基团，取代的或未经取代的脂环基，取代的或未经取代的杂环基，取代的或未经取代的杂芳基烷基，取代的或未经取代的芳基，取代的或未经取代的芳烷基；更优选为氢，非环状取代的或未经取代的C 1-10脂族基团，取代的或未经取代的C 3-10脂环基，取代的或未经取代的C 2-10杂环基，取代的或未经取代的C 2-20杂芳基烷基，取代的或未经取代的C 6-12芳基，取代的或未经取代的C 6-12芳烷基。进一步优选为氢或C 1～6烷基；在本发明的一些具体实施例中，所述R 9具体为甲基、乙基、丙基、异丙基或丁基。
本发明中，所述Y选自卤素、氨基、硝基、羟基或氰基。更优选为F、Cl、Br或氨基。
本发明中，所述R 5独立地选自-NR 2R 3，-OR 2或-SR 2；更优选为-NR 2R 3。
上述R 2、R 3的范围同上，在此不再赘述。
进一步优选的，R 2、R 3独立的选自氢、甲基、乙基或己基。
本发明中，所述R 6独立地选自氢，氘，非环状取代的或未经取代的脂族基团，取代的或未经取代的脂环基，取代的或未经取代的杂环基，取代的或未经取代的杂芳基烷基，取代的或未经取代的芳基，取代的或未经取代的芳烷基；更优选为氢，氘，非环状取代的或未经取代的C 1-10脂族基团，取代的或未经取代的C 3-10脂环基，取代的或未经取代的C 2-10杂环基，取代的或未经取代的C 2-20杂芳基烷基，取代的或未经取代的C 6-20芳基，取代的或未经取代的C 6-12芳烷基。进一步优选为氢、C 1-6烷基、C 6-14芳基或C 3-8环烷基。
本发明优选的，所述U 1选自以下结构：
Figure PCTCN2022082973-appb-000005
其中，R 6选自氢，取代或非取代的C 1-6烷基，取代或非取代的C 6-14芳基，取代或非取代的C 3-8环烷基；所述取代的基团优选为卤素、氨基、硝基、羟基、酰基取代的氨基、脲基或胍基。
进一步优选的，所述R 6选自氢，取代或未取代的甲基、乙基、丙基、异丙基、丁基、异丁基或叔丁基。
所述取代的基团选自卤素、氨基、硝基、羟基、甲酰胺基、乙酰胺基、丙酰胺基、丁酰胺基、脲基或胍基。
R 7和R 8独立地优选为氢，C 1-6烷基，C 6-14芳基，C 3-8环烷基，C 2-10酰基；更优选为氢、甲基、乙基、丙基、异丙基、丁基、异丁基、叔丁基、甲酰基、乙酰基、丙酰基或丁酰基。
本发明优选的，所述U 1选自以下任一结构：
Figure PCTCN2022082973-appb-000006
其中，R 10、R 11独立地优选为氢、氨基、硝基、羟基、卤素、氰基、胺甲基、胺乙基、胺丙基或胺丁基。
本发明优选的，所述U 2为单键，或选自以下任一结构(即4-氨基-4-哌啶甲酸(Apc)或赖氨酸(Lys)，鸟氨酸(Orn)，精氨酸(Arg)残基)，且U 2的羰基端与W连接：
Figure PCTCN2022082973-appb-000007
本发明中，当U 2为单键时，W直接和母核的羰基相连接。
本发明中，所述m 1和m 2独立地选自0，1，2或3；特别地，当X为N时，m 1为2，m 2则独立地选自0，1，3；m 2为2，m 1则独立地选自0，1，3；即所述含m 1和m 2的结构不选自吡啶基团。
本发明中，m 3和m 4独立地选自0，1，2或3。
本发明中，n 1，n 2，n 3和n 4独立地选自0，1，2或3。
本发明中，p为0，1，2，3，4或5。
本发明中，当p为0时，所述N原子直接和C原子相连接。
本发明优选的，所述多肽化合物，具有以下任一结构，或其立体异构体、 混合物、药学上可接受的盐：
Figure PCTCN2022082973-appb-000008
Figure PCTCN2022082973-appb-000009
Figure PCTCN2022082973-appb-000010
Figure PCTCN2022082973-appb-000011
Figure PCTCN2022082973-appb-000012
在本发明的实施方案中，如本文所述的多肽化合物、它们的立体异构体、其混合物、它们的药学上可接受的盐的合成能够根据现有技术已知的任何常规 方法进行，比如用固相肽合成方法[Stewart J.M.y Young J.D.,“Solid Phase Peptide Synthesis,2nd edition”,(1984),Pierce Chemical Company,Rockford,Illinois；Bodanzsky M.y Bodanzsky A.,“The practice of Peptide Synthesis”,(1994),Springer Verlag,Berlin；Lloyd Williams P.et al.,“Chemical Approaches to the Synthesis of Peptides and Proteins”,(1997),CRC,Boca Raton,FL,USA],溶液中的合成、酶促合成[Kullmann W.“Proteases as catalysts for enzymic syntheses of opioid peptides”,(1980),J.Biol.Chem.,255(17),8234-8238]或其任意组合。化合物还能够通过经过目的在于产生所希望序列的基因工程修饰的或未修饰的细菌菌株的发酵来获得，或者通过动物、真菌或优选植物来源的蛋白的游离含有至少所希望序列的肽段的受控水解来获得。例如，可以使用编码加本文所述的多肽氨基酸序列的核酸序列以及任选地进行适当的氨基酸修饰来产生本发明的化合物。
仅作为示例，获得本发明的多肽化合物、它们的立体异构体和其混合物的方法可以包括下述阶段：
-将N-末端保护的和C-末端游离的氨基酸与N-末端游离的和C-末端保护或结合至固体载体的氨基酸偶联；
-消除保护N-末端的基团；
-重复偶联程序并且消除保护N-末端的基团直至获得所希望的肽序列；
-消除保护C-末端的基团或裂解固体载体；
优选地，C-末端结合至固体载体并且该过程在固相中进行，因此包括将N-末端保护的和C-末端游离的氨基酸与N-末端游离的和C-末端结合至聚合物载体的氨基酸偶联；消除保护N-末端的基团；和按必要次数重复该程序以获得所希望长度的化合物，最终随后从最初的聚合物载体的裂解合成的化合物。
在整个合成当中，氨基酸侧链的官能团保持用暂时或永久保护性基团方便地保护，并且能够与从聚合物载体裂解肽的过程同时地或正交地脱保护。
另选地，固相合成能够用会聚策略进行：将肽与聚合物载体偶联或者与预先结合至聚合物载体的肽或氨基酸偶联。会聚合成策略是本领域技术人员广泛已知的并且描述于Lloyd-Williams P.et al.,“Convergent Solid-Phase Peptide Synthesis”,(1993),Tetrahedron,49(48),11065-11133。
在采用现有技术已知的标准程序和条件的情况下，本发明过程能够以无差 别顺序包括额外的C-末端脱保护和/或从聚合物载体裂解肽的阶段；在此之后这些末段官能团能够加以修饰。在式(I)所示多肽化合物固定至聚合物载体时或一旦多肽化合物已从聚合物载体分开，则能够进行C-末端的任选修饰。
任选地和/或额外地，R 1残基能够这样引入：在适当溶剂和碱比如N,N-二异丙基乙胺(DIEA)或三乙胺或添加剂比如1-羟基苯并三唑(HOBt)或1-羟基氮杂苯并三唑(HOAt)和脱水剂比如碳二亚胺、脲鎓盐、鏻盐或脒鎓盐等存在下，将化合物HR 1，其中R 1是-OR 2、-NR 2R 3或-SR 2，与互补片段相应于式(I)化合物发生反应，其中R 1是-NH 2；或者通过先将互补片段相应于式(I)化合物与例如亚硫酰氯预先形成酰卤，再与HR 1发生反应，由此获得化学式(I)的根据本发明的肽，其中片段具有并不牵涉于N-C键形成中官能团用暂时或永久保护性基团加以适宜保护；或者另选地其它R 1残基可以通过同时掺入肽从聚合物载体裂解的过程来引入。
本领域技术人员会容易地理解C-末端和N-末端的脱保护/裂解步骤和它们随后的衍生化能够根据现有技术已知的过程以不同顺序进行。
本发明提供了一种组合物，包括上述多肽化合物，和可接受的辅剂。
本发明中，上述组合物可以为药物组合物，或保健品组合物。
本发明中，所述辅剂包括但不限于本领域技术人员熟知的载体、稀释剂、赋形剂或辅助剂等。
本发明优选的，所述载体包括但不限于无菌水、盐水、缓冲液、磷酸缓冲盐水、缓冲氯化钠、植物盐、最小必需培养基(MEM)、具有HEPES的MEM，等等。
本发明上述组合物中，多肽化合物可以单独存在，或两种及两种以上混合存在，或通过复合、结晶或离子键合或共价键合更紧密地缔合。
本发明提供的多肽化合物C端的氨基酸残基中刚性结构或柔性结构的大小对于维持序列中肽键的构型至关重要，进而，本发明通过在序列C端进行不同结构类型官能团的引入，使多肽化合物可以有效地与GHSR-1a结合，并适合治疗、预防、减轻或诊断由GHSR-1a介导的紊乱所致相关疾病。
基于此，本发明提供了上述多肽化合物，或上述制备方法制备的多肽化合物，或上述组合物，作为生长激素促分泌素受体的激动剂的应用，或在制备用于治疗、预防、减轻和/或诊断由生长激素促分泌素受体介导的紊乱所致相关 疾病的药物，或作为促进生长发育的保健品中的应用。
本发明中，所述生长激素促分泌素受体，也可以称之为胃饥饿素受体、生长激素释放肽受体或GHSR-1a受体。
本发明优选的，所述由生长激素促分泌素受体介导的紊乱所致相关疾病为生长激素缺乏症。
具体的，本发明提供了上述多肽化合物，或上述制备方法制备的多肽化合物，或上述组合物作为GHSR-1a激动剂的用途。
具体的，本发明提供了上述多肽化合物，或上述制备方法制备的多肽化合物，或上述组合物在制备GHSR-1a激动剂的用途。
本发明提供的上述多肽化合物、组合物，或生长激素促分泌素受体的激动剂可以以多种方式施用，这取决于期望局部施用还是全身性施用并取决于待治疗的区域。在一些实施方案中，可经过以下方式向患者施用所述多肽化合物或其组合物或其GHSR-1a激动剂：口腔或直肠、或经粘膜、或肠内、或肌内、或皮下、或髓内、或鞘内、或直接心室内、或静脉内、或玻璃体内、或腹膜内、或鼻内、或眼内。
本发明中，所述术语“保护性基团”涉及阻塞有机官能团的和能够在受控条件下除去的基团。保护性基团、它们的相对反应性和它们保持惰性的条件是本领域技术人员已知的。
氨基基团的代表性保护性基团的实例尤其是酰胺乙酸酯，酰胺苯甲酸，酰胺特戊酸脂；氨基甲酸酯类比如苄氧基羰基(Cbz或Z)，2-氯苄基(CIZ)，对-硝基苄氧基羰基(pNZ)，叔丁氧基羰基(Boc)，2,2,2-三氯乙氧羰基(Troc)，2-(三甲基甲硅烷基)乙基氧基羰基(Teoc)，9-芴基甲基氧基羰基(Fmoc)或烯丙基氧基羰基(Alloc)，三苯甲基(Trt)，甲氧基三苯甲基(Mtt)，2,4-二硝基苯基(Dnp)，N-1-(4,4-二甲基-2,6-二氧代环己-1-亚基)乙基(Dde)，1-(4,4-二甲基-2,6-二氧代-亚环己基)-3-甲基丁基(ivDde)，1-(1-金刚烷基)-1-甲基乙氧基羰基(Adpoc)，优选Boc或Fmoc。
羧基基团代表性保护性基团的实例是酯，比如叔丁基酯(tBu)，烯丙基酯(All)，三苯基甲基酯(Trt酯)，环己基酯(cHx)，苄基酯(Bzl)，邻硝基苄基酯，对硝基苄基酯，对甲氧基苄基酯，三甲基甲硅烷基乙基酯，2-苯基异丙基酯，芴基甲基酯(Fm)，4-(N-[1-(4,4-二甲基-2,6-二氧代-亚环己基)-3-甲基丁 基]氨基)苄基酯(Dmab)，优选All，tBu，cHx，Bzl和Trt酯。
三官能氨基酸的侧链能够在合成过程期间用与N-末端和C-末端保护性基团正交的暂时或永久保护性基团加以保护。
色氨酸侧链的吲哚基团能够通过甲酰基基团(For)，Boc，Mts保护或者能够不加保护地使用。4-氨基-4-哌啶甲酸侧链的哌啶基团通过Boc或Fmoc保护。精氨酸侧链可以通过以下保护性基团保护：Tos，4-甲氧基-2,3,6-三甲基苯磺酰基(Mtr)，Alloc，硝基，2,2,4,6,7-五甲基二氢苯并呋喃-5-磺酰基(Pbf)和2,2,5,7,8-五甲基色满-6-磺酰基(Pmc)。为了保护赖氨酸和鸟氨酸侧链的氨基基团能够使用酰胺，比如乙酸酰胺，苯甲酸酰胺，特戊酸酰胺；氨基甲酸酯类比如Cbz或Z，CIZ，pNZ，Boc，Troc，Teoc，Fmoc或Alloc，Trt，Mtt，Dnp，Dde，ivDde，Adpoc等。
在优选实施方式中，所用的保护性基团策略是如下的策略：氨基基团通过Boc保护，羧基基团通过Bzl、cHx或All保护，精氨酸侧链通过Tos保护，4-氨基-4-哌啶甲酸侧链的哌啶基团通过Fmoc保护，色氨酸侧链通过For或Mts保护并且Apc赖氨酸和鸟氨酸侧链通过CIZ、Fmoc或Alloc保护。
在又一优选的实施方式中，所用的保护性基团策略是如下的策略：氨基基团通过Fmoc保护，羧基基团通过tBu、All或Trt酯保护，精氨酸侧链通过Pmc或Pbf保护，4-氨基-4-哌啶甲酸侧链的哌啶基团通过Boc保护，色氨酸侧链通过Boc保护或未加保护地使用，并且赖氨酸和鸟氨酸侧链通过Boc、Trt或Alloc保护。
这些和其它保护性基团的实例，它们的引入和除去，能够参考文献[Atherton B.and Sheppard R.C.,“Solid Phase Peptide Synthesis:A practical approach”,(1989),IRL Oxford University Press]。术语“保护性基团”也包括固相合成中的聚合物载体。
在合成完全或部分发生在固相中的情况下，用于本发明过程中的可能固体载体涉及聚苯乙烯载体，接枝至聚苯乙烯的聚乙二醇等，比如且不限于对甲基二苯甲基桉树酯(MBHA)[Matsueda G.R.et al.,“A p-methyl benzhydrylamine resin for improved solid-phase synthesis of peptide amides”,(1981),Peptides,2,4550],2-氯三苯甲基树脂[Barlos K.et al.,“Darstellung geschützter PeptidFragmente unter Einsatz substituierter Triphenylmethyl Harze”,(1989), Tetrahedron Lett.,30,3943-3946；Barlos K.et al.,“Veresterung von partiell geschützten PeptidFragmenten mit Harzen Einsatz von 2-Chlorotritylchlorid zur Synthese von LeulGastrin I”,(1989),Tetrahedron Lett.,30,39473951]，
Figure PCTCN2022082973-appb-000013
树脂(Rapp Polymere GmbH)，
Figure PCTCN2022082973-appb-000014
树脂(Matrix Innovation,Inc)等，其可以包括或可以不包括不稳定连接体，比如5-(4-氨甲基-3,5-二甲氧基苯氧基)戊酸(PAL)[Albericio F.et al.,“Preparation and application of the5-(4-(9-fluorenylmethyloxycarbonyl)aminomethyl-3,5-dimethoxy-phenoxy)valeric acid(PAL)handle for the solid-phase synthesis of C-terminal peptide amides under mild conditions”,(1990),J.Org.Chem.,55,3730-3743]，2-[4-氨甲基-(2,4-二甲氧基苯基)]苯氧基乙酸(AM)[Rink H.,“Solid-phase synthesis of protected peptide fragments using a trialkoxy-diphenyl-methylester resin”,(1987),Tetrahedron Lett.,28,3787-3790],Wang[Wang S.S.,“p-Alkoxybenzyl Alcohol Resin and p-Alkoxybenzyl oxycarbonylhydrazide Resin for Solid Phase Synthesis of Protected Peptide Fragments”,(1973),J.Am.Chem.Soc.,95,1328-1333]等，其使得可以同时脱保护和从聚合物载体裂解肽。
定义
在本发明中使用的缩写具有下面的含义：
Ala(Alanine，A)丙氨酸
Aba(2-aminobutyric acid)2-氨基丁酸
Apc(4-Amino-4-piperidine formic acid)4-氨基-4-哌啶甲酸
Arg(Arginine,R)精氨酸
Bal(3-Benzothienylalanine)3-苯并噻吩基丙氨酸
Boc(butyloxycarboryl)叔丁氧羰基
Cit(Citrulline)瓜氨酸
DCM(Dichloromethane)二氯甲烷
DIEA(N,N-Diisopropylethylamine)N,N-二异丙基乙胺
DMF(N,N-Dimethylformamide)N,N-二甲基甲酰胺
Fmoc(Fluorenylmethoxycarbonyl)芴甲氧羰基
Gly(Glycine，G)甘氨酸
HBTU(O-Benzotriazole-N,N,N',N'-tetraMethyl-uroniuM-hexafluorophosphat  e)O-苯并三氮唑-四甲基脲六氟磷酸酯
HOBt(N-Hydroxybenzotrizole)1-羟基苯并三唑
HPLC(High performance liquid chromatography)高效液相色谱
Leu(Leucine，L)亮氨酸
Lys(Lysine,K)赖氨酸
Nle(Norleucine)正亮氨酸
Orn(Ornithine)鸟氨酸
Phe(3-Amino-4-phenylbutyric acid,F)苯丙氨酸
Pro(Proline，P)脯氨酸
TFA(Trifluoroacetic acid)三氟乙酸
Tle(Tertiary leucine)叔亮氨酸
Trp(Tryptophan,W)色氨酸
Val(Valine，V)缬氨酸
如本文所定义的，术语“多肽”，“肽”和“氨基酸序列”在本文中可互换使用，是指任何长度的氨基酸残基的聚合物。该聚合物可以是直链或支链的，它可以包含修饰的氨基酸或氨基酸类似物，并且可以被非氨基酸的化学部分打断。该术语还包括已被天然或人工修饰(例如二硫键形成，糖基化，脂化，乙酰化，磷酸化或任何其他操作或修饰，例如与标记或生物活性组分缀合)的氨基酸聚合物。术语“肽”包括通过共价键(例如酰胺键)连接的两个或更多个天然存在的或合成的氨基酸。
在本公开内容的上下文中，术语“氨基酸”被定义为具有至少一个伯、仲、叔或季氨基和至少一个酸基，其中酸基可以是羧酸、磺酸或磷酸或其混合物。氨基相对于酸基可以是“α”、“β”、“γ”至“ω”。合适的氨基酸包括但不限于在肽中发现的20种常见天然存在的氨基酸(例如丙氨酸，精氨酸，天冬酰胺，半胱氨酸，谷氨酰胺，天冬氨酸，谷氨酸，甘氨酸，组氨酸，异亮氨酸，亮氨酸，赖氨酸，甲硫氨酸，苯丙氨酸，脯氨酸，丝氨酸，苏氨酸，色氨酸，酪氨酸，缬氨酸)的D-和L-异构体以及通过有机合成或其他代谢途径制备的天然存在的和非天然存在的氨基酸。
“氨基酸的主链”可以被选自卤素、羟基、胍基、杂环基团的一个或多个基团取代。因此术语“氨基酸”在其范围内还包括甘氨酸，丙氨酸，缬氨酸，亮氨 酸，异亮氨酸，正亮氨酸，甲硫氨酸，脯氨酸，苯丙氨酸，色氨酸，丝氨酸，苏氨酸，半胱氨酸，酪氨酸，天冬酰胺，谷氨酰胺，天冬氨酸，谷氨酸，赖氨酸，组氨酸，高半胱氨酸，牛磺酸，甜菜碱，N-甲基丙氨酸等。(L)和(D)形式的氨基酸被包括在内。
术语“氨基酸侧链”是指连接至氨基酸的α-碳的部分。例如，丙氨酸的氨基酸侧链是甲基，苯丙氨酸的氨基酸侧链是苯基甲基，半胱氨酸的氨基酸侧链是硫代甲基，天冬氨酸的氨基酸侧链是羧甲基，酪氨酸的氨基酸侧链是4-羟基苯甲基，等等。还包括其他非天然存在的氨基酸侧链，例如天然存在的(例如氨基酸代谢物)或合成制备的(例如α-取代的氨基酸)。
如本文所用，术语“非环状脂族基团”涵盖线性或支化的烷基，烯基和炔基基团。
术语“烷基”是指线性或支化的饱和基团，其具有1至24，优选1至16，更优选1至14，甚至更优选1至12，还更优选1、2、3、4、5或6个碳原子和通过简单键结合至分子其余部分，包括例如且不限于，甲基，乙基，异丙基，异丁基，叔丁基，庚基，辛基，癸基，十二烷基，月桂基，十六烷基，十八烷基，戊基，2-乙基己基，2-甲基丁基，5-甲基己基等。
术语“烯基基团”是指线性或支化的基团，其具有2至24，优选2至16，更优选2至14，甚至更优选2至12，还更优选2、3、4、5或6个碳原子，具有一个或多个碳-碳双键，优选具有1，2或3个碳-碳双键，键是共轭或不共轭的，其通过简单键合至分子其余部分，包括例如且不限于乙烯基(-CH 2＝CH 2)，烯丙基(-CH 2-CH＝CH 2)，油烯基，亚油烯基等基团。
术语“炔基基团”是指线性或支化的基团，其具有2至24，优选2至16，更优选2至14，甚至更优选2至12，还更优选2、3、4、5或6个碳原子，具有一个或多个碳-碳三键，优选具有1，2或3个碳-碳三键，键是共轭或不共轭的，其通过简单键合至分子其余部分，包括例如且不限于乙炔基基团，1-丙炔基，2-丙炔基，1-丁基，2-丁基，3-丁基，戊基，比如1-戊基等。炔基基团还能够含有一个或多个碳-碳双键，包括例如且不限于基团丁-1-烯-3-炔基，戊-4-烯-1-炔基等。
术语“脂环基基团”在本发明中用于涵盖例如且不限于环烷基或环烯基或环炔基基团。
术语“环烷基”是指饱和的单环或多环脂族基团，其具有3至24，优选3至16，更优选3至14，甚至更优选3至12，还更优选3、4、5或6个碳原子并且通过简单键结合至分子其余部分，包括例如且不限于，环丙基，环丁基，环戊基，环己基，环庚基，甲基环己基，二甲基环己基，八氢茚，十氢萘，十二氢非那烯等。
术语“环烯基”是指非芳族单环或多环脂族基团，其具有5至24，优选5至16，更优选5至14，甚至更优选5至12，还更优选5至6个碳原子，具有一个或多个碳-碳双键，优选1，2或3个碳-碳双键，键是共轭或不共轭的，其通过简单键合至分子其余部分，包括例如且不限于环戊-1-烯-1-基基团等。
术语“环炔基”是指非芳族单环或多环脂族基团，其具有8至24，优选8至16，更优选8至14，甚至更优选8至12，还更优选8或9个碳原子，具有一个或多个碳-碳三键，优选1，2或3个碳-碳三键，键是共轭或不共轭的，其通过简单键合至分子其余部分，包括例如且不限于环辛-2-炔-1-基基团等。环炔基基团还能够含有一个或多个碳-碳双键，包括例如且不限于环辛-4-烯-2-炔基基团等。
术语“芳基基团”是指芳族基团，其具有6至30，优选6至18，更优选6至10，还更优选6或10个碳原子，其包含1、2、3或4个芳族环，通过碳-碳键结合或稠合，包括例如且不限于苯基，萘基，二苯基，茚基，菲基或蒽基等；或者芳烷基基团。
术语“芳烷基”是指被芳族基团取代的烷基，具有7至24个碳原子和包括例如且不限于-(CH 2) 1-6-苯基、-(CH 2) 1-6-(1-萘基)、-(CH 2) 1-6-(2-萘基)、-(CH 2) 1-6-CH(苯基) 2以及类似物。
术语“杂环基基团”是指3-10元的烃化的环，其中环中原子的一个或多个，优选环中原子的1、2或3个是不同于碳的元素比如氮、氧或硫，并且可以是饱和或不饱和的。出于本发明意图，杂环能够是单环、双环或三环系统，其可以包括稠环系统；和残基杂环中的氮、碳或硫原子可以被任选地氧化；氮原子可以被任选地季铵化；和残基杂环基可以是部分或完全饱和的或是芳族的。术语杂环基最优选是指5或6元环。饱和的杂环基基团的实例是二噁烷，哌啶，哌嗪，吡咯烷，吗啉和硫吗啉。芳族杂环基基团，也称为杂芳族基团的实例是吡啶，吡咯，呋喃，噻吩，苯并呋喃，咪唑啉，对苯二酚，喹啉和萘啶。
术语“杂芳基烷基基团”是指被取代的或未经取代的芳族杂环基基团取代的烷基，烷基具有1至6个碳原子和芳族杂环基基团具有2至24个碳原子和1至3个非碳的原子，并且包括例如且不限于-(CH 2) 1-6-咪唑基、-(CH 2) 1-6-三唑基、-(CH 2) 1-6-噻吩基、-(CH 2) 1-6-呋喃基、-(CH 2) 1-6-吡咯烷基等。
本文使用的术语“卤素”或变体如“卤化物”或“卤代”是指氟，氯，溴和碘。
如本文所用的术语“杂原子”或变体如“杂-”是指O、N、NH和S。
如本文所用的术语“烷氧基”是指直链或支链烷氧基。实例包括甲氧基，乙氧基，正丙氧基，异丙氧基，叔丁氧基等。
如本文所用的术语“氨基”是指-NR aR b形式的基团，其中R a和R b独立地选自包括但不限于氢、任选取代的烷基、任选取代的烯基、任选取代的炔基和任选取代的芳基的组。
应理解，如本文所述的本发明的化合物均可被任何数目的取代基或官能团部分取代。通常，术语“(经)取代(的)”(无论其是否在术语“任选地”之后)和含于本发明的式中的取代基都是指用指定取代基的基团对给定结构中的氢基的替代。当任何给定结构中的超过一个位置可被超过一个选自指定基团的取代基取代时，所述取代基在每个位置可相同或不同。如本文所用的术语“经取代的”预期包括用有机化合物的所有可允许取代基、本文所述的任何取代基进行的取代。
例如，取代基包括但不限于导致形成稳定部分的以下基团：脂肪族基、烷基、烯基、炔基、杂脂肪族基、杂环基、芳基、杂芳基、酰基、氧代、亚氨基、硫羰基、氰基、异氰基、氨基、叠氮基、硝基、羟基、硫醇基和卤代以及它们的任何组合，包括但不限于以下基团：脂肪族氨基、杂脂肪族氨基、烷基氨基、杂烷基氨基、芳基氨基、杂芳基氨基、烷基芳基、芳基烷基、脂肪族氧基、杂脂肪族氧基、烷氧基、杂烷氧基、芳氧基、杂芳氧基、脂肪族硫基、杂脂肪族硫基、烷硫基、杂烷硫基、芳硫基、杂芳硫基、酰氧基等。本发明涵盖任何和所有的此类组合以得到稳定的取代基/部分。出于本发明的目的，例如氮的杂原子可具有氢取代基和/或满足杂原子的化合价并且导致形成稳定部分的如本文所述的任何合适的取代基。
化合物可含有一个或多个不对称中心，因此以外消旋体和外消旋混合物、单一对映异构体、个体非对映异构体和非对映异构体混合物存在。本文明确包 括这些化合物的所有这些异构形式。化合物也可以以多种互变异构形式表示，在这种情况下，本文明确地包括本文所述的化合物的所用互变异构形式(例如，环体系的烷基化可导致多个位点的烷基化，本文明确地包括所有这样的反应产物)。本文明确地包括了这种化合物的所有这些异构形式。本文明确包括在此描述的化合物的所有晶体形式。
本发明化合物能够作为立体异构体或立体异构体的混合物存在；例如，构成它们的氨基酸能够相互独立地具有构型L-、D-或外消旋。因此，可能的是，获得异构混合物以及外消旋混合物或非对映体的混合物，或纯的非对映体或对映体，取决于不对称碳的数目和异构体或异构混合物所存在的不对称碳。本发明化合物的优选结构纯异构体，也即对映体或非对映体。
例如，在描述U 2能够是-Lys-的情况下，应理解U 2选自-L-Lys-，-D-Lys-或两者的混合物，是外消旋或非外消旋的。描述于该文献的制备程序使得本领域技术人员可以通过选择正确构型的氨基酸获得本发明化合物各自的立体异构体。
本发明的肽的药学上可接受的盐也属于本发明的领域之内。术语“药学上可接受的盐”意指其在动物中和更特别地在人类中的用途得到承认的盐，并且包括用以形成碱加成盐的盐，无论它们是无机盐还是有机盐，无机盐比如且不限于锂、钠、钾、钙、镁、锰、铜、锌或铝等，或有机盐比如且不限于乙胺、二乙胺、乙醇胺、二乙醇胺、精氨酸、赖氨酸、组氨酸或哌嗪等；或者酸加成盐，无论它们是有机盐还是无机盐，有机盐比如且不限于乙酸盐，柠檬酸盐，乳酸盐，丙二酸盐，马来酸盐，酒石酸盐，富马酸盐，苯甲酸，天冬氨酸盐，谷氨酸盐，琥珀酸盐，油酸酯，三氟乙酸盐，草酸盐，双羟萘酸盐或葡糖酸盐等，或无机盐比如且不限于盐酸盐，硫酸盐，磷酸盐，硼酸盐或碳酸盐等。盐的性质不是关键，条件是它是化妆上或药学上可接受的。本发明肽的药学上可接受的盐能够通过现有技术中熟知的常规方法获得(Berge S.M.等人，“Pharmaceutical Salts”,(1977),J.Pharm.Sci.,66,119，其通过引用整体并入本文)。
与现有技术相比，本发明提供了一种多肽化合物，具有式Ⅰ所示结构，或其立体异构体、混合物、药学上可接受的盐。实验结果表明，本发明提供的多肽化合物能够有效地对GHSR-1a表现出较高的激动活性。
具体实施方式
为了进一步说明本发明，下面结合实施例对本发明提供的多肽化合物及其应用进行详细描述。
本领域技术人员将会理解，下列实施例仅用于说明本发明，而不应视为限定本发明的范围。实施例中未注明具体条件者，按照常规条件或制造商建议的条件进行。所用试剂或仪器未注明生产厂商者，均为可以通过市购获得的常规产品。
多肽合成采用标准的Fmoc固相方法。选用Rink Amide树脂，肽链由C端向N端延长。保护氨基酸包括：Fmoc-Apc(Boc)-OH，Fmoc-D-Lys(Boc)-OH，Fmoc-D-Orn(Boc)-OH，Fmoc-Phe-OH，Fmoc-D-Trp(Boc)-OH，Fmoc-D-Bal-OH，Fmoc-D-Cit-OH，Fmoc-D-Arg(Pbf)-OH，Boc-D-Aba-OH，Fmoc-Lys(Boc)-OH，Fmoc-Lys(Alloc)-OH，Fmoc-Gly-OH，Fmoc-D-Ala-OH，Fmoc-D-Val-OH，Fmoc-D-Leu-OH，Fmoc-Pro-OH，Fmoc-β-Ala-OH，Fmoc-D-Tle-OH，Fmoc-Tle-OH，Fmoc-D-Nle-OH，Fmoc-Nle-OH，顺-2-(叔丁氧羰酰胺)-1-环戊烷羧酸，Boc-甲基-1-(氨基甲基)环丁烷羧酸，1-N-Boc-3-吖丁啶羧酸，Boc-3-氨基氧杂环丁烷-3-甲酸，1-Boc-D-吖啶-2-羧酸，(S)-1-Boc-吡咯烷-3-甲酸，Boc-2-吗啉甲酸，(Boc-3-氨基-1-金刚烷)乙酸，(1R,3S,4S)-N-Boc-2-氮杂双环[2.2.1]庚烷-3-羧酸，3-Boc-3-氮杂双环[3.1.0]己烷-1-羧酸。缩合剂为HBTU/HOBt/DIEA。脱保护试剂为哌啶/DMF溶液。粗肽水溶解后冻干保存。用中压液相色谱法或高效液相色谱法(HPLC)分离纯化，纯肽含量大于90％。基质辅助激光解析飞行时间质谱(MALDI-TOF-MS)确定肽序列分子量。
肽序列的合成：
合成条件如下：
保护氨基酸(天然或非天然)：0.2M的DMF溶液，
缩合剂：0.45M HBTU/HOBt的DMF溶液，
活化碱：2M DIEA的DMF溶液，
脱保护试剂：20％v/v哌啶的DMF溶液。
实施例1
化合物1-5，9，11-35，39，41-80的制备：
1.脱保护：称取Rink Amide树脂0.23g(0.1mmol)置于多肽合成反应器中，然后将脱保护试剂按上述浓度配置好后，加入到树脂中，室温条件下反应，抽干，再次加入哌啶/DMF，室温条件下反应后抽干，并用DMF洗涤，直至检测合格。
2.缩合反应：在冰浴条件下分别将氨基酸，缩合剂加入DMF活化，再加入活化碱反应获得活化液，最后将活化液加入树脂中，室温条件下反应后，用5％的茚三酮显色试剂使树脂显色，树脂变色，抽干溶剂并用DMF洗涤，检测合格后抽干溶剂，此时缩合反应完全。
3.重复上述脱保护和缩合反应直到肽链合成结束，得到包含完整多肽序列结构的肽树脂。
4.肽树脂的裂解：称取合成好的肽树脂1.25g，放入250ml茄形瓶中，冰浴，电磁搅拌。按1g肽树脂加入10ml的量配制裂解液【裂解液(体积百分比)：三氟乙酸：苯甲硫醚：水＝90：5：5】。TFA需预先冰浴降温30min或者预先存放于冰箱中使用；将配好的裂解液加入到冰浴条件下的肽树脂中，电磁搅拌，树脂变成黑色，冰浴条件下反应30min，然后撤掉冰浴，室温下继续搅拌反应180min，反应完成，剧烈搅拌下加入冰乙醚200ml，析出白色晶体，继续搅拌30min；用G4砂芯漏斗滤出析出物，用冷乙醚反复洗涤3遍，晾干。加入双蒸水50ml，乙腈5ml使固体充分溶解，抽滤，滤液冻干得粗肽1.04g。
5.粗肽的纯化：粗肽用中压或高效液相色谱进行纯化。色谱柱为C18柱，洗脱剂为乙腈，水和少量乙酸。具体操作步骤：称取粗肽1.00g，加水20ml，乙腈5ml使固体溶解，离心10min(5000转/分钟)，取上清液上样。色谱柱预先用15％乙腈/水/0.1％冰乙酸溶液200ml平衡。上样后继续用15％乙腈/水/0.1％冰乙酸溶液200ml冲洗，高效液相检测洗脱液成分。根据液相检测结果逐渐升高乙腈含量，直至所纯化的多肽主峰被洗脱出来。合并洗脱液，旋转蒸发取出大部分溶剂，冻干纯的多肽，HPLC检测含量大于90％，MALDI-TOF-MS确证分子量。
实施例2
化合物10，40的制备：
本实施例基于实施例1，与实施例1的区别在于，肽链N端最后一个保护 氨基酸为Fmoc-Lys(Alloc)-OH，其侧链保护基脱除方法为：抽干的树脂中加入三苯基膦钯：苯硅烷＝1：10(v/v)，避光，N 2保护反应3h，经检测树脂变色，脱保护完全；将树脂洗净抽干之后，进行乙酰化反应：加入乙酸2ml，DIEA 2ml，反应30min，洗净抽干。最后在20％v/v哌啶的DMF溶液中脱除Fmoc保护基，经裂解和纯化得到。
实施例3
化合物6-8，36-38的制备：
本实施例基于实施例1，与实施例1的区别在于，肽链N端最后一个氨基酸脱保护完成后，加入乙酸2ml，DIEA 2ml，反应30min，进行乙酰化封端，最后经裂解和纯化得到。
由本文所公开的实施方案的合成方法制备的多肽化合物如下表1所示。
表1合成多肽化合物列表
Figure PCTCN2022082973-appb-000015
Figure PCTCN2022082973-appb-000016
Figure PCTCN2022082973-appb-000017
Figure PCTCN2022082973-appb-000018
Figure PCTCN2022082973-appb-000019
Figure PCTCN2022082973-appb-000020
Figure PCTCN2022082973-appb-000021
Figure PCTCN2022082973-appb-000022
Figure PCTCN2022082973-appb-000023
Figure PCTCN2022082973-appb-000024
Figure PCTCN2022082973-appb-000025
Figure PCTCN2022082973-appb-000026
Figure PCTCN2022082973-appb-000027
Figure PCTCN2022082973-appb-000028
实施例4
多肽化合物对GHSR-1a的激动活性评价实验(IC 50)
GHSR活性化合物的筛选是通过重组表达受体来完成的。利用重组表达GHSR提供了几种优点，例如能够在确定的细胞系统中表达受体从而能够更容易区分化合物对GHSR的反应与对其它受体的反应。例如，可利用通常不用表达载体表达GHSR的诸如HEK293、COS7、以及CHO等细胞系中表达GHSR，而利用没有表达载体的相同细胞系作为对照。
可利用不同的技术来测量GHSR-1a的活性，例如，可通过检测GHSR细胞内构象的变化、G-蛋白偶联活性的变化、和/或细胞内信使的变化等来进行测量。优选采用诸如测定细胞内Ca 2+的技术来测量GHSR-1a活性。本领域公知的可用于测量Ca 2+的技术的例子包括使用
Figure PCTCN2022082973-appb-000029
钙离子检测试剂盒等。
Figure PCTCN2022082973-appb-000030
钙离子检测试剂盒采用了钙离子敏感的指示剂以及屏蔽染料以确保研究者进行高灵敏的用于G蛋白偶联受体、离子通道和其它钙离子敏感的靶标的荧光筛选。本实验采用FLIPR钙6检测试剂盒和FLIPR钙6-QF检测试剂盒。
1.试验过程
1.1细胞培养及试剂配制
a)细胞系：Flp In-CHO-GHSR Stable Pool；
b)完全培养基：F12K+10％胎牛血清+1x盘尼西林-链霉素(PS)+600μg/ml潮霉素B；
c)细胞播种培养基：F12K+10％胎牛血清。
d)检测缓冲液：1X HBSS+20mM HEPES。
e)10X A组分：取试验缓冲液和A组分至室温(RT)，加10ml缓冲液于A组分中，涡旋1-2min，保存于-20℃；
1.2化合物管理
a)化合物库存溶液：按照标准协议，将来自内部合成的粉末制成10mM DMSO溶液库存。
b)化合物存储：DMSO中的所有化合物均存放在室温干燥器中进行短期存储(最多4个月)。剩余的化合物在-20℃下长期保存。
1.3激动剂活性试验
a)用完整培养基培养Flp In-CHO-GHSR Stable Pool细胞。
b)将7K细胞/孔置于384孔细胞培养板(Corning,3764)的25磅/英寸细胞播种培养基中，37℃，5％CO 2培养过夜。
c)在室温条件下解冻20X A组分，用试验缓冲液将A组分稀释至2X，置于RT条件下。
d)从培养箱中取出培养皿，室温平衡10min。将培养基改为杏色缓冲液，最后洗涤后各孔保留缓冲液20μl，然后各孔加入2XA组分20μl,37℃孵育3-5s,。
e)在384孔细胞培养板上添加5X化合物10μl，用FLIPR Tetra立即采集数据。
2.数据分析
1)Z’factor＝1-3*(SD Max+SD Min)/(Mean Max-Mean Min)；
2)CV Max＝(SD Max/Mean Max)*100％；
3)CV Min＝(SD Min/Mean Min)*100％；
4)S/B＝Singal/Background；
5)IC 50值的计算公式:
Y＝Bottom+(Top-Bottom)/(1+10^((LogIC 50-X)*HillSlope))
X：log value of compound concentration；Y：Activation％or Inhibition％
根据上述方法，活性测试结果如表2所示。
表2多肽化合物对GHSR-1a的活性(IC 50)
SEQ ID NO. IC 50(nM)
1 149.7
2 3.5
3 17.2
4 1119.0
5 >1000
6 >1000
7 935.2
8 >1000
9 101.3
10 223.2
11 20.3
12 53.7
13 43.5
14 33.2
15 2.3
16 0.7
17 31.1
18 23.5
19 42.6
20 57.4
21 23.0
22 2.3
23 3.3
24 5.4
25 18.1
26 19.5
27 32.8
28 54.1
29 13.3
30 13.4
31 20.8
32 2.4
33 3.1
34 78.4
35 102.3
36 122.1
37 291.3
38 400.5
39 272.2
40 510.3
41 31.4
42 40.1
43 22.3
44 13.5
45 5.1
46 2.8
47 10.7
48 12.1
49 59.4
50 67.2
51 3.2
52 3.8
53 11.2
54 13.9
55 20.0
56 5.5
57 13.4
58 103.9
59 16.8
60 22.4
61 20.5
62 30.3
63 27.0
64 38.1
65 34.3
66 12.0
67 23.3
68 330.2
69 40.2
70 39.7
71 3.3
72 4.0
73 15.9
74 17.3
75 12.4
76 3.9
77 13.6
78 78.4
79 33.1
80 35.1
ghrelin(对照) 43.1
由表2的结果可知，本发明提供的多肽化合物显示了对GHSR-1a的激动活性。
实施例5
多肽化合物对细胞色素P450氧化酶的抑制
将含有细胞色素P450的人肝微粒体(0.253mg/mL蛋白)与测试化合物(0.05-50μM)、CYPs底物(10μM对乙酰氨基酚、5μM双氯芬酸、30μM美芬妥因、5μM氢溴酸右美沙芬、2μM米达唑仑)、1.0mM NADP在37℃温育10分钟。将萘黄酮、磺胺苯吡唑、N-3-苄基尼凡、奎尼定、酮康唑作作为参比抑制剂。结果如表3所示。
表3化合物的细胞色素P450CYP同工酶抑制活性(IC 50)
Figure PCTCN2022082973-appb-000031
Figure PCTCN2022082973-appb-000032
Figure PCTCN2022082973-appb-000033
由表3的结果可知，本发明提供的多肽化合物对细胞色素P450氧化酶的抑制IC 50值均大于50μM。
实施例6
多肽化合物81-84的制备以及活性检测
采用上述实施例1-80中所述相同的方法制备下表4所示的化合物81-84，并根据实施例4所述的方法测试这些化合物对GHSR的活性(IC 50)结果如表4所示。
表4多肽化合物对GHSR的活性(IC 50)
Figure PCTCN2022082973-appb-000034
Figure PCTCN2022082973-appb-000035
从表4的结果可知，在化合物1-80的五肽化合物的C端添加额外的氨基酸不会显著影响其GHSR激动活性。
以上实施例的说明只是用于帮助理解本发明的方法及其核心思想。应当指出，对于本技术领域的普通技术人员来说，在不脱离本发明原理的前提下，还可以对本发明进行若干改进和修饰，这些改进和修饰也落入本发明权利要求的保护范围内。

Claims (10)

  1. 一种多肽化合物，具有式Ⅰ所示结构，或其立体异构体、混合物、药学上可接受的盐：
    Figure PCTCN2022082973-appb-100001
    其中，R 1选自-NR 2R 3，-OR 2或-SR 2；
    并且，R 1不为D型或L型氨基酸；
    R 2和R 3独立地选自氢，氘，衍生自聚乙二醇的聚合物，非环状取代的或未经取代的脂族基团，取代的或未经取代的脂环基，取代的或未经取代的杂环基，取代的或未经取代的杂芳基烷基，取代的或未经取代的芳基，取代的或未经取代的芳烷基；
    W选自单键、D型氨基酸或L型氨基酸；
    U 1选自以下任一结构：
    Figure PCTCN2022082973-appb-100002
    其中，X和Z独立地选自CH-R 4，N-R 4，O，S，Se，S＝O或O＝S＝O；
    R 4，R 7和R 8独立地选自氢，氘，氨基、保护基，衍生自聚乙二醇的聚合物，非环状取代的或未经取代的脂族基团，取代的或未经取代的脂环基，取代的或未经取代的杂环基，取代的或未经取代的杂芳基烷基，取代的或未经取代 的芳基，取代的或未经取代的芳烷基或R 9CO-；
    Y选自卤素、氨基、硝基、羟基或氰基；
    R 5选自-NR 2R 3，-OR 2或-SR 2；
    R 6选自氢，氘，非环状取代的或未经取代的脂族基团，取代的或未经取代的脂环基，取代的或未经取代的杂环基，取代的或未经取代的杂芳基烷基，取代的或未经取代的芳基，取代的或未经取代的芳烷基；
    m 1和m 2独立地选自0，1，2或3；特别地，当X为N时，m 1为2，m 2则独立地选自0，1，3；m 2为2，m 1则独立地选自0，1，3；
    m 3和m 4独立地选自0，1，2或3；
    n 1，n 2，n 3和n 4独立地选自0，1，2或3；
    p为0，1，2，3，4或5；
    U 2为单键，或选自以下任一结构，且U 2的羰基端与W连接：
    Figure PCTCN2022082973-appb-100003
    R 9选自氢，非环状取代的或未经取代的脂族基团，取代的或未经取代的脂环基，取代的或未经取代的杂环基，取代的或未经取代的杂芳基烷基，取代的或未经取代的芳基，取代的或未经取代的芳烷基。
  2. 根据权利要求1所述的多肽化合物，其特征在于，所述R 7和R 8选自氢，C 1-6烷基，C 6-14芳基，C 3-8环烷基或C 2-10酰基；
    所述R 1选自-NR 2R 3或-OR 2，其中R 2和R 3独立地选自氢，甲基，乙基，己基，十二烷基或十六烷基。
  3. 根据权利要求1所述的多肽化合物，其特征在于，所述W选自单键，丙氨酸，精氨酸，天冬酰胺，半胱氨酸，谷氨酰胺，天冬氨酸，谷氨酸，甘氨酸，组氨酸，异亮氨酸，亮氨酸，赖氨酸，甲硫氨酸，苯丙氨酸，脯氨酸，丝氨酸，苏氨酸，色氨酸，酪氨酸，缬氨酸残基中的一种或多种。
  4. 根据权利要求1所述的多肽化合物，其特征在于，所述U 1选自以下结 构：
    Figure PCTCN2022082973-appb-100004
    其中，R 6选自氢，取代或非取代的C 1-6烷基，取代或非取代的C 6-14芳基，取代或非取代的C 3-8环烷基；所述取代的基团选自卤素、氨基、硝基、羟基、酰基取代的氨基、脲基或胍基；
    R 7和R 8独立地选自氢，C 1-6烷基，C 6-14芳基，C 3-8环烷基，C 2-10酰基；
    p为0，1，2，3，4或5。
  5. 根据权利要求4所述的多肽化合物，其特征在于，所述R 6选自氢，取代或未取代的甲基、乙基、丙基、异丙基、丁基、异丁基或叔丁基；
    所述取代的基团选自卤素、氨基、硝基、羟基、甲酰胺基、乙酰胺基、丙酰胺基、丁酰胺基、脲基或胍基；
    R 7和R 8独立地选自氢、甲基、乙基、丙基、异丙基、丁基、异丁基、叔丁基、甲酰基、乙酰基、丙酰基或丁酰基；
    p为0，1，2，3，4或5。
  6. 根据权利要求1所述的多肽化合物，其特征在于，所述U 1选自以下任一结构：
    Figure PCTCN2022082973-appb-100005
    Figure PCTCN2022082973-appb-100006
    其中，R 10、R 11独立地选自氢、氨基、硝基、羟基、卤素、氰基、胺甲基、胺乙基、胺丙基或胺丁基。
  7. 根据权利要求1所述的多肽化合物，其特征在于，具有以下任一结构，或其立体异构体、混合物、药学上可接受的盐：
    Figure PCTCN2022082973-appb-100007
    Figure PCTCN2022082973-appb-100008
    Figure PCTCN2022082973-appb-100009
    Figure PCTCN2022082973-appb-100010
    Figure PCTCN2022082973-appb-100011
    Figure PCTCN2022082973-appb-100012
  8. 一种组合物，包括权利要求1～7任一项所述的多肽化合物，和可接受的辅剂。
  9. 权利要求1～7任一项所述的多肽化合物，或权利要求8所述的组合物，作为生长激素促分泌素受体的激动剂的应用，或在制备用于治疗、预防、减轻和/或诊断由生长激素促分泌素受体介导的紊乱所致相关疾病的药物，或作为促进生长发育的保健品中的应用。
  10. 根据权利要求9所述的应用，其特征在于，所述相关疾病为生长激素缺乏症。
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