Novel LRP1-binding peptide

Abstract

To provide a novel peptide that binds to LRP1.SOLUTION: The present invention provides a linear peptide or a cyclic peptide having an amino acid sequence represented by the following formula (1): XN-X5-X6-X7-X8-X9-X10-X11-X12-X13-X14-X15 (1), or a pharmaceutically acceptable salt thereof.SELECTED DRAWING: Figure 6

Description

Cross-reference

[0001] This application claims priority based on Japanese Patent Application No. 2022-125238 filed in Japan on August 5, 2022, and all of the contents described in the application are hereby incorporated herein by reference in their entirety.

Technical Field

[0002] The present invention relates to a peptide that binds to LRP1 and the like. The present invention relates to a peptide or the like that can pass through a barrier tissue that separates peripheral tissue and brain tissue, such as the blood-brain barrier (BBB), through binding to LRP1. Specifically, it relates to a linear peptide or cyclic peptide having a specific amino acid sequence or the like.

Background Art

[0003] Central nervous system diseases (mental diseases) such as dementia, schizophrenia, and autism spectrum disorder are said to affect up to 1 billion people worldwide, accounting for the largest number of disability-adjusted life years, so the total social burden is enormous. For example, the economic burden in the United States and Europe in 2030 is estimated to reach $6 trillion combined (Non-Patent Document 1). Against this backdrop, drug discovery for central nervous system diseases is expected to lead to a huge market if successful, but it is also known to be the drug discovery area with the highest failure rate (Non-Patent Document 1). One of the reasons is the existence of barrier tissues that separate peripheral tissues from brain tissues in the body: the blood-brain barrier, the blood-cerebrospinal fluid barrier, the blood-spinal cord barrier, and the blood-arachnoid barrier. Among these, the blood-brain barrier (BBB) in particular is an obstacle to drug discovery for central nervous system diseases. The blood-brain barrier is a physical barrier formed by pericytes and further astrocytes covering the periphery of brain capillary endothelial cells connected by tight junctions, and strictly controls the influx and efflux of substances between peripheral tissues and brain tissues (Non-Patent Document 2). For example, substances that migrate from peripheral tissues to brain tissues by passive diffusion are known to preferably have a molecular weight of 400 g / mol or less and high lipophilicity. Therefore, many drug discovery attempts for central nervous system diseases have been made with low-molecular compounds, but there are concerns about side effects due to off-target effects, and the drug discovery targets are also limited.

[0004] In particular, for drug discovery targets such as protein-protein interactions that have attracted attention in recent years, there are few clear pockets to which conventional low-molecular compounds can bind, and medium-molecules such as peptides and high-molecules such as antibodies are superior in their control. In addition, since medium-molecules and high-molecules have high specificity for drug discovery targets, a reduction in side effects due to off-target effects can also be expected. However, as described above, due to the existence of barrier tissues, it is a problem that it is extremely difficult for medium-molecules and high-molecules to migrate from peripheral tissues to brain tissues by passive diffusion.

[0005] So far, as one of the methods for transferring medium- and high-molecular substances from peripheral tissues to brain tissues, receptor-mediated transcytosis (RMT) via receptors expressed in the blood-brain barrier has been studied to transfer specific nutrients and essential components to the brain. For example, transporters using glucose and amino acids as substrates, receptors for insulin, transferrin, and lipoproteins can be mentioned (Non-Patent Document 2). Regarding the attempt to conjugate an antibody that binds to the human transferrin receptor with a drug efficacy molecule and transport the drug efficacy molecule to the central tissue, there is already a marketed drug, and the concept of RMT has been demonstrated in human clinical trials (Non-Patent Document 3).

[0006] Low density lipoprotein receptor-related protein 1 (LRP1) is also one of the receptors for which RMT is expected. The expression level of LRP1 in the blood-brain barrier is well correlated among humans, monkeys, and mice. In addition, the amino acid sequences of cluster 2 (CL2) and cluster 4 (CL4), which are ligand-binding domains that interact with various endogenous ligands, are extremely highly conserved among species. And a molecule in which paclitaxel, an anticancer drug, is conjugated to Angiopep-2 (ANG2), known as an LRP1-binding peptide, has completed the second phase of a human clinical trial, and the safety of RMT by LRP1 in humans has been demonstrated (Non-Patent Document 4).

Prior Art Documents

Non-Patent Documents

[0007]

Non-Patent Document 1

Non-Patent Document 2

Non - Patent Document 3

Non - Patent Document 4

Non - Patent Document 5

Non - Patent Document 6

Non - Patent Document 7

Non - Patent Document 8

Non - Patent Document 9

Summary of the Invention

Problems to be Solved by the Invention

[0008] The invention described herein aims to provide a novel peptide that binds to LRP1. Furthermore, by combining this peptide with "a molecule" and transporting "a molecule" to the brain tissue via RMT by LRP1, it aims to provide a method for using "a molecule" as a pharmaceutical, diagnostic, and / or research reagent.

Means for Solving the Problems

[0009] The inventor searched for a molecule that binds to LRP1 and completed the present invention. That is, one aspect of the present invention is represented by the following formula (1): X N -X 5 -X 6 -X 7 -X 8 -X 9 -X 10 -X 11 -X 12 -X 13 -X 14 -X 15 a linear peptide or cyclic peptide having the amino acid sequence represented by (1), or a pharmaceutically acceptable salt thereof.

[0010] Here, in formula (1), X N is 1 to 4 arbitrary amino acid residues, and X 5 and X 15 each independently represent serine, threonine, cysteine, D-cysteine, or proline, and X 6 represents serine, homoserine, threonine, allothreonine, 2,3-diaminopropionic acid, 2,4-diaminobutyric acid, ornithine, lysine, arginine, proline, cis-4-hydroxy-proline, or trans-4-hydroxy-proline, and X7 represents histidine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, and X 8 represents ornithine, lysine, homolysine, arginine, or homoarginine, and X 9 represents methionine, norleucine, lysine, arginine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, and X 10 represents methionine, leucine, norleucine, isoleucine, valine, lysine, or argin, and X 11 and X 14 each independently represents methionine, leucine, norleucine, isoleucine, valine, 2-aminoheptanoic acid, or 2-aminooctanoic acid, and X 12 represents alanine, D-alanine, or 2-aminoisobutyric acid, and X 13 represents glycine, alanine, asparagine, aspartic acid, glutamine, or glutamic acid, and the N-terminal amino group and the C-terminal carboxy group may be modified or deleted, and X 5 and X 15 is cysteine or D-cysteine when involved in cyclization, and a covalent bond is formed between the side-chain -SH groups thereof via a linker of a disulfide bond, a methylene group, an acetylmethylene group, an ethylene group, or a propylene group, whereby the peptide of formula (1) may have one cyclic structure in the molecule.

[0011] X in the peptide of the above formula (1) 1 from X 15 The preferred embodiments of each amino acid residue of are described in detail in the following embodiments, and each preferred embodiment can be arbitrarily combined independently of each other.

[0012] The present invention further specifically includes the following embodiments. [1] The above formula (1): XN -X 5 -X 6 -X 7 -X 8 -X 9 -X 10 -X 11 -X 12 -X 13 -X 14 -X 15 (1) (wherein X N is any one to four amino acid residues, and X 5 and X 15 each independently represent serine, threonine, cysteine, D-cysteine, or proline, and X 6 represents serine, homoserine, threonine, allothreonine, 2,3-diaminopropionic acid, 2,4-diaminobutyric acid, ornithine, lysine, arginine, proline, cis-4-hydroxy-proline, or trans-4-hydroxy-proline, and X 7 represents histidine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, and X 8 represents ornithine, lysine, homolysine, arginine, or homoarginine, and X 9 represents methionine, norleucine, lysine, arginine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, and X 10 represents methionine, leucine, norleucine, isoleucine, valine, lysine, or argin, and X 11 and X 14 each independently represent methionine, leucine, norleucine, isoleucine, valine, 2-aminoheptanoic acid, or 2-aminooctanoic acid, and X 12 represents alanine, D-alanine, or 2-aminoisobutyric acid, and X 13 represents glycine, alanine, asparagine, aspartic acid, glutamine, or glutamic acid, and the N-terminal amino group and the C-terminal carboxy group may be modified or deleted, and X 5and X 15 When involved in cyclization, it is cysteine or D-cysteine, and a covalent bond is formed between the -SH groups of their respective side chains via a linker of a disulfide bond, a methylene group, an acetylmethylene group, an ethylene group, or a propylene group, whereby the peptide of formula (1) may have one cyclic structure in the molecule.) A linear peptide or cyclic peptide having the amino acid sequence represented by, or a pharmacologically acceptable salt thereof. [2] The following formula (3): X N -X 5 -X 6 -X 7 -X 8 -X 9 -X 10 -X 11 -X 12 -X 13 -X 14 -Pro-X 16 (3) (In the formula, X N is 1 to 4 arbitrary amino acid residues, X 5 and X 16 each independently represent serine, threonine, cysteine, D-cysteine, or proline, X 6 represents serine, homoserine, threonine, allothreonine, 2,3-diaminopropionic acid, 2,4-diaminobutyric acid, ornithine, lysine, arginine, proline, cis-4-hydroxy-proline, or trans-4-hydroxy-proline, X 7 represents histidine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, X 8 represents ornithine, lysine, homolysine, arginine, or homoarginine, X 9 represents methionine, norleucine, lysine, arginine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, X 10represents methionine, leucine, norleucine, isoleucine, valine, lysine, or arginine, X 11 and X 14 each independently represent methionine, leucine, norleucine, isoleucine, valine, 2-aminoheptanoic acid, or 2-aminooctanoic acid, X 12 represents alanine, D-alanine, or 2-aminoisobutyric acid, X 13 represents glycine, alanine, asparagine, aspartic acid, glutamine, or glutamic acid, and the N-terminal amino group and the C-terminal carboxy group may be modified or deleted, X 5 and X 16 is cysteine or D-cysteine when involved in cyclization, and a covalent bond is formed between the side-chain -SH groups of each through a linker of a disulfide bond, a methylene group, an acetylmethylene group, an ethylene group, or a propylene group, whereby the peptide of formula (3) may have one cyclic structure in the molecule. A linear peptide or cyclic peptide having the amino acid sequence represented by) or a pharmaceutically acceptable salt thereof. [3] The following formula (3): X N -X 5 -X 6 -X 7 -X 8 -X 9 -X 10 -X 11 -X 12 -X 13 -X 14 -Pro-X 16 (3) (In the formula, X N is 1 to 4 arbitrary amino acid residues, X 6 and X 16 each independently represent serine, threonine, cysteine, D-cysteine, or proline, X 5 represents serine, homoserine, threonine, allothreonine, 2,3-diaminopropionic acid, 2,4-diaminobutyric acid, ornithine, lysine, arginine, proline, cis-4-hydroxy-proline, or trans-4-hydroxy-proline, X 7represents histidine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, X 8 represents ornithine, lysine, homolysine, arginine, or homoarginine, X 9 represents methionine, norleucine, lysine, arginine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, X 10 represents methionine, leucine, norleucine, isoleucine, valine, lysine, or argin, X 11 and X 14 each independently represents methionine, leucine, norleucine, isoleucine, valine, 2-aminoheptanoic acid, or 2-aminooctanoic acid, X 12 represents alanine, D-alanine, or 2-aminoisobutyric acid, X 13 represents glycine, alanine, asparagine, aspartic acid, glutamine, or glutamic acid, and the N-terminal amino group and C-terminal carboxy group may be modified or deleted, X 6 and X 16 is cysteine or D-cysteine when involved in cyclization, and a covalent bond is formed between the side-chain -SH groups thereof via a linker of a disulfide bond, a methylene group, an acetylmethylene group, an ethylene group, or a propylene group, whereby the peptide of formula (3) may have one cyclic structure in the molecule. A linear peptide or cyclic peptide having an amino acid sequence represented by), or a pharmaceutically acceptable salt thereof. [4] X N is Lys-Gly-Thr-Pro or Gly-Thr-Pro, the linear peptide or cyclic peptide according to any one of [1] to [3], or a pharmaceutically acceptable salt thereof. [5] A linear or cyclic peptide having an LRP1 binding activity, comprising an amino acid sequence in which one or several amino acids are deleted, added, and / or substituted in the amino acid sequence according to any one of [1] to [4], or a pharmaceutically acceptable salt thereof. 〔6〕 A derivative and / or modified form of the peptide according to any one of [1] to [5], and the following

[14] to

[21] . 〔7〕 The derivative or modified form according to [6], wherein the derivative or modified form has an azide group, an alkyne group, a cyclooctyne group, or a tetrazine group directly or via a linker at the N-terminus, C-terminus, or side chain of an amino acid. 〔8〕 A medicine, diagnostic agent, and / or reagent comprising the peptide according to any one of [1] to [7], and the following

[14] to

[21] , or a derivative or modified form thereof. 〔9〕 A peptide-drug conjugate, wherein the derivative or modified form of the peptide according to [6] is linked directly or via a linker to a drug. 〔10〕 The peptide-drug conjugate according to [9], wherein the drug is an antibody, a nucleic acid, or a bioactive peptide. 〔11〕 A polynucleotide encoding an amino acid sequence consisting only of natural amino acids among the amino acid sequences according to any one of [1] to [5], and the following

[14] to

[21] , or a polynucleotide having a base sequence with 70% or more sequence identity to the polynucleotide and encoding a peptide having an LRP1 binding activity. 〔12〕 An expression vector containing the polynucleotide according to

[11] . 〔13〕 A cell, yeast, bacterium, virus, liposome, or nanoparticle presenting the amino acid sequence according to any one of [1] to [5], and the following

[14] to

[21] . 〔14〕 The following formula (6): X 1 -X 2 -X 3 -X 4-X L -X 7 -X 8 -X 9 -X 10 -X 11 -X 12 -X 13 -X 14 -X M (6) (X L is X 5 -X 6 and X M is X 15 -X 16 and, when involved in cyclization within the peptide molecule, each independently represents an amino acid residue having an amino group, a carboxy group, a thiol group, an allyl group, an alkynyl group, an azide group, or a halogen atom, or a derivative thereof; when not involved in cyclization within the peptide molecule, X 5 X 6 or X 15 each independently represents an arbitrary amino acid residue, or a derivative thereof, and X 16 represents an arbitrary amino acid residue, a derivative thereof, or a deletion, and X 7 represents an amino acid residue having a hydrocarbon group which may have a substituent, an amino acid residue having an aromatic carbocyclic group which may have a substituent, an amino acid residue having an aromatic heterocyclic group which may have a substituent, or a derivative thereof, and X 8 represents an amino acid residue having a positive charge, or a derivative thereof, and X 11 X 12 and X 14 each independently represents an amino acid residue having a hydrocarbon group which may have a substituent, or a derivative thereof, and X 13 represents an amino acid residue having a hydrocarbon group which may have a substituent, an amino acid residue having a negative charge, or a derivative thereof, and X 9 X 10 each independently represents an arbitrary amino acid residue, or a derivative thereof, and X 1 X 2 X 3 X 4 each independently represents an arbitrary amino acid residue, a derivative thereof, or a deletion, and X L and XM Between them, the peptide of formula (6) may have one cyclic structure in the molecule by forming a direct or indirect covalent bond via a linker, and in that case, X L and X M The covalent bond between them may be any of their main chain-main chain, main chain-side chain, side chain-main chain, or side chain-side chain bonds, and the N-terminal amino group and C-terminal carboxy group may be modified or deleted.) A linear peptide and a cyclic peptide consisting of the amino acid sequence represented by, or a pharmacologically acceptable salt thereof.

[15] X 5 and X 15 The bond between the Cα carbon atoms of, X 6 and X 15 The bond between the Cα carbon atoms of, X 5 and X 16 The bond between the Cα carbon atoms of, or X 6 and X 16 The bond between the Cα carbon atoms of are each independently the following formula (7): Cα L -(CH2) A -Z-(CH2) B -Cα M (7) (wherein L is 4 or 5, M is 14 or 15, A and B each independently represent an integer of any one of 0 to 2, the sum of A and B is an integer of any one of 0 to 4, and Z is S-S, CH2-CH2, S-CH2, CH2-S, O-CH2, CH2-O, CH=CH, NH-C(=O), N(CH3)-C(=O), NH-C(=S), N(CH3)-C(=S), O-C(=O), C(=O)-NH, C(=O)-N(CH3), C(=S)-NH, C(=S)-N(CH3), C(=O)-O, CH2-CH2-CH2, S-CH2-CH2, CH2-CH2-S, CH2-S-CH2, O-CH2-CH2, CH2-CH2-O, CH2-O-CH2, S-CH2-S, S-C(=CH2)-S, S-(C(=O)-CH3)-S, S-(CH2)2-S, S-(CH2)3-S, S-CH2-C(=O)-CH2-S, S-CH2-C(=CH2)-CH2-S, S-(CH2)4-S, S-CH2-CH=CH-CH2-S, S-CH2-C6H4-CH2-S (the bonding site of the methylene group to the phenylene ring may be any of ortho, meta, and para), S-CH2-C(=O)-CH2-S, or S-CH2-C(=CH2)-CH2-S.) The linear peptide and cyclic peptide according to

[14] containing the structure represented by , or a pharmacologically acceptable salt thereof.

[16] X 8 The linear peptide and cyclic peptide according to claim

[14] or

[15] , or a pharmacologically acceptable salt thereof, wherein X 8 is 2,3-diaminopropionic acid, 2,4-diaminobutyric acid, ornithine, lysine, homolysine, arginine, homoarginine, their D-form amino acids, their N-methylated amino acids, their α-methylated amino acids, or derivatives thereof.

[17] X 12is a linear peptide and a cyclic peptide according to any one of

[14] to

[16] , which is alanine, D-alanine, 2-aminoisobutyric acid, 2-aminobutyric acid, D-2-aminobutyric acid, isovaline, D-isovaline, valine, D-valine, isoleucine, D-isoleucine, leucine, D-leucine, their N-methylated amino acids, their α-methylated amino acids, or their derivatives, or a pharmacologically acceptable salt thereof.

[18] X 11 and X 14 are each independently methionine, the following formula (8):

Chemical formula

Chemical formula

Chemical formula

[14] to

[17] , which is an amino acid residue represented by or a derivative thereof, or a pharmacologically acceptable salt thereof.

[19] X 7 is the following formula (8):

Chemical formula

Chemical formula

Chemical formula

Chemical formula

Chemical formula

Chemical formula

[14] to

[18] , or a pharmaceutically acceptable salt thereof.

[20] X 13which is a linear peptide and a cyclic peptide according to any one of

[14] to

[19] , or a pharmacologically acceptable salt thereof, said peptide being glycine, alanine, aspartic acid, glutamic acid, asparagine, glutamine, D-alanine, D-aspartic acid, D-glutamic acid, D-asparagine, D-glutamine, their N-methylated amino acids, their α-methylated amino acids, or their derivatives.

[21] X which is not involved in cyclization within the peptide molecule L and X M each independently being alanine, isovaline, norvaline, serine, homoserine, threonine, allothreonine, O-methylated-serine, O-methylated-homoserine, O-methylated-threonine, O-methylated-allothreonine, 2,3-diaminopropionic acid, 2,4-diaminobutyric acid, ornithine, lysine, arginine, proline, cis-4-hydroxy-proline, trans-4-hydroxy-proline, azetidine-2-carboxylic acid, pipecolic acid, D-serine, D-homoserine, D-threonine, D-allothreonine, D-O-methylated-serine, D-O-methylated-homoserine, D-O-methylated-threonine, D-O-methylated-allothreonine, D-2,3-diaminopropionic acid, D-2,4-diaminobutyric acid, D-ornithine, D-lysine, D-arginine, D-proline, D-cis-4-hydroxy-proline, D-trans-4-hydroxy-proline, D-azetidine-2-carboxylic acid, D-pipecolic acid, their N-methylated amino acids, their α-methylated amino acids, or their derivatives, being a linear peptide and a cyclic peptide according to any one of

[14] to

[20] , or a pharmacologically acceptable salt thereof.

Advantages of the Invention

[0013] According to the present invention, there are provided useful and novel linear peptides and cyclic peptides for use as pharmaceuticals, diagnostic agents, and / or research reagents by transferring "a certain molecule" combined with the present peptide into brain tissue by RMT via binding to LRP1.

Brief Description of the Drawings

[0014]

Figure 1

Figure 2

Figure 3

Figure 4

Figure 5

Figure 6

Figure 7

Figure 8

BEST MODE FOR CARRYING OUT THE INVENTION

[0015] Next, each embodiment of the present invention will be described with reference to the drawings. It should be noted that each embodiment described below does not limit the invention according to the claims, and not all of the elements and combinations thereof described in each embodiment are essential for the solution means of the present invention.

[0016] (Definition) As used herein, a peptide refers to a compound in which two or more amino acids are linked by an amide bond (peptide bond), and can be, for example, a compound in which 2 to 20 amino acids are amide-bonded. Also, according to the convention of peptide labeling, the left end is the N-terminus (amino terminus) and the right end is the C-terminus (carboxy terminus). The first carbon atom adjacent to the carbonyl group forming the peptide bond is referred to as the Cα carbon atom.

[0017] As used herein, "any amino acid residue, or derivatives thereof" is used in its broadest sense and includes, in addition to natural amino acids, artificial amino acids having an unnatural structure, chemically synthesized compounds having properties known in the art that are characteristic of amino acids, and carboxylic acids having a functional group. Examples of unnatural amino acids include D-amino acids, α / α-disubstituted amino acids (such as α-methylated amino acids like 2-aminoisobutyric acid) with a main-chain structure different from the natural form, N-alkyl-amino acids (such as N-methylated amino acids), N-substituted glycines (peptoids), amino acids with an extended main chain (β-homoamino acids and γ-homoamino acids), amino acids with a side-chain structure different from the natural form (such as cyclohexylalanine, allylglycine, 2-(2-pyridyl)-glycine, 3-(1H-benzimidazol-2-yl)-alanine, etc.), amino acids with a partially substituted side chain (such as norleucine, diaminopropanoic acid, 3-(2-pyridyl)-alanine, etc.), amino acids with an extra functional group in the side chain; amino acids with an extra C, alkyl group, or methyl group in the side chain (such as homonorleucine, γ-methylleucine, etc.), amino acids with a halogen atom (F, Cl, Br, I) in the side chain (such as 3-chloro-alanine, etc.), carboxylic acids with a halogen atom (F, Cl, Br, I) in the side chain (such as 3-chloropropanoic acid, etc.), carboxylic acids with a functional group in the side chain (such as 3-butenoic acid, etc.), amino acids with an extra N or amino group in the side chain (such as β-azidoalanine, ornithine, etc.), amino acids with an extra O or methoxy group in the side chain (such as O-methyl-serine, O-methyl-threonine, etc.), amino acids with an extra hydroxy group in the side chain (such as 3-hydroxy-phenylalanine, etc.), amino acids with an extra carboxy group (-COOH) in the side chain (such as 3-carboxy-phenylalanine, etc.), amino acids with an extra S in the side chain (such as ethionine, etc.), amino acids with a carboxylic acid functional group in the side chain protected by an ester (such as aspartic acid-4-methyl ester, etc.), amino acids with a thio group (-S-) in the side chain oxidized to a sulfinyl group (-S(=O)-) or a sulfonyl group (-S(=O)2-) (such as methionine sulfoxide), etc., but are not limited thereto.

[0018] As used herein, "LRP1" refers to LRP1 of mammals such as mice, rats, dogs, monkeys, and humans.

[0019] As used herein, when it is said that "having LRP1 binding activity", in an in vitro test, the peptide of the present invention or a combination of the peptide of the present invention and "a certain molecule" binds in a concentration-dependent manner to the full-length protein of LRP1, a partial protein such as cluster 4, or a chimeric form of these and other proteins, etc. The presence or absence of LRP1 binding activity can be confirmed by those skilled in the art according to known methods with reference to Non-Patent Document 5 and the like, but is not limited thereto.

[0020] As used herein, "blood-brain barrier tissue" refers to blood-brain barrier tissue of mammals such as mice, rats, dogs, monkeys, and humans.

[0021] As used herein, when it is said that "passing through the blood-brain barrier tissue", (1) in a test using an in vitro blood-brain barrier model, the peptide of the present invention or a combination of the peptide of the present invention and "a certain molecule" migrates from the vascular side well to the brain side well, (2) in a pharmacokinetic test using LC-MS / MS or the like, the peptide of the present invention or a combination of the peptide of the present invention and "a certain molecule" administered from the periphery is detected in the brain tissue, (3) in a pharmacokinetic test using an in vivo imaging system or the like, the peptide of the present invention or a combination of the peptide of the present invention and "a certain molecule" administered from the periphery is detected in the brain, etc., and when any one of these effects is shown, it is said that "passing through the blood-brain barrier tissue". The presence or absence of the ability to pass through the blood-brain barrier tissue can be confirmed by those skilled in the art according to known methods with reference to Non-Patent Documents 6 and 7 and the like, but is not limited thereto.

[0022] (The peptide of the present invention) [1] The linear peptide and cyclic peptide in one embodiment of the present disclosure have the following formula (1): X N -X 5 -X 6 -X 7 -X8 -X 9 -X 10 -X 11 -X 12 -X 13 -X 14 -X 15 (1) has an amino acid sequence represented by In formula (1), X N is any one to four amino acid residues, and X 5 and X 15 each independently represent serine, threonine, cysteine, D-cysteine, or proline, and X 6 represents serine, homoserine, threonine, allothreonine, 2,3-diaminopropionic acid, 2,4-diaminobutyric acid, ornithine, lysine, arginine, proline, cis-4-hydroxy-proline, or trans-4-hydroxy-proline, and X 7 represents histidine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, and X 8 represents ornithine, lysine, homolysine, arginine, or homoarginine, and X 9 represents methionine, norleucine, lysine, arginine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, and X 10 represents methionine, leucine, norleucine, isoleucine, valine, lysine, or argin, and X 11 and X 14 each independently represent methionine, leucine, norleucine, isoleucine, valine, 2-aminoheptanoic acid, or 2-aminooctanoic acid, and X 12 represents alanine, D-alanine, or 2-aminoisobutyric acid, and X 13 represents glycine, alanine, asparagine, aspartic acid, glutamine, or glutamic acid. The N-terminal amino group and the C-terminal carboxy group may be modified or deleted, and X 5 and X 15When involved in cyclization, it is cysteine or D-cysteine, and a covalent bond is formed between the -SH groups of their respective side chains via a linker of a disulfide bond, a methylene group, an acetylmethylene group, an ethylene group, or a propylene group, whereby the peptide of formula (1) may have one cyclic structure in the molecule.

[0023] In the peptide of formula (1) above, X N is preferably Lys-Gly-Thr-Pro or Gly-Thr-Pro. When the N-terminus is lysine, it may be bound to a molecule via the ε-amino group of its side chain. When the N-terminus is glycine, it may be bound to a molecule via its α-amino group. X 5 and X 15 are cysteine or D-cysteine, and a covalent bond is formed between the -SH groups of their respective side chains via a disulfide bond or a linker of a methylene group, whereby it is preferable to form one cyclic structure in the molecule.

[0024] As a more preferred embodiment in the peptide of formula (1) above, for example, the amino acid sequence represented by the following formula (2) can be mentioned. A covalent bond is formed between the -SH groups of their respective side chains via a disulfide bond or a linker of a methylene group, whereby it may have one cyclic structure in the molecule, and it may also be bound to a molecule via the ε-amino group of Lys1.

[0025] Ac-Lys-Gly-Thr-Pro-Cys-X 6 -X 7 -Lys-X 9 -X 10 -Leu-Ala-Glu-X 14 -Cys-OH (2) In formula (2), X 6 represents serine, threonine, or 2,3-diaminopropionic acid, X 7 represents histidine, tyrosine, phenylalanine, or tryptophan, X 9 represents tyrosine, methionine, norleucine, or arginine, X10 represents methionine, norleucine, arginine, or lysine, and X 14 represents leucine, isoleucine, methionine, or norleucine.

[0026] [2] The linear peptides and cyclic peptides in other embodiments of the present disclosure have the following formula (3): X N -X 5 -X 6 -X 7 -X 8 -X 9 -X 10 -X 11 -X 12 -X 13 -X 14 -Pro-X 16 (3) and has an amino acid sequence represented by In the above formula (3), X N is 1 to 4 arbitrary amino acid residues, and X 5 and X 16 each independently represent serine, threonine, cysteine, D-cysteine, or proline, and X 6 represents serine, homoserine, threonine, allothreonine, 2,3-diaminopropionic acid, 2,4-diaminobutyric acid, ornithine, lysine, arginine, proline, cis-4-hydroxy-proline, or trans-4-hydroxy-proline, and X 7 represents histidine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, and X 8 represents ornithine, lysine, homolysine, arginine, or homoarginine, and X 9 represents methionine, norleucine, lysine, arginine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, and X 10 represents methionine, leucine, norleucine, isoleucine, valine, lysine, or arginine, and X 11 and X14 each independently represents methionine, leucine, norleucine, isoleucine, valine, 2-aminoheptanoic acid, or 2-aminooctanoic acid, and X 12 represents alanine, D-alanine, or 2-aminoisobutyric acid, and X 13 represents glycine, alanine, asparagine, aspartic acid, glutamine, or glutamic acid. The N-terminal amino group and the C-terminal carboxy group may be modified or deleted, and X 5 and X 16 are cysteine or D-cysteine when involved in cyclization, and form a covalent bond via a linker of a disulfide bond, a methylene group, an acetylmethylene group, an ethylene group, or a propylene group between the respective side-chain -SH groups, whereby the peptide of formula (3) may have one cyclic structure in the molecule.

[0027] In the peptide of formula (3) above, X N is preferably Lys-Gly-Thr-Pro or Gly-Thr-Pro. When the N-terminus is lysine, it may be bound to a molecule via the ε-amino group of its side chain. When the N-terminus is glycine, it may be bound to a molecule via its α-amino group. X 5 and X 16 are cysteine or D-cysteine, and form a covalent bond via a disulfide bond or a linker of a methylene group between the respective side-chain -SH groups, whereby it is preferable to form one cyclic structure in the molecule.

[0028] As a more preferred embodiment in the peptide of formula (3) above, for example, the amino acid sequence represented by the following formula (4) can be mentioned. A covalent bond is formed via a disulfide bond or a linker of a methylene group between the respective side-chain -SH groups, whereby it may have one cyclic structure in the molecule, and it may be bound to a molecule via the ε-amino group of Lys1.

[0029] Ac-Lys-Gly-Thr-Pro-Cys-X 6 -X 7-Lys-X 9 -X 10 -Leu-Ala-Glu-X 14 -Pro-Cys-OH (4) In formula (4), X 6 represents serine, threonine, or 2,3-diaminopropionic acid, and X 7 represents histidine, tyrosine, phenylalanine, or tryptophan, and X 9 represents tyrosine, methionine, norleucine, or arginine, and X 10 represents methionine, norleucine, arginine, or lysine, and X 14 represents leucine, isoleucine, methionine, or norleucine.

[0030] [3] The linear peptides and cyclic peptides in another embodiment of the present disclosure are represented by the following formula (3): X N -X 5 -X 6 -X 7 -X 8 -X 9 -X 10 -X 11 -X 12 -X 13 -X 14 -Pro-X 16 (3) and have an amino acid sequence represented thereby. In the above formula (3), X N is any one to four amino acid residues, and X 6 and X 16 each independently represent serine, threonine, cysteine, D-cysteine, or proline, and X 5 represents serine, homoserine, threonine, allothreonine, 2,3-diaminopropionic acid, 2,4-diaminobutyric acid, ornithine, lysine, arginine, proline, cis-4-hydroxy-proline, or trans-4-hydroxy-proline, and X 7 represents histidine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine,8 represents ornithine, lysine, homolysine, arginine, or homoarginine, and X 9 represents methionine, norleucine, lysine, arginine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, and X 10 represents methionine, leucine, norleucine, isoleucine, valine, lysine, or argin, and X 11 and X 14 each independently represents methionine, leucine, norleucine, isoleucine, valine, 2-aminoheptanoic acid, or 2-aminooctanoic acid, and X 12 represents alanine, D-alanine, or 2-aminoisobutyric acid, and X 13 represents glycine, alanine, asparagine, aspartic acid, glutamine, or glutamic acid, and the N-terminal amino group and the C-terminal carboxy group may be modified or deleted, and X 6 and X 16 is cysteine or D-cysteine when involved in cyclization, and a covalent bond is formed between the respective side-chain -SH groups via a linker of a disulfide bond, a methylene group, an acetylmethylene group, an ethylene group, or a propylene group, whereby the peptide of formula (3) may have one cyclic structure in the molecule.

[0031] In the peptide of the above formula (3), X N is preferably Lys-Gly-Thr-Pro or Gly-Thr-Pro. When the N-terminus is lysine, it may be bound to a molecule via the ε-amino group of its side chain. When the N-terminus is glycine, it may be bound to a molecule via the α-amino group of it. X 6 and X 16 are cysteine or D-cysteine, and a covalent bond is formed between the respective side-chain -SH groups via a disulfide bond or a linker of a methylene group, whereby it is preferable to form one cyclic structure in the molecule.

[0032] In a more preferred embodiment of the peptide of the above formula (3), for example, the amino acid sequence represented by the following formula (5) can be mentioned. A disulfide bond or a covalent bond through a methylene group linker may be formed between the side chain -SH groups, and thereby it may have one cyclic structure in the molecule, and it may be bound to a molecule through the ε amino group of Lys1.

[0033] Ac-Lys-Gly-Thr-Pro-X 5 -Cys-X 7 -Lys-X 9 -X 10 -Leu-Ala-Glu-X 14 -Pro-Cys-OH (5) In formula (5), X 5 represents serine, threonine, or 2,3-diaminopropionic acid, and X 7 represents histidine, tyrosine, phenylalanine, or tryptophan, and X 9 represents tyrosine, methionine, norleucine, or arginine, and X 10 represents methionine, norleucine, arginine, or lysine, and X 14 represents leucine, isoleucine, methionine, or norleucine.

[0034] [4] The peptide according to this embodiment includes a peptide having homology in which 1 to several amino acids are deleted, added, and / or substituted in the amino acid sequence represented by the above [1] to [3], as long as it has binding activity to LRP1. In this specification, in the case of "a peptide in which 1 to several amino acids are deleted, added, and / or substituted", the number of those amino acids is not particularly limited as long as the peptide has LRP1 binding activity, but is preferably 1 to 5, more preferably 1 or 2. The location where deletion, addition, and / or substitution occur may be at the end of the peptide or in the middle, and may be at one location or at two or more locations.

[0035] As an amino acid sequence in which one to several amino acids are deleted, added, and / or substituted in the above amino acid sequence, when calculated using the above amino acid sequence and BLAST (Basic Local Alignment Search Tool at the National Center for Biological Information) or the like (for example, using default or initial parameters), those having an identity of at least 50% or more, preferably 70% or more, more preferably 80% or more, and particularly preferably 90% or more are exemplified.

[0036] It is also known that there are peptides and protein domains with highly similar three-dimensional structures even if they have low primary structure homology. Therefore, peptides having a three-dimensional structure homology of at least 50% or more, preferably 70% or more, more preferably 80% or more, and particularly preferably 90% or more with the above amino acid sequence and having Ras binding activity are also included in this embodiment. The homology of the three-dimensional structures of such peptides can be predicted as follows from the amino acid sequence of a peptide with an unknown three-dimensional structure using a homology modeling method or the like. For example, when any amino acid sequence (target sequence) having a sequence similar to the cyclic peptide (reference peptide) of this embodiment is given, an alignment (a juxtaposed sequence) between the target sequence and the reference sequence is given. By using the alignment calculated by FASTA, PSI-BLAST, LIBRA, etc., the correspondence relationship between the amino acids of the target sequence and the reference sequence is determined. Based on this relationship, the three-dimensional coordinates of each amino acid on the target sequence are created from the three-dimensional coordinates of the reference peptide. In the construction of the three-dimensional coordinates, there may be structurally inappropriate gaps, collisions, or distortions between amino acid residues. Therefore, these structural distortions are eliminated by energy minimization calculations. Depending on the modeling software, in order to smoothly eliminate these structural distortions, some perform it not simultaneously for all atoms of the peptide but step by step. That is, first, it is performed for the α-carbon atoms forming the peptide backbone, then for the main chain atoms including the α-carbon atoms, and finally for the entire peptide including the side chain atoms. If an alignment for the target sequence is obtained in this way, its three-dimensional structure can be predicted and constructed. As an index of three-dimensional structure homology, it can be compared using, for example, RMSD (Root Mean Square Deviation), which is the difference in XYZ coordinates when optimally superimposed.

[0037] The peptide of this embodiment also includes salts of the peptide. As the salts of the peptide, salts with physiologically acceptable bases or acids are used. For example, addition salts of inorganic acids (hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, phosphoric acid, etc.), addition salts of organic acids (p-toluenesulfonic acid, methanesulfonic acid, oxalic acid, p-bromophenylsulfonic acid, carboxylic acid, succinic acid, citric acid, benzoic acid, acetic acid, etc.), inorganic bases (ammonium hydroxide, or alkali or alkaline earth metal hydroxides, carbonates, bicarbonates, etc.), addition salts of amino acids, and the like can be mentioned.

[0038] (Derivatives and modified forms of the peptide) The peptide of the present embodiment includes various derivatives and / or modified forms thereof as long as they solve the problems of the present invention. Such derivatives include those in which the saturated fatty chain of the peptide is replaced with an unsaturated fatty chain, those in which some of the atoms of the peptide are replaced with other atoms containing radioactive or non-radioactive isotope atoms, those in which the amide bond of the peptide is replaced with a thioamide bond (-NH-C(=S)-), those in which the amide bond of the peptide is replaced with an alkene (-C=C-), those in which the amide bond of the peptide is replaced with an alkyl (-C-C-), those in which the amide bond of the peptide is replaced with a hydroxyethylene (-C(-OH)-C-), those in which the amide bond of the peptide is replaced with an ester (-O-C(=O)-), those in which the amide bond of the peptide is replaced with an alkene (-C=C-), those in which the amide bond of the peptide is replaced with (-C-NH-), or those in which the amide bond of the peptide is replaced with (-C(=O)-C-), etc. Such modified forms include those in which the α-carbon of the peptide is disubstituted, those in which the amide bond of the peptide is N-alkylated, those in which some of the functional groups of the peptide are modified by halogenation, cyanation, nitration, oxidation, hydroxylation, amination, deamination, dehydrogenation, amidation, acetylation, methoxylation, prenylation, alkylation, etc. (for example, those in which some of the amino groups of the peptide are acetylated, alkylated, or deaminated, those in which some of the carboxy groups of the peptide are amides or esters, etc.), those in which the S of the peptide becomes sulfoxide S(=O) or sulfone S(=O)2, those in which the peptide is multimerized via a chemical linker, those in which the peptide is biotinylated, those in which the peptide is fluorescently labeled, those in which the peptide is luminescently labeled, and further those in which the peptide is fused or conjugated with an alkyl chain, polyethylene glycol, antibody, lectins, sugar chain, enzyme, peptide, peptoid, membrane-permeable peptide, low molecular compound, nucleic acid, or a molecule that induces ubiquitination of a protein, etc. directly or via a linker, but are not limited thereto.

[0039] According to some embodiments of the present disclosure, the peptide of the present invention contains an amino acid residue having an azide group at its N-terminus or C-terminus. Accordingly, a "certain molecule" having an alkyne group can be covalently bonded to the N-terminus or C-terminus of the peptide of the present invention by a "copper(I)-catalyzed alkyne-azide cycloaddition (CuAAC) reaction" (or abbreviated as "click" reaction). According to other embodiments of the present disclosure, an azide group is introduced by reacting a linker having an azide group and an N-hydroxysuccinimide (NHS) ester group with the α-amino group or ε-amino group of the peptide of the present invention. Accordingly, a "certain molecule" having an alkyne group can be covalently bonded to the peptide of the present invention by the CuAAC reaction. According to other embodiments of the present disclosure, the peptide of the present invention contains an amino acid residue having an alkyne group at its N-terminus or C-terminus. Thereby, a "certain molecule" having an azide group can be covalently bonded to the N-terminus or C-terminus of the peptide of the present invention by the CuAAC reaction. According to other embodiments of the present disclosure, an alkyne group is introduced by reacting a linker having an alkyne group and an NHS ester group with the α-amino group or ε-amino group of the peptide of the present invention. Accordingly, a "certain molecule" having an azide group can be covalently bonded to the peptide of the present invention by the CuAAC reaction. The CuAAC reaction is as shown in Scheme 1 below.

[0040] [Chemical Formula]

[0041] By the CuAAC reaction, 1,5-disubstituted-1,2,3-triazole is obtained. The reaction between an alkyne and an azide is very selective, and alkyne groups and azide groups do not exist in natural biomolecules. Furthermore, this reaction is rapid and pH-insensitive.

[0042] The above alkyne group may be replaced with a cyclooctyne group. According to some embodiments of the present disclosure, the peptide of the present invention contains an amino acid residue having an azide group at its N-terminus or C-terminus. Therefore, a "certain molecule" having a cyclooctyne group can be covalently bonded to the N-terminus or C-terminus of the peptide of the present invention by a "strain-promoted azide-alkyne click chemistry (SPAAC) reaction". According to other embodiments of the present disclosure, a linker having an azide group and an NHS ester group is reacted with the α-amino group or ε-amino group of the peptide of the present invention to introduce an azide group. Therefore, a "certain molecule" having a cyclooctyne group can be covalently bonded to the peptide of the present invention by a SPAAC reaction. According to other embodiments of the present disclosure, the peptide of the present invention contains an amino acid residue having a cyclooctyne group at its N-terminus or C-terminus. Thereby, a "certain molecule" having an azide group can be covalently bonded to the N-terminus or C-terminus of the peptide of the present invention by a SPAAC reaction. According to other embodiments of the present disclosure, a linker having a cyclooctyne group and an NHS ester group is reacted with the α-amino group or ε-amino group of the peptide of the present invention to introduce a cyclooctyne group. Therefore, a "certain molecule" having an azide group can be covalently bonded to the peptide of the present invention by a SPAAC reaction. The SPAAC reaction is as shown in Scheme 2 below.

[0043] [Chemical formula]

[0044] Alternatively, in certain embodiments, a "certain molecule" having a tetrazine group or a cyclooctyne group is bonded to the corresponding cyclooctyne group or tetrazine group of a peptide or linker by an "inverse electron demand Diels-Alder (iEDDA) reaction" as shown in Scheme 3 below.

[0045] [Chemical formula]

[0046] Examples of amino acid residues having an azide group include L-azidohomoalanine (AHA), 4-azido-L-phenylalanine, 4-azido-D-phenylalanine, 3-azido-L-alanine, 3-azido-D-alanine, 4-azido-L-homoalanine, 4-azido-D-homoalanine, 5-azido-L-ornithine, 5-azido-d-ornithine, 6-azido-L-lysine, and 6-azido-D-lysine. Examples of amino acid residues having an alkyne group include, for example, L-homopropargylglycine (L-HPG), D-homopropargylglycine (D-HPG), and β-homopropargylglycine (β-HPG).

[0047] The strained alkyne group at the free end of the linker can be a cyclooctene group, for example, a trans-cyclooctene (TCO) group; or a cyclooctyne group, for example, a dibenzocyclooctyne (DBCO) group, a difluorocyclooctyne (DIFO) group, a bicyclononine (BCN) group, and a dibenzocyclooctyne (DICO) group. Alternatively, the tetrazine group at the free end of the linker can be a 1,2,3,4-tetrazine group, a 1,2,3,5-tetrazine group, and a 1,2,4,5-tetrazine group, and derivatives thereof, for example, a 6-methyltetrazine group, but are not limited thereto.

[0048] In a preferred embodiment of the present disclosure, derivatives and modified forms of the peptide have an azide group, an alkyne group, or a tetrazine group directly or via a linker at the N-terminus, C-terminus, or side chain of the amino acid. Examples of such derivatives include the following examples. For example, when the epsilon amino group of the N-terminal lysine of the peptide is reacted with DBCO-PEG4-NHS ester (D5922) or BCN-PEG4-NHS ester (BP-22851), peptides (SEQ ID NOs: 49 and 50) with DBCO or BCN added via a PEG linker are obtained.

[0049]

Chemical formula

[0050] Alternatively, a peptide (SEQ ID NO: 51 and SEQ ID NO: 52) with a DBCO group introduced into the side chain of an amino acid can also be obtained by reacting DBCO-PEG5-DBCO (BP-22450) with an N-terminal lysine into which an azide group has been introduced with Fmoc-Lys(εN3)-OH, as in the following Scheme 4.

[0051]

Chemical formula

[0052] Furthermore, in the following Scheme 5, an N3-PEG4-NHS (N-Succinimidyl 15-azido-4,7,10,13-tetraoxapentadecanoate) is reacted with the α-amino group of the N-terminal glycine of the peptide represented by SEQ ID NO: 53 to synthesize a peptide (SEQ ID NO: 54) into which an azide group has been introduced, and further, a peptide (SEQ ID NO: 55) with a DBCO group introduced at the N-terminal can be obtained using DBCO-PEG5-DBCO.

[0053]

Chemical formula

[0054] (Peptide drug conjugate) The peptide of the present disclosure may be a conjugate (complex). This complex is a complex including the peptide of the present disclosure, a linker bound to this peptide, and "a certain molecule" bound to the linker. This complex is preferably a complex in which at least "a certain molecule" can pass through the fenestrated tissue. The entire complex may also be able to pass through the fenestrated tissue.

[0055] Examples of the linker include, but are not limited to, those composed of about 1 to 15 residues of amino acids, those composed of fatty acids, those composed of polyethylene glycol, or derivatives thereof. The linker may be such that it dissociates or separates under specific environments or conditions, or may maintain a stable structure.

[0056] As used herein, the "certain molecule" can be any molecule desired by those skilled in the art, as long as it is a molecule intended to be delivered to the brain. However, since the mechanism for passing through the blood-brain barrier is receptor-mediated transcytosis (RMT) via binding to LRP1, molecules that are too large to be transported by this mechanism are not preferred. Examples of the "certain molecule" include, but are not limited to, fluorescent dyes, low molecular weight compounds, peptides, peptoids, nucleic acids, sugar chains, lipids, antibodies, proteins, enzymes, liposomes, micelles, nanoparticles, carbon nanotubes, carbon nanohorns, carbon nanosheets, and their radioisotope-labeled forms. In a preferred embodiment, the "certain molecule" is a drug, which may be, for example, an antibody, a nucleic acid, or a bioactive peptide.

[0057] The complex of the peptide of the present disclosure and the "certain molecule" may be a fusion or a graft. For example, the peptide of the present invention may be fused to the N-terminus and / or C-terminus of an antibody or a protein, or grafted into a loop region.

[0058] The above-mentioned fusion or graft may be expressed as part of an external domain of cells, fungi, viruses, etc., or conjugated to an external domain of cells, fungi, viruses, etc.

[0059] The complex of the peptide of the present disclosure and the "certain molecule" may be via a "molecule having binding activity to the certain molecule". The peptide of the present disclosure and the "molecule having binding activity to the certain molecule" may be any of a conjugate, a fusion, and / or a graft. The binding between the "certain molecule" and the "molecule having binding activity to the certain molecule" may be reversible or irreversible.

[0060] The complex of the peptide of the present disclosure and the "certain molecule" may be a crystal, and the present invention encompasses peptides having a single crystal form or a mixture of crystal forms. The crystal can be produced by applying a crystallization method known per se to effect crystallization.

[0061] The complex of the peptide of the present disclosure and "a certain molecule" may be a pharmaceutically acceptable cocrystal or cocrystal salt. Here, the cocrystal or cocrystal salt means a crystalline substance composed of two or more unique solids at room temperature, each having different physical properties (e.g., structure, melting point, heat of fusion, hygroscopicity, solubility, and stability, etc.). The cocrystal or cocrystal salt can be produced according to the cocrystallization method known per se.

[0062] (Action and effect of cyclic peptide) As shown in the examples described later, sequences 1 to 46, which are representative examples of the amino acid sequence group represented by this embodiment, have LRP1 binding activity. And sequence 2 has the ability to pass through the blood-brain barrier because it migrates from the blood-side well to the brain-side well in the in vitro blood-brain barrier model. Since the amino acid sequence group represented by this embodiment has amino acid sequences and conformational features similar to these representative examples, it is considered highly likely that they also have LRP1 binding activity and the ability to pass through the blood-brain barrier. And it is considered highly likely that the conjugate, fusion, and graft of the peptide of the present invention also have LRP1 binding activity and the ability to pass through the blood-brain barrier.

[0063] As shown in the examples described later, sequences 1 to 46, which are representative examples of the amino acid sequence group represented by this embodiment, have LRP1 binding activity. This indicates the following. X 5 and X 15 The bond between the Cα carbon atoms of, X 6 and X 15 The bond between the Cα carbon atoms of, X 5 and X 16 The bond between the Cα carbon atoms of, or X 6 and X 16 The bond between the Cα carbon atoms of accepts a wide range of structures such as amide bonds, disulfide bonds, thioether bonds, C=C bonds, C-C bonds, bonds via triazole, bonds via dithiotetrafluorobenzene, and bonds via a dimethylphenylene group (the bonding site of the methylene group to the phenylene ring may be ortho, meta, or para).

[0064] The amino acid sequence of LRP1 cluster 4 is highly conserved in mammals. Since SEQ ID NO: 2, a representative example of the peptide of the present invention, exhibits the ability to cross the blood-brain barrier in in vitro blood-brain barrier models of rat and monkey types, it is highly likely that the peptide group consisting of the amino acid sequence represented in this embodiment, and the complex of the peptide group and "a certain molecule" bind to LRP1 of non-human mammals such as mouse, rat, dog, and monkey, and exhibit the ability to cross the barrier tissue.

[0065] (Method for producing cyclic peptide) The peptide of this embodiment can be produced by known peptide production methods such as chemical synthesis methods such as the liquid phase method, the solid phase method, or a hybrid method combining the liquid phase method and the solid phase method.

[0066] For the solid phase method, a commercially available automatic synthesizer can be used. For example, the hydroxyl group of a resin having a hydroxyl group and the carboxyl group of a first amino acid (usually the C-terminal amino acid of the target peptide) whose α-position amino group is protected by a protecting group such as an Fmoc group are esterified. As the esterification catalyst, known dehydrating condensing agents such as 1-mesitylenesulfonyl-3-nitro-1,2,4-triazole (MSNT), dicyclohexylcarbodiimide (DCC), and diisopropylcarbodiimide (DIPCDI) can be used. Next, the protecting group of the α-position amino group of the first amino acid is removed, and a second amino acid in which all functional groups other than the carboxyl group of the main chain are protected is added, and the carboxyl group is activated to bond the first and second amino acids. Further, the α-position amino group of the second amino acid is deprotected, a third amino acid in which all functional groups other than the carboxyl group of the main chain are protected is added, and the carboxyl group is activated to bond the second and third amino acids. This is repeated to synthesize a peptide of the desired length. The linear peptide is cleaved from the resin, purified, the functional group for cyclizing the peptide is deprotected, and the peptide is cyclized according to a conventional method.

[0067] Examples of the resin for solid-phase synthesis include Merrifield resin, MBHA resin, Cl-Trt resin, SASRIN resin, Wang resin, Rink amide resin, HMFS resin, Amino-PEGA resin (Merck), HMPA-PEGA resin (Merck), etc. These resins may be used after washing with a solvent (dimethylformamide (DMF), 2-propanol, methylene chloride, etc.). Examples of the protecting group for the α-position amino group include benzyloxycarbonyl (Cbz) group, tert-butoxycarbonyl (Boc) group, fluorenylmethoxycarbonyl (Fmoc) group, benzyl group, allyl group, allyloxycarbonyl (Alloc) group, etc. The Cbz group can be deprotected by hydrofluoric acid, hydrogenation, etc., the Boc group can be deprotected by trifluoroacetic acid (TFA), and the Fmoc group can be deprotected by treatment with piperidine. For the protection of the α-position carboxy group, methyl ester, ethyl ester, benzyl ester, tert-butyl ester, cyclohexyl ester, etc. can be used. As other functional groups of amino acids, hydroxy groups such as serine and threonine can be protected with a benzyl group or a tert-butyl group, and hydroxy groups such as tyrosine can be protected with a 2-bromobenzyloxycarbonyl group or a tert-butyl group. Amino groups in side chains such as lysine, carboxy groups such as glutamic acid and aspartic acid can be protected in the same manner as the α-position amino group and the α-position carboxy group.

[0068] The activation of the carboxy group can be carried out using a condensing agent. Examples of the condensing agent include dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIPCDI), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC or WSC), (1H-benzotriazol-1-yloxy)tris(dimethylamino)phosphonium hexafluorophosphate (BOP), 1-[bis(dimethylamino)methyl]-1H-benzotriazolium-3-oxide hexafluorophosphate (HBTU), and the like. Cleavage of the peptide chain from the resin can be performed by treatment with an acid such as TFA or hydrogen fluoride (HF).

[0069] In one aspect, the peptide of the present disclosure may be cyclized. As used herein, cyclization means that in one peptide, two or more amino acids separated by one or more amino acids are directly or indirectly covalently bonded via a linker to form one or more cyclic structures within the molecule. Cyclization can be carried out according to the methods described in Non-Patent Documents 8 and 9. For example, an amide bond between an amino group and a carboxy group, a disulfide bond between a thiol group and a thiol group, a thioether bond between a thiol group and a halogen group, a thioether bond by a thiol-ene reaction between a thiol group and an allyl group, a C=C bond by an olefin metathesis reaction between an allyl group and an allyl group (the C=C bond may be converted to a C-C bond by a reduction reaction), a bond via a triazole by a click reaction between an alkynyl group and an azide group, a thioether bond between a linker having a halogen group and two thiol groups, and the like, but is not limited thereto. The direct or linker-mediated indirect covalent bond for cyclization may be any of main chain-main chain, main chain-side chain, side chain-main chain, and side chain-side chain.

[0070] For the cyclization of the peptides of the present disclosure, for example, (1) cysteine, D-cysteine, homocysteine, D-homocysteine having a thiol group can be used as amino acid 1 and amino acid 2 respectively, and a disulfide bond formed between their thiol groups can be used; (2) an amino acid having a nucleophilic halogen atom (chloro, bromo, or iodo) (e.g., 3-chloroalanine), or a carboxylic acid having a halogen atom (chloro, bromo, or iodo) (e.g., 3-chloropropanoic acid) can be used as amino acid 1, and a thioether bond formed between it and an amino acid having a thiol group can be used; (3) amino acid 1 and amino acid 2 having a thiol group, and a linker having a nucleophilic halogen atom (chloro, bromo, or iodo) (e.g., 1,1-diiodomethane, 1,1-dichloroacetone, 1,2-diiodoethane, 1,3-diiodopropane, 1,4-diiodobutane, α,α'-dibromo-o-xylene, α,α'-dibromo-m-xylene, α,α'-dibromo-p-xylene, hexafluorobenzene, etc.) can be used to form a thioether bond; (4) β-azidoalanine having an azide group can be used as amino acid 1, 2-amino-5-hexynoic acid having an alkynyl group can be used as amino acid 2, and a covalent bond via a triazole formed by a click reaction between them can be used; (5) an amino acid having an allyl group (e.g., allylglycine, D-allylglycine, homoallylglycine, D-homoallylglycine, etc.), or a carboxylic acid having an allyl group (e.g., 3-butenoic acid) can be used as amino acid 1, and a thioether bond formed by a thiol-ene reaction between it and an amino acid having a thiol group can be used; (6) an amino acid having an allyl group, or a carboxylic acid having an allyl group can be used as amino acid 1 and amino acid 2 respectively, and a C=C bond formed by an olefin metathesis reaction between their allyl groups can be used; (7) a C-C formed by reducing a C=C bond formed by an olefin metathesis reaction can be used; (8) amino acid 1 having an amino group (e.g., 2,3-diaminopropanoic acid, 2,The amide bond between an amino acid having an N-terminal amino group and a carboxy group or a C-terminal carboxy group (such as 4-diaminobutyric acid, ornithine, lysine, their D-form amino acids, β-alanine, etc.) and an amino acid 2 having a carboxy group (such as aspartic acid, glutamic acid, their D-form amino acids, etc.) or a C-terminal carboxy group can be used. The amide bond between an amino acid having an N-terminal amino group and a carboxy group or a C-terminal carboxy group can be used. Amino acid 1 and amino acid 2 can be on either the N-terminal side.,

[0071] (Medicines, diagnostic agents, research reagents including cyclic peptides) The pharmaceutical composition according to the present disclosure contains, as an active ingredient, a peptide composed of the above-described amino acid sequence, and the peptide can bind to LRP1 and migrate to the central nervous system tissue via the RMT of LRP1. The administration form of the above pharmaceutical composition is not particularly limited, and may be oral administration or parenteral administration. Examples of parenteral administration include injection administration such as intramuscular injection, intravenous injection, subcutaneous injection, transdermal administration, transmucosal administration (nasal, oral, ocular, pulmonary, vaginal, or rectal administration), etc. In view of the property that the peptide in the pharmaceutical composition is easily metabolized and excreted, various modifications can be made. For example, by adding an alkyl chain, polyethylene glycol, or sugar chain to the peptide, the residence time in the blood can be prolonged and the antigenicity can be reduced. In addition, biodegradable polymers such as polylactic acid-glycolic acid (PLGA), porous hydroxyapatite, liposomes, surface-modified liposomes, emulsions prepared with unsaturated fatty acids, nanoparticles, microparticles, nanospheres, etc. can be used as sustained-release bases, and the peptide can be encapsulated therein. In the case of transdermal administration, a weak current can also be passed through the skin surface to permeate the stratum corneum (iontophoresis method).

[0072] The above pharmaceutical composition may use the active ingredient as it is, or may be formulated by adding pharmaceutically acceptable carriers, excipients, additives, etc. Examples of dosage forms include, for example, liquid preparations (such as injections), dispersants, suspensions, tablets, pills, powders, suppositories, powders, fine granules, granules, capsules, syrups, troches, inhalants, ointments, eye drops, nasal drops, ear drops, poultices, etc. These preparations may be controlled-release preparations such as immediate-release preparations or sustained-release preparations (such as sustained-release microcapsules). Formulation can be carried out, for example, by appropriately using excipients, binders, disintegrants, lubricants, solvents, solubilizers, coloring agents, flavoring and odor-correcting agents, stabilizers, emulsifiers, absorption promoters, surfactants, pH adjusters, preservatives, antioxidants, etc. according to conventional methods. Examples of components used in formulation include pharmaceutically acceptable organic solvents such as purified water, saline, phosphate buffer, dextrose, glycerol, ethanol, animal and vegetable oils, lactose, mannitol, glucose, sorbitol, crystalline cellulose, hydroxypropyl cellulose, starch, corn starch, anhydrous silicic acid, magnesium aluminum silicate, collagen, polyvinyl alcohol, polyvinyl pyrrolidone, carboxyvinyl polymer, sodium carboxymethyl cellulose, sodium polyacrylate, sodium alginate, water-soluble dextran, sodium carboxymethyl starch, pectin, methyl cellulose, ethyl cellulose, xanthan gum, gum arabic, tragacanth, casein, agar, polyethylene glycol, diglycerin, glycerin, propylene glycol, petrolatum, paraffin, octyldodecyl myristate, isopropyl myristate, higher alcohols, stearyl alcohol, stearic acid, human serum albumin, etc., but are not limited thereto.When a peptide is poorly absorbed transmucosally, as absorption enhancers for improving the absorption of poorly absorbable drugs, surfactants such as polyoxyethylene lauryl ethers, sodium lauryl sulfate, and saponins; bile salts such as glycolic acid, deoxycholic acid, and taurocholic acid; chelating agents such as EDTA and salicylic acids; fatty acids such as caproic acid, capric acid, lauric acid, oleic acid, linoleic acid, and mixed micelles; enamine derivatives, N-acyl collagen peptides, N-acyl amino acids, cyclodextrins, chitosans, nitric oxide donors, etc. may be used.

[0073] The pills or tablets can also be coated with sugar coating, gastric-soluble, or enteric-soluble substances. The injections can contain distilled water for injection, physiological saline, propylene glycol, polyethylene glycol, vegetable oil, alcohols, etc. Furthermore, wetting agents, emulsifiers, dispersants, stabilizers, solubilizers, solubilization aids, preservatives, etc. can be added. If necessary, additives such as ordinary preservatives, antioxidants, colorants, sweeteners, adsorbents, wetting agents, etc. can be appropriately used in appropriate amounts.

[0074] The pharmaceutical composition of the present disclosure transports a certain molecule to the central nervous system tissue through binding to LRP1 and is effective for the treatment or prevention of various diseases targeted by the certain molecule. For example, when the certain molecule is an anti-cancer agent, brain tumors (such as pineal astrocytoma, pilocytic astrocytoma, diffuse astrocytoma, anaplastic astrocytoma, etc.), schwannoma, etc. can be mentioned. When the certain molecule targets a molecule related to central nervous system diseases, mental disorders (such as schizophrenia, schizoaffective disorder, schizophreniform disorder, delusional disorder, etc.), pediatric mental disorders (such as attention deficit disorder, attention deficit / hyperactivity disorder, conduct disorder, autism, etc.), neurodegenerative disorders, neural stem cell disorders, neural progenitor disorders, ischemic disorders, traumatic nerve disorders, mood disorders, psychomotor disorders, sleep disorders (such as hypersomnia, circadian rhythm sleep disorder, insomnia, abnormal sleep behavior, sleep apnea, etc.), mental disorders such as anxiety (such as acute stress disorder, generalized anxiety disorder, social anxiety disorder, panic disorder, post-traumatic stress disorder, agoraphobia, obsessive-compulsive disorder, etc.), factitious mental disorders (such as acute hallucinatory mania, etc.), impulse control disorders (such as compulsive gambling, intermittent explosive disorder, etc.), mood disorders (such as bipolar I disorder, bipolar II disorder, mania, mixed mood state, etc.), major depression, chronic depression, seasonal depression, psychotic depression, seasonal depression, cognitive disorders (such as amnesia, senile dementia, HIV-related dementia, Alzheimer's disease, Huntington's disease, Lewy body dementia, vascular dementia, drug-related dementia, tardive dyskinesia, intergenerational myoclonus, dystonia, delirium, Pick's disease, Creutzfeldt-Jakob disease, HIV disease, Gilles de la Tourette syndrome, epilepsy, muscle spasm, mild cognitive impairment, etc.), mental retardation (such as spasticity, Down syndrome, fragile X syndrome, etc.); premenstrual syndrome (PMS), premenstrual dysphoric disorder (PDD), postpartum depression, neuron injury disorders (such as eye injury, retinopathy or macular degeneration of the eye, tinnitus, hearing impairment, cerebral edema, etc.), Parkinson's disease, Parkinson's disease-like disorders, migraine, epilepsy, Alzheimer's disease, brain injury, stroke, cerebrovascular diseases (such as cerebral arteriosclerosis, cerebral amyloid angiopathy, hereditary cerebral hemorrhage, cerebral hypoxia-ischemia, etc.), drug dependence (such as narcotic dependence, alcoholism, amphetamine dependence, cocaine addiction, nicotine dependence, drug withdrawal syndrome, etc.), eating disorders (such as anorexia nervosa, bulimia nervosa, binge eating disorder, polyphagia, obesity, compulsive overeating disorder, pica, etc.), etc. are mentioned, but not limited thereto.

[0075] The pharmaceutical composition of the present disclosure may be administered in combination with other pharmaceuticals or therapies useful for the above diseases. For example, in the case of malignant tumors, it may be combined with various chemotherapy, surgical treatment, and radiotherapy.

[0076] When the pharmaceutical composition of the present disclosure is administered to mammals (e.g., humans, mice, rats, guinea pigs, rabbits, dogs, horses, monkeys, pigs, etc.), particularly humans, the dosage varies depending on symptoms, the age, sex, weight, sensitivity difference, administration method, administration interval, type of active ingredient, and type of formulation, and is not particularly limited. For example, 30 μg to 1000 mg, 100 μg to 500 mg, or 100 μg to 100 mg can be administered once or divided into several times.

[0077] (Polynucleotide encoding the peptide of the present disclosure) In one embodiment, a polynucleotide encoding the peptide of the present disclosure is provided. This polynucleotide can be a polynucleotide encoding an amino acid sequence composed only of natural amino acids among the amino acid sequences of the peptide of the present disclosure or a polynucleotide having a base sequence having a predetermined sequence identity with the polynucleotide. The degree of sequence identity of the base sequence can be about 70% or more, preferably about 80% or more, more preferably about 90% or more, and even more preferably about 95% or more. The base sequence identity can be determined by a method known per se. For example, the base sequence identity (%) can be determined in the same manner as the amino acid sequence identity (%) described above.

[0078] In another embodiment, the polynucleotide can be a base sequence in which one or more nucleotides are modified by one or more selected from substitution, addition, deletion, and insertion in the base sequence of the polynucleotide encoding the amino acid sequence composed only of natural amino acids among the amino acid sequences of the peptide of the present disclosure. The number of nucleotides to be modified is not particularly limited as long as it is one or more, but can be, for example, 1 to about 50, preferably 1 to about 30, more preferably 1 to about 10, and even more preferably 1 to about 5 (e.g., 1 or 2).

[0079] (Expression Vector) The expression vector according to an embodiment of the present disclosure may include a polynucleotide encoding a target polypeptide to be expressed or a target polynucleotide to be expressed, and a promoter functionally linked to the polynucleotide. "The promoter is functionally linked to the polynucleotide" means that the promoter is bound to the polynucleotide encoding the gene so as to enable the expression of the polynucleotide itself under its control or the expression of the polypeptide encoded by the polynucleotide.

[0080] The backbone of the expression vector of this embodiment is not particularly limited as long as it can produce a target substance in a predetermined cell. Examples include plasmid vectors and viral vectors. When the expression vector is used as a medicine, viral vectors such as adenovirus, retrovirus, adeno-associated virus, herpes virus, vaccinia virus, poxvirus, poliovirus, Sindbis virus, and Sendai virus are suitable vectors for administration to mammals.

[0081] When a prokaryotic cell is used as the host cell, an expression vector that can utilize the prokaryotic cell as the host cell can be used. Such an expression vector may include elements such as a promoter-operator region, a start codon, a polynucleotide encoding the polypeptide or a partial peptide thereof of this embodiment, a stop codon, a terminator region, and an origin of replication. The promoter-operator region for expressing the polypeptide of the present invention in bacteria includes a promoter, an operator, and a Shine-Dalgarno (SD) sequence. For these elements, those known per se can be used.

[0082] In addition, when eukaryotic cells are used as host cells, an expression vector that can utilize the eukaryotic cells as host cells can be used. In this case, the promoter to be used is not particularly limited as long as it can function in eukaryotes such as mammals. When aiming for the expression of a polypeptide, examples of such promoters include viral promoters such as the SV40-derived early promoter, cytomegalovirus LTR, Rous sarcoma virus LTR, MoMuLV-derived LTR, and adenovirus-derived early promoter, as well as mammalian constitutive protein gene promoters such as the β-actin gene promoter, PGK gene promoter, and transferrin gene promoter. When aiming for the expression of a polynucleotide, the promoter can be a polIII promoter (e.g., tRNA promoter, U6 promoter, H1 promoter).

[0083] The expression vector of the present invention may further include sites for transcription initiation and termination, a ribosome binding site that may be required for translation in the transcription region, an origin of replication, and a selection marker gene (e.g., ampicillin, tetracycline, kanamycin, spectinomycin, erythromycin, chloramphenicol), etc. The expression vector of the present invention can be prepared by a method known per se (e.g., see Molecular Cloning cited above).

[0084] (Cells, liposomes, or nanoparticles presenting the peptide of the present disclosure) In another aspect of the present disclosure, cells, liposomes, or nanoparticles presenting the peptide of the present disclosure are provided. The cells can be prepared by isolating a desired cell from a living body and conjugating the peptide of the present disclosure to this cell in vitro, or by introducing a vector expressing the peptide of the present disclosure into the cell to present it on the cell surface. The liposomes or nanoparticles can be prepared by conjugating the peptide of the present disclosure, preparing a conjugate of the peptide of the present disclosure and a fatty chain, and inserting it into the lipid membrane via the fatty chain to present it on the surface of the liposomes or nanoparticles.

[0085] The following examples are merely illustrative and are intended only to explain the present invention in detail together with the above-described embodiments, and do not limit the present invention. Those skilled in the art can modify the present invention in various ways without departing from the meaning of the present invention, and such modifications are also included in the scope of the present invention.

Examples

[0086] The abbreviations used in this specification represent the following meanings. BSA: Bovine serum albumin RP-HPLC: Reverse-phase high performance liquid chromatography HRP: Horseradish peroxidase SA: Streptavidin D-PBS: Dulbecco’s Phosphate buffered saline ELISA: Enzyme-linked immunosorbent assay Ac: Acetyl Cys: L-Cysteine Gly: Glycine Ala: L-Alanine Ser: L-Serine Thr: L-Threonine Pro: L-Proline cHyp: cis-4-Hydroxy-L-Proline tHyp: trans-4-Hydroxy-L-Proline Tyr: L-Tyrosine Trp: L-Tryptophan Phe: L-Phenylalanine 4fF: 4―fluoro―L-Phenylalanine 4cF: 4―chloro―L-Phenylalanine His: L-Histidine Lys: L-Lysine Orn: L - Ornithine Arg: L - Arginine Val: L - Valine Leu: L - Leucine Ile: L - Isoleucine Nle: L - Norleucine Ahep: (S) - 2 - Aminoheptanonic acid Aoc: (S) - 2 - Aminooctanonic acid Aib: 2 - Aminoisobutyric acid Asp: L - Aspartic acid Glu: L - Glutamic acid 5 - FAM: 5 - Carboxyfluorescein DX: D - amino acid X c(XY): Cyclization between amino acid X to amino acid Y

[0087] (Peptide synthesis) The chemical synthesis of all the peptides used in this example was entrusted to Scrum Co., Ltd. (Tokyo, Japan), and was carried out using an automatic synthesizer Syro II (manufactured by Biotage) with a standard solid-phase synthesis method using the 9-fluorenylmethoxycarbonyl group (Fmoc group) as a protecting group for the α-amino group. The side-chain protected amino acid-resin located at the C-terminus was placed in a synthesis column, and the apparatus was set up. Subsequently, 1-[bis(dimethylamino)methylene]-1H-1,2,3-triazol[4,5-b]pyridinium 3-oxide hexafluorophosphate (HATU) / diisopropylethylamine (DIEA) was added to the next amino acid protected with the Fmoc group for activation, and then placed in the column for reaction. After completion of the reaction, it was washed, and the Fmoc group was deprotected using 20% piperidine. By repeating this step, the peptide chain was extended. After deprotecting the Fmoc group of the final amino acid, the peptide-resin was taken out from the apparatus. 30% hexafluoro-2-propanol (HFIP) / dichloromethane (DCM) was added to the peptide-resin to cleave the linear side-chain protected peptide from the resin, and the side-chain protected peptide was recovered by ether precipitation. The side-chain protected peptide was purified by RP-HPLC using a SunFire C18 column (10×150 mm) (manufactured by Waters), and then lyophilized. The cyclization of the peptide was referred to the methods described in Non-Patent Document 8 and Non-Patent Document 9. The theoretical molecular weight, measured molecular weight, purity, cyclization type, and amino acid sequence of the peptides synthesized in this example are shown in Table 1 and Table 2. Also, the structural formulas of Sequence 2 and Sequence 46 among these are shown in Figure 7. In Table 1 and Table 2, amino acid residues without D notation indicate the L form.

[0088]

Table 1

[0089]

Table 2

[0090] (Construction of Binding Test by ELISA Method) To evaluate the binding activity of the peptide to LRP1, a binding assay by ELISA was constructed. The following briefly describes this ELISA method. First, goat anti-human IgG-Fcγ polyclonal antibody (Catalog No. 109 - 0005 - 008, manufactured by Jackson ImmunoResearch) was added to a 96-well Maxisorp plate (Catalog No. 439454, manufactured by Nunc) at 1 μg / 50 μL / well using D-PBS and coated overnight at 4°C. Then, 0.5% BSA / D-PBS was further added at 300 μL / well and blocked at room temperature for 30 minutes. After washing the plate with 0.1% Tween20 / D-PBS, LRP1 cluster 2-Fc chimeric protein (Catalog No. 2368 - L2 - 050, manufactured by R&D system), LRP1 cluster 3-Fc chimeric protein (Catalog No. 48248 - L3 - 050, manufactured by R&D system), or LRP1 cluster 4-Fc chimeric protein (Catalog No. 5395 - L4 - 050, manufactured by R&D system), which are recombinant proteins of LRP1, was added at 100 ng / 50 μL / well using 0.5% BSA / D-PBS and reacted with the coated antibody at room temperature for 30 minutes. After washing the plate with 0.1% Tween20 / PBS, sequences 45 and 46, which are biotinylated peptides, were prepared at arbitrary concentrations using 0.5% BSA / D-PBS and added at 50 μL / well. After reacting at room temperature for 30 minutes, the plate was washed with 0.1% Tween20 / D-PBS. Sequences 45 and 46 that bound to the recombinant protein of LRP1 captured on the plate were detected with SA-HRP (Catalog No. ab7403, manufactured by Abcam). For the quantification of HRP, the absorbance at 450 nm was measured using TMB-ELISA Substrate Solution (Catalog No. 34028, manufactured by Thermo Fisher).

[0091] Figure 1 shows the selective and peptide concentration-dependent binding of the LRP1 cluster 4-Fc chimeric proteins of sequences 45 and 46 used in this binding assay (n = 4, ±SEM). The specific binding activity was estimated by subtracting the absorbance obtained when the biotinylated peptide was added to wells not capturing the recombinant protein of LRP1 from the absorbance obtained when the biotinylated peptide was added to wells capturing the recombinant protein of LRP1. As a result, both sequence 45 and sequence 46 bound to the LRP1 cluster 4-Fc chimeric protein selectively and in a peptide addition concentration-dependent manner. Little or only slight binding to the LRP1 cluster 2-Fc chimeric protein and the LRP1 cluster 3-Fc chimeric protein was shown. The binding EC50 values of sequence 45 and sequence 46 were calculated to be 10.5 nM and 11.0 nM, respectively, and showed equivalent binding activity regardless of the presence or absence of a cyclic structure.

[0092] (Construction of competitive binding assay by ELISA method) To compare the binding activity of the peptide with amino acid substitutions introduced into Array 1 to LRP1 and its resistance to degradation by proteases with those of Array 2, a competitive binding assay by the ELISA method shown in Figure 2 was constructed. Immediately after mixing (Plasma incubation) Array 2 or the amino acid-substituted peptide with mouse plasma (0 hours) or after 24 hours at 37°C, it was diluted to an arbitrary final concentration so that the plasma concentration became sufficiently low and then mixed with Array 46 (500 nM) and subjected to the above-described competitive binding assay. Since the binding of Array 46 to the LRP1 cluster 4-Fc chimeric protein competes with the binding of Array 2 or the amino acid-substituted peptide present in the solution to the LRP1 cluster 4-Fc chimeric protein, it is inhibited in a concentration-dependent manner by Array 2 or the amino acid-substituted peptide. That is, the binding activity of Array 2 or the amino acid-substituted peptide to the LRP1 cluster 4-Fc chimeric protein is detected as a competitive inhibitory activity against the binding of Array 46. Using the binding value of Array 46 to the wells not capturing the LRP1 cluster 4-Fc chimeric protein as 100% inhibitory activity and the binding value of Array 46 to the wells without the addition of Array 2 or the amino acid-substituted peptide as 0% inhibitory activity, the inhibitory activity values of Array 2 and the amino acid-substituted peptide were calculated in %.

[0093] (Evaluation of the binding activity of the amino acid-substituted peptide to LRP1) The results of the competitive binding assay for the LRP1 cluster 4-Fc chimeric protein, examined with n = 4 (±SEM), are shown in Figure 3. Sequence 2 at 0 hours of plasma and culture inhibited the binding of sequence 46 in an added concentration-dependent manner. The binding of sequence 46 (500 nM) in the wells to which 500 nM of sequence 2 was added was 52.4% on average, indicating that sequence 2 and sequence 46 compete for binding to LRP1 cluster 4 in a 1:1 ratio. Sequence 2 at 24 hours of plasma and culture also inhibited the binding of sequence 46 in an added concentration-dependent manner. However, the binding of sequence 46 (500 nM) in the wells to which the equivalent of 2000 nM of sequence 2 was added was 50.6% on average. That is, despite the addition of the equivalent of 2000 nM to the wells, only the inhibitory activity equivalent to 500 nM was shown, indicating that approximately 75% of sequence 2 was degraded during the 24-hour incubation with mouse plasma. As a result of subjecting the same test to DMSO immediately after mixing with mouse plasma, the binding of sequence 46 (500 nM) was not inhibited at all, indicating that the plasma mixed into the binding system did not cause system inhibition.

[0094] Sequences 1 to 44 were mixed with mouse plasma immediately or after 24 hours at 37°C, and then diluted to a final concentration of 2000 nM so that the plasma concentration would be sufficiently low, and then mixed with sequence 46 (500 nM) and subjected to the above-described competitive binding assay. The results are shown in Figure 4. The inhibitory activities of sequence 1 and sequence 2 were estimated to be 70.7% and 79.4%, respectively. The inhibitory activities of sequence 6, sequence 7, sequence 9, sequence 19, sequence 24, sequence 34, and sequence 43 were 91.9%, 87.6%, 85.8%, 79.1%, 77.4%, 82.0%, and 83.5%, respectively, showing inhibitory activities equal to or higher than those of sequence 1 and sequence 2. The other sequences tended to show attenuated inhibitory activities. Similar to sequence 2, many amino acid-substituted peptides also showed uniformly attenuated inhibitory activities after 24 hours of mixing with mouse plasma compared to when subjected to the competition test immediately after mixing with mouse plasma, indicating that they were degraded in mouse plasma.

[0095] (Evaluation by in vitro blood-brain barrier model) Figure 5 shows an overview of an in vitro blood-brain barrier model for evaluating the ability to cross the blood-brain barrier. Tests were conducted using a rat-type BBB kit (Catalog No. RBT-24H, manufactured by Pharmacell) and a monkey-type BBB kit (Catalog No. MBT-24H, manufactured by Pharmacell). These kits retain in vivo BBB characteristics (such as the presence or absence of the expression of various receptors and transporters and the ability to form tight junctions) and are widely used as an evaluation system for estimating the brain permeability of drugs (Non-Patent Documents 6 and 7). The insert well was used as the vascular side, and the bottom well was used as the brain side. The amount of the peptide added to the insert well that permeated through the bottom surface of the three-layer structure composed of endothelial cells / membrane filter / pericytes was evaluated. In addition, the TEER value (trans-epithelial electrical resistance value), which represents the BBB function, was measured before and after the test to confirm that the addition of the peptide did not affect the BBB function.

[0096] Figure 6 shows the results of evaluating the BBB permeability of SEQ ID NO: 2, with ANG2 (Non-Patent Document 4) and L57 (Non-Patent Document 5), for which LRP1 binding activity and BBB permeability have been reported, as comparison targets. 5-FAM-labeled ANG (SEQ ID NO: 47), 5-FAM-labeled L57 (SEQ ID NO: 48), and SEQ ID NO: 2 were added to the insert well at final concentrations of 10 μM, 3 μM, and 1 μM, respectively. After culturing at 37°C for 24 hours, the culture supernatant in the bottom well was collected. A calibration curve was created by serially diluting each 5-FAM-labeled peptide in the medium to an arbitrary concentration, and the concentration of the 5-FAM-labeled peptide contained in each collected culture supernatant was calculated. The amount of peptide (number of moles) added to the insert well (vascular side) was taken as 100%, and the amount of peptide (number of moles) that migrated to the bottom well (brain side) was calculated as a percentage. The BBB permeability of SEQ ID NO: 2 was significantly higher than that of 5-FAM-labeled ANG and L57 at any concentration, regardless of whether it was the rat BBB shown in the upper graph or the monkey BBB shown in the lower graph (n = 4, ±SD, *p < 0.01, Dunnett's test against SEQ ID NO: 2). After 24 hours of culture, the TEER value (trans-epithelial electrical resistance value) of each well was 150 Ω×cm 2The above was achieved, and it was shown that the BBB function was maintained. From these results, it was shown that the peptide of the present invention can pass through the barrier tissue, particularly the blood-brain barrier. And it was confirmed that the BBB permeability of SEQ ID NO: 2, which is a representative example of the peptide of the present invention, is superior to that of existing ANG2 and L57.

[0097] (Synthesis and Evaluation of Peptide Modification DBCO-KS-487) <Synthesis Method> The synthesis was outsourced to Scrum Co., Ltd. (Tokyo, Japan). First, the following linear peptide precursor was synthesized by performing a standard solid-phase synthesis method using 9-fluorenylmethoxycarbonyl group (Fmoc) as a protecting group for the α-amino group with an automatic synthesizer SyruII (manufactured by Biotage).

[0098] NH2-Gly-Thr(tBu)-Pro-Cys(Trt)-Thr(tBu)-Tyr(tBu)-Lys(Boc)-Tyr(tBu)-Nle-Leu-Ala-Glu(OtBu)-Nle-Cys(Trt)-Trt(2-Cl)-Resin

[0099] Next, the resin was immersed in dimethylformamide (DMF), and diisopropylethylamine (DIEA) was added to adjust the pH to 8. At this pH of 8, N3-PEG4-NHS (Cas#944251-24-5, catalog #J64834, manufactured by Alfa Aesar) dissolved in DMF was added and reacted to introduce an azide group (N3-) to the N-terminal α-amino group, and the following was obtained.

[0100] N3-PEG4-Gly-Thr(tBu)-Pro-Cys(Trt)-Thr(tBu)-Tyr(tBu)-Lys(Boc)-Tyr(tBu)-Nle-Leu-Ala-Glu(OtBu)-Nle-Cys(Trt)-Trt(2-Cl)-Resin

[0101] Subsequently, trifluoroacetic acid (TFA) / ultrapure water (Water) / Thioanisole / 1,2-ethanedithiol (EDT) / triisopropylsilyl chloride (Tips) (83 / 5 / 5 / 5 / 2) was added to deprotect the side chains of each amino acid residue and cleave the peptide from the resin. After the addition, the reaction was carried out at room temperature for 2 hours. After the reaction, the resin was filtered, and the resulting filtrate was precipitated by adding it to cooled ether. The precipitate was dissolved in a small amount of ultrapure water and lyophilized to obtain the following.

[0102] N3-PEG4-Gly-Thr-Pro-Cys-Thr-Tyr-Lys-Tyr-Nle-Leu-Ala-Glu-Nle-Cys-OH (SEQ ID NO: 56)

[0103] The lyophilized product was dissolved in DMSO, and a small amount of 0.1 M NH4CO3 aqueous solution was added dropwise, and the oxidation reaction was carried out with stirring at room temperature for 1 to 2 days to form an S-S bond between the side chains of the Cys residues. It was separated and purified by RP-HPLC and lyophilized to obtain the following cyclic peptide.

[0104] N3-PEG4-Gly-Thr-Pro-c(Cys-Thr-Tyr-Lys-Tyr-Nle-Leu-Ala-Glu-Nle-Cys)-OH (SEQ ID NO: 54)

[0105] The purified cyclic peptide and DBCO-PEG5-DBCO (Cas#2363130-04-3, Catalog#BP-22450, manufactured by BROADPHARM) were each dissolved in dimethyl sulfoxide (DMSO), mixed, and then 50 mM phosphate buffer was added. After the addition, the reaction was carried out at room temperature for 2 hours. It was separated and purified by RP-HPLC and lyophilized to obtain the following DBCO-KS-487 (theoretical molecular weight 2701.3, measured molecular weight 2702.0, purity 95.0%). Here, the cyclic peptide portion excluding the N-terminal modification portion (DBCO-PEG5-Taz-PEG4) is referred to as KS-487.

[0106] DBCO-PEG5-Taz-PEG4-Gly-Thr-Pro-c(Cys-Thr-Tyr-Lys-Tyr-Nle-Leu-Ala-Glu-Nle-Cys)-OH (SEQ ID NO: 55)

[0107] <Confirmation of Reactivity of DBCO> DBCO-KS-487 and 6-azidohexanoic acid were mixed in an appropriate amount in DMSO / PBS (50 / 50) (pH 7.5) and reacted at room temperature. Then, the shift of the peak position of DBCO-KS-487 was evaluated by RP-HPLC. The results are shown in Figure 8. The graph before the reaction in Figure 8 shows a peak (10.200 min) derived from DBCO-KS-487, while the peak (10.248 min) derived from DBCO-KS-487 almost disappeared in the graph after the reaction in Figure 8. As shown in the graph after the reaction in Figure 8, new peaks (7.904 min and 9.285 min) emerged. From the results shown in Figure 8 (before and after the reaction), it was shown that the DBCO group added to the cyclic peptide retains the activity for click reaction.

Industrial Applicability

[0108] The peptide according to the present invention has LRP1 binding activity, and when the peptide binds to LRP1, it can pass through a barrier tissue that separates peripheral tissue and brain tissue via the RMT of LRP1, for example, the blood-brain barrier (BBB). Therefore, the peptide according to the present invention is considered to be useful as a means for transporting any molecule to the brain tissue by combining it with any molecule.

Claims

1. The following formula (1): X Ｎ -X ５ -X ６ -X ７ -X ８ -X ９ -X １０ -X １１ -X １２ -X １３ -X １４ -X １５ (1) (wherein X Ｎ is an optional amino acid residue of 1 to 4, X ５ and X １５ each independently represent serine, threonine, cysteine, D-cysteine, or proline, X ６ represents serine, homoserine, threonine, allothreonine, 2,3-diaminopropionic acid, 2,4-diaminobutyric acid, ornithine, lysine, arginine, proline, cis-4-hydroxy-proline, or trans-4-hydroxy-proline, X ７ represents histidine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, X ８ represents ornithine, lysine, homolysine, arginine, or homoarginine, X ９ represents methionine, norleucine, lysine, arginine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, X １０ represents methionine, leucine, norleucine, isoleucine, valine, lysine, or argin, X １１ and X １４ each independently represent methionine, leucine, norleucine, isoleucine, valine, 2-aminoheptanoic acid, or 2-aminooctanoic acid, X １２ represents alanine, D-alanine, or 2-aminoisobutyric acid, X １３ represents glycine, alanine, asparagine, aspartic acid, glutamine, or glutamic acid, the N-terminal amino group and the C-terminal carboxy group may be modified or deleted, X ５ and X １５ When involved in cyclization, it is cysteine or D-cysteine, and a covalent bond is formed between the side-chain -SH groups of each through a linker of a disulfide bond, a methylene group, an acetylmethylene group, an ethylene group, or a propylene group, whereby the peptide of formula (1) may have one cyclic structure in the molecule.) A linear peptide or cyclic peptide having the amino acid sequence represented by, or a pharmacologically acceptable salt thereof.

2. The following formula (2): X Ｎ -X ５ -X ６ -X ７ -X ８ -X ９ -X １０ -X １１ -X １２ -X １３ -X １４ -Pro-X １６ (3) (wherein X Ｎ is 1 to 4 arbitrary amino acid residues, and X ５ and X １６ each independently represent serine, threonine, cysteine, D-cysteine, or proline, and X ６ represents serine, homoserine, threonine, allothreonine, 2,3-diaminopropionic acid, 2,4-diaminobutyric acid, ornithine, lysine, arginine, proline, cis-4-hydroxy-proline, or trans-4-hydroxy-proline, X ７ represents histidine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, X ８ represents ornithine, lysine, homolysine, arginine, or homoarginine, X ９ represents methionine, norleucine, lysine, arginine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, X １０ represents methionine, leucine, norleucine, isoleucine, valine, lysine, or argin, X １１ and X １４ each independently represent methionine, leucine, norleucine, isoleucine, valine, 2-aminoheptanoic acid, or 2-aminooctanoic acid, X １２ represents alanine, D-alanine, or 2-aminoisobutyric acid, X １３ represents glycine, alanine, asparagine, aspartic acid, glutamine, or glutamic acid, and the N-terminal amino group and the C-terminal carboxy group may be modified or deleted, X ５ and X １６ When involved in cyclization, it is cysteine or D-cysteine, and a covalent bond is formed between the side-chain -SH groups of each through a linker of a disulfide bond, a methylene group, an acetylmethylene group, an ethylene group, or a propylene group, whereby the peptide of formula (3) may have one cyclic structure in the molecule.) A linear peptide or cyclic peptide having the amino acid sequence represented by, or a pharmacologically acceptable salt thereof.

3. The following formula (3): X Ｎ -X ５ -X ６ -X ７ -X ８ -X ９ -X １０ -X １１ -X １２ -X １３ -X １４ -Pro-X １６ (3) (wherein X Ｎ is one to four arbitrary amino acid residues, X ６ and X １６ each independently represent serine, threonine, cysteine, D-cysteine, or proline, X ５ represents serine, homoserine, threonine, allothreonine, 2,3-diaminopropionic acid, 2,4-diaminobutyric acid, ornithine, lysine, arginine, proline, cis-4-hydroxy-proline, or trans-4-hydroxy-proline, X ７ represents histidine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, X ８ represents ornithine, lysine, homolysine, arginine, or homoarginine, X ９ represents methionine, norleucine, lysine, arginine, tyrosine, O-methyl-tyrosine, phenylalanine, 4-amino-phenylalanine, 4-fluoro-phenylalanine, or 4-chloro-phenylalanine, X １０ represents methionine, leucine, norleucine, isoleucine, valine, lysine, or argin, X １１ and X １４ each independently represent methionine, leucine, norleucine, isoleucine, valine, 2-aminoheptanoic acid, or 2-aminooctanoic acid, X １２ represents alanine, D-alanine, or 2-aminoisobutyric acid, X １３ represents glycine, alanine, asparagine, aspartic acid, glutamine, or glutamic acid, the N-terminal amino group and the C-terminal carboxy group may be modified or deleted, X ６ and X １６ When involved in cyclization, it is cysteine or D-cysteine, and a covalent bond is formed between the side-chain -SH groups of each through a linker of a disulfide bond, a methylene group, an acetylmethylene group, an ethylene group, or a propylene group, whereby the peptide of formula (3) may have one cyclic structure in the molecule.) A linear peptide or cyclic peptide having the amino acid sequence represented by, or a pharmacologically acceptable salt thereof.

4. X Ｎ The linear peptide or cyclic peptide according to any one of claims 1 to 3, or a pharmacologically acceptable salt thereof, wherein X is Lys-Gly-Thr-Pro or Gly-Thr-Pro.

5. A linear or cyclic peptide having LRP1 binding activity, or a pharmacologically acceptable salt thereof, comprising an amino acid sequence in which one or several amino acids are deleted, added, and / or substituted in the amino acid sequence according to any one of Claims 1 to 3.

6. A derivative and / or modified form of the peptide according to any one of Claims 1 to 3.

7. The derivative or modified form of the peptide according to Claim 6, wherein the derivative or modified form has an azide group, an alkyne group, a cyclooctyne group or a tetrazine group directly or via a linker at the N-terminus, C-terminus, or side chain of the amino acid.

8. A medicament, diagnostic agent, and / or reagent comprising the peptide according to any one of Claims 1 to 3, or a derivative or modified form thereof.

9. A peptide-drug conjugate in which the derivative or modified form of the peptide according to Claim 6 is linked directly or via a linker to a drug.

10. The peptide-drug conjugate according to Claim 9, wherein the drug is an antibody, a nucleic acid, or a bioactive peptide.

11. A polynucleotide encoding an amino acid sequence composed only of natural amino acids among the amino acid sequences according to any one of Claims 1 to 3, or a polynucleotide having a nucleotide sequence with 70% or more sequence identity to the polynucleotide and encoding a peptide having LRP1 binding activity.

12. An expression vector containing the polynucleotide according to Claim 11.

13. A cell, yeast, bacterium, virus, liposome, or nanoparticle presenting the amino acid sequence according to any one of Claims 1 to 3.

14. The following formula (6): X １ -X ２ -X ３ -X ４ -X Ｌ -X ７ -X ８ -X ９ -X １０ -X １１ -X １２ -X １３ -X １４ -X Ｍ (6) (X Ｌ is X ５ - X ６ and X Ｍ is X １５ - X １６ and when involved in the cyclization within the peptide molecule, each independently represents an amino acid residue having an amino group, a carboxy group, a thiol group, an allyl group, an alkynyl group, an azide group, or a halogen atom, or a derivative thereof; when not involved in the cyclization within the peptide molecule, X ５ , X ６ , or X １５ each independently represents an arbitrary amino acid residue, or a derivative thereof; X １６ represents an arbitrary amino acid residue, a derivative thereof, or a deletion; X ７ represents an amino acid residue having a hydrocarbon group which may have a substituent, an amino acid residue having an aromatic carbocyclic group which may have a substituent, an amino acid residue having an aromatic heterocyclic group which may have a substituent, or a derivative thereof; X ８ represents an amino acid residue having a positive charge, or a derivative thereof; X １１ , X １２ and X １４ each independently represents an amino acid residue having a hydrocarbon group which may have a substituent, or a derivative thereof; X １３ represents an amino acid residue having a hydrocarbon group which may have a substituent, an amino acid residue having a negative charge, or a derivative thereof; X ９ , X １０ each independently represents an arbitrary amino acid residue, or a derivative thereof; X １ , X ２ , X ３ , X ４ each independently represents an arbitrary amino acid residue, a derivative thereof, or a deletion; X Ｌ and X Ｍ may form a direct or indirect covalent bond via a linker between them, and the peptide of formula (6) may have one cyclic structure in the molecule. In that case, X Ｌ and X Ｍ The covalent bond between them may be any of the bonds between the main chains, between the main chain and the side chain, between the side chain and the main chain, or between the side chains, and the N-terminal amino group and the C-terminal carboxy group may be modified or deleted.) A linear or cyclic peptide consisting of the amino acid sequence represented by the formula, or a pharmacologically acceptable salt thereof.

15. X ５ and X １５ the bond between the Cα carbon atoms of, X ６ and X １５ the bond between the Cα carbon atoms of, X ５ and X １６ the bond between the Cα carbon atoms of, or X ６ and X １６ the bond between the Cα carbon atoms of is each independently represented by the following formula (7): Cα Ｎ -(CH ２ ) Ａ -Z-(CH ２ ) Ｂ -Cα Ｍ (7) (wherein, N is 4 or 5, M is 14 or 15, A and B each independently represent an integer of 0 to 2, the sum of A and B is an integer of 0 to 4, and Z is S-S, CH ２ -CH ２ , S-CH ２ , CH ２ -S, O-CH ２ , CH ２ -O, CH=CH, NH-C(=O), N(CH ３ )-C(=O), NH-C(=S), N(CH ３ )-C(=S), O-C(=O), C(=O)-NH, C(=O)-N(CH ３ ), C(=S)-NH, C(=S)-N(CH ３ ), C(=O)-O, CH ２ -CH ２ -CH ２ , S-CH ２ -CH ２ , CH ２ -CH ２ -S, CH ２ -S-CH ２ , O-CH ２ -CH ２ , CH ２ -CH ２ -O, CH ２ -O-CH ２ , S-CH ２ -S, S-C(=CH ２ )-S, S-(C(=O)-CH ３ ), S-(CH ２ ) ２ -S, S-(CH ２ ) ３ -S, S-CH ２ -C(=O)-CH ２ -S, S-CH ２ -C(=CH ２ )-CH ２ -S, S-(CH ２ ) ４ -S, S-CH ２ -CH=CH-CH ２ -S, S-CH ２ -C ６ H ４ -CH ２ -S (the bonding site of the methylene group to the phenylene ring may be any of ortho, meta and para), S-CH ２ -C(=O)-CH ２ -S, or S-CH ２ -C(=CH ２ )-CH ２ -S. The linear or cyclic peptide according to Claim 14, or a pharmacologically acceptable salt thereof, comprising the structure represented by the formula.

16. X ８ is 2,3-diaminopropionic acid, 2,4-diaminobutyric acid, ornithine, lysine, homolysine, arginine, homoarginine, their D-form amino acids, their N-methylated amino acids, their α-methylated amino acids, or derivatives thereof, the linear peptide and cyclic peptide according to claim 14 or 15, or a pharmacologically acceptable salt thereof.

17. X １２ The linear and cyclic peptides according to claim 14 or 15, or a pharmacologically acceptable salt thereof, wherein X is alanine, D-alanine, 2-aminoisobutyric acid, 2-aminobutanoic acid, D-2-aminobutanoic acid, isovaline, D-isovaline, valine, D-valine, isoleucine, D-isoleucine, leucine, D-leucine, their N-methylated amino acids, their α-methylated amino acids, or derivatives thereof.

18. X １１ and X １４ are each independently methionine, the following formula (8): 【Chemical 1】 (In the formula, the wavy line represents the attachment point to the carbonyl group or nitrogen atom forming the amide bond of the main chain, and R １ , R ２ and R ６ each independently represent a hydrogen atom or a methyl group, and R ３ , R ４ , R ５ each independently represent a hydrogen atom, a methyl group, or a halogen atom (F, Cl, Br, I), and n represents any integer from 0 to 10); or The following formula (9) 【Chemical 2】 (In the formula, Z represents C or N, and R ６ represents a hydrogen atom or a methyl group, and n represents an integer of any one of 1 to 6); or The following formula (10) 【Chemical Formula 3】 (In the formula, the wavy line represents the bonding site with the carbonyl group or nitrogen atom forming the amide bond of the main chain, Z represents C or N, R ６ represents a hydrogen atom or a methyl group, and n represents an integer of any one of 1 to 6.) The linear or cyclic peptide according to Claim 14 or 15, or a pharmacologically acceptable salt thereof, which is an amino acid residue represented by the formula or a derivative thereof.

19. X ７ is represented by the following formula (8): [Chemical Formula 4] (wherein the wavy line represents the attachment point to the carbonyl group or nitrogen atom forming the amide bond of the main chain, R １ , R ２ and R ６ each independently represents a hydrogen atom or a methyl group, and R ３ , R ４ , R ５ each independently represents a hydrogen atom, a methyl group, or a halogen atom (F, Cl, Br, I), and n represents any integer from 0 to 10); or The following formula (9) [Chemical Formula 5] (wherein, Z represents C or N, and R ６ represents a hydrogen atom or a methyl group, and n represents an integer of 1 to 6); or The following formula (10) 【Chemical Formula 6】 (In the formula, the wavy line represents the bonding site with the carbonyl group or nitrogen atom forming the amide bond of the main chain, Z represents C or N, and R ６ represents a hydrogen atom or a methyl group, and n represents an integer of any one of 1 to 6.); or The following formula (11) 【Chemical Formula 7】 (wherein, R １２ represents a 2-thienyl group, 3-thienyl group, 2-pyridyl group, 3-pyridyl group, 4-pyridyl group, 2-thiazolyl group, 4-thiazolyl group, 2-oxazolyl group, 4-oxazolyl group, 1-pyrazolyl group, 1H-imidazol-4-yl group, 1-imidazolyl group, 1,2,3-triazol-1-yl group, or 1,2,4-triazol-1-yl group, and R ６ represents a hydrogen atom or a methyl group); or The following formula (12) 【Chemical Formula 8】 (In the formula, the wavy line represents the attachment point to the carbonyl group or nitrogen atom that forms the amide bond of the main chain, and R ７ , R ８ , R ９ , R １０ , R １１ each independently represents a hydrogen atom, a hydroxy group, a methoxy group, a methyl group, a tert-butyl group, a methyl halide group, or a halogen atom (F, Cl, Br, I), and R ６ represents a hydrogen atom or a methyl group.); or The following formula (13) 【Chemical Formula 9】 (wherein R １３ , R １４ , R １５ , R １６ each independently represents a hydrogen atom, a hydroxy group, a methoxy group, a methyl group, a tert-butyl group, a methyl halide group, or a halogen atom (F, Cl, Br, I), and R ６ represents a hydrogen atom or a methyl group.) The linear and cyclic peptides according to claim 14 or 15, or a pharmacologically acceptable salt thereof, which are amino acid residues represented by , or derivatives thereof.

20. X １３ is glycine, alanine, aspartic acid, glutamic acid, asparagine, glutamine, D-alanine, D-aspartic acid, D-glutamic acid, D-asparagine, D-glutamine, their N-methylated amino acids, their α-methylated amino acids, or derivatives thereof, the linear peptide and cyclic peptide according to claim 14 or 15, or a pharmacologically acceptable salt thereof.

21. X that is not involved in the cyclization within the peptide molecule Ｌ and X Ｍ are each independently alanine, isovaline, norvaline, serine, homoserine, threonine, allothreonine, O-methylated-serine, O-methylated-homoserine, O-methylated-threonine, O-methylated-allothreonine, 2,3-diaminopropionic acid, 2,4-diaminobutyric acid, ornithine, lysine, arginine, proline, cis-4-hydroxy-proline, trans-4-hydroxy-proline, azetidine-2-carboxylic acid, pipecolic acid, D-form serine, D-form homoserine, D-form threonine, D-form allothreonine, D-form O-methylated-serine, D-form O-methylated-homoserine, D-form O-methylated-threonine, D-form O-methylated-allothreonine, D-form 2,3-diaminopropionic acid, D-form 2,4-diaminobutyric acid, D-form ornithine, D-form lysine, D-form arginine, D-form proline, D-form cis-4-hydroxy-proline, D-form trans-4-hydroxy-proline, D-form azetidine-2-carboxylic acid, D-form pipecolic acid, their N-methylated amino acids, their α-methylated amino acids, or derivatives thereof, the linear peptide and cyclic peptide according to claim 14 or 15, or a pharmacologically acceptable salt thereof.

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