Alpha helix cell-penetrating peptide multimer, preparation method therefor and use therefor

Patent metadata

Field	Value
Publication number	US-2016229894-A1
Application number	US-201415030051-A
Country	US
Kind code	A1
Filing date	Oct 17, 2014
Priority date	Oct 17, 2013
Publication date	Aug 11, 2016
Grant date	—

Abstract

The present invention relates to an α-helical cell-penetrating peptide multimer, a preparation method thereof and the use thereof, and more particularly, to a peptide multimer comprising a plurality of amphipathic peptides, a method for preparing the peptide multimer, a composition for preventing or treating HIV, which comprises the peptide multimer as an active ingredient, and a composition for intracellular delivery of a biologically active substance, which comprises the peptide multimer and the biologically active substance.

First claim

1 . A peptide multimer comprising a plurality of homogeneous or heterogeneous α-helical amphipathic peptides.

2 . The peptide multimer of claim 1 , wherein further comprising a linker located at one or more amino acid positions selected from the group consisting of i, i+3, i+4, i+7, i+8, i+10 and i+11 (where i is an integer).

3 . The peptide multimer of claim 2 , wherein the linker is located at two or more amino acid positions selected from the group consisting of i, i+3, i+4, i+7, i+8, i+10 and i+11 (where i is an integer).

4 . The peptide multimer of claim 1 , wherein the amphipathic peptide comprises one or more hydrophilic amino acids selected from the group consisting of arginine, lysine and histidine.

5 . The peptide multimer of claim 1 , wherein the amphipathic peptide comprises one or more hydrophobic amino acids selected from the group consisting of leucine, valine, tryptophan, phenylalanine, tyrosine and isoleucine.

6 . The peptide multimer of claim 1 , wherein the peptide comprises 5-50 amino acids.

7 . The peptide multimer of claim 1 , wherein the peptide comprises 7-23 amino acids.

8 . The peptide multimer of claim 1 , wherein one to three hydrophilic amino acids and hydrophobic amino acids are arrayed respectively in the peptide.

9 . The peptide multimer of claim 1 , wherein the amphipathic peptide comprises one or more of seven amino acid sequences represented by the following formulas:

XYXXYYX YXYYXXY

XYYXYYX YXXYXXY

XYYXXYX YXXYYXY

XYYXXYY YXXYYXX

XXYXXYY YYXYYXX

XXYYXYY YYXXYXX

XXYYXXY YYXXYYX

wherein X is a hydrophilic amino acid and Y is a hydrophobic amino acid.

10 . The peptide multimer of claim 1 , wherein the hydrophilic amino acid in the amphipathic peptide comprises one or more positively charged amino acid residue selected from the group consisting of arginine, lysine and histidine, in an amount of 33% or more.

11 . The peptide multimer of claim 1 , wherein the hydrophobic amino acid of the amphipathic peptide comprises one or more residues selected from the group consisting of leucine, tryptophan, valine, phenylalanine, tyrosine and isoleucine, in an amount of 25% or more.

12 . The peptide multimer of claim 1 , wherein the amphipathic peptide comprises a sequence represented by the following SEQ ID NO: 11:
KLLKLLK (SEQ ID NO: 11).

13 . The peptide multimer of claim 1 , wherein the peptide comprises one or more of amino acid sequences represented by the following formulas:

CYYXXYXCYYXXYXZW (1)

XYYCXYXXYYCXYXZW (2)

XYYXCYXXYYXCYXZW (3)

XYYXXYCXYYXXYCZW (4); 

and

XYYXXYXCYYXXYXCW (5)

wherein X, Z and W are hydrophobic amino acids, Y is a hydrophilic amino acid, and C is cysteine.

14 . The peptide multimer of claim 13 , wherein the peptide comprises at least one sequence selected from the group consisting of SEQ ID NOs: 1 to 10.

15 . The peptide multimer of claim 1 , wherein the α-helical content of the peptide is at least 80% in a cell membrane condition in which trifluoroethanol and buffer are mixed at a ratio of 1:1.

16 . The peptide multimer of claim 1 , wherein the linker comprises a covalent bond that connects between peptides.

17 . The peptide multimer of claim 16 , wherein the covalent bond is at least one selected from the group consisting of a disulfide bone between cysteines, a maleimide bond, an ester bond, a thioether bond, and a bond formed by a click reaction.

18 . The peptide multimer of claim 1 , wherein the multimer is a dimer, a trimer, or a tetramer.

19 . A method for preparing a peptide multimer, comprising the steps of:
constructing α-helical peptides comprising hydrophilic and hydrophobic amino acids; selecting a plurality of homogeneous or heterogeneous α-helical peptides; and connecting the plurality of selected α-helical peptides at one or more amino acid positions selected from the group consisting of i, i+3, i+4, i+7, i+8, i+10 and i+11 (where i is an integer).

20 . The method of claim 19 , wherein the plurality of selected α-helical peptides is connected to each other at two or more amino acid positions selected from the group consisting of i, i+3, i+4, i+7, i+8, i+10 and i+11 (where i is an integer).

21 . The method of claim 19 , wherein the plurality of α-helical peptides is connected to each other by a covalent bond that connects between peptides.

22 . The method of claim 21 , wherein the covalent bond is at least one selected from the group consisting of a disulfide bone between cysteines, a maleimide bond, an ester bond, a thioether bond, and a bond formed by a click reaction.

23 . A method for preventing or treating HIV, comprising administering a composition that comprises the α-helical peptide multimer of claim 1 as an active ingredient to a subject in need.

24 . The method of claim 23 , wherein the peptide multimer binds to the TAR (trans-activating region) of HIV.

25 . A method of delivering a biologically active substance intracellularly, comprising using peptide multimer of claim 1 and the biologically active substance.

26 . The method of claim 24 , where the biologically active substance is DNA, RNA, siRNA, an aptamer, a protein, an antibody or a low molecular compound.

Assignees

• Univ Seoul Nat R & Db Found

Inventors

• Yu Jaehoon
• Lee Yan
• Hyun Soonsil
• Jang Sangmok

Classifications

• C07K2319/00

  Fusion polypeptide · CPC title

• C12N15/113

  Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; {Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing (when used in plants C12N15/8218)} · CPC title

• A61K38/00

  Medicinal preparations containing peptides (peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00) · CPC title

• A61K47/64

  Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent (peptidic linkers A61K47/65) · CPC title

• A61K47/42

  Proteins; Polypeptides; Degradation products thereof; Derivatives thereof, e.g. albumin, gelatin or zein (oligopeptides having up to five amino acids {A61K47/183}; polyamino acids A61K47/34) · CPC title