WO2019085926A1 - Polymyxin analogue and preparation method therefor - Google Patents

Abstract

The present invention belongs to the field of medicines, and relates to a polymyxin analogue having high antibacterial activity, and a preparation method therefor and an application thereof. The polymyxin analogue has a broad-spectrum and high-efficiency bactericidal effect on drug-resistant bacteria and fungi.

Description

Polymyxin analog and preparation method thereof Technical field

The invention belongs to the field of medicine, relates to an antibacterial peptide and a preparation method and application thereof, and particularly relates to a group of polymyxin analogs having high antibacterial activity and a synthetic preparation and application thereof.

Background technique

With the increasing use of antibiotics, the problem of bacterial resistance has become increasingly prominent. Bacterial resistance is becoming a major problem that threatens human health, especially multi-drug resistance to almost all existing antibiotics. The emergence of bacteria has made this problem even more prominent. However, there is still no major breakthrough in the development of new antibiotics capable of killing multi-drug resistant bacteria. At present, polymyxin, which has the strongest antibacterial ability, is regarded as the "last line of defense" against resistant bacteria, which makes it necessary to use polymyxin as an antibacterial agent.

Polymyxin is a class of cyclic peptide antibiotics produced by Bacillus polymyxa discovered in 1947. It consists of various components, including polymyxin A, polymyxin B, polymyxin C, and more. Colistin D, polymyxin E, usually have a narrow antibacterial spectrum and are only resistant to Gram-negative bacteria. It is commonly used clinically to treat infections caused by Gram-negative bacteria. However, due to its severe nephrotoxicity and neurotoxicity, it is gradually replaced by other antibiotics (aminoglycosides, etc.) with low toxicity and good antibacterial effect. Until the emergence of multi-drug resistant bacteria in the past 20 years, polymyxin has once again entered the field of vision as a clinical therapeutic drug. The polymyxin products currently on the market are mainly polymyxin B sulfate, polymyxin E sulfate and sodium polymyxin E methanesulfonate. The antibacterial activity of polymyxin B and polymyxin E is slightly different. In general, the activity of polymyxin B is stronger than that of polymyxin E (Chen Guanrong, the clinical application of polymyxin and the response to super bacteria [J]. Medical Herald, 2011, 30 (2): 135-140).

Natural polymyxin is generally composed of a fatty acyl chain and a cyclic heptapeptide linked by a linear tripeptide, wherein the 4-position amino acid L-Dab (α, γ-diaminobutyric acid) is condensed with the 10-position amino acid L-Thr to form a heptapeptide. ring. The structures of polymyxin B and polymyxin E are very similar, with only differences in the 6 amino acids. Among them, the 6-position amino acid of polymyxin B is D-Phe, and the 6-position amino acid of polymyxin E is D-Leu (Cui, et.al. Research Development of Polymyxins [J]. World Notes on Antibiotics, 2015 , 36(5): 205-210).

However, polymyxin nephrotoxicity and neurotoxicity remain a major concern. Therefore, the search for polymyxin analogs with low toxicity and high bactericidal activity has become an urgent need.

Summary of the invention

The invention relates to a polypeptide and a preparation method and application thereof, in particular to a group of polymyxin analogs having high antibacterial activity, and synthetic preparation and application thereof.

In one aspect, the application relates to polypeptides and pharmaceutically acceptable salts thereof, which polypeptides have at least 70% sequence identity to the sequences of Formula I. In some embodiments, the polypeptide has the following sequence: R1-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-Xaa7-Xaa8-Xaa9-Xaa10-Xaa11-Xaa12 (Formula I), wherein R1 is: Fat a straight or branched C ₆ -C ₂₀ acyl group or deletion; Xaa1 is: Leu, Ala, AEEAc, Phe, IPhe, BrPhe, or 3FPhe, or a deletion; Xaa2 is: Leu, Ala, Arg, or AEEAc , or deletion; Xaa3 is: Dab, Ala, Leu, or Orn, or deletion; Xaa4 is: Thr, Ala, Dab, or Lys; Xaa5 is: Dab, Lys, Orn, or Ala, or deletion; Xaa6 is: Lys , Glu, or Dab; Xaa7 is: Dab, Ala, Lys, or Orn, or a deletion; Xaa8 is: D-Leu, D-Phe, Lys, or D-Ala; Xaa9 is: Leu, or Ala; Xaa10 is: Dab, Leu, Orn, Ala, or D-Leu, or a deletion; Xaa11 is: Dab, Leu, Orn, Ala, or D-Leu, or a deletion; Xaa12 is: Glu, Lys, or Asp.

In some aspects, the polypeptide is a cyclic peptide. In certain embodiments, the polypeptide is formed by the condensation of Xaa6 with a side chain group of Xaa12 to form an amide linkage, and optionally, the pendant group comprises an amino group and/or a carboxyl group. Alternatively, the polypeptide is obtained by dehydration of an amino group with a carboxyl group to form an amide bond.

In some embodiments, the R 1 is selected from the group consisting of heptanoyl, methylheptanoyl, octanoyl, methyloctanoyl, decanoyl, methyl decanoyl, decanoyl, methyl decanoyl, lauroyl, myristoyl, Palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, preferably n-octanoyl, decanoyl, lauroyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, more preferably n-octanoyl or a deletion Xaa1 is preferably Leu or deleted, Xaa2 is preferably Leu, Xaa3 is preferably Dab, Xaa3 is preferably Dab, Xaa4 is preferably Thr, Xaa4 is preferably Dab, Xaa6 is preferably Dab, Xaa6 is preferably Lys or Dab, Xaa7 is preferably Dab, Xaa8 is preferably D-Phe or D-Leu, Xaa9 is preferably Leu, Xaa10 is preferably Dab Xaa11 is preferably Dab, and/or Xaa12 is preferably Glu.

In some aspects, the polypeptide has the formula:

Wherein R1 is: an aliphatic straight-chain or branched C ₆ -C ₂₀ acyl group or a deletion; Xaa1 is: Leu, Ala, AEEAc, Phe, IPhe, BrPhe, 3FPhe or a deletion; Xaa2 is: Leu, Ala, Arg, AEEAc or deletion; Xaa3 is: Dab, Ala, Leu, Orn or deletion; Xaa4 is: Thr, Ala, Dab or Lys; Xaa5 is: Dab, Lys, Orn, Ala or deletion; Xaa6 is: Lys or Dab; Xaa7 is: Dab, Ala, Lys, Orn or deletion; Xaa8 is: D-Leu, D-Phe, Lys or D-Ala; Xaa9 is: Leu or Ala; Xaa10 is: Dab, Leu, Orn, Ala, D- Leu or deletion; Xaa11 is: Dab, Leu, Orn, Ala, D-Leu or deletion. In some embodiments, the R1 is selected from the group consisting of heptanoyl, methylheptanoyl, octanoyl, methyloctanoyl, decanoyl, methyldecanoyl, decanoyl, methyldecanoyl, lauroyl, myristoyl , palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, preferably n-octanoyl, decanoyl, lauroyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, more preferably n-octanoyl or Deletion; Xaa1 is preferably Leu or deleted, Xaa2 is preferably Leu, Xaa3 is preferably Dab, Xaa3 is preferably Dab, Xaa4 is preferably Thr, Xaa5 is preferably Dab, Xaa6 is preferably Lys or Dab, Xaa7 is preferably Dab, Xaa8 is preferably D-Phe or D-Leu, Xaa9 is preferably Leu, Xaa10 is preferred Dab, Xaa11 is preferably Dab.

In some aspects, the polypeptide has the formula:

Wherein R1 is: an aliphatic straight-chain or branched C ₆ -C ₂₀ acyl group or a deletion; Xaa1 is: Leu, Ala, AEEAc, Phe, IPhe, BrPhe, 3FPhe or a deletion; Xaa2 is: Leu, Ala, Arg, AEEAc or deletion; Xaa3 is: Dab, Ala, Leu, Orn or deletion; Xaa4 is: Thr, Ala, Dab, Lys; Xaa5 is: Dab, Lys, Orn, Ala or deletion; Xaa7 is: Dab, Ala, Lys, Orn or deletion; Xaa8 is: D-Leu, D-Phe, Lys, D-Ala; Xaa9 is: Leu, Ala; Xaa10 is: Dab, Leu, Orn, Ala, D-Leu or deletion; Xaa11 is: Dab, Leu, Orn, Ala, D-Leu or deletion; Xaa12 is: Glu, Asp. In some embodiments, the R1 is selected from the group consisting of heptanoyl, methylheptanoyl, octanoyl, methyloctanoyl, decanoyl, methyldecanoyl, decanoyl, methyldecanoyl, lauroyl, myristoyl , palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, preferably n-octanoyl, decanoyl, lauroyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, more preferably n-octanoyl or Xaa1 is preferably Leu or deleted, Xaa2 is preferably Leu, Xaa3 is preferably Dab, Xaa3 is preferably Dab, Xaa4 is preferably Thr, Xaa5 is preferably Dab, Xaa7 is preferably Dab, Xaa7 is preferably Dab, Xaa8 is preferably D-Phe or D-Leu, Xaa9 is preferably Leu, Xaa10 is preferably Dab, Xaa11 is preferably Dab, And/or Xaa12 is preferably Glu.

In some aspects, the polypeptide has the formula:

Wherein R1 is: an aliphatic straight-chain or branched C ₆ -C ₂₀ acyl group or a deletion; Xaa1 is: Leu, Ala, AEEAc, Phe, IPhe, BrPhe, 3FPhe or a deletion; Xaa2 is: Leu, Ala, Arg, AEEAc or deletion; Xaa3 is: Dab, Ala, Leu, Orn or deletion; Xaa4 is: Thr, Ala, Dab or Lys; Xaa5 is: Dab, Lys, Orn, Ala or deletion; Xaa7 is: Dab, Ala, Lys, Orn or deletion; Xaa8 is: D-Leu, D-Phe, Lys or D-Ala; Xaa9 is: Leu or Ala; Xaa10 is: Dab, Leu, Orn, Ala, D-Leu or deletion; Xaa11 is: Dab, Leu, Orn, Ala, D-Leu or missing. In some embodiments, the R1 is selected from the group consisting of heptanoyl, methylheptanoyl, octanoyl, methyloctanoyl, decanoyl, methyldecanoyl, decanoyl, methyldecanoyl, lauroyl, myristoyl , palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, preferably n-octanoyl, decanoyl, lauroyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, more preferably n-octanoyl or Deletion; Xaa1 is preferably Leu or deleted, Xaa2 is preferably Leu, Xaa3 is preferably Dab, Xaa4 is preferably Dab, Xaa4 is preferably Thr, Xaa5 is preferably Dab, Xaa7 is preferably Dab, Xaa8 is preferably D-Phe or D-Leu, Xaa9 is preferably Leu, Xaa10 is preferably Dab, and Xaa11 is preferably Dab.

In one embodiment, the polypeptide is a peptide or cyclic peptide consisting of the sequence represented by R1-Leu-Dab-Thr-Dab-Xaa6-Dab-Xaa8-Leu-Dab-Dab-Glu, wherein R1 is preferably n-octanoyl , decanoyl, lauroyl, palmitoyl, 17-carboxyheptadecanoyl or a deletion, more preferably n-octanoyl or a deletion; Xaa6 is preferably Lys or Dab, and Xaa8 is preferably D-Phe or D-Leu. In another embodiment, the polypeptide is a peptide or cyclic peptide consisting of the sequence represented by R1-Dab-Thr-Dab-Xaa6-Dab-Xaa8-Leu-Dab-Dab-Glu, wherein R1 is preferably n-octane Acyl, decanoyl, lauroyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, more preferably n-octanoyl or a deletion; Xaa6 is preferably Lys or Dab, and Xaa8 is preferably D-Phe or D-Leu.

In some specific embodiments, the polypeptide comprises the sequence H ₂ N-Leu-Dab-Thr-Dab-Lys-Dab-D-Phe-Leu-Dab-Dab-Glu, or the sequence H ₂ N-Dab- A peptide or cyclic peptide of Thr-Dab-Lys-Dab-D-Phe-Leu-Dab-Dab-Glu.

In some embodiments, the polypeptide of the invention comprises at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95 with the above polypeptide. %, 96%, 97%, 98%, 99% sequence identity amino acid sequence. In other embodiments, the polypeptide of the invention has an amino acid sequence of at least 1, 2, 3, 4, 5, 6 amino acid additions, substitutions, and/or deletions as compared to the polypeptide described above. In other embodiments, the polypeptide of the invention has at least 1, 2, 3, 4, 5, 6 amino acid substitutions compared to the above polypeptide, wherein the substitution can be a conservative substitution, And/or approximate replacement. In other embodiments, the polypeptide of the invention is a truncated amino acid of six, seven, eight, nine, ten, eleven amino acid lengths of the above polypeptide.

In some aspects, the polypeptides provided herein are a low toxicity polypeptide.

In one aspect, the application also provides the use of the aforementioned polypeptide or a pharmaceutically acceptable salt thereof. The polypeptides of the present invention can be used for bacteriostatic, antibacterial or antibacterial substances.

In some examples, the antimicrobial substance is an antimicrobial or antimicrobial agent. In some examples, the bacterium is a bacterium or a fungus. It is particularly surprising that the polypeptides of the invention can be used against anti-negative or anti-resistant bacteria, or for the preparation of anti-negative or anti-resistant bacteria. In some examples, the bacterium, fungus or drug resistant bacterium is Acinetobacter baumanii, drug resistant Acinetobacter baumanii, Pseudomonas aeruginosa, resistant One or more of Pseudomonas aeruginosa, Bacillus subtilis, Staphylococcus aureus, E. coli, and/or candida Albicans, More preferably, it is Pseudomonas aeruginosa, a drug-resistant Acinetobacter baumannii, and a drug-resistant Pseudomonas aeruginosa.

In another aspect, the invention relates to an antimicrobial substance. In some embodiments, the antimicrobial substance is an antimicrobial or antimicrobial agent. The antibacterial substance or the microbial agent of the present invention includes the polypeptide of the present invention, and one or more of a pharmaceutically acceptable salt and/or a carrier and an excipient.

In another aspect, the present application is directed to a pharmaceutical composition comprising a polypeptide of the present application, or a pharmaceutically acceptable salt thereof. In some embodiments, the pharmaceutical compositions of the present application further comprise a pharmaceutically acceptable excipient, diluent or carrier.

In another aspect, the present application relates to the use of the polypeptide or a pharmaceutically acceptable salt thereof for the manufacture of a medicament and/or a pharmaceutical composition for preventing or treating an infectious disease.

In another aspect, the present application relates to a method of treating a disease in a mammal comprising administering to a mammal in need of such treatment, preferably a human, a therapeutically effective amount of a polypeptide of the invention, or a pharmaceutically acceptable salt thereof, or a pharmaceutical combination thereof Things.

The above combination therapies can be administered in the following manner: (a) a single pharmaceutical composition comprising a polypeptide of the present application, at least one additional therapeutic agent described herein, and a pharmaceutically acceptable excipient, diluent or carrier; Or (b) two separate pharmaceutical compositions comprising (i) a first composition comprising a polypeptide of the present application and a pharmaceutically acceptable excipient, diluent or carrier, and (ii) comprising at least one An additional therapeutic agent and a second composition of a pharmaceutically acceptable excipient, diluent, or carrier. The pharmaceutical compositions can be administered simultaneously or sequentially and in any order.

In one aspect, the invention relates to a set of polypeptides or pharmaceutically acceptable salts thereof, the amino acid sequences of which are as follows:

R1-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-Xaa7-Xaa8-Xaa9-Xaa10-Xaa11-Xaa12 (Formula I).

In some embodiments, the polypeptide is a cyclic peptide having the amino acid sequence shown below:

Wherein R1 is: an aliphatic straight or branched C ₆ -C ₂₀ acyl group or a deletion;

Xaa1 is: Leu, Ala, AEEAc, Phe, IPhe, BrPhe, 3FPhe or deletion;

Xaa2 is: Leu, Ala, Arg, AEEAc or deletion;

Xaa3 is: Dab, Ala, Leu, Orn or missing;

Xaa4 is: Thr, Ala, Dab, Lys;

Xaa5 is: Dab, Lys, Orn, Ala or missing;

Xaa6 is: Lys, Glu, Dab;

Xaa7 is: Dab, Ala, Lys, Orn or missing;

Xaa8 is: D-leu, D-Phe, Lys, D-Ala;

Xaa9 is: Leu, Ala;

Xaa10 is: Dab, Leu, Orn, Ala, D-Leu or deletion;

Xaa11 is: Dab, Leu, Orn, Ala, D-Leu or deletion;

Xaa12 is: Glu, Lys, Asp;

As a preferred embodiment of the present invention, the R1 is selected from the group consisting of heptanoyl, methylheptanoyl, octanoyl, methyloctanoyl, decanoyl, methyldecanoyl, decanoyl, methyldecanoyl, lauroyl, meat Myristoyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, Xaa1 is preferably Leu or deleted, Xaa2 is preferably Leu, Xaa3 is preferably Dab, Xaa4 is preferably Thr, Xaa5 is preferably Dab, Xaa6 is preferably Lys or Dab, Xaa7 is preferably Dab, Xaa8 is preferably D-Phe or D-Leu, Xaa9 is preferably Leu, Xaa10 is preferably Dab, Xaa11 is preferably Dab, and Xaa12 is preferably Glu.

The amino acid residues of the polypeptides involved in the present invention include natural amino acids and unnatural amino acids, wherein the three-letter codes corresponding to the natural amino acids involved are shown in Table 1, the unnatural amino acids involved and the corresponding letter codes and The structure is shown in Table 2.

The amino acid according to the present invention is an L-form amino acid by default unless it is specifically limited.

In some embodiments, the polypeptide of the present invention is a cyclic peptide which is obtained by dehydration of an amino group (or a carboxyl group) of a Xaa6 and Xaa12 side chain with a carboxyl group (or an amino group) to form an amide bond.

In some embodiments, the peptides of the present invention may have a C-terminus either in the form of a carboxyl group or in the form of an amide, preferably in the form of an amide.

Table 1: Natural amino acid three letter code table

Table 2: Unnatural amino acid and fatty acid code and structure table

In a preferred embodiment of the invention, a cyclic peptide (ZAMP18) is disclosed, the amino acid sequence of which is:

A further preferred embodiment of the invention discloses a cyclic peptide with a fatty chain (ZAMP29) having an amino acid sequence of:

In another preferred embodiment of the invention, a group of cyclic peptides CTAMP-41, CTAMP-42, CTAMP-43, CTAMP-44, CTAMP-45, CTAMP-46, CTAMP-47, ZAMP-3, ZAMP-4 are disclosed. , ZAMP-5, ZAMP-6, ZAMP-7, ZAMP-8, ZAMP-9, ZAMP-10, ZAMP-11, ZAMP-12, ZAMP-13, ZAMP-14, ZAMP-15, ZAMP-16, ZAMP -17, ZAMP-19, ZAMP-20, ZAMP-21, ZAMP-22, ZAMP-23, ZAMP-24, ZAMP-25, ZAMP-26, ZAMP-27, ZAMP-28, ZAMP-30, ZAMP-31 , ZAMP-32, ZAMP-33, ZAMP-34, ZAMP-35, ZAMP-36, ZAMP-37, ZAMP-38, ZAMP-39, ZAMP-40 have the amino acid sequences:

In one aspect, the polypeptide of the invention may be a peptide having a linear primary structure, and in another aspect, the polypeptide of the invention may be a cyclic peptide. In some embodiments, the polypeptide of the invention comprises at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92 of the peptide or cyclic peptide shown in any of the above sequences. %, 93%, 94%, 95%, 96%, 97%, 98%, 99% sequence identity amino acid sequence. In other embodiments, the polypeptide of the invention comprises a addition, substitution and/or deletion of at least 1, 2, 3, 4, 5, 6 amino acids with a polypeptide as set forth in any of the above sequences. Amino acid sequence. In other embodiments, the polypeptide of the invention comprises a substitution of at least 1, 2, 3, 4, 5, 6 amino acids with a polypeptide as set forth in any of the above sequences, wherein the substitution may be Conservative substitutions, and/or approximate replacements. In other embodiments, the polypeptide of the invention is a continuous truncation of six, seven, eight, nine, ten, eleven amino acid lengths of the polypeptide shown in any of the above sequences.

In another aspect, the invention also provides a process for the preparation of the above peptide or cyclic peptide.

In some aspects, the method comprises: synthesizing the polypeptide.

In some aspects, the above polypeptides can be prepared using solid phase synthesis techniques, including:

(1) solid phase synthesis of a polypeptide on a resin;

(2) The product of the step (1) is cleaved with a strong acid; a side chain protecting group scavenger is added, filtered, and the polypeptide is precipitated with 5-20 volumes of an organic solvent, centrifuged, and the organic solvent is repeatedly washed and precipitated, and dried to obtain a crude peptide. .

Optionally, the step (1) comprises the following steps:

(a) soaking resin - removal of amino protecting group - washing - monitoring - coupling of amino acids - monitoring - washing - removal of amino protecting groups - sequential coupling of residual amino acids;

(b) The resin peptide is further cyclized.

Wherein, the amino protecting group refers to a chemical group introduced to protect the amino group involved in the condensation reaction. The amino protecting group includes, but is not limited to, t-butoxycarbonyl (Boc), benzyloxycarbonyl (Z) or 9-fluorenyl-methoxycarbonyl (Fmoc), preferably 9-fluorenyl-methoxycarbonyl (Fmoc).

In solid phase peptide synthesis, the side chain of some amino acids can be protected by introducing chemical groups. For example, Arg can be protected with pentamethylbenzofuran-5-sulfonyl (Pbf); Dab, Orn can be tert-butoxycarbonyl ( Boc) protection; Thr can be protected with tert-butyl (tBu). In the present invention, Fmoc-L-Dab(Boc)-OH has the structure shown below:

The protecting group includes, but is not limited to, a reasonable selection according to a specific situation.

The solvent used in the liquid phase environment of the step (a) is selected from the group consisting of dimethylformamide (DMF), dichloromethane (DCM), N-methylpyrrolidone (NMP), preferably DMF.

The removal of the amino protecting group in the step (a) requires the removal of the amino protecting group, and the removal agent of the amino protecting group is selected from piperidine (PIP) solution at a concentration of 10-40% (PIP/DMF). The time is 20-50 min; the preferred concentration is 20-25% (PIP/DMF) and the removal time is 25-35 min.

The coupling of the amino acid in the step (a) requires the addition of a coupling reagent consisting of a carbodiimide type reagent or a benzotriazole salt type reagent and 1-hydroxybenzotriazole (HOBt). .

The carbodiimide type reagent includes, but is not limited to, dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC) or N-diaminopropyl-N-ethylcarbodiimide. (EDC).

The benzotriazolium salt type reagent includes, but is not limited to, 2-(1H-benzotriazo L-1-yl)-1,1,3,3-tetramethyluronium tetrafluoroborate ( TBTU), O-benzotriazole-N,N,N',N'-tetramethylurea hexafluorophosphate (HBTU), benzotriazole-1-oxotris(dimethylamino) hexafluorophosphate Phosphorus (BOP) or benzotriazol-1-yl-oxytripyrrolidinylphosphonium hexafluorophosphate (PyBOP).

The coupling reagent is preferably diisopropylcarbodiimide (DIC) and 1-hydroxybenzotriazole (HOBt), or 2-(1H-benzotriazo L-1-yl)-1,1. , 3,3-tetramethyluronium tetrafluoroborate (TBTU) and 1-hydroxybenzotriazole (HOBt), further preferably DIC (diisopropylcarbodiimide) and 1-hydroxybenzotriazole (HOBt).

The "monitoring" in step (a) uses a ninhydrin assay to monitor the condensation reaction of the polypeptide.

The sequential linking of amino acids in the step (a) means that the amino acids are linked one by one from the C-terminus to the N-terminus according to the amino acid sequence of the polypeptide.

The cyclized amino acid in the step (b) is Xaa6 and Xaa12, and a ring is formed by forming an amide bond between a side chain amino group and a carboxyl group.

In the step (b), an orthogonal protection strategy is employed for the ring formation, the amino protecting group on the side chain of the ring-forming amino acid is allyloxycarbonyl (Alloc), and the carboxyl protecting group is allyl (OAll).

The removal of Alloc and OAll in step (b) requires the addition of a remover, and the remover used is tetrakis(triphenylphosphine)palladium ((Pd(PPh ₃ ) ₄ )).

The use of (Pd(PPh ₃ ) ₄ ) for the removal of Alloc and OAll in step (b) requires the addition of a scavenger, and the scavenger may be selected from H ₃ N·BH ₃ , Me ₂ NH·BH ₃ or PhSiH ₃ , preferably PhSiH ₃ .

The cyclization reaction in the step (b) requires the addition of a coupling reagent consisting of a carbodiimide type reagent or a tertiary amine type and a benzotriazole salt type reagent or a pyridine triazolium salt type.

Wherein the carbodiimide type reagent includes, but is not limited to, dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC) or N-diaminopropyl-N-ethylcarbamate Imine (EDC).

The tertiary amine type reagents include, but are not limited to, N,N-diisopropylethylamine (DIEA).

The benzotriazolium salt type reagent includes, but is not limited to, 2-(1H-benzotriazo L-1-yl)-1,1,3,3-tetramethyluronium tetrafluoroborate ( TBTU), O-benzotriazole-N,N,N',N'-tetramethylurea hexafluorophosphate (HBTU), benzotriazole-1-oxotris(dimethylamino) hexafluorophosphate Phosphorus (BOP) or benzotriazol-1-yl-oxytripyrrolidinylphosphonium hexafluorophosphate (PyBOP).

The pyridine triazolium salt type includes, but is not limited to, (3H-1,2,3-triazolo[4,5-b]pyridin-3-yloxy)tri-1-pyrrolidinyl hexafluorophosphate (PyAOP), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU), 7-azabenzotriazole-1 - methoxytris(dimethylamino)phosphine hexafluorophosphate (AOP).

The coupling reagent is preferably diisopropylcarbodiimide (DIC) and 1-hydroxybenzotriazole (HOBt), or 2-(1H-benzotriazo L-1-yl)-1,1. , 3,3-tetramethylurea tetrafluoroborate (TBTU) and 1-hydroxybenzotriazole (HOBt) or N,N-diisopropylethylamine (DIEA) and 2-(7-azo Benzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU). Further preferred are N,N-diisopropylethylamine (DIEA) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU) ).

Optionally, the step (1) further comprises: (c) the resin peptide obtained in the step (b) is linked to a fatty acid chain, which may be a mono-, di- or other polybasic fatty acid chain.

The coupling agent in the step (c) is diisopropylcarbodiimide (DIC) and 1-hydroxybenzotriazole (HOBt), and the reaction solvent is DMF.

The side chain protecting group scavenger according to the step (2) includes, but not limited to, thioanisole, triisopropylsilane, phenol, water, 1,2-ethanedithiol, m-cresol or any two or two of the above The above combination is prepared and prepared by trifluoroacetic acid or hydrofluoric acid at 5-20% (V/V). Preferred is trifluoroacetic acid (TFA): thioanisole: 75% phenol: water = 85:5:5:5 (V/V/V/V).

If the C-terminus of the polypeptide of the present invention is in the form of a carboxylic acid, the step (1) is carried out by using Wang resin; if the C-terminus of the polypeptide of the present invention is in the form of an amide, the step (1) is synthesized by using Rink Amide MBHA resin. .

It is particularly advantageous that the polypeptide preparation method provided by the present invention may further comprise a purification step in order to meet the quality requirements for medical use. Purification methods employed include, but are not limited to, reverse phase chromatography or ion exchange chromatography, preferably reverse phase chromatography.

The in vitro antibacterial activity of the polypeptides of the invention can be identified by determining its minimum inhibitory concentration (MIC). The American Clinical Laboratory Standardization Committee (NCCLS) recommends the use of micro-broth dilution to determine the minimum inhibitory concentration (MIC) of each antimicrobial peptide. The medium is cultured in Mueller-Hinton (MH) broth and modified RPMI-1640. base. Amphotericin B, polymyxin E sulfate and vancomycin hydrochloride were used as positive controls. In vitro activity assays indicate that the antimicrobial peptides provided by the present invention have higher antibacterial activity, particularly anti-negative bacteria (such as Pseudomonas aeruginosa) or anti-resistant bacteria (such as drug-resistant Acinetobacter baumannii, drug-resistant patina) The activity of the bacterium was significantly improved.

definition

Unless otherwise stated, the following terms as used in this application have the following meanings. A particular term should not be considered as undefined or unclear without a particular definition, and should be understood in the ordinary meaning of the art. When a trade name appears in this document, it is intended to refer to its corresponding commodity or its active ingredient.

The term "aliphatic straight-chain or branched C ₆ -C ₂₀ acyl group" means a substituent containing a carbonyl group moiety and a non-carbonyl group moiety, and the non-carbonyl group includes an aliphatic straight chain or a branch. Chain groups include, but are not limited to, alkyl groups, cycloalkyl groups, alkene groups, and alkynyl groups. The invention includes, but is not limited to, acyl groups found in known polymyxin compounds, including heptanoyl, methylheptanoyl (including (S)-6-methylheptanoyl), octanoyl, methyl octyl Acyl (including (S)-6-methyloctanoyl, (S)-7-methyloctanoyl), decanoyl, methyldecanoyl (including (S)-6-methyldecanoyl, (S)- 7-methyldecanoyl and (S)-8-methyldecanoyl) and decanoyl. In the present invention, lauroyl, palmitoyl, myristoyl and 17-carboxylic acid-heptadecanoyl are also included. The "lauroyl", "palmitoyl" and "myristoyl" refer to a hydroxy group in lauric acid, palmitic acid, and myristic acid substituted. The 17-carboxylic acid-heptadecanoyl group generally means:

The term "fatty acid chain" refers to a substituent comprising an aliphatic straight or branched chain moiety and one, two or more carboxyl group moieties including, but not limited to, an alkyl group, a ring An alkyl group, an alkene group, an alkynyl group or the like, the one, two or more carboxyl group moieties are meant to comprise a monocarboxyl group, a dicarboxyl group or more, and the term "monobasic fatty acid chain" means a substituent comprising an aliphatic straight or branched chain moiety and a carboxyl group moiety, the term "dibasic fatty acid chain" means a substituent comprising an aliphatic straight or branched chain moiety and two carboxyl group moieties, the term " "polybasic fatty acid chain" means a substituent comprising an aliphatic straight or branched chain moiety and a plurality of carboxyl group moieties

The term "substituted" means that any one or more hydrogen atoms on a particular atom are replaced by a substituent as long as the valence of the particular atom is normal and the substituted compound is stable. When the substituent is a keto group (i.e., =0), it means that two hydrogen atoms are substituted, and the ketone substitution does not occur on the aryl group.

The term "optional" or "optionally" means that the subsequently described event or circumstance may or may not occur, the description including the occurrence of the event or condition and the occurrence of the event or condition. For example, an ethyl group "optionally" substituted with halo, refers to an ethyl group may be unsubstituted (CH ₂ CH _3), monosubstituted (e.g., CH ₂ CH ₂ F), polysubstituted (e.g. CHFCH ₂ F, CH ₂ CHF _2, etc.) or completely substituted (CF ₂ CF ₃ ). It will be understood by those skilled in the art that for any group containing one or more substituents, no substitution or substitution pattern that is sterically impossible to exist and/or which cannot be synthesized is introduced.

As used herein, C _mn means having mn carbon atoms in this moiety. For example, "carbon _3-10 cycloalkyl" means that the cycloalkyl has 3 to 10 carbon atoms. "Carbon _0-6 alkylene" means that the alkylene group has 0 to 6 carbon atoms, and when the alkylene group has 0 carbon atoms, the group is a bond.

The numerical range in this document refers to each integer in a given range. For example, " _C1-6 " means that the group may have 1 carbon atom, 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, 5 carbon atoms or 6 carbon atoms.

When any variable (eg, R) occurs more than once in the composition or structure of a compound, its definition in each case is independent. Thus, for example, if a group is replaced by 2 R, then each R has an independent option.

The term "halo" or "halogen" refers to fluoro, chloro, bromo and iodo.

The term "acyl" refers to a -CO- group.

The term "carboxy" refers to a -COOH group.

The term "hydroxy" refers to an -OH group.

The term "cyano" refers to a -CN group.

The term "mercapto" refers to a -SH group.

The term "amino" means -NH ₂ group.

The term "nitro" refers to a -NO ₂ group.

The term "alkyl" refers to _a hydrocarbon group of the formula C _n H _{2n +.} The alkyl group can be straight or branched. For example, the term "hydrocarbon ₁ - ₆ alkyl" refers to a monovalent straight or branched aliphatic group containing from 1 to 6 carbon atoms (eg, methyl, ethyl, n-propyl, isopropyl, n-butyl, Isobutyl, sec-butyl, tert-butyl, n-pentyl, 1-methylbutyl, 2-methylbutyl, 3-methylbutyl, neopentyl, 3,3-dimethylpropyl , hexyl, 2-methylpentyl, etc.). Similarly, the alkyl moiety (i.e., alkyl) of an alkoxy group, an alkylamino group, a dialkylamino group, an alkylsulfonyl group, and an alkylthio group has the same definition as defined above.

The term "alkoxy" refers to -O-alkyl.

The term "alkylthio" refers to -S-alkyl.

The term "alkylamino" refers to -NH(alkyl).

The term "dialkylamino" refers to -N(alkyl) ₂ .

The term "alkenyl" refers to a straight or branched unsaturated aliphatic hydrocarbon group having at least one double bond consisting of a carbon atom and a hydrogen atom. Non-limiting examples of alkenyl groups include, but are not limited to, ethenyl, 1-propenyl, 2-propenyl, 1-butenyl, isobutenyl, 1,3-butadienyl, and the like.

The term "alkynyl" means a straight or branched unsaturated aliphatic hydrocarbon group having at least one triple bond composed of a carbon atom and a hydrogen atom. Non-limiting examples of alkynyl groups include, but are not limited to, ethynyl (-C a carbon atom), 1-propynyl (-C group (atoms, and _3), 2-propynyl (-CH ₂ -CH (original) And 1,3-butadiynyl (-C alkynyl (sub and hydrogen) and the like.

The term "cycloalkyl" refers to a carbocyclic ring that is fully saturated and can exist as a single ring, fused ring or spiro ring. Unless otherwise indicated, the carbocyclic ring is typically a 3 to 10 membered ring, preferably a 3 to 8 membered ring. Non-limiting examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, norbornyl (bicyclo[2.2.1]heptyl), bicyclo[2.2.2]octyl, diamond Alkyl and the like.

The term "aryl" refers to an all-carbon monocyclic or fused polycyclic aromatic ring group having a conjugated π-electron system. For example, an aryl group can have 6-20 carbon atoms, 6-14 carbon atoms or 6-12 carbon atoms. Non-limiting examples of aryl groups include, but are not limited to, phenyl, naphthyl, anthracenyl, and the like.

The term "heteroaryl" refers to a monocyclic or fused polycyclic ring system containing at least one ring atom selected from N, O, S, the remaining ring atoms being C, and having at least one aromatic ring. Preferred heteroaryl groups have a single 4 to 8 membered ring, especially a 5 to 8 membered ring, or a plurality of fused rings containing from 6 to 14, especially from 6 to 10 ring atoms. Non-limiting examples of heteroaryl groups include, but are not limited to, pyrrolyl, furyl, thienyl, imidazolyl, oxazolyl, pyrazolyl, pyridyl, pyrimidinyl, pyrazinyl, quinolinyl, isoquinolinyl , tetrazolyl, triazolyl, triazinyl, benzofuranyl, benzothienyl, fluorenyl, isodecyl and the like.

The term "treating" means administering a compound or formulation described herein to prevent, ameliorate or eliminate a disease or one or more symptoms associated with the disease, and includes:

(i) preventing a disease or disease state from occurring in a mammal, particularly when such a mammal is susceptible to the disease state but has not been diagnosed as having the disease state;

(ii) inhibiting the disease or disease state, ie, curbing its development;

(iii) alleviating the disease or condition, even if the disease or condition resolves.

The term "therapeutically effective amount" means (i) treating or preventing a particular disease, condition or disorder, (ii) alleviating, ameliorating or eliminating one or more symptoms of a particular disease, condition or disorder, or (iii) preventing or delaying The amount of a compound of the present application in which one or more symptoms of a particular disease, condition, or disorder are described herein. The amount of a compound of the present application which constitutes a "therapeutically effective amount" will vary depending on the compound, the condition and severity thereof, the mode of administration, and the age of the mammal to be treated, but can be routinely determined by those skilled in the art It is determined by its own knowledge and the present disclosure.

The term "pharmaceutically acceptable" is for those compounds, materials, compositions and/or dosage forms that are within the scope of sound medical judgment and are suitable for use in contact with human and animal tissues without Many toxic, irritating, allergic reactions or other problems or complications are commensurate with a reasonable benefit/risk ratio.

As the pharmaceutically acceptable salt, for example, a metal salt, an ammonium salt, a salt with an organic base, a salt with an inorganic acid, a salt with an organic acid, a salt with a basic or acidic amino acid, or the like can be mentioned. . Non-limiting examples of metal salts include, but are not limited to, salts of alkali metals such as sodium salts, potassium salts, and the like; salts of alkaline earth metals such as calcium salts, magnesium salts, barium salts, and the like; aluminum salts and the like. Non-limiting examples of salts formed with organic bases include, but are not limited to, with trimethylamine, triethylamine, pyridine, picoline, 2,6-lutidine, ethanolamine, diethanolamine, triethanolamine, cyclohexylamine, A salt formed by dicyclohexylamine or the like. Non-limiting examples of salts formed with inorganic acids include, but are not limited to, salts formed with hydrochloric acid, hydrobromic acid, nitric acid, sulfuric acid, phosphoric acid, and the like. Non-limiting examples of salts formed with organic acids include, but are not limited to, with formic acid, acetic acid, trifluoroacetic acid, fumaric acid, oxalic acid, malic acid, maleic acid, tartaric acid, citric acid, succinic acid, methanesulfonic acid, benzene. a salt formed of a sulfonic acid, p-toluenesulfonic acid or the like. Non-limiting examples of salts formed with basic amino acids include, but are not limited to, salts formed with arginine, lysine, ornithine, and the like. Non-limiting examples of salts formed with acidic amino acids include, but are not limited to, salts formed with aspartic acid, glutamic acid, and the like.

The term "pharmaceutical composition" refers to one or more compounds of the present application, or salts thereof, and excipients, diluents, or agents generally accepted in the art for delivery of a biologically active compound to an organism (eg, a human), or Formulation of the carrier. The purpose of the pharmaceutical composition is to facilitate administration of the compounds of the present application to an organism.

The term "pharmaceutically acceptable excipient, diluent, or carrier" refers to those excipients, diluents, or carriers that do not significantly irritate the organism and which do not impair the biological activity and properties of the active compound. Suitable carriers, diluents and excipients are well known to those skilled in the art and include, for example, carbohydrates, waxes, water soluble and/or water swellable polymers, hydrophilic or hydrophobic materials, gelatin, oil, Raw materials such as solvents and water.

The present application also includes isotopically labeled compounds of the present application which are identical to those described herein, but wherein one or more atoms are replaced by an atom having a different atomic weight or mass than the atomic or mass number normally found in nature. Examples of isotopes that may be incorporated into the compounds of the present application include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, iodine, and chlorine, such as ² H, ³ H, ¹¹ C, ¹³ C, ¹⁴ C, ^{13 respectively.} N, ¹⁵ N, ¹⁵ O, ¹⁷ O, ¹⁸ O, ³¹ P, ³² P, ³⁵ S, ¹⁸ F, ¹²³ I, ¹²⁵ I and ³⁶ Cl, and the like.

Certain isotopically-labeled compounds of the present application (such as those labeled with ³ H and ¹⁴ C) can be used in compound and/or substrate tissue distribution assays. Deuterated (i.e., ³ H) and carbon-14 (i.e., ¹⁴ C) isotopes are especially preferred for their ease of preparation and detectability. Furthermore, substitution with heavier isotopes such as deuterium (ie, ² H) can provide certain therapeutic advantages resulting from higher metabolic stability (eg, increased in vivo half-life or reduced dosage requirements), and thus in some cases The following may be preferred. Positron emitting isotopes such as ¹⁵ O, ¹³ N, ¹¹ C and ¹⁸ F can be used in positron emission tomography (PET) studies to determine substrate occupancy. Isotopically labeled compounds of the present application can generally be prepared by substituting an isotopically labeled reagent for an unisotopically labeled reagent by procedures similar to those disclosed in the schemes and/or examples disclosed below.

Specific implementation

The compounds of the present application can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments listed below, combinations thereof with other chemical synthesis methods, and those well known to those skilled in the art. Equivalent alternatives, preferred embodiments include, but are not limited to, embodiments of the present application.

The chemical reactions of the specific embodiments of the present application are accomplished in a suitable solvent which is suitable for the chemical changes of the present application and the reagents and materials required thereof. In order to obtain the compounds of the present application, it is sometimes necessary for those skilled in the art to modify or select the synthetic steps or reaction schemes based on the prior embodiments.

The following examples are merely representative of one aspect of the invention and are not a limitation of the subject matter of the invention. For example, the invention includes, but is not limited to, the following:

A polypeptide or a pharmaceutically acceptable salt thereof, which polypeptide has at least 70% sequence identity to the sequence of Formula I:

R1-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-Xaa7-Xaa8-Xaa9-Xaa10-Xaa11-Xaa12 (Formula I),

Wherein R1 is: an aliphatic straight-chain or branched C ₆ -C ₂₀ acyl group or a deletion;

Xaa1 is: Leu, Ala, AEEAc, Phe, IPhe, BrPhe, 3FPhe or deletion;

Xaa2 is: Leu, Ala, Arg, AEEAc or deletion;

Xaa3 is: Dab, Ala, Leu, Orn or missing;

Xaa4 is: Thr, Ala, Dab or Lys;

Xaa5 is: Dab, Lys, Orn, Ala or missing;

Xaa6 is: Lys, Glu or Dab;

Xaa7 is: Dab, Ala, Lys, Orn or missing;

Xaa8 is: D-Leu, D-Phe, Lys or D-Ala;

Xaa9 is: Leu or Ala;

Xaa10 is: Dab, Leu, Orn, Ala, D-Leu or deletion;

Xaa11 is: Dab, Leu, Orn, Ala, D-Leu or deletion;

Xaa12 is: Glu, Lys or Asp.

2. The polypeptide of claim 1, or a pharmaceutically acceptable salt thereof, wherein the polypeptide is a cyclic peptide.

3. The polypeptide according to any one of items 1 to 2, wherein the polypeptide is obtained by dehydration of Xaa6 and a side chain group of Xaa12 to form an amide bond, optionally, the side chain group. The group contains an amino group and/or a carboxyl group.

4. The polypeptide according to any one of items 1 to 3, wherein R1 is selected from the group consisting of heptanoyl, methylheptanoyl, octanoyl, methyloctanoyl, decanoyl, methyl decanoyl, hydrazine Acyl, methyl decanoyl, lauroyl, myristoyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, preferably n-octanoyl, decanoyl, lauroyl, palmitoyl, 17-carboxylic acid-ten Heptanoyl or deletion, more preferably n-octanoyl or deletion; Xaa1 is preferably Leu or deleted, Xaa2 is preferably Leu, Xaa3 is preferably Dab, Xaa4 is preferably Thr, Xaa5 is preferably Dab, Xaa6 is preferably Lys or Dab, Xaa7 is preferably Dab, Xaa8 is preferably D -Phe or D-Leu, Xaa9 is preferably Leu, Xaa10 is preferably Dab, Xaa11 is preferably Dab, and/or Xaa12 is preferably Glu.

5. The polypeptide of any one of clauses 1 to 3, wherein the polypeptide has the following formula:

Wherein R1 is: an aliphatic straight-chain or branched C ₆ -C ₂₀ acyl group or a deletion;

Xaa1 is: Leu, Ala, AEEAc, Phe, IPhe, BrPhe, 3FPhe or deletion;

Xaa2 is: Leu, Ala, Arg, AEEAc or deletion;

Xaa3 is: Dab, Ala, Leu, Orn or missing;

Xaa4 is: Thr, Ala, Dab or Lys;

Xaa5 is: Dab, Lys, Orn, Ala or missing;

Xaa6 is: Lys or Dab;

Xaa7 is: Dab, Ala, Lys, Orn or missing;

Xaa8 is: D-Leu, D-Phe, Lys or D-Ala;

Xaa9 is: Leu or Ala;

Xaa10 is: Dab, Leu, Orn, Ala, D-Leu or deletion;

Xaa11 is: Dab, Leu, Orn, Ala, D-Leu or deletion.

6. The polypeptide according to item 5, wherein R1 is selected from the group consisting of heptanoyl, methylheptanoyl, octanoyl, methyloctanoyl, decanoyl, methyldecanoyl, decanoyl, methyl Decanoyl, lauroyl, myristoyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, preferably n-octanoyl, decanoyl, lauroyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or Deletion, more preferably n-octanoyl or deletion; Xaa1 is preferably Leu or deletion, Xaa2 is preferably Leu, Xaa3 is preferably Dab, Xaa4 is preferably Dab, Xaa4 is preferably Thr, Xaa5 is preferably Dab, Xaa6 is preferably Lys or Dab, Xaa7 is preferably Dab, Xaa8 is preferably D-Phe or D -Leu, Xaa9 is preferably Leu, Xaa10 is preferably Dab, and Xaa11 is preferably Dab.

7. The polypeptide of any one of clauses 1 to 3, wherein the polypeptide has the following formula:

Wherein R1 is: an aliphatic straight-chain or branched C ₆ -C ₂₀ acyl group or a deletion;

Xaa1 is: Leu, Ala, AEEAc, Phe, IPhe, BrPhe, 3FPhe or deletion;

Xaa2 is: Leu, Ala, Arg, AEEAc or deletion;

Xaa3 is: Dab, Ala, Leu, Orn or missing;

Xaa4 is: Thr, Ala, Dab or Lys;

Xaa5 is: Dab, Lys, Orn, Ala or missing;

Xaa7 is: Dab, Ala, Lys, Orn or missing;

Xaa8 is: D-Leu, D-Phe, Lys or D-Ala;

Xaa9 is: Leu or Ala;

Xaa10 is: Dab, Leu, Orn, Ala, D-Leu or deletion;

Xaa11 is: Dab, Leu, Orn, Ala, D-Leu or deletion;

Xaa12 is: Glu or Asp.

8. The polypeptide according to item 7, or a pharmaceutically acceptable salt thereof, wherein R1 is selected from the group consisting of heptanoyl, methylheptanoyl, octanoyl, methyloctanoyl, decanoyl, methyldecanoyl, decanoyl, methylhydrazine Acyl, lauroyl, myristoyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, preferably n-octanoyl, decanoyl, lauroyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion More preferably, it is n-octanoyl or a deletion; Xaa1 is preferably Leu or a deletion, Xaa2 is preferably Leu, Xaa3 is preferably Dab, Xaa3 is preferably Dab, Xaa4 is preferably Thr, Xaa5 is preferably Dab, Xaa7 is preferably Dab, Xaa8 is preferably D-Phe or D-Leu, Xaa9 is preferably Leu, Xaa10 is preferably Dab, Xaa11 is preferably Dab, and/or Xaa12 is preferably Glu.

The polypeptide of any one of items 1 to 3, wherein the polypeptide has the following formula:

Wherein R1 is: an aliphatic straight-chain or branched C ₆ -C ₂₀ acyl group or a deletion;

Xaa1 is: Leu, Ala, AEEAc, Phe, IPhe, BrPhe, 3FPhe or deletion;

Xaa2 is: Leu, Ala, Arg, AEEAc or deletion;

Xaa3 is: Dab, Ala, Leu, Orn or missing;

Xaa4 is: Thr, Ala, Dab or Lys;

Xaa5 is: Dab, Lys, Orn, Ala or missing;

Xaa7 is: Dab, Ala, Lys, Orn or missing;

Xaa8 is: D-Leu, D-Phe, Lys or D-Ala;

Xaa9 is: Leu or Ala;

Xaa10 is: Dab, Leu, Orn, Ala, D-Leu or deletion;

Xaa11 is: Dab, Leu, Orn, Ala, D-Leu or deletion.

10. The polypeptide according to item 9, or a pharmaceutically acceptable salt thereof, wherein R1 is selected from the group consisting of heptanoyl, methylheptanoyl, octanoyl, methyloctanoyl, decanoyl, methyldecanoyl, decanoyl, methylhydrazine Acyl, lauroyl, myristoyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, preferably n-octanoyl, decanoyl, lauroyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion More preferably, it is n-octanoyl or a deletion; Xaa1 is preferably Leu or a deletion, Xaa2 is preferably Leu, Xaa3 is preferably Dab, Xaa3 is preferably Dab, Xaa4 is preferably Thr, Xaa5 is preferably Dab, Xaa7 is preferably Dab, Xaa8 is preferably D-Phe or D-Leu, Xaa9 is preferably Leu, Xaa10 is preferably Dab, and Xaa11 is preferably Dab.

The polypeptide according to any one of items 1 to 4, wherein the polypeptide is represented by R1-Leu-Dab-Thr-Dab-Xaa6-Dab-Xaa8-Leu-Dab-Dab-Glu a sequence consisting of a peptide or a cyclic peptide, wherein R1 is preferably n-octanoyl, decanoyl, lauroyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, more preferably n-octanoyl or a deletion; Xaa6 is preferably Lys Or Dab, Xaa8 is preferably D-Phe or D-Leu.

12. The polypeptide according to item 11, or a pharmaceutically acceptable salt thereof, which is a peptide consisting of H ₂ N-Leu-Dab-Thr-Dab-Lys-Dab-D-Phe-Leu-Dab-Dab-Glu Or a cyclic peptide.

13. The polypeptide of claim 12, or a pharmaceutically acceptable salt thereof, the amino acid sequence of the polypeptide is as follows:

The polypeptide according to any one of items 1 to 4, wherein the polypeptide is a sequence represented by R1-Dab-Thr-Dab-Xaa6-Dab-Xaa8-Leu-Dab-Dab-Glu A peptide or cyclic peptide consisting of wherein R1 is preferably n-octanoyl, decanoyl, lauroyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, more preferably n-octanoyl or a deletion; Xaa6 is preferably Lys or Dab Xaa8 is preferably D-Phe or D-Leu.

The polypeptide according to item 14, or a pharmaceutically acceptable salt thereof, which is a peptide or a ring consisting of n-octanoyl-Dab-Thr-Dab-Lys-Dab-D-Phe-Leu-Dab-Dab-Glu Peptide.

16. The polypeptide of claim 15 or a pharmaceutically acceptable salt thereof, the amino acid sequence of the polypeptide being as follows:

17. The polypeptide of clause 1-3, or a pharmaceutically acceptable salt thereof, the amino acid sequence of the polypeptide having at least 70% sequence identity to one of the following sequences:

18. A method of producing the polypeptide of any one of 18.1, or a pharmaceutically acceptable salt thereof, comprising: synthesizing the polypeptide.

19. The method of clause 18, comprising:

(1) solid phase synthesis of a polypeptide on a resin;

(2) The product of the step (1) is cleaved with a strong acid; a side chain protecting group scavenger is added, filtered, and the polypeptide is precipitated with 5-20 volumes of an organic solvent, centrifuged, washed with an organic solvent, and dried to obtain a crude peptide.

20. The method of item 19, wherein the step (1) is performed in a liquid phase environment, and specifically comprises:

(a) soaking resin - removal of amino protecting group - washing - monitoring - coupling of amino acids - monitoring - washing - removal of amino protecting groups - sequential coupling of residual amino acids;

(b) The resin peptide is further cyclized.

21. The method of item 20, wherein the solvent used in the liquid phase environment of step (a) is selected from the group consisting of: dimethylformamide (DMF), dichloromethane (DCM), N-methylpyrrolidone (NMP), Preference is given to DMF; the amino protecting group is selected from the group consisting of tert-butoxycarbonyl (Boc), benzyloxycarbonyl (Z) or 9-fluorenyl-methoxycarbonyl (Fmoc), preferably 9-fluorenyl-methoxycarbonyl (Fmoc).

The method according to any one of items 20 to 21, wherein the coupling of the amino acid in the step (a) requires the addition of a coupling reagent consisting of a carbodiimide type reagent or a benzotriazine. Imidazolium salt type reagent and 1-hydroxybenzotriazole (HOBt); preferably diisopropylcarbodiimide (DIC) and 1-hydroxybenzotriazole (HOBt), or 2-(1H-benzene And trisazo-l-1-yl)-1,1,3,3-tetramethyluronium tetrafluoroborate (TBTU) and 1-hydroxybenzotriazole (HOBt), further preferably DIC (diisopropyl) Carbodiimide) and 1-hydroxybenzotriazole (HOBt).

The method according to any one of items 20 to 22, wherein the cyclized amino acid in the step (b) is Xaa6 and Xaa12, and a ring is formed by forming an amide bond between a side chain amino group and a carboxyl group.

The method according to any one of items 20 to 23, wherein the step (b) is carried out by orthogonal protection strategy, and the amino protecting group on the side chain of the ring-forming amino acid is allyloxycarbonyl (Alloc). The carboxy protecting group is an allyl group (OAll).

25. The method according to item 24, wherein the step (b) comprises deamination protection, and the removal of Alloc and OAll requires the addition of a degreasing agent, which is tetrakis(triphenylphosphine)palladium (Pd(PPh) ₃ ) ₄ ).

26. The method of item 25, wherein the step (b) further requires the addition of a scavenger selected from the group consisting of H ₃ N·BH ₃ , Me ₂ NH·BH ₃ or PhSiH ₃ , preferably PhSiH ₃ .

27. The method according to any one of items 20-26, wherein the cyclization reaction in the step (b) requires the addition of a coupling reagent consisting of a carbodiimide type reagent or a tertiary amine type and a benzotriene a razoloxyl salt type reagent or a pyridine triazolium salt type; preferably diisopropylcarbodiimide (DIC) and 1-hydroxybenzotriazole (HOBt), or 2-(1H-benzotriazo) L-1-yl)-1,1,3,3-tetramethyluronium tetrafluoroborate (TBTU) and 1-hydroxybenzotriazole (HOBt) or N,N-diisopropylethylamine ( DIEA) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU); further preferably N,N-diisopropyl B Amine (DIEA) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU).

28. The method of any one of clauses 20 to 27, wherein the step (1) further comprises: (c) linking the resin peptide obtained in the step (b) to a fatty acid chain or a polybasic fatty acid chain.

29. The method according to item 28, wherein the step (c) is carried out in the presence of a coupling agent in a DMF solvent, the coupling agent being diisopropylcarbodiimide (DIC) and 1-hydroxyl Benzotriazole (HOBt).

30. Use of a polypeptide according to any of the preceding claims, or a pharmaceutically acceptable salt thereof, for use in bacteriostatic, antibacterial or antibacterial, antibacterial substances.

31. The use of claim 30, wherein the antimicrobial substance is an antibacterial or antibacterial agent.

32. The use of item 31, wherein the bacterium is a bacterium or a fungus.

33. The use of claim 32, wherein the bacterium or fungus is Acinetobacter baumanii, drug resistant Acinetobacter baumanii, Pseudomonas aeruginosa, Drug resistant Pseudomonas aeruginosa, Bacillus subtilis, Staphylococcus aureus, E. coli, and/or candida Albicans One or several.

Example 1: Preparation and Purification of Z18

(1) Materials and reagents

Rink Amide MBHA resin with a substitution value of 0.65 mmol/g.

The amino acids are: Fmoc-Glu(OAll)-OH, Fmoc-Dab(Boc)-OH, Fmoc-D-Phe-OH, Fmoc-Leu-OH, Fmoc-D-Leu-OH, Fmoc-Lys(Alloc)- OH, Fmoc-Thr(tBu)-OH

Synthetic reagents: HOBt, DIC, DMF, DCM, piperidine, tetrakis(triphenylphosphine)palladium, phenylsilane, sodium diethyldithiocarbamate.

(2) Instrument

PSI300 peptide synthesis instrument, Waters600 semi-preparative high performance liquid chromatography, centrifuge.

(3) Operation steps (taking 0.15mmol as an example)

a. Solid phase chemical synthesis of peptides

Weigh 0.23g of Rink Amide MBHA resin, put it into the peptide synthesizer reactor, add 10ml DMF, soak for 2h, add 10ml of 20% PIP/DMF solution, mix for 30min to remove amino protecting group, DMF wash resin 7 times; weigh three Amount of Fmoc-Glu(OAll)-OH, DIC, HOBT was dissolved in 10 ml of DMF, and the reaction was carried out at room temperature. The reaction progress was monitored by ninhydrin reaction, and the reaction was completed after monitoring colorlessness. The resin was washed with DMF. Times. After the first amino acid is coupled to the resin, the next amino acid coupling reaction can be continued as described above and cycled until all amino acid coupling is complete. To the reactor, tetrakis(triphenylphosphine)palladium and phenylsilane were added, and the ratio of the molar amount to the resin was 1:0.1:10. After being dissolved in 10 ml of DCM, the reaction was carried out under the protection of N ₂ and the temperature was room temperature. The reaction was carried out by ninhydrin reaction, black purple was regarded as completion of the reaction, the resin was washed 5 times with DCM, the resin was washed 3 times with sodium diethyldicarboformate/DMF, 5 times with DMF, and 10 ml was added to the reactor. DMF dissolved 3 times the amount of HATU and 6 times the amount of DIEA were reacted, the reaction temperature was room temperature, the reaction progress was monitored by ninhydrin reaction, and the reaction was completed after monitoring colorless, and the resin was washed 5 times with DMF. Finally, 10 ml of 20% PIP/DMF solution was added and mixed for 30 min to remove the last amino acid terminal Fmoc protecting group.

b. Cracking and precipitation

After the peptide synthesis was completed, the resin was dried under vacuum and weighed. The lysis reagent was added in a ratio of 1 ml of the lysis reagent/100 mg of the resin, and the reagent ratio was TFA: thioanisole: 75% phenol: water = 85:5:5:5 (V: V: V: V), and the reaction was stirred at room temperature. Hours, suction filtration. The peptide was precipitated by adding 10 volumes of ice diethyl ether to the lysate, centrifuged, and the supernatant was discarded, and the precipitate was repeatedly washed with ice diethyl ether for 4 to 5 times, dried in vacuo, and the crude peptide was weighed.

c. separation and purification

The crude peptide was purified by semi-preparative RP-HPLC.

Purification

Column: YMC-pack ODS-AQ C18 preparative column (10mm × 250mm, 10μm)

Flow rate: 5ml/min

Detection wavelength: 215nm

Mobile phase: Phase A: 0.1% TFA/water solution

Phase B: 0.1% TFA / acetonitrile

The gradient elution procedure is shown in Table 3:

Table 3 gradient elution table

2. Analysis method

Column: YMC-pack ODS-AQ C18 analytical column (4.6mm × 250mm, 5μm)

Flow rate: 1ml/min

Detection wavelength: 215nm

Mobile phase: Phase A: 0.05% TFA / water

Phase B: 0.05% TFA / acetonitrile

The gradient elution procedure is shown in Table 4:

Table 4 gradient elution table

The target component with a purity greater than 95% was collected, steamed under low pressure, and lyophilized. Molecular weight confirmation by ESI-MS, m/z=1232.05 (M+H) ⁺ is consistent with the theoretical molecular weight of 1231.32.

Example 2: Preparation and purification of Z29

Amino acid sequence:

(1) Materials and reagents

Rink Amide MBHA resin with a substitution value of 0.65 mmol/g.

The amino acids are: Fmoc-Glu(OAll)-OH, Fmoc-Dab(Boc)-OH, Fmoc-D-Phe-OH, Fmoc-Leu-OH, Fmoc-D-Leu-OH, Fmoc-Lys(Alloc)- OH, Fmoc-Thr(tBu)-OH

Synthetic reagents: HOBt, DIC, DMF, DCM, piperidine, tetrakis(triphenylphosphine)palladium, phenylsilane, sodium diethyldithiocarbamate, n-octanoic acid.

(2) Instrument

PSI300 peptide synthesis instrument, Waters600 semi-preparative high performance liquid chromatography, centrifuge.

(3) Operation steps (taking 0.15mmol as an example)

a. Solid phase chemical synthesis of peptides

Weigh 0.23g of Rink Amide MBHA resin, put it into the peptide synthesizer reactor, add 10ml DMF, soak for 2h, add 10ml of 20% PIP/DMF solution, mix for 30min to remove amino protecting group, DMF wash resin 7 times; weigh three Amount of Fmoc-Glu(OAll)-OH, DIC, HOBT was dissolved in 10 ml of DMF, and the reaction was carried out at room temperature. The reaction progress was monitored by ninhydrin reaction, and the reaction was completed after monitoring colorlessness. The resin was washed with DMF. Times. After the first amino acid is coupled to the resin, the next amino acid coupling reaction can be continued as described above and cycled until all amino acid coupling is complete. To the reactor, tetrakis(triphenylphosphine)palladium and benzenesilane were added, and the ratio of the molar amount to the resin was 1:0.1:10. After adding 10 ml of DCM, the solution was dissolved in the dark and protected under N ₂ at a temperature of At room temperature, the reaction was carried out by ninhydrin reaction, black purple was regarded as completion of the reaction, the resin was washed 5 times with DCM, the resin was washed 3 times with sodium diethyldicarboformate/DMF, 5 times with DMF, and added to the reactor. 10 ml of DMF dissolved 3 times the amount of HATU and 6 times the amount of DIEA were reacted, the reaction temperature was room temperature, the reaction progress was monitored by ninhydrin reaction, and the reaction was completed after monitoring colorless, and the resin was washed 5 times with DMF. Finally, 10 ml of 20% PIP/DMF solution was added and mixed for 30 min to remove the last amino acid terminal Fmoc protecting group. Three times the amount of n-octanoic acid was weighed, and DIC and HOBT were dissolved in 10 ml of DMF, and the reaction was carried out at room temperature. The reaction progress was monitored by ninhydrin reaction, and the reaction was completed after monitoring colorlessness, and the resin was washed 5 times with DMF.

b. Cracking and precipitation

After the peptide synthesis was completed, the resin was dried under vacuum and weighed. The lysis reagent was added in a ratio of 1 ml of the lysis reagent/100 mg of the resin, and the reagent ratio was TFA: thioanisole: 75% phenol: water = 85:5:5:5 (V: V: V: V), and the reaction was stirred at room temperature. Hours, suction filtration. The peptide was precipitated by adding 10 volumes of ice diethyl ether to the lysate, centrifuged, and the supernatant was discarded, and the precipitate was repeatedly washed with ice diethyl ether for 4 to 5 times, dried in vacuo, and the crude peptide was weighed.

c. separation and purification

The crude peptide was purified by semi-preparative RP-HPLC.

Purification

Column: YMC-pack ODS-AQ C18 preparative column (10mm × 250mm, 10μm)

Flow rate: 5ml/min

Detection wavelength: 215nm

Mobile phase: Phase A: 0.1% TFA/water solution

Phase B: 0.1% TFA / acetonitrile

The gradient elution procedure is shown in Table 5:

Table 5 gradient elution table

2. Analysis method

Column: YMC-pack ODS-AQ C18 analytical column (4.6mm × 250mm, 5μm)

Flow rate: 1ml/min

Detection wavelength: 215nm

Mobile phase: Phase A: 0.05% TFA / water

Phase B: 0.05% TFA / acetonitrile

The gradient elution procedure is shown in Table 6:

Table 6 gradient elution table

The target component with a purity greater than 95% was collected, steamed under low pressure, and lyophilized. Molecular weight confirmation by ESI-MS, m/z = 606.25 (M+2H) ²⁺ is consistent with the theoretical molecular weight of 1210.34.

Example 3: Preparation and purification of the remaining CAMP series antimicrobial peptides in Table 7

(1) Materials and reagents

Rink Amide MBHA resin with a substitution value of 0.65 mmol/g.

The amino acids are: Fmoc-Glu(OAll)-OH, Fmoc-Asp(OAll)-OH, Fmoc-Dab(Boc)-OH, Fmoc-Leu-OH, Fmoc-D-Leu-OH, Fmoc-D-Phe- OH, Fmoc-Lys(Alloc)-OH, Fmoc-Thr(tBu)-OH, Fmoc-Phe-OH, Fmoc-Orn(Boc)-OH, Fmoc-Arg(Pbf)-OH, Fmoc-Phe(4- CF ₃ )-OH, Fmoc-Phe(4-Br)-OH, Fmoc-Phe(4-I)-OH, Fmoc-Ala-OH, Fmoc-D-Ala-OH, Fmoc-AEEA-OH,

Reagents: HOBt, DIC, DMF, piperidine, n-octanoic acid, citric acid, lauric acid, palmitic acid, t-butyl monodecanoate.

(2) Instrument

PSI300 peptide synthesizer, Waters600 semi-preparative high performance liquid chromatography, Q-Tof MicroYA019 mass spectrometer, magnetic stirrer.

(3) Operation steps (taking 0.15mmol as an example)

The polypeptide in Table 7 was prepared and purified in a manner similar to the procedure of ac in Example 1, and if it was a fatty chain polypeptide, it was prepared according to the procedure ac in Example 2, and the fraction having a purity greater than 95% was collected, and steamed, Freeze-dried. The measured values of ESI-MS are shown in Table 7.

Table 7 polymyxin analogs and their molecular weight

Example 4: Determination of in vitro antibacterial activity

The minimum inhibitory concentration (MIC) of each antimicrobial peptide was determined according to the micro-broth dilution method recommended by the American Committee for Clinical Laboratory Standards (NCCLS). The bacterial culture medium was Mueller-Hinton (MH) broth medium, Candida albicans medium. Hyclone modified RPMI-1640 medium was used.

The specific steps are:

(1) Preparation of antibacterial drug stock solution:

The above antimicrobial peptide at a concentration of 320 μg/ml and 80 μg/ml positive control amphotericin B, polymyxin E sulfate and vancomycin hydrochloride were precisely prepared. The prepared stock solutions were stored in an environment of -20 ° C for use.

(2) Preparation of the medium:

21 g of MH broth medium was weighed, dissolved in distilled water and made up to 1 L, and sterilized at 121 ° C for 30 min.

(3) Preparation of inoculum:

Use the inoculating loop to pick 3 to 5 colonies of similar morphology to be inoculated, inoculate them in 4 to 5 ml of MH broth, and incubate at 35 ° C for 12 to 16 h. In the logarithmic growth phase, the bacterial liquid was corrected to a MH turbidity standard with MH broth, and contained about 1 to 2 × 10 ⁸ CFU/ml. The above bacterial suspension was diluted 1:1000 with MH broth and set aside.

(4) Preparation of diluted antibacterial drugs and bacterial inoculation:

Take a 96-well plate, add 160 μL of MH broth to the first well, add 100 μL of MH broth to each of the 2-12 wells, then add 40 μL of the antibacterial drug solution (320 μg/ml) to the first well, and mix. Pipette 100 μL to the second well, mix and then take 100 μL to the third well from the second well, so as to serially dilute to the 10th well, and take 100 μL from the 10 well to discard, then to the 1-10 100 μL of each of the prepared inoculum was added to the wells and the 12th well, so that the final bacterial concentration per well was about 0.5 × 10 ⁵ CFU/ml. The drug concentrations in the first 1-10 well were 32 μg/ml, 16 μg/ml, 8 μg/ml, 4 μg/ml, 2 μg/ml, 1 μg/ml, 0.5 μg/ml, 0.25 μg/ml, 0.125 μg/ml, and 0.0625 μg, respectively. /ml, the 11th hole is a blank control without antibacterial drugs and inoculum, and the 12th hole is a negative control without antibacterial drugs.

(5) Incubation

The 96-well plates inoculated with bacteria were incubated in a normal air incubator at 35 ° C for 16-20 h, and the 96-well plates inoculated with fungi were incubated in a 28 ° C air incubator for 40-50 h.

(6) Results

The lowest drug concentration without bacterial growth was observed by the naked eye as the minimum inhibitory concentration (MIC) of the sample. The MIC measurement results of the respective antimicrobial peptides are shown in Table 8.

All of the compositions and/or methods disclosed and claimed herein can be prepared and carried out without undue experimentation in light of the present disclosure. While the compositions and methods of the present invention are described in accordance with the preferred embodiments, the compositions and/or compositions described herein may be practiced without departing from the spirit, scope and scope of the invention. Or the method and the order of the steps or steps of the method are changed.

The disclosures of all of the documents cited herein are hereby incorporated by reference in their entirety to the extent of the extent of the disclosure of the disclosure of the disclosure of the disclosure.

Table 8 MIC results of polymyxin analogs

Example 5: Determination of the median lethal dose

Median Lethal Dose (LD50) refers to the exposure dose of harmful substances or radiation that can kill half of the test subjects. It is a common parameter for describing the toxicity of harmful substances or radiation. LD50 is usually characterized by the ratio of the mass of the hazardous substance to the weight of the test organism and can be used to compare the relative toxicity of different substances.

The specific steps are:

(1) Selection and culture of experimental animals

ICR mice, male, weighing 20-22 g, were from Shanghai Xipuer-Beikai Experimental Animal Co., Ltd. The mice were housed in an animal laboratory at 20 ± 2 ° C for 12 hours, and the experimental animals were given free access to water and food. Randomly grouped by weight.

(2) Preparation of test substance

Accurately weigh CTAMP43, ZAMP18 and polymyxin E, add sodium chloride injection to prepare a solution containing 10mg/ml of sample, and chill at -20 °C for use.

(3) Route of administration and volume of administration

According to the route of administration of the clinical study recommended by the test drug, intravenous administration was carried out at a dose of 10 ml/kg.

(4) Experimental method

Before the experiment, a small number of mice were used for pre-experiment, and the lethal dose range of 0% and 100% mortality of the test substance was measured, and then the formal test was conducted. The number of formal test groups was 5 dose groups, and the number of animals in each group was 10.

(5) Pre-experiment

One mouse was administered, and the test sample was administered intravenously at 5 mg/kg (administered volume 0.2 ml). The injection is completed in about 20s. If the animal does not die, another mouse is taken and a higher dose is given. If the animal dies or is in a state of sudden death, take another one and give a lower dose. The dose of animal survival was supplemented to 3, and the survival state of the animals was observed. Find 0% and 100% death doses.

(6) Formal test

In the range of 0% and 100% lethal doses, the inter-group dose ratio was set, and the number of groups was 5, and the number of animals in each group was 10. The immediate response of the animals after administration was observed and recorded. Body weight changes were monitored and the status of the animals after administration was recorded. If an animal dies, it should be recorded, and the experiment is terminated 7 days after administration, and all animals are sacrificed. Based on the experimental results, the LD50 was determined.

(7) Results

In this test, the LD50 of CTAMP43 and ZAMP18 with high activity was measured and compared with polymyxin E sulfate. The test results are shown in Table 9 below. The results showed that the LD50 of CTAMP43 and ZAMP18 were 34.6 mg/kg and 35.0 mg/kg, respectively, while the LD50 of polymyxin E was 9.3-10 mg/ml, indicating that CTAMP43 and ZAMP18 were less toxic than polymyxin E. -4 times.

Table 9 Half lethal dose measurement results

In view of the present disclosure, while the compositions and methods of the present invention have been described in accordance with the preferred embodiments, those skilled in the art may, without departing from the concept, the spirit and the scope of the present invention, The compositions and/or methods described herein, as well as the order of the steps or steps of the methods, are varied.

The disclosures of all of the documents cited herein are hereby incorporated by reference in their entirety to the extent of the extent of the disclosure of the disclosure of the disclosure of the disclosure.

Claims (15)

1. A polypeptide or a pharmaceutically acceptable salt thereof, which polypeptide has at least 70% sequence identity to the sequence of Formula I:

   R1-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-Xaa7-Xaa8-Xaa9-Xaa10-Xaa11-Xaa12 (Formula I),

   Wherein R1 is: an aliphatic straight-chain or branched C ₆ -C ₂₀ acyl group or a deletion;

   Xaa1 is: Leu, Ala, AEEAc, Phe, IPhe, BrPhe, 3FPhe or deletion;

   Xaa2 is: Leu, Ala, Arg, AEEAc or deletion;

   Xaa3 is: Dab, Ala, Leu, Orn or missing;

   Xaa4 is: Thr, Ala, Dab or Lys;

   Xaa5 is: Dab, Lys, Orn, Ala or missing;

   Xaa6 is: Lys, Glu or Dab;

   Xaa7 is: Dab, Ala, Lys, Orn or missing;

   Xaa8 is: D-Leu, D-Phe, Lys or D-Ala;

   Xaa9 is: Leu or Ala;

   Xaa10 is: Dab, Leu, Orn, Ala, D-Leu or deletion;

   Xaa11 is: Dab, Leu, Orn, Ala, D-Leu or deletion;

   Xaa12 is: Glu, Lys or Asp.
2. The polypeptide of claim 1 or a pharmaceutically acceptable salt thereof, wherein the polypeptide is a cyclic peptide.
3. A polypeptide according to any one of claims 1 to 2, or a pharmaceutically acceptable salt thereof, which is obtained by dehydration of Xaa6 and a side chain group of Xaa12 to form an amide bond, optionally, the side chain group Contains amino and/or carboxyl groups.
4. The polypeptide according to any one of claims 1 to 3, wherein R1 is selected from the group consisting of heptanoyl, methylheptanoyl, octanoyl, methyloctanoyl, decanoyl, methyldecanoyl, decanoyl , methyl decanoyl, lauroyl, myristoyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, preferably n-octanoyl, decanoyl, lauroyl, palmitoyl, 17-carboxylic acid - seventeen Alkanoyl group or deletion, more preferably n-octanoyl or deletion; Xaa1 is preferably Leu or deleted, Xaa2 is preferably Leu, Xaa3 is preferably Dab, Xaa4 is preferably Dab, Xaa4 is preferably Thr, Xaa5 is preferably Dab, Xaa6 is preferably Lys or Dab, Xaa7 is preferably Dab, Xaa8 is preferably D- Phe or D-Leu, Xaa9 is preferably Leu, Xaa10 is preferably Dab, Xaa11 is preferably Dab, and/or Xaa12 is preferably Glu.
5. The polypeptide or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 3, which has the general formula: R1-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Lys-Xaa7-Xaa8-Xaa9-Xaa10-Xaa11 -Glu,

   Wherein R1 is: an aliphatic straight-chain or branched C ₆ -C ₂₀ acyl group or a deletion;

   Xaa1 is: Leu, Ala, AEEAc, Phe, IPhe, BrPhe, 3FPhe or deletion;

   Xaa2 is: Leu, Ala, Arg, AEEAc or deletion;

   Xaa3 is: Dab, Ala, Leu, Orn or missing;

   Xaa4 is: Thr, Ala, Dab or Lys;

   Xaa5 is: Dab, Lys, Orn, Ala or missing;

   Xaa7 is: Dab, Ala, Lys, Orn or missing;

   Xaa8 is: D-Leu, D-Phe, Lys or D-Ala;

   Xaa9 is: Leu or Ala;

   Xaa10 is: Dab, Leu, Orn, Ala, D-Leu or deletion;

   Xaa11 is: Dab, Leu, Orn, Ala, D-Leu or deletion.
6. The polypeptide according to claim 5 or a pharmaceutically acceptable salt thereof, wherein R1 is selected from the group consisting of heptanoyl, methylheptanoyl, octanoyl, methyloctanoyl, decanoyl, methyldecanoyl, decanoyl, methyldecanoyl , lauroyl, myristoyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, preferably n-octanoyl, decanoyl, lauroyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, More preferably, it is n-octanoyl or a deletion; Xaa1 is preferably Leu or a deletion, Xaa2 is preferably Leu, Xaa3 is preferably Dab, Xaa3 is preferably Dab, Xaa4 is preferably Thr, Xaa5 is preferably Dab, Xaa7 is preferably Dab, Xaa8 is preferably D-Phe or D-Leu, Xaa9 is preferably Leu, Xaa10 Preferably, Dab, Xaa11 is preferably Dab.
7. The polypeptide of any one of claims 1 to 4, or a pharmaceutically acceptable salt thereof, which is represented by R1-Leu-Dab-Thr-Dab-Xaa6-Dab-Xaa8-Leu-Dab-Dab-Glu A peptide or cyclic peptide consisting of a sequence, wherein R1 is preferably n-octanoyl, decanoyl, lauroyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, more preferably n-octanoyl or a deletion; Xaa6 is preferably Lys or Dab, Xaa8 is preferably D-Phe or D-Leu.
8. The polypeptide according to claim 7 or a pharmaceutically acceptable salt thereof, which is a peptide consisting of H ₂ N-Leu-Dab-Thr-Dab-Lys-Dab-D-Phe-Leu-Dab-Dab-Glu or Cyclic peptide.
9. The polypeptide of claim 8 or a pharmaceutically acceptable salt thereof, the amino acid sequence of the polypeptide being as follows:

   
10. The polypeptide of any one of claims 1 to 4, or a pharmaceutically acceptable salt thereof, which comprises a sequence represented by R1-Dab-Thr-Dab-Xaa6-Dab-Xaa8-Leu-Dab-Dab-Glu Peptide or cyclic peptide, wherein R1 is preferably n-octanoyl, decanoyl, lauroyl, palmitoyl, 17-carboxylic acid-heptadecanoyl or a deletion, more preferably n-octanoyl or a deletion; Xaa6 is preferably Lys or Dab, Xaa8 is preferably D-Phe or D-Leu.
11. The polypeptide according to claim 10 or a pharmaceutically acceptable salt thereof, which is a peptide or cyclic peptide consisting of n-octanoyl-Dab-Thr-Dab-Lys-Dab-D-Phe-Leu-Dab-Dab-Glu. .
12. The polypeptide of claim 11 or a pharmaceutically acceptable salt thereof, the amino acid sequence of the polypeptide being as follows:

    
13. The polypeptide of any one of claims 1 to 3, or a pharmaceutically acceptable salt thereof, having an amino acid sequence of at least 70% sequence identity to one of the following sequences:

    

    

    
14. A method of producing a polypeptide according to any one of claims 1 to 13, or a pharmaceutically acceptable salt thereof, comprising: synthesizing the polypeptide.
15. Use of the polypeptide according to any one of claims 1 to 13, or a pharmaceutically acceptable salt thereof, for use in bacteriostatic, antibacterial or antibacterial or antibacterial substances.