Peptide with antibacterial activity

Abstract

The invention relates to organic chemistry, pharmacology and medicine and concerns a novel peptide. The claimed peptide is characterized by a high level of broad-spectrum antibacterial activity and is promising for use in the treatment of infectious diseases caused by bacterial infections, including diseases caused by antibiotic-resistant (including multidrug-resistant) bacterial strains. The compound according to the invention is characterized by the ability to inhibit strains of pathogens, including strains of pathogens which are not responsive to widely-used drugs, in particular carbapenem-resistant bacterial isolates and vancomycin-resistant bacterial isolates..

Description

[0001] Peptide with antibacterial activity

[0002] Field of technology

[0003] The invention relates to the chemistry of organic compounds, pharmacology and medicine and concerns a new peptide characterized by antibacterial activity, which, in particular, can be used for the prevention and treatment of infectious diseases in a subject.

[0004] State of the art

[0005] In recent decades, humanity has faced a global problem of antimicrobial resistance, the main cause of which is natural evolutionary selection due to the excessive use of antimicrobials in medicine and agriculture. It is estimated that by the middle of the 21st century, more than 10 million deaths annually may be associated with antibiotic resistance. Recent work has shown that now worldwide more than 1.2 million deaths annually are directly caused by antibiotic-resistant infections [Murray, CJ; Ikuta, KS; Sharara, F.; Swetschinski, L.; Robles Aguilar, G.; Gray, A.; Han, C.; Bisignano, C.; Rao, P.; Wool, E.; et al. Global Burden of Bacterial Antimicrobial Resistance in 2019: A Systematic Analysis. Lancet 2022, 399, 629-655]. The widespread spread of antibiotic resistance requires a search for new effective antibacterial compounds.However, in recent decades, the number of registered antibiotics has been limited [Browne, K.; Chakraborty, S.; Chen, R.; Willcox, MD; Black, DS; Walsh, WR; Kumar, N. A New Era of Antibiotics: The Clinical Potential of Antimicrobial Peptides. Int. J. Mol. Sci. 2020, 21 , 7047].

[0006] In recent years, so-called antimicrobial peptides (hereinafter referred to as AMPs), which are the most important component of the innate immune system, have attracted great interest. AMPs are known to have pronounced antibacterial, antiviral, antifungal, antiparasitic and antitumor effects [Huan Y, Kong Q, Moi H, Yi H. Antimicrobial Peptides: Classification, Design, Application and Research Progress in Multiple Fields. Front Microbiol. 2020; 11:582779].

[0007] It is important to note that the probability of developing resistance to AMPs is significantly lower than to small molecule antibiotics [Yu, G.; Baeder, DY; Regoes, RR; Rolff, J. Predicting Drug Resistance Evolution: Insights from Antimicrobial Peptides and Antibiotics. Proc. R. Soc. In Biol. Sci. 2018, 285]. This is due to the mechanism of action of AMPs, which is based on direct damage to the bacterial cell wall through membrane permeabilization [The Mechanism of Membrane Permeabilization by Peptides: Still an Enigma. Aust. J. Chem. 2019, 73, 96-103]. This makes AMPs equally effective against sensitive and multidrug-resistant bacteria [Browne, K.; Chakraborty, S.; Chen, R.; Willcox, MD; Black, DS; Walsh, WR; Kumar, N. A New Era of Antibiotics: The Clinical Potential of Antimicrobial Peptides. Int. J. Mol. Sci. 2020, 21, 7047].

[0008] De novo developed peptides and modified peptides with antimicrobial activity are known from the prior art (US9707282B2; RU2468033, RU2702661, WO2013124436). A number of antimicrobial peptides are currently at the clinical trial stage, and some have already been approved for use in clinical practice [Dijksteel, Gabrielle S et al. “Review: Lessons Learned From Clinical Trials Using Antimicrobial Peptides (AMPs).” Frontiers in microbiology vol. 12 616979. 22 Feb. 2021].

[0009] Previously, the strategy of using AMPs in combination with traditional small molecule antibiotics was studied, and it was shown that the antibiotic resistance phenotype of bacteria does not affect the effectiveness of AMPs [Bolatchiev, A. Antibacterial Activity of Human Defensins against Staphylococcus Aureus and Escherichia Coli. PeerJ 2020, 8, e10455].

[0010] Thus, attempts to create new peptide molecules with pharmacological activity seem appropriate and relevant.

[0011] Disclosure of invention

[0012] The objective of the present invention is to develop and create new effective antibacterial agents that are promising for use in clinical practice for the treatment and / or prevention of infectious diseases.

[0013] The technical result of the invention is the development and production of a new peptide compound that has high antibacterial activity, a broad spectrum of action and is promising for use in the therapy of infectious diseases in a subject caused by a bacterial infection, including diseases caused by antibiotic-resistant (including multiresistant) bacterial strains. It has been unexpectedly established that the peptide according to the invention is characterized by the ability to inhibit pathogen strains, including pathogen strains that are not sensitive to widely used drugs, in particular carbapenem-resistant bacterial isolates, vancomycin-resistant bacterial isolates.

[0014] In addition, the peptide according to the invention expands the arsenal of available antibacterial agents for the treatment of infectious diseases caused, in particular, by multidrug-resistant bacterial strains.

[0015] The specified technical result is achieved by developing and creating a peptide with antibacterial activity, or its pharmaceutically acceptable salt, having the sequence SEQ ID NO: 1.

[0016] The present invention also includes the use of the peptide of the invention as an antibacterial agent. The subject of the present invention is also the use of the peptide of the invention for the preparation of a pharmaceutical composition with antibacterial activity for the treatment and / or prevention of an infectious disease in a subject caused by a bacterial infection.

[0017] In particular embodiments of the invention, the disease is caused by a gram-negative or gram-positive bacterium.

[0018] In particular embodiments of the invention, the disease is caused by an antibiotic-resistant bacterial strain.

[0019] In particular embodiments of the invention, the disease is caused by bacteria from the genus Acinetobacter, Pseudomonas, Staphylococcus, Proteus or Klebsiella.

[0020] In particular embodiments of the invention, the disease is caused by Pseudomonas aeruginosa, Acinetobacter baumannii, Klebsiella pneumoniae, Proteus vulgaris, Enterococcus faecalis or Staphylococcus aureus.

[0021] The subject of the present invention is also a pharmaceutical composition with antibacterial activity for the treatment and / or prevention of an infectious disease in a subject caused by a bacterial infection, containing an effective amount of a peptide according to the invention and at least one pharmaceutically acceptable excipient.

[0022] In particular embodiments of the invention, the pharmaceutically acceptable excipient is a carrier, filler and / or solvent.

[0023] In particular embodiments of the invention, the disease is caused by a gram-negative or gram-positive bacterium.

[0024] In particular embodiments of the invention, the disease is caused by an antibiotic-resistant bacterial strain.

[0025] In particular embodiments of the invention, the disease is caused by a bacterium from the genus Acinetobacter, Pseudomonas or Klebsiella.

[0026] In particular embodiments of the invention, the disease is caused by Pseudomonas aeruginosa, Acinetobacter baumannii, Klebsiella pneumoniae, Proteus vulgaris, Enterococcus faecalis or Staphylococcus aureus.

[0027] In particular embodiments of the invention, the subject is a human being.

[0028] The present invention also includes the preparation of compounds of the invention.

[0029] The present invention also includes a method for treating and / or preventing an infectious disease in a subject caused by a bacterial infection by administering to the subject a peptide of the invention or a pharmaceutical composition of the invention. The present invention also includes a method for blocking and / or inhibiting and / or suppressing the growth of bacteria, including pathogenic bacteria with multiple drug resistance (in particular, carbapenem-resistant bacteria, vancomycin-resistant bacteria), using a peptide of the invention or pharmaceutical compositions of the invention.

[0030] The present invention relates to a peptide or its pharmaceutically acceptable salts that exhibit antibacterial activity. These molecules exhibit a broad spectrum of activity against various pathogens (including antibiotic-resistant (including multiresistant) bacterial strains, etc.). The claimed peptide, in particular, exhibits pronounced antimicrobial activity against a broad spectrum of gram-negative or gram-positive antibiotic-resistant bacterial strains.

[0031] Detailed disclosure of the invention

[0032] Brief description of the drawings

[0033] Figure 1. Kaplan-Meier curve constructed based on the results of a study of the peptide of the invention in a mouse model of lethal generalized infection in vivo.

[0034] Figure 2. Mass spectrum (MS) of the peptide of the invention.

[0035] Figure 3. HPLC chromatogram of the peptide of the invention.

[0036] Definitions and terms

[0037] For a better understanding of the present invention, some terms used in the present description of the invention are provided below. The following definitions apply herein unless otherwise explicitly stated.

[0038] In the description of this invention, the terms "includes" and "including" are interpreted to mean "includes, among other things." These terms are not intended to be construed as "consists only of."

[0039] The term "and / or" means one, more, or all of the listed elements.

[0040] Also here, listing numeric ranges by endpoints includes all numbers within that range.

[0041] The term "optional" or "optional" or "optionally" as used herein means that the subsequently described event or circumstance may, but need not, occur and that the description includes instances in which the event or circumstance occurs and instances in which it does not occur.

[0042] The term "amino acid" also refers to naturally occurring amino acids (including both L-amino acids and D-amino acids). Amino acids are designated by standard abbreviations: arginine (Arg; R), leucine (Leu; L), lysine (Lys; K), phenylalanine (Phe; F), tryptophan (Trp; W). The compounds that are the essence of this invention may exist in radioisotope-labeled form, i.e., said compounds may contain one or more atoms whose atomic mass or mass number differs from the atomic mass or mass number of the most common natural isotopes. Radioisotopes of hydrogen, carbon, phosphorus, chlorine include 3 H, 14WITH, 32 R, 35 S, and 36 C1, respectively. Compounds of the present invention that contain such radioisotopes and / or other radioisotopes of other atoms are within the scope of the present invention. Tritiated, i.e. 3 H and carbon, i.e. 14 Radioisotopes are particularly preferred due to their ease of preparation and detection.

[0043] The radiolabeled compounds of the present invention can be prepared by methods well known to those skilled in the art. The labeled compounds can be prepared by the procedures described herein by simply replacing the unlabeled reagents with the appropriate labeled reagents.

[0044] The compounds of the present invention may exist in free form or, if desired, in the form of a pharmaceutically acceptable salt or other derivative. The term "pharmaceutically acceptable salt" as used herein refers to those salts which, within the limits of medical judgment, are suitable for use in contact with human and animal tissues without undue toxicity, irritation, allergic reaction, etc., and which meet a reasonable benefit-risk ratio. Pharmaceutically acceptable salts of amines, carboxylic acids, phosphonates and other types of compounds are well known in the medical arts. Salts may be prepared in situ during the isolation or purification of the compounds of the invention, or may be prepared separately by reacting a free acid or a free base of a compound of the invention with a suitable base or acid, respectively.Examples of pharmaceutically acceptable, non-toxic acid salts include salts of the amino group formed with inorganic acids such as hydrochloric, hydrobromic, phosphoric, sulfuric and perchloric acids, or organic acids such as acetic, oxalic, maleic, tartaric, succinic or malonic acids, or obtained by other methods used in the art, such as ion exchange.Other pharmaceutically acceptable salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecyl sulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanate, hydroiodide, 2-hydroxyethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valeriate, and the like. Typical alkali and alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and others.In addition, pharmaceutically acceptable salts may contain, if desired, non-toxic ammonium, quaternary ammonium and amine cations derived from such counterions as halides, hydroxides, carboxylates, sulfates, phosphates, nitrates, lower alkyl sulfonates and aryl sulfonates.

[0045] By "therapeutically effective amount" is meant that amount of the compound administered or delivered to a patient that is most likely to produce the desired response to treatment. The exact amount required may vary from subject to subject depending on the age, weight, and general condition of the patient, the severity of the disease, the method of administration, combination therapy with other drugs, etc.

[0046] By "prophylactically effective amount" is meant that amount of the compound administered or delivered to a patient that is most likely to produce the desired response to the prophylaxis of infectious diseases caused by bacterial infection. The exact amount required may vary from subject to subject depending on the age, body weight, and general condition of the patient, the severity of the disease, the method of administration of the drug, combination therapy with other drugs, etc. For prophylactic treatment, a therapeutically or prophylactically effective amount is that amount that will be effective in preventing a microbial (e.g., bacterial) infection.

[0047] The term "patient" ("subject") includes all mammalian species, preferably humans, that utilize the compounds of the present invention either by self-administration and / or administration to the patient by another person for the treatment and / or prevention of a disease or medical condition.

[0048] The terms "treatment" and "therapy" cover the treatment of pathological conditions in mammals, preferably in humans, and include: a) blocking (stopping) the course of the disease, b) alleviating the severity of the disease, i.e. inducing regression of the disease.

[0049] The term "prophylaxis", "prevention", "preventive therapy" covers the elimination of risk factors, as well as preventive treatment of subclinical stages of the disease in humans, aimed at reducing the likelihood of the clinical stages of the disease. Patients for preventive therapy are selected based on factors that, based on known data, entail an increased risk of developing clinical stages of the disease compared to the general population. Preventive therapy includes a) primary prevention and b) secondary prevention. Primary prevention is defined as preventive treatment in patients who have not yet reached the clinical stage of the disease. Secondary prevention is the prevention of recurrence of the same or a similar clinical state of the disease.

[0050] The term risk reduction refers to therapy that reduces the incidence of clinical disease. Examples of risk reduction include primary and secondary disease prevention.

[0051] The term "antibacterial agent" as used herein refers to a compound or combination of compounds capable of: (i) inhibiting, reducing, or preventing the growth of bacteria; (ii) inhibiting or reducing the ability of bacteria to cause infection in a subject; or (iii) inhibiting or reducing the ability of bacteria to multiply or remain infectious in the environment. The term "antibacterial agent" also refers to compounds capable of reducing the infectivity or virulence of bacteria.

[0052] It will be understood by one skilled in the art that the peptide described herein can exist and is often used in the form of its pharmaceutically acceptable derivatives, such as salts, prodrugs, metabolites, esters, ethers, hydrates, polymorphs, solvates, complexes, enantiomers or other pharmaceutically acceptable derivatives. Therefore, reference to a peptide described herein is intended to include such pharmaceutically acceptable salts, prodrugs, metabolites, esters, ethers, hydrates, polymorphs, solvates, complexes, enantiomers or any other pharmaceutically acceptable derivatives thereof.

[0053] Pharmaceutical compositions

[0054] The invention also relates to pharmaceutical compositions that comprise a peptide of the invention (or a prodrug, a pharmaceutically acceptable salt or another pharmaceutically acceptable derivative) and one or more pharmaceutically acceptable carriers, adjuvants, solvents and / or excipients, such that can be administered to a patient together with the compound that is the essence of this invention and that do not destroy the pharmacological activity of this compound and are non-toxic when administered in doses sufficient to deliver a therapeutic amount of the compound.

[0055] The pharmaceutical compositions specified in this invention comprise the peptides of the present invention together with pharmaceutically acceptable carriers, which may include any solvents (in particular water), diluents, dispersions or suspensions, surfactants, isotonic agents, thickeners and emulsifiers, preservatives, binders, lubricants, etc., suitable for a particular dosage form.Materials that can serve as pharmaceutically acceptable carriers include, but are not limited to, mono- and oligosaccharides and derivatives thereof; gelatin; talc; excipients such as cocoa butter and suppository wax; oils such as peanut, cottonseed, sesame, olive, corn and soybean oils and others; glycols such as propylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffers such as magnesium hydroxide and aluminum hydroxide; alginic acid; depyrogenated water; isotonic solution, Ringer's solution; alcohol and phosphate buffer solutions. The composition may also contain other non-toxic compatible lubricants such as sodium lauryl sulfate and magnesium stearate, as well as colorants, release agents, film agents, sweeteners, flavorings and aromas, preservatives and antioxidants.

[0056] The subject of the present invention is also dosage forms - a class of pharmaceutical compositions, the structure of which is optimized for a specific method of administration into the body in a therapeutically effective dose, for example, for administration into the body intravenously, orally, intramuscularly, subcutaneously, intraocularly, by inhalation, intranasally and sublingually, in recommended dosages.

[0057] The dosage forms of this invention may contain structures obtained by methods of using liposomes, microencapsulation methods, methods of obtaining nanoforms of a drug or other methods known in pharmaceuticals.

[0058] For parenteral administration, aqueous suspensions, isotonic saline solutions or sterile injection solutions are used, the compatible agents of which contain pharmacological agents, for example, propylene glycol or butylene glycol.

[0059] In particular embodiments of the invention, the compositions of the invention may include a peptide of the present invention conjugated to polyethylene glycol, namely conjugation of the N-terminus of the peptide to polyethylene glycol (hereinafter referred to as PEG) [ACS Appl. Mater. Interfaces 2020, 12, 41, 46991-47001; https: / / doi.org / 10.1021 / acsami.0c13492]. There are various variations of PEG - depending on its molecular weight.

[0060] This approach allows to reduce immunogenicity, increase solubility and half-life of protein drugs [Pasut G. Pegylation of biological molecules and potential benefits: pharmacological properties of certolizumab pegol. BioDrugs. 2014;28 Suppl 1:S15-S23. doi:10.1007 / s40259-013-0064-z], PEG has high solubility in water and many organic solvents, is non-toxic and non-immunogenic. PEGylation of biologically active compounds allows to increase metabolic stability by creating steric hindrances that protect the molecule from proteases, and thus increase the circulation time in vivo.

[0061] Methods of therapeutic use The compound of the present invention is an antibacterial agent and therefore it is a useful agent for the treatment and / or prevention of a bacterial infection of a subject, including an infection caused by antibiotic-resistant (including multi-resistant) bacterial strains.

[0062] In another aspect, the invention also relates to methods of treating or preventing a bacterial infection in a subject, comprising administering to the subject an effective amount of a peptide of the invention. Subjects to be treated include dogs, cats, horses, cattle, sheep, pigs, poultry, primates (e.g., rhesus and cynomolgus macaques, monkeys, marmosets, tamarins, chimpanzees, macaques), rabbits and rodents (rats, mice, guinea pigs and the like). In one embodiment, the subject is a human, and the antimicrobial peptide of the invention can be delivered topically, intranasally, intraocularly. The antimicrobial peptide can be delivered in the form of drops, spray, cream, gel, ointment and the like.

[0063] Infections that can be treated with the disclosed compounds include infections of the outer ear, middle ear infections such as acute otitis media, cranial sinus infections, eye infections, oral cavity infections such as tooth, gum and mucous membrane infections, upper respiratory tract infections, lower respiratory tract infections, genitourinary infections, gastrointestinal infections, gynecological infections, sepsis, bone and joint infections, skin and skin structure infections, burns, antibacterial prophylaxis in surgery and antibacterial prophylaxis of immunocompromised patients, such as those undergoing cancer chemotherapy or after organ transplantation. These infections can be treated in hospital settings or in the community using the various routes of administration described in this invention.

[0064] The compound or compositions described herein can also be used for prophylactic purposes. Accordingly, the compound or composition can be administered to a subject potentially at risk of developing a microbial infection. Subjects at risk of developing a microbial infection include individuals exposed to a particular microorganism belonging to pathogenic bacterial species; individuals with impaired immune systems, or subjects particularly vulnerable to infections due to impaired natural defenses (e.g., skin at risk due to burns or cuts).

[0065] The antimicrobial peptide described in the present invention can be used for the treatment or prevention of infectious diseases caused by various bacterial organisms, including infection with pathogenic bacterial species. Examples of bacterial infection include, but are not limited to, gram-positive and gram-negative aerobic and anaerobic bacteria, such as staphylococci, such as S. aureus; enterococci, such as E. faecalis; streptococci, such as S. pyogenes and S. pneumoniae; Escherichia species, such as E. coli, including enterotoxigenic, enteropathogenic, enteroinvasive, enterohemorrhagic and enteroaggregative strains of E. coli; propionibacterium strains, such as P. acnes; Haemophilus, such as H. influenzae; Moraxella, such as M. catarrhalis. Other examples include mycobacteria such as M. tuberculosis, M. avian-intracellulare, M. kansasii, M. bovis, M. africanum, M. genavense, M. leprae, M. xenopi, M. simiae, M. scrofulaceum, M. malmoense, M. celatum, M.abscessus, M. chelonae, M. szulgai, M. gordonae, M. haemophilum, M. fortuni and M. marinum; corynebacteria, such as C. diphtheriae; pseudomonas species, such as P. aeruginosa; Borrelia species, such as B. burgdorferi; Listeria species, such as L. monocytogenes, Bacillus species, such as B. cereus; Bordetella species, such as B. bronchiseptica; Klebsiella species, Clostridium species, such as C. perfringens, C. tetani; Chlamydia species, such as C. psittaci; Rickettsia species, such as R. rickettsii and R. prowazekii; Salmonella species, such as S. typhimurium; Yersinia species, such as Y. enterocolitica and Y. pseudotuberculosis; Klebsiella species, such as K. pneumoniae or K. aerogenes; and Mycoplasma species, such as M. pneumonia; Actinobacteria species, H. parasuis; and Trueperella pyogenes.

[0066] In certain aspects of the invention, staphylococcal bacteria have been selected, for example, S. pseudintermedius, S. aureus, S. schleiferi, S. chromogenes, S. simulans, S. xylosus. Also, streptococcal bacteria can be selected, for example, S. uberis, S. agalactiae, S. dysgalactiae, S. suis. In addition, bacteria of the Pasteurellaceae family are also suitable for treatment with the compositions described in this invention. Suitable bacteria of the Pasteurellaceae family include M. haemolytica, P. multocida, H. somni, Escherichia species, for example, E. coli and Klebsiella species.

[0067] In certain embodiments of the invention, the bacteria are S. pseudintermedius and / or P. aeruginosa.

[0068] In certain embodiments of the invention, the bacteria are bacteria of the genus Acinetobacter, in particular Acinetobacter baumannii.

[0069] In certain embodiments of the invention, the bacteria are Proteus vulgaris.

[0070] For therapeutic use, the compounds of the invention may be administered by means of a pharmaceutical composition in any pharmaceutical dosage form by any route of administration. Dosage forms typically include a pharmaceutically acceptable carrier suitable for the particular dosage form selected. In particular, a compound of the invention may be administered daily for a period of time necessary to treat and / or prevent diseases relevant to the patient, including a course of therapy lasting days, months, years or the patient's lifetime. Routes of administration include, but are not limited to, intravenous, intramuscular, oral, subcutaneous, intraocular, inhalation, intranasal and sublingual. The preferred route of administration is intravenous.

[0071] The invention also relates to a pharmaceutical composition containing a daily dose of said compound in the form of a fixed dosage unit, and to a combination containing said pharmaceutical composition or said compound. In a preferred embodiment, said composition for use according to the invention is administered once a day in a dosage of 1 mg or more of the selected compound according to the invention. The preferred dosage is 1-500 mg. The most preferred dosage is 10-200 mg.

[0072] One or more additional pharmacologically active agents may be administered in combination with the peptide of the invention. In general, any additional single or multiple active agents other than the compounds of the invention, including but not limited to other antibacterial drugs, may be used in any combination with the compound of the invention in one or a separate dosage form, allowing simultaneous or sequential therapeutic action of the active agents.

[0073] Implementation of the invention

[0074] Although the invention has been described with reference to the disclosed embodiments, it will be apparent to those skilled in the art that the specific experiments described in detail are provided merely for the purpose of illustrating the present invention and should not be considered as limiting the scope of the invention in any way. It will be understood that various modifications can be made without departing from the spirit of the present invention.

[0075] Obtaining a peptide according to the invention

[0076] General methods for obtaining the peptide according to the invention

[0077] A person skilled in the art can easily synthesize the peptide of the invention. Standard methods for preparing synthetic peptides are well known in the art. The peptide of the invention can be synthesized using such commonly used methods as 1-BOC or FMOC protection of alpha-amino groups. Both methods involve stepwise synthesis, wherein one amino acid is added at each step, starting from the carboxyl end of the peptide. The peptide of the invention can also be synthesized by solid-phase peptide synthesis methods well known in the art.

[0078] In addition, the peptide of the invention can be obtained not only by chemical synthesis, but also by using biotechnological methods: nucleotide sequences encoding the amino acid sequence of the claimed peptide can be synthesized - these nucleotide sequences can be introduced into cells (using vectors or native nucleic acids), and the cells transformed in this way can be used to express the claimed peptides. The said vectors and nucleotide sequences can be introduced into the human body (and other living organisms) for expressing the claimed peptides directly in vivo. The said methods are also well known to specialists in this field of technology.

[0079] Synthesis of the peptide according to the invention

[0080] The peptide of the invention was synthesized by solid-phase Fmoc synthesis using a JBMS-96-A automatic synthesizer (Jianbang Pharmacy Technology Co., Ltd.). The peptide was purified (purity>93%) using reversed-phase high-performance liquid chromatography (Table 2).

[0081] Some particular embodiments of the invention are disclosed in Table 1 below.

[0082] Table 1. Examples of peptides according to the invention.

[0083] HPLC of the peptide of the invention was performed on a YMC-Triart C18 column (4.6*250mm*5um), eluting with 0.1% trifluoroacetic acid in 100% water (solvent A) and 0.1% trifluoroacetic acid in 100% acetonitrile (solvent B), at a flow rate of 1 ml / min (Fig. 3).

[0084] Table 2. HPLC and mass spectroscopy data for the peptide of the invention.

[0085] The claimed peptide may be included in the composition of other large molecules (proteins, peptides, nucleic acids, carbohydrates, lipids) without changing their pharmacological activity or with the purpose of imparting new properties to them. In addition, derivatives of the claimed peptide may be obtained by chemical modification of the terminal sections of amino acids.

[0086] Characteristics of biological activity Spectrum of antibacterial activity of the peptide according to the invention in vitro

[0087] The in vitro study of the efficacy of the claimed peptide was conducted using serial dilutions. Pure bacterial cultures were grown on a solid nutrient medium (mannitol-salt agar, BioMedia, St. Petersburg, Russia). A suspension was prepared from the fresh morning culture in sterile saline, which corresponded to the turbidity standard of 0.5 McFarland (equivalent to 1-2x10 8CFU / mL). The suspension was dissolved in Mueller Hinton Broth (BBL™ Mueller Hinton Broth, Becton, Dickinson and Company, USA) to obtain an inoculum with an approximate concentration of 5x10 5CFU / mL. The inoculum (100 μl) was then added to the wells of sterile microplates (Medpolymer, St. Petersburg, Russia), into which 100 μl of the peptide of the invention were then added at various concentrations (from 0 to 32 μg / ml). Controls: sterility control (only Mueller-Hinton broth, without inoculum) and growth control (bacterial inoculum without the claimed peptide). The microplates were then incubated in an incubator at 37 °C. After 18-20 h, the minimum inhibitory concentration (MIC) values ​​were estimated. The MIC was considered to be the minimum concentration of the peptide compound at which there was no visual growth in the corresponding well [Wiegand, Irith et al. “Agar and broth dilution methods to determine the minimal inhibitory concentration (MIC) of antimicrobial substances.” Nature protocols vol. 3.2 (2008): 163-75].

[0088] The study of the antibacterial activity of the claimed compound was conducted against clinical multidrug-resistant gram-negative carbapenem-resistant bacteria and gram-positive vancomycin-resistant bacteria, the results are presented in Table 3 below.

[0089] Table 3. Minimum inhibitory concentrations (MIC; μg / ml) of the ABP5 peptide according to the invention in relation to clinical isolates of bacteria (MIC values ​​are presented as median; n is the number of bacterial isolates studied). Thus, the obtained research results show that the peptide according to the invention has high antibacterial activity, including against antibiotic-resistant (including multi-resistant) bacterial strains.

[0090] Study of the activity of the peptide according to the invention in vivo

[0091] A murine model of lethal generalized infection was used for screening the efficacy of the ABP5 peptide in vivo. To model sepsis, ICR (CD-1) mice (males, average weight = 30 g) were injected intraperitoneally with a bacterial suspension of a carbapenem-resistant isolate of K. pneumoniae (~6.75*10 8 CFU / mouse). There were 2 groups of mice in the experiment (12 individuals in each group): Group 1 — control (received saline), Group 2 — received ABP5, at an empirical dose of 100 μg / mouse; ABP5 was administered 30 minutes after infection, once intraperitoneally. Survival was assessed every 24 hours for 5 days. For statistical analysis, the survival function was estimated using the Kaplan-Meier method (https: / / www.statskingdom.com / kaplan-meier.htnnl).

[0092] The study showed that the introduction of the ABP5 peptide significantly increased the probability of survival compared to the control group (p=0.0004074). By the end of the experiment, 120 hours after infection, the percentage of surviving animals in the ABP5 group was 58%, versus 0% in the control group. Interestingly, only 48 hours after infection, only 16% of animals survived in the control group, while in the ABP5 group, 91% (Fig. 1).

[0093] Although the invention has been described with reference to the disclosed embodiments, it will be apparent to those skilled in the art that the specific experiments described in detail are provided merely for the purpose of illustrating the present invention and should not be considered as limiting the scope of the invention in any way. It will be understood that various modifications can be made without departing from the spirit of the present invention.

Claims

Invention formula 1. A peptide with antibacterial activity, or a pharmaceutically acceptable salt thereof, having the sequence SEQ ID NO:

1.

2. Use of the peptide according to paragraph 1 as an antibacterial agent.

3. Use of a peptide according to claim 1, for obtaining a pharmaceutical composition with antibacterial activity for the treatment and / or prevention of an infectious disease in a subject caused by a bacterial infection.

4. Use according to paragraph 3, characterized by the fact that the disease is caused by a gram-negative or gram-positive bacterium.

5. Use according to paragraph 3, characterized by the fact that the disease is caused by an antibiotic-resistant bacterial strain.

6. Use according to paragraph 4, characterized in that the disease is caused by a bacterium from the genus Acinetobacter, Pseudomonas, Staphylococcus, Proteus or Klebsiella.

7. The use according to claim 4, characterized in that the disease is caused by Pseudomonas aeruginosa, Acinetobacter baumannii, Klebsiella pneumoniae, Proteus vulgaris, Enterococcus faecalis or Staphylococcus aureus.

8. A pharmaceutical composition with antibacterial activity for the treatment and / or prevention of an infectious disease in a subject caused by a bacterial infection, containing an effective amount of the peptide according to claim 1 and at least one pharmaceutically acceptable excipient.

9. The pharmaceutical composition according to claim 8, characterized in that the pharmaceutically acceptable excipient is a carrier, filler and / or solvent.

10. The pharmaceutical composition according to claim 8, characterized in that the disease is caused by a gram-negative or gram-positive bacterium.

11. The pharmaceutical composition according to claim 8, characterized in that the disease is caused by an antibiotic-resistant bacterial strain.

12. A pharmaceutical composition according to claim 8, characterized in that the disease is caused by a bacterium from the genus Acinetobacter, Pseudomonas or Klebsiella.

13. The pharmaceutical composition according to claim 8, characterized in that the disease is caused by Pseudomonas aeruginosa, Acinetobacter baumannii, Klebsiella pneumoniae, Proteus vulgaris, Enterococcus faecalis or Staphylococcus aureus.

14. The pharmaceutical composition of claim 8, wherein the subject is a human.

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