WO2021211006A1

WO2021211006A1 - Inhaled hexapeptide for treating interleukin-6 related respiratory diseases

Info

Publication number: WO2021211006A1
Application number: PCT/RU2020/000385
Authority: WO
Inventors: Vladislav Nikolaevich KARKISCHENKO; Veronika Igorevna SKVORTSOVA; Igor Anatolievich Pomytkin; Aleksandr Sergeevich SAMOYLOV; Tatyana Alekseevna ASTRELINA; Yurij Dmitrievich UDALOV
Original assignee: Federal State Budgetary Institution Center Of Biomedical Technologies Of Federal Medical And Biological Agency
Priority date: 2020-04-13
Filing date: 2020-07-27
Publication date: 2021-10-21

Abstract

The present invention is in the field of healthcare. More specifically, the invention relates to hexapeptide of formula H-Tyr-D-Ala-Gly-Phe-Leu-Arg-OH (I) or a pharmaceutically acceptable salt thereof and an aqueous pharmaceutical compositions thereof for use in the treatment of an interleukin-6 related respiratory disease by pulmonary administration to a mammal in need thereof. Preferably, the pharmaceutically acceptable salt is H-Tyr-D-Ala-Gly-Phe-Leu-Arg-OH diacetate. Preferably, the composition is administered pulmonary in form of aerosol having particle size from 1 to 5 microns. Further, the invention relates to a method of treating an interleukin-6 related respiratory disease, which comprises a step of pulmonary administration of an effective amount of a hexapeptide of formula H-Tyr-D-Ala-Gly-Phe-Leu-Arg-OH (I) or a pharmaceutically acceptable salt thereof to a mammal in need thereof. Preferably, the effective amount of a hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof is from 0.001 to 1.000 mg/kg per kg body weight of the mammal. Preferably, the interleukin-6 related respiratory disease is selected from the group consisting of acute respiratory distress syndrome, acute lower respiratory infection, pneumonia, pulmonary edema, bronchitis, tracheobronchitis, chronic obstructive pulmonary disease, and asthma.

Description

Inhaled hexapeptide for treating interleukin-6 related respiratory diseases

Field of the Invention

[0001] The present invention is in the field of healthcare. More specifically, the invention relates to use of inhaled hexapeptide in the treatment of interleukin-6 related respiratory diseases.

Background of the invention

[0002] Respiratory diseases, also known as lung diseases, are group of diseases affecting the respiratory system. According to World Health Organization (WHO) International Classification of Diseases 10th Revision (ICD-10), diseases of the respiratory system are classified as acute upper respiratory infections (J00-J06), influenza and pneumonia (J09-J18), other acute lower respiratory infections (J20-J22), other diseases of upper respiratory tract (J30-J39), chronic lower respiratory diseases (J40-J47), lung diseases due to external agents (J60-J70), other respiratory diseases principally affecting the interstitium (J80-J84), suppurative and necrotic conditions of lower respiratory tract (J85-J86), other diseases of pleura (J90-J94), and other diseases of the respiratory system (J95-J99). Respiratory diseases are leading causes of death and disability in the world. Chronic obstructive pulmonary disease (COPD) affects more than 200 million people in the world. Asthma afflicts up to 334 million people worldwide. Lower respiratory tract infection and pneumonia are two of the leading causes of death, accounting for more than 4 million fatalities annually. Forum of International Respiratory Societies. The Global Impact of Respiratory Disease. 2nd Edition. Sheffield, European Respiratory Society, 2017.

[0003] lnterleukin-6 (IL-6) is a small size cytokine (21 KDa). It is produced by cells of the innate immune system and pulmonary epithelial cells. IL-6 is a major regulator of acute phase immune process and antibodies production. However, IL-6 overproduction contributes to the development of a variety of respiratory diseases and predicts its severity. Overexpression of IL- 6 in pulmonary epithelial cells was observed in patients with asthma and other respiratory diseases. Rincon M, Irvin CG. Role of IL-6 in asthma and other inflammatory pulmonary diseases. Int J Biol Sci. 2012;8(9):1281-90. Increased levels of IL-6 were found in asthmatic patients. Yokoyama A et at. Am J Respir Crit Care Med. 1995 May;151 (5):1354-8. Elevated IL-6 levels were found to be predictive of increase mortality in COPD patients. Celli BR et al.. Am J Respir Crit Care Med. 2012, 185(10): 1065-72. Elevated IL-6 levels were associated with an acute respirtory distress-syndrome (ARDS) and pneumonia caused by a variety of virus infections. McGonagle D et al. Autoimmun Rev. 2020, 19(6): 102537. Liu B et al. J Autoimmun. 2020, 111:102452. Elevated IL-6 levels were observed in influenza A/H1N1 virus pneumonia. Davey RT Jr et al. PLoS One. 2013;8(2):e57121. Elevated IL-6 levels predicted the severity of the disease in patients with community-acquired pneumonia. Ramirez P et al. Crit Care Med. 2011, 39(10):2211-7. High IL-6 concentrations in serum and bronchoalveolar lavage fluid were associated with the disease severity and poor outcome in pneumonia patients. Bordon J et al. Int J Infect Dis. 2013, 17(2):e76-83. Higher levels of IL-6 were associated with early mortality of patients with community-acquired pneumonia. Bacci MR et al. J Med Biol Res. 2015, 48(5):427- 32. The elevation of IL-6 concentrations reflected the severity of disease in patients with community-acquired pneumonia. Zobel K et al. BMC Pulm Med. 2012, 12:6. The concentrations of IL-6 were higher in nonsurvivors than in survivor patients with community-acquired pneumonia. Lee YL et al. J Crit Care. 2010, 25(1):176.e7-13. Increased IL-6 levels were associated with pulmonary fibrosis. Papiris SA et al. Cytokine. 2018, 102:168-172. Elevated IL-6 levels increased risk of death in hospitalized patients with pneumonia. Andrijevic I et al. Ann Thorac Med. 2014, 9(3):162-7. Thus, IL-6 overexpression contributes to severity of respiratory diseases. Therefore, there is a great need in safe and effective approaches to antagonize IL-6 activity in patients with respiratory diseases.

[0004] The coronavirus disease 2019 (COVID-19) is caused by SARS-CoV-2 virus and is characterized by diffuse alveolar damage of the lung, microthrombosis, and cytokine release syndrome. Elevated IL-6 levels were found to be associated with an acute respirtory distress- syndrome (ARDS) and pneumonia caused by SARS-CoV-2 virus. McGonagle D et al. Autoimmun Rev. 2020, 19(6):102537. Early elevated IL-6 levels predicted in-hospital mortality for patients wth coronavirus disease 2019 (COVID-19). Luo M et al. JCI Insight. 2020:139024. Elevated IL-6 levels in patients with COVID-19 were the predictor of most severe course of the disease and the need for intensive care. Gubernatorova EO et al. Cytokine Growth Factor Rev. 2020, 53:13-24. Meta-analysis of numerous studies on COVID-19 has demonstrated that a threshold IL-6 level of 1.7 pg/ml can be used as a discriminator between severe and non-severe forms of the disease and IL-6 level of 4.6 pg/ml can be used as a discriminator between nonsurvivors vs. survivors prognosis. Henry BM et al. Clin Chem Lab Med. 2020, 58(7):1021-1028. Therefore, there is a great need in safe and effective approaches to antagonize IL-6 activity in patients with COVID-19.

[0005] Anti-interleukin-6 agents are a class of therapeutic agents that directed against IL-6 itself (e.g. siltuximab) or IL-6 receptor (e.g. tocilizumab). Tocilizumab is a recombinant humanized anti-human interleukin 6 receptor monoclonal antibody of the immunoglobulin IgGl subclass with a molecular weight of approximately 148 kDa. European patent 0628639B; US patent 5,795,965; Japanese patent 3370324B. Tocilizumab binds to both membrane (mlL-6R) and soluble (slL-6R) interleukin-6 receptors (IL-6R), thereby antagonizing action of IL-6. China's National Health Commission included the use of tocilizumab in guidelines to treat COVID-19 patients. Tocilizumab has been shown to reduce the severity of COVID-19 disease and mortality in a group of COVID-19 patients. Zhang C et al. Int J Antimicrob Agents. 2020, 55(5): 105954. Xu X et al. Proc Natl Acad Sci USA. 2020, 117(20):10970-10975. However, efficacy and safety of anti-interleukin-6 agents in the treatment of COVID-19 has not yet been demonstrated in multi-centered phase 3 clinical trials.

[0006] The hexapeptide H-Tyr-D-Ala-Gly-Phe-Leu-Arg-OH is an analogue of dynorphin A (1-6) with Gly to D-Ala substitution at position 2 of the amino acid sequence. The hexapeptide is a strong base and is used in form of non-toxic salts with acids. Diacetate salt of hexapeptide is a a commercially available drug in Russian Federation in form of injections under the name dalargin for the treatment of diseases of the digestive system, specifically acute pancreatitis, gastric ulcers, and duodenal ulcers, with history of clinical use for more than 30 years. The hexapeptide demonstrated a broad spectrum of biological activities within a dose range from 1 to 1000 pg/kg of the body weight of a mammal. The use of hexapeptide in form of intramuscular injections or intravenous infusions are disclosed in RU patents 2032422, 2241488, 2198641 , 2104717, 2270025, 2351334, 2343885, 2405534, 2363455, 2515550, 2180598, 2635083,

2436588, 2473325, 2144831 , 2646569, 2200026, 2146530, 2155608, 2185176, 2139725,

2266130, 2326661, 2299742, 2218896, 2416398, 2318503, 2430753, 2299438, 2258529,

2099077, 2122415, 2261713, 2006039, 2366416, 2228762, 2185849, 2496493, 2366417,

2017488, 2167671, 2181564, 2284192, 2290203, 2286793, 2366432, 2299065, 2429002,

2196603, 2142814, 2285522, 2203693, 2230549, 2180591, 2142736, 2113856, 2217139,

2266752, 2261722, 2362580, 2223741 , 222814869, 2217186, 2266130, 2326661 , 2299742, 2218896, 2657416, 2416398, 2318503, 2430753, 2299438, 2258529, 2099077, 2122415, 2261713, 2006039, 2366416. It has been shown that injections of the hexapeptide are useful for reducing IL-6 levels in plasma of patients suffering from the coronary heart disease. Dontsov AV. Klin Med (Mosk). 2017, 95(2):127-31 [in RUSJ. RU patent 2672888 disclosed the use of the intranasal hexapeptide as an agent having direct antiviral activity in the treatment of acute respiratory virus infections. The intramuscular injections of hexapeptide has been used for treating pneumonia in adults. Borovskaya et al. Bulletin physiology and pathology of respiration. 2004, 17:85-88 [in Rus]. The intravenous infusions of hexapepetide has been used for treating respiratory distess-syndrome in newborns. Bichenov RG. The role of dalargin in the complex treatment of newborn children with respiratory distress syndrome. 14.00.37. Anesthesiology and resuscitation. PhD thesis. Rostov-on-Don, 2000 [in Rus]. However, efficacy of the treatment of respiratory diseases using injectable forms of the hexapeptide is limited by, at first, a very fast elimination of the hexapeptide from circulation with half-life of few minutes due to fast degradation by peptidases and, at second, a very low absorption of the hexapeptide by lung tissues, accounting for 0.4% of the total dose of the drug administered by intravenous injection. Kalenikova El et al. Pharmacokinetics of dalargin. Problems of Biological, Medical and Pharmaceutical Chemistry. 1988, 34 (1): 75-83 [in Rus].

[0007] Pulmonary drug delivery represents a drug administration method that provides direct topical treatments for respiratory diseases. Compared to intravenous or intramuscular injections, it provides a painless and safer alternative. However, the elevation of concentrations of peptidases such as elastase and cathepsin G from neutrophils in the lungs occurring at inflammation has been identified as a potential barrier for delivery of peptides by inhalation to treat respiratory diseases, because these enzymes facilitate a rapid hydrolytic degradation of the peptides in lungs. Fellner RC et al. Mol Cell Pediatr. 2016, 3(1):16. Pathological high IL-6 concentrations were found to cause neutrophil degranulation and a significant release of neutrophil peptidases in a concentration-dependent manner. Bank U et al. Inflammation. 1995, 19(1):83-99. In this context, the elevated neutrophil peptidases activity in the lungs caused by elevated IL-6 concentrations is the potential barrier for delivery of peptides by pulmonary route to treat IL-6 related respiratory diseases.

[0008] Nothing is known from prior art about efficacy of pulmonary administered hexapeptide of formula H-Tyr-D-Ala-Gly-Phe-Leu-Arg-OH or salts thereof in the treatment of IL-6 related respiratory diseases.

Summary of the invention

[0009] The inventors surprisingly found that pulmonary administered hexapeptide of formula (I):

H-T yr-D-Ala-Gly-Phe-Leu-Arg-OH (I) is much more effective for preventing, diminishing, or eliminating symptoms of IL-6 related respiratory diseases in a mammal compared to the hexapeptide administered by injections in the same dose.

[0010] A first aspect of the invention relates to a hexapeptide of formula (I):

H-Tyr-D-Ala-Gly-Phe-Leu-Arg-OH (I) or a pharmaceutically acceptable salt thereof for use in the treatment of an interleukin-6 related respiratory disease by pulmonary administration to a mammal in need thereof.

[0011] According to another aspect, the invention relates to an aqueous pharmaceutical composition comprising a hexapeptide of formula (I):

H-Tyr-D-Ala-Gly-Phe-Leu-Arg-OH (I) or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable excipient for use in the treatment of an interleukin-6 related respiratory disease by pulmonary administration to a mammal in need thereof.

[0012] According to another aspect, the invention relates to a method of treating an interleukin-6 related respiratory disease, which comprises a step of pulmonary administration of an effective amount of a hexapeptide of formula (I):

H-Tyr-D-Ala-Gly-Phe-Leu-Arg-OH (I) or a pharmaceutically acceptable salt thereof to a mammal in need thereof.

[0013] In a preferred embodiment, the pharmaceutically acceptable salt of the hexapeptide of formula (I) is diacetate.

[0014] In preferred embodiments, the interleukin-6 related respiratory disease is selected from the group consisting of acute respiratory distress syndrome, acute lower respiratory infection, pneumonia, pulmonary edema, bronchitis, tracheobronchitis, chronic obstructive pulmonary disease, and asthma. More preferably, the acute lower respiratory infection is COVID-19.

[0015] In preferred embodiments, the hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof is administered pulmonary in form of aerosol having particle size from 0.1 to 10 microns, more preferably, from 1 to 5 microns.

[0016] In preferred embodiments, the aqueous pharmaceutical composition comprising a hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof is administered pulmonary in form of aerosol having particle size from 0.1 to 10 microns, more preferably, from 1 to 5 microns.

[0017] In preferred embodiments, the effective amount of hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof for use in the method of the invention is from 0.001 to 1.000 mg per kg body weight of the mammal.

[0018] Because of pulmonary route of administration of the hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof, this invention provides a painless and safer alternative for the hexapeptide injections during the course of the treatment of interleukin-6 related respiratory diseases. [0019] Because of direct topical delivery of the hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof to the lung, this invention provides particularly advantageous method for achieving the therapeutically effective levels of the hexapeptide (I) in the lung, which levels cannot been achieved with the hexapeptide (I) injections.

[0020] Because of the enhanced delivery of the hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof to the lung by pulmonary administration, this invention provides particularly advantageous compositions and methods over the hexapeptide injections for preventing, diminishing, or eliminating symptoms of an interleukin-6 related respiratory disease in a mammal in need thereof.

Brief Description of the Drawings

[0021] FIG.1 shows representative hematoxylin-eosin stained histological preparations of mouse lungs obtained before (0 h) and after (24, 72, and 120 h) induction of acute lung injury, whch injury is characterized by elevated levels of interleukin-6 in the lungs.

[0022] FIG.2 shows meaniSEM mRNA levels of interleukin-6 in lungs of healthy mice (“Intact”), mice with acute lung injury induced by sequential administration of a-galactosylceramide and lipopolysaccharide (“LPS”), mice with acute lung injury induced by sequential administration of a-galactosylceramide and lipopolysaccharide and treated with intramuscular injections of hexapeptide (I) diacetate (“LPS+HP(I) i.m.”), and mice with acute lung injury induced by sequential administration of a-galactosylceramide and lipopolysaccharide and treated with pulmonary administered hexapeptide (I) diacetate (“LPS+HP(I) inh.”).

[0023] FIG.3 shows the Kaplan-Meier survival curves for mice with acute lung injury (ALI) induced by sequential administration of a-galactosylceramide and lipopolysaccharide (“LPS”), mice with acute lung injury induced by sequential administration of a-galactosylceramide and lipopolysaccharide and treated with intramuscular injections of hexapeptide (I) diacetate (“LPS+HP(I) i.m.”), and mice with acute lung injury (ALI) induced by sequential administration of a-galactosylceramide and lipopolysaccharide and treated with pulmonary administered hexapeptide (I) diacetate (“LPS+HP(I) inh.”).

[0024] FIG.4 shows the response rates in patients with severe COVID-19 treated with standard therapy (“Control”) or pulmonary administered hexapeptide (I) diacetate (“HP(I) inh”). A patient having serum IL-6 level less than 1.7 pg/ml after the treatment was considered to be a responder, and having serum IL-6 level higher than 1.7 pg/ml was considered to be a nonresponder. Detailed description of the invention

[0025] The inventors surprisingly found that pulmonary administered hexapeptide of formula (I):

[0026] A first aspect of the invention relates to a hexapeptide of formula (I):

[0027] The hexapeptide of formula (I), short name hexapeptide (I), has the chemical name Tyrosyl-D-Alanyl-Glycyl-Phenylalanyl-Leucyl-Arginine, chemical formula of C₃₅H₅₁N₉O8, molecular weight of 725.84 daltons, and CAS registry number 81733-79-1 as an identifier.

[0028] The present invention relates to all pharmaceutically acceptable salts and solvates of the hexapeptide of formula (I). Non-exclusive examples of such pharmaceutically acceptable salts include chloride, bromide, sulfate, acetate, pyruvate, malate, fumarate, and citrate.

[0029] In a preferred embodiment, the pharmaceutically acceptable salt of the hexapeptide of formula (I) is diacetate, the chemical name Tyrosyl-D-Alanyl-Glycyl-Phenylalanyl-Leucyl- Arginine diacetate.

[0030] The hexapeptide of formula (I) can be obtained by any method known in the art, in particular by the solid-phase synthesis. The hexapeptide of formula (I) and its salts and solvates are commercially available, for example, in the Bachem catalog No. 4030569.0100 (https://shop.bachem.com/4030569.htmn. The hexapeptide (I) diacetate is a commercially available active pharmaceutical ingredient (API) that is produced by several manufacturers in the Russian Federation, e.g. by Berachim Ltd (Obninsk, Russia).

[0031] As used herein, the term “interleukin-6”, short name “IL-6”, refers to native interleukin-6 from any species, including mouse, rat, bovine, and human, preferably human.

[0032] As used herein, the term “respiratory disease" refers to a disease affecting the respiratory system. The term “respiratory system” relates to the airway and the lung. [0033] As used herein, the term “interleukin-6 related respiratory disease” refers to a respiratory disease associated with interleukin-6 overproduction. Such respiratory diseases include acute respiratory distress syndrome (ARDS), COVID-19 related ARDS, acute lower respiratory infection such as COVID-19, pneumonia, a virus related pneumonia, COVID-19 related pneumonia, pulmonary edema, bronchitis, tracheobronchitis, chronic obstructive pulmonary disease (COPD), and asthma.

[0034] In preferred embodiments, the interleukin-6 related respiratory disease is selected from the group consisting of acute respiratory distress syndrome, acute lower respiratory infection, pneumonia, pulmonary edema, bronchitis, tracheobronchitis, chronic obstructive pulmonary disease, and asthma. More preferably, the acute lower respiratory infection is COVID-19.

[0035] As used herein, the term “pulmonary administration” refers to administration of a drug through the lungs by inhalation. The term “inhalation” used with respect to formulations and compositions of the invention is synonymous with “pulmonary administration.” As used herein, the term “inhalation” refers to inhaling the vapor or dispersion of solid or liquid particles with added medication. In specific examples, inhaling can occur through a nebulizer or other aerosol- delivery device.

[0036] In some embodiments, hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof can be administered pulmonary in form of aerosol. Numerous methods and devices are well-known from the art that can be employed to generate said aerosols in therapeutically useful size ranges and concentrations. Specifically, these are nebulizers, metered-dose inhalers (MDIs), and dry powder inhalers (DPIs). Pilcer G at at. Int J Pharm. 2010, 392(1 -2):1 -19. Nebulizers such as jet nebulizers or ultrasonic nebulizers are used for the delivery of aqueous pharmaceuticals. MDIs suspends or dissolves drug powders into liquid propellants and when a metered quantity of the propellant is released from the storage canister, the propellant evaporates and expands quickly to disperse the powdered drug or liquid droplet drug. Such propellants include, but are not limited to, chlorofluorocarbon, a hydrochlorofluorocarbon, or a hydrocarbon. DPIs delivers a precisely measured dose medicine into the lungs in dry powder form. It is designed to generate a drug powder aerosol onto or via the inspiratory air flow. In practicing the invention, any methods and devices, including nebulizers, MDIs, and DPIs may be used for pulmonary delivery of hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof to a patient in need thereof.

[0037] As used herein, the term “aerosol” refers to suspension of liquid or solid particles in a gaseous medium. In practicing the invention, hexapeptide (I) or a pharmaceutically acceptable salt thereof can be administed pulmonary as aerosol in form of liquid formulations and dry powders.

[0038] In a preferred embodiment, hexapeptide (I) or a pharmaceutically acceptable salt thereof is pulmonary administered in form of aerosol with particle size from 0.1 to 10.0 microns, more preferably from 1 to 5 microns.

[0039] As used herein, the term “treatment” or “treating” means to prevent, diminish, or inhibit the progress of a disease to which this term is applied, or one or more symptoms of this disease. The term “treatment” refers to both therapeutic treatment and prophylactic or preventative measures. Those in need of treatment include those already with the disorder as well as those prone to having the disorder or diagnosed with the disorder or those in which the disorder is to be prevented.

[0040] As used herein, “mammal" refers to any animal classified as a mammal, e.g. mouse, rat, bovine, and human. The preferred mammal herein is a human.

[0041] In practicing the invention, the hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof can be used as an active pharmacutical ingredient (API) of a pharmaceutical composition, which can be formulated in a solid, a semi-solid, or a liquid form.

[0042] According to another aspect, the invention relates to an aqueous pharmaceutical composition comprising a hexapeptide of formula (I):

H-T yr-D-Ala-Gly-Phe-Leu-Arg-OH (I) or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable excipient for use in the treatment of an interleukin-6 related respiratory disease by pulmonary administration to a mammal in need thereof.

[0043] In practicing the invention, the aqueous pharmaceutical composition contains from 0.1 to 100 mg/ml of the hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof.

[0044] In a preferred embodiment, the aqueous pharmaceutical composition contains 0.9 to 1.1 mg/ml of hexapeptide (I) diacetate.

[0045] As used herein, the term "excipient" describes any ingredient other than hexapeptide (I) or a pharmaceutically acceptable salt thereof. In some embodiments, the compositions may contain an excipient in an amount from 0.001 to 99.999%. [0046] Non-exclusive examples of excipients for use in the aqueous pharmaceutical composition of the invention include purified water, water for injection, buffer systems to maintain a pH of 3.0-8.5, regulators of tonicity of solutions, antimicrobial preservatives, surfactants such as no-ionic and amphiphilic surfactants, stabilizing agents, and emulsifiers.

[0047] As used herein, the term “aqueous pharmaceutical composition” means that a pharmaceutical water, e.g. purified water or water for injections, is the essential excipient of the composition of the invention.

[0048] As used herein, the term “pharmaceutical water” refers to water that meet requirements of regulatory documents of the Russian Federation and/or other countries, e.g. the requirements of FS.2.2.0020.15 “purified water” and the requirements of FS.2.2.0019.15 “water for injection” of the pharmacopoeia of the Russian Federation; and/or CPMP/QWP/158/01 «Note for Guidance on Quality of Water for Pharmaceutical Use», EMEA, 2002. Preferably, the pharmaceutical water is selected from the group consisting of purified water and water for injection.

[0049] As used herein, the term “buffer system” describes a combination of an acid and a base in amounts sufficient to maintain the desired pH level. Nonexclusive examples of pharmaceutically acceptable buffer systems for maintaining a pH in the range from 3.0 to 8.5 are described in the Pharmacopoeia of the Russian Federation (OFS.1.3.0003.15 “Buffer Solutions”) and include acetate buffer, citrate buffer, succinate buffer, phosphate buffer, and imidazole buffer.

[0050] Non-exclusive examples of regulators of tonicity include suitable osmotically active inorganic agent such as chlorides, sulfates or phosphates of sodium, calcium or magnesium; suitable osmotically active organic agent such as sugars and sugar alcohols, in particular trehalose, mannitol, and sorbitol. A preferred tonicity regulator is sodium chloride in an amount of from 0.1 to 0.9% of said composition.

[0051] In practicing the invention, the aqueous pharmaceutical composition is administered pulmonary in form of vapors or dispersions of liquid particles in a gaseous medium, thereby delivering from 0.1 to 50 mg of hexapeptide (I) or a pharmaceutically acceptable salt thereof into the lung of a mammal. Preferably, the composition is administered pulmonary in an amount that delivers 10 mg of hexapeptide (I) diacetate into the lung of a mammal. [0052] In practicing the invention, the aqueous pharmaceutical composition is administered pulmonary using a nebulizer or a metered-dose inhaler (MDI).

[0053] As used herein, the term “nebulizer” refers to an inhalation device that converts a liquid drug for spraying into a dispersion in a gaseous medium to deliver the active pharmaceutical ingredient to the lungs. Non-exclusive examples of nebulizers include compressor, ultrasound, or other type of nebulizers.

[0054] In some embodiments, the aqueous pharmaceutical composition is under pressure in a package with a metering valve-spray system (aerosols and sprays) for subsequent spraying.

[0055] In a preferred embodiment, the aqueous pharmaceutical composition is pulmonary administered in form of aerosol with particle size from 0.1 to 10.0 microns, more preferably from 1 to 5 microns.

[0056] In practicing the invention, the aqueous pharmaceutical composition may be in a unit dosage form suitable for a single pulmonary administration of a precise dose. As used herein, the term "unit dosage form" refers to a physically discrete unit of the composition of the invention suitable for a mammal to be treated by pulmonary route of administration. The level of a particular effective dose for any particular mammal depends on many factors, including the kind of the mammal; disorder being treated and the severity of the disorder; the particular composition used; age, weight, general health, and gender of the subject; time of administration; duration of treatment; drugs and/or additional therapies combined with or in combination with the composition of the present invention, and similar factors well known in medical technology.

[0057] In some embodiments, the unit dosage form contains hexapeptide (I) or a pharmaceutically acceptable salt thereof in an amount of 1.0 to 10.0 mg.

[0058] In practicing the present invention, the composition of the invention can be prepared by procedures well-known from the art. Such procedures include, but are not limited to, mixing hexapeptide (I) or a pharmaceutically acceptable salt thereof with other ingredients of the composition in conventional manner. Guidance for the preparation of compositions of the invention can be found in "Remington: The science and practice of pharmacy" 20th ed. Mack Publishing, Easton PA, 2000 ISBN 0-912734-04-3 and " Encyclopaedia of Pharmaceutical Technology", edited by Swarbrick, J. & J. C. Boylan, Marcel Dekker, Inc., New York, 1988 ISBN 0-8247-2800-9 or a newer edition. [0059] According to another aspect, the invention relates to a method of treating an interleukin-6 related respiratory disease, which comprises a step of pulmonary administration of an effective amount of a hexapeptide of formula (I):

[0060] As used herein, the term “effective amount” refers to an amount of an active pharmaceutical ingredient that is sufficient to cause a reduction, reverse, alleviation, or inhibition of progress of a disease to which this term is applied, or one or more symptoms of this disease.

[0061] In a preferred embodiment, the effective amount of a hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof is from 0.001 to 1.000 mg/kg per kg body weight of the mammal.

[0062] In practicing the invention, the effective amount of a hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof can be administered pulmonary by continous inhalation for period of from 1 minute to 2 hours, preferably from 30 to 60 minutes.

[0063] In practicing the invention, the effective amount of a hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof can be administered pulmonary once- or twice-a-day for one day or longer.

[0064] Because of pulmonary route of administration of the hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof, this invention provides a painless and safer alternative for the hexapeptide injections during the course of the treatment of interleukin-6 related respiratory diseases.

[0065] Because of direct topical delivery of the hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof to the lung, this invention provides particularly advantageous method for achieving the therapeutically effective levels of the hexapeptide (I) in the lung, which levels cannot been achieved with the hexapeptide (I) injections.

[0066] Because of the enhanced delivery of the hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof to the lung by pulmonary administration, this invention provides particularly advantageous compositions and methods over the hexapeptide injections for preventing, diminishing, or eliminating symptoms of an interleukin-6 related respiratory disease in a mammal in need thereof.

[0067] The following examples are presented to demonstrate the invention. The examples are illustrative only and are not intended to limit the scope of the invention in any way.

Example 1

[0068] This example shows the aqueous pharmaceutical composition for pulmonary administration. [0069] Table 1.

[0070] The aqueous pharmaceutical composition is prepared as follows. A commercially available hexapeptide (I) diacetate, API grade (Berachim Ltd., Obninsk, Russia), is dissolved in purified water in amounts as indicated in table 1 and the solution is adjusted to pH 3.0-8.5 using phosphate buffer. The solution is filtered through a filter with a pore size of 0.22 microns to prepare a sterile, preservative-free solution. The solution is filled into bottles. The aqueous pharmaceutical composition is administered pulmonary in a recommended dose to treat a IL-6 related respiratory disease in a subject in need thereof by inhalation using a nebulizer.

Example 2 [0071] This example shows the preferred aqueous pharmaceutical composition for pulmonary administration.

[0072] Table 2.

[0073] The aqueous pharmaceutical composition is prepared as follows. A commercially available hexapeptide (I) diacetate, API grade (Berachim Ltd., Obninsk, Russia), is dissolved in purified water in amounts as indicated in table 2. The solution is filtered through a filter with a pore size of 0.22 microns to prepare a sterile, preservative-free solution with pH of about 4.8. The solution is filled into ampoules and sealed in a sterile atmosphere. The aqueous pharmaceutical composition is administered pulmonary in the dose of 10 mg per subject using nebulizer to treat a IL-6 related respiratory disease.

Example 3

[0074] This example shows the aqueous pharmaceutical composition for pulmonary administration in a unit dosage form.

[0075] Table 3.

[0076] The aqueous pharmaceutical composition is prepared as follows. A commercially available hexapeptide (I) diacetate, API grade (Berachim Ltd., Obninsk, Russia), is dissolved in water for injections in an amount as indicated in table 3 to obtain a solution of the hexapeptide (I) diacetate at concentration of about 1 mg/ml, yield 100%. The solution is filtered through a filter with a pore size of 0.22 microns (Merck Millipore) to prepare a sterile, preservative-free solution, yield 99.9%. The solution is filled into a sterile 10 ml bottles in a stream of nitrogen or any other oxygen-free inert gas and sealed with a sterile butyl-rubber stopper and aluminum crimp. The obtained aqueous pharmaceutical composition in a unit dosage form is administered pulmonary to a subject suffering from a IL-6 related respiratory disease, e.g. COVID-19.

Example 4

[0077] This example illustrates a model for the assessment of efficacy of the hexapeptide (I) or a pharmaceutically acceptable salt thereof in the treatment of interleukin-6 related respiratory diseases.

[0078] An efficacy of the hexapeptide (I) diacetate was tested in a model of an acute lung injury (ALI) acompanied with an acute respiratory distress-syndrome (ARDS), which ALI/ARDS model is induced by intratracheal administration of lipopolysaccharide (LPS) as described in D'Alessio FR. Mouse Models of Acute Lung Injury and ARDS. Methods Mol Biol. 2018, 1809:341-350 with some modifications aimed to make the model more fatal. For this purpose, C57BI/6 male mice were pre-treated with 1 pg/mouse of a-Galactosyl Ceramide (a-GalCer, KRN7000, Sigma- Aldrich), as described in Aoyagi T et al. Int Immunol. 2011, 23(2):97-108, and after 24 hours mice received intratracheal LPS (E.coli; 300 pg/mouse) with addition of 10 pL/mouse Freund's complete adjuvant and 100 pg/mouse muramyl peptide. This model is characterized by dramatic overproduction of interleukin-6 in the lungs, which IL-6 overproduction is associated with acute lung injury, pulmonary edema, acute respiratory distress-syndrome, pneumonia, and a high lethality. Other symptoms of the modelled IL-6 related respratory disease were an inhibition of motor activity, impairment of coordination, insuffitiently wide opening of the eyelids and ptosis, low muscle tonus, decrease in appetite, water intake, and urination; and disturbance of the respiration. FIG.1 shows hematoxylin-eosin stained histological preparations of healthy lung tissue before a-GalCer/LPS administration (0 h) and lung preparations obtained from animals after 24, 72, and 120 h of LPS administration. The lung preparations from healthy animals are characterized by moderate blood supply, the absence of pathological changes in the stroma and parenchyma, as well as in the walls of the bronchi and bronchioles. In contrast, the lung preparations from a-GalCer/LPS-induced animals are characterized by of bronchopneumonia with areas of destruction of the walls of the bronchus and bronchioles, desquamated epithelium, and accumulation of inflammatory infiltrate, i.e. by signs of acute lung injury caused by IL-6 related inflammation.

[0079] To evaluate the dynamics of IL-6 production in the lungs, the ALI/ARDS were induced in C57BI/6 male mice by the sequential a-GalCer and LPS administration as described above. The levels of IL-6 were determined in the aqueous extract of the lung homogenates obtained from animals at different times (n = 6 per every time point) after the LPS injection (“LPS” group). For reference, levels of IL-6 in the aqueous extracts of the lung homogenates were determined in six healthy naive mice (“Intact” group). Measurements were performed on a Bio-Plex® MAGPIX™ Multiplex Reader (Bio-Rad, SN: 12250707) using a commercially available standard panel (Bio-Plex Pro™ Mouse Cytokine Th17 Panel A 6-Plex # M6000007NY) in accordance with the manufacturer’s protocol. Analysis of the obtained data was carried out in the Bio-Plex Data Pro program (version 1.0.0.06) of the Bio-Rad company. Results are presented in Table 4 as mean ± SEM of IL-6 concentrations in the lung homogenates. For reference, levels of IL-6 in the aqueous extracts of the lung homogenates of intact mice (“Intact”) was 119 ± 9 pg/ml.

[0080] Table 4.

Differs significantly from Intact (p<0.05).

[0081] Table 4 shows that a-GalCer/LPS-induced mice are characterized by on average 15.7- fold increase in the production of interleukin-6 in the lungs compared to healthy intact animals. Therefore, this model of ALI/ARDS is useful for evaluation of efficacy of an agent in the treatment of respiratory diseases associated with IL-6 overproduction.

[0082] In overall, the ALI/ARDS model reproduces the essential signs of interleukin-6 related respiratory diseases in humans such as overproduction of IL-6 in lungs, alveolar diffuse injury of the lungs, acute respiratory distress syndrome, pulmonary edema, and pneumonia.

Example 5

[0083] This example demonstrates that pulmonary administration provides a better delivery of hexapeptide (I) diacetate to the lungs as compared to hexapeptide (I) diacetate injections.

[0084] C57BI/6 male mice were randomized into two groups of eighteen animals each. In the first group (“HP(I) i.m.”), mice received 10 mg/kg hexapeptide (I) diacetate in 50 pi of water solution by intramuscular injection. In the second group (ΉR(I) inh.”), mice received 10 mg/kg hexapeptide (I) diacetate in 50 mI of water solution by inhalation. Lungs were extracted before (0 min) and after (1, 5, 15, 60 min) the hexapeptide (I) diacetate administration, from three animals at every time point in both groups. Concentrations of hexapeptide (I) in lung homogenates at every time point (0, 1, 5, 15, 60 min) were measured using Aglient HPLC 1260 with Agilent 6545XT Accurate mass Q-TOF LC/MS detector. Areas under pharmacokinetic curves (AUCs) for absorption of hexapeptide (I) diacetate by lungs for 60 min were 6446 ng/ml and 2991 ng/ml for pulmonary route of administration and injections, respectively. Thus, the pulmonary administration provides at least 2 fold higher delivery of hexapeptide (I) diacetate to the lung compared to injections of the same dose.

Example 6

[0085] This example demonstrates that inhaled hexapeptide (I) diacetate inhibits overexpression of IL-6 in the lungs. .

[0086] C57BI/6 male were randomized into four groups of six animals each. The first group was intact (“Intact”). The ALI/ARDS were induced by the sequential a-GalCer and LPS administration in groups 2 through 4 as described in the example 4 of the invention. In the second group (“LPS”), mice received 50 mI of saline by inhalation one hour after the LPS administration. In the third group (“LPS+HP(I) i.m.”), mice received 100 pg/kg hexapeptide (I) diacetate in 50 mI of water solution by intramuscular injection one hour after the LPS administration. In the fourth group (“LPS+HP(I) inh.”), mice received 100 pg/kg hexapeptide (I) diacetate in 50 mI of water solution by inhalation one hour after the LPS administration. After 3 hours, lungs were extracted from mice of all groups and levels of IL-6 mRNA were measured in the lung homogenates by quantitative real-time PCR. RNA was isolated using the "RNA Extran Kit" (Sintol, Russia), according to the manufacturer's protocol. cDNA was synthesized using Revepta-L reagent supermix (AmpliSens, Russia). Real-time PCR was performed using CFX96 Touch Real-Time PCR Detection System (BioRad, CA) and specific primers IL-6-F-5'- AT GAAGTT CCT CT CT GCAAG-3' and I L-6-R-5'-GT GT AATT AAGCCT CCGACT -3' . Data were normalized to GAPDH. Results are presented in FIG.2 as fold change for mRNA of IL-6 meantSEM in the lungs. One-way ANOVA followed by Tukey’s post-test demonstrates a statistically significant difference between groups. A significant 6.3-fold increase in the IL-6 expression was found in mice of “LPS” group compared to mice of “Intact” group (p<0.01). The inhaled hexapeptide (I) diacetate significantly inhibited the LPS-induced IL-6 overproduction in mice of “LPS+HP(I) inh.” group as compared to mice of “LPS” group (p<0.01), whereas the effect of hexapeptide (I) diacetate injection was non-significant (p=0.21). Thus, pulmonary administered hexapeptide (I) diacetate inhibits overexpression of IL-6 in the lungs associated with the respiratory disease.

Example 7

[0087] This example shows effect of inhaled hexapeptide (I) on IL-6 levels in lungs and serum.

[0088] C57BI/6 male mice were randomized into three groups of six animals each. The first group was intact (“Intact"). The ALI/ARDS were induced in groups 2 and 3 by the sequential a- GalCer and LPS administration as described in the example 4 of the invention. In the second group (“LPS”), mice received 50 pi of saline by inhalation one hour after the LPS administration. In the third group (“LPS+HP(I) inh.”), mice received 100 pg/kg hexapeptide (I) diacetate in 50 mI of water solution by inhalation one hour after the LPS administration. Levels of IL-6 were measured in the aqueous extracts of the lung homogenates and serum obtained from animals seven hours after saline or LPS injection using a commercially available standard panel (Bio- Plex Pro™ Mouse Cytokine Th17 Panel A 6-Plex # M6000007NY) as described in example 4 of the invention. Results are presented in table 5 as mean ± SEM of IL-6 concentrations (pg/ml) in the lung homogenates and serum.

[0089] Table 5.

^•Differs significantly from LPS (p<0.05).

[0090] Table 5 shows that LPS induced 7.8- (p<0.05) and 2.9-fold increase (p<0.05) in IL-6 levels in lungs and serum, respectively, compared to intact control. The inhalation of hexapeptide (I) diacetate into the lungs of LPS-induced mice resulted in reduction of IL-6 levels by 12.3 (p<0.05) and 6.5 times (p<0.05) in the lungs and serum, respectively, compared to LPS- induced control. Thus, the pulmonary administration of hexapeptide (I) diacetate significantly reduces IL-6 levels in lungs and serum of mammals suffering from the respiratory disease associated with interleukin-6 overproduction.

Example 8

[0091] This example illustrates the dose-response of inhaled hexapeptide (I) diacetate in the treatment of the interleukin-6 related respiratory disease.

[0092] C57BI/6 male mice were randomized into five groups of six animals each. The ALI/ARDS were induced in all groups by the sequential a-GalCer and LPS administration as described in the example 4 of the invention. In the first group, mice received by inhalation 50 mI of saline. In other groups, mice received hexapeptide (I) diacetate in doses of 1, 10, 100, and 1000 pg/kg as solutions in 50 mI of water. Levels of IL-6 were measured in the aqueous extracts of the lung homogenates obtained from animals six hours after LPS injection using a commercially available standard panel (Bio-Plex Pro™ Mouse Cytokine Th17 Panel A 6-Plex # M6000007NY) as described in example 4 of the invention. Results are presented in table 6 as mean ± SEM of IL-6 concentrations in the lung homogenates.

[0093] Table 6.

‘Differs significantly of saline (p<0.05).

[0094] Table 6 shows that pulmonary administration of hexapeptide (I) diacetate in doses of 1 to 1000 pg/kg (0.001 to 1.000 mg/kg) significantly reduces overproduction of IL-6 in the lung of animals suffering from the IL-6 related respiratory disease.

Example 9

[0095] This example illustrates a therapeutic efficacy of inhaled hexapeptide (I) diacetate over hexapeptide (I) diacetate injections.

[0096] C57BI/6 male mice were randomized into three groups of 15 animals each. ALI/ARDS has been induced in all mice by the sequential a-GalCer and LPS administration as described in the example 4 of the invention. In the first group (“LPS”), mice received intramuscularly 50 pi of saline daily, once-a-day, starting from the day of the LPS injection. In the second group (“LPS+HP(I) i.m.”), mice received intramuscularly 100 pg/kg hexapeptide (I) diacetate in 50 mI of water solution daily, once-a-day, starting from the day of the LPS injection. In the third group (“LPS+HP(I) inh.”), mice received by inhalation 100 pg/kg hexapeptide (I) diacetate in 50 mI of water solution daily, once-a-day, starting from the day of the LPS injection. After 72 hours, the LPS-induced ALI/ARDS resulted in the death of 87% (13/15), 27% (4/15), and 7% (1/15) of animals in the “LPS” group (control), the “LPS+HP(I) i.m.” group, and the “LPS+HP(I) inh.” group, respectively. FIG.3 shows the Kaplan-Meier survival curves for 144 hours of observations after the LPS administration. The Kaplan-Meier analysis shows that there was a significant difference between the groups (p <0.0001 ; Mantel-Cox log rank test). Median survival was 24 hours in the “LPS” group. Hexapeptide (I) diacetate significantly reduced the death of animals in “LPS+HP(I) i.m.” (p<0.0001) as well as in “LPS+HP(I) inh.” (p<0.0001) groups compared to “LPS” group. However, the therapeutic effect of inhaled hexapeptide (I) diacetate was higher than the effect of hexapeptide (I) diacetate injections. The hazard ratio for the “LPS+HP(I) inh.7“LPS+HP(l) i.m.” was 0.2295 (95% Cl 0.0397 to 1.327; logrank), i.e. the rate of deaths in group of mice treated with inhaled hexapeptide (I) was 4.4 times less than the rate of deaths in the group of mice treated with hexapeptide (I) injections. Thus, pulmonary administered hexapepide (I) diacetate is more effective than hexapeptide (I) diacetate injections in the reduction of mortality associated with IL-6 overproduction.

Example 10

[0097] This example illustrates that inhaled hexapeptide (I) diacetate are useful for use in the treatment of COVID-19.

[0098] Meta-analysis of clinical studies on COVID-19 revealed that the interleukin-6 level of 1.7 pg/ml in the serum is strong discriminator between severe (>1.7 pg/ml) and non-severe forms of COVID-19 (<1.7 pg/ml). Henry BM et al. Clin Chem Lab Med. 2020, 58(7):1021 -1028.

[0099] Inhalations of hexapeptide (I) diacetate was used in the treatment of COVID-19 patients during the course of clinical trials approved by the human research ethics committee. Each study protocol conforms to the ethical guidelines of the 1975 Declaration of Helsinki as reflected in a prior approval by institution’s human research committee. Written informed consent was obtained from each participant included in the study. Patients with confirmed diagnosis of COVID-19, who had blood IL-6 levels >1.7 pg/ml, were randomized to two groups. In the first group (control; n=75), patients received daily standard therapy recommended by the Ministry of Health of the Russian Federation (5^th through 7^th versions of the guidelines for the treatment of COVID-19). In the second group (HP(I); n=11), patients received 10 mg hexapeptide (I) diacetate by inhalation once-a-day daily. The levels of IL-6 in serum were assessed with commercially available ELISA assay before the start of the treatment (at day 0) and after treatment at day 5. At day 0, there was no statistically significant difference between groups (p=0.8495, unpaired two-tailed t-test) and the levels of IL-6 were 21.2±3.3 and 19.5±6.2 pg/ml (meanlSEM) in groups 1 and 2, respectively. The threshold value of 1.7 pg/ml for serum IL-6 was used as the criterion of discrimination between severe and non-severe forms of COVID-19 to assess an individual’s response to the treatment. A patient was considered a responder if serum IL-6 level at day 5 was less than 1.7 pg/ml, and a non-responder if serum IL-6 level at day 5 was higher than 1.7 pg/ml. The five-day treatment with inhaled hexapeptide (I) diacetate resulted in 2.8 fold decrease in IL-6 levels from 19.5±6.2 to 6.9±2.3 pg/ml. Levels of interleukin- 6 less than 1.7 pg/ml were achieved in 54.5% of patients treated with inhaled hexapeptide (I) diacetate for 5 days, and only in 29.3% of patients in control group. Therefore, the response rates were 54.5% and 29.3% of patients in the group treated with inhaled hexapeptide (I) diacetate and control group, respectively (FIG.4). Since the threshold level of 1.7 pg/ml discriminates between severe and non-severe forms of COVID-19, these results indicate that during the same treatment period the COVID-19 severity was weakened in 54.5% of patients treated with inhaled hexapeptide (I) diacetate compared to 29.3% of patients on standard therapy. Thus, pulmonary administered hexapeptide (I) diacetate is useful in the treatment of COVID-19.

Claims

Claims:

1. A hexapeptide of formula (I):

2. The hexapeptide as claimed in claim 1 , wherein the pharmaceutically acceptable salt is diacetate.

3. The hexapeptide as claimed in any claims 1 to 2, wherein the interleukin-6 related respiratory disease is selected from the group consisting of acute respiratory distress syndrome, acute lower respiratory infection, pneumonia, pulmonary edema, bronchitis, tracheobronchitis, chronic obstructive pulmonary disease, and asthma.

4. The hexapeptide as claimed in claim 3, wherein the acute lower respiratory infection is COVID-19.

5. An aqueous pharmaceutical composition comprising a hexapeptide of formula (I):

6. The composition as claimed in claim 5, wherein the pharmaceutically acceptable salt is diacetate.

7. The composition as claimed in any claims 5 to 6, wherein the interleukin-6 related respiratory disease is selected from the group consisting of acute respiratory distress syndrome, acute lower respiratory infection, pneumonia, pulmonary edema, bronchitis, tracheobronchitis, chronic obstructive pulmonary disease, and asthma.

8. The composition as claimed in claim 7, wherein the acute lower respiratory infection is COVID-19.

9. The aqueous pharmaceutical composition as claimed in any claims 5 to 8, wherein said composition is administered pulmonary in form of aerosol having particle size from 1 to 5 microns.

10. A method of treating an interleukin-6 related respiratory disease, which comprises a step of pulmonary administration of an effective amount of a hexapeptide of formula (I):

H-T yr-D-Ala-Gly-Phe-Leu-Arg-OH (I) or a pharmaceutically acceptable salt thereof to a mammal in need thereof.

11. The method as claimed in claim 10, wherein the pharmaceutically acceptable salt is diacetate.

12. The method as claimed in claim 10, wherein the interleukin-6 related respiratory disease is selected from the group consisting of acute respiratory distress syndrome, acute lower

' respiratory infection, pneumonia, pulmonary edema, bronchitis, tracheobronchitis, chronic obstructive pulmonary disease, and asthma.

13. The method as claimed in claim 12, wherein the acute lower respiratory infection is COVID-19.

14. The method as claimed in claim 10, wherein the hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof is administered in form of aqueous aerosol having particle size from 1 to 5 microns.

15. The method as claimed in claim 10, wherein the effective amount of a hexapeptide of formula (I) or a pharmaceutically acceptable salt thereof is from 0.001 to 1.000 mg/kg per kg body weight of the mammal.