WO2021204861A1

WO2021204861A1 - Compositions for use in the inhibition of coronavirus replication

Info

Publication number: WO2021204861A1
Application number: PCT/EP2021/059039
Authority: WO
Inventors: Torsten BJØRN
Original assignee: Nordic Pharma Lab Aps
Priority date: 2020-04-09
Filing date: 2021-04-07
Publication date: 2021-10-14

Abstract

The present invention provides compositions comprising fosfomycin and/or doxycycline for use to impair coronavirus replication and prevent and treat diseases caused by coronaviruses.

Description

COMPOSITIONS FOR USE IN THE INHIBITION OF CORONAVIRUS REPLICATION

Field of invention

The present invention provides compositions of medicinal agents that are currently available and approved by health authorities for human use to inhibit the replication of coronaviruses that can cause severe pneumonias and hence abort the infection at an early stage before pneumonia supervenes, and if pneumonia develops, can be used to treat it. It is relevant to the fields of infectious diseases, respiratory medicine and intensive care, as well as to general medical practice.

Background of invention

The current pandemic of COVID-19 disease (2019-2020) caused by the new coronavirus SARS-CoV-2 has added impetus and urgency to the discovery and testing of existing drugs, available at the present time and approved for human use by one or more national health authorities, to treat this disease. The focus is one the diseases caused by the viruses of the Betacoronavirus genus that cause the severe pneumonias of severe acute respiratory syndrome SARS (SARS-CoV), Middle East respiratory syndrome (EMC/2012) as well as the current COVID-19 (SARS-CoV-2). These viruses enter host cells through the binding of their spike (S) protein to the angiotensin converting enzyme 2 (ACE-2) expressed on the cell surface. The trans-membrane serine protease 2 (TMPRSS2) is required for processing the S protein for fusion with ACE2. In the host cells the viral RNA is replicated, the RNA encoding a long precursor viral protein sequence that is processed to form the viral proteins so that new complete virus particles can be formed. The emphasis in screening existing drugs for activity against the betacoronaviruses has been on those that may impair the entry of the virus into the host cells by interfering with the S protein-ACE2 interaction, and those that may impair the synthesis of viral proteins inside the host cell. Either mechanism will impair the replication of the virus in in-vitro cell cultures or in the human body.

Drugs that impair viral replication can be used prophylactically to prevent the development of disease after exposure to the virus, and can be used to abort mild disease or prevent it from becoming severe. They can also be used to lessen the progression of viral pneumonia and shorten its duration, and by preventing early death give more time for the host immune defense and repair processes to act against the infection and its consequences. The present invention concerns the use of two currently available antibiotics that are authorized for human use to impair betacoronavirus replication.

Summary of the invention Accordingly, the present invention makes use of two antibiotics, separately or preferably in combination, to impair the replication of coronaviruses capable of causing severe human disease. The antibiotics are fosfomycin and doxycycline. The invention thus comprises: 1. A composition comprising the antibiotic fosfomycin as an active ingredient for use in impairing the replication of a betacoronavirus and hence to prevent the development of disease after exposure to the virus, abort an infection or shorten its duration or prevent its progression to severe disease and to treat severe disease resulting from such infection.

2. A composition comprising the antibiotic doxycycline as an active ingredient for use in impairing the replication of a betacoronavirus and hence to prevent the development of disease after exposure to the virus, abort an infection or shorten its duration or prevent its progression to severe disease and to treat severe disease resulting from such infection.

3. A composition comprising a combination of the antibiotics fosfomycin and doxycycline as active ingredients for use in impairing the replication of a betacoronavirus and hence to prevent the development of disease after exposure to the virus, abort an infection or shorten its duration or prevent its progression to severe disease and to treat severe disease resulting from such infection.

4. A composition for use according to items 1 to 3 above, wherein the active ingredient or ingredients are taken orally. 5. A composition for use according to items 1 to 3 above, wherein the active ingredient or ingredients are given intravenously.

6. A composition for use according to items 1 to 3 above, wherein the active ingredient or ingredients are given by the inhalation of an aerosol.

In the following detailed description of the invention, details of the scope of the invention and its practical performance will be given. Detailed description of the invention

The purpose of the present invention is to provide compositions for use to impair betacoronavirus replication and thus prevent the development of disease after exposure to the virus, abort mild disease or shorten its duration or prevent it from becoming severe, and to treat severe disease such as viral pneumonia.

Active ingredients of the compositions of the invention

Compositions according to the present invention comprise the antibiotics fosfomycin and doxycycline, either alone or, preferably, in combination with each other.

Fosfomycin

Fosfomycin is the international non-proprietary name of a broad-spectrum antibiotic isolated and characterized in 1969 from Streptomyces fradiae strains under the name phosphomycin or phosphonomycin. Its structure was determined to be (-)(IR, 2S)-1,2- epoxypropylphosphonic acid with the systematic (lUPAC) name [(2R,3S)-3- methyloxiran-2-yl]phosphonic acid and a formula weight of 138.1 Da. It is not protein bound and has a mean elimination half-life of 5.7 hours. Fosfomycin is bactericidal and inhibits bacterial cell wall biosynthesis by inactivating the enzyme UDP-N- acetylglucosamine-3-enolpyruvyltransferase, also known as MurA The r. This enzyme catalyzes the committed step in peptidoglycan biosynthesis, the ligation of phosphoenolpyruvate to the 3'-hydroxyl group of UDP-N-acetylglucosamine to form N- acetylmuramic acid. Fosfomycin is a phosphoenolpyruvate analogue that inhibits MurA by alkylating an active site cysteine residue. The antibiotic enters the bacterial cell via the glycerophosphate transporter. The feature of fosfomycin that is relevant to the present invention is that it is a highly diffusible epoxide that can functionally inactivate an exposed cysteine residue. There is such a residue in S1 attachment region of the coronavirus S protein that binds to ACE2. This is the conserved cysteine residue in position 474, between other residues that are crucial for S protein attachment: -FNCY- (residues 472-475, denoted by the internationally accepted single-letter amino-acid code) of the SARS-CoV-2 S protein sequence.

In addition to this action, fosfomycin has been shown to have the capacity to favor phagocytosis and act as an immunomodulator. It is accumulated by polymorphonuclear leukocytes to reach concentrations that are up to twice those of the extracellular fluid, but does not affect their cellular functions, while exerting a bactericidal effect on bacteria such as Staphylococcus aureus. The chief adverse effects of fosfomycin are gastric irritation from orally administered fosfomycin disodium, evidence of allergy in the form of transient rashes (0.3% of cases) and eosinophilia (0.2%), as well as transiently raised liver enzymes (0.3% of cases) from systemically administered fosfomycin.

The principal forms (i.e. salts) of fosfomycin that come within the scope of this invention are: i) Fosfomycin disodium, formula weight 182.0 Da, pH of 5% solution 9.0-10.5. This salt is hygroscopic, highly soluble in water and shows a high bioavailability, but is locally irritant if un-neutralized. This is the preferred form for intravenous and inhalational administration, in pH neutralized solutions. ii) Fosfomycin calcium monohydrate, formula weight 194.1 Da, pH of 0.4% solution 8.1-9.6. This salt is sparingly soluble in water and not hygroscopic, but is less irritating to the stomach when used for oral treatment, when its bioavailability in terms of entering the systemic circulation may be as low as 12%. iii) Fosfomycin trometamol, formula weight 259.2 Da, pH of 5% solution 3.5- 5.5. This salt is highly soluble in water and is well tolerated when given orally, when it shows a bioavailability of about 40%. This is the preferred form for oral use. When the name “fosfo ycin” is used herein, it refers to an inorganic or organic salt of fosfomycin as exemplified by the principal forms above, and the dose of fosfomycin refers to the amount of the free acid form of fosfomycin present in the salt.

Doxycycline

Doxycycline is a second-generation tetracycline antibiotic, first described in 1957 and in general use since 1967, being included in the World Health Organization's List of Essential Medicines. Its chemical name is alpha-6-deoxy-5-oxytetracycline. It has a formula weight of 444.4, is highly (90%) protein bound and has an elimination half-life of 15-25 hours. It is a bacteriostatic antibiotic with a very broad antibacterial spectrum, acting by preventing the binding of transfer RNA to messenger RNA at the 30S ribosomal subunit of the bacteria. The features of doxycycline that are relevant to the present invention is that it also shows a range of activities that are also shown by one or more agents of a long list of other known agents that are known to impair coronavirus replication, including chloroquine, hydroxychloroquine, azithromycin, teicoplanin, celastrol, pristimerin, and ivermectin. Like these molecules, doxycycline is a large molecule with a fundamentally planar structure and a paucity of reactive side groups. Apart from its antibacterial activity, doxycycline shares many other activities shown to a different degree by other members of the list. These include antimalarial activity (chloroquine, hydroxychloroquine, ivermectin), activity against other parasites, including ectoparasites (ivermectin), and an anti-inflammatory effect in diseases such as rheumatoid arthritis and systemic lupus erythematosus (hydroxychloroquine). An action of doxycycline on coronavirus replication is attributable to an inhibition of the intracellular synthesis of viral protein.

In addition to these effects, doxycycline has a beneficial effect on lung function in chronic inflammatory lung diseases such as bronchial asthma and chronic obstructive pulmonary disease, blocking the action matrix metalloproteinases, including MMP-9, that contribute to the inflammatory response, reducing allergen-induced eosinophilic inflammation, and also reducing glandular hyperplasia, airway wall thickening, smooth muscle hyperplasia and subepithelial collagen deposition. These features imply an additional benefit from the use of doxycycline to treat viral pneumonia. The chief adverse effects of doxycycline are gastrointestinal, occasionally causing nausea, vomiting and/or diarrhea. It only rarely gives rise to Clostridium difficile colitis. Oral doxycycline can cause painful esophageal irritation if it is not adequately washed into the stomach. On prolonged use, it can cause photosensitivity manifested as an erythematous rash on exposed parts of the body. Doxycycline has little effect on hypoplasia of dental enamel or on staining of teeth and its use in children with severe diseases is permitted. Synergic action

Because fosfomycin and doxycycline act by different mechanisms on viral replication their effects show synergism.

Medical indications The indications for the use of the compositions described are as follows: i) Prophylactic use in subjects who through their occupation or daily life are subject to exposure to coronavirus capable of causing severe disease, especially during epidemics. ii) Prophylactic use in subjects who are known to have been exposed to coronavirus capable of causing severe disease. iii) Therapeutic use in subjects who are experiencing early symptoms of coronavirus infection, to abort the infection or prevent progression to serious disease. iv) Therapeutic use in subjects who are experiencing moderate symptoms of coronavirus infection, to shorten the duration of infection and prevent progression to serious disease. v) Therapeutic use is subjects with serious disease due to coronavirus, to shorten the duration of the disease and prevent further progression.

Formulations

In a preferred embodiment for oral use, the composition comprises fosfomycin trometamol supplied as a dry granulate to be dissolved in a supplied pharmaceutically acceptable carrier, preferably an aqueous carrier or diluent. The carrier may be simply water and the formulation may include pharmaceutically acceptable auxiliary substances or adjuvants, including, without limitation, buffering, sweetening and flavoring agents. Similarly, the doxycycline formulation comprises doxycycline as a dry doxycycline hydrochloride hemiethanolate hemihydrate powder, formulated in the same way for dissolution in water.

In a further embodiment for oral use, the composition comprises both fosfomycin trometamol and doxycycline hydrochloride hemiethanolate hemihydrate, both being supplied as a dry granulate or powder formulated as above to be dissolved in water. In a preferred embodiment for intravenous use, the fosfomycin is presented as a dry powder of fosfomycin disodium containing dry succinic acid to achieve a near neutral pH on reconstitution with sterile water for injection and subsequent dilution with a suitable diluent for intravenous infusion. The doxycycline is presented as a dry powder of doxycycline hydrochloride hemiethanolate hemihydrate for reconstitution with sterile water for injection and subsequent dilution with a suitable diluent to a volume between 100 ml_ and 1000 ml_ for intravenous infusion, said suitable diluents for fosfomycin and doxycycline comprising Sodium Chloride Injection (USP), or 5% Dextrose Injection (USP), or Ringer’s Injection (USP) or Invert Sugar, 10% in Water, or Lactated Ringer’s Injection (USP), or Dextrose 5% in Lactated Ringer’s, or a number of approved proprietary diluents for intravenous injection.

In a further embodiment for intravenous use, the fosfomycin disodium, succinic acid, and doxycycline hydrochloride hemiethanolate hemihydrate are supplied together as a mixed dry powder for reconstitution with sterile water for injection and subsequent dilution with one of the above approved diluents for intravenous infusion. Because of the relatively short elimination half-life of fosfomycin, it is necessary to adjust the doses of the fosfomycin and doxycycline so that each infusion containing one third of the total daily dose is given three times a day, at approximately 8-hour intervals. In one embodiment for intravenous use, a dried preparation of a composition according to the present invention may be pre-packaged, for example in single dose units, to be reconstituted in sterile water for injection and diluted with one of the above-mentioned diluents before intravenous infusion. In an embodiment for giving the active ingredients by direct inhalation into the lungs, the fosfomycin is presented as the solid powder for intravenous use, dissolved to a concentration of fosfomycin of 40 mg/ml_ in sterile 0.17% NaCI. The doxycycline is formulated with an excipient to make it suitable for aerosol administration, a non- limiting example of which is formulation as a complex with hydroxypropyl-gamma- cyclodextrin to a doxycycline concentration of 10-15 mg/ml_.

Dosage

The composition is formulated such that a single unit dose of fosfomycin in the present compositions for oral or intravenous use is in the range of 1 gram to 8 gram, given three times a day to achieve a total daily dose of 3 gram to 24 gram. The total daily dose of doxycycline is 100 mg to 200 mg. If given three times a day at about 8-hour intervals because of being mixed with fosfomycin, a formulation must be used that provides one-third of the total daily dose of doxycycline in a single unit dose. This applies whether the single unit dose of the mixed formulation of fosfomycin and doxycycline is for oral or intravenous administration.

The single unit dose of fosfomycin for inhalational use is in the range of 80 mg to 200 mg, administered two or three times a day, resulting in a total daily dose of 160 mg to 600 mg. The single unit dose of doxycycline for inhalational use is in the range of 20 mg to 60 mg, administered two or three times a day, resulting in a total daily dose of 40 mg to 180 mg.

Prophylactic administration will last for as many days as the person subject to viral exposure has need of it, subject to the judgement of the attending clinician.

Prophylactic administration to a person who is known to have been exposed to the virus will last for at least 14 days to cover the expected maximum incubation period, and again subject to the judgement of the attending clinician. Prophylactic administration will preferably be by the oral route. Therapeutic administration will last from the time of inception of therapy until the end of the illness, as determined by the attending clinician, and will preferably be by oral administration for as long as the patient is capable of taking it, and by intravenous administration if this should become necessary. If the clinician should so determine, therapeutic administration may also be carried out by the inhalation of an aerosol directly into the lungs. This method of administration may be especially suitable for patients who have been admitted to intensive care for pneumonia die to coronavirus, whether or not they are undergoing assisted ventilation.

It is not recommended to exceed a total daily dose of fosfomycin of 24 gram, or of doxycycline of 200 mg, whatever the routes or combined routes of administration may be.

Administration

Oral administration is by the subject drinking the solutions described in the section on formulations for oral use.

Intravenous administration is by slow intravenous administration of the diluted solutions described in the section on formulation for intravenous use. Administration by inhalation is by giving the solutions for inhalation by means of a nebulizer capable of producing aerosol particles of less 5 pm in diameter, allowing the aerosol particles to reach the lower airways.

Embodiments

1. A composition comprising the antibiotic fosfomycin as an active ingredient for use in impairing the replication of a betacoronavirus and hence to prevent the development of disease after exposure to the virus, abort an infection or shorten its duration or prevent its progression to severe disease and to treat severe disease resulting from such infection.

4. A composition for use according to embodiments 1 to 3 above, wherein the active ingredient or ingredients are taken orally.

5. A composition for use according to embodiments 1 to 3 above, wherein the active ingredient or ingredients are given intravenously. 6. A composition for use according to embodiments 1 to 3 above, wherein the active ingredient or ingredients are given by the inhalation of an aerosol.

7. A method for impairing the replication of a betacoronavirus in a subject, said method comprising administering to said subject a composition comprising fosfomycin as an active ingredient and hence prevent the development of disease after exposure to the virus, abort an infection or shorten its duration or prevent its progression to severe disease and to treat severe disease in said subject resulting from such infection.

8. A method for impairing the replication of a betacoronavirus in a subject, said method comprising administering to said subject a composition comprising doxycycline as an active ingredient and hence prevent the development of disease after exposure to the virus, abort an infection or shorten its duration or prevent its progression to severe disease and to treat severe disease in said subject resulting from such infection. 9. A method for impairing the replication of a betacoronavirus in a subject, said method comprising administering to said subject a composition comprising fosfomycin and doxycycline as active ingredients and hence prevent the development of disease after exposure to the virus, abort an infection or shorten its duration or prevent its progression to severe disease and to treat severe disease in said subject resulting from such infection.

10. A method for impairing the replication of a betacoronavirus in subject according to embodiments 7 to 9 above, wherein the active ingredient or ingredients administered to said subject are taken orally. 11. A method for impairing the replication of a betacoronavirus in subject according to embodiments 7 to 9 above, wherein the active ingredient or ingredients administered to said subject are given intravenously. 12. A method for impairing the replication of a betacoronavirus in subject according to embodiments 7 to 9 above, wherein the active ingredient or ingredients administered to said subject are given by the inhalation of an aerosol.

Claims

4. A composition for use according to any of the preceding claims, wherein the active ingredient or ingredients are taken orally.

5. A composition for use according to claims 1 to 3, wherein the active ingredient or ingredients are given intravenously.

6. A composition for use according to claims 1 to 3, wherein the active ingredient or ingredients are given by the inhalation of an aerosol.

7. A method of impairing the replication of a betacoronavirus in a subject, comprising administering the antibiotic fosfomycin to the subject to prevent the development of disease after exposure to the virus, abort an infection or shorten its duration or prevent its progression to severe disease and to treat severe disease resulting from such infection.

8. A method of impairing the replication of a betacoronavirus in a subject, comprising administering the antibiotic doxycycline to the subject to prevent the development of disease after exposure to the virus, abort an infection or shorten its duration or prevent its progression to severe disease and to treat severe disease resulting from such infection.

9. A method of impairing the replication of a betacoronavirus in a subject, comprising administering a combination of the antibiotics fosfomycin and doxycycline to the subject to prevent the development of disease after exposure to the virus, abort an infection or shorten its duration or prevent its progression to severe disease and to treat severe disease resulting from such infection.

10. A method of impairing the replication of a betacoronavirus in a subject according to claims 7-9, wherein the antibiotics fosfomycin and/or doxycycline are administered orally.

11. A method of impairing the replication of a betacoronavirus in a subject according to claims 7-9, wherein the antibiotics fosfomycin and/or doxycycline are administered intravenously.

12. A method of impairing the replication of a betacoronavirus in a subject according to claims 7-9, wherein the antibiotics fosfomycin and/or doxycycline are administered by the inhalation of an aerosol.