DESC:FIELD OF THE INVENTION
The present invention relates to a synergistic combination administered in a form of pharmaceutical composition comprising Chloroquine and/ or derivatives such as Hydroxychloroquine (HCQ) and synthetic minerals such as Zinc salts (e.g. zinc acetate, zinc oxide), a process for preparing such pharmaceutical composition, and use of the said pharmaceutical composition for the prevention, treatment and prophylaxis of diseases caused by viruses.

BACKGROUND AND PRIOR ART
Coronavirus disease 2019 (COVID-19) is an emerging infectious disease of potential zoonotic origin. Severe acute respiratory syndrome (SARS) corona virus – 2, (SARS-CoV-2), the etiological agent of COVID-19, can cause a serious or life-threatening disease or condition, including severe respiratory illness. The public health concerns caused by the outbreak of COVID-19 was declared as a “public health emergency of international concern” on 31 January, 2020 and “pandemic” on 11 March 2020 by World Health Organization. Impacts of the COVID-19 pandemic are currently ‘skyrocketing’ across the globe, causing enormous public health havoc and a colossal economic crisis.
The spread of COVID-19 is so accelerating that it needed sixty seven days from the first diagnosed case to reach the first 100,000 cases, eleven days for the second 100,000 cases, four days for the third 100,000 cases, three days for the fourth 100,000 cases and just extra seven days for additional 400,000 cases around the globe. A similar pattern has been seen in the fatality as it required seventeen days from the first death reported on 11 January 2020 to reach the first 100 deaths, thirty days for the 1,000 deaths, sixty one days for the 5,000 deaths and sixty eight days for 10,000 deaths and just nine additional days for additional 20,000 deaths. By 1 April 2020, the number of confirmed cases skyrocketed to approximately 823,000 and 40,598 deaths from 205 countries and territories around the globe. In the single day of 31 March, 2020, there were approximately 72,700 new confirmed cases and 4,193 deaths reported across the world. The reported mortality rate for COVID-19 is around 3% and the mortality rates vary depending on geographical regions, age distributions and co-morbidities. Recently, the Chinese Government included chloroquine phosphate in the list of antiviral therapies for the treatment of novel coronavirus pneumonia or COVID19 caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in its 6th and 7th versions of official Guidelines for the Prevention, Diagnosis, and Treatment of Pneumonia caused by COVID-19. Immediate research on the potential therapeutic application for the disease can save lives and prevent from the rapid viral transmission. Chloroquine phosphate is an old drug used for the treatment of malaria caused by Plasmodium species. Chloroquine phosphate can also be replaced with another member of its family, hydroxychloroquine. The active compound chloroquine constitutes 60% in chloroquine phosphate and 77.5% in hydroxychloroquine respectively.
Mechanism of actions of chloroquine or hydroxychloroquine has been studied for both SARS-CoV and SARS-CoV-2. Hydroxychloroquine inhibits terminal glycosylation of Angiotensin Converting Enzyme 2 (ACE2), the receptor that SARS-CoV and SARS-CoV-2 target for cell entry (Vincent MJ, Bergeron E, Benjannet S, Erickson BR, Rollin PE, Ksiazek TG, Seidah NG, Nichol ST: Chloroquine is a potent inhibitor of SARS coronavirus infection and spread. Virol J. 2005 Aug 22;2:69). ACE2 that is not in the glycosylated state may less efficiently interact with the SARS-CoV-2 spike protein, further inhibiting viral entry (Wang M, Cao R, Zhang L, Yang X, Liu J, Xu M, Shi Z, Hu Z, Zhong W, Xiao G: Remdesivir and chloroquine effectively inhibit the recently emerged novel coronavirus (2019-nCoV) in vitro. Cell Res. 2020 Mar;30(3):269-271).
The synthesis reference has been discussed in the U.S. Patent 2,546,658. A 200mg oral dose of hydroxychloroquine has a half-life of 537 hours or 22.4 days in blood, and 2963 hours or 123.5 days in plasma. A 155mg intravenous dose has a half-life of 40 days. This is the prescribed dose to be taken twice a day for 7 days as a prophylaxis for Malaria. In the journal, pre-proof paper by Fantini et. al. (“Structural and molecular modeling studies reveal a new mechanism of action of chloroquine and hydroxychloroquine against SARS-CoV-2 infection”, International Journal of Antimicrobial Agents), authors explain the mechanism of action of HCQ on SARS- COV 2. The first step of the viral replication cycle, i.e. the attachment to the surface of respiratory cells mediated by the spike (S) viral protein, offers several potential therapeutic targets. The S protein uses the ACE-2 receptor for entry, but also sialic acids linked to host cell surface gangliosides. Using a combination of structural and molecular modelling approaches, the authors showed that chloroquine (CLQ), one of the drugs currently under investigation for SARS-CoV- 2 treatment, binds sialic acids and gangliosides with high affinity. In vitro studies for SARS-CoV-2 also found chloroquine and hydroxychloroquine, a less-toxic derivative of chloroquine to be able to effectively inhibit viral growth. While the reported 50% effective concentration of the drugs are higher than the normal serum concentration reported in people taking these drugs, the concentrations in lungs and other vital organs are higher. The most common side-effect of these are retinopathy and other side-effects precipitating due to higher concentrations of the compounds in blood.
The recommended 3-day chloroquine phosphate dosage for malaria treatments for adults is 1 gm orally followed by 500 mg after 6-8 hours in the first day and 500 mg per day for second and the third day. Similarly, the 3-day dosage for hydroxychloroquine is 800 mg orally followed by 400 mg after 6-8 hours in the first day and 400 mg per day for second and third day. This study was specifically conducted to examine the various dosages of chloroquine and hydroxychloroquine utilized in previous clinical trials registered and underwent for the treatment of pneumonia caused by SARS-CoV-2. The guidelines by the Chinese government have postulated the dose of 500 mg twice a day for 7 days for adults aged 18-65 years and weighing more than 50 kg; and initial dose of 500 mg twice a day for first two days and 500 mg per day for days 3 through 7 for adults of weight less than 50 kg.
With the advent of the devastating pandemic, many clinical trials have been initiated, and wherein, most of trials used oral chloroquine (68.2%, 15/33) or hydroxychloroquine (68.2%, 15/33) singly as treatment options, followed by chloroquine in combination with other antivirals ((13.6%, 3/33), hydroxychloroquine with azithromycin (9.1%, 2/33), chloroquine as aerosols (9.1%, 2/33), chloroquine with azithromycin (4.5%, 1/33) and hydroxychloroquine with other antivirals (4.5%, 1/33). Among the trials using chloroquine or HCQ as prophylactic agent, 45400 healthy at-risk population are to be enrolled with the UK study by the University of Oxford, enrolling 40,000 individuals for a duration of three months with 250 mg chloroquine daily (NCT04303507) and the least dose is planned to be given in a study in Turkey where 200 mg hydroxychloroquine is planned to be given every three weeks along with daily vitamin C and zinc tablets) (NCT04326725).
Separately, combination of chloroquine with zinc ions have their own association in targeting killing of cancer cells (Jing Xue et al “Chloroquine is a Zinc ionophore” Department of Pathology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma, United States of America, Department of Radio-Oncology, Nanjing Medical University Affiliated Suzhou Hospital, Suzhou, China), however, based on the clinical trial being conducted involving administration of HCQ with zinc tablets indicates as one particular proposal towards combating the COVID-19 symptoms.
While several options are being tried and tested to target symptomatic association of the disease, it becomes important to take cognizance of certain patient pool which ultimately develop severe lung related complications as a prime site of infection in COVID-19. Targeting lung related treatments through inhalation is well known medical paradigm, and having reviewed the recent clinical trial which involve around chloroquine inhalation administration, there is no sufficient knowledge over the success of such inhalation route of administering drugs in COVID-19 either alone or in combination as a cocktail therapy.
Hence, there is an instant need to develop an appropriate inhalation composition involving Chloroquine and/ or derivatives such as Hydroxychloroquine (HCQ) in combination with other drugs and/ or synthetic minerals such as zinc acetate or zinc oxide, which is cost effective, which can be produced by simple manufacturing techniques, and which also exhibits better or improved patient compliance.

OBJECTIVES OF THE INVENTION
An object of the present invention is to provide a pharmaceutical composition administered by inhalation comprising Chloroquine and/ or derivatives such as Hydroxychloroquine (HCQ) in combination with other drugs.
Another object of the present invention is to provide a pharmaceutical composition administered by inhalation comprising Hydroxychloroquine (HCQ) and synthetic minerals such as zinc acetate or zinc oxide.
Yet another object of the present invention is to provide a pharmaceutical composition administered by inhalation comprising Hydroxychloroquine (HCQ) and synthetic minerals such as zinc acetate or zinc oxide which shows synergistic action.
Yet another object of the present invention is to provide a process of preparing a pharmaceutical composition administered by inhalation comprising Hydroxychloroquine (HCQ) and synthetic minerals such as zinc acetate or zinc oxide.
Still another object of the present invention is to provide a pharmaceutical composition administered by inhalation comprising Hydroxychloroquine (HCQ) and synthetic minerals such as zinc acetate or zinc oxide for the treatment of infection caused by Coronaviridae virus (including COVID-19).
Still another object is to provide a method of alleviating or treating infection caused by Coronaviridae virus (including COVID-19) by administering inhalable pharmaceutical composition comprising Hydroxychloroquine (HCQ) and synthetic minerals such as zinc acetate or zinc oxide.
Yet another object is to provide the use of inhalable pharmaceutical composition comprising Hydroxychloroquine (HCQ) and synthetic minerals such as zinc acetate or zinc oxide in the treatment of infection caused by Coronaviridae virus (including COVID-19).

SUMMARY OF THE INVENTION
According to one aspect of the present invention there is provided a pharmaceutical composition administered by inhalation comprising Chloroquine and/ or derivatives such as Hydroxychloroquine (HCQ) in combination with other drugs.
According to another aspect of the present invention there is provided a pharmaceutical composition administered by inhalation comprising Hydroxychloroquine (HCQ) and synthetic minerals such as zinc acetate or zinc oxide.
According to another aspect of the present invention there is provided a process of preparing a pharmaceutical composition administered by inhalation comprising Hydroxychloroquine (HCQ) and synthetic minerals such as zinc acetate or zinc oxide.
According to yet another aspect of the present invention there is provided a pharmaceutical composition administered by inhalation comprising Hydroxychloroquine (HCQ) and synthetic minerals such as zinc acetate or zinc oxide for the treatment of infection caused by Coronaviridae virus (including COVID-19).
According to still another aspect of the present invention there is provided a method of alleviating or treating infection caused by Coronaviridae virus (including COVID-19) by administering inhalable pharmaceutical composition comprising Hydroxychloroquine (HCQ) and synthetic minerals such as zinc acetate or zinc oxide.
According to yet another aspect of the present invention there is provided a use of inhalable pharmaceutical composition comprising Hydroxychloroquine (HCQ) and synthetic minerals such as zinc acetate or zinc oxide in the treatment of infection caused by Coronaviridae virus (including COVID-19).

BRIEF DESCRIPTION OF DRAWINGS
Figure 1 presents plasma concentration and lung concentration for Oral tablet-200mg of HCQ for inhalation dose estimation for HCQ.
Figure 2 presents plasma concentration and lung concentration for Inhalation route - 80 mg for HCQ.
Figure 3 presents plasma concentration and lung concentration for Inhalation route - 100 mg for HCQ.
Figure 4 presents plasma concentration and lung concentration for Inhalation route – 160 mg for HCQ.
Figure 5 presents plasma concentration and lung concentration for Oral tablet 30 mg of Zinc for inhalation dose estimation for Zinc.
Figure 6 presents plasma concentration and lung tissue concentration for Inhalation route - 8 mg for Zinc (Cell Permeability 1.1*10^-7 cm/s).
Figure 7 presents lung tissue concentration with oral dose regimen of HCQ.
Figure 8 presents lung tissue concentration with proposed dose of HCQ via inhalation route i.e. administration through nebulization.
Figure 9 presents lung tissue concentration with proposed dose of HCQ via inhalation route i.e. administration through dry powder inhaler (DPI).
Figure 10 presents lung tissue concentration with oral dose regime of Zinc (equivalent units of Zinc from zinc acetate).
Figure 11 presents lung tissue concentration with proposed dose of Zinc Acetate via inhalation route of nebulization with 8 mg dose.
Figure 12 presents lung tissue concentration with proposed dose of Zinc Acetate via inhalation route of nebulization with 4 mg dose.
Figure 13 presents lung tissue concentration with proposed dose of Zinc Acetate via inhalation route of DPI formulation.
Figure 14 presents efficacy comparison for HCQ oral vs nebulized inhalation route against the viral load for 8 days of dosing regime.
Figure 15 presents efficacy comparison for Zinc oral vs nebulized inhalation route against the viral load for 8 days of dosing regime.
Figure 16 presents synergistic effect of HCQ on Zinc lung dose increase in presence of HCQ.
Figure 17 presents synergistic effect of HCQ + Zn in treatment of COVID 19 infection.

DETAILED DESCRIPTION OF THE INVENTION
Hydroxychloroquine is an analogue of chloroquine. In the previous SARS outbreak, hydroxychloroquine was reported to have anti-SARS-CoV activity in vitro. HCQ acts as RNA polymerase inhibitor as well as ACE 2 receptor binder – thus limiting cell binding as well as replication engine of the virus. HCQ also acts as ionophore which will introduce Zn into the cell and augment the action of stopping of virus replication. This suggests that hydroxychloroquine may be a potential pharmacological agent for the treatment of COVID-19 infection
Zinc is an essential trace element that is crucial for growth, development, and the maintenance of immune function. Increasing the intracellular Zn2+ concentration with zinc-ionophores (like hydroxychloroquine) can efficiently impair the replication of a variety of RNA viruses, including poliovirus and influenza. Literature (Xue J, Moyer A, Peng B, Wu J, Hannafon BN, Ding WQ. Chloroquine is a zinc ionophore. PLoS One. 2014;9(10)) shows that ionophoric action of HCQ brings about 1000 fold increase in intracellular zinc concertation (few micromoles to millimoles) It has been demonstrated in some studies (te Velthuis AJ, van den Worm SH, Sims AC, Baric RS, Snijder EJ, van Hemert MJ. Zn(2+) inhibits coronavirus and arterivirus RNA polymerase activity in vitro and zinc ionophores block the replication of these viruses in cell culture. PLoS Pathog. 2010;6(11)) that the combination of Zn2+ and Zinc ionophore inhibits the replication of SARS-coronavirus (SARS-CoV) and in cell culture. Further in the recent pre-print paper before peer-review by Derwand et. al. (“Does zinc supplementation enhance the clinical efficacy of chloroquine/hydroxychloroquine to win todays battle against COVID-19?”, 8th April 2020 ) authors hypothesize that the combination of CQ/HCQ with parenteral zinc in the treatment of hospitalized COVID-19 patients may help to improve clinical outcomes and to limit the COVID-19 fatality rates
The inventors of the present invention have developed pharmaceutical inhalation composition comprising hydroxychloroquine in combination with zinc salts which not only acts as a targeted solution showcasing synergistic action for infection caused by Coronaviridae virus (including COVID 19) but also improves patient compliance.
Chloroquine is a well known lysosomotropic agent, currently attracting many hopes in terms of antiviral therapy as well as in antitumoral effect because of its pH-dependent inhibiting action on the degradation of cargo delivered to the lysosome, thus effectively disabling this final step of the autophagy pathway.
Hydroxychloroquine (HCQ) is a chemical derivative of chloroquine (CQ) which features a hydroxyethyl group instead of an ethyl group.
According to an aspect of some embodiments of the present invention there are provided pharmaceutical inhalable compositions in unit dosage form. The compositions according to this aspect of the present invention comprise an amount of HCQ or a pharmaceutically acceptable salt thereof, an amount of zinc salt, and a pharmaceutically acceptable carrier.
As used in this specification, whether in a transitional phrase or in the body of the claim, the terms “comprise(s)” and “comprising” are to be interpreted as having an open-ended meaning. That is, the terms are to be interpreted synonymously with the phrases “having at least” or “including at least”. When used in the context of a process, the term “comprising” means that the process includes at least the recited steps, but may include additional steps. When used in the context of a compound or composition, the term “comprising” means that the compound or composition includes at least the recited features or components, but may also include additional features or components.
As used in this specification, the singular forms “a,” “an” and “the” specifically also encompass the plural forms of the terms to which they refer, unless the content clearly dictates otherwise.
The term “about” is used herein to mean approximately, in the region of, roughly, or around. When the term “about” is used in conjunction with a numerical range, it modifies that range by extending the boundaries above and below the numerical values set forth. In general, the term “about” is used herein to modify a numerical value above and below the stated value by a variance of 20%.
It will be well appreciated that the term “hydroxychloroquine or HCQ” as used herein is denoted in broad sense to include not only “hydroxychloroquine or HCQ” per se but also its pharmaceutically acceptable derivatives thereof. Suitable pharmaceutically acceptable derivatives include pharmaceutically acceptable salts, pharmaceutically acceptable solvates, pharmaceutically acceptable hydrates, pharmaceutically acceptable anhydrates, pharmaceutically acceptable enantiomers, pharmaceutically acceptable esters, pharmaceutically acceptable isomers, pharmaceutically acceptable polymorphs, pharmaceutically acceptable prodrugs, pharmaceutically acceptable tautomers, pharmaceutically acceptable complexes etc.
As used herein, the term "hydroxychloroquine or HCQ" refers to the racemic hydroxychloroquine, which is 2-[[4-[(7~chloro~4~quinolinyl)amino]pentyl]ethylamino]ethanol as disclosed in U.S. Patent No. 2,546,658, which is incorporated herein by reference, or any of the single enantiomers "(S)-(+) hydroxychloroquine" or "( )-(-) hydroxychloroquine" as disclosed in U.S. Patent No. 5,31 4,894, also incorporated herein by reference. This term may relate either to the free form of hydroxychloroquine or to any pharmaceutically acceptable salt thereof, such as hydroxychloroquine sulfate.
The term “zinc” refers either to envisage an ionic form of zinc (ZN+2) or salt form of zinc which includes but is not limited to zinc acetate, zinc oxide, zinc chloride
The term "combination" as used herein, refers either a fixed combination in one dosage unit form, a non- fixed combination or a kit of parts for the combined administration.
The term "treating" or "treatment" as used herein refers to relieving, reducing or alleviating at least one symptom in a subject or effecting a delay of progression of a disease. For example, treatment can be the diminishment of one or several symptoms of a disorder or complete eradication of a disorder, such as cancer. Within the meaning of the present invention, the term "treat" also denotes to arrest, delay the onset (i.e., the period prior to clinical manifestation of a disease) and/or reduce the risk of developing or worsening a disease.
The term "pharmaceutically acceptable salt" refers to a charged species of the parent compound and its counter ion, which is typically used to modify the solubility characteristics of the parent compound and/or to reduce any significant irritation to an organism by the parent compound, while not abrogating the biological activity and properties of the administered compound. Examples, without limitation, of pharmaceutically acceptable salts include salts comprising an anion such as a carboxylate or sulfate anion, and/or a cation such as, but not limited to, ammonium, sodium, potassium, and the like. Suitable salts are described in, e.g., Birge et ai. [J Pharma Sci 1977, 66: 1-19], An example of pharmaceutically acceptable salt of hydroxychloroquine is hydroxychloroquine sulfate.
The term "pharmaceutical composition" refers to a preparation of one or more of the active agents described herein (e.g., HCQ and zinc) with other chemical components such as pharmaceutically acceptable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate administration of a compound to an organism.
The term "pharmaceutically acceptable carrier" refers to a carrier, adjuvant, or a diluent that does not cause significant irritation to an organism and does not abrogate the biological activity and properties of the administered compound.
The term “excipient" refers to an inert substance added to a pharmaceutical composition to further facilitate administration of an active ingredient. Examples, without limitation, of excipients include surfactants, isotonicity agents, pH adjusters, buffers, various sugars and types of starch, cellulose derivatives, gelatin, vegetable oils, and polyethylene glycols.
According to the present invention, there is provided a pharmaceutical composition administered by inhalation comprising Hydroxychloroquine (HCQ) and synthetic minerals such as zinc acetate or zinc oxide along with one or more carriers.
The present invention provides an inhalable pharmaceutical composition comprising:
(A) chloroquine or its pharmaceutically acceptable salt thereof or a derivative of chloroquine or its salt;
(B) zinc or its pharmaceutically acceptable salts and derivatives; and
(C) pharmaceutically acceptable excipients.
In an embodiment of the present invention, the chloroquine or its pharmaceutically acceptable salt, derivative thereof is hydroxychloroquine or its pharmaceutically acceptable salts.
In an embodiment of the present invention, the zinc or its derivatives is selected from zinc acetate, zinc oxide, zinc chloride.
In an embodiment of the present invention, the composition is an atomizable composition.
In an embodiment of the present invention, the composition is a nebulizable composition comprising a dispersion of hydroxychloroquine or its pharmaceutically acceptable salt thereof; and zinc or its pharmaceutically acceptable salts and derivatives in an aqueous, organic or aqueous/organic medium.
In an embodiment of the present invention, the particle size of hydroxychloroquine or its pharmaceutically acceptable salt, and zinc acetate or its pharmaceutically acceptable salt is from about 0.1 micron to about 5 micron.
In an embodiment of the present invention, the amount of hydroxychloroquine or its pharmaceutically acceptable salt is from about 0.005 mg/ml to about 50 mg/ml of total formulation.
In an embodiment of the present invention, the amount of zinc or its salts is from about from about 0.001 mg/ml to 5 mg/ ml of total formulation.
In an embodiment of the present invention, volume of the nebulization composition is from about 0.2 ml to about 6 ml.
In an embodiment of the present invention, the pharmaceutically acceptable excipients are surfactants, isotonicity agents, pH adjusters, buffers, various sugars and types of starch, cellulose derivatives, gelatin, vegetable oils, and polyethylene glycols.
In an embodiment of the present invention, the composition is a dry powder comprising finely divided hydroxychloroquine or its pharmaceutically acceptable salt thereof; and zinc or its pharmaceutically acceptable salts and derivatives; optionally together with a pharmaceutically acceptable carrier in finely divided form.
In an embodiment of the present invention, the composition is an aerosol comprising hydroxychloroquine or its pharmaceutically acceptable salt; and zinc or its pharmaceutically acceptable salts and derivatives in solution or dispersion in a propellant.
In an embodiment of the present invention, the composition comprising chloroquine or its pharmaceutically acceptable salt thereof or a derivative of chloroquine or its salt; zinc or its pharmaceutically acceptable salts and derivatives; and pharmaceutically acceptable excipients is for use in treatment of an infection caused by COVID-19.
Suitably, the pharmaceutical composition, according to the present invention are presented either in the form of inhalation solutions or suspensions to be administered with the help of a suitable nebulizer or in the form of dry powder inhaler to be administered either through capsule based inhalers or through blister and/ or cartridge comprising multiple pockets of storing the dry powder or reservoir based multidose inhalers. Alternatively, propellant driven aerosol compositions are also envisaged within the scope of the present invention which are administered either through conventional press-and-breathe inhalers or through breath actuated inhalers (both associated with dose counters/ indicators) well known in the art.
According to a preferred embodiment, the pharmaceutical inhalable composition is presented in the form of liquid solution/ suspension comprising a therapeutically effective amount of Hydroxychloroquine sulfate and Zinc acetate dihydrate along with one or more excipients for administration through a suitable nebulizer (also known as “nebulization composition”). The liquid solution/ suspension may be packed in an ampoule (to be presented as single dose) or vial (to be presented as multidose).
The nebulization composition of the present invention is provided in sterile unit dose treatments. The nebulization composition comprises an aqueous suspension comprising therapeutically effective amount of Hydroxychloroquine sulfate and Zinc acetate dihydrate and wherein the composition comprises one or more pharmaceutically acceptable excipients.
The nebulization composition includes from about 0.005 mg/ml to about 50 mg/ml of Hydroxychloroquine sulfate equivalent to Hydroxychloroquine and from about 0.001 mg/ml to 5 mg/ ml of Zinc acetate dihydrate equivalent to Zinc. Preferably, the nebulization composition comprises about 33 mg/ml of Hydroxychloroquine sulfate equivalent to Hydroxychloroquine of 26.7 mg/ml and 2.66 mg/ml of Zinc acetate dihydrate equivalent to Zinc. The volume of the nebulization composition is about 0.2 ml to about 6 ml.
The nebulization composition of the present invention contains Hydroxychloroquine sulfate and Zinc acetate dihydrate in micronized form. Suitable micronization techniques like Microfluidizer, High pressure homogenizer, Ball mill, Sonication and other such techniques commonly known in the art can be employed to effectively size reduce the mometasone or its pharmaceutically acceptable salt thereof. The particle size of Hydroxychloroquine sulfate and Zinc acetate dihydrate desired for effective nebulization of the compositions of the present invention can range from about 0.1 micron to about 5 micron.
The nebulization composition of the present invention have a pH of about 2 to about 8. The pH may be adjusted by the addition of one or more pharmaceutically acceptable acids. Examples of suitable pharmaceutically acceptable acids include inorganic acids, such as hydrochloric acid, hydrobromic acid, nitric acid, sulfuric acid, and phosphoric acid, and combinations thereof. Examples of other suitable pharmacologically acceptable acids include organic acids, such as ascorbic acid, citric acid, malic acid, maleic acid, tartaric acid, succinic acid, fumaric acid, acetic acid, formic acid, and/or propionic acid. In one embodiment, the pH is adjusted with 1N hydrochloric acid or 1N sulfuric acid. In another embodiment, the pH is adjusted with one or more organic acids selected from ascorbic acid, fumaric acid and citric acid. A preferred organic acid is citric acid. If desired, mixtures of the abovementioned acids may also be used, particularly in the case of acids which have other properties in addition to their acidifying properties, e.g., those which act as flavorings or antioxidants, such as for example citric acid or ascorbic acid. The nebulization composition may contain 0.0001% to 0.53% w/v of citric acid, or 0.0001% to 2.20% w/v of sodium citrate, or 0.00937% to 9.5% w/v of monosodium phosphate dehydrate, or 0.0017% to 1.7% w/v of Dibasic sodium phosphate anhydrous.
The nebulization composition of the present invention may optionally include a buffer. General and biological buffers in the pH range of about 2.0 to about 8.0 include, but are not limited to, acetate, borate, citrate, collidine, formate, maleate, Mcllvaine, phosphate, Prideaux-Ward, succinate, citrate -phosphate -borate buffers.
Suitable tonicity adjusting agents include, but are not limited to, ammonium carbonate, ammonium chloride, ammonium lactate, ammonium nitrate, ammonium phosphate, ammonium sulfate, ascorbic acid, bismuth sodium tartrate, boric acid, calcium chloride, calcium disodium edetate, calcium gluconate, calcium lactate, citric acid, dextrose, diethanolamine, dimethyl sulfoxide, edetate disodium, edetate trisodium monohydrate, fluorescein sodium, fructose, galactose, glycerin, lactic acid, lactose, magnesium chloride, magnesium sulfate, mannitol, polyethylene glycol, potassium acetate, potassium chlorate, potassium chloride, potassium iodide, potassium nitrate, potassium phosphate, potassium sulfate, propylene glycol, silver nitrate, sodium acetate, sodium bicarbonate, sodium biphosphate, sodium bisulfite, sodium borate, sodium bromide, sodium cacodylate, sodium carbonate, sodium chloride, sodium citrate, sodium iodide, sodium lactate, sodium metabisulfite, sodium nitrate, sodium nitrite, sodium phosphate, sodium propionate, sodium succinate, sodium sulfate, sodium sulfite, sodium tartrate, sodium thiosulfate, sorbitol, sucrose, tartaric acid, triethanolamine, urea, urethan, uridine, zinc sulfate, and mixtures thereof. The nebulization composition of the present invention may contain 0.0001% to 1.13% w/v of sodium chloride, preferably 0.9% w/v.
Suitable osmotic adjusting agents include, but are not limited to, sodium chloride, potassium chloride, zinc chloride, calcium chloride and mixtures thereof. Other osmotic adjusting agents may also include, but are not limited to, mannitol, glycerol, dextrose and mixtures thereof.
Any cosolvent that is suitable for inhalation and capable of dissolving or dispersing the Hydroxychloroquine sulfate and Zinc acetate dihydrate in the mixture of cosolvent and water can be used. Examples of suitable cosolvents include, for example, alcohols, ethers, hydrocarbons, and perfluorocarbons. Preferably, the cosolvent is a short chain polar alcohol. More preferably, the cosolvent is an aliphatic alcohol having from one to six carbon atoms, such as ethanol or isopropanol. The most preferred cosolvent is ethanol. Examples of suitable hydrocarbons include n-butane, isobutane, pentane, neopentane and isopentanes. Examples of suitable ethers include dimethyl ether and diethyl ether. Examples of suitable perfluorocarbons include perfluoropropane, perfluorobutane, perfluorocyclobutane, and perfluoropentane.
Suitable nonionic surfactants include all substances of this type that can normally be used in such nebulization compositions. Preferably mentioned are made of polyethylene glycol ethers of linear alcohols, reaction products of fatty acids with ethylene oxide and/or propylene oxide, and also polyvinyl alcohol, polyvinylpyrrolidone, copolymers of polyvinyl alcohol and polyvinylpyrrolidone, polyethylene oxide -polypropylene oxide block copolymers and also copolymers of (meth)acrylic acid and (meth)acrylic esters, and also alkyl ethoxylates and alkylaryl ethoxylates, which optionally phosphated and optionally neutralized with bases, it being possible for mention to be made, by way of example, of sorbitol ethoxylates. Suitable anionic surfactants include all substances of this type that can normally be used in agrochemical compositions. Preference is given to alkali metal salts and alkaline earth metal salts of alkylsulphonic acids or alkylarylsulphonic acids. Examples of suitable cationic surfactants include but are not limited to ammnonium bromides; nonionic surfactants include but are not limited to alkyl ethoxylates, alkyl gluccosides and alkyl phenol ethoxylates; and amphoteric surfactants include but are not limited to betaines, amphoacetates, and amphodiacetates. A preferred anionic surfactant is sodium dodecyl sulfate (SDS) and a preferred cationic surfactant is dodecyl trimethyl ammonium bromide; SDS is most preferred for the methods of the present invention. Also possible is the incorporation of amphoteric surfactants, such as alkyl betaines, alkyl amido betaines, and alkyl amphoacetates. Preferably, Polysorbates and Sorbitans are used as surfactants in the nebulization composition.
Suitable antioxidants include, but are not limited to, ascorbic acid, for example, provided that it has not already been used to adjust the pH, vitamin A, vitamin E, tocopherols and similar vitamins or pro-vitamins occurring in the human body.
The nebulization composition also contains a complexing agent, examples include but not limited to EDTA and salts thereof, such as edetate disodium. The nebulization composition comprising a complexing agent provides an improved stability with reduced levels of impurities and degradation products.
According to a preferred embodiment, the pharmaceutical inhalable composition is presented in the form of dry powder inhaler comprising a therapeutically effective amount of Hydroxychloroquine sulfate and Zinc acetate dihydrate along with one or more excipients to be administered either through capsule based inhalers or through blister and/ or cartridge comprising multiple pockets of storing the dry powder or reservoir based multidose inhalers (also known as dry powder compositions).
Representative non-limiting examples of carriers for use in the pharmaceutical inhalable composition presented as dry powder inhaler include, without limitation, lactose, glucose, or sodium starch glycolate. In some embodiments, the particulate carrier is lactose. The particulate lactose is in some instances alpha lactose monohydrate. In general, the particle size of the lactose should be such that it can be entrained in an air stream but not deposited in the key target sites of the lung. Apart from these carriers, there is optional presence of lubricants/ anti-adherents such as stearates or stearic acid derivatives for e.g. magnesium stearate.
The dry powder compositions, according to the invention, is conveniently presented in unit dosage form and is prepared by conventional pharmaceutical techniques. Such techniques include the step of bringing into association the compound of the invention and the pharmaceutically acceptable carrier(s), or an excipient. In general, the formulations are prepared by uniformly and intimately bringing into association the active ingredient with finely divided solid carriers, and then, if necessary, preparing discrete dosage units of the product.
The dry powder composition is metered and filled into capsules, e.g., gelatin or hydroxypropyl methylcellulose capsules, such that the capsule contains a unit dose of active ingredient.
When the dry powder is in a capsule containing a unit dose of active ingredient, the total amount of composition will depend on the size of the capsules and the characteristics of the inhalation device with which the capsules are being used. However, representative characteristic total fill weights of dry powder per capsule are between 1 and 25 mg, e.g., 5, 10, 15 or 20 mg.
Alternatively, the dry powder composition according to the invention is filled into the reservoir of a multidose dry powder inhaler (MDPI), for example of the kind illustrated in WO 92/10229.
Another aspect of the invention provides for a dry powder inhaler comprising the inhaler and a composition according to the invention.
Another aspect of the invention provides a method for the administration of a particulate medicament, comprising inhalation of a composition of the invention from a multidose dry powder inhaler.
In some embodiments, the amount of HCQ in the pharmaceutical inhalable compositions of the invention ranges between about 1 mg to about 200 mg, including any integer within this range. In some embodiments, the amount of HCQ in the pharmaceutical inhalable composition ranges between about 5 mg to about 20 mg, including any integer within this range. In some embodiments, the amount of HCQ in the pharmaceutical inhalable composition is about 10 mg.
In some embodiments, the amount of a pharmaceutically acceptable salt of HCQ in the pharmaceutical inhalable compositions of the invention ranges between about 10 mg to about 150 mg, including any integer within this range. In some embodiments, the amount of a pharmaceutically acceptable salt of HCQ in the pharmaceutical inhalable composition ranges between about 20 mg to about 100 mg, including any integer within this range. In some embodiments, the amount of a pharmaceutically acceptable salt of HCQ in the pharmaceutical inhalable composition is about 80 mg.
Herein throughout, whenever an amount of HCQ is indicated, it corresponds to an equivalent amount of an HCQ pharmaceutically acceptable salt as described herein, or an equimolar amount of an HCQ pharmaceutically acceptable salt. Thus, for example, weight amounts of hydroxychloroquine refer to an amount of hydroxychloroquine sulfate which includes the intended amount of hydroxychloroquine per se, in accordance with the widespread use of the sulfate salt, in the art. The skilled person will be readily capable of determining an amount of a pharmaceutically acceptable salt of HCQ, which will comprise the same amount of HCQ per se as in the amounts recited herein.
In some embodiments, the amount of zinc in the pharmaceutical inhalable compositions of the invention ranges between about 1mg to about 20 mg, including any integer within this range. In some embodiments, the amount of zinc in the pharmaceutical inhalable composition is about 2 mg. In some embodiments, the amount of zinc in the pharmaceutical inhalable composition is about 4 mg. In some embodiments, the amount of zinc in the pharmaceutical inhalable composition is about 8 mg.
According to the present invention, there is provided a process of preparing a pharmaceutical composition administered by inhalation comprising Hydroxychloroquine (HCQ) and synthetic minerals such as zinc acetate or zinc oxide with one or more carriers or excipients.
According to preferred embodiment of the present invention, a nebulization composition involves dispersion of Hydroxychloroquine (HCQ) and zinc acetate in suitable excipients like surfactants, followed by other excipients such as buffers, tonicity agents and making up of the volume with suitable solvent and filling in suitable containers like ampoule or vials. The containers may or may not require any form of sterilization. The nebulization composition of the present invention is administered by a suitable nebulizer. Suitable nebulizers include, but are not limited to, a jet nebulizer, an ultrasonic nebulizer, vibrating mesh nebulizer and a breath actuated nebulizer. Preferably, the nebulizer is a jet nebulizer connected to an air compressor with adequate airflow. The nebulizer being equipped with a mouthpiece or suitable face mask. The nebulization compositions are administered by nebulizers manufactured, designed or sold by Omron, such as the Omron MICRO AIR.TM. Ultrasonic Nebulizer. Other nebulizers also include those manufactured, designed, or sold by Aerogen. Additionally, the compositions described herein can also be nebulized using inhalers other than those described above, for example jet-stream inhalers or by breath actuated jet nebulizers.
According to a preferred embodiment of the present invention, a dry powder composition involves mixing and/ or homogenization of Hydroxychloroquine (HCQ) and zinc acetate followed by addition of lactose (or a specific grade). The lactose used has uniform particle size or is used in two different particle size grades i.e. coarser grade and a finer grade. Alternatively, lubricant such as magnesium stearate is added to the dry powder composition. The resulting powder blend is either filed in capsules for delivery through capsule based inhalers or is filled in blister strips or multi-pocket cartridges which will eventually be loaded in respective multidose inhalers or is filled in reservoir containers which will be loaded in respective multidose reservoir inhalers known in the art.
According to the invention, there is provided a pharmaceutical inhalable composition administered by inhalation comprising Hydroxychloroquine (HCQ) and synthetic minerals such as zinc acetate or zinc oxide for the treatment of infection caused by Coronaviridae virus (including COVID-19). Preferably, the pharmaceutical inhalable composition is in the form of nebulization composition or dry powder composition as described hereinabove. Preferably, such pharmaceutical inhalable composition comprises combination of Hydroxychloroquine (HCQ) sulfate and zinc acetate.
It may be well appreciated by a person skilled in the art that the pharmaceutical inhalable composition comprising Hydroxychloroquine (HCQ) and zinc may require specific dosage amounts and specific frequency of administrations specifically considering their individual established doses, the dosing frequency, patient adherence and the regimen adopted. As described herein, considering that there are various parameters to govern the dosage and administration of the combination composition as per the present invention, it would be well acknowledged by a person skilled in the art to exercise caution with respect to the dosage, specifically, for special populations associated with other disorders.
In order that this invention be more fully understood, the following preparative and testing methods and examples are set forth. The following examples are for the purpose of illustration of the invention only and are not intended in any way to limit the scope of the present invention.
EXAMPLES
(a) Following are representative label contents of the nebulization composition, as envisaged per the invention:
S.no. Label claim of nebulized solution
1 Each ampoule contains:
Hydroxychloroquine sulfate equivalent to Hydroxychloroquine………..40 mg
Zinc acetate dihydrate equivalent to Zinc…...…………………………….. 2mg
2 Each ampoule contains:
Hydroxychloroquine sulfate equivalent to Hydroxychloroquine………..60 mg
Zinc acetate dihydrate equivalent to Zinc…...…………………………….. 4mg
3 Each ampoule contains:
Hydroxychloroquine sulfate equivalent to Hydroxychloroquine………..80 mg
Zinc acetate dihydrate equivalent to Zinc…...…………………………….. 6 mg
4 Each ampoule contains:
Hydroxychloroquine sulfate equivalent to Hydroxychloroquine………..100 mg
Zinc acetate dihydrate equivalent to Zinc…...…………………………….. 8 mg

(b) Following are representative label contents of the dry powder composition, as envisaged per the invention
QTPP Elements Capsule Based Contents
Dosage strength (A) Each Capsule contains 10 mg of Hydroxychloroquine sulphate and 2.5 mg equivalent of Zinc in the form of Zinc acetate, with lactose. (2 doses at a time)

(B) Each Capsule contains 7.5 mg of Hydroxychloroquine sulphate and 2.5 mg equivalent of Zinc in the form of Zinc acetate, with lactose. (3 doses at a time)
(C) Each Capsule contains 5 mg of Hydroxychloroquine sulphate and 2.5 mg equivalent of Zinc in the form of Zinc acetate, with lactose. (4 doses at a time)
Net filled content per unit 30 mg ± 5% with Lactose as carrier.
18mg ± 5% without any carrier
Pack configuration (A) Capsules are supplied in HDPE bottles (10-30 capsules) along with device in carton with the patient information leaflet.

Capsules are supplied in Alu-Alu blisters along with device in carton with the patient information leaflet.

The inventors have studied the design of the dose and prediction of therapeutic effect of Hydroxychloroquine (HCQ) and zinc acetate formulations through In-silico modelling. According to the inventors, it is believed that inhaled HCQ has more efficacy against COVID-19 virus at ~20-40% of that of solid oral dose, and this consideration was determined through testing of nebulization and dry powder composition against the solid oral dose.
A Physiologically based pharmacokinetic (PBPK) model was developed and used for HCQ dose estimation through inhalation route (Nebulizer). Systemic Parameters were derived using Intravenous PK data which were then used along with other drug parameters to simulate oral PK. Lung tissue concentrations were predicted for standard 200mg oral HCQ. Simulations were performed for 80mg, 100mg and 160 mg dose through oral inhalation route. Lung tissue concentrations were compared to that of 200mg oral tablet. PBPK model was developed and used for Zinc dose estimation through inhalation route (Nebulizer). Systemic Parameters were derived using Intravenous PK data of Zinc Sulfate (ZnSO4) which were then used along with other Zinc parameters to simulate oral PK. Lung tissue Zn concentrations were predicted for 30mg oral Zinc (NutriSet tablets). Simulations were performed for 8mg dose through oral inhalation route. Lung tissue concentrations were compared to that of 30mg oral tablet. Besides this, simulation showed the following outcomes.
Results of PBPK Model:
The PBPK model for single dose was developed for HCQ on Oral route in Preludium TM software using the literature data on IV absorption based compartmental analysis, followed by the PK of oral tablet dose. This model was then validated based on inhalation PK route for the dose proposed above (nebulizer 40 mg / DPI 7.5 mg). This validated model was then used to predict the lung dose in Oral route and Inhalation route. This is shown in figures 1 to 6.
The PBPK model for single dose was developed for Zinc Acetate on Oral route in Preluidum TM software using the literature data on IV absorption based compartmental analysis, followed by the PK of oral tablet dose. This model was then validated based on inhalation PK route for the dose proposed above (nebulizer 8 mg / DPI 2.5 mg). This validated model was then used to predict the lung dose in Oral route and Inhalation route.
This provided the inhalation dose regime required to match the plasma concentration as that of oral route while providing much higher lung dose.
Then, a steady state dose regime model was developed in MATLABTM Simbiology software which provides the PBPK model when HCQ and Zn are administered daily including the loading dose. In this process, first, solid oral dose of HCQ sulphate (loading dose of 600 mg followed by daily 200mg twice) was simulated and blood plasma concentration as a function of time was simulated up to 9 days. Then, the lung dose (tissue concentration) for this profile was estimated for 9day period as shown in the figures 7 to 10 below.
The second consideration, as per the inventors, was that inhaled zinc (in acetate form) has more efficacy against COVID 19 virus at ~20-40% of that of solid oral dose, and this consideration was determined through testing of nebulization and dry powder composition against the solid oral dose. The results of this simulation were as follows-
Observations:
From figures 7 to 9, it could be seen that with nebulization and dry powder composition respectively at the 1/5th and 1/10th of the solid oral route HCQ, inhalation route provides same plasma concentration and gives about 150- 2000% more lung dose and thus, is very targeted in therapeutic action against COVID-19 and other RNA viruses where the lung is primary site of infection.
From figures 10 to 12, it could be seen that with nebulization and dry powder composition respectively at the 1/4th and 1/8th of the solid oral route Zinc, inhalation route provides same plasma concentration and gives about 150- 200% more lung dose and thus is very targeted in therapeutic action against COVID-19 and other RNA viruses where the lung is primary site of infection.
Pharmacokinetic Pharmacodynamic (PK-PD) model to predict synergistic effect of HCQ-Zn:
The third consideration, as per the inventors, was that in presence of HCQ targeted to the lung via inhalation route, the HCQ acts on the lung epithelial cells creating ionophoric effect and facilities more entry of Zn into the cells thereby the HCQ and Zinc inhaled combination has more efficacy than individual HCQ and Zinc. This effect was modelled as enhanced permeability of Zinc into the lung cells which causes higher intracellular zinc concentration, and hence provides higher efficacy as compared to efficacy of standalone zinc given.
To predict if the efficient lung dose delivery in nebulization route can provide improved therapeutic effect for HCQ and Zn, the inventors modelled the PK-PD based action mechanism. As per the knowledge available in the prior art, the information on viral load reduction for various concentrations of HCQ in a cell culture environment for various viral loading [X Yao, F Ye, M Zhang, C Cui, B Huang, P Niu, et al. In Vitro Antiviral Activity and Projection of Optimized Dosing Design of Hydroxychloroquine for the Treatment of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2).Clin Infect Dis (2020 Mar 9)] was taken as a base data for developing an E-Max model using MATLABTM software. Basis the E-Max and EC-50% fitted values, the Pk concentration curve was given as input from the Figure 7 and Figure 8 for solid oral and nebulized inhalation route respectively.
Figure 14 shows efficacy comparison of HCQ for oral vs nebulized inhalation route against the viral load for 8 days of dosing regimen. This clearly showed that inhalation HCQ is about 120% to 140% superior in its antiviral action against COVID-19 and also the viral load reduces very rapidly from the onset and day 1 of the loading day itself as against average 6 days in the case of solid oral route.
Figure 15 shows efficacy comparison of Zinc for oral vs nebulized inhalation route against the viral load for 8 days of dosing regimen. This clearly showed that Inhalation Zinc is about 20% (steady state) to 200% (first two days of dosing) superior in its antiviral action against COVID-19 and also the viral load reduces very rapidly from the onset and day 1 of the loading day itself as against average 6 days in the case of solid oral route.
Figure 16 shows enhancement of zinc permeability due to presence of HCQ. It could be seen from the graph that steady state lung dose increases by about 1.5 to 1.8 times as against the standalone Zinc inhalation administration proposed.
Further, Figure 17 shows combined efficacy of Zinc and HCQ together against the individual content and the quantum of synergic effect was to the tune of 20-40 %
The inventors thus concluded that simulation results showed lung tissue concentration using oral tablet (200mg) route was almost equivalent to Inhalation (2*40 mcg nebulization). Similarly, for the Dry powder capsule based formulation 10 mg of Hydroxychloroquine equivalent and 2.5 mg Zinc equivalent zinc, with lactose. (2 doses at a time). The efficacy data showed that HCQ and Zinc via inhalation route is 120-140% more effective in reducing the viral load while reducing the dose to 1/5th and 1/3rd respectively. Finally, the HCQ and Zinc together had a synergistic effect to the tune of 150-300% than that of HCQ and Zinc individually (taken separately) through the inhalation route.
,CLAIMS:1. An inhalable pharmaceutical composition comprising:
(A) chloroquine or its pharmaceutically acceptable salt thereof or a derivative of chloroquine or its salt;
(B) zinc or its pharmaceutically acceptable salts and derivatives; and
(C) pharmaceutically acceptable excipients.
2. The composition as claimed in claim 1, wherein the chloroquine or its pharmaceutically acceptable salt, derivative thereof is hydroxychloroquine or its pharmaceutically acceptable salts.
3. The composition as claimed in claim 1, wherein the zinc or its derivatives is selected from zinc acetate, zinc oxide, zinc chloride.
4. The composition as claimed in claim 1 is an atomizable composition.
5. The composition as claimed in claim 4 is a nebulizable composition comprising a dispersion of hydroxychloroquine or its pharmaceutically acceptable salt thereof; and zinc or its pharmaceutically acceptable salts and derivatives in an aqueous, organic or aqueous/organic medium.
6. The composition as claimed in claim 5, wherein the particle size of hydroxychloroquine or its pharmaceutically acceptable salt, and zinc acetate or its pharmaceutically acceptable salt is from about 0.1 micron to about 5 micron.
7. The composition as claimed in claim 5, wherein the amount of hydroxychloroquine or its pharmaceutically acceptable salt is from about 0.005 mg/ml to about 50 mg/ml of total formulation.
8. The composition as claimed in claim 5, wherein the amount of zinc or its salts is from about from about 0.001 mg/ml to 5 mg/ ml of total formulation.
9. The composition as claimed in claim 5, wherein volume of the nebulization composition is from about 0.2 ml to about 6 ml.
10. The composition as claimed in claim 1, wherein the pharmaceutically acceptable excipients are surfactants, isotonicity agents, pH adjusters, buffers, various sugars and types of starch, cellulose derivatives, gelatin, vegetable oils, and polyethylene glycols.
11. The composition as claimed in claim 4, wherein the composition is a dry powder comprising finely divided hydroxychloroquine or its pharmaceutically acceptable salt thereof; and zinc or its pharmaceutically acceptable salts and derivatives; optionally together with a pharmaceutically acceptable carrier in finely divided form.
12. The composition as claimed in claim 4, wherein the composition is an aerosol comprising hydroxychloroquine or its pharmaceutically acceptable salt; and zinc or its pharmaceutically acceptable salts and derivatives in solution or dispersion in a propellant.
13. The composition as claimed in claim 1 comprising chloroquine or its pharmaceutically acceptable salt thereof or a derivative of chloroquine or its salt; zinc or its pharmaceutically acceptable salts and derivatives; and pharmaceutically acceptable excipients is for use in treatment of an infection caused by COVID-19.