A delayed-release pharmaceutical composition for the inhibition of SARS-CoV-2 activity

Abstract

Delayed-release pharmaceutical composition for inhibiting the main protease of SARS-CoV-2, comprising: an effective amount of at least one natural bioactive compound selected from the group consisting of: 5,7-dihydroxy-2-(3-hydroxy-4-(((2R,3R,4R,5R,6R)-3,4,6-trihydroxy-2-(hydroxymethyl)-5-(((2S,3R,4S,5S,6R)-3, 4,5-Trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-3-yl)oxy)phenyl)-4H-chromen-4-one; ((2S,3R,4R,5S,6R)-6-((2-(3,4-dihydroxyphenyl)-5,7-dihydroxy-4-oxo-4H-chromen-3-yl)oxy)-3,4,5-trihydroxy-tetrahydro-2H-pyran-2-yl)methyl(E)-3-(4-hydroxyphenyl)acrylate; 2-(2,4-Dihydroxyphenyl)-3,5,7-trihydroxy-4H-chromen-4-one; 1,6-Dihydroxy-3-methyl-8-(((2R,3S,4R,6S)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)anthracene-9,10-dione; 1,3,8-Trihydroxy-6-(((2R,3R,4S,5S,6R)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)methyl)anthracene-9,10-dione; in combination with one or more pharmaceutically acceptable excipients selected from hydrophilic polymers, hydrophobic polymers, diluents, binders and lubricants; wherein the composition is formulated as a sustained-release oral dosage form providing controlled release of the bioactive compound(s) over a period of up to 12 hours.

Description

The present invention relates to the field of pharmaceutical compositions for antiviral therapy, in particular to sustained release oral dosage forms containing natural bioactive compounds capable of inhibiting the main protease (Mpro) of SARS-CoV-2. More particularly, the invention provides a sustained release pharmaceutical composition which delivers one or more natural compounds having strong Mpro inhibitory activity in a controlled manner over a prolonged period of time, thereby improving the therapeutic efficacy and compliance of patients.The worldwide outbreak of SARS-CoV-2 presents a significant public health threat and has resulted in severe airway disorders and high mortality rates worldwide. Despite the development of vaccines and synthetic antiviral drugs, problems still exist such as restricted accessibility, possible side effects, the appearance of drug resistant virus strains and lower efficacy against new variants. This emphasizes the urgent need for alternative therapeutic strategies which are safe, effective and affordable for broad use.Natural materials have long been known for their broad spectrum of antiviral activities and their minimal side effects due to their biocompatibility. Several plant derived compounds have shown that they can inhibit the entry, proliferation and construction of viruses by targeting important viral proteins and host receptors. However, scientifically validated, standardized natural compositions developed specifically for effective control of SARS-CoV-2 are lacking.The present invention addresses these limitations by providing a novel composition of natural bioactive compounds with demonstrated antiviral activity against SARS-CoV-2. The formulation is intended to inhibit virus replication and prevent the interaction between virus and host cell, thereby reducing the severity of the infection and promoting recovery. This natural product-based approach provides a safer, more persistent and potentially more versatile solution to handling current and newly occurring coronavirus infections.The main object of the present invention is to provide an oral sustained release pharmaceutical composition containing one or more natural bioactive compounds capable of inhibiting the SARS-CoV-2 main protease (Mpro) and ensuring a controlled release for up to 12 hours.The present disclosure further provides for improving the bioavailability of natural Mpro inhibitors.Another object of the present disclosure is to reduce the frequency of dosing, thereby improving patient compliance.Another object of the present disclosure is to minimize variations in plasma concentration to ensure consistent antiviral activity.Another object of the present disclosure is to provide aids in the use of pharmaceutically acceptable excipients that allow sustained release of bioactive compounds.The present invention relates generally to a sustained release pharmaceutical composition for inhibiting the major protease (Mpro) of SARS-CoV-2. The composition comprises an effective amount of at least one natural bioactive compound selected from the group consisting of:CP1: 5,7-dihydroxy-2-(3-hydroxy-4-(((2R,3R,4R,5R,6R)-3,4,6-trihydroxy-2-(hydroxymethyl)-5-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-3-yl)oxy)phenyl)-4H-chromen-4-oneCP2: ((2S,3R,4R,5S,6R)-6-((2-(3,4-Dihydroxyphenyl)-5,7-dihydroxy-4-oxo-4H-chromen-3-yl)oxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)methyl(E)-3-(4-hydroxyphenyl)acrylateCP3: 2-(2,4-dihydroxyphenyl)-3,5,7-trihydroxy-4H-chromen-4-onesCP4: 1,6-dihydroxy-3-methyl-8-(((2R,3S,4R,6S)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)anthracene-9,10-dioneCP5: 1,3,8-trihydroxy-6-(((2R,3R,4S,5S,6R)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)methyl)anthracene-9,10-dioneThe composition further contains one or more pharmaceutically acceptable adjuvants selected from hydrophilic polymers, hydrophobic polymers, diluents, lubricants and binders. The formulation is designed as a sustained release oral dosage form that allows for controlled release of the bioactive compound(s) over a period of up to 12 hours.These compounds were selected based on their demonstrated or potential ability to inhibit SARS-CoV-2Pro by molecular docking. Each compound has structural features such as multiple hydroxyl groups and glycosidic bonds that contribute to its binding affinity to the active site of Mpro. The use of flavonoids (e.g. CP1, CP2, CP3) and anthraquinone derivatives (e.g. CP4, CP5) extends the spectrum of antiviral activity and at the same time offers antioxidant and anti-inflammatory advantages which may assist concomitant therapy in COVID-19.EXAMPLE 2; Pharmaceutically Acceptable Excipients:Hydrophilic polymers (e.g. hydroxypropyl methylcellulose [HPMC], carboxymethylcellulose [CMC]) are used to form a gel barrier upon hydration which regulates the release of the active agents.Hydrophobic polymers (e.g., ethyl cellulose, polymethacrylates) may be included to retard diffusion of bioactive agents and prolong release.Diluents (e.g., lactose, microcrystalline cellulose) provide an adequate volume in tablet compression.Binders (e.g. povidone, starch) promote cohesion during granulation.Lubricating agents (e.g. magnesium stearate, stearic acid) aid in the manufacturing process.EXAMPLE 3; Formulation and ProcedureThe sustained release oral dosage form may be prepared by methods such as, but not limited to, wet granulation, dry granulation, or direct compression. The selection of polymers and their concentrations is optimized to achieve the desired release kinetics that the active agents seek to release over a period of 6 to 12 hours.The formulation strategy comprises:Sustained release tablets of the matrix type wherein the bioactive substances are embedded in a polymer matrix.coated granules or pellets encapsulated in a capsule and enabling modified release profiles.multilayer tablets having an outer immediate release layer and a sustained release coreEXAMPLE 4: Sustained Release Tablet CompositionActive ingredient in an amount of: 100 mgHPMC K15M: 150 mgmicrocrystalline cellulose: 100 mgPovidone K30: 20 mg- Magnesium stearate: 5 mglactose monohydrate: 125 mgwherein the active ingredients are selected from the group consisting of Cp1-Cp5, wherein the IUPAC names of the active ingredients are as follows:CP1: 5,7-dihydroxy-2-(3-hydroxy-4-(((2R,3R,4R,5R,6R)-3,4,6-trihydroxy-2-(hydroxymethyl)-5-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-3-yl)oxy)phenyl)-4H-chromen-4-one; CP2: ((2S,3R,4R,5S,6R)-6-((2-(3,4-dihydroxyphenyl)-5,7-dihydroxy-4-oxo-4H-chromen-3-yl)oxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)methyl(E)-3-(4-hydroxyphenyl)acrylate; CP3: 2-(2,4-dihydroxyphenyl)-3,5,7-trihydroxy-4H-chromen-4-one; CP4: 1,6-dihydroxy-3-methyl-8-(((2R,3S,4R,6S)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)anthracene-9,10-dione; CP5: 1,3,8-trihydroxy-6-(((2R,3R,4S,5S,6R)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)methyl)anthracene-9,10-dione.EXAMPLE 5: Wet Granulation MethodIn the wet granulation method, the process begins by weighing precisely all necessary ingredients, including the natural bioactive compound (e.g., CP1-CP5), the hydrophilic polymer such as hydroxypropyl methylcellulose (HPMC K15M), microcrystalline cellulose (MCC), lactose monohydrate and other adjuvants. These ingredients are sieved through a 40 mesh sieve to ensure uniform particle size and remove agglomerates. The sieved powders are then dry blended in a rapid mix granulator or planetary mixer for 5 to 10 minutes to achieve uniform distribution. Separately, a binder solution is prepared by dissolving povidone K30 in purified water. This binder solution is slowly added to the dry mix with mixing to form a uniform wet mass. The wet mass is then passed through a 16 or 20 mesh screen to form wet granules which are subsequently dried in a tray dryer or a fluidized bed dryer at a controlled temperature of 40 to 50°C until the moisture content is reduced to an acceptable level, typically 1 to 2%. After drying, the granulate is sieved through a 20-mesh sieve to break up any lumps. The dried granules are then mixed with magnesium stearate as a lubricant (and optionally with colloidal silica as a lubricant) for about 3 to 5 minutes to assure uniform distribution without admixture. Finally, the oiled granulate is pressed into tablets with the aid of a rotary tablet press.EXAMPLE 6: EvaluationsWeight variation testThe weight variation test is performed to ensure that the weight of the individual tablets is uniform, reflecting a uniform drug content. Twenty tablets are randomly selected from each batch and individually weighed with an electronic balance. The average weight is calculated and the individual tablet weights are compared to the average. According to the Pharmacopoeia standards, if the tablet weighs more than 250 mg, no more than two tablets should deviate by ± 5% from the average weight.The average tablet weight was 500 mg. The weight variation for all tablets was within the permissible range, with the maximum variation being ±2.1%. None of the tablets exceeded the pharmacopoeia limits.Hardness TestTablet hardness or breaking strength is measured to determine the mechanical integrity of the tablet during handling and transport. A Monsanto or Pfizer hardness tester measures the force required to break the tablet. Ten tablets are selected randomly and tested individually. The optimum hardness for sustained release tablets is generally in the range of 5 to 8 kg / cm 2, to maintain integrity and control the release profile of the active ingredient.The tablets had an average hardness of 6.5 kg / cm 2 with a range of 6.0 to 7.0 kg / cm 2. This indicates sufficient mechanical strength for handling and packaging without breaking the tablets.Friability TestThe friability test evaluates the ability of the tablet to resist attrition during transport and handling. A sample of 6.5 g of tablets is placed in a Roche Friabilator and rotated at 25 U / min for 4 minutes. After the passage, the tablets are dusted off and reweighed. The weight loss percentage is calculated. According to the pharmacopoeia limits, friability should be less than 1% to achieve acceptable mechanical strength.The friability of the sustained release tablets was 0.45% and thus within the permissible range, which indicates good fracture and chipping resistance.Thickness and DiameterThe thickness and diameter of ten randomly selected tablets are measured with a caliper or digital micrometer. Uniform tablet thickness ensures uniform tablet appearance, packaging and drug release. Variations in thickness may indicate problems with compression force during tableting.The tablets had an average thickness of 4.2 ± 0.1 mm and an average diameter of 12.0 ± 0.2 mm. All tablets were of uniform size and were within the design parameters set.Disintegration TestAlthough sustained release tablets are designed not to disintegrate rapidly, disintegration tests for quality control are sometimes carried out. Six tablets are placed in water at 37 ± 0.5°C in a disintegration tester. The test checks whether formulation errors lead to premature decomposition.The tablets remained intact in the disintegration medium for more than 12 hours, confirming their sustained release and robustness.Uniformity of Drug ContentThe uniformity of drug content ensures that each tablet contains the correct amount of pharmaceutical drug (API). Ten tablets are individually comminuted and an accurately weighed portion corresponding to one tablet is dissolved in a suitable solvent. The solution is filtered, diluted accordingly and analysed by UV spectrophotometry or HPLC in order to determine the concentration of active compound.The active ingredient content in the tablets was between 98.5% and 101.2% of the stated amount, which proves excellent uniformity of the content and compliance with the pharmaceutical standards.In vitro Release StudyIn the in vitro release assay, the release profile of the drug over time is evaluated. The test is carried out with a USP type II (paddle) dissolution apparatus in 900 mL pH 6.8 phosphate buffer at 37 ± 0.5°C, with the paddle rotating at 50 U / min. The samples are taken at specific time intervals (1, 2, 4, 6, 8, 10 and 12 hours), filtered and assayed for the active ingredient content by UV-Vis spectrophotometry or HPLC. Release kinetics were then examined to confirm sustained release of drug.The tablets exhibited cumulative drug release of 25% after 2 hours, 60% after 6 hours and 95% after 12 hours, following zero order kinetics, confirming controlled and sustained drug release over 12 hours.Examples1. The present invention relates generally to a sustained release pharmaceutical composition for inhibiting the major protease (Mpro) of SARS-CoV-2. The composition comprises an effective amount of at least one natural bioactive compound selected from the group consisting of:CP1: 5,7-dihydroxy-2-(3-hydroxy-4-(((2R,3R,4R,5R,6R)-3,4,6-trihydroxy-2-(hydroxymethyl)-5-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-3-yl)oxy)phenyl)-4H-chromen-4-oneCP2: ((2S,3R,4R,5S,6R)-6-((2-(3,4-Dihydroxyphenyl)-5,7-dihydroxy-4-oxo-4H-chromen-3-yl)oxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)methyl(E)-3-(4-hydroxyphenyl)acrylateCP3: 2-(2,4-dihydroxyphenyl)-3,5,7-trihydroxy-4H-chromen-4-onesCP4: 1,6-dihydroxy-3-methyl-8-(((2R,3S,4R,6S)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)anthracene-9,10-dioneCP5: 1,3,8-trihydroxy-6-(((2R,3R,4S,5S,6R)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)methyl)anthracene-9,10-dioneThe composition further contains one or more pharmaceutically acceptable adjuvants selected from hydrophilic polymers, hydrophobic polymers, diluents, lubricants and binders. The formulation is designed as a sustained release oral dosage form that allows for controlled release of the bioactive compound(s) over a period of up to 12 hours.

Claims

A sustained release pharmaceutical composition for inhibiting the major protease of SARS-CoV-2, comprising: an effective amount of at least one natural bioactive compound selected from the group consisting of: 5,7-dihydroxy-2-(3-hydroxy-4-(((2R,3R,4R,5R,6R)-3,4,6-trihydroxy-2-(hydroxymethyl)-5-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-3-yl)oxy)phenyl)-4H-chromen-4-one; (2S,3R,4R,5S,6R)-6-((2-(3,4-Dihydroxyphenyl)-5,7-dihydroxy-4-oxo-4H-chromen-3-yl)oxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)methyl(E)-3-(4-hydroxyphenyl)acrylate; 2-(2,4-Dihydroxyphenyl)-3,5,7-trihydroxy-4H-chromen-4-one; 1,6-Dihydroxy-3-methyl-8-(((2R,3S,4R,6S)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)anthracene-9,10-dione; 1,3,8-Trihydroxy-6-(((2R,3R,4S,5S,6R)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)methyl)anthracene-9,10-dione; in combination with one or more pharmaceutically acceptable excipients selected from hydrophilic polymers, hydrophobic polymers, diluents, binders and lubricants; wherein the composition is formulated as a sustained release oral dosage form providing a controlled release of the bioactive compound(s) over a period of up to 12 hours.The sustained release pharmaceutical composition of claim 1, wherein the hydrophilic polymer is selected from hydroxypropyl methylcellulose (HPMC), preferably HPMC K15M, to modulate the release rate of the bioactive compound.The sustained release pharmaceutical composition according to claim 1, wherein the oral dosage form is a tablet prepared by a wet granulation method comprising the steps of mixing, granulating using a binder solution, drying, sizing, lubricating and compressing.The sustained release pharmaceutical composition of claim 1, wherein the composition has a drug release profile characterized by an initial release of 20-30% of the drug within 2 hours, followed by a sustained release that achieves a cumulative drug release of 90-95% over 12 hours.