WO2024051793A1

WO2024051793A1 - METHODS ANS KITS FOR TREATING SARS-CoV-2 INFECTION

Info

Publication number: WO2024051793A1
Application number: PCT/CN2023/117619
Authority: WO
Inventors: Zhen Wang; Haji Akber Aisa; Yuanchao XIE; Bo Zhao; Yu Wang; Zhuobing Zhang; Ning Li
Original assignee: Shanghai Vinnerna Biosciences Co., Ltd.
Priority date: 2022-09-09
Filing date: 2023-09-08
Publication date: 2024-03-14

Abstract

A method and kit for treating, preventing, or alleviating a SARS-CoV-2 infection or a condition or symptom associated with the SARS-CoV-2 infection. The method comprises administering to the subject a therapeutically effective amount of a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof, wherein the nucleoside analog is VV116 or VV116-3.

Description

METHODS ANS KITS FOR TREATING SARS-CoV-2 INFECTION

TECHNICAL FIELD

The present invention relates to methods and kits for treating SARS-CoV-2infection.

BACKGROUND

Coronaviruses are a group of positive-sense single-strand RNA viruses in the family Coronaviridae in the order Nidovirales. Coronaviruses can be further classified by genera: alphacoronavirus, betacoronavirus, gammacoronavirus, and deltacoronavirus. Coronaviruses can cause a wide range of disease in both humans and animals, including the common cold. In some cases, coronavirus infection can be more severe. Examples of coronaviruses include SARS-CoV (causing Severe Acute Respiratory Syndrome (SARS) ) , MERS-CoV (Middle East Respiratory Syndrome (MERS) , also sometimes called camel flu) .

Coronavirus disease 2019 (Covid-19) pandemic continues to spread rapidly worldwide. With the selective pressure of host-immunity, vaccines and antivirals, the wild-type severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) identified back in 2020 has now been fueled to the evolution of variants with increasing transmissibility and capability of evading human immunity, e.g., the Omicron variant, currently circulating worldwide. As estimated in a modeling transmission study, the outbreak of Omicron would overwhelm a local healthcare system with a burden of 15.6 times the available intensive care unit capacity. In addition to high coverage of vaccination and effective implementation of non-pharmaceutical interventions, a widespread and timely distributable efficacious antiviral therapy is the key strategy to effective epidemic control.

SUMMARY

In a first aspect, the present application provides a method of treating, preventing, or alleviating a SARS-CoV-2 infection or a condition or symptom associated with the SARS-CoV-2 infection in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof, or a pharmaceutical composition comprising a therapeutically effective amount of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof.

In a second aspect, the present application provides a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof for use in a method for treating, preventing, or alleviating a SARS-CoV-2 infection or a condition or symptom associated with the SARS-CoV-2 infection.

In a third aspect, the present application provides use of a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof in the preparation of a medicament or a kit for treating, preventing, or alleviating a SARS-CoV-2 infection or a condition or symptom associated with the SARS-CoV-2 infection.

In a fourth aspect, the present application provides a method of treating, preventing, or alleviating COVID-19 in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof, or a pharmaceutical composition comprising a therapeutically effective amount of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof.

In a fifth aspect, the present application provides a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof for used in a method for treating, preventing, or alleviating COVID-19.

In a sixth aspect, the present application provides use of a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof in the preparation of a medicament or a kit for treating, preventing, or alleviating COVID-19.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows the CONSORT diagram of the study. Participants were recruited between April 4, 2022 and May 2, 2022 from 7 sites in Shanghai, China. Consort diagram were summarized by data cut-off date on May 13, 2022.

FIG. 2 shows the time to sustained clinical recovery. Shown are estimated time to sustained clinical recovery of COVID-19 related symptom by Kaplan-Meier method for each treatment group in the full analysis population (n = 771) (A) , per-protocol population (n = 729) (B) and participants who commenced trial regimen within 5 days after symptom onset (n = 596) (C) . 95%CI is estimated using normal approximation Brookmeyer-Crowley method based on log-log transform. Hazard ratio is calculated using the COX proportional hazards model. P value is calculated using log-rank test. The non-inferiority margin δ = 0.8. CI = confidence interval, NE = not estimable.

FIG. 3 shows the in vitro activity of VV116 and VV116-3 against SARS-CoV-2 Omicron variant BA. 5.2.

FIG. 4 shows the in vitro activity of PF-07321332 against SARS-CoV-2 Omicron variant BA. 5.2.

DETAILED DESCRIPTION

The present invention relates to a method for treating SARS-CoV-2 infection. The method of the present invention comprises administering to a subject a therapeutically effective amount of a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof.

Terminology

In order to facilitate the understanding of the present invention, some technical and scientific terms are specifically defined below. Unless otherwise specifically defined herein, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which the present invention belongs.

As used in this specification and the appended claims, the singular forms "a" , "an" and "the" include plural referents, unless otherwise specified. Thus, for example, reference to "a polypeptide" includes a combination of two or more polypeptides and the like.

As used herein, when referring to measurable values (e.g., amounts, durations, etc. ) , "about" is intended to encompass variations of ±20%or ±10%based on the particular value, including ±5%, ±1%and ±0.1%, as such variations are suitable for implementing the disclosed methods.

The term “effective amount” as used herein refers to an amount of an active pharmaceutical ingredient, e.g., VV116, that produces an acute or chronic therapeutic effect upon appropriate dose administration. The effect includes prevention, correction, inhibition, or reversal of symptoms, signs and underlying pathology of a disease/condition (e.g., COVID-19) and related complications to any detectable extent.

“A therapeutically effective amount” means an amount of a pharmaceutically active ingredient, e.g., VV116, that, when administered to a subject (e.g., a human or an animal) for treating a disease, is sufficient to affect such treatment for the disease. The "therapeutically effective amount" will vary depending on the active pharmaceutical ingredient, the disease and its severity and the age, weight, etc., of the subject to be treated. A therapeutically effective amount can refer to a dose as set forth herein.

“Treating” includes any effect, e.g., lessening, slowing down, reducing, reversing, modulating, or eliminating, that results in the improvement of the condition, disease, disorder, etc. "Treating" or "treatment" of a disease state includes: arresting the development of the disease state or its clinical symptoms; or relieving the disease state, i.e., causing temporary or permanent regression of the disease state or its clinical symptoms.

“Preventing” the disease state includes causing the clinical symptoms of the disease state not to develop in a subject that may be exposed to or predisposed to the disease state, but does not yet experience or display symptoms of the disease state.

The term “alleviating” does not necessarily require curative results. As used therein, “delaying” the development of a target disease or disorder means to defer, hinder, slow, re-tard, stabilize, and/or postpone progression of the disease. This delay can be of varying lengths of time, depending on the history of the disease and/or individuals being treated. A method that “delays” or alleviates the development of a disease, or delays the onset of the disease, is a method that reduces probability of developing one or more symptoms of the dis-ease in a given time frame and/or reduces extent of the symptoms in a given time frame, when compared to not using the method. Such comparisons are typically based on clinical studies, using a number of subjects sufficient to give a statistically significant result.

“Disease state” means any disease, disorder, condition, symptom, or indication.

The term "administering" refers to the placement of a compound as disclosed herein into a subject by a method or a route which results in at least partial delivery of the compound at a desired site. Pharmaceutical compositions comprising the compound (s) disclosed herein can be administered by any appropriate route which results in an effective treatment in the subject.

An "adverse event" (AE) described herein is any adverse and often unintended or undesirable sign, symptom, or disease associated with the use of medical treatment. For example, an adverse event may be associated with the activation of the immune system or the expansion of immune system cells in response to treatment. The medical treatment may have one or more related AEs, and each AE may have the same or a different severity level.

The term "subject" , "patient" and "individual" include any organism, preferably an animal, more preferably a mammal (such as rat, mouse, dog, cat and rabbit) , and most preferably a human. The terms "subject" , "patient" and "patient" are used interchangeably herein.

The term "in need thereof" means that a subject has a disease, is diagnosed with a disease, or is in need of treating or preventing a disease. A subject in need thereof can be a subject in need of treating or preventing a disease.

The term "pharmaceutically acceptable" refers to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.

The term "pharmaceutically acceptable carrier" means a pharmaceutically-acceptable material, composition or vehicle, such as a liquid or solid filler, diluent, excipient, manufacturing aid (e.g., lubricant, talc magnesium, calcium or zinc stearate, or steric acid) , or solvent encapsulating material necessary or used in formulating an active ingredient or agent for delivery to a subject. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not injurious to the patient.

The term “coronavirus” or “CoV” as used herein refers to any member of Coronaviridae family, including, but not limited to, the wild-type coronavirus, naturally-occurring coronavirus variants and coronavirus variants generated in a laboratory. In some embodiments, the term “coronavirus” or “CoV” refers to SARS-CoV or SARS-CoV-2. In some embodiments, the term “SARS-CoV-2” refers to wild-type SARS-CoV-2 or SARS-CoV-2 variants, including, but not limited to, Alpha variant (BA 0.1.7) , Beta variant (B. 1.351) , Gamma variant (P. 1) , Delta variant (B. 1.617.2) , and Omicron variant (B. 1.1.529) . In some embodiments, the term “SARS-CoV-2 Omicron variants” refers to wild-type Omicron or Omicron variants

Nucleoside analogs

"Nucleoside analog" as used herein refers to any one of the nucleoside analogs described in WO2021213288A1, or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof, which is incorporated herein by reference in its entirety.

In a particularly preferred embodiment, the nucleoside analog that can be used in any of the uses, therapies, and methods of the present invention is VV116, and the chemical structural formula of VV116 is as follows:

In a particularly preferred embodiment, the nucleoside analog that can be used in any of the uses, therapies, and methods of the present invention is VV116-3, and the chemical structural formula of VV116-3 is as follows:

In some embodiments, the pharmaceutically acceptable salt is a conventional non-toxic salt formed by reaction of the nucleoside analog with an inorganic or organic acid, preferably selected from the group consisting of hydrochloride, hydrobromide and mesylate.

Uses and methods

In a first aspect, the present application provides a method of treating, preventing, or alleviating a SARS-CoV-2 infection or a condition or symptom associated with the SARS-CoV-2 infection in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof.

In one or more embodiments, the condition or symptom of the SARS-CoV-2 infection is COVID-19.

In one or more embodiments, the condition or symptom of the SARS-CoV-2 infection is mild-to-moderate COVID-19.

In one or more embodiments, the condition or symptom of the SARS-CoV-2 infection is fever or chill, cough, shortness of breath or difficult breathing, fatigue, muscle or body aches, headache, new loss of taste or smell, sore throat, congestion or runny nose, nausea or vomiting, or diarrhea.

In one or more embodiments, the condition or symptom of the SARS-CoV-2 infection is fever or chill, cough, shortness of breath, or loss of smell.

In one or more embodiments, the subject has or is suspected of having a SARS-CoV-2 infection.

In one or more embodiments, the subject has been or thought to have been exposed to a SARS-CoV-2 and has not yet developed a symptom of a SARS-CoV-2 infection.

In a fourth aspect, the present application provides a method of treating, preventing, or alleviating COVID-19 in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof.

In one or more embodiments, the COVID-19 is mild-to-moderate COVID-19.

In one or more embodiments, the COVID-19 is caused by SARS-CoV-2.

In one or more embodiments, the SARS-CoV-2 includes, but is not limited to: SARS-CoV-2 alpha, beta, gamma, delta, lambda, Epsilon, Kappa, Eta, Iota, Theta, Zeta and Omicron variants.

In one or more embodiments, the SARS-CoV-2 includes, but is not limited to: A. 1-A. 6, B. 3-B. 7, B. 9, B. 10, B. 13-B. 16, B. 2, B. 1 lineage (including, but not limited to, B. 1, B. 1.1, B. 1.1.7, B. 1.1.7 with E484K, B. 1.2, B. 1.5-B. 1.72, B. 1.9, B. 1.13, B. 1.22, B. 1.26, B. 1.37, B. 1.3-B. 1.66, B. 1.177, B. 1.243, B. 1.313, B. 1.351, B. 1.427, B. 1.429, B. 1.525, B. 1.526, B. 1.526.1, B. 1.526.2, B. 1.617, B. 1.617.1, B. 1.617.2, B. 1.617.3, B. 1.619, B. 1.620, and B. 1.621) , P. 1, P. 2, P. 3, and R. 1.

In one or more embodiments, the SARS-CoV-2 is a SARS-CoV-2 Omicron variant.

In one or more embodiments, the SARS-CoV-2 Omicron variant include, but are not limited to: AY. 100, AY. 3, AY. 39.1.2, AY. 39.1.4, AY. 43, B. 1, B. 1.1.528, B. 1.1.529, B. 1.617.2, BA. 1, BA. 1.1, BA. 1.1.1, BA. 1.1.10, BA. 1.1.11, BA. 1.1.12, BA. 1.1.13, BA. 1.1.14, BA. 1.1.15, BA. 1.1.16, BA. 1.1.17, BA. 1.1.18, BA. 1.1.2, BA. 1.1.3, BA. 1.1.4, BA. 1.1.5, BA. 1.1.6, BA. 1.1.7, BA. 1.1.8, BA. 1.1.9, BA. 1.10, BA. 1.12, BA. 1.13, BA. 1.13.1, BA. 1.14, BA. 1.14.1, BA. 1.14.2, BA. 1.15, BA. 1.15.1, BA. 1.15.2, BA. 1.15.3, BA. 1.16, BA. 1.16.1, BA. 1.16.2, BA. 1.17, BA. 1.17.1, BA. 1.17.2, BA. 1.18, BA. 1.19, BA. 1.2, BA. 1.20, BA. 1.21, BA. 1.21.1, BA. 1.22, BA. 1.23, BA. 1.24, BA. 1.3, BA. 1.4, BA. 1.5, BA. 1.6, BA. 1.7, BA. 1.8, BA. 1.9, BA. 2, BA. 2.1, BA. 2.10, BA. 2.10.1, BA. 2.10.2, BA. 2.10.3, BA. 2.11, BA. 2.12, BA. 2.12.1, BA. 2.12.2, BA. 2.13, BA. 2.14, BA. 2.15, BA. 2.16, BA. 2.17, BA. 2.18, BA. 2.19, BA. 2.2, BA. 2.2.1, BA. 2.20, BA. 2.21, BA. 2.22, BA. 2.23, BA. 2.23.1, BA. 2.24, BA. 2.25, BA. 2.25.1, BA. 2.26, BA. 2.27, BA. 2.28, BA. 2.29, BA. 2.3, BA. 2.3.1, BA. 2.3.10, BA. 2.3.11, BA. 2.3.12, BA. 2.3.13, BA. 2.3.14, BA. 2.3.15, BA. 2.3.16, BA. 2.3.17, BA. 2.3.18, BA. 2.3.2, BA. 2.3.3, BA. 2.3.4, BA. 2.3.5, BA. 2.3.6, BA. 2.3.7, BA. 2.3.8, BA. 2.3.9, BA. 2.30, BA. 2.31, BA. 2.32, BA. 2.33, BA. 2.34, BA. 2.35, BA. 2.36, BA. 2.37, BA. 2.38, BA. 2.38.1, BA. 2.39, BA. 2.4, BA. 2.40, BA. 2.40.1, BA. 2.41, BA. 2.42, BA. 2.43, BA. 2.44, BA. 2.45, BA. 2.46, BA. 2.47, BA. 2.48, BA. 2.49, BA. 2.5, BA. 2.50, BA. 2.51, BA. 2.52, BA. 2.53, BA. 2.54, BA. 2.55, BA. 2.56, BA. 2.56.1, BA. 2.57, BA. 2.58, BA. 2.59, BA. 2.6, BA. 2.60, BA. 2.61, BA. 2.62, BA. 2.63, BA. 2.64, BA. 2.65, BA. 2.66, BA. 2.67, BA. 2.68, BA. 2.69, BA. 2.7, BA. 2.70, BA. 2.71, BA. 2.72, BA. 2.73, BA. 2.74, BA. 2.75, BA. 2.76, BA. 2.77, BA. 2.78, BA. 2.79, BA. 2.79.1, BA. 2.8, BA. 2.80, BA. 2.81, BA. 2.9, BA. 2.9.1, BA. 2.9.2, BA. 2.9.3, BA. 2.9.4, BA. 2.9.5, BA. 3, BA. 3.1, BA. 4, BA. 4.1, BA. 4.1.1, BA. 4.1.2, BA. 4.1.3, BA. 4.1.4, BA. 4.2, BA. 4.3, BA. 4.4, BA. 4.5, BA. 4.6, BA. 4.7, BA. 5, BA. 5.1, BA. 5.1.1, BA. 5.1.2, BA. 5.1.3, BA. 5.1.4, BA. 5.2, BA. 5.2.1, BA. 5.2.2, BA. 5.2.3, BA. 5.2.4, BA. 5.3, BA. 5.3.1, BA. 5.3.2, BA. 5.3.3, BA. 5.3.4, BA. 5.5, BA. 5.6, BC. 1, BC. 2, BD. 1, BE. 1, BE. 1.1, BE. 2, BE. 3, BF. 1, BF. 1.1, BF. 2, BF. 3, BF. 4, BF. 5, BF. 15, BG. 1, BG. 2, BG. 3, BG. 4, XAA, XAB, XAC, XAD, XAE, XAF, XAG, XAH, XD, XE, XF, XG, XH, XJ, XK, XL, XM, XN, XP, XQ, XR, XS, XT, XU, XV, XW, XY, XZ; preferably, BA. 5.2.

In one or more embodiments, the nucleoside analog is a compound of formula I as described herein or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof.

In one or more embodiments, the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is administered at a single dose of about 100 to about 800 mg, preferably from about 300 to about 600 mg. Preferably, non-limiting examples of the aforementioned dose is about 100 mg, about 200 mg, about 300 mg, about 400 mg, about 500 mg, about 600 mg, about 700 mg, about 800 mg.

In one or more embodiments, the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is administered at a frequency of about once a day, twice a day, thrice a day, once every two days.

In one or more embodiments, the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is administered for 1-10 days or longer, preferably 1 day, 3 days, 5 days, 6 days, 10 days.

In one or more embodiments, the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is administered orally on an empty stomach or on a regular diet.

In one or more embodiments, the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is administered orally at a single dose of about 600 mg on day 1 and about 300 mg on day 2 to 5, twice a day.

In one or more embodiments, the VV116 or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is in a formulation comprising about 80-120mg or about 280-320mg VV116 hydrobromide; preferably, the formulation is a tablet.

In one or more embodiments, the VV116 or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is in a formulation comprising about 80-120mg VV116 hydrobromide, about 80-120mg aminoalkyl methacrylate copolymer, about 80-100mg lactose monohydrate, about 80-100mg microcrystalline cellulose, about 14-18mg croscarmellose sodium, about 3-5mg magnesium stearate, about 10-15mg Gastric soluble film coating premix; or about 280-320mg VV116 hydrobromide, about 280-320mg aminoalkyl methacrylate copolymer, about 250-300mg lactose monohydrate, about 250-300mg microcrystalline cellulose, about 45-50mg croscarmellose sodium, about 10-15mg magnesium stearate, about 32-40mg Gastric soluble film coating premix.

In one or more embodiments, the VV116 or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is in a tablet comprising about 100mg VV116 hydrobromide, about 100mg aminoalkyl methacrylate copolymer, about 90mg lactose monohydrate, about 90mg microcrystalline cellulose, about 16mg croscarmellose sodium, about 4mg magnesium stearate, about 12mg Gastric soluble film coating premix; or about 300mg VV116 hydrobromide, about 300mg aminoalkyl methacrylate copolymer, about 270mg lactose monohydrate, about 270mg microcrystalline cellulose, about 48mg croscarmellose sodium, about 12mg magnesium stearate, about 36mg Gastric soluble film coating premix. As used herein, the aminoalkyl methacrylate copolymer purchased from Evonik Operations GmbH (pharmaceutical excipient registration number: F20180001249, Center for Drug Evaluation, National Medical Products Administration, China) ; the Gastric soluble film coating premix can be purchased from Evonik Operations GmbH, JRS Pharma GmbH &Co. KG, etc.

In one or more embodiments, the therapeutically effective amount of a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof achieves a median time from randomization to sustained clinical recovery 4 days post administration.

In one or more embodiments, lower than about 3%, 2.5, or 2.3%patients exhibit grade 3 or higher treatment emergent adverse event (TEAEs) from the treatment.

In a particularly preferred embodiment, provided herein is a method of treating, preventing, or alleviating a SARS-CoV-2 infection, especially the SARS-CoV-2 infection is SARS-CoV-2 Omicron variants infection, wherein the method comprises administering to the subject in need thereof of a nucleoside analog VV116, wherein the dosage regimen is: administered orally at a single dose of about 600 mg on day 1 and about 300 mg on day 2 to 5, twice a day.

In a particularly preferred embodiment, provided herein is a method of treating, preventing, or alleviating the COVID-19 caused by SARS-CoV-2, especially the COVID-19 caused by SARS-CoV-2 Omicron variants, the method comprising administering to the subject in need thereof of a nucleoside analog VV116, wherein the dosage regimen is: administered orally at a single dose of about 600 mg on day 1 and about 300 mg on day 2 to 5, twice a day; In a particularly preferred embodiment, the COVID-19 is mild-to-moderate COVID-19.

Kits

The present application also provides a kit comprising doses of a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof disclosed herein for oral administration for at least 5 days; wherein the doses include:

one dose for oral administration on day 1 in a frequency of once a day, comprising one or several formulations of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof, wherein the total amount of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof in the dose is in a range of from about 400 mg to about 800 mg; and

doses for oral administration for the rest of the days in a frequency of twice a day, each of which comprises one or several formulations of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof, wherein the total amount of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof in each dose is in a range of from about 80mg to 300mg.

Preferably, the dose for oral administration on day 1 in a frequency of once a day comprises about 600mg of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof, and each of the doses for oral administration for the rest of the days in a frequency of twice a day comprises about 300mg of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof.

Preferably, the kit comprises one dose for administration at day 1 and 8 doses for administration at day 2 to day 5. Preferably, the dose for administration at day 1 comprises one or more formulations comprising about 600mg of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof in total, and each of the 8 doses for administration at day 2 to day 5 comprises one or more formulations comprising about 300mg of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof in total. For example, each formulation may comprise about 100mg of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof, and thus the one dose for administration at day 1 should contain 6 formulations so as that the subject could receive about 600 mg of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof at day 1 in one administration.

Preferably, the formulation of the present application is a pharmaceutical composition comprising the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof and one or more pharmaceutically acceptable carriers or excipients. Preferably, the one or more pharmaceutically acceptable carriers or excipients are selected from a group consisting of aminoalkyl methacrylate copolymer, lactose monohydrate, microcrystalline cellulose, croscarmellose sodium, magnesium stearate, and Gastric soluble film coating premix. In some embodiments, the formulation comprises the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof and aminoalkyl methacrylate copolymer, lactose monohydrate, microcrystalline cellulose, croscarmellose sodium, magnesium stearate, and Gastric soluble film coating premix. In some embodiments, the formulation comprises, based on the total weight of the formulation, 20～26wt%of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof, 20～26wt%of aminoalkyl methacrylate copolymer, 18～23wt%of lactose monohydrate, 18～23wt%of microcrystalline cellulose, 3～5wt%of croscarmellose sodium, 0.5～1.5wt%of magnesium stearate and 2～4wt%of Gastric soluble film coating premix.

Preferably, the formulations independently comprise:

about 80-120mg VV116 hydrobromide, about 80-120mg aminoalkyl methacrylate copolymer, about 80-100mg lactose monohydrate, about 80-100mg microcrystalline cellulose, about 14-18mg croscarmellose sodium, about 3-5mg magnesium stearate, and about 10-15mg Gastric soluble film coating premix; or

about 280-320mg VV116 hydrobromide, about 280-320mg aminoalkyl methacrylate copolymer, about 250-300mg lactose monohydrate, about 250-300mg microcrystalline cellulose, about 45-50mg croscarmellose sodium, about 10-15mg magnesium stearate, and about 32-40mg Gastric soluble film coating premix.

More preferably, the formulations independently comprise:

about 100mg VV116 hydrobromide, about 100mg aminoalkyl methacrylate copolymer, about 90mg lactose monohydrate, about 90mg microcrystalline cellulose, about 16mg croscarmellose sodium, about 4mg magnesium stearate, and about 12mg Gastric soluble film coating premix; or

about 300mg VV116 hydrobromide, about 300mg aminoalkyl methacrylate copolymer, about 270mg lactose monohydrate, about 270mg microcrystalline cellulose, about 48mg croscarmellose sodium, about 12mg magnesium stearate, and about 36mg Gastric soluble film coating premix.

Formulations described above are also contemplated in the subject application.

The kit as described herein can be used in any one of the treatment, prevention and alleviation methods and uses as disclosed herein.

In some embodiments, the present application also provides use of the above-mentioned formulations in preparation of a kit for treating or preventing or alleviating a SARS-CoV-2 infection or a condition or symptom associated with the SARS-CoV-2 infection or COVID-19 in a subject in need thereof as disclosed herein. Preferably, the kit thus prepared is as described in any one of the above-mentioned embodiments.

The present invention is further illustrated by the following examples, which should not be construed as limiting the present invention. The contents of all references cited throughout the present application are explicitly incorporated herein by reference.

Examples

Example 1: VV116 vs. nirmatrelvir-ritonavir for mild-to-moderate Covid-19 with high risk of progression: a phase 3, non-inferiority, single blind, randomized study

1 Study design

In this multicenter, randomized, single-blind, parallel-controlled, phase 3 trial, eligible participants were randomly assigned in a 1: 1 ratio to orally receive either VV116 (600 mg on day 1 and 300 mg on day 2 to 5 every 12 hours) or nirmatrelvir-ritonavir (300 mg of nirmatrelvir plus 100 mg of ritonavir for 5 days every 12 hours) using a centralized, interactive web response system. VV116 is manufactured by Vigonvita Life Sciences Co., Ltd. Randomization was performed without stratification as designed by statistician together with the principal investigators. All site investigators, site staff (except for those who performed randomization and administration of the study drug) , and those who were involved in outcome assessments were unaware of the intervention assignments. Participants remained unblinded throughout the study. Data cutoff for the primary analysis occurred on May 13, 2022 when the target number of primary end point events was met (n >724) in the full analysis set. The study design schematic is outlined in FIG. 1.

2 Participants

After written informed consent was obtained from participants or participants-designated decision-maker, participants from 7 national designated Covid-19 treatment hospitals in Shanghai, China were assessed for eligibility between April 4, 2022, and May 2, 2022.

Adults aged 18 years or older were eligible if they had mild or moderate Covid-19 with total symptoms scores ≥2 as determined based on definitions adapted from Food and Drug Administration.

Other key inclusion criteria were 1) SARS-CoV-2 infection confirmed by laboratory reverse-transcriptase-polymerase-chain-reaction (RT-PCR) test within 7 days, or, the onset of signs or symptoms within 5 days prior to the first dose, or, the cycle threshold value ≤20 on RT-PCR test on the day of first dose; and 2) with at least one risk factor for progression to severe illness of Covid-19 (age ≥60 years, cardiovascular disease, hypertension, chronic lung diseases, diabetes, obesity or overweight [BMI>25 kg/m²] , active cancer, chronic kidney disease, current smoker, etc. ) ; and 3) agree to adhere to contraception restrictions.

Key exclusion criteria were 1) confirmed or suspected severe/critical COVID-19 or with an anticipated need for mechanical ventilation prior to randomization; or 2) confirmed eye disease with evidence of inflammation, vessel deformity, retinal hemorrhage or decollement, optic nerve lesion, or fundus lesion) ; or 4) have alanine transaminase or aspartate transaminase >1.5 upper limit of normal; or 5) estimated glomerular filtration rate (eGFR) < 30 ml per minute; or 6) participants who have known allergies to any of the components used in the formulation of the VV116 or nirmatrelvir-ritonavir; or 8) with previous or ongoing therapeutic therapy for COVID-19 including the investigational drugs; or 9) use of contraindicated drugs listed in the package insert of nirmatrelvir-ritonavir. Although nirmatrelvir-ritonavir is not contradicted in participants with eGFR ≥ 30 to < 60 ml per minute, we excluded these participants to avoid overdose exposure in the updated protocol Ver. 3.0 (Version date April 10, 2022) .

3 Assessment

After eligibility assessment including baseline lab results confirmation completed, randomization number with information on drug allocation was assigned by the unblinded site staff. Drug was administrated by unblinded study nurse who was not involved in the evaluation of drug efficacy and safety and was prohibited to discuss the administered drug with the trial investigator and clinical staff. To decrease the possibilities of the recognition of received drug by participants, both study drugs were re-packaged to mask the drug information. Outcome assessors and clinical care givers were unaware of the treatment assignment.

Covid-19 related symptoms and the WHO 11-point Clinical Progression Scale were used for participants’ evaluation and scored by investigators on day 1 prior to the study drug administration followed by daily assessment at approximately the same time every day until the resolution of COVID-19-related target symptoms or day 28, whichever was earlier. Assessment of vital signs, oxygen support, and physical examinations were also performed during the study. SARS-CoV-2 RNA from nasopharyngeal swabs was measured by RT-PCR assay at each site on days 1, 3, 5 (end-of-treatment visit) , 7, 10, 14, 21 and 28. Both qualitative (positive/negative) and quantitative data (cycle threshold value) reported from the RT-PCR assay were collected if available. Other data collection included lab results, patient-reported adverse events, demographics, pre-existing conditions and current medical history, pre-specified medical history for COVID-19, prior SARS-CoV-2 vaccinations, or prophylactic antibodies.

4 End Points

The primary efficacy end point was time from randomization to sustained clinical recovery through day 28. Sustained clinical recovery was defined as the sustained alleviation of all targeted COVID-19 related symptoms/signs score to ≤1 for 2 consecutive days. The first day of the 2 consecutive-day period will be considered the event date. Secondary efficacy end point included percentage of the participants who had progression to severe/critical COVID-19 or death from any cause; changes of WHO progression scale through Day 28; time to sustained resolution of symptoms through Day 28; percentage of participants with negative SARS-CoV-2 at Day 3, 5, 7, 10, 14. Sustained resolution of symptoms was defined as the absence of all targeted COVID-19 related symptoms/signs for 2 consecutive days. The median time from randomization to sustained negative SARS-CoV-2 was initially not planned in the protocol and was added in the statistical analysis plan before database lock. Definition for the time to sustained negative SARS-CoV-2 was the duration from randomization to the first day of the 2 consecutive-day with negative SARS-CoV-2. Safety end points included adverse events and serious adverse events as coded according to the Medical Dictionary for Regulatory Activities (MedDRA) , version 25.0. The severity of AEs was determined in accordance with the NCI CTCAE version v5.0. Any adverse event that emerged from the time the participant provided informed consent through Day 28 was actively recorded and reported for participants who underwent randomization and had received at least one dose of VV116 or nirmatrelvir-ritonavir.

5 Statistical Analysis

The primary efficacy hypothesis was that VV116 would be noninferior to nirmatrelvir-ritonavir with respect to sustained clinical recovery. Due to the lack of data on the time to clinical recovery in participants with Omicron infection treated by nirmatrelvir-ritonavir, the reference duration of 5.5 days was estimated according to the duration of acute symptoms in individuals infected with SARS-CoV-2 during Omicron wave and an overall vaccination rate of more than 90%in the general population in Shanghai. To satisfy the noninferiority hypothesis, the lower boundary of the two-sided 95%confidence interval for the hazard ratio of the primary end point had to be above 0.8. The noninferiority margin corresponds to 6.875 days to sustained clinical recovery, which is 25%longer than 5.5 days. A minimum 724 events are required to ensure a statistical power of 85%.

The noninferiority hypothesis was tested in the Full Analysis Set (FAS) (participants who received at least one pill of VV116 or nirmatrelvir-ritonavir) . Sensitivity analyses in the FAS2 population (participants who commenced trial regimen within 5 days after symptom onset) and per-protocol population (participants without protocol deviation that potentially influenced the efficacy of the trial regimen) were tested. For all the other efficacy analyses, data were analyzed in the FAS population. By the time of data cutoff (May 13, 2022) , 771 were included in the FAS population in which, sustained clinical recovery events occurred in 755 participants. The Kaplan-Meier method was used to estimate the median time to sustained clinical recovery for each group, and the 95%CI was estimated using the BrookMeyer-Crowley method with log-log transformation. For those without efficacy end point events, data were censored on the last day that their status was known.

Subgroup analyses of the primary end point were prespecified to assess the consistency of the intervention effect across the following subgroups: age (< 60 vs. ≥ 60 years) , sex (female vs. male) , SARS-CoV-2 vaccination status (unvaccinated vs. routine vaccine vs. booster vaccine) .

6 Results

6.1 Participants

In the current analysis, of the 997 participants screened from April 4, 2022 through May 2, 2022, 822 participants had been randomized for oral treatment of VV116 (n = 411) or nirmatrelvir-ritonavir (n = 411) . Fifty-one randomized participants did not receive the study (n = 27) or control drugs (n = 24) and were excluded, leaving 771 participants in the FAS population; among them, 755 (97.9%) had achieved sustained clinical recovery and 256 (33.2%) completed follow-up (day 28) , 19 (2.5%) discontinued the trial and the remaining 496 (64.3%) were being actively followed-up.

In the FAS population, baseline demographic and disease characteristics were balanced between treatment groups (Table 1) . The median age was 53 years (IQR, 38-66 years) with half female participants (50.2%) , mostly being Han nationality (99.7%) and mild Covid-19 (92.1%) . A total of 75.7%of participants had received SARS-CoV-2 vaccination, 30.9%and 44.9%had completed routine course or boost shot, respectively. The most common prespecified characteristics and coexisting conditions associated with a risk of progression to severe Covid-19 at baseline were aged 60 years or older (37.7%) , cardiovascular disease or hypertension (35.1%) , a BMI of 25 or above (32.9%) , current smoking (12.5%) , diabetes (10.1%) . Most participants (77.3%) received the oral treatment of VV116 or nirmatrelvir-ritonavir within 5 days of symptom onset and 43.6%within 3 or fewer days. Most participants (80.0%) received VV116 or nirmatrelvir-ritonavir within 5 days of first positive results confirmed by RT-PCR with a median Ct value of 21.7 (IQR, 18.6-25.8) .

Table 1. Demographic and Clinical Characteristics of the Full Analysis Population*

*Participants are those who underwent randomization and received at least one dose of VV116
or Nirmatrelvir-ritonavir. Participants were grouped according to assignments.

IQR denotes interquartile range; RT-PCR, reverse transcriptase-polymerase chain reaction; Covid-19, coronavirus disease 2019; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Race or ethnic group was reported by the patient.
Obesity was defined by a body-mass index of 25 or higher in accordance with WHO criteria
for adult Asians
§ Missing data, invalid samples, tests not done, or results reported as “unknown” were all
categorized as unknown and are not shown individually.

6.2 Primary end point

In the full analysis set, the median time from randomization to sustained clinical recovery was 4 days (95%confidence interval [CI] , 4-5 days) in the VV116 group and 5 days [95%CI, NE-NE days] in the nirmatrelvir-ritonavir group (hazard ratio, 1.17; 95%CI, 1.012 to 1.349; lower boundary, 1.012>1.0; one-sided P=0.039 for superiority) (FIG. 2) . Sensitivity analyses produced similar results showing superiority of VV116 over nirmatrelvir-ritonavir in the per-protocol analysis population (hazard ratio, 1.16; 95%CI, 1.002 to 1.346) and among participants who commenced treatment within 5 days after symptom onset (hazard ratio, 1.20; 95%CI, 1.020 to 1.415) (FIG. 2) .

In most prespecified subgroups, the time to sustained clinical recovery in the VV116 group was noninferior towards superior to the nirmatrelvir-ritonavir group regardless of age, sex, and vaccination status. The point estimate for the difference in the time to sustained clinical recovery indicates uncertainty about VV116 being noninferior to nirmatrelvir-ritonavir only in participants with moderate Covid-19, participants who commenced treatment beyond 5 days of symptom onset or first confirmed date of positive SARS-CoV-2.

6.3 Secondary end points

By the time of data cut-off date on May 13, 2022, no participants in this trial progressed to severe Covid-19 or death. The median time from randomization to resolution of target Covid-19 related symptoms was 7 days (95%CI, 7-8 days) in both the VV116 group and the nirmatrelvir-ritonavir group (hazard ratio, 1.06; 95%CI, 0.914 to 1.225; lower boundary, 0.914>0.8; one-sided P=0.44 for noninferiority) (Table 2) . There was a higher proportion of participants with sustained clinical recovery in the VV116 than in the nirmatrelvir-ritonavir group by each prespecified time point: 5-day (66.4%vs. 57.6%) , 7-day (86.2%vs. 81.7%) , 10-day (94.3%vs. 92.0%) , 14-day (97.4%vs. 96.6%) , 21-day (98.2%vs. 97.4%) , 28-day (98.2%vs. 97.7%) (Table 2) . The median time from randomization to sustained negative SARS-CoV-2 was 7 days (95%CI, 6-7 days) in either group (hazard ratio, 0.98; 95%CI, 0.894 to 1.138; lower boundary, 0.894>0.8; one-sided P=0.82 for noninferiority) . Percentages of participants with negative SARS-CoV-2 were similar between the VV116 and the nirmatrelvir-ritonavir group by 5-day (48.4%vs. 47.3%) , 7-day (75.0%vs. 71.1%) , 10-day (87.8%vs. 89.1%) , 14-day (94.8%vs. 92.5%) after randomization as was the changes of viral Ct values from baseline to day 5, 7, 10 and 14 (Table 2) .

Table 2. Primary and Secondary Efficacy Outcomes (Full Analysis Population) *

This value is a hazard ratio that was calculated by means of a Cox proportional-hazards
model.
SD denotes standard deviation.

6.4 Safety

As compared with nirmatrelvir-ritonavir, participants who received VV116 reported fewer AEs (66.1%vs 77.0%) and grade 3 or 4 AEs (2.3%vs 5.7%) (Table 3) . The percentage of participants with AEs considered by the site investigator to be related to the trial regimen was also lower in the VV116 group than in the nirmatrelvir-ritonavir group (51.0%vs 66.9%) , as was the grade 3 or 4 AEs (1.8%vs 5.2%) . Two serious AEs (one was acute cerebral infarction and the other was the deterioration of the pre-existed interstitial lung disease) were reported from 2 participants in the nirmatrelvir-ritonavir group, but none were considered by the investigators to be related to nirmatrelvir-ritonavir (Table 3) .

The most frequently reported adverse events (occurring in ≥5%of participants in either group) considered by investigators to be related to the trial regimen were dysgeusia (VV116 group: 3.4%vs nirmatrelvir-ritonavir group: 24.8%) , triglyceride increase (10.2%vs 19.1%) , hyperlipemia (2.3%vs 7.2%) , urinary tract infection (6.0%vs 5.7%) , dizzy (4.2%vs 6.2%) , hepatic function abnormal (5.5%vs 2.8%) , bradycardia (5.5%vs 4.1%) ; all these AEs were nonserious and mostly grade 1 or 2.

Table 3. Adverse Events (Safety Population) *

*Participants were those received at least one dose of VV116 or Nirmatrelvir-ritonavir. Participants were grouped
according to actual intervention.
Related events were those determined by the investigators to be related to the assigned regimen.
Severity grades were defined according to the NCI CTCAE version v5.0. For those not listed in NCI-CTCAE v
5.0, severity was determined according to prespecified criteria listed in the protocol.

7 Analysis

In this trial in symptomatic adults hospitalized with mild-to-moderate COVID-19 at high risk for severe disease, a 5-day course of oral treatment with VV116 was superior to nirmatrelvir-ritonavir in shortening the time to sustained clinical recovery. This efficacy superiority was established in the full analysis population, the per-protocol population as well as participants who commenced treatment within 5 days of symptom onset. Prespecified subgroup analyses of the primary end point showed that VV116 was noninferior towards superior to nirmatrelvir-ritonavir, regardless of age, sex, and vaccination status. Statistical testing of the secondary end points showed that VV116 was noninferior to nirmatrelvir-ritonavir regarding the time to sustained symptom resolution and negative SARS-CoV-2. No participants in either group progressed to severe/critical COVID-19 or death. VV116 was well tolerated with a lower incidence of AEs than those of the nirmatrelvir-ritonavir.

As compared to remdesivir or monoclonal antibodies that require injection or infusion in ready-to-access healthcare facilities, oral antivirals like VV116 or nirmatrelvir-ritonavir are more feasible to be administrated in early infection and more likely to be accepted by patients. Such therapies if given promptly would have a broad impact on public health in the pandemic in many ways including the mitigation of the overall hospitalization burden, saving ward resources, facilitating post-exposure prophylaxis, minimizing household transmissions, and reducing post-acute sequelae of COVID-19 in long-term.

Genetic analysis of SARS-CoV-2 viral genomes from the 129 patients in this period showed that all the analyzed samples were clustered into the BA. 2.2 sub-lineage, suggesting that the Omicron variant was most likely responsible for the majority of infections involved in this trial. To our best knowledge, this is the first RCT performed in patients infected with Omicron variant. In this population, the median time to sustained clinical recovery was 4 and 5 days in the VV116 and nirmatrelvir-ritonavir group, respectively. The median time to sustained symptom resolution was 7 days in either group, shorter than those reported from other trials evaluating Ivermectin (10 days) , REGEN-COV antibody (14 days) , and bamlanivimab with or without etesevimab (8 days) . Another unique feature of this trial is the inclusion of 75.7%of participants who had received SARS-CoV-2 vaccines which is the main population in the real world but has been excluded from almost all the other previous trials. The results generated from our trial demonstrated that VV116 was noninferior to nirmatrelvir-ritonavir in participants with or without vaccination and showed superiority in the subgroups who received boosted vaccination. Based on the quick resolution of targeted Covid-19-related symptoms observed in either group, both drugs are likely to have potent efficacy in vaccinated populations.

In this trial, fewer AEs occurred in the VV116 group than in the nirmatrelvir-ritonavir group. This might be related to the fact that unlike nirmatrelvir-ritonavir which requires careful assessment of concomitant medications individually to avoid drug-drug interaction, VV116 does not inhibit or induce major drug metabolizing enzymes and is not an inhibitor of major drug transporters, so interaction with concomitant medications is less likely. Although being transient, dysgeusia was reported in one-quarter of the participants receiving nirmatrelvir-ritonavir in this study, higher than that previously reported in the EPIC-HR trial (5.6%) , thus warranting more attentions in future trials. In addition, incidence of dyslipidemia was relatively high in both nirmatrelvir-ritonavir and VV116 recipients. Although such adverse reaction has been noted in the long-term treatment of ritonavir in human immunodeficiency virus infected patients, the effect of nirmatrelvir or VV116 on lipid metabolism remains further investigations.

Example 2: In Vitro Activity of VV116 and VV116-3 Against SARS-CoV-2 (Omicron variant) in Vero E6 cells

Cell culture medium: Cell growth medium was DMEM medium containing 10%FBS; cell maintenance medium (DMEM maintenance medium) was DMEM medium containing final concentration of 2%FBS.

Sample dilution: DMEM medium containing 10%DMSO, 2%FBS.

2.1 Cell Culture and Treatment

Vero E6 cells (ATCC) were cultured in T75 cell culture flasks in 37℃, 5%CO₂ and passaged at a 1: 3 ratio every 48 hours with Cell growth medium.

The day before the test, the cell culture medium was aspirated, the cells were rinsed with PBS, 2 mL Trypsin solution was added, and digested for 2-3 minutes at room temperature. Culture medium was added to neutralize trypsin and the cells were counted and transferred to a 96-well plate (10,000 cells/well) .

2.2 Sample preparation and dilution of virus

The test compounds VV116, VV116-3 and the positive control compound PF-07321332 (Nirmatrelvir) were dissolved in DMSO to prepare a 10 mM stock solution for storage. First, the 10 mM stock solution of the test compound was diluted 10 times to 1 mM with DMEM medium containing 2%FBS. Then a compound dilution medium (DMEM medium containing 10%DMSO, 2%FBS) was used for three-fold serial dilution, to produce a total of 8 gradients (the initial concentration of the compound is 300 μM) . The experiment was set up in 3 parallels. Meanwhile, in a biological safety cabinet, the SARS-CoV-2 Omicron variant was diluted to 100 TCID50/0.05mL with DMEM maintenance solution.

2.3 Compound inoculated cells

Hank's solution was discarded from cell culture wells, then 45 μl DMEM maintenance medium containing 2%FBS were added to each well, 50 μl diluted virus were added to fix the virus amount in each well of the white 96 well plate to 100 TCID50, and finally 5 μl diluted vv116 /vv116-3 /pf-07321332 compound samples were suck to inoculate on Vero-E6 monolayer cells. Inoculate 3 wells of each dilution of each compound, so that the final volume of each well is 0.1ml, the initial concentration of the test compound and the positive control compound tested in the cultured cells was 15 μ m, and the final concentration of DMSO in each well of cells was 0.5%. Place the white 96 well culture plate inoculated with samples in the 5%CO₂ incubator at 37 ℃. The normal cell control contained no virus and only DMEM maintenance solution. No compound was added to the virus control, and only DMEM maintenance solution containing 100 TCID50 /0.05ml was added.

2.4 Inspection of test results

After the test sample and virus were cultured on the cells for 3 days, the cell viability was detected using the CellTiter Glo Chemiluminescence Live Cell Detection Kit. 50μl of CellTiter-Glo reagent was added to each well of a white 96-well plate, and then the microplate was placed on the plate. Gently shake and mix on a shaker for 2 minutes, incubate at room temperature for 10 minutes, and detect by LumiStation 1800 chemiluminescence microplate reader. For details, refer to the instruction manual of the CellTiter Glo kit.

2.5 Test data analysis

The antiviral activity of the compound is represented by Inhibition %, and the calculation formula is as follows:

Inhibition (%) = (Raw data_cpd -Average_VC) / (Average_CC -Average_VC) *100

wherein Raw data_cpd represents the readings of compound-treated wells; Average_VC and Average_CC represent the mean values of virus control wells (cells infected with virus only, no compound treated) and cell control wells (cells not treated with compound and virus infected) , respectively.

The IC₅₀ value of each compound against SARS-CoV-2 Omicron variant was calculated using Graphpad prism 8.0 software, and the VV116, VV116-3, PF-07321332 concentration values (μM) and inhibition rate (%) were entered into X and in Graphpad prism 8.0 software. In column Y, select the Transform concentration (X) program in the Analyze Data option, select transform to logarithms, and finally select the Log (inhibitor) vs. response-Variable slope (four parameters) program under the Dose-response-Inhibition option to get the corresponding compound pair IC50 value of SARS-CoV-2 Omicron variant.

2.6 SARS-CoV-2 Virus

The SARS-CoV-2 virus is SARS-CoV-2 Omicron variant, the SARS-CoV-2 Omicron variant includes, but is not limited to: AY. 100, AY. 3, AY. 39.1.2, AY. 39.1.4, AY. 43, B. 1, B. 1.1.528, B. 1.1.529, B. 1.617.2, BA. 1, BA. 1.1, BA. 1.1.1, BA. 1.1.10, BA. 1.1.11, BA. 1.1.12, BA. 1.1.13, BA. 1.1.14, BA. 1.1.15, BA. 1.1.16, BA. 1.1.17, BA. 1.1.18, BA. 1.1.2, BA. 1.1.3, BA. 1.1.4, BA. 1.1.5, BA. 1.1.6, BA. 1.1.7, BA. 1.1.8, BA. 1.1.9, BA. 1.10, BA. 1.12, BA. 1.13, BA. 1.13.1, BA. 1.14, BA. 1.14.1, BA. 1.14.2, BA. 1.15, BA. 1.15.1, BA. 1.15.2, BA. 1.15.3, BA. 1.16, BA. 1.16.1, BA. 1.16.2, BA. 1.17, BA. 1.17.1, BA. 1.17.2, BA. 1.18, BA. 1.19, BA. 1.2, BA. 1.20, BA. 1.21, BA. 1.21.1, BA. 1.22, BA. 1.23, BA. 1.24, BA. 1.3, BA. 1.4, BA. 1.5, BA. 1.6, BA. 1.7, BA. 1.8, BA. 1.9, BA. 2, BA. 2.1, BA. 2.10, BA. 2.10.1, BA. 2.10.2, BA. 2.10.3, BA. 2.11, BA. 2.12, BA. 2.12.1, BA. 2.12.2, BA. 2.13, BA. 2.14, BA. 2.15, BA. 2.16, BA. 2.17, BA. 2.18, BA. 2.19, BA. 2.2, BA. 2.2.1, BA. 2.20, BA. 2.21, BA. 2.22, BA. 2.23, BA. 2.23.1, BA. 2.24, BA. 2.25, BA. 2.25.1, BA. 2.26, BA. 2.27, BA. 2.28, BA. 2.29, BA. 2.3, BA. 2.3.1, BA. 2.3.10, BA. 2.3.11, BA. 2.3.12, BA. 2.3.13, BA. 2.3.14, BA. 2.3.15, BA. 2.3.16, BA. 2.3.17, BA. 2.3.18, BA. 2.3.2, BA. 2.3.3, BA. 2.3.4, BA. 2.3.5, BA. 2.3.6, BA. 2.3.7, BA. 2.3.8, BA. 2.3.9, BA. 2.30, BA. 2.31, BA. 2.32, BA. 2.33, BA. 2.34, BA. 2.35, BA. 2.36, BA. 2.37, BA. 2.38, BA. 2.38.1, BA. 2.39, BA. 2.4, BA. 2.40, BA. 2.40.1, BA. 2.41, BA. 2.42, BA. 2.43, BA. 2.44, BA. 2.45, BA. 2.46, BA. 2.47, BA. 2.48, BA. 2.49, BA. 2.5, BA. 2.50, BA. 2.51, BA. 2.52, BA. 2.53, BA. 2.54, BA. 2.55, BA. 2.56, BA. 2.56.1, BA. 2.57, BA. 2.58, BA. 2.59, BA. 2.6, BA. 2.60, BA. 2.61, BA. 2.62, BA. 2.63, BA. 2.64, BA. 2.65, BA. 2.66, BA. 2.67, BA. 2.68, BA. 2.69, BA. 2.7, BA. 2.70, BA. 2.71, BA. 2.72, BA. 2.73, BA. 2.74, BA. 2.75, BA. 2.76, BA. 2.77, BA. 2.78, BA. 2.79, BA. 2.79.1, BA. 2.8, BA. 2.80, BA. 2.81, BA. 2.9, BA. 2.9.1, BA. 2.9.2, BA. 2.9.3, BA. 2.9.4, BA. 2.9.5, BA. 3, BA. 3.1, BA. 4, BA. 4.1, BA. 4.1.1, BA. 4.1.2, BA. 4.1.3, BA. 4.1.4, BA. 4.2, BA. 4.3, BA. 4.4, BA. 4.5, BA. 4.6, BA. 4.7, BA. 5, BA. 5.1, BA. 5.1.1, BA. 5.1.2, BA. 5.1.3, BA. 5.1.4, BA. 5.2, BA. 5.2.1, BA. 5.2.2, BA. 5.2.3, BA. 5.2.4, BA. 5.3, BA. 5.3.1, BA. 5.3.2, BA. 5.3.3, BA. 5.3.4, BA. 5.5, BA. 5.6, BC. 1, BC. 2, BD. 1, BE. 1, BE. 1.1, BE. 2, BE. 3, BF. 1, BF. 1.1, BF. 2, BF. 3, BF. 4, BF. 5, BF. 15, BG. 1, BG. 2, BG. 3, BG. 4, XAA, XAB, XAC, XAD, XAE, XAF, XAG, XAH, XD, XE, XF, XG, XH, XJ, XK, XL, XM, XN, XP, XQ, XR, XS, XT, XU, XV, XW, XY, XZ.

The SARS-CoV-2 Omicron variant (e.g. BA. 5.2) was obtained from the Hubei Provincial Center for Disease Control and Prevention (China) , Item number: YJ20220704-03.

2.7 Study Results

This test evaluated the inhibitory effect of the test compounds VV116 and VV116-3 on the cytopathic effect of SARS-CoV-2 Omicron BA. 5.2 strain at the cellular level. The results showed that the positive control compound (PF-07321332) showed the expected antiviral activity, the antiviral activities of the test compounds VV116 and VV116-3 are shown in Table 4 and Table 5, and the antiviral inhibition rates of the test compounds VV116 and VV116-3 are shown in FIG. 3, and the antiviral inhibition rate of PF-07321332 are shown in FIG. 4.

Table 4. IC₅₀ of Test Compounds and Positive Control Compounds

Table 5. Inhibitory rates of test compounds and positive control compounds against SARS-CoV-2 Omicron BA. 5.2 strain.

2.8 CONCLUSIONS

The test compounds VV116 and VV116-3 can significantly inhibit the cytopathic effect (CPE) of SARS-CoV-2 Omicron BA. 5.2 strain on susceptible cells Vero E6 at the cellular level, and their half-inhibitory concentration IC₅₀ values are respectively were 0.189 ± 0.011 μM and 0.310 ± 0.046 μM.

Claims

A method of treating, preventing, or alleviating a SARS-CoV-2 infection or a condition or symptom associated with the SARS-CoV-2 infection in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof, or a pharmaceutical composition comprising a therapeutically effective amount of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof;

wherein the nucleoside analog is VV116 or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof, and the chemical structural formula of VV116 is as follows:

or the nucleoside analog is VV116-3 or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof, and the chemical structural formula of VV116-3 is as follows:
The method according to claim 1, wherein the condition or symptom of the SARS-CoV-2 infection is fever or chill, cough, shortness of breath or difficult breathing, fatigue, muscle or body aches, headache, new loss of taste or smell, sore throat, congestion or runny nose, nausea or vomiting, or diarrhea; preferably, the condition or symptom of the SARS-CoV-2 infection is fever or chill, cough, shortness of breath, or loss of smell.
A method of treating, preventing, or alleviating COVID-19 in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof, or a pharmaceutical composition comprising a therapeutically effective amount of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof;

wherein the nucleoside analog is VV116 or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof, and the chemical structural formula of VV116 is as follows:

or the nucleoside analog is VV116-3 or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof, and the chemical structural formula of VV116-3 is as follows:
The method according to claim 3, wherein the COVID-19 is mild-to-moderate COVID-19.
The method according to claim 3, wherein the COVID-19 is caused by SARS-CoV-2.
The method according to any one of claims 1-5, wherein the SARS-CoV-2 includes, but is not limited to: SARS-CoV-2 alpha, beta, gamma, delta, lambda, Epsilon, Kappa, Eta, Iota, Theta, Zeta and Omicron variants.
The method according to any one of claims 1-6, wherein the SARS-CoV-2 is a SARS-CoV-2 Omicron variant.
The method according to any one of claims 1-7, wherein the SARS-CoV-2 Omicron variant includes, but is not limited to: AY. 100, AY. 3, AY. 39.1.2, AY. 39.1.4, AY. 43, B. 1, B. 1.1.528, B. 1.1.529, B. 1.617.2, BA. 1, BA. 1.1, BA. 1.1.1, BA. 1.1.10, BA. 1.1.11, BA. 1.1.12, BA. 1.1.13, BA. 1.1.14, BA. 1.1.15, BA. 1.1.16, BA. 1.1.17, BA. 1.1.18, BA. 1.1.2, BA. 1.1.3, BA. 1.1.4, BA. 1.1.5, BA. 1.1.6, BA. 1.1.7, BA. 1.1.8, BA. 1.1.9, BA. 1.10, BA. 1.12, BA. 1.13, BA. 1.13.1, BA. 1.14, BA. 1.14.1, BA. 1.14.2, BA. 1.15, BA. 1.15.1, BA. 1.15.2, BA. 1.15.3, BA. 1.16, BA. 1.16.1, BA. 1.16.2, BA. 1.17, BA. 1.17.1, BA. 1.17.2, BA. 1.18, BA. 1.19, BA. 1.2, BA. 1.20, BA. 1.21, BA. 1.21.1, BA. 1.22, BA. 1.23, BA. 1.24, BA. 1.3, BA. 1.4, BA. 1.5, BA. 1.6, BA. 1.7, BA. 1.8, BA. 1.9, BA. 2, BA. 2.1, BA. 2.10, BA. 2.10.1, BA. 2.10.2, BA. 2.10.3, BA. 2.11, BA. 2.12, BA. 2.12.1, BA. 2.12.2, BA. 2.13, BA. 2.14, BA. 2.15, BA. 2.16, BA. 2.17, BA. 2.18, BA. 2.19, BA. 2.2, BA. 2.2.1, BA. 2.20, BA. 2.21, BA. 2.22, BA. 2.23, BA. 2.23.1, BA. 2.24, BA. 2.25, BA. 2.25.1, BA. 2.26, BA. 2.27, BA. 2.28, BA. 2.29, BA. 2.3, BA. 2.3.1, BA. 2.3.10, BA. 2.3.11, BA. 2.3.12, BA. 2.3.13, BA. 2.3.14, BA. 2.3.15, BA. 2.3.16, BA. 2.3.17, BA. 2.3.18, BA. 2.3.2, BA. 2.3.3, BA. 2.3.4, BA. 2.3.5, BA. 2.3.6, BA. 2.3.7, BA. 2.3.8, BA. 2.3.9, BA. 2.30, BA. 2.31, BA. 2.32, BA. 2.33, BA. 2.34, BA. 2.35, BA. 2.36, BA. 2.37, BA. 2.38, BA. 2.38.1, BA. 2.39, BA. 2.4, BA. 2.40, BA. 2.40.1, BA. 2.41, BA. 2.42, BA. 2.43, BA. 2.44, BA. 2.45, BA. 2.46, BA. 2.47, BA. 2.48, BA. 2.49, BA. 2.5, BA. 2.50, BA. 2.51, BA. 2.52, BA. 2.53, BA. 2.54, BA. 2.55, BA. 2.56, BA. 2.56.1, BA. 2.57, BA. 2.58, BA. 2.59, BA. 2.6, BA. 2.60, BA. 2.61, BA. 2.62, BA. 2.63, BA. 2.64, BA. 2.65, BA. 2.66, BA. 2.67, BA. 2.68, BA. 2.69, BA. 2.7, BA. 2.70, BA. 2.71, BA. 2.72, BA. 2.73, BA. 2.74, BA. 2.75, BA. 2.76, BA. 2.77, BA. 2.78, BA. 2.79, BA. 2.79.1, BA. 2.8, BA. 2.80, BA. 2.81, BA. 2.9, BA. 2.9.1, BA. 2.9.2, BA. 2.9.3, BA. 2.9.4, BA. 2.9.5, BA. 3, BA. 3.1, BA. 4, BA. 4.1, BA. 4.1.1, BA. 4.1.2, BA. 4.1.3, BA. 4.1.4, BA. 4.2, BA. 4.3, BA. 4.4, BA. 4.5, BA. 4.6, BA. 4.7, BA. 5, BA. 5.1, BA. 5.1.1, BA. 5.1.2, BA. 5.1.3, BA. 5.1.4, BA. 5.2, BA. 5.2.1, BA. 5.2.2, BA. 5.2.3, BA. 5.2.4, BA. 5.3, BA. 5.3.1, BA. 5.3.2, BA. 5.3.3, BA. 5.3.4, BA. 5.5, BA. 5.6, BC. 1, BC. 2, BD. 1, BE. 1, BE. 1.1, BE. 2, BE. 3, BF. 1, BF. 1.1, BF. 2, BF. 3, BF. 4, BF. 5, BF. 15, BG. 1, BG. 2, BG. 3, BG. 4, XAA, XAB, XAC, XAD, XAE, XAF, XAG, XAH, XD, XE, XF, XG, XH, XJ, XK, XL, XM, XN, XP, XQ, XR, XS, XT, XU, XV, XW, XY, XZ.
The method according to any one of claims 1-8, wherein the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is administered at a single dose of about 100 to about 800 mg.
The method according to claim 9, wherein the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is administered at a single dose of about 300 to about 600 mg.
The method according to claim 9, wherein the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is administered at a single dose of about 100 mg, about 200 mg, about 300 mg, about 400 mg, about 500 mg, about 600 mg, about 700 mg or about 800 mg.
The method according to any one of claims 1-11, wherein the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is administered at a frequency of about once a day, twice a day, thrice a day, or once every two days.
The method according to any one of claims 1-12, wherein the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is administered for 1-10 days or longer.
The method according to claim 13, wherein the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is administered for 1 day, 3 days, 5 days, 6 days, or 10 days.
The method according to any one of claims 1-14, wherein the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is administered orally on an empty stomach or on a regular diet.
The method according to any one of claims 1-15, wherein the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is administered orally at a single dose of about 600 mg on day 1, once a day, and about 300 mg on day 2 to 5, twice a day.
The method according to any one of claims 1-16, wherein the VV116 or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is in a formulation comprising about 80-120mg VV116 hydrobromide, about 80-120mg aminoalkyl methacrylate copolymer, about 80-100mg lactose monohydrate, about 80-100mg microcrystalline cellulose, about 14-18mg croscarmellose sodium, about 3-5mg magnesium stearate, and about 10-15mg Gastric soluble film coating premix; or about 280-320mg VV116 hydrobromide, about 280-320mg aminoalkyl methacrylate copolymer, about 250-300mg lactose monohydrate, about 250-300mg microcrystalline cellulose, about 45-50mg croscarmellose sodium, about 10-15mg magnesium stearate, and about 32-40mg Gastric soluble film coating premix.
The method according to claim 17, wherein the VV116 or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof is in a tablet comprising about 100mg VV116 hydrobromide, about 100mg aminoalkyl methacrylate copolymer, about 90mg lactose monohydrate, about 90mg microcrystalline cellulose, about 16mg croscarmellose sodium, about 4mg magnesium stearate, and about 12mg Gastric soluble film coating premix; or about 300mg VV116 hydrobromide, about 300mg aminoalkyl methacrylate copolymer, about 270mg lactose monohydrate, about 270mg microcrystalline cellulose, about 48mg croscarmellose sodium, about 12mg magnesium stearate, and about 36mg Gastric soluble film coating premix.
The method according to any one of claims 1-18, wherein the therapeutically effective amount of a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof achieves a median time from randomization to sustained clinical recovery 4 days post administration.
The method according to any one of claims 1-19, wherein lower than about 3%patients exhibit grade 3 or higher treatment emergent adverse event (TEAEs) from the treatment.
The method according to claim 20, wherein lower than about 2.3%patients exhibit grade 3 or higher treatment emergent adverse event (TEAEs) from the treatment.
A kit comprising doses of a nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof for oral administration for at least 5 days; wherein the doses comprise:

a dose for oral administration on day 1 in a frequency of once a day, comprising one or several formulations of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof, wherein the total amount of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof in the dose is in a range of from about 400 mg to about 800 mg; and

doses for oral administration for the rest of the days in a frequency of twice a day, each of which comprises one or several formulations of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof, wherein the total amount of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof in each dose is in a range of from about 80mg to 300mg;

wherein the nucleoside analog is VV116 or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof, and the chemical structural formula of VV116 is as follows:

or the nucleoside analog is VV116-3 or a pharmaceutically acceptable salt, stereoisomer, or tautomer thereof, and the chemical structural formula of VV116-3 is as follows:
The kit according to claim 22, wherein the dose for oral administration on day 1 in a frequency of once a day comprises about 600mg of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof, and each of the doses for oral administration for the rest of the days in a frequency of twice a day comprises about 300mg of the nucleoside analog or a pharmaceutically acceptable salt, stereoisomer or tautomer thereof.
The kit according to claim 22, wherein the formulation comprises:

about 80-120mg VV116 hydrobromide, about 80-120mg aminoalkyl methacrylate copolymer, about 80-100mg lactose monohydrate, about 80-100mg microcrystalline cellulose, about 14-18mg croscarmellose sodium, about 3-5mg magnesium stearate, and about 10-15mg Gastric soluble film coating premix; or

about 280-320mg VV116 hydrobromide, about 280-320mg aminoalkyl methacrylate copolymer, about 250-300mg lactose monohydrate, about 250-300mg microcrystalline cellulose, about 45-50mg croscarmellose sodium, about 10-15mg magnesium stearate, and about 32-40mg Gastric soluble film coating premix; or

about 100mg VV116 hydrobromide, about 100mg aminoalkyl methacrylate copolymer, about 90mg lactose monohydrate, about 90mg microcrystalline cellulose, about 16mg croscarmellose sodium, about 4mg magnesium stearate, and about 12mg Gastric soluble film coating premix; or

about 300mg VV116 hydrobromide, about 300mg aminoalkyl methacrylate copolymer, about 270mg lactose monohydrate, about 270mg microcrystalline cellulose, about 48mg croscarmellose sodium, about 12mg magnesium stearate, and about 36mg Gastric soluble film coating premix.