CN117120057A

CN117120057A - Novel medicine for treating coronavirus infectious disease

Info

Publication number: CN117120057A
Application number: CN202380009597.3A
Authority: CN
Inventors: 清水亮辅; 松尾裕美子; 福原章浩; 深尾圭太; 黑田隆之; 登治谦; 石桥彻
Original assignee: Shionogi and Co Ltd
Current assignee: Shionogi and Co Ltd
Priority date: 2022-01-19
Filing date: 2023-01-18
Publication date: 2023-11-24

Abstract

The present invention provides a pharmaceutical composition comprising a coronavirus 3CL protease inhibitor for the treatment of novel coronavirus infectious disease (covd-19). Provided is a pharmaceutical composition for treating a novel coronavirus infectious disease (COVID-19), which contains a complex comprising a compound represented by formula (I) and fumaric acid as active ingredients.

Description

Novel medicine for treating coronavirus infectious disease

Technical Field

The present invention relates to the treatment of diseases in which coronavirus 3CL protease participates. In particular to a pharmaceutical composition containing coronavirus 3CL protease inhibitor for treating novel coronavirus infectious disease (COVID-19).

Background

After 60 s of the twentieth century until 2018, 6 coronaviruses were reported. Among them, the symptoms caused by 4 kinds (OC 43, 229E, NL and HKU 1) are mild symptoms such as common cold symptoms and gastrointestinal symptoms, but infections caused by severe acute respiratory syndrome coronavirus (severe acute respiratory syndrome coronavirus (SARS-CoV)) occurring in asia and canada in 2002 to 2003 and infections caused by middle east respiratory syndrome coronavirus (Middle East respiratory syndrome coronavirus (MERS-CoV)) occurring in the middle east after 2012 are zoonotic infections, high infectivity and high lethality to humans, and thus, raise serious concerns on public health (non-patent document 1).

The pneumonia of unknown cause is collectively produced in several places, and epidemiological investigation results confirm that the pneumonia is caused by SARS-CoV-2 (non-patent document 1).

SARS-CoV-2 invades host cells by specifically binding spike proteins located on their outer surface to angiotensin converting enzyme 2 (ACE 2) expressed on the cell membrane surface of the host cell targeted. ACE2 is expressed in nasal mucosa epithelial cells, lung cell epithelial cells, small intestine epithelial cells, and the like (non-patent document 2).

The infectious disease caused by SARS-CoV-2 is named as COVID-19, and the World Health Organization (WHO) announced "International sudden Public Health Event of Interest (PHEIC)" about COVID-19 at 30 months of 2020. Then, according to the world infection and the degree of illness, the use of covd-19 as a pandemic (world-wide pandemic) was declared on day 11, 3 in 2020 (non-patent document 3). It was originally thought that SARS-CoV-2 infects humans only from animals, but it is becoming clear that not only infects humans from symptomatic infectors, but also infectors who are asymptomatic and not ill (non-patent document 1).

Among the main symptoms of covd-19, there are many symptoms similar to cold, such as fever, cough, runny nose, nasal obstruction, sore throat, muscle pain, diarrhea, and the like, and symptoms such as disappearance of taste and smell are reported. The young people have a low serious proportion, but elderly people and high-risk patients suffering from cardiovascular diseases, respiratory diseases, kidney diseases, diabetes, obesity, immunodeficiency and the like are liable to be serious, pneumonia rapidly progresses, and fall into shortness of breath, dyspnea and the like, oxygen inhalation is sometimes required, and further treatment by artificial respirators and extracorporeal membrane type artificial lung (ECMO) is required. Thromboembolism is also a characteristic of severe cases caused by COVID-19, and has a high mortality rate. Therefore, the medical system is squeezed and crashed in various countries worldwide (non-patent document 1).

The cumulative number of infectious agents (death number) is 1,700,000 cases (18,000 cases) or more in japan and 276,000,000 cases (5,300,000 cases) or more in the world, and is extended to 223 countries and regions in 2021, 12 months. The number of infected countries has been reported to rise to 60 countries in excess of 30 tens of thousands (non-patent document 4).

At month 12 of 2021, vaccination was advancing worldwide, but spike protein variants were also found to cause an increase in infectivity and pathogenicity, and immune escape, and may become a problem in the future. As covd-19 therapeutic agents, in japan, adefovir as a direct acting antiviral agent, desquamation Mi Songji baratinib as an immunomodulator immunosuppressant, casirizumab (casiriumab) and idevidimab (imdevimab), and sotovimumab (sotrovimab), mo Nupi lasvir (Molnupiravir) as an RNA protease inhibitor, are approved, but it is not possible to achieve the object of terminating infection expansion, and development of new therapeutic agents is desired.

At month 2021, at the website of the feijie company, it was reported that in high risk patients for adults, PAXLOVID (TM) (PF-07321332; ritonavir) reduced the risk of hospitalization or death by 89% compared to placebo (non-patent document 5).

Furthermore, in 2021, month 12, PAXLOVID (TM) has obtained emergency use permission in the united states, and in 2022, month 2, 10, PAXLOVID (registered trademark) combination package has obtained special approval in japan.

The structural formula of PAXLOVID (TM) (PF-07321332) is shown below, and the chemical structure is different from that of the compound of the present invention (non-patent document 6).

[ chemical formula 1]

The non-clinical test result of S-217622 targeting 3CL protease and its chemical structural formula are published on day 11 and 29 of 2021. In addition, it was disclosed that a single dose escalation test for healthy adults in japan confirmed good safety and tolerability. However, there is no disclosure about the therapeutic effect and the dosage for use of the novel coronavirus infectious disease (covd-19) infected person (non-patent document 7).

Prior art literature

Non-patent literature

Non-patent document 1: j Clin Med 2020;9:1225

Non-patent document 2: j Pathol 2004;203:631-7

Non-patent document 3: WHO Health organization.WHO direct-General's opening remarks at the media briefing on COVID-19.11March 2020 [2021, 12 th month 19 th day inspection ], website < URL https:// www.who.int/dg/speches/deltatail/who-direct-General-s-open g-remarks-at-the-medium-briefing-on-covid-19- - -11-march-2020 >

Non-patent document 4: world Health organization.Coronavir Disease (COVID-19) Dashboard.5October 2020 [ 12 months 19 days check out 2022 ], website < https:// COVID 19.who.int/>

Non-patent document 5: "Pfizer's Novel COVID-19Oral Antiviral Treatment Candidate Reduced Risk Of Hospitalization Or Death By 89%In Interim Analysis Of Phase 2/3EPIC-HR Study", [ online ], 5 th day of 2021, pfizer Press Release th day of 2022, 9 th day of 21 st inspection, website < URL https:// www.pfizer.com/news/press-release-detail/Pfizer-Novel-COVID-19-oral-anti-treatment-truck >)

Non-patent document 6:261st Am Chem Soc (ACS) Natl Meet 2021-04-05/2021-04-16. Virtual, N/A. Abst 243

Non-patent document 7: AIMECS2021 (AFMC International Medicinal Chemistry Symposium 2021), online symposium (2021), 11/29/12/2/2021

Disclosure of Invention

Problems to be solved by the invention

The present invention provides a pharmaceutical composition for treating a novel coronavirus infectious disease (covd-19) which has excellent coronavirus 3CL protease inhibitory activity.

Means for solving the problems

The present inventors have conducted studies to solve the above problems, and as a result, have found that the coronavirus 3CL protease inhibitor described in non-patent document 7 is useful for the treatment of novel coronavirus infectious disease (covd-19) by administering to a human in a specific administration amount, thereby maintaining an effective blood level without exceeding NOAEL (No observable adverse effect level (without an observable adverse effect level); non-toxic amount), and thus have completed the present application.

The present application relates to the following (1) to (36).

(1) A pharmaceutical composition for treating novel coronavirus infectious disease (COVID-19) comprising a complex comprising a compound represented by formula (I) and fumaric acid as active ingredients.

[ chemical formula 2]

(2) The pharmaceutical composition according to item (1) above, wherein the patient suffering from novel coronavirus infectious disease (covd-19) is classified as an asymptomatic patient, a mild patient or a moderate patient.

(3) The pharmaceutical composition according to item (1) or (2) above, which is used for shortening the course of a novel coronavirus infectious disease (COVID-19).

(4) The pharmaceutical composition according to any one of the above items (1) to (3), which is used for inhibiting the viral proliferation of SARS-CoV-2.

(5) The pharmaceutical composition according to any one of the items (1) to (4), wherein the amount of the active ingredient administered per 1 day of the pharmaceutical composition is 20mg to 2000mg.

(6) The pharmaceutical composition according to any one of items (1) to (5) above, wherein the amount of the active ingredient administered on the treatment day 1 is 375mg, and the amounts of the active ingredient administered on the treatment day two to the treatment day five are 125mg, respectively.

(7) The pharmaceutical composition according to any one of items (1) to (5) above, wherein the amount of the active ingredient administered on the treatment day 1 is 750mg, and the amounts of the active ingredient administered on the treatment day two to the treatment day five are 250mg, respectively.

(8) The pharmaceutical composition according to any one of the above items (1) to (7), wherein the composition is administered once a day.

(9) The pharmaceutical composition according to any one of the items (1) to (8), which is an oral preparation.

(10) A method of treating a novel coronavirus infectious disease (covd-19), comprising the steps of: an effective amount of a complex comprising a compound represented by formula (I) and fumaric acid as an active ingredient is administered to a subject in need of treatment for a novel coronavirus infectious disease (COVID-19).

[ chemical formula 3]

(11) The method of treating according to item (10) above, wherein the patient suffering from novel coronavirus infectious disease (covd-19) is classified as an asymptomatic patient, a mild patient or a moderate patient.

(12) The method of treating according to item (10) or (11) above, which is used for shortening the course of a novel coronavirus infectious disease (COVID-19).

(13) The therapeutic method according to any one of the above items (10) to (12), which is used for inhibiting the viral proliferation of SARS-CoV-2.

(14) The method of treatment according to any one of the above items (10) to (13), wherein the amount of the active ingredient administered per 1 day is 20mg to 2000mg.

(15) The treatment method according to any one of the above items (10) to (14), wherein the amount of the active ingredient administered on the treatment day 1 is 375mg, and the amounts of the active ingredient administered on the treatment day two to the treatment day five are 125mg, respectively.

(16) The therapeutic method according to any one of the above items (10) to (14), wherein the amount of the active ingredient administered on the treatment day 1 is 750mg, and the amounts of the active ingredient administered on the treatment day two to the treatment day five are 250mg, respectively.

(17) The therapeutic method according to any one of the above items (10) to (16), wherein the administration is performed once a day.

(18) The therapeutic method according to any one of the above items (10) to (17), which is an oral preparation.

(19) Use of a complex comprising a compound represented by formula (I) and fumaric acid as active ingredients in the manufacture of a medicament for the treatment of novel coronavirus infectious disease (covd-19).

[ chemical formula 4]

(20) The use according to item (19) above, wherein the patient suffering from novel coronavirus infectious disease (covd-19) is classified as an asymptomatic patient, a mild patient or a moderate patient.

(21) The use as set forth in item (19) or (20) above, for shortening the course of a novel coronavirus infectious disease (COVID-19).

(22) The use according to any one of the above items (19) to (21), for inhibiting the viral proliferation of SARS-CoV-2.

(23) The use according to any one of the above items (19) to (22), wherein the amount of the active ingredient administered per 1 day is 20mg to 2000mg.

(24) The use according to any one of the above items (19) to (23), wherein the amount of the active ingredient administered on the treatment day 1 is 375mg, and the amounts of the active ingredient administered on the treatment day two to the treatment day five are 125mg, respectively.

(25) The use according to any one of the above items (19) to (23), wherein the amount of the active ingredient administered on the treatment day 1 is 750mg, and the amounts of the active ingredient administered on the treatment day two to the treatment day five are 250mg, respectively.

(26) The use according to any one of the above items (19) to (25), wherein the administration is once a day.

(27) The use according to any one of the above items (19) to (26), which is an oral preparation.

(28) A complex comprising a compound represented by formula (I) and fumaric acid as active ingredients for use in the treatment of novel coronavirus infectious disease (covd-19).

[ chemical formula 5]

(29) The complex comprising a compound and fumaric acid according to item (28) above, wherein the patient suffering from novel coronavirus infectious disease (covd-19) is classified as an asymptomatic patient, a mild patient or a moderate patient I.

(30) The complex comprising a compound and fumaric acid as described in the above item (28) or (29), which is used for shortening the course of a novel coronavirus infectious disease (COVID-19).

(31) The complex comprising a compound and fumaric acid according to any one of items (28) to (30) above, which is useful for inhibiting the viral proliferation of SARS-CoV-2.

(32) The complex comprising a compound and fumaric acid according to any one of items (28) to (31), wherein the amount of the active ingredient administered per 1 day is 20mg to 2000mg.

(33) The complex comprising a compound and fumaric acid according to any one of items (28) to (32) above, wherein the amount of the active ingredient administered on treatment day 1 is 375mg and the amounts of the active ingredient administered on treatment days two to five are 125mg, respectively.

(34) The complex comprising a compound and fumaric acid according to any one of items (28) to (32) above, wherein the amount of the active ingredient administered on treatment day 1 is 750mg, and the amounts of the active ingredient administered on treatment days two to five are 250mg, respectively.

(35) The complex comprising a compound and fumaric acid according to any one of items (28) to (34) above, wherein the complex is administered once a day.

(36) The complex comprising a compound and fumaric acid according to any one of items (28) to (35) above, which is an oral preparation.

The present invention also relates to the following (1 ') to (10').

(1') the complex comprising a compound and fumaric acid according to any one of items (28) to (32), wherein the amount of the active ingredient administered on treatment day 1 is 375mg and the amounts of the active ingredient administered on treatment days two to five are 125mg, respectively.

(2') the complex comprising a compound and fumaric acid according to any one of items (28) to (32), wherein the amount of the active ingredient to be administered on the treatment day 1 is 250mg, and the amounts of the active ingredient to be administered on the treatment day two to the treatment day five are 125mg, respectively.

(3') the complex comprising a compound and fumaric acid according to any one of items (28) to (32), wherein the amount of the active ingredient to be administered on the treatment day 1 is 250mg, and the amounts of the active ingredient to be administered on the treatment day two to the treatment day five are 100mg, respectively.

(4') the complex comprising a compound and fumaric acid according to any one of items (28) to (32), wherein the amount of the active ingredient to be administered on the treatment day 1 is 125mg, and the amounts of the active ingredient to be administered on the treatment day two to the treatment day five are 75mg, respectively.

(5') the complex comprising a compound and fumaric acid according to any one of items (28) to (32), wherein the amount of the active ingredient to be administered on treatment day 1 is 150mg, and the amounts of the active ingredient to be administered on treatment days two to five are 75mg, respectively.

(6') the complex comprising a compound and fumaric acid according to any one of items (28) to (32), wherein the amount of the active ingredient to be administered on treatment day 1 is 75mg, and the amounts of the active ingredient to be administered on treatment days two to five are 50mg, respectively.

(7') the complex comprising a compound and fumaric acid according to any one of items (28) to (32), wherein the amount of the active ingredient to be administered on the treatment day 1 is 100mg, and the amounts of the active ingredient to be administered on the treatment day two to the treatment day five are 50mg, respectively.

(8') the complex comprising a compound and fumaric acid according to any one of items (28) to (32), wherein the amount of the active ingredient to be administered on the treatment day 1 is 6mg/kg, and the amounts of the active ingredient to be administered on the treatment day two to the treatment day five are 3mg/kg, respectively.

(9 ') the complex comprising a compound and fumaric acid according to any one of the above items (1 ') to (8 '), wherein administration is performed once a day.

(10 ') the complex comprising a compound and fumaric acid according to any one of items (1 ') to (9 ') above, which is an oral preparation.

ADVANTAGEOUS EFFECTS OF INVENTION

The complex comprising the compound represented by the formula (I) and fumaric acid contained in the pharmaceutical composition of the present invention exhibits an excellent effect of being effective for the treatment of a novel coronavirus infectious disease (COVID-19). The complex containing the compound represented by the formula (I) and fumaric acid contained in the pharmaceutical composition of the present invention is a highly safe pharmaceutical agent which can be administered to a human in a specific dose to maintain an effective blood level without exceeding NOAEL.

Drawings

FIG. 1 shows a powder X-ray diffraction pattern of fumaric acid co-crystal Form I crystals (Form I) of the compound represented by formula (I). The horizontal axis represents 2θ (°), and the vertical axis represents intensity (Count).

FIG. 2 is a diagram showing the structure of an asymmetric unit of a fumaric acid co-crystal Form I crystal (Form I) of the compound of formula (I).

FIG. 3 shows a simulation of the change in plasma drug concentration at various doses. The vertical axis represents the drug concentration in the simulated plasma. The horizontal axis represents the elapsed time after the first administration of the drug.

[ FIG. 4 ] ]Light/medium symptoms representing the 2a Phase fraction (Phase 2a Part) and asymptomatic SARS-CoV-2 infected patients (Modified intention-to-treatment of the intended treatment population) [ mITT ]]Human population) at various time points. The vertical axis represents the amount of change (log) in viral potency from baseline ₁₀ (TCID ₅₀ /mL)), the horizontal axis represents the evaluation time point.

FIG. 5 shows the time from the first confirmation of SARS-CoV-2 virus titer negative in the mild/moderate and asymptomatic SARS-CoV-2 infected persons (mITT population) in the 2a stage fraction (Phase 2a Part). The vertical axis represents the ratio (unit:%) of those with negative SARS-CoV-2 virus titer. The horizontal axis represents time (unit: hours) from the start of treatment.

FIG. 6 shows the amount of change in the total score of 12 symptoms of COVID-19 from baseline at each time point in the light/medium SARS-CoV-2 infected person (treatment-to-treatment population) [ ITT ] population in the 2a Phase section (Phase 2a Part). The vertical axis represents the amount of change from baseline in the total score of 12 symptoms of covd-19. The horizontal axis represents the number of days from the start of administration.

FIG. 7 shows the time until recovery of 5 symptoms of COVID-19 in the mild/moderate SARS-CoV-2 infected persons (persons less than 72 hours from the onset of COVID-19 in the ITT group) in the 3-stage fraction (Phase 3 Part). The vertical axis represents the proportion (unit:%) of subjects recovering from 5 symptoms of covd-19. The horizontal axis represents time (unit: hours) from the start of treatment.

FIG. 8 shows the time until recovery of 5 symptoms of COVID-19 in the mild/moderate SARS-CoV-2 infected person (ITT population) at stage 3 (Phase 3 Part). The vertical axis represents the proportion (unit:%) of subjects recovering from 5 symptoms of covd-19. The horizontal axis represents time (unit: hours) from the start of treatment.

FIG. 9 shows the time until the first confirmation of virus titer negativity in the febrile/moderate SARS-CoV-2 infected person (the group of mITT groups whose time from onset of COVID-19 to random grouping is less than 72 hours) at stage 3 (Phase 3 Part). The vertical axis represents the ratio (unit:%) of those with negative SARS-CoV-2 virus titer. The horizontal axis represents time (unit: hours) from the start of treatment.

FIG. 10 shows the time until the first confirmation of virus titer negative in the febrile/moderate SARS-CoV-2 infected person (mITT population) in the 3-stage fraction (Phase 3 Part). The vertical axis represents the ratio (unit:%) of those with negative SARS-CoV-2 virus titer. The horizontal axis represents time (unit: hours) from the start of treatment.

FIG. 11 shows the time until recovery of 12 symptoms of COVID-19 in the mild/moderate SARS-CoV-2 infected persons (persons less than 72 hours from the onset of COVID-19 in the ITT group) in the 3-stage fraction (Phase 3 Part). The vertical axis represents the proportion (unit:%) of subjects recovering from 12 symptoms of covd-19. The horizontal axis represents time (unit: hours) from the start of treatment.

FIG. 12 shows the time until recovery of 14 symptoms of COVID-19 in the mild/moderate SARS-CoV-2 infected persons (persons less than 72 hours from the onset of COVID-19 in the ITT group) in the 3-stage fraction (Phase 3 Part). The vertical axis represents the proportion of subjects recovering from 14 symptoms of COVID-19 (unit:%). The horizontal axis represents time (unit: hours) from the start of treatment.

Detailed Description

The term "consisting of … …" means having only the constituent elements. The term "comprising" means not limited to the constituent elements, but not excluding the elements not described.

Hereinafter, the present invention will be described with reference to the embodiments. Throughout this specification, unless specifically stated otherwise, the expression in the singular is to be understood as also encompassing the plural concepts thereof. Accordingly, unless specifically indicated otherwise, singular forms (e.g., "a," "an," "the," etc., in the context of English) are to be construed to also include plural forms of the concepts thereof.

In addition, in the present specification, unless specifically stated otherwise, terms used should be understood to be used in the meaning commonly used in the art. Accordingly, unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. In case of conflict, the present specification, including definitions, will control.

The pharmaceutical composition for treating a novel coronavirus infectious disease (covd-19) of the present invention is characterized by comprising a complex comprising a compound represented by the formula (I) and fumaric acid as active ingredients.

[ chemical formula 6]

In the present specification, the pharmaceutical composition for treating a novel coronavirus infectious disease (covd-19) according to the present invention is sometimes referred to as a novel coronavirus infectious disease (covd-19) therapeutic agent.

The compound represented by the formula (I) is (6E) -6- [ (6-chloro-2-methyl-2H-indazol-5-yl) imino ] -3- [ (1-methyl-1H-1, 2, 4-triazol-3-yl) methyl ] -1- [ (2, 4, 5-trifluorophenyl) methyl ] -1,3, 5-triazin-E (triazinane) -2,4-dione ((6E) -6- [ (6-chloro-2-methyl-2H-indazol-5-yl) imino ] -3- [ (1-methyl-1H-1, 2, 4-triazol-3-yl) methyl ] -1- [ (2, 4, 5-trifluorophenyl) methyl ] -1,3,5-triazinane-2,4-dione, and has the coronavirus 3CL protease inhibitory activity. In addition, the compound represented by the formula (I) includes the following tautomers.

[ chemical formula 7]

Complexes comprising a compound represented by formula (I) and fumaric acid may sometimes form solvates (e.g., hydrates, etc.), co-crystals, and/or clathrates, and these are referred to as "complexes" in this specification.

As used herein, "co-crystal" refers to a regular arrangement of counter molecules within the same lattice, and may include any number of counter molecules. The co-crystal means: the intermolecular interaction between the compound and the counter molecule is a substance that chemically interacts non-covalently and non-ionically, such as hydrogen bonds and van der waals forces.

For example, as a co-crystal of the compound represented by the formula (I), a compound represented by the formula (I) and a counter molecule may be included in any number of counter molecules. Preferably consisting of a compound represented by formula (I) and fumaric acid, may comprise any number of fumaric acids. Further preferred are compounds represented by formula (I) and fumaric acid are 1:1 in molar ratio.

The co-crystal is distinguished from salt in the following ways: the compound remains essentially charge-free or neutral.

The co-crystal is distinguished from the hydrate or solvate in the following ways: the counter molecule is not water or solvent.

As used herein, "crystal" refers to a solid in which constituent atoms, ions, molecules, and the like are regularly arranged in three dimensions, and is distinguished from an amorphous solid that does not have such a regular internal structure. The crystals of the present invention may be single crystal, bicrystal, polycrystal, etc.

In addition, "crystals" sometimes exist in "polymorphs" which are identical in composition but differ in arrangement in the crystals, and they are collectively referred to as "crystal forms".

The crystal form and crystallinity can be measured by various techniques including powder X-ray diffraction measurement, raman spectroscopy, infrared absorption spectrometry, moisture absorption/desorption measurement, differential scanning calorimetry measurement, and dissolution characteristics, for example.

In addition, a "polymorph" may be formed by recrystallizing a complex comprising the compound represented by formula (I) and fumaric acid.

In the preparation of the present invention, various salts, complexes (hydrates, solvates, co-crystals, clathrates) and polymorphs thereof may be used, and a mixture of two or more of them may be used.

(powder X-ray diffraction (XRPD))

Powder X-ray diffraction (XRPD) is one of the most sensitive analytical methods for determining the crystalline form and crystallinity of solids. When the crystal is irradiated with X-rays, reflection occurs at the lattice plane, and mutual interference occurs, and ordered diffraction lines corresponding to the period of the structure are displayed. On the other hand, amorphous solids generally do not have an ordered repetition period in their structure, and therefore, do not undergo diffraction phenomena, exhibiting a broad XRPD pattern without features (also known as a halo pattern).

The crystalline form of the compound represented by formula (I) can be identified by a powder X-ray diffraction pattern and a characteristic diffraction peak. The crystalline form of the compound represented by formula (I) can be distinguished from other crystalline forms by the presence of characteristic diffraction peaks.

The characteristic diffraction peak used in the present specification is a peak selected from the observed diffraction patterns. The characteristic diffraction peaks are preferably selected from about 10, more preferably about 5, and even more preferably about 3 of the diffraction patterns.

In distinguishing a plurality of crystals, a peak that is confirmed in the crystal and is not confirmed in other crystals becomes a characteristic peak preferable for determining the crystal, compared with the intensity of the peak. If such characteristic peaks are present, one or both peaks may also characterize the crystal. Comparing the measured images, it can be said that the powder X-ray diffraction patterns are substantially identical if the characteristic peaks are identical.

In general, since an error may occur in the diffraction angle (2θ) in powder X-ray diffraction in the range of ±0.2°, it is to be understood that the value of the diffraction angle in powder X-ray diffraction also includes a value in the range of about ±0.2°. Therefore, not only crystals in which diffraction angles of peaks in powder X-ray diffraction are completely uniform, but also crystals in which diffraction angles of peaks are uniform with an error of about ±0.2° are included in the present invention.

It is known that the intensities of the peaks shown in the following tables and figures may generally vary depending on various factors, such as the effect of the preferred orientation of the crystal with respect to the X-ray beam, the influence of coarse particles, the purity of the substance being analyzed, or the crystallinity of the sample. In addition, the peak position may be shifted based on the variation in the sample height. In addition, when measurements are made using different wavelengths, different offsets can be obtained according to the bragg equation (nλ=2dsinθ), but other XRPD patterns obtained by using other wavelengths are also included within the scope of the present invention.

(analysis of Single Crystal Structure)

By one of the methods for specifying a crystal, parameters of crystallography, atomic coordinates (values indicating spatial positional relationships of atoms) and a three-dimensional structure model in the crystal can be obtained. Reference is made to sakura's guidelines for X-ray structural analysis "skirt chinese house release (1983), stout & Jensen, X-Ray Structure Determination: a Practical Guide, macmillan co., new York (1968), etc. Single crystal structure analysis is useful in identifying the structure of crystals of such complexes, salts, optical isomers, tautomers, geometric isomers of the present invention.

The compound represented by the formula (I) has a coronavirus 3CL protease inhibitory activity and is therefore useful as a therapeutic and/or prophylactic agent for diseases in which coronavirus 3CL proteases are involved. In the present invention, the term "therapeutic agent and/or prophylactic agent" also includes a symptom-improving agent. The disease in which coronavirus 3CL protease participates may be a viral infectious disease, and preferably a coronavirus infectious disease.

As one embodiment, coronaviruses that infect humans are exemplified. As coronaviruses infecting humans, HCoV-229E, HCoV-NL63, HCoV-HKU1, HCoV-OC43, SARS-CoV, MERS-CoV, and/or SARS-CoV-2 can be mentioned.

In one embodiment, the coronavirus may be an α coronavirus and/or a β coronavirus, more preferably a β coronavirus, and still more preferably sarbecovirus.

As one embodiment, examples of the alpha coronavirus include HCoV-229E and HCoV-NL63. HCoV-229E is particularly preferred.

As one embodiment, examples of the beta coronavirus include HCoV-HKU1, HCoV-OC43, SARS-CoV, MERS-CoV, and/or SARS-CoV-2. Preferably, HCoV-OC43 or SARS-CoV-2, and particularly preferably SARS-CoV-2.

As one embodiment, as the beta coronavirus, there may be mentioned beta coronavirus lineage A (beta-coronavirus lineage A), beta coronavirus lineage B (beta-coronavirus lineage B), and beta coronavirus lineage C (beta-coronavirus lineage C). More preferably, the beta coronavirus lineage A (beta-coronavirus lineage A) and the beta coronavirus lineage B (beta-coronavirus lineage B), and particularly preferably, the beta coronavirus lineage B (beta-coronavirus lineage B) are exemplified.

As the beta coronavirus lineage A (. Beta. -coronavirus lineage A), for example, HCoV-HKU1 and HCoV-OC43, preferably HCoV-OC43, can be mentioned. As the beta coronavirus spectrum B (. Beta. -coronavirus lineage B), for example, SARS-CoV and SARS-CoV-2, preferably SARS-CoV-2, can be mentioned. As the beta coronavirus lineage C (. Beta. -coronavirus lineage C), MERS-CoV is preferably mentioned.

As one embodiment, the coronavirus may be HCoV-229E, HCoV-OC43 and/or SARS-CoV-2, and particularly preferably SARS-CoV-2.

Examples of infectious diseases caused by coronaviruses include HCoV-229E, HCoV-NL63, HCoV-OC43, HCoV-HKU1, SARS-CoV, MERS-CoV, and/or SARS-CoV-2. Preferably, the infectious disease is caused by HCoV-229E, HCoV-OC43 and/or SARS-CoV-2, and particularly preferably, the infectious disease is caused by SARS-CoV-2.

As coronavirus infectious diseases, a novel coronavirus infectious disease (COVID-19) is particularly preferred.

The classification of the severity of a disease in a patient infected with a novel coronavirus (a patient) is exemplified below. ( Reference is made to: new type coronavirus infectious disease COVID-19 diagnosis and treatment guide 6.0 edition (Japanese Ministry of Thick students) )

(light symptoms)

The oxygen saturation is 96% or more. The clinical state is: no symptoms of respiratory organs, or cough alone, no dyspnea, and in any case no manifestations of pneumonia were found.

(middle-aged condition I)

The oxygen saturation is less than 96% -more than 93%. Dyspnea and pneumonia are manifested.

(middle disease II)

The oxygen saturation is 93% or less. There is a hypopnea requiring oxygen administration.

(severe cases)

Into the ICU or requires a respirator.

The above classification is a definition of the severity classification in japan, and for example, the severity classification in NIH in china and usa can be cited.

In addition, the asymptomatic SARS-CoV-2 infected person (asymptomatic person) refers to an asymptomatic pathogen carrier. For example, 14 kinds of people whose symptoms of COVID-19 are not recognized can be mentioned.

( 14 symptoms of covd-19: debilitation (fatigue), muscle pain or body pain, headache, shivering/sweating, fever or fever, abnormal taste, abnormal smell, runny nose or stuffy nose, sore throat, cough, shortness of breath (dyspnea), nausea, vomiting, diarrhea )

In the present specification, the 12 symptoms of covd-19 include debilitation (fatigue), muscle pain or body pain, headache, shivering/sweating, feeling fever or fever, runny nose or nasal obstruction, sore throat, cough, shortness of breath (dyspnea), nausea, vomiting, and diarrhea. The 5 symptoms of covd-19 include runny nose, nasal obstruction, sore throat, cough, fever, and weakness (tiredness).

In the present specification, the term "inhibiting severe cases" means inhibiting the increase in the asymptomatic SARS-CoV-2 infection to a degree that the disease is mild and severe, and is classified as mild, moderate, or severe.

In the present specification, the term "inhibiting severe cases" means inhibiting the increase in the severity of an asymptomatic SARS-CoV-2 infection or a mild SARS-CoV-2 infection, which is classified as a moderate disease I, a moderate disease II or a severe disease.

In the present specification, the term "inhibiting severe cases" means that the cases of asymptomatic SARS-CoV-2 infection, mild cases of SARS-CoV-2 infection, or severe cases of SARS-CoV-2 infection of moderate disease I are elevated to such an extent that they are classified as moderate disease II or severe.

In the present specification, inhibiting severe cases means inhibiting the increase of asymptomatic SARS-CoV-2 infected persons, mild SARS-CoV-2 infected persons, moderate SARS-CoV-2 infected persons or moderate SARS-CoV-2 infected persons of moderate symptoms II to a degree that is classified as severe.

In the present specification, the term "inhibiting severe infection" means that the risk of hospitalization and death of a patient suffering from SARS-CoV-2 infection is reduced by the virus growth inhibitory effect of the agent of the present invention.

In the present specification, the term "inhibiting severe infection" means reducing inflammation in the lung of a patient suffering from SARS-CoV-2 infection by the virus growth inhibitory effect of the agent of the present invention.

In the present specification, the term "inhibiting severe infection" means inhibiting pneumonia caused by a viral infection of SARS-CoV-2 by the virus growth inhibitory effect of the agent of the present invention.

In the present specification, the term "inhibiting the occurrence of severe disease" means that the viral growth inhibitory effect of the agent of the present invention suppresses an excessive immune response of a host caused by viral infection of SARS-CoV-2.

In the present specification, the term "inhibiting severe disease" means lowering the rate of deterioration of the order Scale (order Scale) which classifies the severity of clinical illness into 8 orders. Specifically, it means that the first inhibition of the deterioration of the order scale to 3 or more after the start of the treatment.

In the present specification, the term "reduction of the course of a disease" means shortening the time until the symptoms recover or the recovery of the symptoms, and includes an improvement in the total score of the symptoms (14 symptoms) of covd-19 indirectly.

In the present specification, the term "reduction of the course of a disease" means shortening the time until the symptoms recover or the recovery of the symptoms, and includes an improvement in the total score of the symptoms (12 symptoms) of covd-19 indirectly.

In the present specification, the term "reduction of the course of disease" means shortening the time until the symptoms recover or the recovery of symptoms, and includes an improvement in the total score of the symptoms (5 symptoms) of covd-19 indirectly.

As one embodiment, the pharmaceutical composition of the present invention is for administration to an infected person having at least one risk factor for severe symptoms of covd-19.

One embodiment of the present invention is to administer the pharmaceutical composition of the present invention to an infected patient having at least one of the following severe risk factors among SARS-CoV-2 infected patients.

Over 50 years old

Obesity (BMI of 30 kg/m) ² Above, the method comprises

Cardiovascular diseases (cardiovascular system diseases including hypertension and congenital heart disease)

Chronic lung disease (including asthma, interstitial lung disease)

Type 1 or type 2 diabetes

Chronic kidney disease (including dialysis patients)

Chronic liver disease

Immunosuppressed state (e.g., malignancy treatment, bone marrow or organ transplantation, immunodeficiency, poorly controlled HIV, AIDS, sickle cell anemia, thalassemia, long-term administration of immunosuppressant)

Chronic Obstructive Pulmonary Disease (COPD)

Dyslipidemia (He)

Smoke-absorbing

Immunodeficiency after solid organ transplantation

Pregnancy (gestation)

Patients suffering from a neurological disease or complex condition (e.g., cerebral palsy, congenital disease)

Patients with high medical dependence (e.g. tracheotomy, gastrostomy, positive pressure ventilation)

In one embodiment, the pharmaceutical composition of the present invention is used for patients suffering from SARS-CoV-2-induced pneumonia.

One embodiment of the present invention is to administer the pharmaceutical composition of the present invention to an infected patient having at least one clinical state as shown below among SARS-CoV-2 infected patients.

Patients fitted with ECMO (extracorporeal membrane type artificial lung)

Patient fitted with artificial respirator

Patient entering ICU

Oxygen saturation (SpO) ₂ ) Patients with less than 93% (indoor air) or requiring oxygen inhalation

Oxygen saturation (SpO 2) lower than 94% (indoor air, sea level)

PaO2/FiO2 of less than 300mmHg

Respiratory rate of 30 times/min or more

Lung infiltration of 50% or more

(Process for producing Compound of formula (I))

The compound represented by the formula (I) can be produced, for example, by the synthetic method shown below. The extraction, purification, and the like may be performed by performing a treatment performed in a usual organic chemistry experiment. Reference may be made to methods known in the art for synthesis. The extraction, purification, and the like may be performed by performing a treatment performed in a usual organic chemistry experiment.

The compounds represented by formula (I) may be manufactured by referring to methods known in the art. For example, reference may be made to WO2010092966, WO2012020749, WO2013089212, WO2014200078, WO2012020742, WO2013118855 and the like.

(method for identifying Compound)

NMR analysis was performed at 400MHz using DMSO-d ₆ 、CDCl ₃ The measurement was performed. In addition, when NMR data is expressed, all peaks measured may not be described.

In the specification, RT represents LC/MS: retention time in liquid chromatography/mass spectrometry was measured under the following conditions.

(measurement condition 1)

Column: ACQUITY UPLC (registered trademark) BEH C18 (1.7 μm i.d.2.1X105 mm) (Waters)

Flow rate: 0.8 mL/min

UV detection wavelength: 254nm

Mobile phase: [A] an aqueous solution containing 0.1% formic acid, and [ B ] an acetonitrile solution containing 0.1% formic acid

Gradient: after a linear gradient of 5% to 100% solvent [ B ] was performed for 3.5 minutes, 100% solvent [ B ] was maintained for 0.5 minutes.

(measurement of powder X-ray diffraction Pattern)

Powder X-ray diffraction measurement of the obtained crystals was performed according to the powder X-ray diffraction measurement method described in the general test method of the Japanese drug administration. The measurement conditions are as follows.

(apparatus)

Rigaku Corporation SmartLab

(method of operation)

Assay: reflection method

The wavelength is used: cuK alpha line

Tube current: 200mA

Tube voltage: 45kV

Sample plate: aluminum (Al)

Incidence angle of X-rays: 2.5 degree

Sampling width: 0.02 degree

A detector: hyPix-3000 (two-dimensional detection mode)

(determination and analysis method for analysis of Single Crystal Structure)

The measurement conditions and the analysis method of the single crystal structure analysis are as follows.

(apparatus)

Rigaku Corporation XtaLAB P200 MM007

(measurement conditions)

Measuring temperature: 25 DEG C

The wavelength is used: cuK alpha line

Software: crysalisPro 1.171.39.46e (Rigaku Oxford Diffraction, 2018)

(data processing)

Software: crysalisPro 1.171.39.46e (Rigaku Oxford Diffraction, 2018)

And carrying out Lorentz and polarization correction and absorption correction on the data.

(analysis of Crystal Structure)

Phase determination was performed using the direct method program sheldxt (sheldlick, g.m., 2015), and refinement was performed using the full-matrix least squares method using sheldxl (sheldlick, g.m., 2015). The shift parameters of non-hydrogen atoms were all refined with anisotropy. The hydrogen atom was introduced by calculation using the default parameters of ShellXL, treated as riding atom. All hydrogen atoms were refined with isotropic parameters.

PLUTON (Spek, 1991)/ORTEP (Johnson, 1976) is used in the drawing of FIG. 2.

The synthesis of the compound represented by formula (I) is shown below, for example.

[ chemical formula 8]

Process 1 Synthesis of Compound 18

Compound 4 (926 mg,4.04 mmol), acetonitrile (7.41 mL), potassium carbonate (726 mg,5.25 mmol) and 2,4, 5-trifluorobenzyl bromide (1000 mg,4.44 mmol) were mixed. The reaction solution was stirred at 80℃for 40 minutes, allowed to stand for cooling, and then diluted with ethyl acetate. After filtration of insoluble matter, the filtrate was concentrated to give a crude product of compound 18 (1.51 g,4.04mmol, yield: quantitative).

LC/MS (ESI): m/z=374, rt=2.54 min, lc/MS assay condition 1

Process 2 Synthesis of Compound 19

Compound 18 (1.51 g,4.04 mmol) and TFA (3.02 mL) were mixed. The reaction solution was stirred at room temperature for 4 hours and allowed to stand overnight. TFA was distilled off under reduced pressure, and toluene was added to the residue to conduct azeotropy. Isopropyl ether was added to the residue, which was then suspended and filtered to give compound 19 (1.22 g,3.84mmol, yield 95%).

LC/MS (ESI): m/z=318, rt=1.68 min, lc/MS assay condition 1

Process 3 Synthesis of Compound 20

Compound 19 (200 mg,0.63 mmol), DMF (1.8 mL), potassium carbonate (261 mg,1.89 mmol) and 3- (chloromethyl) -1-methyl-1H-1, 2, 4-triazole hydrochloride (1599 mg,0.946 mmol) were mixed. The reaction solution was stirred at 60℃for 2 hours, and a saturated aqueous ammonium chloride solution was added. The aqueous layer was extracted with ethyl acetate, and the organic layer was washed with saturated brine. The organic layer was dried over magnesium sulfate, filtered, and concentrated. The residue was suspended in a mixed solvent of isopropyl ether, hexane, ethyl acetate and chloroform and filtered. The residue was mixed with DMF (1.8 mL), potassium carbonate (261 mg,1.89 mmol) and 3- (chloromethyl) -1-methyl-1H-1, 2, 4-triazole hydrochloride (1599 mg,0.946 mmol). The reaction solution was stirred at 60℃for 6 hours, and a saturated aqueous ammonium chloride solution was added. The aqueous layer was extracted with ethyl acetate, and the organic layer was washed with saturated brine. The organic layer was dried over magnesium sulfate, filtered, and concentrated. The residue was suspended in a mixed solvent of isopropyl ether, hexane, ethyl acetate and chloroform, and filtered to give compound 20 (116 mg,0.281mmol, yield 45%).

LC/MS (ESI): m/z=413, rt=1.84 min, lc/MS measurement conditions: 1

Process 4 Synthesis of Compound represented by formula (I)

Compound 20 (115 mg,0.279 mmol), THF (2.30 mL) and 6-chloro-2-methyl-2H-indazol-5-amine (60.8 mg,0.335 mmol) were mixed. LHMDS (558. Mu.L, 0.558 mmol) was added dropwise to the reaction solution at 0deg.C. The reaction solution was stirred at 0℃for 2.5 hours, at room temperature for 40 minutes, and a saturated aqueous ammonium chloride solution was added. The organic layer was concentrated by extraction with chloroform. The residue was purified by silica gel column chromatography (chloroform/methanol) to give a compound represented by the formula (I) (61.8 mg,0.116mmol, yield 42%).

¹ H-NMR(CDCl ₃ )δ：7.96(s,1H),7.82(d,J＝2.5Hz,2H),7.48(br s,1H),7.45-7.37(m,1H),7.08(s,1H),6.97-6.88(m,1H),5.35(s,2H),5.17(s,2H),4.21(s,3H),3.89(s,3H).

LC/MS (ESI): m/z=532, rt=1.70 min, lc/MS assay condition 1

To 1170mg of the compound represented by the compound (I) were added 278mg (1.1 eq) of fumaric acid and 5.85mL of ethyl acetate, and the mixture was stirred at room temperature for 45 minutes. The solid was filtered and dried, whereby fumaric acid co-crystal form I crystals (1369.4 mg, 94.6%) of the compound represented by the formula (I) were obtained.

The results of single crystal structure analysis of fumaric acid co-crystal type I crystals of the compound represented by formula (I) are shown below.

R1 (I > 2.00s (I)) was 0.0470, and it was confirmed by the final difference Fourier that there was neither a loss of electron density nor a misplacement.

The crystallographic data are shown in table 1.

TABLE 1

Here, the volume represents the unit cell volume, and Z represents the number of molecules in the unit cell.

The atomic coordinates of the non-hydrogen atom are shown in tables 2 to 3. Here, U (eq) refers to an equivalent isotropic displacement parameter.

Atomic fraction coordinates (×10) ⁴ ) And equivalent isotropic displacement parameterU (eq) is 1/3of the trace of the orthogonalized Uij tensor. (Fractional Atomic Coordinates (. Times.10) ⁴ )and Equivalent Isotropic Displacement Parameters/>U (eq) is defined as 1/3of the trace of the orthogonalized UIJ tensor.) [ Table 2]

Atoms	x	y	z	U(eq)
					Cl36	8115.3(9)	8341.6(8)	5010.7(5)	79.9(3)
F32	8958.5(19)	7981.3(17)	307.5(9)	78.5(5)
					O35	7267(2)	5961.4(16)	1399.9(10)	56.3(5)
O34	5322(3)	4254.8(16)	4098.2(11)	63.3(5)
					O38	3536(2)	9367.5(19)	8936.3(12)	64.2(5)
N12	6506(2)	7056.8(18)	2611.0(12)	44.2(5)
					F33	13870(2)	7642(2)	1402.1(13)	100.3(7)
N16	5475(2)	6174.4(18)	3988.1(12)	48.2(5)
					N14	6120(3)	5115.3(18)	2713.0(12)	47.3(5)
N9	2815(3)	8924(2)	7397.8(13)	55.4(6)
					N10	5772(3)	8146(2)	3856.1(13)	55.1(6)
N1	1276(3)	8864(2)	7324.6(14)	60.2(6)
					F31	12197(3)	7751(3)	3084.6(13)	124.9(9)
N23	3644(3)	4434(2)	1818.7(15)	64.5(6)
					N20	3122(3)	4249(2)	1061.4(15)	64.9(6)
C11	6673(3)	6043(2)	2193.8(15)	44.7(6)
					C9	5879(3)	7178(2)	3527.6(15)	44.2(6)
C10	5619(3)	5119(2)	3639.3(15)	48.4(6)
					N22	5784(3)	3621(2)	814.1(15)	67.8(7)
039	6151(3)	8893(3)	8285.8(15)	109.2(10)
					C12	6985(3)	8068(2)	2049.4(15)	47.2(6)
C20	5248(3)	4044(2)	1633.9(16)	50.7(6)
					C7	5022(3)	8298(2)	4770.9(15)	50.8(6)
C4	3693(3)	8762(2)	6554.3(16)	49.4(6)
					C13	8823(3)	7976(2)	1872.6(16)	48.8(6)
C5	5385(3)	8700(2)	6267.8(17)	56.4(7)
					C19	6380(3)	4009(2)	2279.5(17)	54.5(7)
C14	9741(3)	7934(2)	1013.2(16)	54.7(7)
					C3	2685(3)	8593(2)	5965.2(17)	54.4(7)
C6	6015(3)	8469(2)	5392.0(16)	54.3(7)
					C23	5121(4)	9287(3)	8898.3(18)	62.1(7)
O41	1842(3)	4874(3)	3529.1(18)	119.8(10)
					C8	3370(3)	8376(2)	5054.8(17)	57.4(7)
C24	5542(3)	9730(3)	9679.7(17)	61.9(7)
					C18	9684(4)	7917(3)	2570.7(18)	67.1(8)
C15	11431(3)	7827(3)	831.3(19)	67.8(8)
					C16	12217(3)	7760(3)	1541(2)	67.9(8)
C2	1134(4)	8667(3)	6497.1(18)	67.4(8)

TABLE 3

Atoms	x	y	z	U(eq)
					C17	11360(4)	7806(3)	2405(2)	75.0(9)
C21	4400(4)	3767(3)	485.7(19)	70.6(8)
					O43	-464(4)	4618(4)	3203.2(19)	154.2(15)
C1	9(4)	8943(3)	8139(2)	81.7(10)
					C26	307(4)	4766(4)	3745(2)	93.6(12)
C25	-384(4)	4909(4)	4700(2)	92.1(11)
					C22	1397(4)	4562(4)	963(3)	102.7(13)

Next, the atoms of the hydrogen atoms are noted in table 4. Here, U (iso) refers to an isotropic displacement parameter. The numbers of hydrogen atoms in table 4 are associated with the numbers of bonded non-hydrogen atoms.

Coordinates of hydrogen atomsAnd isotropic displacement parameter->(Hydrogen Atom Coordinates/>and Isotropic Displacement Parameters/>)

TABLE 4

Atoms	x	y	z	U(iso)
					H38	3370.9	9206.88	8452.86	96
H16	5092.25	6215.55	4554.71	58
					H12A	6452.59	8783.45	2347.49	57
H12B	6603.63	8119.01	1479.7	57
					H5	6053.99	8811.71	6658.45	68
H19A	6229.72	3381.57	2741.61	65
					H19B	7509.94	3824.57	1962.58	65
H41	2202.36	4700.41	3007.94	180
					H8	2702.01	8287.11	4656.27	69
H24	6652.83	9619.42	9719.44	74
					H18	9115.24	7953.15	3160.4	81
H15	12010.7	7800.84	243.55	81
					H2	176.44	8593.16	6310.6	81
H21	4344.44	3553.57	-79.51	85
					H1A	260.69	8258.79	8539.89	122
H1B	-1049.48	8985.26	7978.29	122
					H1C	-14.15	9635.57	8433.78	122
H25	-1486.76	4863.66	4886.06	110
					H22A	719.4	4375.73	1521.91	154
H22B	1225.91	4127.33	499	154
					H22C	1105.77	5390.24	801.98	154

In addition, the interatomic bond length (unit: angstrom) is shown in Table 5.

TABLE 5

Co-crystal form I crystals of fumaric acid of the compound represented by formula (I) there are a compound represented by formula (I) 1 and 1 molecule of fumaric acid in an asymmetric unit. The molecular structure obtained by X-ray structural analysis of fumaric acid co-crystal form I crystals of the compound represented by formula (I) is shown in fig. 2.

The numbers of the non-hydrogen atoms in tables 2 to 3 and 5 correspond to the numbers shown in fig. 2, respectively.

As shown in Table 5, the bond length of N10-C9 is aboutThe bond length of N16-C9 is approximately +.>

Due to the bond length of N10-C9 (about) Longer than the bond of N16-C9 (about +.>) Short, therefore, the compound represented by formula (I) of fumaric acid co-crystal form I is identified as an imino structure:

[ chemical formula 9]

In addition, the results of powder X-ray diffraction of fumaric acid co-crystal form I crystals of the compound represented by formula (I) are shown.

In the powder X-ray diffraction pattern, at the diffraction angle (2θ): peaks were confirmed at 7.8.+ -. 0.2 °, 9.5.+ -. 0.2 °, 10.1.+ -. 0.2 °, 10.9.+ -. 0.2 °, 13.8.+ -. 0.2 °, 14.7.+ -. 0.2 °, 18.6.+ -. 0.2 °, 22.6.+ -. 0.2 °, 23.5.+ -. 0.2 ° and 24.6.+ -. 0.2 °.

In the powder X-ray diffraction pattern, diffraction angle (2θ): peaks of 9.5.+ -. 0.2 °, 10.9.+ -. 0.2 °, 18.6.+ -. 0.2 °, 23.5.+ -. 0.2 ° and 24.6.+ -. 0.2 ° are particular characteristics of the fumaric acid co-crystal form I crystal as the compound represented by formula (I).

The compound represented by the formula (I) has a coronavirus 3CL protease inhibitory activity and is therefore useful as a therapeutic and/or prophylactic agent for viral infectious diseases.

Further, the compound represented by the formula (I) has usefulness as a medicament, and preferably has any one or more of the following excellent characteristics.

a) The inhibition of CYP enzymes (e.g., CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP3A4, etc.) is weak.

b) Shows good pharmacokinetics with high bioavailability, moderate clearance rate and the like.

c) High metabolic stability.

d) The CYP enzyme (e.g., CYP3 A4) does not exhibit an irreversible inhibitory effect in the concentration range of the measurement conditions described in the present specification.

e) Has no mutagenicity.

f) The risk of cardiovascular system is low.

g) Showing high solubility.

h) The protein unbound fraction (fu value) was high.

i) Has high selectivity of coronavirus 3CL protease.

j) Has high coronavirus proliferation inhibiting activity. For example, it has high coronavirus proliferation inhibitory activity when added to Human Serum (HS) or Human Serum Albumin (HSA).

Examples of the coronavirus proliferation inhibitor include, for example, EC in the CPE inhibition effect confirmation test (SARS-CoV-2) described later ₅₀ Is 10. Mu.M or less, preferably 1. Mu.M or less, more preferably 100nM or less.

The crystal/complex (co-crystal) of the compound represented by formula (I) is useful as a medicine, and preferably has any one or more of the following excellent characteristics.

A) Shows high bioavailability, moderate clearance, high AUC, high maximum blood concentration, etc., and good pharmacokinetics.

B) Shows high solubility, high chemical stability and low hygroscopicity.

The pharmaceutical composition of the present invention may be administered by any method orally, parenterally. Examples of the parenteral administration include percutaneous, subcutaneous, intravenous, intraarterial, intramuscular, intraperitoneal, transmucosal, inhalation, nasal, eye drop, ear drop, and intravaginal administration.

In the case of oral administration, the composition may be formulated into any of commonly used dosage forms such as solid preparations for internal use (for example, tablets, powders, granules, capsules, pills, films, etc.), solutions for internal use (for example, suspensions, emulsions, elixirs, syrups, lemonades, alcoholic solutions, aromatic solutions, extracts, decoction, tinctures, etc.), and the like, according to a conventional method. The tablet can be sugar-coated tablet, film coated tablet, enteric coated tablet, delayed release tablet, buccal tablet, sublingual tablet, buccal tablet, chewable tablet or orally disintegrating tablet, the powder and granule can be dry syrup, and the capsule can be soft capsule, microcapsule or delayed release capsule.

In the case of parenteral administration, it can be suitably administered by any of commonly used dosage forms such as injections, drops, external preparations (e.g., eye drops, nose drops, ear drops, aerosols, inhalants, lotions, injections, coating agents, cough-containing agents, enemas, ointments, plasters, jellies, creams, patches, cataplasms, external powders, suppositories, etc.). The injection can be O/W, W/O, O/W/O, W/O/W emulsion.

If necessary, various pharmaceutical additives such as excipients, binders, disintegrants, lubricants and the like, which are suitable for the dosage form of the compound represented by the formula (I), may be mixed with an effective amount of the compound to prepare a pharmaceutical composition. The pharmaceutical composition can be prepared into a pharmaceutical composition for children, the elderly, patients of severe disease or for surgery by appropriately changing the effective amount, dosage form and/or various pharmaceutical additives of the compound represented by the formula (I). For example, the pharmaceutical composition for children may be administered to a newborn (less than 4 weeks after birth), an infant (4 weeks after birth to less than 1 year old), a young child (more than 1 year old and less than 7 years old), a child (more than 7 years old and less than 15 years old), or a patient of 15 years to 18 years old. For example, a pharmaceutical composition for elderly people may be administered to patients over 65 years old.

The administration amount of the pharmaceutical composition of the present invention (pharmaceutical composition comprising fumaric acid co-crystal form I crystals of the compound represented by formula (I)) is preferably set in consideration of the age, weight, kind, degree of disease, administration route, etc. of the patient, and in the case of oral administration, is usually in the range of 0.05 to 200 mg/kg/day, preferably 0.1 to 100 mg/kg/day. In the case of parenteral administration, although there are large differences depending on the route of administration, it is usually in the range of 0.005 to 200 mg/kg/day, preferably 0.01 to 100 mg/kg/day. It is administered 1 to 1 time a day to several times.

For example, 375mg may be orally administered 1 day 1, and 125mg may be orally administered 1 day 2 to 5 days 1, based on the compound represented by the formula (I), to children over 6 years and less than 12 years, and to children over 12 years and adults.

For example, 250mg may be orally administered 1 time on day 1 to 1 and 125mg may be orally administered 1 time on day 2 to 5 days 1 for children weighing 30kg or more and less than 40kg, and 6 years or more and less than 12 years, based on the compound represented by the formula (I).

For example, 250mg may be orally administered 1 time on day 1 to 100mg orally administered 1 time on day 2 to 5 days 1 for a child weighing 30kg or more and less than 40kg, 6 years or more and less than 12 years old, based on the compound represented by the formula (I).

For example, for a child weighing 20kg or more and less than 30kg, 6 years or more and less than 12 years old, 125mg may be orally administered 1 time on day 1, and 75mg may be orally administered 1 time on day 2 to day 5, 1 time on day 1, based on the compound represented by the formula (I).

For example, 150mg may be orally administered 1 time on day 1 to 1 time on day 2 to 5 days 1 to 75mg orally administered 1 time on day 1, based on the compound represented by the formula (I), for children weighing 20kg or more and less than 30kg, and 6 years or more and less than 12 years.

For example, for a child weighing 15kg or more and less than 20kg, 75mg may be orally administered 1 time on day 1, and 50mg may be orally administered 1 time on day 2 to day 5, 1 time on day 1, based on the compound represented by the formula (I).

For example, 100mg may be orally administered 1 day 1 and 50mg may be orally administered 1 day 2 to 5 days 1 for children weighing 15kg or more and less than 20kg, based on the compound represented by the formula (I).

For example, for children weighing 15kg or more and less than 20kg, 6mg/kg may be orally administered 1 time on day 1, and 3mg/kg may be orally administered 1 time on day 2 to day 5, 1 time on day 1, based on the compound represented by the formula (I).

The compound represented by the formula (I) may be used in combination with other novel therapeutic agents for coronavirus infectious disease (covd-19) (including agents that have been approved as such therapeutic agents, and agents that are in development or later developed) (hereinafter, referred to as combination agents) for the purpose of enhancing the action of the compound or reducing the administration amount of the compound. In this case, the compound represented by formula (I) and the combination agent may be administered simultaneously or with a time difference therebetween, without limitation on the timing of administration. The compound represented by the formula (I) and the combination agent may be administered in the form of a preparation containing 2 or more kinds of each active ingredient, or may be administered in the form of a single preparation containing the active ingredients.

The administration amount of the combination agent may be appropriately selected based on the clinically used amount. The mixing ratio of the compound represented by formula (I) and the pharmaceutical combination may be appropriately selected according to the administration subject, the administration route, the disease, the symptoms, the combination, and the like of the subject. For example, when the subject is a human, 0.01 to 100 parts by weight of the combination agent may be used per 1 part by weight of the compound represented by the formula (I).

As illustrated in the examples below, the present invention provides methods for treating a novel coronavirus infectious disease (COVID-19) and/or alleviating symptoms caused by the novel coronavirus infectious disease (COVID-19).

In addition, the present invention has an advantage that substantially no side effects are generated when the pharmaceutical composition of the present invention is administered to a human.

In addition, methods are provided for alleviating symptoms (Long covd, e.g., tiredness, dyspnea, etc.) that persist after recovery from a novel coronavirus infectious disease (covd-19).

Examples

Hereinafter, the present invention will be described based on examples, but the present invention is not limited to these examples and the like.

Test example 1: cytopathic Effect (Cytopathic effect) (CPE) inhibition Effect confirmation assay Using human TMPRSS 2-expressing Vero E6 cells (Vero E6/TMPRSS2 cells)

< operation procedure >

Dilution and dispensing of the test sample

Test samples were diluted to appropriate concentrations in DMSO in advance, and after 3-fold gradient dilution series were prepared, they were dispensed into 96-well plates.

Dilution and dispensing of cells and SARS-CoV-2

VeroE6/TMPRSS2 cells (JCRB 1819, 1.5X10) ⁴ Individual cells/well) and SARS-CoV-2hCoV-19/Japan/TY/WK-521/2020, hCoV-19/Japan/QK002/2020, hCoV-19/Japan/QHN/2020, hCoV-19/Japan/QHN002/2020, hCoV-19/Japan/TY7-501/2021, hCoV-19/Japan/TY7-503/2021, hCoV-19/Japan/TY8-612/2021, hCoV-19/Japan/TY11-927-P1/2021, hCoV-19/Japan/TY33-456/2021, hCoV-19/Japan/28-444/2021 hCoV-19/Japan/TY26-717/2021, hCoV-19/Japan/TY38-871/2021, hCoV-19/Japan/TY40-385/2022, hCoV-19/Japan/TY41-721/2022, hCoV-19/Japan/TY41-716/2022, hCoV-19/Japan/TY41-703/2022, hCoV-19/Japan/TY41-702/2022, hCoV-19/Japan/TY41-796/2022, hCoV-19/Japan/TY41-795/2022, hCoV-19/Japan/TY41-686/2022 (30-3000 TCID) ₅₀ Well) in a culture medium (MEM, 2% FBS, penicillin-streptomycin) and was dispensed into a well containing a test sample, and then mixed with a sample in CO ₂ Culturing in an incubator for 3 days or 4 days.

Measurement of a light-emitting Signal by dispensing CellTiter-Glo (registered trademark) 2.0

After plates after 3 days of incubation were returned to room temperature, cellTiter-Glo (registered trademark) 2.0 was dispensed into each well and mixed with a plate mixer. After a certain time, the luminescence signal (Lum) was measured with a plate reader.

< calculation of measured item values >)

50% concentration of inhibition of death of SARS-CoV-2 infected cells (EC ₅₀ ) Calculation of

Let x be the pair of compound concentrationsWhen the value is given as% Efficacy (efficiency), the inhibition curve is fitted by the following Logistic regression equation, and the value of x substituted into y=50 (%) is calculated as EC ₅₀ 。

y＝min+(max-min)/{1+(X50/x)^Hill}

% potency = { (sample-virus control)/(cell control-virus control) } 100%

(％Efficacy＝{(Sample-virus control)/(cell control-virus control)}*100％)

Cell control (cell control): average Lum (the average of Lum of cell control wells) of cell control wells

Virus control (virus control): average Lum (the average of Lum of virus control wells) of virus control wells

min: y-axis lower limit, max: y-axis upper limit, X50: the x-coordinate of the inflection point, hill: slope of curve at midpoint of min and max

The fumaric acid co-crystal form I crystals of the compound represented by formula (I) were tested essentially as described above. The results are shown below.

TABLE 6

Test example 2

Inhibition Activity assay for SARS-CoV-23CL protease

< Material >

Commercially available recombinant SARS-CoV-23CL protease

Commercially available substrate peptides

Dabcyl-Lys-Thr-Ser-Ala-Val-Leu-Gln-Ser-Gly-Phe-Arg-Lys-Met-Glu (Edans) -NH2 (SEQ ID NO: 1)

Internal standard peptide (Internal Standard peptide)

Dabcyl-Lys-Thr-Ser-Ala-Val-Leu (13C6,15N) -Gln (SEQ ID NO: 2)

Dabcyl-Lys-Thr-Ser-Ala-Val-Leu (13C6,15N) -Gln can be synthesized by reference (Athereton, E.; sheppard, R.C., "In Solid Phase Peptide Synthesis, A Practical Approach", IRL Press from the university of oxford (IRL Press at Oxford University Pres) 1989, and Bioorg. Med. Chem., vol.5, no. 9, 1997, pages 1883-1891, etc.). Examples are shown below.

H-Lys-Thr-Ser-Ala-Val-Leu (13C6,15N) -Glu (resin) -OαOtBu (Lys side chain protected by Boc, thr side chain protected by tert-butyl, ser side chain protected by tert-butyl, C-terminal OH of Glu protected by tert-butyl, carboxylic acid of Glu side chain condensed with resin) was synthesized by Fmoc solid phase synthesis using Rink amide resin. Modification of the N-terminal Dabcyl group is the condensation of 4-dimethylaminoazobenzene-4' -carboxylic acid (Dabcyl-OH) on the resin with EDC/HOBT. Final deprotection and cleavage from resin was achieved by using TFA/edt=95: 5, performing treatment. Then, purification was performed by reverse phase HPLC.

·RapidFire Cartridge C4 typeA

< operation procedure >

Preparation of assay buffer

In this test, a detection buffer consisting of 20mM Tris-HCl, 1mM EDTA, 10mM DTT, 0.01% BSA was used.

Dilution and dispensing of the test sample

Test samples were diluted in DMSO to moderate concentrations in advance, and after making a 3-fold gradient dilution series, dispensed into 384-well plates.

Addition of enzyme and substrate, enzymatic reaction

To the prepared compound plate, 8. Mu.M substrate and 6nM enzyme solution were added and incubated at room temperature for 3 hours. Then, a reaction termination solution (0.072. Mu.M internal standard (Internal Standard), 0.1% formic acid, 10% acetonitrile) was added to terminate the enzyme reaction.

Determination of the reaction products

For the plates after completion of the reaction, the measurement was performed using a Rapid FireSystem 360 and a mass spectrometer (Agilent, 6550iFunnel Q-TOF). As mobile phases for the measurement, a solution A (75% isopropanol, 15% acetonitrile, 5mM ammonium formate) and a solution B (0.01% trifluoroacetic acid, 0.09% formic acid) were used.

The reaction Product detected by the mass spectrometer was calculated using RapidFire Integrator as a Product area (area) value. In addition, the detected internal standard (Internal Standard) is also calculated as an internal standard (Internal Standard) area (area) value.

< calculation of measured item values >)

Calculation of P/IS

The area value obtained in the previous item was calculated by the following equation, and P/IS was calculated.

P/is=product area (Product area) value/internal standard area (Internal Standard area) value

50% SARS-CoV-2 3CL protease inhibition concentration (IC ₅₀ ) Calculation of

Let x be the logarithmic value of the compound concentration, let y be% Inhibition (Inhibition), the Inhibition curve was fitted by the following Logistic regression equation, and the value of x substituted into y=50 (%) was calculated as IC ₅₀ 。

y＝min+(max-min)/{1+(X50/x)＾Hill}

% inhibition = {1- (sample-control (-))/control (+) -control (-)) } 100

(％Inhibition＝{1-(Sample-Control(-))/Control(+)-Control(-))}*100)

Control (-): average P/IS ratio of wells without SARS-CoV-2 3CL protease and test substrate

(Control(-)：the average of P/IS ratio in the wells without SARS-CoV-2 3CL protease and test substance)

Control (+): average P/IS ratio of wells with SARS-CoV-2 3CL protease and no test substrate

(Control(+)：the average of P/IS ratio in the wells with SARS-CoV-2 3CL protease and without test substance)

min: y-axis lower limit, max: y-axis upper limit, X50: the x-coordinate of the inflection point, hill: slope of curve at intermediate point of min and max

Fumaric acid co-crystal form I crystals of a compound represented by formula (I): 0.0132 mu M

Test example 3-1 first phase clinical trial

All tests were performed based on the clinical practice standard (GCP) of pharmaceuticals. The first-stage clinical trial consisted of parts 1 to 7, and in part 1, safety, tolerability, and pharmacokinetics were confirmed in single administration and repeated administration of the fumaric acid co-crystal form I crystal of the compound represented by formula (I).

Hereinafter, the secondary object and a part of the secondary evaluation items of section 1 are shown.

TABLE 7

The Day before the administration Day of the test drug (Day-1), subjects judged to be acceptable by the screening test were randomly assigned to either one of the active drug group (the amount of fumaric acid co-crystal type I crystals of the compound represented by formula (I) is referred to in the following table) or the placebo group. Day 1 administered the test drug (active drug or placebo) in a single administration on an empty stomach.

(results)

T at single administration in the first phase _1/2,z The results of (2) are shown below.

TABLE 8

Geometric mean (coefficient of variation)

As shown in the above table, the half-life exceeded 40 hours in all of the administration amount groups, indicating that the half-life of the compound represented by formula (I) was long.

In addition, in the first-stage trial, serious adverse events that could not negate causal relationships were not reported.

Test example 3-2 prediction of single administration on empty stomach of 500mg

For the administration of 500mg (suspension) of the test agent in the 1 st stage test in Japan to 6 healthy adult men in Japanese in a single oral administration in a fasting state, 336 hours agoAssuming that the pharmacokinetics is linear, the change in plasma drug concentration in the dose of the regimen of figure 3 was simulated using an additive method. From pharmacokinetic/pharmacodynamic analysis using a mouse model of SARS-CoV-2 infection, it was estimated that the viral titers were reduced by 2log and 3log of the highest human plasma concentration (C _max ) 14.1. Mu.g/mL and 23.0. Mu.g/mL, respectively. In addition, the loading dose is set so as to achieve the concentration of the drug in the plasma required to exert the desired drug effect from the first day of administration. According to the above, as the usage amount for the 2/3 stage test, set as: it is assumed that the dose 375mg (day 1) and the maintenance dose 125mg (day 2 to day 5) for obtaining the exposure required to reduce the virus titer by 2log or more are repeatedly administered for 5 days 1 time each day; it is assumed that the exposure load amount of 750mg (day 1) and the maintenance amount of 250mg (day 2 to day 5) required for reducing the virus titer by 3log are sufficiently obtained and the administration is repeated for 5 days 1 time. It is considered that the concentration in plasma required for exhibiting the drug effect can be maintained at the set dose without exceeding NOAEL (75.2 to 77.6. Mu.g/mL) obtained in the toxicity test using a monkey.

Test example 4 clinical trial (Ph 2/3 trial)

Evaluation of the efficacy and safety of repeated oral administration of the test agent (active ingredient: fumaric acid co-crystal form I of the compound represented by formula (I)) to SARS-CoV-2 infected persons having only mild/moderate and no/mild symptoms was carried out by a double-blind comparative test of random grouping with placebo as a comparative subject.

The main evaluation items of the 2a stage Part (Phase 2a Part) were the amount of change of the viral titer of SARS-CoV-2 from baseline at each time point common to both the febrile/moderate and asymptomatic SARS-CoV-2 infectors, and the antiviral effect of the test agent was confirmed.

The main evaluation items of the 2b stage Part (Phase 2b Part) are: the clinical symptom improvement effect and antiviral effect of the test agent were confirmed by the change in the total score of 12 symptoms of covd-19 per unit time from the start of the first administration (Day 1) to 120 hours (Day 6) and the change in the viral titer of Day 4 for SARS-CoV-2 relative to the baseline in the mild/moderate patients suffering from SARS-CoV-2.

The time until recovery of 5 symptoms of covd-19 was evaluated by the subject himself with 4 scales (0: none, 1: mild, 2: moderate, 3: severe) for 5 symptoms of covd-19, the main evaluation item of the 3-stage Part (Phase 3 Part) being the time for mild/moderate SARS-CoV-2 infected persons, and the effectiveness was verified.

The main evaluation item of the 2b/3 stage Part (Phase 2b/3 Part) was the proportion of subjects with onset/worsening of the symptoms of covd-19 among the asymptomatic/only mildly symptomatic SARS-CoV-2 infected subjects, and the subjects themselves evaluated 12 symptoms of covd-19 on 4 scales (0: none, 1: mild, 2: moderate, 3: severe) and the subjects themselves evaluated dysgeusia and dysolfactory on 3 scales (0: common, 1: imperceptible, 2: completely imperceptible) to verify effectiveness.

The major secondary evaluation items of the 3-stage Part (Phase 3 Part) and the 2 b/3-stage Part (Phase 2b/3 Part) were the amount of change in the viral RNA amount of SARS-CoV-2 from Day 4 relative to the baseline, and the time until the virus titer of SARS-CoV-2 was first confirmed to be negative, and the antiviral effect was verified.

Patients with mild/moderate SARS-CoV-2 infection were selected to fully meet the criteria described below.

(a) Male or female patients over 12 years old and less than 70 years old.

(b) People diagnosed as SARS-CoV-2 positive within 120 hours prior to registration.

(c) The time from onset of covd-19 to registration was less than 120 hours.

(d) A person who has any one of the following symptoms (12 symptoms of COVID-19) caused by COVID-19 at the time of registration and has symptoms (excluding symptoms existing before the onset of COVID-19) of a moderate level (COVID-19 symptom score: 2) or more.

Systemic symptoms: tiredness, muscle pain or body pain, headache, chills/sweating, sensation of fever

Respiratory symptoms: runny nose or stuffy nose, sore throat, cough, shortness of breath (dyspnea)

Digestive organ symptoms: nausea, vomiting and diarrhea

Asymptomatic SARS-CoV-2 infected individuals were selected to fully meet the criteria described below.

(a) Male or female patients over 12 years old and below 70 years old.

(c) Asymptomatic (Phase 2a Part): within 2 weeks prior to registration, no following symptoms of covd-19 (excluding symptoms present prior to SARS-CoV-2 infection) were found.

Systemic symptoms: tiredness, muscle pain or body pain, headache, chills/sweating, fever, abnormal taste, abnormal smell

Digestive organ symptoms: nausea, vomiting and diarrhea

Asymptomatic/only mild symptoms (2 b/3 stage Part (Phase 2b/3 Part)): persons (excluding symptoms present before the onset of COVID-19) who did not have the following symptoms (12 symptoms of COVID-19) caused by COVID-19 above a moderate (COVID-19 symptom score: 2) within 2 weeks prior to random grouping.

Systemic symptoms: debilitation (fatigue), muscle pain or body pain, headache, chills/sweating, feeling fever or fever

Digestive organ symptoms: nausea, vomiting and diarrhea

Method for administering test agent

(i) Test agent

250mg tablet: the tablet contains fumaric acid co-crystal form I crystals of the compound represented by the formula (I) and 250mg of the fumaric acid co-crystal form I crystals are calculated based on the compound represented by the formula (I).

125mg tablet: the tablet contains fumaric acid co-crystal form I crystals of the compound represented by the formula (I), and contains 125mg based on the compound represented by the formula (I).

(ii) Placebo

placebo-D tablets: a tablet which is indistinguishable in appearance from the 250mg tablet and does not contain fumaric acid co-crystal form I crystals of the compound represented by the formula (I).

placebo-B tablets: a tablet which is indistinguishable in appearance from the 125mg tablet and does not contain fumaric acid cocrystal form I crystals of the compound represented by the formula (I).

Administration amount and administration method

In Phase 2a Part, subjects who were judged to be qualified mild/moderate or asymptomatic SARS-CoV-2 infected subjects were treated with 1:1:1 was randomly assigned to one of 375/125mg group of test agent, 750/250mg group of test agent, placebo group.

In the 2b Phase Part (Phase 2b Part) and the 3 Phase Part (Phase 3 Part), subjects of SARS-CoV-2 infected subjects who were judged to be eligible for mild/moderate disease were treated with 1:1:1 to a 375/125mg group of the test agent, a 750/250mg group of the test agent, a placebo group, in the 2b/3 Phase 2b/3Part, subjects of SARS-CoV-2 infected subjects who will be judged to be eligible/only with mild symptoms at 1:1:1 was randomly assigned to one of 375/125mg group of test agent, 750/250mg group of test agent, placebo group.

The administration of the test agent was repeated 1 day for 5 days with 1 time. In 375/125mg group, 375mg was administered as load dose at Day 1 and 125mg was administered as maintenance dose at Day 2-5. In the 750/250mg group, 750mg was administered as the loading dose to Day 1, and 250mg was administered as the maintenance dose to Day 2 to 5.

Test drugs for each administration group

375/125mg group

In Day 1, 3 125mg tablets and placebo-D tablets were administered, and in Day 2 to 5, 1 Day 1 tablet was administered with 125mg tablets and placebo-D tablets, respectively.

750/250mg group

At Day 1, 3 250mg and placebo-B tablets were administered, respectively, and at Day 2 to 5, 1 Day 1, 250mg and placebo-B tablets were administered, respectively.

Placebo group

3 placebo-D and placebo-B tablets were administered at Day 1, and 1 placebo-D and placebo-B tablets were administered at Day 2-5, respectively, for 1 Day 1.

"Day 1" means the first Day of administration, and "Day 2 to Day 5" means the 2 nd to 5 th days from the first Day of administration.

Main evaluation item of effectiveness (2 a Phase 2a Part)

The main item for evaluating the effectiveness of the 2a Phase 2a Part is the amount of change in the viral titer of SARS-CoV-2 from baseline at each time point common to both mild/moderate and asymptomatic SARS-CoV-2 infected individuals. Defined as the absolute change from baseline in observed values of SARS-CoV-2 virus titer at each time point.

Main evaluation item of efficacy (Phase 2b Part): SARS-CoV-2 infected with mild/moderate disease)

The main evaluation items of the effectiveness of the 2b stage Part (Phase 2b Part) for the mild/moderate SARS-CoV-2 infected person are the amount of change per unit time from the first administration start (Day 1) to 120 hours (Day 6) of the 12 symptom total score of COVID-19, and the amount of change in the viral titer of SARS-CoV-2 of Day 4 from the baseline. The total score of 12 symptoms of covd-19 was a score obtained by adding the scores obtained by evaluating the 12 symptoms of covd-19 (debilitation [ tiredness ], muscle pain or body pain, headache, shivering/sweating, feeling fever or fever, runny nose or nasal obstruction, sore throat, cough, shortness of breath [ dyspnea ], nausea, vomiting, diarrhea) in the subject's log on 4 scales (0: none, 1: mild, 2: moderate, 3: severe), respectively. For the change amount of 12 symptom total scores of the covd-19 observed at each time point from the baseline, the result obtained by dividing the AUC of 120 hours (Day 6) from the start of the first administration (Day 1) by the evaluation time (unit: hour) at the time of AUC calculation was calculated as the change amount per unit time.

Main evaluation item of efficacy (Phase 3 Part): SARS-CoV-2 infected with mild/moderate disease)

The main item of evaluation of the effectiveness of the 3-Phase Part (Phase 3 Part) of SARS-CoV-2 infected patients against mild/moderate symptoms is the time until recovery of the 5 symptoms of COVID-19. Is defined as the time from the start of administration of the test drug to the point at which 5 symptoms of covd-19 (runny nose or nasal congestion, sore throat, cough, feeling hot or fever, weakness (tiredness)) are completely recovered.

The symptoms of covd-19 were evaluated in the subject log on 4 scales (0: none, 1: mild, 2: moderate, 3: severe) and their recovery was judged according to the following rules, and their status continued for at least 24 hours.

Regarding the original symptoms that exist just before the onset of covd-19 and that the subject judges to be worsening at the baseline (pre-administration examination) time point, improvement or maintenance of the severity of the condition relative to the baseline time point is required.

The severity of the disease at baseline is severe to moderate, mild, or none

The disease is mild to moderate at baseline, or no

The disease is mild to mild or absent at baseline

Regarding the original symptoms that exist just before the onset of covd-19 and that the subject judged not to have deteriorated at the baseline (pre-administration examination) time point, it is necessary to maintain or improve the severity of the condition at the baseline time point.

The disease degree at baseline is severe to severe, moderate, mild, or none

The disease is mild to moderate, mild, or none at baseline

The disease is mild to mild or absent at baseline

For symptoms other than those described above, i.e., symptoms that do not exist before onset of covd-19, appear at or after the baseline (pre-administration test) time point, it is desirable that the symptoms disappear.

The disease is severe, moderate, or mild to no at baseline

Main evaluation item of effectiveness (2 b/3 Phase Part (Phase 2b/3 Part): asymptomatic/SARS-CoV-2 infected with only mild symptoms)

The main item of evaluation of the effectiveness of the 2b/3 stage fraction (Phase 2b/3 Part) in the 2/3 stage trial for asymptomatic/only mildly symptomatic SARS-CoV-2 infected subjects is the proportion of subjects with onset/exacerbation of the symptoms of COVID-19. Is defined as the proportion of subjects who develop/worsen any of 12 symptoms or dysgeusia and dysolfaction of covd-19 during evaluation of the subject's journal.

In the subject log recorded for the symptoms of COVID-19, the symptoms were evaluated on 3 scales of 0 to 2 (0: as usual, 1: imperceptible as compared with usual, 2: completely imperceptible), and the symptoms were evaluated on 4 scales of 0 to 3 (0: none, 1: mild, 2: moderate, 3: severe) for 12 symptoms of COVID-19, and the onset was judged according to the following rule.

Abnormal taste and smell

Score for dysgeusia and olfactory abnormalities from "0: as usual, "worsen to" 1: compared with the usual, no or 2 is felt: completely imperceptible ", or from" 1: in comparison with usual, no "deterioration to" 2: completely go unnoticed "

Feeling 3 symptoms of fever, cough, shortness of breath (dyspnea)

Score for a symptom worsens from baseline by more than 1 grade:

the disease degree at the baseline is no- & gtworsening to mild, moderate or severe

The disease is mild to moderate or severe at baseline

The severity of the disease at baseline was moderate to severe

(cases where the severity of the condition is severe when the baseline is not included in the subjects for diagnosis of symptoms of COVID-19)

Fatigue, muscle pain or body pain, headache, shivering/sweating, runny nose or nasal obstruction, sore throat, nausea, vomiting, diarrhea 9 kinds of symptoms

At the same time point, the score of 2 or more symptoms worsens by 1 or more grades from baseline, the state of which persists for at least 24 hours:

The disease is mild to moderate or severe at baseline

The severity of the disease at baseline was moderate to severe

Primary secondary evaluation item of effectiveness

The primary secondary evaluation items of the effectiveness of the 3-Phase Part (Phase 3 Part) and the 2 b/3-Phase Part (Phase 2b/3 Part) in the 2/3-Phase test were the amount of change in the viral RNA amount of SARS-CoV-2 from Day 4 relative to the baseline, and the time until the virus titer of SARS-CoV-2 was first confirmed to be negative. The time until the virus titer of SARS-CoV-2 is confirmed to be negative for the first time is defined as the time from the start of administration of the test drug to the time when the virus titer of SARS-CoV-2 does not reach the detection limit for the first time.

Analysis of Main evaluation items (2 a Phase section)

Aiming at the light/middle-disease SARS-CoV-2 infected person group, asymptomatic SARS-CoV-2 infected person group and the combined group, respectively taking the mITT group as the object, calculating the summary statistics of the variation of the virus titer of SARS-CoV-2 relative to the baseline at each time point. In addition, the virus titers of SARS-CoV-2 at each time point between the dose group and placebo group of the test agent were compared at a bilateral significance level of 5% using the van Elteren test for the population after combining the mild/moderate and asymptomatic SARS-CoV-2 infectors. The layer used in the van Elteren test used a population of mild/moderate SARS-CoV-2 infected persons, and a population of asymptomatic SARS-CoV-2 infected persons.

Analysis of Main evaluation items (Phase 2b Part: SARS-CoV-2 infected person with mild/moderate disease)

In the ITT1 group, a covariance analysis was performed in which the total score of 12 symptoms of the first-administration start (Day 1) of the COVID-19 was 1 or more and the total score of 12 symptoms of the first-administration start (Day 1) of the COVID-19 was a CoV-2 infection, and the time from the onset of the COVID-19 to the random grouping (less than 72 hours, 72 hours or more) and the total score of 12 symptoms of the COVID-19 at the baseline was used as a covariate, and the dose groups of the test agent and the placebo group were compared in pairs. In addition, as a main analysis of the amount of change in the viral titer of SARS-CoV-2 against Day 4 from the baseline, a covariance analysis was performed in which the amount of change in the viral titer of SARS-CoV-2 against the baseline in the patient 1 population was determined by using the time (less than 72 hours, 72 hours or more) from the onset of the COVID-19 to the random grouping and whether or not the SARS-CoV-2 vaccine was inoculated and the viral titer of SARS-CoV-2 against the baseline as a covariate, and the dose groups of the test drugs were compared with the placebo group in pairs.

Analysis of Main evaluation items (Phase 3 Part: SARS-CoV-2 infected person with mild/moderate disease)

As a main analysis, among ITT populations, the mild/moderate SARS-CoV-2 infected population, whose time from the onset of covd-19 to the random grouping was less than 72 hours, was compared between 125mg group of test drug and placebo group for the time until recovery of 5 symptoms of covd-19, using the stratified generalized Wilcoxon test of Peto-predce, layered with or without inoculation of SARS-CoV-2 vaccine, at a single-sided significance level of 0.025. The same analysis as the main analysis was performed with respect to the main analysis of the minor/moderate SARS-CoV-2 infectious agent population in the ITT population as the main evaluation item. In this case, the time (less than 72 hours and 72 hours or more) from the onset of the COVID-19 to the random grouping is included in the layer, in addition to whether or not the SARS-CoV-2 vaccine is inoculated.

Analysis of Main evaluation items (2 b/3 stage Part (Phase 2b/3 Part): asymptomatic/SARS-CoV-2 infected with only mild symptoms)

As a main analysis, the proportion of subjects with 5% bilateral significance level for onset/exacerbation of the covd-19 symptoms was compared between 125mg group of test agent and placebo group using Mantel-Haenszel test as to whether or not to vaccinate with SARS-CoV-2 vaccine, targeting the asymptomatic/only mildly symptomatic SARS-CoV-2 infected population in ITT population.

Analysis of major secondary evaluation items (3-Phase Part (Phase 3 Part) and 2 b/3-Phase Part (Phase 2b/3 Part))

Variation of the viral RNA amount of SARS-CoV-2 from baseline of Day 4:

the main analysis was performed for the mild/moderate SARS-CoV-2 infected person, the asymptomatic/only mild SARS-CoV-2 infected person, respectively. Among the light/middle-disease SARS-CoV-2 infected patients, the 3-Phase partial (Phase 3 Part) ITT group was subjected to a covariance analysis using, as a main analysis of the amount of change of the viral RNA amount of SARS-CoV-2 relative to the baseline, and the amount of the viral RNA of SARS-CoV-2 at the baseline as a covariate, and the 125mg group of the test drug was compared with the placebo group. As a major secondary analysis, the same analysis was performed on ITT population. In this case, the time (less than 72 hours and 72 hours or more) from the onset of the COVID-19 to the random grouping was included in the covariate, in addition to the amount of SARS-CoV-2 virus RNA at the time of the inoculation of the SARS-CoV-2 vaccine and the baseline. Among asymptomatic/only slightly symptomatic SARS-CoV-2 infected individuals, the ITT population of stage 2b/3 (Phase 2b/3 Part) was used as a main analysis of the amount of viral RNA of SARS-CoV-2 against Day 4 relative to the baseline, and a covariance analysis using whether or not to inoculate SARS-CoV-2 vaccine and the amount of viral RNA of SARS-CoV-2 at the baseline as a covariate was used as a main analysis of the amount of viral RNA of SARS-CoV-2 against Day 4 relative to the baseline, and the comparison was made between the 125mg group of the test agent and the placebo group.

Time until the virus titer of SARS-CoV-2 was first confirmed to be negative:

the main analysis was performed for the mild/moderate SARS-CoV-2 infected person, the asymptomatic/only mild SARS-CoV-2 infected person, respectively. Among the mild/moderate SARS-CoV-2 infected individuals, the 3-Phase Part (Phase 3 Part) of the mITT population was compared between the 125mg group of the test agent and the placebo group for the time until the virus titer of SARS-CoV-2 was first confirmed to be negative, using the stratified log-rank test for whether or not the SARS-CoV-2 vaccine was vaccinated as a layer, at a single-sided significance level of 0.025, with the time from the onset of COVID-19 to the time of random grouping of less than 72 hours. As a major secondary analysis, the same analysis was performed on the mITT population. In this case, the time (less than 72 hours and 72 hours or more) from the onset of the COVID-19 to the random grouping is included in the layer, in addition to whether or not the SARS-CoV-2 vaccine is inoculated. Among asymptomatic/only mildly symptomatic SARS-CoV-2 infected individuals, the mITT population of the 2b/3 stage fraction (Phase 2b/3 Part) was compared between the 125mg group of the test agent and the placebo group at a single-sided significance level of 0.025 using a stratified log-rank test with a layer of SARS-CoV-2 vaccine.

Results of the Main evaluation project

(1) Variation of SARS-CoV-2 viral titer from baseline at each time point (2 a Phase 2a Part)

In the 2a Phase section (Phase 2a Part), 69 cases were randomly grouped, of which 22 cases (of which 1 case was not administered) were assigned to 375/125mg group, 23 cases were assigned to 750/250mg group, and 24 cases were assigned to placebo group. Of 69 cases, the number of cases in which the baseline RT-PCR was positive was 44, and the number of cases in which the baseline virus titer was detected was 40. The 40 cases consisted of 375/125mg group 14 cases, 750/250mg group 13 cases and placebo group 13 cases. The number of these individuals and their details were calculated based on the RT-PCR measurement results and the virus titer measurement results obtained before the time point of day 17 of 1 month 2022.

As a final result of the 2a stage Part (Phase 2a Part), the number of cases in which the baseline RT-PCR was positive was 47 out of 69, and the number of cases in which the baseline virus titer was detected was 43. The 43 cases consisted of 375/125mg group 15 cases, 750/250mg group 14 cases and placebo group 14 cases.

The hospital Day specified in the test protocol is denoted by Visit, and the correspondence and tolerance between the Day of administration (Day) are as follows. Op V refers to an Optional hospital date (optionvisual) indicating an Optional hospital date.

TABLE 9

The mean value of each group of SARS-CoV-2 viral titers versus baseline variability is shown in FIG. 4, with respect to the modified intent-to-treat population (all subjects who were randomized and both baseline RT-PCR and viral titers positive). In this analysis, the SARS-CoV-2 infected person with mild/moderate symptoms and the asymptomatic SARS-CoV-2 infected person are included in the analysis, and only Visit, which is the necessary date of hospital, is shown. In addition, the viral titers were below the lower detection limit (0.8 log ₁₀ (TCID ₅₀ Per mL)) as 0.8log ₁₀ (TCID ₅₀ /mL). In the 375/125mg group, at the time point of Visit 3 (day 4 after start of dosing), the viral titers were statistically significantly reduced in the 750/250mg group at the time points of Visit 2 (day 2 after start of dosing) and Visit 3 (day 4 after start of dosing) at a significance level of 0.05 compared to the placebo group. Among the RT-PCR measurement results and the virus titer measurement results obtained before the time point of day 17 of 1 month 2022, the virus titer was found to be decreased in the 375/125mg group at all time points after Visit 3 (4 th day after initiation of administration) and in the 750/250mg group at all time points after Visit 2 (2 nd day after initiation of administration) compared with the placebo group.

In addition, the proportion of positive patients with viral titers at each time point is shown below.

At the Day4 time point, the ratio of positive patients with a viral titer of 0.8 or higher in the placebo group was reduced by 80% in the 750/250mg group and by 63% in the 375/125mg group. At the Day6 time point, the proportion of positive patients with a viral titer above 0.8 in the placebo group was reduced by 54% in the 750/250mg group and by 100% in the 375/125mg group.

At the time points of Day4 and Day6, a tendency was observed that the proportion of patients positive for the viral titer was low compared to the placebo group in both the 750/250mg group and the 375/125mg group. Thus, it has been shown that patients who are expelling infectious viruses can be rapidly reduced by administering the pharmaceutical composition of the present invention.

Results of the secondary evaluation items

(1) Time until the first confirmation of virus titer was negative (Phase 2a Part)

The time until the virus titer of SARS-CoV-2 was confirmed to be negative for the first time is shown in Table 10 and FIG. 5 below.

The results in the 375/125mg group 15, 750/250mg group 13, placebo group 14 are shown in 69 cases of the 2a Phase section (Phase 2a Part).

TABLE 10

[ stratification log-rank test, subject layer (mild/moderate, asymptomatic/mild only) was used as stratification factor ]

As shown in table 10, in the 375/125mg group, the median was short compared to the placebo group, 49.8 hours, and a significant difference was observed for the time until the virus titer negative was confirmed for the first time (p= 0.0159). In the 750/250mg group, the median was short, 48.4 hours compared to the placebo group, and a significant difference was observed for the time until the first confirmation of virus titer negative (p=0.0205).

In addition, as shown in fig. 5, the time until 50% of patients became negative in virus titer in placebo group was about 4.6 days (111.1 hours) after the start of treatment. In contrast, in the 750/250mg group and 375/125mg group, the time until the virus titer of 50% of patients became negative was about 2.6 days after the start of the treatment (61.3 to 62.7 hours). Thus, it was confirmed that the time until 50% of patients were negative for the first time was shortened by about 2 days.

(2) The amount of change from baseline in the total score of 12 symptoms of COVID-19 at each time point (Phase 2a Part)

The amount of change from baseline in the total score for 12 symptoms of covd-19 at each time point is shown in fig. 6.

Of 69 cases in the 2a Phase section (Phase 2a Part), 375/125mg group 13 cases, 750/250mg group 12 cases, and placebo group 14 cases of subjects with mild/moderate symptoms are shown.

As shown in FIG. 6, there was found a tendency that the values of the total scores of 12 symptoms of COVID-19 improved in the 375/125mg group and 750/250mg group, compared to the placebo group, at all time points after Day 2 (after 1 administration).

(3) Inhibition of severe Effect (2 a Phase 2a Part)

The proportion of subjects whose scores were first deteriorated to 3 or more in 69 cases of the 2a stage Part (Phase 2a Part) at any time point after the start of administration on the order scale (order scale) of the order of 8 levels of clinical severity (table 11) is shown in table 12.

TABLE 11

TABLE 12

	375/125mg group	750/250mg group	Placebo group
				Deterioration rate	0.0％	0.0％	14.3％
Number of deterioration cases/number of analysis target cases	0/13	0/12	2/14

As shown in Table 12, in 375/125mg group and 750/250mg group, no cases were found in which the order scale (Ordinal scale) was deteriorated to 3 or more for the first time after the start of administration.

Results of the Main evaluation project

(1) Variation of SARS-CoV-2 viral titer from baseline at Day4 time point (2 b Phase 2b Part)

In Phase 2b, part 428 cases were randomized and 341 cases of SARS-CoV-2 virus titer were detected at baseline as the ITT population. The 341 cases consisted of 375/125mg group 114 cases, 750/250mg group 116 cases, and placebo group 111 cases.

The amount of change in SARS-CoV-2 virus titer (ITT population) at time point of Day4 from baseline is shown in the following table by group.

TABLE 13

As shown in the above table, in 375/125mg group and 750/250mg group, a statistically significant decrease in viral titers (p < 0.0001) was confirmed at Day4 time point compared to placebo group. It is thus shown that by administering the pharmaceutical composition of the present invention, a remarkably excellent antiviral effect is exhibited.

The proportion of positive patients with a viral titer of 1.1 or more at Day4 time point is shown below.

In the placebo group, the proportion of positive patients with viral titers was 50.5%, whereas in the 375/125mg group, the proportion of positive patients with viral titers was 1.9%, and in the 750/250mg group, the proportion of positive patients with viral titers was 5.4%.

Day4 time points, in both the 375/125mg group and the 750/250mg group, found a significantly lower trend for the proportion of patients positive for viral titers than the placebo group. Thus, it has been shown that patients who are expelling infectious viruses can be rapidly reduced by administering the pharmaceutical composition of the present invention.

(2-a) amount of change per unit time of 12 symptom total score of COVID-19 from baseline to 120 hours (2 b Phase 2b Part)

Of 341 cases in the 2b Phase Part (Phase 2b Part), 12 symptom total scores of covd-19 were observed at the base line and after the start of administration, and 332 cases were counted in which 12 symptom total scores of covd-19 at the base line were 1 or more, and the 332 cases were constituted by 375/125mg group 109 cases, 750/250mg group 113 cases, and placebo group 110 cases.

The change per unit time (ITT population) of the total score of 12 symptoms of COVID-19 from baseline to 120 hours is shown in the table below.

TABLE 14

As shown in the above table, in 375/125mg group and 750/250mg group, the decrease trend of the numerical value was confirmed but no statistically significant difference was confirmed as compared with placebo group.

(2-b) amount of change per unit time of sub-total score of symptoms of COVID-19 from baseline to 120 hours (2 b Phase 2b Part)

The 12 symptoms of the covd-19 shown in (2-a) above were classified into 5 (acute symptoms, main clinical symptoms, respiratory symptoms, systemic symptoms and digestive organ symptoms), and the results of the analysis (ITT population) are shown in the following table.

Acute symptoms: sore throat, cough, fever or fever

Major clinical symptoms: runny nose or stuffy nose, sore throat, cough, chills/sweating, feeling hot or fever

Systemic symptoms: tiredness (tiredness), muscle pain or body pain, headache, chills/sweating, feeling fever or fever

Digestive organ symptoms: nausea, vomiting and diarrhea

TABLE 15-1

[ Table 15-2]

[ Table 15-3]

[ tables 15-4]

[ tables 15-5]

As shown in the above table, among the 12 symptoms, among respiratory symptoms (runny nose or nasal obstruction, sore throat, cough, shortness of breath (dyspnea)) which are characteristic symptoms in the group aggregated in the test, a significant improvement effect was confirmed in 375/125mg group and 750/250mg group compared with placebo group (375/125 mg group: p=0.0153, 750/250mg group: p=0.0032).

The results of the validity are illustrated centering on the 375/125mg group results used in the validation of the validity.

Results of the Main evaluation project

(1) Time until recovery of 5 symptoms of COVID-19 (Phase 3 Part)

Of the ITT population of the mild/moderate SARS-CoV-2 infected persons in the 3-stage fraction (Phase 3 Part), 375/125mg group 347 cases and placebo group 343 cases, 375/125mg group 603 cases and placebo group 600 cases were the population of less than 72 hours from the onset of COVID-19 as the main analysis target population.

The time to recovery of 5 symptoms of COVID-19 and its Kaplan-Meier curves in light/medium SARS-CoV-2 infected persons (persons less than 72 hours from the onset of COVID-19 in the ITT group) are shown in Table 16 and FIG. 7 below.

TABLE 16

5 symptoms (runny nose or stuffy nose, sore throat, cough, feeling of fever or fever, hypodynamia (tiredness))

[a] The maximum value contains the deleted data.

[b] Layering factor whether to vaccinate SARS-CoV-2 vaccine [ Yes, no ]

As shown in the above table, a reduction of 24.3 hours (about 1 day) was observed in the median value of 375/125mg group as compared with placebo group, in the time until recovery of 5 symptoms of COVID-19 in the light/medium SARS-CoV-2 infected persons (the persons group of ITT groups, whose time from onset of COVID-19 to random group was less than 72 hours). In addition, statistically significant differences were shown (Peto-predce's hierarchical generalized Wilcoxon test: unilateral p=0.0204).

It was thus shown that by administering the pharmaceutical composition of the present invention, a remarkably excellent effect of improving clinical symptoms is exhibited.

The time to recovery of 5 symptoms of covd-19 in mild/moderate SARS-CoV-2 infected individuals (ITT population) and their Kaplan-Meier curves are shown in table 17 below and fig. 8.

TABLE 17

[a] The maximum value contains the deleted data.

[b] Layering factor-time from onset of COVID-19 to random grouping [ less than 72 hours, 72 hours or more ], whether SARS-CoV-2 vaccine is vaccinated [ Yes, no ]

As shown in the above table, no significant difference was confirmed in the time to recovery of 5 symptoms of COVID-19 in the mild/moderate SARS-CoV-2 infected person (ITT population). The time until recovery of 5 symptoms of covd-19 was found to be 10.6 hours shorter than that of the placebo group, which was a median value in 375/125mg group.

Results of primary secondary evaluation items

(1) Variation of the amount of SARS-CoV-2 viral RNA from Day 4 from baseline (Phase 3 Part)

The amount of change from baseline in the viral RNA amount in Day 4 in the light/medium disease SARS-CoV-2 infected person (the group of ITT groups whose time from onset of COVID-19 to random grouping was less than 72 hours) is shown in Table 18 below.

TABLE 18

Units log ₁₀ copy/mL (copies/mL)

The lower limit of the amount of viral RNA of SARS-CoV-2 is 2.08log ₁₀ copy/mL

When the RT-PCR of SARS-CoV-2 is negative, the treatment is carried out according to 2.27log10 copies/mL, and when the viral RNA amount of SARS-CoV-2 is lower than the lower limit value of quantification, the treatment is carried out according to 2.08log ₁₀ copy/mL processing

[a] Covariates-whether or not the baseline viral RNA amount of SARS-CoV-2 was vaccinated with SARS-CoV-2 vaccine [ Yes, no ]

[b] Layering factor whether to vaccinate SARS-CoV-2 vaccine [ Yes, no ]

As shown in the above table, the modified estimate of the amount of change in the viral RNA amount of SARS-CoV-2 in Day 4 showed about 1.5log in 375/125mg group as compared to placebo group ₁₀ Reduction in (copy/mL) confirmed significant differences (covariance analysis: one-sided p)<0.0001)。

It was thus shown that by administering the pharmaceutical composition of the present invention, a remarkably excellent antiviral effect is exhibited.

The amount of change from baseline in the amount of viral RNA in Day 4 in the febrile/moderate SARS-CoV-2 infected person (ITT population) is shown in Table 19 below.

TABLE 19

Units log ₁₀ copy/mL

When the RT-PCR of SARS-CoV-2 was negative, the sample was taken at 2.27log ₁₀ copy/mL treatment, when the amount of viral RNA of SARS-CoV-2 is below the lower limit of quantification, the amount is 2.08log ₁₀ copy/mL processing

[a] Covariates-the amount of viral RNA of SARS-CoV-2 at baseline, the time from onset of COVID-19 to random grouping [ less than 72 hours, 72 hours or more ], whether to vaccinate with SARS-CoV-2 vaccine [ yes, no ]

As shown in the above table, the modified estimate of the amount of change in the viral RNA amount of SARS-CoV-2 in Day 4 showed 1.2log in 375/125mg group as compared to placebo group ₁₀ (copy/mL) reduction.

(2) Time until the first confirmation of virus titer was negative (Phase 3 Part)

The time until the first confirmation of virus titer negativity among the febrile/moderate SARS-CoV-2 infected persons (persons having a time period from onset of COVID-19 to random grouping of less than 72 hours in the mITT group) and the Kaplan-Meier curves thereof are shown in Table 20 and FIG. 9 below.

TABLE 20

[a] The maximum value contains the deleted data.

[b] Layering factor whether to vaccinate SARS-CoV-2 vaccine [ Yes, no ]

As shown above, a reduction of 29.1 hours (about 1 day) was observed in 375/125mg group as a median value, and a statistically significant difference was confirmed (stratified log-rank test: single-sided p < 0.0001) in the time until the virus titer of SARS-CoV-2 was first confirmed to be negative, as compared with placebo group.

The time until the first confirmation of the negative viral titers in the febrile/moderate SARS-CoV-2 infected individuals (mITT population) and their Kaplan-Meier curves are shown in Table 21 below and FIG. 10.

TABLE 21

[a] The maximum value contains the deleted data.

As shown above, a reduction of 37.5 hours (about 1.5 days) was observed in 375/125mg group as a median value, as compared to placebo group, until the virus titer of SARS-CoV-2 was first confirmed to be negative.

Results of the secondary evaluation items

(1) Time until recovery of 12 symptoms of COVID-19 (Phase 3 Part)

The time to recovery of 12 symptoms of COVID-19 and its Kaplan-Meier curve in the light/medium SARS-CoV-2 infected persons (persons less than 72 hours from the onset of COVID-19 to the random grouping in ITT group) are shown in Table 22 and FIG. 11 below.

TABLE 22

Symptoms of symptoms (debilitation (tiredness), muscle or body pain, headache, chills/sweating, fever or fever, runny nose or stuffy nose, sore throat, cough, shortness of breath (dyspnea), nausea, vomiting, diarrhea)

[a] The maximum value contains the deleted data.

[b] Layering factor whether to vaccinate SARS-CoV-2 vaccine [ Yes, no ]

As shown above, a tendency to shorten by 34.0 hours (about 1.5 days) was observed in 375/125mg group as a median value in the time until recovery of 12 symptoms of COVID-19, compared with placebo group.

(2) Time until recovery of 14 symptoms of COVID-19 (Phase 3 Part)

The time to recovery of 14 symptoms of COVID-19 and its Kaplan-Meier curve among the feeble/moderate SARS-CoV-2 infected persons (persons less than 72 hours from the onset of COVID-19 in the ITT group) are shown in Table 23 and FIG. 12 below.

TABLE 23

14 symptoms (fatigue, muscle or body pain, headache, chills/sweating, fever or fever, runny nose or stuffy nose, sore throat, cough, shortness of breath (dyspnea), nausea, vomiting, diarrhea, abnormal taste, abnormal smell)

[a] The maximum value contains the deleted data.

[b] Layering factor whether to vaccinate SARS-CoV-2 vaccine [ Yes, no ]

As shown above, a reduction of 44.1 hours (about 2 days) was observed in 375/125mg group as a median value, as compared to placebo group, until recovery of 14 symptoms of COVID-19.

Frequency of occurrence of adverse events

In the 2a Phase Part (Phase 2a Part), there are no deaths, serious adverse events and adverse events that lead to discontinuation of administration.

In addition, in the 2b Phase Part (Phase 2b Part), there are no adverse events leading to death. In the placebo group, 2 severe adverse events were observed, but in the active drug group no one was observed. The adverse events leading to discontinuation of administration were confirmed in 2 of 375/125mg groups (active drug groups), and were judged to be causally related to the test drug.

In the 3-Phase Part (Phase 3 Part), there are no adverse events leading to death. 1 of the severe adverse events were confirmed in 375/125mg group and placebo group, respectively, and were judged to have no causal relationship with the test drug. Among adverse events leading to discontinuation of administration, 375/125mg of eczema and vomiting, 750/250mg of rash (2 pieces), placebo of muscle weakness and hypoesthesia were judged to be causally related to the test drug, but all abated or recovered after discontinuation of administration of the test drug.

Test example 5 clinical trials for children

Evaluation of safety and pharmacokinetics in repeated oral administration of a test agent (active ingredient: fumaric acid co-crystal form I of a compound represented by formula (I)) to a patient suffering from SARS-CoV-2 infection in a mild/moderate state over 6 years and less than 12 years was carried out by a double-blind comparison test based on a random group of placebo as a comparison subject.

The main evaluation items of the test were adverse events, clinical examinations, vital signs (vital signs) as safety, and Day 2 and Day 6 plasma concentrations as pharmacokinetics.

Patients were selected that fully met the criteria described below.

(a) Male or female patients over 6 years old and less than 12 years old.

(b) People diagnosed as SARS-CoV-2 positive within 72 hours prior to registration.

(c) The time from onset of covd-19 to registration was less than 72 hours.

(d) A person who has one or more of the following symptoms (12 symptoms of COVID-19) caused by COVID-19 and has a moderate (COVID-19 symptom score: 2) or more (excluding symptoms existing before the onset of COVID-19) at the time of registration.

Digestive organ symptoms: nausea, vomiting and diarrhea

(e) A person weighing more than 20 kg.

Method for administering test agent

(i) Test agent

25mg tablet: the tablet contains fumaric acid co-crystal form I crystals of the compound represented by the formula (I), and 25mg of the fumaric acid co-crystal form I crystals are contained in the tablet based on the compound represented by the formula (I).

(ii) Placebo

Placebo-a tablets: a tablet which is indistinguishable in appearance from the above 25mg tablet and does not contain fumaric acid co-crystal form I crystals of the compound represented by the formula (I).

Administration amount and administration method

Subjects to be judged as eligible were classified as 2 in each weight class: the ratio of 1 was randomly assigned to one of the test, placebo groups.

The administration of the test agent was repeated 1 day for 5 days with 1 time. In the weight category of 40kg or more, 375mg was administered as a load dose to Day 1, and 125mg was administered as a maintenance dose to Day 2 to 5. In the weight category of 30kg or more and less than 40kg, 250mg was administered as a load dose in Day 1, and 125mg was administered as a maintenance dose in Day 2 to 5. In the weight category of 20kg or more and less than 30kg, 125mg was administered as the load amount in Day 1, and 75mg was administered as the maintenance amount in Day 2 to 5.

Further, the following amounts are also given.

In the weight category of 40kg or more, 375mg was administered as a load dose to Day 1, and 125mg was administered as a maintenance dose to Day2 to 5. In the weight category of 30kg or more and less than 40kg, 250mg is taken at Day 1 as a load amount, and 100mg is taken at Day2 to 5 as a maintenance amount. In the weight category of 20kg or more and less than 30kg, 150mg was administered as the load dose in Day 1, and 75mg was administered as the maintenance dose in Day2 to 5.

Test agent by weight class

Test agent group

40kg or more

In Day 1, 3 125mg tablets were administered, and in Day2 to 5,1 Day 1, 125mg tablets were administered.

30kg or more and less than 40kg

In Day 1, 2 125mg tablets were administered, and in Day2 to 5,1 Day 1, 125mg tablets were administered.

20kg or more and less than 30kg

In Day 1, 1 tablet of 125mg was administered, and in Day2 to 5, 3 tablets of 25mg were administered.

Further, the following amounts are also given.

40kg or more

30kg or more and less than 40kg

In Day 1, 2 pieces of 125mg were applied, and in Day2 to 5, 4 pieces of 25mg were applied.

20kg or more and less than 30kg

In Day 1, 1 tablet of 125mg and 25mg each was applied, and in Day2 to 5, 3 tablets of 25mg were applied.

Placebo group

40kg or more

At Day 1, 3 placebo-B tablets were administered and at Day 2-5, 1 Day 1 placebo-B tablets were administered.

30kg or more and less than 40kg

At Day 1, 2 placebo-B tablets were administered and at Day 2-5, 1 Day 1 placebo-B tablets were administered.

20kg or more and less than 30kg

At Day 1, 1 placebo-B tablet was administered and at Day 2-5, 3 placebo-a tablets were administered for 1 Day.

Further, the following amounts are also given.

40kg or more

30kg or more and less than 40kg

At Day 1, 2 placebo-B tablets were administered and at Day 2-5, 4 placebo-a tablets were administered for 1 Day.

20kg or more and less than 30kg

At Day 1, placebo-B and placebo-a were administered in 1 tablet, and at Day 2-5, 3 days, placebo-a was administered.

Main evaluation item of safety

Adverse events, clinical examinations, vital signs

Principal evaluation items of pharmacokinetics

Plasma concentration of Day 2 and Day 6 in the test agent

Analysis of the main evaluation items: safety of

Adverse events were classified according to the organ class large classification and primitive based on ICH med dra. Among the adverse events reported by eCRF, adverse events occurring after the administration of the first test drug were used for safety analysis.

TEAE, death, other severe TEAE, the number of occurrences of TEAE that led to discontinuation of administration, and their proportions were pooled for each administration amount. The 95% confidence interval for their occurrence ratio was calculated by the Clopper-Pearson method. The number of reported adverse events is also shown. Adverse events that were judged to be "related" to the causal relationship of the test drug were summarized as side effects, as well as TEAE.

After randomization, summary statistics are calculated for each administration group for observations at predetermined measurement time points and amounts of change from baseline at each time point. For qualitative clinical exam data, a table of baseline and predetermined measured time points offset is made for the exam category.

After randomization, summary statistics are calculated for each administration group for observations at predetermined measurement time points and amounts of change from baseline at each time point.

Analysis of the main evaluation items: a list of the plasma concentrations of the compound represented by formula (I) was prepared for the pharmacokinetic PK concentration population, based on the body weight type and Day x (Day), together with the elapsed time from the start of administration of the test drug immediately before blood collection until blood collection. In Day 2 and Day 6, the concentrations of the compound represented by formula (I) in plasma at the time point of 24 hours after administration were accumulated, and N, mean, SD, the coefficient of variation, the geometric mean, the coefficient of variation, the median, the minimum, and the maximum were summarized.

Test example 6 in-home preventive administration clinical trial

The test is a randomized, double-blind, multi-center, parallel-group, placebo-controlled comparative test targeting about 2040 symptomatic COVID-19 first-onset patients in the home with co-occupants. Subjects were treated with 1:1 were randomly assigned to 375/125mg or placebo groups of the test drug. For the proportion of subjects who had developed symptoms of COVID-19 who had been infected with SARS-CoV-2 (confirmed to be RT-PCR positive at the central inspection agency) before Day 10.

Symptomatic covd-19 first onset patients select patients who meet the criteria described below.

Age-free restriction

The first COVID-19 patient in the co-resident in the home within 6 months before the consent was obtained

Is a person who confirms 1 or more symptoms of COVID-19 within 24 hours from the time of consent

Human being SARS-CoV-2 positive in airway samples taken 72 hours prior to randomization

The co-resident in the home selects a subject that meets the criteria described below.

Over 12 years old

SARS-CoV-2 negative person when screening by SARS-CoV-2 assay Using airway sample

People who are not judged by the experimentally responsible physician to be infected with SARS-CoV-2

Living together with the first onset patient, living together with the same living family to the end of the trial

Method for administering test agent

(i) Test agent

(ii) Placebo

Placebo tablets: a tablet which is indistinguishable in appearance from the 125mg tablet described above and does not contain fumaric acid co-crystal form I crystals of the compound represented by the formula (I).

Administration amount and administration method

Symptomatic covd-19 patients with first onset were treated with 1: the ratio of 1 was randomly assigned to one of the test, placebo groups.

The administration of the test agent was repeated 1 day for 5 days with 1 time. 375mg was administered at Day 1 as the loading dose, and 125mg was administered at Day 2 to 5 as the maintenance dose.

After 5 days of trial administration, a follow-up period of 10 days was used.

Test drugs for each administration group

Test agent group

In Day 1, 3 125mg tablets were administered, and in Day 2 to 5,1 Day 1, 125mg tablets were administered.

Placebo group

At Day 1, 3 placebo tablets were administered, and at Day 2-5, 1 Day 1 placebo tablet was administered.

The main evaluation items were the proportion of subjects who had been infected with SARS-CoV-2 (confirmed to be RT-PCR positive at the central inspection agency) before Day 10, who developed symptoms of COVID-19.

A subject who had developed symptoms of COVID-19, who had been infected with SARS-CoV-2 (confirmed by the central inspection agency to be positive by RT-PCR) before Day 10, was defined as a subject in the mITT population who met two of the following 2 criteria.

Confirmation of infection with SARS-CoV-2 before Day 10 after first administration of test drug by RT-PCR positive judgment by the Central inspection agency

Symptoms of COVID-19 appear before Day 10 after the first administration of the test drug (see definition below)

The definition of the occurrence of a symptom of covd-19 in a subject meets any 1 or more of the following criteria:

confirming any 1 or more of the following symptoms (or worsening of the same in case of confirming a covd-19 like symptom at baseline): (a) Fever or (b) shortness of breath or dyspnea

Confirming more than 2 of the following 7 types of covd-19 symptoms for more than 24 hours (or worsening in case of a baseline confirming covd-19 like symptoms): (1) runny nose or nasal obstruction, (2) sore throat, (3) cough, (4) fever, 5) debilitation, (6) abnormal gustatory sensation, and (7) abnormal smell

Analysis of the main evaluation items: with respect to the principal analysis, a corrected poisson regression model using the robust variance is utilized. Estimating the risk ratio of the test drug group to the placebo group, calculating the p-value of null hypothesis (null hypothesis) with the risk ratio of 1, and the 95% Confidence Interval (CI) of the risk ratio using a corrected poisson regression model. The model contained as covariates the time (less than 48 hours, more than 48 hours) until the subject group was entered after the first onset of the patient's symptoms of covd-19, and the geographic region (north america, south america, europe, africa, japan, asia outside japan).

Preparation example

The formulation examples shown below are merely examples and are not intended to limit the scope of the invention in any way.

Formulation example 1 suspension

To a raw material of the compound represented by the formula (I), for example, water for injection is added to prepare a suspension.

Formulation example 2 tablet

To a raw material of a compound represented by the formula (I), for example, D-mannitol or magnesium stearate is added as an additive to prepare a tablet.

Formulation example 3 tablet

To 125mg (152.3 mg of fumaric acid co-crystal form I of the compound represented by the formula (I)) of the compound represented by the formula (I) was added D-mannitol, croscarmellose sodium, hydroxypropylcellulose, light silicic anhydride, crystalline cellulose, and magnesium stearate as additives, and the mixture was prepared into tablets.

Formulation example 4 tablet

To 25mg (30.46 mg of fumaric acid co-crystal form I of the compound represented by the formula (I)) of the compound represented by the formula (I) was added D-mannitol, croscarmellose sodium, hydroxypropylcellulose, light silicic anhydride, crystalline cellulose, and magnesium stearate as additives, and the mixture was prepared into tablets.

Preparation example 5 granule

The raw material of the compound represented by the formula (I) is added with, for example, D-mannitol and magnesium stearate as additives to prepare granules.

Preparation example 6 granule

To 125mg (152.3 mg of fumaric acid co-crystal form I of the compound represented by the formula (I)) of the compound represented by the formula (I) was added D-mannitol, powder reduced maltose syrup (maltitol), croscarmellose sodium, hydroxypropylcellulose, light silicic anhydride, magnesium stearate, and sucralose as additives, and the mixture was prepared into granules.

Industrial applicability

The novel method for treating coronavirus infectious disease (covd-19) and the therapeutic pharmaceutical composition used therein according to the present invention are considered to exhibit excellent therapeutic effects by administering a prescribed amount of a complex comprising a compound represented by formula (I) and fumaric acid as active ingredients to a patient. In addition, the present invention provides a highly safe pharmaceutical composition for covd-19 therapy which can maintain an effective blood level without exceeding NOAEL by administration in a specific administration amount.

Claims

1. A pharmaceutical composition for treating novel coronavirus infectious disease (COVID-19) comprising a complex comprising a compound represented by formula (I) and fumaric acid as active ingredients,

[ chemical formula 1]

2. The pharmaceutical composition of claim 1, wherein the patient suffering from novel coronavirus infectious disease (covd-19) is a patient classified as asymptomatic, mild or moderate I.

3. A pharmaceutical composition according to claim 1 or 2 for use in shortening the course of a novel coronavirus infectious disease (covd-19).

4. A pharmaceutical composition according to any one of claims 1 to 3 for inhibiting viral proliferation of SARS-CoV-2.

5. The pharmaceutical composition according to any one of claims 1 to 4, wherein the amount of the active ingredient administered per 1 day of the pharmaceutical composition is 20mg to 2000mg.

6. The pharmaceutical composition according to any one of claims 1 to 5, wherein the amount of the active ingredient administered on day 1 of treatment is 375mg and the amounts of the active ingredient administered on the second to fifth days of treatment are 125mg, respectively.

7. The pharmaceutical composition according to any one of claims 1 to 5, wherein the amount of the active ingredient administered on day 1 of treatment is 750mg and the amounts of the active ingredient administered on the second to fifth days of treatment are 250mg, respectively.

8. The pharmaceutical composition of any one of claims 1-7, wherein administration is once a day.

9. The pharmaceutical composition of any one of claims 1-8, which is an oral formulation.