RU2825648C1 - Methods of treating coronavirus infections - Google Patents

Abstract

FIELD: chemistry; pharmaceutics.

SUBSTANCE: invention relates to a method of inhibiting a viral infection in a patient in need thereof, comprising administering to the patient a therapeutically effective amount of compound 1 or a pharmaceutically acceptable salt thereof, wherein the viral infection is a coronavirus infection caused by SARS-CoV-2. Invention also relates to use of a pharmaceutical composition for inhibiting coronavirus infection in a person in need thereof, to the use of compound 1 in inhibiting infection caused by SARS-CoV-2in humans, to the use of compound 1 in the manufacture of a medicinal product for inhibiting coronavirus infection.

EFFECT: inhibition of viral infection caused by SARS-CoV-2.

15 cl, 10 dwg, 4 tbl, 3 ex

Description

FIELD OF TECHNOLOGY TO WHICH THE INVENTION RELATES

[0001] The present invention relates to methods for treating patients with coronavirus infections, including patients suffering from coronavirus disease 2019 (COVID-19).

BACKGROUND OF THE INVENTION

[0002] Coronaviruses are a large family of viruses characterized by club-shaped spikes protruding from their surfaces, unusually large RNA genomes, and a unique replication strategy. Fehr et al., Methods Mol. Biol. , 2015, 1282: 1–23. Coronaviruses typically cause a range of diseases in mammals and birds ranging from mild to moderate upper respiratory tract illness, including those caused by coronaviruses 229E, NL63, OC43, and HKU1. Common Human Coronaviruses, Centers for Disease Control and Prevention, available at https://www.cdc.gov/coronavirus/general-information.html (last accessed April 27, 2020). However, in the past two decades, new coronaviruses have emerged from animal reservoirs that cause severe and widespread illness and death in humans. Coronaviruses, National Institute of Allergy and Infectious Diseases, available at https://www.niaid.nih.gov/diseases-conditions/coronaviruses (last accessed April 27, 2020).

[0003] In 2002 and 2003, outbreaks of highly pathogenic severe acute respiratory syndrome coronavirus (SARS-CoV) caused over 8,000 cases worldwide with a case fatality rate of ~10%. In 2012, Middle East respiratory syndrome coronavirus (MERS-CoV) caused 2,500 confirmed cases with a case fatality rate of 36%. Wit et al., Nature Reviews Microbiology , 2016, 14, 523–534.

[0004] In 2019, a novel coronavirus named SARS-CoV-2 emerged and caused an acute respiratory syndrome in humans called coronavirus disease 2019 (COVID-19). The World Health Organization declared a global pandemic on March 11, 2020. Coronaviruses, National Institute of Allergy and Infectious Diseases, available at https://www.niaid.nih.gov/diseases-conditions/coronaviruses (last accessed April 27, 2020). As of April 26, 2020, there were a total of 2,992,970 confirmed cases of COVID-19 and 207,518 confirmed deaths caused by SARS-CoV-2. John Hopkins University Coronavirus Resource Center, available at https://coronavirus.jhu.edu/map.html (last updated April 27, 2020, 10:31:17 AM).

[0005] Treatment of coronavirus infection, especially COVID-19, is an urgent need. However, there are currently no clinically available antiviral drugs that can effectively treat diseases caused by SARS-CoV, SARS-CoV-2, or MERS-CoV. An effective approach to discovering new drugs is to test existing antiviral drugs or drug candidates for new indications. Given the development timeframe and manufacturing requirements for new drugs, repurposing existing drugs or drug candidates is a promising solution for outbreaks caused by emerging viruses.

ESSENCE OF THE INVENTION

[0006] In one aspect, the present invention relates to a method of treating a viral infection in a patient, comprising administering to the patient a therapeutically effective amount of compound 1:

or a pharmaceutically acceptable salt thereof.

[0007] In another aspect, the present invention relates to a method of treating a viral infection in a patient, comprising administering to the patient a therapeutically effective amount of compound 2:

or a pharmaceutically acceptable salt thereof.

[0008] In another aspect, the present invention relates to a method of treating a viral infection in a patient, comprising administering to the patient a therapeutically effective amount of compound 3:

or a pharmaceutically acceptable salt thereof.

[0009] In another aspect, the present invention relates to methods of treating a viral infection in a patient comprising administering to the patient a therapeutically effective amount of compound 4:

or a pharmaceutically acceptable salt thereof.

[0010] In another aspect, the present invention relates to a method of treating a viral infection in a patient, comprising administering to the patient a therapeutically effective amount of compound 5:

Compound 5,

or a pharmaceutically acceptable salt thereof.

[0011] In certain embodiments, the viral infection is caused by a coronavirus.

[0012] In certain embodiments, the viral infection is a coronavirus infection caused by a coronavirus.

[0013] In certain embodiments, the coronavirus infection is caused by SARS-CoV-2, SARS-CoV, MERS-CoV, 229E, NL63, OC43, or HKU1.

[0014] In certain embodiments, the coronavirus infection is caused by SARS-CoV-2.

[0015] In certain embodiments, the coronavirus infection is caused by SARS-CoV.

[0016] In certain embodiments, the coronavirus infection is caused by MERS-CoV.

[0017] In certain embodiments, the present invention provides a method of treating a patient suffering from COVID-19.

[0018] In certain embodiments, COVID-19 is caused by SARS-CoV-2.

[0019] In certain embodiments, the present invention provides a medicinal or pharmaceutical composition comprising compound 1 or a pharmaceutically acceptable salt thereof for the treatment of a viral infection, in particular a coronavirus infection.

[0020] In certain embodiments, the present invention provides a medicinal or pharmaceutical composition comprising Compound 2 or a pharmaceutically acceptable salt thereof for the treatment of a viral infection, in particular a coronavirus infection.

[0021] In certain embodiments, the present invention provides a medicinal or pharmaceutical composition comprising compound 3 or a pharmaceutically acceptable salt thereof for the treatment of a viral infection, in particular a coronavirus infection.

[0022] In certain embodiments, the present invention provides a medicinal or pharmaceutical composition comprising compound 4 or a pharmaceutically acceptable salt thereof for the treatment of a viral infection, in particular a coronavirus infection.

[0023] In certain embodiments, the present invention provides a medicinal or pharmaceutical composition comprising compound 5 or a pharmaceutically acceptable salt thereof for the treatment of a viral infection, in particular a coronavirus infection.

[0024] In certain embodiments, the present invention relates to the use of compound 1 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for the treatment of a viral infection, in particular a coronavirus infection.

[0025] In certain embodiments, the present invention relates to the use of compound 2 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for the treatment of a viral infection, in particular a coronavirus infection.

[0026] In certain embodiments, the present invention relates to the use of compound 3 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for the treatment of a viral infection, in particular a coronavirus infection.

[0027] In certain embodiments, the present invention relates to the use of compound 4 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for the treatment of a viral infection, in particular a coronavirus infection.

[0028] In certain embodiments, the present invention relates to the use of compound 5 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for the treatment of a viral infection, in particular a coronavirus infection.

BRIEF DESCRIPTION OF GRAPHIC MATERIALS

[0029] Fig. 1 shows the effect of compound 2 on the induction of apoptosis, IAP expression, and HBV replication and gene expression in the liver cancer cell line HepG2.2.15.

[0030] Figure 2 shows that Compound 2 promotes viral clearance in the pAAV-HBV 1.3 C57BL/6J mouse model.

[0031] Figure 3 shows that compound 2 specifically induces apoptosis in HBV-infected (HBsAg+) hepatocytes in mouse HBV models.

[0032] Figure 4 shows that Compound 2 promotes the expression of IFN-γ and TNF-α and increases the number of CD4+ and CD8+ T cells in liver tissue in HBV models.

[0033] Fig. 5 shows the procedure for testing the antiviral activity of compound 1 and compound 2 using the SARS-CoV-2 immunofluorescence assay (IFA). Imatinib, remdesivir, chloroquine, and lopinavir were used as standards.

[0034] Fig. 6 shows the results of the antiviral activity of compound 1 and compound 2 using the SARS-CoV-2 immunofluorescence assay (IFA). Imatinib, remdesivir, chloroquine, and lopinavir were used as standards.

[0035] Fig. 7 shows a method for inhibiting cytokines associated with influenza virus-induced cytokine storm.

[0036] Fig. 8 shows the results of inhibition of cytokines associated with the cytokine storm induced by the influenza virus.

[0037] Fig. 9 shows the survival of mice infected with influenza virus (IFV) treated with compound 1 or reference compounds (ponatinib, oseltamivir).

[0038] Figure 10 shows the results for compound 1 and ponatinib in the RIPK1 kinase assay. GSK2982772 was used as a reference.

DETAILED DESCRIPTION OF THE INVENTION

[0039] All published documents cited in this document are incorporated herein by reference in their entirety.

[0040] Unless defined otherwise, 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.

[0041] The term "approximately" is used herein to mean approximately, on the order of, roughly, or about. When used in conjunction with a numerical range, the term "approximately" modifies the range by extending the limits upward and downward from the stated numerical values. Generally, the term "approximately" is used herein to modify a numerical value upward and downward from the stated value by a deviation of 10%.

[0042] The term "comprises" means "including without limitation."

[0043] The terms "treating," "treating," and "treating" as used herein refer to reversing, alleviating, delaying the onset of, or suppressing the progression of a disease or disorder or one or more symptoms thereof, including, but not limited to, therapeutic benefit. In some embodiments, treatment is performed after the development of one or more symptoms. In some embodiments, treatment can be performed in the absence of symptoms. For example, treatment can be performed on a subject before symptoms develop (e.g., in light of a history of symptoms and/or in light of genetic or other predisposing factors). Treatment can also be continued after symptoms have resolved, for example, to prevent or delay their recurrence.

[0044] A therapeutic benefit includes eradicating and/or ameliorating the manifestations of the underlying disorder being treated, such as cancer; it also includes eradicating and/or ameliorating the manifestations of one or more symptoms associated with the underlying disorder, such that the subject experiences an improvement, although the subject may still be affected by the underlying disorder. In some embodiments, "treating" or "treating" includes one or more of the following: (a) suppressing the disorder (e.g., reducing one or more symptoms caused by the disorder and/or reducing the severity of the disorder); (b) slowing or halting the progression of one or more symptoms associated with the disorder (e.g., stabilizing the disorder and/or delaying the worsening or progression of the disorder); and/or (c) alleviating the disorder (e.g., causing regression of clinical symptoms, reducing the intensity of manifestations of the disorder, delaying the progression of the disorder, and/or improving quality of life).

[0045] As used herein, "administering" or "administering" a compound of formula (I) or formula (I-A) or a pharmaceutically acceptable salt thereof encompasses delivering the compound or a pharmaceutically acceptable salt or prodrug or other pharmaceutically acceptable derivative thereof to a patient using any suitable formulation or route of administration, such as described herein.

[0046] The term "therapeutically effective amount" or "effective amount" as used herein refers to an amount that is effective to elicit a desired biological or medical response, including an amount of a compound that, when administered to a subject to treat a disorder, is sufficient to effect such treatment of the disorder. An effective amount will vary depending on the disorder and its severity, as well as the age, weight, etc. of the subject to be treated. An effective amount may be presented in a single or multiple doses (e.g., a single dose or multiple doses may be required to achieve the desired treatment endpoint). An effective amount may be considered to be administered in an effective amount if, in combination with one or more other agents, a desired or beneficial result can be achieved or is achieved. Suitable doses of any co-administered compounds may optionally be reduced due to the combined effect, additive, or synergistic effect of the compound.

[0047] As used herein, a "patient" to whom administration is intended includes, without limitation, humans (i.e., a male or female of any age group, such as a pediatric subject (e.g., an infant, a child, an adolescent) or an adult subject (e.g., a young adult, a middle-aged adult, or an elderly adult)) and/or other primates (e.g., cynomolgus monkeys, rhesus monkeys).

[0048] As used herein, "pharmaceutically acceptable" or "physiologically acceptable" refers to compounds, salts, compositions, dosage forms, and other materials that are useful in the preparation of a pharmaceutical composition suitable for pharmaceutical use in veterinary or human medicine.

[0049] The term "pharmaceutically acceptable salt" as used herein refers to those salts that are suitable, within the limits of sound medical judgment, for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic reaction, etc., commensurate with a reasonable benefit/risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, SM Berge et al. describe pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences , 1977, 66, 1-19. Pharmaceutically acceptable salts of compound 1 include salts prepared from suitable inorganic and organic acids and bases. Examples of pharmaceutically acceptable non-toxic acid addition salts are salts at the amino group formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid, or with organic acids such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid, or using other methods used in the art such as ion exchange. Other pharmaceutically acceptable salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecyl sulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxyethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate salts, etc. Although pharmaceutically acceptable counterions will be preferred for the preparation of pharmaceutical compositions, other anions are quite acceptable as intermediates in the synthesis. Thus, these may be pharmaceutically undesirable anions such as iodide, oxalate, trifluoromethanesulfonate, etc., if such salts are intermediate chemical compounds.

[0050] The term "pharmaceutically acceptable carrier" is used herein to mean a material that is compatible with the recipient subject, preferably a mammal, more preferably a human, and is suitable for delivering the active agent to the target site without terminating the activity of the agent. The toxicity or undesirable effects associated with the carrier, if any, should preferably be proportionate to a reasonable risk/benefit ratio for the intended use of the active agent.

[0051] The term "orally" refers to the administration of a composition intended for oral administration. Examples of oral forms include, but are not limited to, tablets, pills, capsules, powders, granules, solutions or suspensions, and drops. Such forms may be swallowed whole, or they may be presented in chewable form.

[0052] The term "alkyl" means a branched chain or straight chain alkyl group with a specified number of carbon atoms. For example, the definition "C ₁ -C ₅ " in "C ₁ -C ₅ alkyl" means a straight chain or branched chain alkyl group with 1, 2, 3, 4 or 5 carbon atoms. For example, "C ₁ -C ₅ alkyl" includes methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, isobutyl, pentyl, etc.

[0053] The term "cycloalkyl" refers to a specific single saturated alkyl ring with a specified number of carbon atoms. For example, "cycloalkyl" includes cyclopropyl, methylcyclopropyl, 2,2-dimethylcyclobutyl, 2-ethylcyclopentyl, cyclohexyl, etc.

[0054] The term "heterocycle" refers to an aromatic or non-aromatic ring containing 5-6 atoms in which 1-4 heteroatoms such as O, N, S are contained. "Heterocycle" includes the heteroaromatic ring mentioned above; it also includes the dihydro and tetrahydro analogs. "Heterocycles" further include, without limitation: imidazolyl, indolyl, isothiazolyl, isoxazolyl, oxadiazolyl, oxazolyl, oxetanyl, pyranyl, pyrazinyl, pyrazolyl, pyridazinyl, pyridyl, pyrimidinyl, pyrrolyl, quinazolinyl, tetrazolyl, thiadiazolyl, thiazolyl, thienyl, triazolyl, 1,4-alkyldioxinyl, alkylpyrrolidinyl, dihydroimidazolyl, dihydroisoxazolyl, dihydroisothiazolyl, dihydrooxadiazolyl, dihydrooxazolyl, dihydropyrazinyl, dihydropyrazolyl, dihydropyridyl, dihydropyrimidinyl, dihydropyrrolyl, dihydrotetrazolyl, dihydrothiadiazolyl, dihydrothiazolyl, dihydrothienyl, dihydrotriazolyl, methylenedioxybenzophenonacyl, tetrahydrofuranyl, tetrahydrothiophenyl and their N-oxides, etc. The bond to the substituent of the heterocycle may be via a C atom or a heteroatom. In one embodiment, the heterocycle is selected from imidazolyl, pyridyl, 1-pyrrolidone, 2-piperidone, 2-pyrimidone, 2-pyrrolidone, thienyl, oxazolyl, triazolyl, isoxazolyl, etc.

[0055] The term "aryl" refers to a monocyclic or polycyclic aromatic group, preferably a monocyclic or bicyclic aromatic group, for example phenyl or naphthyl. Unless otherwise specified, an aryl group may be unsubstituted or substituted by one or more, in particular one to four, groups, including, for example, halogen, alkyl, alkenyl, _—OCF3 , _—CF3 , _—NO2 , —CN, —NC, _—OH , alkoxy, amino, alkylamino, —CO2H, _—CO2alkyl , —OCO-alkyl, aryl and heteroaryl.

[0056] The term "heteroaryl" means a stable monocyclic ring containing up to six atoms or a stable bicyclic ring in which each ring contains up to six atoms. At least one of the rings is an aromatic ring containing 1-4 atoms selected from O, N or S. Heteroaryl groups include, but are not limited to: imidazolyl, triazolyl, pyrazolyl, furanyl, thienyl, oxazolyl, isoxazolyl, pyrazinyl, pyridazinyl, pyridyl, pyrimidinyl, pyrrolyl. With respect to the definition of heteroaryl, any N-oxidized derivatives of heteroaryl containing an N atom should also be added. When the heteroaryl substituted group is a bicyclic ring and one of the two rings is a non-aromatic ring or a ring that does not contain a heteroatom, the bicyclic ring is fused through the aromatic ring or heteroatoms.

[0057] The term "halogen" or "halogen radical" means chlorine, fluorine, bromine and iodine.

[0058] In one aspect, the present invention relates to a method of treating a viral infection, including a coronavirus infection, in a human in need thereof, comprising administering to the patient a therapeutically effective amount of a compound of formula (I):

or a pharmaceutically acceptable salt thereof, where:

R ₁ is hydrogen, C _1-4 alkyl, C _3-6 cycloalkyl, C _1-4 alkyloxy or phenyl; and

R ₂ is hydrogen, C _1-4 alkyl, C _3-6 cycloalkyl or halogen.

[0059] In one embodiment of the compound of formula (I), R ₁ is hydrogen or C _1-4 alkyl.

[0060] In another embodiment of the compound of formula (I), R ₁ is hydrogen.

[0061] In another embodiment of the compound of formula (I), R ₂ is hydrogen or C _1-4 alkyl.

[0062] In another embodiment of the compound of formula (I), R ₂ is C _1-4 alkyl.

[0063] In another embodiment of the compound of formula (I), R ₂ is methyl or ethyl. In another embodiment, R ₂ is methyl.

[0064] In another embodiment of the compound of formula (I), R ₁ is hydrogen and R ₂ is C _1-4 alkyl.

[0065] In one embodiment, the compound of formula (I) is compound 1:

or a pharmaceutically acceptable salt thereof.

[0066] The compounds of formula (I) and compound 1 are small molecule inhibitors of Abelson tyrosine kinase (ABL). Among the drugs or drug candidates screened in an attempt to repurpose existing compounds for the treatment of coronavirus infections, ABL inhibitors have demonstrated promising forms of activity against coronaviruses, including SARS-CoV and MERS-CoV, through various mechanisms of action.

[0067] First and second generation ABL inhibitors imatinib and dasatinib have been reported to inhibit SARS-CoV and MERS-CoV with low cytotoxicity in Vero E6 cells. Dyall et al., Antimicrob Agents Chemother , 58 (8), 4885–93.

[0068] In another study of the effect of imatinib against coronaviruses, imatinib demonstrated suppression of SARS-CoV virus production in Vero E6 and Calu-3 cells or MERS-CoV virus production in Vero E6 and MRC5 cells. Coleman et al., Journal of Virology , 90 (19), 8924–33.

[0069] It has also been reported that imatinib can inhibit viral membrane fusion. Coleman et al., Journal of Virology , 90 (19), 8924-33. Since viral replication requires entry of viruses into host cells by fusion with cellular membranes, blocking the fusion induced by the coronavirus spike protein may lead to suppression of viral replication and treatment of coronavirus infections.

[0070] Following the COVID-19 outbreak, it has been reported that a subset of patients with severe COVID-19 appear to have cytokine storm syndrome (CSS), which is a major cause of death. Mehta et al., The Lancet , 395, 10229, 1033-1034. Cytokine storm is caused by excessive or uncontrolled levels of cytokines released by the immune system. This causes more immune cells to be activated and leads to hyperinflammation. This can cause severe harm to the patient or even death. Thus, therapeutic options that result in immunosuppression, including selective cytokine blockade, may be beneficial.

[0071] The ABL kinase inhibitors nilotinib and ponatinib have been reported to suppress the production of multiple proinflammatory cytokines. Chen et al., Front. Immunol . 10: 1393. The study reported that nilotinib and ponatinib suppressed the production of three important cytokines, interleukin 8 (IL-8), interferon-gamma-inducible protein 10 (IP-10), and monocyte chemoattractant protein 1 (MCP-1), all of which are associated with the influenza virus-induced cytokine storm.

[0072] As a third-generation ABL kinase inhibitor, ponatinib has additionally demonstrated prolongation of survival in mice with lethal influenza virus infection in animal models. Chen et al., Front. Immunol . 10: 1393. Suppression of cytokine production and prolonged survival further suggest the potential use of ABL inhibitors for the treatment of COVID-19 patients with severe symptoms by suppressing the cytokine storm.

[0073] In another study, ponatinib was shown to inhibit receptor-interacting serine-threonine protein kinase 1 (RIPK1). Gordon et al., BioRxiv , 2020.03.22, 002386. In this study, RIPK1 was identified as an interactor with the SARS-CoV-2 Nsp12 protein using a protein-protein interaction map of human SARS-CoV-2. Since ponatinib is an inhibitor of RIPK1, it was therefore suggested that ponatinib could be repurposed as a potential antiviral drug for the treatment of COVID-19.

[0074] Without being limited to the above mechanisms of action of ABL inhibitors against coronaviruses and viral infections, the present invention provides methods of treating coronavirus infections using compounds of formula (I) or compound 1 or a pharmaceutically acceptable salt thereof, which are potent ABL inhibitors and thus are potential antiviral compounds for the treatment of coronavirus infections.

[0075] The chemical name of compound 1 is 3-(2-(1H-pyrazolo[3,4-b]pyridin-5-yl)ethynyl)-4-methyl-N-(4-((4-methylpiperazin-1-yl)methyl)-3-(trifluoromethyl)phenyl)benzamide.

[0076] As used herein, compounds of formula (I) and compound 1 include any tautomeric forms. As a non-limiting example, tautomerization can occur at pyrazole and pyrimidine groups.

[0077] Compounds of formula (I) or compound 1, or a pharmaceutically acceptable salt thereof, can be prepared according to the preparation methods described in U.S. Patent No. 8,846,671 B2, issued September 30, 2014, which is incorporated herein by reference in its entirety and for all purposes, or in a manner analogous thereto.

[0078] In certain embodiments, the viral infection is caused by a coronavirus.

[0079] In certain embodiments, the coronavirus infection is caused by a coronavirus, including SARS-CoV-2, SARS-CoV, MERS-CoV, 229E, NL63, OC43, and HKU1.

[0080] In certain embodiments, the coronavirus infection is caused by a coronavirus, including SARS-CoV-2, SARS-CoV, and MERS-CoV.

[0081] In certain embodiments, the coronavirus infection is caused by SARS-CoV-2.

[0082] In certain embodiments, the patient has COVID-19.

[0083] In certain embodiments, the coronavirus infection is caused by SARS-CoV.

[0084] In certain embodiments, the coronavirus infection is caused by MERS-CoV.

[0085] In certain embodiments, replication of the coronavirus is suppressed.

[0086] In certain embodiments, coronavirus polymerase is inhibited.

[0087] In certain embodiments, fusion of the coronavirus with the membrane is inhibited.

[0088] In certain embodiments, the ABL signaling pathway is inhibited.

[0089] In certain embodiments, a specific ABL2 signaling pathway is inhibited.

[0090] In certain embodiments, receptor-interacting serine-threonine protein kinase 1 (RIPK1) is inhibited.

[0091] In certain embodiments, the method comprises treating a patient with cytokine storm syndrome.

[0092] In certain embodiments, cytokine production in a patient is suppressed.

[0093] In certain embodiments, the production of IL-8, IP-10, and MCP-1 in a patient is suppressed.

[0094] In certain embodiments, the inhibition occurs in vitro or in vivo.

[0095] Similar to ponatinib, compound 1 of the present invention is a third-generation ABL inhibitor that equally inhibits both ABL1 and ABL2 kinase, and has higher activity compared to other ABL inhibitors, including imatinib, dasatinib, nilotinib, and ponatinib. Table 1 shows the ABL kinase inhibitory activity of compound 1.

Table 1. Kinase inhibition activity of compound 1

Compound 1, Kinase % of control at 10 nM % of control at 100 nM % of control at 1000 nM Unphosphorylated ABL1 4.4 0.1 0 Phosphorylated ABL1 3 0 0 ABL2 2.6 0 0

[0096] Table 2 below shows that Compound 1 has higher activity against ABL kinases compared to other ABL inhibitors, which demonstrates that Compound 1 may have higher potential as an antiviral compound against coronaviruses, including SARS-CoV-2.

Table 2. Activity forms of compound 1

IC ₅₀ (nM) Inhibition of kinases
(BCR-ABL1-kinase) Activity in cells
(Ba/F3 expressing BCR-ABL1) Imatinib 210 ± 21 565 ± 656 Dasatinib 0.13 ± 0.0039 31 ± 4 Nilotinib 8.8 ± 0.45 10 ± 3 Ponatinib 0.33 ± 0.03 11 ± 3 Compound 1, 0.30 ± 0.0045 6 ± 3

[0097] In another aspect, the present invention relates to a method of treating a viral infection, including a coronavirus infection, in a human in need thereof, comprising administering to the patient a therapeutically effective amount of a compound of formula (II):

or a pharmaceutically acceptable salt thereof, where:

X is selected from -C(=O)-, -C(=S)-, -C(=NH)- and -SO ₂ -;

Y is selected from -NH-, -O-, -S- or absent;

R is selected from -C _3-6 cycloalkylene And ;

R ₁ is selected from And

Z represents O, S or NH;

n is 0, 1, or 2;

ring A is a C _4-8 aliphatic ring; and

Ring B is phenyl, naphthyl, pyridyl, pyridazinyl, pyrazinyl or pyrimidinyl, and Ring B is optionally substituted.

[0098] In one embodiment of the compound of formula (II), X is C=O or -SO ₂ -.

[0099] In one embodiment of the compound of formula (II), X is -SO ₂ -.

[0100] In one embodiment of the compound of formula (II), Y is absent.

[0101] In another embodiment of the compound of formula (II), R is

[0102] In another embodiment of the compound of formula (II), R is .

[0103] In another embodiment of the compound of formula (II), R ₁ is selected from

[0104] In another embodiment of the compound of formula (II), R ₁ is .

[0105] In another embodiment of the compound of formula (II), R ₁ is .

[0106] In another embodiment of the compound of formula (II), n is 0 or 1.

[0107] In another embodiment of the compound of formula (II), ring A is a C5-6 aliphatic ring.

[0108] In another embodiment of the compound of formula (II), ring B is phenyl, pyridyl, pyridazinyl, pyrazinyl or pyrimidinyl.

[0109] In another embodiment of the compound of formula (II), ring B is phenyl or pyridyl.

[0110] In another embodiment of the compound of formula (II), ring B is phenyl.

[0111] In another embodiment of the compound of formula (II), R is ; R ₁ is ; and ring B is phenyl.

[0112] In another embodiment of the compound of formula (II), X is C=O or -SO ₂ -; Y is absent; R is ; R ₁ is ; and ring B is phenyl or pyridyl.

[0113] In one embodiment, the compound of formula (II) is compound 2:

Compound 2,

or a pharmaceutically acceptable salt thereof.

[0114] The compounds of formula (II) and compound 2 are small molecule inhibitors of proteins that are inhibitors of apoptosis proteins (IAPs). Apoptosis, or programmed cell death, is a cellular process that is critical for homeostasis, normal development, and immune defense of the host organism. Inappropriate regulation of apoptosis has been implicated in many human diseases, including viral infections. D. W. Nicholson,Nature2000, 407, 810-816. Campbell GR et al., 2019 Autophagy, 15(4): 744–746. As a consequence, targeting key regulators of apoptosis has great potential for developing new approaches to the treatment of viral infections.

[0115] One class of apoptosis regulators are the IAPs. This class includes proteins such as XIAP, cIAP1, cIAP2, ML-IAP, HIAP, KIAP, TSIAP, NAIP, survivin, livin, ILP-2, Apollon, and BRUCE. IAP proteins potently suppress cellular apoptosis. Among the IAPs, cellular IAP1 (cIAP1) and cIAP2 play key roles in regulating death receptor-mediated apoptosis, whereas X-linked IAP (XIAP) suppresses both death receptor-mediated apoptosis and mitochondria-mediated apoptosis by binding to and inhibiting caspase-3/7 and caspase-9, three cysteine proteases critical for apoptosis. GS Salvesen et al., Nat. Rev. Mol. Cell. Biol . 2002, 3, 401-410.

[0116] Thus, IAP inhibitors are useful tools for elucidating the antiapoptotic function of IAP. IAP inhibitors have been reported to have promising activity against viral infections, including those caused by HIV and hepatitis B virus (HBV).

[0117] Recent studies have reported that IAP inhibition is a novel target for viral clearance. Campbell GR et al., 2019, Autophagy , 15(4): 744-746. The result showed that IAP inhibitor selectively killed HIV-1-infected resting memory CD4+ T cells but not uninfected T _CM , resulting in selective apoptosis of only HIV-infected T _CM while sparing uninfected “background” cells in the absence of viral infection.

[0118] IAP inhibitors have also been reported to selectively kill HBV-infected hepatocytes. In one study, HBV infection was induced in two mouse models to examine the importance of host factors in controlling infection. Ebert G, et al. Proc Natl Acad Sci USA , 2015, 112(18): 5797-5802. The result showed that cIAPs limit the death of infected hepatocytes and promote HBV viral persistence. Liver-specific cIAP1 and complete cIAP2-deficient animals effectively control HBV infection compared to wild-type mice, suggesting that antagonizing cIAP function may promote clearance of HBV infection.

[0119] In another study, cIAP inhibitors demonstrated potential efficacy in the treatment of HBV infection and other intracellular infections. Ebert G, et al. Proc Natl Acad Sci USA , 2015, 112(18): 5803-5808. In this study, an immunocompetent mouse model of chronic HBV infection was used to examine the in vivo efficacy of cIAP inhibitors in the treatment of HBV infection. The data show that birinapant, a small molecule IPA inhibitor, was able to rapidly reduce serum HBV DNA levels and serum HBV surface antigen levels, and promote the clearance of hepatocytes containing HBV core antigen.

[0120] Without being limited to the above mechanisms of action of IAP inhibitors, the present invention provides a method of treating coronavirus infections using compounds of formula (II) or compound 2 or a pharmaceutically acceptable salt thereof, which are potent IAP inhibitors and thus are potential antiviral compounds for the treatment of coronavirus infections.

[0121] The chemical name of compound 2 is (5S,5'S,8S,8'S,10aR,10a'R)-3,3'-(1,3-phenylenedisulfonyl)bis(N-benzhydryl-5-((S)-2-(methylamino)propanamido)-6-oxodecahydropyrrolo[1,2-a][1,5]diazocin-8-carboxamide.

[0122] As used herein, compounds of formula (II) and compound 2 include any tautomeric forms. As a non-limiting example, tautomerization can occur at pyrazole and pyrimidine groups.

[0123] The compound of formula (II) or compound 2, or a pharmaceutically acceptable salt thereof, can be prepared according to the preparation methods described in U.S. Patent No. 8,883,771 B2, issued November 11, 2014, which is incorporated herein by reference in its entirety and for all purposes, or in a manner analogous thereto.

[0124] In certain embodiments, the viral infection is caused by a coronavirus.

[0125] In certain embodiments, the coronavirus infection is caused by a coronavirus, including SARS-CoV-2, SARS-CoV, MERS-CoV, 229E, NL63, OC43, and HKU1.

[0126] In certain embodiments, the coronavirus infection is caused by a coronavirus, including SARS-CoV-2, SARS-CoV, and MERS-CoV.

[0127] In certain embodiments, the coronavirus infection is caused by SARS-CoV-2.

[0128] In certain embodiments, the patient has COVID-19.

[0129] In certain embodiments, the coronavirus infection is caused by SARS-CoV.

[0130] In certain embodiments, the coronavirus infection is caused by MERS-CoV.

[0131] In certain embodiments, replication of the coronavirus is suppressed.

[0132] In certain embodiments, coronavirus polymerase is inhibited.

[0133] In certain embodiments, IAP proteins are inhibited.

[0134] In certain embodiments, cIAP proteins are inhibited.

[0135] In certain embodiments, apoptosis is selectively induced in cells infected with a coronavirus.

[0136] In certain embodiments, clearance of coronavirus is achieved.

[0137] In certain embodiments, clearance of coronavirus is achieved by enhancing T cell-mediated antiviral immunity.

[0138] In certain embodiments, the inhibition occurs in vitro or in vivo.

[0139] Similar to the above-mentioned IAP inhibitors, compound 2 of the present invention is a potent IAP inhibitor and can promote apoptosis, and thus may be a potential antiviral compound for the treatment of coronavirus infection. Fig. 1A-1C show the effect of compound 2 on the induction of apoptosis, IAP expression, and HBV replication and gene expression in the liver cancer cell line HepG2.2.15.

[0140] As can be seen in Fig. 1, compound 2 suppresses IAP expression in a dose-dependent manner. The combination of compound 2 and TNF-α significantly activates the apoptotic pathway. As shown in Fig. 1D, low concentrations of compound 2 and birinapant result in an increase in viral replication by suppressing cIAP2 expression, and high concentrations of compound 2 and birinapant impair the enhancement of viral replication by suppressing cell proliferation through the inactivation of other IAP proteins.

[0141] Injection of Compound 2 also promotes viral clearance in the pAAV-HBV 1.3 model in C57BL/6J mice, as shown in Figs. 2A-2D.

[0142] Fig. 3A-3D show that Compound 2 specifically induces apoptosis in HBV-infected (HBsAg+) hepatocytes in mouse HBV models.

[0143] Figs. 4A-4E show that Compound 2 promotes the expression of IFN-γ and TNF-α and increases the number of CD4+ and CD8+ T cells in liver tissue in HBV models.

[0144] In another aspect, the present invention relates to a method of treating a viral infection, including a coronavirus infection, in a human in need thereof, comprising administering to the patient a therapeutically effective amount of a compound of formula (III), (IV) or (V):

or a pharmaceutically acceptable salt thereof, where:

Ring A is

X, substituted or unsubstituted, is selected from the group consisting of alkylene, alkenylene, cycloalkylene, cycloalkenylene and heterocycloalkylene;

Y is selected from the group consisting of (CH ₂ ) _n -N(R ^a ) ₂ and

Q is selected from the group consisting of O, O( _CH2 ) _1-3 , ^NRc _, ^NRc ( _C1-3alkylene ), OC(=O)( _C1-3alkylene ), C(=O)O, C(=O)O( _C1-3alkylene ), NHC(=O)( _C1-3alkylene ), C(=O)NH and C(=O)NH( _C1-3alkylene );

Z represents O or NR ^c _;

R ₁ and R ₂ are independently selected from the group consisting of H, CN, NO ₂ , halogen, alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, aryl, heteroaryl, heterocycloalkyl, OR', SR', NR'R'', COR', CO ₂ R', OCOR', CONR'R'', CONR'SO ₂ R'', NR'COR'', NR'CONR''R''', NR'C=SNR''R''', NR'SO ₂ R'', SO ₂ R' and SO ₂ NR'R'';

R ₃ is selected from the group consisting of H, alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, aryl, heteroaryl, heterocycloalkyl, OR', NR'R'', OCOR', CO ₂ R', COR', CONR'R'', CONR'SO ₂ R'', C _1-3 alkylene-CH(OH)CH ₂ OH, SO ₂ R' and SO ₂ NR'R'';

R', R'', and R''' are independently H, alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, aryl, heteroaryl, C _1-3 alkyleneheterocycloalkyl, or heterocycloalkyl;

R' and R'' or R'' and R''' may be taken together with the atom to which they are bonded to form a 3- to 7-membered ring;

R ₄ is hydrogen, halogen, C _1-3 alkyl, CF ₃ or CN;

R ₅ is hydrogen, halogen, C _1-3 alkyl, substituted C _1-3 alkyl, hydroxyalkyl, alkoxy or substituted alkoxy;

R ₆ is selected from the group consisting of H, CN, NO ₂ , halogen, alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, aryl, heteroaryl, heterocycloalkyl, OR', SR', NR'R'', CO ₂ R', OCOR', CONR'R'', CONR'SO ₂ R'', NR'COR'', NR'CONR''R''', NR'C=SNR''R''', NR'SO ₂ R'', SO ₂ R' and SO ₂ NR'R'';

R₇, substituted or unsubstituted, selected from the group consisting of hydrogen, alkyl, alkenyl, (CH₂)_0-3cycloalkyl, (CH₂)_0-3cycloalkenyl, (CH₂)_0-3heterocycloalkyl, (CH₂)_0-3aryl and (CH₂)_0-3heteroaryl;

R ₈ is selected from the group consisting of hydrogen, halogen, NO ₂ , CN, CF ₃ SO ₂ and CF ₃ ;

R _a is selected from the group consisting of hydrogen, alkyl, heteroalkyl, alkenyl, hydroxyalkyl, alkoxy, substituted alkoxy, cycloalkyl, cycloalkenyl, and heterocycloalkyl;

R _b is hydrogen or alkyl;

R _c is selected from the group consisting of hydrogen, alkyl, substituted alkyl, hydroxyalkyl, alkoxy and substituted alkoxy; and

n, r, and s are independently equal to 1, 2, 3, 4, 5, or 6.

[0145] In one embodiment of a compound of formula (III), (IV) or (V), ring A is

[0146] In another embodiment of a compound of formula (III), (IV), or (V), X is alkylene. In another embodiment, X is C _1-4 alkylene.

[0147] In one embodiment of a compound of formula (III), (IV) or (V), Y is

[0148] In another embodiment of a compound of formula (III), (IV) or (V), Q is C(=O)O(C _1-3 alkylene).

[0149] In another embodiment of a compound of formula (III), (IV) or (V), Z is O.

[0150] In another embodiment of a compound of formula (III), (IV), or (V), R ₁ is SO ₂ R', and R' is alkyl. In another embodiment, R ₁ is SO ₂ Me.

[0151] In another embodiment of a compound of formula (III), (IV), or (V), R ₂ is alkyl. In another embodiment, SO ₂ is methyl, propyl, or isopropyl. In another embodiment, SO ₂ is methyl.

[0152] In another embodiment of a compound of formula (III), (IV), or (V), R ₃ is H or alkyl. In another embodiment, R ₃ is C ₁ -C ₄ alkyl. In another embodiment, R ₃ is methyl, propyl, or isopropyl. In another embodiment, R ₃ is isopropyl.

[0153] In another embodiment of the compound of formula (III), (IV), or (V), R ₄ is hydrogen or halogen. In another embodiment, R ₄ is halogen. In another embodiment, R ₄ is -Cl or -F. In another embodiment, R ₄ is -Cl.

[0154] In another embodiment of the compound of formula (III), (IV), or (V), R ₅ is hydrogen or halogen. In another embodiment, R 5 is halogen. In another embodiment, R ₅ is -Cl or -F. In another embodiment, R ₅ is -F.

[0155] In another embodiment of a compound of formula (III), (IV), or (V), R ₆ is H or alkyl. In another embodiment, R ₆ is H.

[0156] In another embodiment of a compound of formula (III), (IV), or (V), R7 is H or alkyl. In another embodiment, R ₇ is H.

[0157] In another embodiment of the compound of formula (III), (IV), or (V), R ₈ is hydrogen, halogen, or CF ₃ SO ₂ . In another embodiment, R ₈ is CF ₃ SO ₂ .

[0158] In another embodiment of a compound of formula (III), (IV), or (V), each of n, r, and s is independently 1 or 2. In another embodiment, n, r, and s are 2. In another embodiment, n is 1; and r and s are 2.

[0159] In another embodiment of a compound of formula (III), (IV) or (V), R ^b is H.

[0160] In one embodiment, the compound of formula (III) or (IV) is compound 3:

Compound 3,

or a pharmaceutically acceptable salt thereof.

[0161] In another embodiment, the present invention relates to a method of treating a viral infection in a patient, comprising administering to the patient a therapeutically effective amount of compound 3.

[0162] In another aspect, the present invention relates to a method of treating a viral infection, including a coronavirus infection, in a human in need thereof, comprising administering to the patient a therapeutically effective amount of a compound of formula (VI):

(VI),

or a pharmaceutically acceptable salt thereof, where:

SO ₂₁ is SO ₂ SO ₂ ',

SO ₂₂ is alkyl, preferably C ₁ -C ₄ alkyl, more preferably methyl, propyl or isopropyl,

SO ₂₃ is alkyl, preferably C ₁ -C ₄ alkyl, more preferably methyl, propyl or isopropyl,

SO ₂₄ is a halogen, preferably fluoride, chloride,

SO ₂₅ is a halogen, preferably fluoride, chloride,

SO ₂₆ is selected from H, halogen, alkyl, preferably fluoride, chloride, C ₁ -C ₄ alkyl, more preferably methyl, propyl, isopropyl,

SO _21b is H or alkyl, preferably C ₁ -C ₄ alkyl, more preferably methyl, propyl or isopropyl,

n ₂ , SO ₂ and s ₂ are independently 1, 2, 3, 4, 5 or 6, more preferably both of SO ₂ and s ₂ are 2, and n ₂ is 3, 4 or 5, more preferably all of n ₂ , SO ₂ and s ₂ are 2, and

SO ₂ ' is alkyl, preferably C ₁ -C ₄ alkyl, more preferably methyl, propyl or isopropyl.

[0163] In one embodiment of the compound of formula (VI), SO ₂₁ is SO ₂ (C ₁ -C ₄ alkyl). In another embodiment, SO ₂₁ is SO ₂ Me.

[0164] In another embodiment of the compound of formula (VI), SO ₂₂ is C ₁ -C ₄ alkyl. In another embodiment, SO ₂₂ is methyl.

[0165] In another embodiment of the compound of formula (VI), SO ₂₃ is C ₁ -C ₄ alkyl. In another embodiment, SO ₂₃ is isopropyl.

[0166] In another embodiment of the compound of formula (VI), SO ₂₄ is fluoride or chloride. In another embodiment, SO ₂₄ is chloride.

[0167] In another embodiment of the compound of formula (VI), SO ₂₅ is fluoride or chloride. In another embodiment, SO ₂₅ is fluoride.

[0168] In another embodiment of the compound of formula (VI), SO ₂₆ is H or C ₁ -C ₄ alkyl. In another embodiment, SO ₂₆ is H.

[0169] In another embodiment of the compound of formula (VI), SO _21b is H.

[0170] In another embodiment of the compound of formula (VI), each of n ₂ , SO ₂ , and s ₂ is independently 1 or 2. In another embodiment, n ₂ , SO ₂ , and s ₂ are 2.

[0171] In another embodiment of the compound of formula (VI), SO ₂₁ is SO ₂ (C ₁ -C ₄ alkyl); SO ₂₂ is C ₁ -C ₄ alkyl; and SO ₂₃ is C ₁ -C ₄ alkyl. In another embodiment, SO ₂₁ is SO ₂ Me; SO ₂₂ is methyl; and SO ₂₃ is isopropyl.

[0172] In another embodiment of the compound of formula (VI), SO ₂₄ is -Cl and SO ₂₅ is -F.

[0173] In one embodiment, the compound of formula (VI) comprises compound 4:

Compound 4,

or a pharmaceutically acceptable salt thereof.

[0174] In another embodiment, the present invention relates to a method of treating a viral infection in a patient, comprising administering to the patient a therapeutically effective amount of compound 4.

[0175] In another aspect, the present invention relates to a method of treating a viral infection in a patient, comprising administering to the patient a therapeutically effective amount of compound 5:

Compound 5,

[0176] Compounds of formula (III), (IV), (V), or (VI), Compound 3, Compound 4, and Compound 5 are inhibitors of BCL-2 proteins. BCL-2 proteins function as critical regulators of apoptosis in cells. DT Chao, et al., Annu Rev Immunol 1998; 16:395-419. Bcl-2 proteins serve to check the state of apoptosis, allowing healthy and beneficial cells to survive. This family of proteins includes anti-apoptotic proteins such as BCL-2, BCL-xL, and MCL-1, and pro-apoptotic molecules including Bid, Bim, Bad, Bak, and Bax. Replication of several important human microorganisms, including coronaviruses, influenza A virus (IAV), HBV, HCV, and HIV, depends on mitochondria-initiated apoptosis mediated by BCL-2, BCL-xL, and BCL-w. Geng et al., Proc Natl Acad Sci USA 2012; 109: 18471–18476. Park J et al., Hepatology 2012; 56: 831–840. Busca A et al., J Biol Chem 2012; 287: 15118–15133. Tan et al., J Virol. 2007; 81: 6346–6355.

[0177] BCL-xL inhibitors such as ABT-737 and its orally available analogue ABT-236 have been reported to accelerate apoptosis in virus-infected cells. Therefore, BCL-xL inhibitors may be potential antiviral compounds for the treatment of viral infections.

[0178] In another study, ABT-263 was found to accelerate apoptosis in virus-infected cells by activating the caspase-mediated apoptosis pathway, thereby initiating cell death in influenza A virus (IAV)-infected cells. Kakkola et al., Cell Death and Disease , 2013, 4, e742.

[0179] Without being limited to the above mechanisms of action of BCL-xL inhibitors, the present invention provides a method of treating coronavirus infections using compounds of formula (III), (IV), (V) or (VI) or compound 3, compound 4 or compound 5 or a pharmaceutically acceptable salt thereof, which are potent BCL-xL inhibitors and thus are potential antiviral compounds for the treatment of coronavirus infections.

[0180] The chemical name of compound 3 is ( R )-2-(1-(3-(4-(N-(4-(4-(3-(2-(4-chlorophenyl)-1-isopropyl-5-methyl-4-(methylsulfonyl)-1H-pyrrol-3-yl)-5-fluorophenyl)piperazin-1-yl)phenyl)sulfamoyl)-2-(trifluoromethylsulfonyl)phenylamino)-4-(phenylthio)butyl)piperidine-4-carbonyloxy)ethylphosphonic acid.

[0181] The chemical name of compound 4 is ( R )-1-(3-(4-(N-(4-(4-(3-(2-(4-chlorophenyl)-1-isopropyl-5-methyl-4-(methylsulfonyl)-1H-pyrrol-3-yl)-5-fluorophenyl)piperazin-1-yl)phenyl)sulfamoyl)-2-(trifluoromethylsulfonyl)phenylamino)-4-(phenylthio)butyl)piperidine-4-carboxylic acid.

[0182] The chemical name of compound 5 is (S)-N-((4-(((1,4-dioxan-2-yl)methyl)amino)-3-nitrophenyl)sulfonyl)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((6-(4-chlorophenyl)spiro[3.5]non-6-en-7-yl)methyl)piperazin-1-yl)benzamide.

[0183] As used herein, compounds of formula (III), (IV), (V), or (VI) and compounds 3, 4, and 5 include any tautomeric forms. As a non-limiting example, tautomerization can occur at pyrazole and pyrimidine groups.

[0184] The compound of formula (III), (IV), or (V) and compounds 3 and 4, or a pharmaceutically acceptable salt thereof, can be prepared according to the preparation methods described in U.S. Patent No. 9,096,625 B2, issued August 4, 2015, which is incorporated herein by reference in its entirety and for all purposes, or in a manner analogous thereto.

[0185] Compound 5 or a pharmaceutically acceptable salt thereof can be prepared according to the preparation methods described in U.S. Patent No. 10,221,174 B, issued March 5, 2019, which is incorporated herein by reference in its entirety and for all purposes, or in a manner analogous thereto.

[0186] In certain embodiments, the viral infection is caused by a coronavirus.

[0187] In certain embodiments, the coronavirus infection is caused by a coronavirus, including SARS-CoV-2, SARS-CoV, MERS-CoV, 229E, NL63, OC43, and HKU1.

[0188] In certain embodiments, the coronavirus infection is caused by a coronavirus, including SARS-CoV-2, SARS-CoV, and MERS-CoV.

[0189] In certain embodiments, the coronavirus infection is caused by SARS-CoV-2.

[0190] In certain embodiments, the patient has COVID-19.

[0191] In certain embodiments, the coronavirus infection is caused by SARS-CoV.

[0192] In certain embodiments, the coronavirus infection is caused by MERS-CoV.

[0193] In certain embodiments, replication of the coronavirus is suppressed.

[0194] In certain embodiments, coronavirus polymerase is inhibited.

[0195] In certain embodiments, BCL-2 proteins are inhibited.

[0196] In certain embodiments, BCL-xL proteins are inhibited.

[0197] In certain embodiments, apoptosis is selectively induced in cells infected with a coronavirus.

[0198] In certain embodiments, the caspase-mediated apoptotic pathway is activated.

[0199] In certain embodiments, the inhibition occurs in vitro or in vivo.

[0200] Also provided herein are pharmaceutical compositions and dosage forms comprising compounds of formulas (I)-(VI) or compounds 1-5, or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable excipients. The compositions and dosage forms provided herein may further comprise one or more additional active ingredients. The compounds of formulas (I)-(VI) or compounds 1-5, or a pharmaceutically acceptable salt thereof, may be administered as part of the described pharmaceutical composition.

[0201] The compound of the present invention can be administered by any suitable route, such as orally, buccally, by inhalation, sublingually, rectally, vaginally, intracisternally or intrathecally by lumbar puncture, transurethral, nasal, transcutaneously, i.e. transdermal, or parenterally (including intravenous, intramuscular, subcutaneous, intracoronary, intradermal, intramammary, intraperitoneal, intra-articular, intrathecal, retrobulbar, intrapulmonary injection and/or surgical implantation at a specific site). Parenteral administration can be accomplished using a needle and syringe or using high pressure techniques.

[0202] Pharmaceutical compositions include compositions in which a compound of the present invention is administered in an amount effective to achieve the intended purpose. The precise composition, route of administration, and dosage are determined by the individual physician based on the condition or disease diagnosed. The dosage and interval can be adjusted individually to provide levels of the compound of the present invention that are sufficient to maintain therapeutic effects.

[0203] The toxicity and therapeutic efficacy of the compounds of the present invention can be determined using standard pharmaceutical procedures in cell cultures or in experimental animals, for example, to determine the maximum tolerated dose (MTD) of the compound, which is defined as the highest dose that does not cause toxicity in animals. The dose ratio between the maximum tolerated dose and the dose that provides therapeutic effects (e.g., tumor growth suppression) is the therapeutic index. The dosage may vary within this range depending on the dosage form used and the route of administration used. Determination of a therapeutically effective amount is within the skill of the art, especially in light of the detailed description provided herein.

[0204] The therapeutically effective amount of a compound of the present invention required for use in therapy varies depending on the nature of the condition being treated, the length of time over which activity is desired, and the age and condition of the subject, and is ultimately determined by the attending physician. Dosage amounts and dosing intervals can be adjusted individually to provide plasma levels of the compound of the present invention sufficient to maintain the desired therapeutic effects. The desired dosage can be administered as a single dose or as multiple doses administered at appropriate intervals, such as one, two, three, four or more sub-doses per day. Multiple doses are often desirable or necessary. For example, a compound of the present invention can be administered at the following frequency: four doses delivered as one dose per day at four-day intervals (q4d x 4); four doses delivered as one dose per day at three-day intervals (q3d x 4); delivered as one dose per day at five-day intervals (qd x 5); one dose per week for three weeks (qwk3); five daily doses with a two-day rest period and a further five daily doses (5/2/5) or any dosing regimen determined to be appropriate in the circumstances.

[0205] In certain embodiments, the compounds of the present invention are typically administered in admixture with a pharmaceutical carrier to provide a pharmaceutical composition selected taking into account the intended route of administration and standard pharmaceutical practice.

[0206] Pharmaceutical compositions for use in accordance with the present invention are formulated in a conventional manner using one or more physiologically acceptable carriers including excipients and/or excipients that facilitate processing of the compound of the present invention.

[0207] These pharmaceutical compositions can be prepared, for example, by conventional mixing, dissolving, granulating, coating, emulsifying, encapsulating, entrapping or lyophilizing methods. The appropriate formulation depends on the selected route of administration. When a therapeutically effective amount of a compound of the present invention is administered orally, the composition is typically in the form of a tablet, capsule, powder, solution or elixir. When administered in the form of a tablet, the composition may further comprise a solid carrier such as gelatin or an excipient. The tablet, capsule and powder contain from about 0.01% to about 95%, and preferably from about 1% to about 50%, of the compound of the present invention. When administered in liquid form, a liquid carrier such as water, petroleum or animal or vegetable oils may be added. The liquid form of the composition may further comprise physiological saline, dextrose or other saccharide solutions or glycols. When administered in liquid form, the composition contains from about 0.1% to about 90%, and preferably from about 1% to about 50%, by weight of the compound of the present invention.

[0208] Solid dosage forms for oral administration include capsules, tablets, pills, powders and granules. In such solid dosage forms, the active ingredient is mixed with at least one inert pharmaceutically acceptable excipient or carrier, such as sodium citrate or dicalcium phosphate, and/or a) fillers or dry diluents, such as starches, lactose, sucrose, glucose, mannitol and silicic acid; b) binders, such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidinone, sucrose and acacia; c) humectants, such as glycerol; d) disintegrating agents, such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates and sodium carbonate; e) dissolution retarders, such as paraffin; f) absorption accelerators such as quaternary ammonium compounds; g) wetting agents such as, for example, cetyl alcohol and glycerol monostearate; h) absorbents such as kaolin and bentonite clay, and i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate and mixtures thereof. In the case of capsules, tablets and pills, the dosage form may also contain buffering agents such as phosphates or carbonates.

[0209] Solid compositions can also be used as fillers in soft and hard gelatin capsules using such excipients as lactose or milk sugar, as well as high molecular weight polyethylene glycols and the like. Solid dosage forms of tablets, dragees, capsules, pills and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulation art. They may optionally contain opacifying agents and can also be formulated such that they release the active ingredient(s) only or preferentially in a certain part of the intestinal tract, optionally in a delayed manner. Examples of filling compositions that can be used include polymeric substances and waxes.

[0210] In solid dosage forms, the active ingredients may be mixed with at least one inert diluent, such as sucrose, lactose or starch. Such dosage forms may also contain, as is common practice, additional substances other than inert diluents, for example tableting lubricants and other tableting additives, such as magnesium stearate and microcrystalline cellulose. In the case of capsules, tablets and pills, the dosage forms may also contain buffering agents.

[0211] The active ingredients may also be presented in microencapsulated form with one or more of the excipients mentioned above.

[0212] When a therapeutically effective amount of a compound of the present invention is administered by intravenous, cutaneous or subcutaneous injection, the composition is in the form of a pyrogen-free parenterally acceptable aqueous solution. The preparation of such parenterally acceptable solutions with appropriate consideration of pH, isotonicity, stability, etc. is within the skill of the art. A preferred composition for intravenous, cutaneous or subcutaneous injection typically comprises an isotonic carrier medium.

[0213] The compounds of the present invention can be readily combined with pharmaceutically acceptable carriers well known in the art. Standard pharmaceutical carriers are described in Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, PA, 19th ed. 1995. Such carriers enable the active agents to be formulated as tablets, pills, dragees, capsules, liquids, gels, syrups, slurries, suspensions, and the like, for oral ingestion by the subject to be treated.

[0214] Pharmaceutical preparations for oral use can be obtained by adding a compound of the present invention to a solid excipient, optionally grinding the resulting mixture and processing the mixture of granules after adding suitable excipients, if desired, to obtain tablet or dragee cores. Suitable excipients include, for example, fillers and cellulose preparations. Disintegrants can be added if desired.

[0215] The compounds of the present invention can be formulated for parenteral administration by injection, for example, by bolus injection or continuous infusion. Formulations for injection can be presented in unit dosage form, for example, in ampoules or in multi-dose containers, with an added preservative. The compositions can take such forms as suspensions, solutions, or emulsions in oily or aqueous carrier media, and can contain formulatory agents such as suspending, stabilizing and/or dispersing agents.

[0216] Pharmaceutical compositions for parenteral administration include aqueous solutions of the active agent in water-soluble form. In addition, suspensions of the compound of the present invention can be prepared as suitable oily injection suspensions. Suitable lipophilic solvents or carrier media include fatty oils or synthetic fatty acid esters. Aqueous injection suspensions can contain substances that increase the viscosity of the suspension. The suspension can also optionally contain suitable stabilizers or agents that increase the solubility of the compounds and allow the preparation of highly concentrated solutions. Alternatively, the composition of the present invention can be presented in powder form for reconstitution with a suitable carrier medium, for example, sterile pyrogen-free water, before use.

[0217] The compounds of the present invention can also be formulated as compositions for rectal administration, such as, for example, suppositories or retention enemas containing conventional suppository bases. In addition to the formulations described above, a compound of the present invention can also be formulated as a depot preparation. Such long-acting formulations can be administered by implantation (e.g., subcutaneously or intramuscularly) or by intramuscular injection. Thus, for example, a compound of the present invention can be formulated with suitable polymeric or hydrophobic materials (e.g., as an emulsion in an acceptable oil) or ion exchange resins.

[0218] In particular, the compounds of the present invention can be administered orally, buccally or sublingually in the form of tablets containing excipients such as starch or lactose, or in capsules or vaginal suppositories, alone or in admixture with excipients, or in the form of elixirs or suspensions containing flavoring or coloring agents. Such liquid preparations can be prepared with pharmaceutically acceptable additives such as suspending agents. A compound of the present invention can also be injected parenterally, for example intravenously, intramuscularly, subcutaneously or intracoronarily.

[0219] For parenteral administration, the compound of the present invention is generally used in the form of a sterile aqueous solution which may contain other substances, for example salts or monosaccharides such as mannitol or glucose, to make the solution isotonic with blood.

[0220] In certain embodiments, a compound of Formula (I)-(VI) or compounds 1-5, or a pharmaceutically acceptable salt thereof, is administered orally to patients in need of such treatment.

[0221] In certain embodiments, a compound of Formula (I)-(VI) or compounds 1-5, or a pharmaceutically acceptable salt thereof, is administered intravenously to patients in need of such treatment.

[0222] A compound of the present invention used in the method of the present invention can be administered in an amount of from about 0.005 to about 500 milligrams per dose, from about 0.05 to about 250 milligrams per dose, or from about 0.5 to about 100 milligrams per dose. For example, a compound of the present invention can be administered in an amount of about 0.005, about 0.05, about 0.5, about 5, about 10, about 20, about 30, about 40, about 50, about 100, about 150, about 200, about 250, about 300, about 350, about 400, about 450, or about 500 milligrams per dose, including all doses from 0.005 to 500 milligrams.

[0223] The dosage of a composition comprising a compound of the present invention or a composition comprising it may be from about 1 μg/kg to about 200 mg/kg, from about 1 μg/kg to about 100 mg/kg, or from about 1 μg/kg to about 50 mg/kg. The dosage of the composition may include any dosage, including, but not limited to, about 1 μg/kg. The dosage of the composition may be any dosage, including, but not limited to, about 1 mcg/kg, about 10 mcg/kg, about 25 mcg/kg, about 50 mcg/kg, about 75 mcg/kg, about 100 mcg/kg, about 125 mcg/kg, about 150 mcg/kg, about 175 mcg/kg, about 200 mcg/kg, about 225 mcg/kg, about 250 mcg/kg, about 275 mcg/kg, about 300 mcg/kg, about 325 mcg/kg, about 350 mcg/kg, about 375 mcg/kg, about 400 mcg/kg, about 425 mcg/kg, about 450 mcg/kg, about 475 mcg/kg, about 500 mcg/kg, approximately 525 mcg/kg, approximately 550 mcg/kg, approximately 575 mcg/kg, approximately 600 mcg/kg, approximately 625 mcg/kg, approximately 650 mcg/kg, approximately 675 mcg/kg, approximately 700 mcg/kg, approximately 725 mcg /kg, approximately 750 mcg/kg, approximately 775 mcg/kg, approximately 800 mcg/kg, approximately 825 mcg/kg, approximately 850 mcg/kg, approximately 875 mcg/kg, approximately 900 mcg/kg, approximately 925 mcg/kg, approximately 950 mcg/kg, approximately 975 mcg/kg, approximately 1 mg/kg, approximately 5 mg/kg, approximately 10 mg/kg, approximately 15 mg/kg, about 20 mg/kg, about 25 mg/kg, about 30 mg/kg, about 35 mg/kg, about 40 mg/kg, about 45 mg/kg, about 50 mg/kg, about 60 mg/kg, about 70 mg/kg, about 80 mg/kg, about 90 mg/kg, about 100 mg/kg, about 125 mg/kg, about 150 mg/kg, about 175 mg/kg, about 200 mg/kg or more.

[0224] In certain embodiments, a compound of formula (I)-(VI) or compounds 1-5, or a pharmaceutically acceptable salt thereof, is administered every day, once daily, twice daily, three times daily, once every two days (QOD), or once every three days. The amount administered is from 0.5 mg to 100 mg, preferably from 1 mg to 80 mg, more preferably from 1 mg to 60 mg. In the most preferred embodiments, this amount is about 1 mg, 2 mg, 4 mg, 6 mg, 8 mg, 10 mg, 12 mg, 14 mg, 16 mg, 18 mg, 20 mg, 22 mg, 24 mg, 26 mg, 28 mg, 30 mg, 32 mg, 34 mg, 36 mg, 38 mg, 40 mg, 42 mg, 44 mg, 46 mg, 48 mg, 50 mg, 52 mg, 54 mg, 56 mg, 58 mg, or 60 mg.

[0225] In certain embodiments, the compound of formula (I)-(VI) or compounds 1-5 are provided in solid dosage form.

[0226] In certain embodiments, the present invention relates to the use of a compound of formula (I)-(VI) or compounds 1-5, or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for the treatment of coronavirus infection.

[0227] Various embodiments are described in the present invention. One of ordinary skill in the art, upon review of the present disclosure, will readily appreciate that the various embodiments may be combined in any variation. For example, embodiments of the present invention include treating various disorders, various patient populations, administering dosage forms in various dosages, minimizing various adverse events, and improving various efficacy rates, etc. Any combination of the various embodiments is within the scope of the present invention.

EXAMPLES

[0228] Example 1. Dose-Response Curve (DRC) Analysis by Immunofluorescence

[0229] Ten-spot DRCs were constructed for each drug. Vero cells were seeded at 1.2 x ¹⁰⁴ cells/well in DMEM supplemented with 2% FBS and 1x antibiotic and antifungal solution (Gibco) in black 384-well µClear plates (Greiner Bio-One) 24 h prior to experiment. Ten-spot DRCs were constructed with compound concentrations ranging from 0.1 to 50 μM. For viral infections, plates were transferred to a BSL3 containment facility and SARS-CoV-2 was added at a multiplicity of infection (MOI) of 0.0125. Cells were fixed 24 h post infection (hpi) with 4% PFA and analyzed by immunofluorescence. The resulting images were analyzed using in-house software to quantify cell numbers and infection rates, and antiviral activity was normalized relative to positive (mock) and negative (0.5% DMSO) controls in each assay plate. DRC was approximated using sigmoidal dose-response models with the following equation: Y = lower value + (upper value − lower value)/[1 + (IC ₅₀ /X) ^{Hill slope} ] using XLFit 4 or Prism7 software.

[0230] IC ₅₀ values were calculated from normalized curves fitting the activity data set. All IC ₅₀ values and 50% cytotoxic concentration (CC ₅₀ ) were measured in duplicate, and the quality of each assay was controlled using the Z'-factor and the percent coefficient of variation (%CV).

[0231] Following the procedure described above, compound 1 and compound 2 were tested in a SARS-CoV-2 immunofluorescence assay (IFA) against imatinib using remdesivir, chloroquine, and lopinavir as standards. The procedure and results are shown in Table 3, Fig. 5, and Figs. 6A-6F.

Table 3

Compound Maximum concentration (µM) Maximum inhibition (%) IC ₅₀ (µM) CC ₅₀ (µM) SI ₅₀ Under test Compound 1, 5 99.71 1.32 > 5 > 3.8 Imatinib 50 99.50 19.59 > 50 > 2.6 Compound 2, 50 25.93 > 50 > 50 NA Reference Remdesivir 50 95.01 12.36 > 50 > 4.0 Chloroquine 150 100.01 13.00 > 150 > 11.5 Lopinavir 50 99.93 17.29 > 50 > 2.9

[0232] Example 2. Inhibition of cytokines associated with influenza virus-induced cytokine storm

[0233] Following the method described in Front Immunol . 2019 June 21;1393 and Am. J. Physiol., Lung Cell Mol. Physiol. 315:L52-L58, 2018, U937 cells were infected with influenza virus at an MOI of 0.1 and treated with the indicated concentrations of compound 1 and compound 2. The cell culture supernatant was collected after 24 hours of treatment, and the production of the indicated cytokines was determined using a Luminex assay. The method and results are shown below in Fig. 7 and Figs. 8A-8C. The survival of mice infected with influenza virus (IFV) is shown in Table 4 and Figs.

Study group Dosage schedule Total number of animals Day 21 Log-rank test (Cox-Mantel), p-value Number of survivors Survival rate (%) Control in the form of a carrier environment QD, 2 hours before infection 9 0 0 NA Oseltamivir, 10 mg/kg 9 9 100 ****, P < 0.0001 Compound 1, 10 mg/kg 9 1 11,11 *, P = 0.0249 Compound 1, 20 mg/kg 9 2 22,22 *, P = 0.0126 Ponatinib, 15 mg/kg 9 2 22,22 *, P = 0.0419 Ponatinib, 15 mg/kg QD, 72 hours after infection 9 1 11,11 NS, P = 0.3412

[0234] As shown in Example 1 and Example 2, Compound 1 suppresses SARS-CoV-2 infection in Vero cells, suppresses the production of cytokines associated with the cytokine storm induced by the influenza virus, and, at 5 mg/kg or 10 mg/kg, improves the survival of mice infected with the influenza virus.

[0235] Example 1 and Example 2 further indicate that Compound 2 suppresses the production of cytokines associated with the cytokine storm induced by the influenza virus.

[0236] Example 3. R1PK1 Kinase Inhibition Assay

[0237] Following the method described in the manufacturer's instructions for the RIPK1 Kinase Enzyme System kit (Promega, V6930), the inhibitory effects of Compound 1 and ponatinib on RIPK1 kinase activity were determined. GSK2982772, a selective RIPK1 inhibitor, was used as a positive control. Figure 10 shows the results of the assay. The results demonstrate that Compound 1 inhibits RIPK1 kinase activity in a cell-free assay with an IC ₅₀ value of 54 nM, indicating higher activity than ponatinib (IC ₅₀ value of 231 nM).

[0238] The foregoing invention has been described in more detail by way of illustration and example for purposes of clarity and understanding. The present invention has been described with reference to various specific and preferred embodiments and methods. However, it should be understood that many changes and modifications can be made without departing from the spirit and scope of the present invention. It will be apparent to one skilled in the art that changes and modifications can be practiced within the scope of the appended claims. It should therefore be understood that the foregoing description is intended to be illustrative and not limiting.

[0239] Accordingly, the scope of the present invention should not be determined by reference to the above description, but should instead be determined by reference to the following appended claims, together with the full scope of equivalents that are entitled to legal protection within the scope of such claims.

Claims (20)

1. A method for inhibiting a viral infection in a patient in need thereof, comprising administering to the patient a therapeutically effective amount of compound 1: or a pharmaceutically acceptable salt thereof; wherein the viral infection is a coronavirus infection caused by SARS-CoV-2. 2. The method according to paragraph 1, wherein the patient suffers from COVID-19. 3. The method according to claim 1, wherein the method provides for inhibition of coronavirus polymerase. 4. The method according to claim 1, wherein the method provides for inhibition of Abelson kinase. 5. The method according to any one of paragraphs 1-4, wherein the method provides for suppression of the fusion of the coronavirus with the membrane. 6. The method according to any one of claims 1-5, wherein the method provides for inhibition of serine-threonine protein kinase 1 interacting with receptors. 7. The method according to any one of paragraphs 1-6, wherein the patient suffers from cytokine storm syndrome. 8. The method according to claim 7, wherein the method provides suppression of cytokine production in the patient. 9. The method according to any one of claims 1-8, wherein compound 1 or a pharmaceutically acceptable salt thereof is administered orally. 10. The method according to any one of claims 1-9, wherein compound 1 or a pharmaceutically acceptable salt thereof is administered once a day, twice a day, or three times a day. 11. The method according to any one of claims 1-10, wherein the therapeutically effective amount is from 0.5 mg to 100 mg, from 1 mg to 80 mg, or from 1 mg to 60 mg. 12. The method according to any one of claims 1-11, wherein compound 1 is presented in a solid dosage form. 13. Use of a pharmaceutical composition for inhibiting coronavirus infection in a person in need thereof, wherein the pharmaceutical composition comprises an effective amount of compound 1 or a pharmaceutically acceptable salt thereof, wherein the coronavirus infection is caused by SARS-CoV-2. 14. Use of compound 1 or a pharmaceutically acceptable salt thereof in inhibiting SARS-CoV-2 infection in humans. 15. Use of compound 1 or a pharmaceutically acceptable salt thereof in the manufacture of a medicinal product for inhibiting coronavirus infection, wherein the coronavirus infection is caused by SARS-CoV-2.