SARS-COV-2 attachment inhibitors and therapeutic use thereof

Abstract

The present disclosure relates to compounds that act as inhibitors against the attachment of the SARS-CoV-2 virus, pharmaceutical formulations thereof, and methods of using the compounds to prevent and treat diseases associated with the SARS-CoV-2 virus.

Description

SARS-COV-2 ATTACHMENT INHIBITORS AND THERAPEUTIC USE THEREOFSTATEMENT OF GOVERNMENT SUPPORT

[0001] This invention was made with government support under U19AI171954 and U19AI171443 awarded by the National Institutes of Health. The government has certain rights in the invention.FIELD

[0002] The present disclosure relates to compounds that act as inhibitors against the attachment of the SARS-CoV-2 virus, pharmaceutical formulations thereof, and methods of using the compounds to prevent and treat diseases associated with the SARS-CoV-2 virus.BACKGROUND

[0003] The severe acute respiratory syndrome coronavirus 2 responsible for COVID-19 remains a persistent threat to mankind, especially for the immunocompromised and elderly for which the vaccine may have limited effectiveness. Attachment of SARS-CoV-2 requires a high affinity interaction of the viral spike protein with the cellular receptor angiotensin-converting enzyme 2 (ACE2). To date there remains no FDA approved commercially available SARS-CoV-2 attachment inhibitors. Novel mutations on the spike protein correlate with the high transmissibility of new variants of SARS-CoV-2, highlighting the need for small molecule inhibitors of virus attachment onto target cells.SUMMARY

[0004] The present disclosure describes several SARS-CoV-2 attachment inhibitors that are selective over other virus entry mechanisms such as those found in the Lassa and Machupo virus lifecycles. These inhibitors exhibit excellent cytotoxicity profiles demonstrating limited toxicity in human lung cells known to express ACE2.

[0005] Provided herein are methods of inhibiting the angiotensin converting enzyme 2 (ACE2) receptor, cathepsin L, or transmembrane protease, serine 2 (TMPRSS2) activity comprising contacting the ACE2, cathepsin-L or TMPRSS2 with a compound having a structure of Formula (I):or a pharmaceutically acceptable salt thereof, wherein: R1and R1' are each independently H, OH, or Ci^alkyl; R2and R3are each independently OH, Ci-ealkyl, Ci-ealkylhydroxy, or Ci-ealkaryl, wherein when both R2and R3are each Ci-ealkyl the alkyl substituents may be taken together to form a 5-7 membered heterocycle, wherein theheterocycle may optionally contain 1, 2, or 3 additional heteroatoms selected from N, 0, and S; R4and R4’ are each independently H or halo; and R5is H, Ci-ealkyl, Ci-ealkylhyrdoxy, or a 5-7 membered aryloxy.

[0006] Provided herein are methods of inhibiting the angiotensin converting enzyme 2 (ACE2) receptor, cathepsin L, or transmembrane protease, serine 2 (TMPRSS2) activity comprising contacting the ACE2, cathepsin-L or TMPRSS2 with a compound having a structure of Formula (II):or a pharmaceutically acceptable salt thereof, wherein: R1is H or halo; R2is a 5-7 membered heteroaryl, wherein the heteroaryl contains 1 , 2, or 3 heteroatoms selected from N, 0, and S, and wherein the heteroaryl is optionally substituted with 1 , 2, or 3 Ra; R3is a Ci salkyl-N(Ra)2 or Co ealkylene-5-7 membered heterocycle, wherein the heterocycle contains 1 , 2, or 3 heteroatoms selected from N, 0, and S; and wherein the heterocycle is optionally substituted with 1 , 2, or 3 Ra; W is CH2 or 0; X is CH2 or 0; Y and Z are each independently CH2, 0, or NRa; Rais H, Ci-ealkyl, Ci-ealkylaryl, or (C(0))o-iCo-6alkyleneN(RN)2; R4and R4’, at each occurrence, is independently H, halo, Ci-ealkyl, Ci-ealkoxy, Ci-ealkaryl, or C(O)NHCi-6alkylene-5-7 membered heterocycle, wherein the heterocycle contains 1 , 2, or 3 heteroatoms selected from N, 0, and S; and RNis H or Ci-ealkyl.

[0007] Provided herein are methods of inhibiting the angiotensin converting enzyme 2 (ACE2) receptor, cathepsin L, or transmembrane protease, serine 2 (TMPRSS2) activity comprising contacting the ACE2, cathepsin-L or TMPRSS2 with a compound having a structure of Formula (III):or a pharmaceutically acceptable salt thereof, wherein: R1is a Co-eal kylene, wherein the alkylene is optionally substituted with 1 , 2, or 3 Ci-ealkyl or NH2; R2is a 5-10 membered aryl or a 5-10 membered heterocycle, wherein the heterocycle contains 1 , 2, or 3 heteroatoms selected from N, 0, and S, and wherein the aryl and the heterocycle are optionally substituted with 1 , 2, or 3 Ra; and Rais halo, Co-ealkyleneCs-ioaryl.

[0008] Provided herein are methods of inhibiting the angiotensin converting enzyme 2 (ACE2) receptor, cathepsin L, or transmembrane protease, serine 2 (TMPRSS2) activity comprising contacting the ACE2, cathepsin-L or TMPRSS2 with a compound having a structure of Formula (IV):or a pharmaceutically acceptable salt thereof, wherein: R1is Ci-ealkylene optionally substituted with Co-ealkaryl; and R2and R2are each independently C ealkoxy or Ci-ealkoxyaryl.

[0009] Additionally, provided herein are methods of treating or preventing SARS-CoV-2 in a subject comprising administering to the subject an effective amount of one or more compounds having a structure of Formula (I), Formula (II), Formula (III), Formula (IV), and pharmaceutically acceptable salts of any of the foregoing.

[0010] Further provided herein are pharmaceutical compositions comprising the compounds as disclosed herein.

[0011] Further aspects and advantages will be apparent to those of ordinary skill in the art from a review of the following detailed description, taken in conjunction with the drawings. While the compounds and methods disclosed herein are susceptible of cases in various forms, the description hereafter includes specific cases with the understanding that the disclosure is illustrative and is not intended to limit the invention to the specific cases described herein.BRIEF DESCRIPTION OF THE DRAWINGS

[0012] FIGURE 1 shows the entry receptors and co-factors of the SARS-CoV-2, MACV, and LASV viruses.

[0013] FIGURE 2 shows the firefly luciferase (Flue) luminescence and NanoLuc® luciferase (NIuc) luminescence of hACE2-H1299 cells that were screened.

[0014] FIGURE 3 shows the multiple-paired screens methodology used to identify compounds of the present disclosure.DETAILED DESCRIPTION

[0015] There is a need for small molecule inhibitors of virus attachment onto target cells to provide new and effective therapies for diseases associated with SARS-CoV-2 viruses.Compounds of the Disclosure

[0016] Compounds having a structure of Formula (I), Formula (II), Formula (III), Formula (IV), and pharmaceutically acceptable salts thereof can inhibit the angiotensin converting enzyme 2 (ACE2) receptor, cathepsin L, or transmembrane protease, serine 2 (TMPRSS2) activity. Disclosed herein are compounds having a structure of Formula (I):R1and R1' are each independently H, OH, or Ci-ealkyl;R2and R3are each independently OH, Ci-ealkyl, Ci-ealkylhydroxy, or Ci-ealkaryl, wherein when both R2and R3are each Ci-ealkyl the alkyl substituents may be taken together to form a 5-7 membered heterocycle, wherein the heterocycle may optionally contain 1 , 2, or 3 additional heteroatoms selected from N, 0, and S;R4and R4’ are each independently H or halo; andR5is H, Ci-ealkyl, C veal kylhyrdoxy, or a 5-7 membered aryloxy, or a pharmaceutically acceptable salt thereof.

[0017] In various cases, R1and R1' are each independently H or CH3. In various cases, R1and R1' are each independently H. In various cases, R1and R1' are each independently CH3. In various cases, R2and R3are each independently Ci-ealkyl or Ci-galkaryl, wherein when both R2and R3are each Ci-ealkyl the alkyl substituents may be taken together to form a 5-7 membered heterocycle. In various cases R2and R3are each independently Cisalkyl. In various cases, the alkyl groups taken together form a 6 membered heterocycle. In various cases, the alkyl groups taken together form a 7 membered heterocycle. In various cases, one of R2and R3can be a Cisalkyl and the other of R2and R3can be a Ci-salkaryl. In various cases, R4and R4’ each independently H or Cl. In various cases, R4and R4’ are each independently H. In various cases, R4and R4’ each independently Cl. In various cases R5is H, Ci-salkyl, or a 6 membered aryloxy. In various cases R5is H. In various cases R5is Cisalkyl. In various cases R5is Csalkyl. In various cases R5is / -propyl. In various cases R5is phenoxy.

[0013] Disclosed herein are compounds having a structure of Formula (II):whereinR1is H or halo;R2is a 5-7 membered heteroaryl, wherein the heteroaryl contains 1, 2, or 3 heteroatoms selected from N, 0, and S, and wherein the heteroaryl is optionally substituted with 1 , 2, or 3 Ra;R3is a Ci-6alkyl-N(Ra)2 or Co-ealkylene-5-7 membered heterocycle, wherein the heterocycle contains 1, 2, or 3 heteroatoms selected from N, 0, and S; and wherein the heterocycle is optionally substituted with 1, 2, or 3 Ra;W is CH2or O;X is CH2 or 0;Y and Z are each independently CH2, 0, or NRS;Rais H.Ci-ealkyl, Ci-ealkylaryl, or (C(0))o-iCo-6alkyleneN(RN)2;R4and R4’, at each occurrence, is independently H, halo, Ci-ealkyl, Ci-ealkoxy, Ci-ealkaryl, or C(O)NHCi. ealkylene-5-7 membered heterocycle, wherein the heterocycle contains 1 , 2, or 3 heteroatoms selected from N, 0, and S; andRNis H or Ci-ealkyl, or a pharmaceutically acceptable salt thereof.

[0014] In various cases R1is H or F. In various cases, R1is H. In various cases, R1is F. In various cases R2is 4-pyridi ne, 4-pyrazole, or 2-methyl-4-imidazole. In various cases R2is 4-pyridine or 4-pyrazole. In various cases, R2is 4-pyrazole. In various cases, R3is N, N-dimethyl-2-ethylene, N-benzyl-N-methyl-2-ethylene, N- methyl-3-pyrrolidine, or 1 -pyrrolidine. In various cases, R3is N, N-dimethyl-2-ethylene. In various cases, W is 0. In various cases, X is CH2. In various cases, Y is CH2 or NRa. In various cases, Y is CH2. In various cases, Y is NRa. In various cases, Rais H or C2alkyl. In various cases Rais H. In various cases Z is CH2. In various cases, Z is 0. In various cases, R4is H and R4’ are both H. In various cases, R4is H and R4’ is F. In various cases, R4is H and R4’ is CH3 In various cases, R4is H and R4’ is OCH3.

[0015] Disclosed herein are compounds having a structure of Formula (III):whereinR1is a Co-galkylene, wherein the alkylene is optionally substituted with 1, 2, or 3 Ci-galkyl or NH2;R2is a 5-10 membered aryl or a 5-10 membered heterocycle, wherein the heterocycle contains 1, 2, or3 heteroatoms selected from N, 0, and S, and wherein the aryl and the heterocycle are optionally substituted with 1, 2, or 3 Ra; andRais halo, Co-ealkyleneCs-ioaryl, or a pharmaceutically acceptable salt thereof.

[0016] In various cases R1is a Coalkylene and R2is 4-piperidine. In various cases, R1is a Cialkylene substituted with a NH2 and R2is a 4-fluorophenyl .

[0017] Disclosed herein are compounds having a structure of Formula (IV):whereinR1is Ci-ealkylene optionally substituted with Co-ealkaryl; andR2and R2’ are each independently Ci-ealkoxy or Ci-ealkoxyaryl, or a pharmaceutically acceptable salt thereof

[0018] In various cases, R1is Ci-2alkylene optionally substituted with a phenyl. In various cases, when R1is C2alkyl each of R2and R2’ are H.

[0019] Compounds as disclosed herein include those as provided in TABLE A or a pharmaceutically acceptable salt thereof.TABLE A

[0020] In Table A, compound numbers 1-9 have structures according to Formula (I), compound numbers 10-21 have structures according to Formula (II), compound numbers 22-23 have structures according to Formula (III), and compound numbers 24-26 have structures according to Formula (IV).

[0021] Unless otherwise indicated, structures depicted herein are also meant to include all isomeric (e.g., enantiomeric, diastereomeric, cis-trans, conformational, and rotational) forms of the structure. For example, the R and S configurations for each asymmetric center, (Z) and (E) double bond isomers, and (Z) and (E) conformational isomers are included in this disclosure, unless only one of the isomers is specifically indicated. Therefore, single stereochemical isomers as well as enantiomeric, diastereomeric, cis / trans, conformational, and rotational mixtures of the present compounds are within the scope of the disclosure. In some cases, the compounds disclosed herein are stereoisomers. "Stereoisomers" refer to compounds that differ in the chirality of one or more stereocenters. Stereoisomers include enantiomers and diastereomers. The compounds disclosed herein can exist as a single stereoisomer, or as a mixture of stereoisomers. Stereochemistry of the compounds shown herein indicate a relative stereochemistry, not absolute, unless discussed otherwise. As indicated herein, a single stereoisomer, diastereomer, or enantiomer refers to a compound that is at least more than 50% of the indicated stereoisomer, diastereomer, or enantiomer, and in some cases, at least 90% or 95% of the indicated stereoisomer, diastereomer, or enantiomer.

[0022] In some cases, the compounds of the disclosure are optically pure. As used herein, "optically pure” refers to the predominant presence of one enantiomer of a compound if multiple stereochemical configurations can exist (e.g., at least 99% enantiomeric excess).

[0023] Unless otherwise indicated, all tautomeric forms of the compounds of the disclosure are within the scope of the disclosure.

[0024] The compounds of the disclosure are defined herein by their chemical structures and / or chemical names. Where a compound is referred to by both a chemical structure and a chemical name, and the chemical structure and chemical name conflict, the chemical structure is determinative of the compound's identity.

[0025] As used herein, the term “alkyl” refers to straight chained and branched saturated hydrocarbon groups containing one to thirty carbon atoms, for example, one to twenty carbon atoms, or one to ten carbon atoms. The term Cnmeans the alkyl group has “n” carbon atoms. For example, Cealkyl refers to an alkyl group that has 6 carbon atoms. Cualkyl refers to an alkyl group having a number of carbon atoms encompassing the entire range (i.e. , 1 to 7 carbon atoms), as well as all subgroups (e.g., 1-5, 2-5, 3-6, 1, 2, 3, 4, 5, 6, and 7 carbon atoms). Nonlimiting examples of alkyl groups include, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl (2- methylpropyl), and t-butyl (1,1 -dimethylethyl). Unless otherwise indicated, an alkyl group can be an unsubstituted alkyl group or a substituted alkyl group.

[0026] As used herein, the term “alkylene” refers to a bivalent saturated aliphatic radical. The term Cnmeans the alkylene group has "n" carbon atoms, e.g., a Ci alkylene is CH2. For example, Ci-ealkylene refers to an alkylene group having a number of carbon atoms encompassing the entire range, as well as all subgroups, as previously described for "alkyl" groups.

[0027] As used herein, the term “cycloalkyl” specifically refers to a non-aromatic ring in which each atom of the ring is carbon, i.e., a carbocycle, and can be monocyclic, bicyclic, bridged, fused or spirocyclic. The term Cnmeans the cycloalkyl group has “n” ring carbon atoms. For example, C5 cycloalkyl refers to a cycloalkyl group that has 5 ring carbon atoms in the ring. C3-8 cycloalkyl refers to cycloalkyl groups having a number of ring carbon atoms encompassing the entire range (i.e., 3 to 8 carbon atoms), as well as all subgroups (e.g., 4-8, 3-7, 4-7, 3-6, 4-6, 3-5, 4-5, 3, 4, 5, 6, 7, and 8 carbon atoms). Nonlimiting examples of cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl.

[0028] As used herein, the term “heterocycle” refers to a ring which contains one to four heteroatoms independently selected from oxygen, nitrogen, and sulfur, and can be aromatic or non-aromatic (e.g., fully saturated or partially unsaturated). Additionally, heterocycles of the disclosure can be monocyclic, bicyclic, bridged, fused or spirocyclic. Nonlimiting examples of heterocycle groups include piperidine, piperazine, tetrahydrofuran, furan, tetrahydropyran, pyran, dihydrofuran, morpholine, oxazepane, oxazole, isoxazole, thiazole, pyrrole, and pyridine. Additional nonlimiting examples of heterocycle groups include benzothiazolyl, quinolyl, indole, isoquinolinyl, or quinazolinyl and the like.

[0029] As used herein, the term “alkoxy” refers to a “— O-alkyl” group.

[0030] As used herein, the term “halo” refers to a fluoro (F), chloro (Cl), bromo (Br), or iodo (I) group.

[0031] As used herein, the term “haloalkyl” refers to an alkyl group in which one or more of the hydrogen atoms are replaced by halogen In some cases, a haloalkyl group is perhalogenated (i.e., all hydrogen atoms are replaced by halogen atoms). Haloalkyl groups include but are not limited to, chloromethyl, fluoromethyl, difluoromethyl, trifluoromethyl, 1 ,1 -difluoroethyl, 2-fluoroethyl, 1 -chloro-2-fluoromethyl and 2-fluoroisobutyl

[0032] As used herein, the term “haloalkoxy” refers to an alkoxy, or O-alkyl” group in which one or more of the hydrogen atoms are replaced by a halo group. Such groups include but are not limited to, fluoromethoxy, chloromethoxy, bromomethoxy, fluoroethoxy, iodoethoxy and the like.

[0033] As used herein, the term “hydroxyalkyl” refers to an alkyl group in which one or more of the hydrogen atoms are replaced by a hydroxyl group (OH). Such groups include but are not limited to, hydroxymethyl, hydroxyethyl, and the like.

[0034] As used herein, a “substituted” functional group is a functional, group having at least one hydrogen radical that is substituted with a non-hydrogen radical (i.e., a substituent). Examples of non-hydrogen radicals (or substituents) include, but are not limited to, alkyl, cycloalkyl, alkenyl, cycloalkyl, alkynyl, ether, aryl, heteroaryl, heterocycle, hydroxyl, oxy (or oxo), alkoxyl, ester, thioester, acyl, carboxyl, cyano, nitro, amino, sulfhydryl, and halo. When a substituted alkyl group includes more than one non-hydrogen radical, the substituents can be bound to the same carbon or different carbon atoms.Pharmaceutically Acceptable Salts

[0035] As used herein, the term "pharmaceutically acceptable salt" refers to salts of a compound which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without undue side effects, such as, toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefi t / risk ratio.

[0036] Pharmaceutically acceptable salts are well known in the art. For example, S. M. Berge et al., describe pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 1977, 66, 1-19, which is incorporated herein by reference. Pharmaceutically acceptable salts of the compounds described herein include those derived from suitable inorganic and organic acids and bases. These salts can be prepared in situ during the final isolation and purification of the compounds.

[0037] Where the compound described herein contains a basic group, or a sufficiently basic bioisostere, acid addition salts can be prepared by 1) reacting the purified compound in its free-base form with a suitable organic or inorganic acid and 2) isolating the salt thus formed. In practice, acid addition salts might be a more convenient form for use and use of the salt amounts to use of the free basic form.

[0038] Examples of pharmaceutically acceptable, non-toxic acid addition salts are salts of an 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 by using other methods used in the art such as ion exchange. Other pharmaceuticallyacceptable salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, glycolate, gluconate, glycolate, hemisulfate, heptanoate, hexanoate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2- naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, palmoate, pectinate, persulfate, 3- phenylpropionate, phosphate, picrate, pivalate, propionate, salicylate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate salts, and the like.

[0039] Where the compound described herein contains a carboxyl group or a sufficiently acidic bioisostere, base addition salts can be prepared by 1) reacting the purified compound in its acid form with a suitable organic or inorganic base and 2) isolating the salt thus formed In practice, use of the base addition salt might be more convenient and use of the salt form inherently amounts to use of the free acid form. Salts derived from appropriate bases include alkali metal (e.g., sodium, lithium, and potassium), alkaline earth metal (e.g., magnesium and calcium), ammonium and N+(C i-4alkyl)4 salts. This disclosure also envisions the quaternization of any basic nitrogen-containing groups of the compounds disclosed herein. Water or oil-soluble or dispersible products may be obtained by such quaternization.

[0040] Basic addition salts include pharmaceutically acceptable metal and amine salts. Suitable metal salts include the sodium, potassium, calcium, barium, zinc, magnesium, and aluminum. The sodium and potassium salts are usually preferred. Further pharmaceutically acceptable salts include, when appropriate, nontoxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulfate, phosphate, nitrate, lower alkyl sulfonate and aryl sulfonate. Suitable inorganic base addition salts are prepared from metal bases which include sodium hydride, sodium hydroxide, potassium hydroxide, calcium hydroxide, aluminum hydroxide, lithium hydroxide, magnesium hydroxide, zinc hydroxide and the like. Suitable amine base addition salts are prepared from amines which are frequently used in medicinal chemistry because of their low toxicity and acceptability for medical use. Ammonia, ethylenediamine, N-methyl-glucamine, lysine, arginine, ornithine, choline, N,N'-dibenzylethylenediamine, chloroprocaine, dietanolamine, procaine, N- benzylphenethylamine, diethylamine, piperazine, tris(hydroxymethyl)-aminomethane, tetramethylammonium hydroxide, triethylamine, dibenzylamine, ephenamine, dehydroabietylamine, N-ethylpiperidine, benzylamine, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, ethylamine, basic amino acids, dicyclohexylamine and the like.

[0041] Other acids and bases, although not in themselves pharmaceutically acceptable, may be employed in the preparation of salts useful as intermediates in obtaining the compounds described herein and their pharmaceutically acceptable acid or base addition salts.

[0042] It should be understood that a compound disclosed herein can be present as a mixture / combination of different pharmaceutically acceptable salts. Also contemplated are mixtures / combinations of compounds in free form and pharmaceutically acceptable salts.Pharmaceutical Formulations, Doses, and Routes of Administration

[0043] Also provided herein are pharmaceutical formulations that include an effective amount of compounds of the disclosure and one or more pharmaceutically acceptable excipients. As used herein, the term "formulation” is used interchangeable with "composition.”

[0044] An "effective amount" includes both a "therapeutically effective amount" and a "prophylactically effective amount." The term "therapeutically effective amount" refers to an amount effective in treating and / or ameliorating a disease or condition in a subject. The term "prophylactically effective amount" refers to an amount effective in preventing and / or substantially lessening the chances of a disease or condition in a subject. As used herein, the terms “patient” and “subject” may be used interchangeably and mean animals, such as dogs, cats, cows, horses, and sheep (i.e., non-human animals) and humans. Particular patients or subjects are mammals (e.g., humans). The terms “patient” and “subject” include males and females.

[0045] As used herein, the term “excipient” means any pharmaceutically acceptable additive, carrier, diluent, adjuvant, or other ingredient, other than the active pharmaceutical ingredient (API), suitably selected with respect to the intended form of administration, and consistent with conventional pharmaceutical practices.

[0046] The compounds of the disclosure can be administered alone or as part of a pharmaceutically acceptable composition or formulation. In addition, the compounds can be administered all at once, as for example, by a bolus injection, multiple times, e.g. by a series of tablets, or delivered substantially uniformly over a period of time, as for example, using transdermal delivery. It is also noted that the dose of the compound can be varied over time.

[0047] The compounds disclosed herein and other pharmaceutically active compounds, if desired, can be administered to a subject or patient by any suitable route, e.g. orally, topically, rectally, parenterally, (for example, subcutaneous injections, intravenous, intramuscular, intradermal, and intrathecal injection, or infusion techniques), or as a buccal, inhalation, or nasal spray. The administration can be to provide a systemic effect (e.g. enteral or parenteral). All methods that can be used by those skilled in the art to administer a pharmaceutically active agent are contemplated. In some cases, the disclosed formulations can be administered orally or topically.

[0048] The compounds for use in the methods of the disclosure can be formulated in unit dosage form. The term "unit dosage form" refers to physically discrete units suitable as unitary dosage for subjects undergoing treatment, with each unit containing a predetermined quantity of active material calculated to produce the desired therapeutic effect, optionally in association with a suitable pharmaceutical carrier. The unit dosage form can befor a single daily dose or one of multiple daily doses (e.g. , about 1 to 4 or more times per day). When multiple daily doses are used, the unit dosage form can be the same or different for each dose.

[0049] The compounds of the disclosure can be administered to a subject or patient at dosage levels in the range of about 0.1 to about 3,000 mg per day. For a normal adult human having a body weight of about 70 kg, a dosage in the range of about 0.01 to about 100 mg per kilogram body weight is typically sufficient. The specific dosage and dosage range that will be used can potentially depend on a number of factors, including the requirements of the subject or patient, the severity of the condition or disease being treated, and the pharmacological activity of the compound being administered. The determination of dosage ranges and optimal dosages for a particular subject or patient is within the ordinary skill in the artMethods of Treatment

[0050] The compounds disclosed herein, and pharmaceutically acceptable salts thereof, can act as inhibitors of the attachment of SARS-CoV-2 viruses. Attachment of SARS-CoV-2 requires a high affinity interaction of the viral spike protein with the cellular receptor angiotensin-converting enzyme 2 (ACE2), cathepsin L, or TMPRSS2 in a subject To date there remains no FDA approved commercially available SARS-CoV-2 attachment inhibitors. Novel mutations on the spike protein correlate with the high transmissibility of new variants of SARS-CoV-2, highlighting the need for small molecule inhibitors of virus attachment onto target cells.

[0051] Thus, the disclosure provides a method of inhibiting the angiotensin converting enzyme 2 (ACE2) receptor, cathepsin L, or transmembrane protease, serine 2 (TMPRSS2) activity comprising contacting the ACE2, cathepsin-L or TMPRSS2 with a compound according to Formula (I), (II), (III), or (IV).

[0052] Further, the disclosure provides a method of preventing and / or treating a SARS-CoV-2 virus in a subject comprising contacting the SARS-CoV-2 with an effective amount of a compound or salt disclosed herein or a formulation thereof, in an amount effective to inhibit any one of ACE2, cathepsin L, and TMPRSS2. In some cases, the contacting occurs in vitro. In some cases, the contacting occurs in vivo. In some cases, the contacting comprises administering an effective amount of a compound or salt disclosed herein or a formulation thereof to a subject in need thereof. As used herein, the terms “patient” and “subject” may be used interchangeably and mean animals, such as dogs, cats, cows, horses, and sheep (i.e , non-human animals) and humans Particular patients are mammals (e.g., humans). In various cases, the subject suffers from a disease or disorder associated with SARS-CoV-2

[0053] As used herein, “administering” of a composition to a human subject or patient shall be includes method practiced on the human body as well as prescribing a composition of the disclosure that a human subject or patient will self-administer by any technique (e.g., orally, inhalation, topical application, injection, insertion, etc.).Synthesis of the Compounds of the Disclosure

[0054] The compounds of the disclosure can be synthesized by any method known in the art. Compounds 1-8 can be commercially obtained from ChemDiv with the following catalog numbers: C073-3656, C073-4521 , C073-4531, C073-2659, C073-4523, C073-4520, C073-2648, and C073-3619. These and related compounds can also be synthesized following a published method (K. Paulvannan, Preparation of Tricyclic Nitrogen Heterocycles via Tandem Four-Component Condensation I Intramolecular Diels-Alder Reaction, Tetrahedron Letters, 1999, 40, 1851-1854) Compounds 9-20 and 40 can be synthesized by published methods (J. Pan, Y Yin, L Zhao, Y. Feng, Discovery of (S)-6-methoxy-chroman-3-carboxylic acid (4-pyridin-4-yl-phenyl)-amide as potent and isoform selective ROCK2 inhibitors Bioorganic & Medicinal Chemistry (2019), 27(7), 1382-1390). Compounds 21-22 can be synthesized by published methods (Benzimidazoles and analogs as Rho kinase inhibitors and their preparation and use in the treatment of diseases, W02009079011 A1). Compounds 23-25 can be commercially obtained from Chembridge with the following catalog numbers: 5422286, 5423202, 5421803. Compound 26 can be synthesized by published methods (X. Fang, Y. Yin, Y-T. Chen, L. Yao, B. Wang, M. Cameron, L. Lin, S. Khan, C. Ruiz, T. Schroeter, W. Grant, A. Weiser, J. Pocas, A. Pachori, S.Schurer, P. LoGrasso, Y. Feng, Tetrahydroisoquinoline Derivatives As Highly Selective and Potent Rho Kinase Inhibitors, J. Medicinal Chem. 2010, 53(15), 5727-5737). Compound 27 can be synthesized by published methods (Preparation of phenylaminopyrimidine derivatives and analogs as protein kinase inhibitors, WG2009032861 A1). Compound 28 can be commercially obtained from Life Chemicals with catalog number F0526-1327. Compound 29 can be synthesized by variations on published methods (X. Song, Y. He, M. Koenig, Y. Shin, R. Noel, W. Chen, Y-Y. Ling, D. Fuerstein, L. Li, C. Ruiz, M. Cameron, D. Duckett, T. Kamenecka, Synthesis and SAR of 2, 4-diami nopyrimidines as potent c-jun N-terminal kinase inhibitors, MedChemComm, 2012, 3(2), 238-243). Compound 30 can be synthesized by variations on published methods (Preparation of benzo[d]oxazoles and benzo[d]thiazoles as kinase inhibitor, WO2010024903 A1). Compound 31 can be synthesized by variations on published methods (Nicotinic acid derivatives, Netherlands Patent NL6604123, 1966). Compound 32 can be commercially obtained from Chembridge with catalog number 11453958. Compounds 33-34 can be commercially obtained from Life Chemicals with catalog numbers F0130-0001 and F0526-1327. Compound 35 can be synthesized by published methods (R. Jiang, B. Frackowiak, Y. Shin, X.Song, W. Chen, M. Cameron. D. Duckett, T. Kamenecka, Bioorg Med Chem Lett 2013, 23(9), 2683-2687). Compounds 36-37 were commercially obtained from Chembridge with catalog numbers 94256426 and 54863471. Compound 38 can be commercially obtained from Life Chemicals with catalog number F1299-0404 Compound 39 can be commercially obtained from Sigma Aldrich with catalog number S-201 . Compound 41 can be commercially obtained from ChemDiv with catalog number 8009-7916 Compounds 42, 45, and 47 can be synthesized by published methods (Bicyclic heteroaryls as kinase inhibitors and their preparation, WG2011050245 A1). Compound 43 can be commercially obtained from Enamine with catalog number Z44611499, formerly T5255047). Compound 44 can be commercially obtained from ChemDiv with catalognumber Y508-3093. Compound 46 can be commercially obtained from Enamine with catalog number Z56915465, formerly T0516-9493).EXAMPLES

[0014] The following examples are provided for illustration and are not intended to limit the scope of the invention.Biological AssaysSARS2 Entry Assay Protocol

[0055] The hACE-2-H1299 cells were seeded at 250 cells 12 piL / well into a 1536 well assay plate (Aurora 2110-11020-s). After addition of 10 nl of test compounds or vehicle into the appropriate well, plates were incubated 24 hours at 37°C and 5% CO2. Following the incubation, 2 piL / well of assay media (RPMI-1640 supplemented with 10 % FBS, 1X Antibiotic-Antimycotic and 1 pg / ml puromycin) was dispensed for column 1-3. For column 4-48, 2 piL / well of SARS2 PV (contains either firefly or nano luciferase) was added at a 1 :10 dilution in assay media. The plates were further incubated for 48 hours at 37°C and 5% CO2. The plates were then removed from that environment and incubated at room temperature for 10 minutes followed by addition of 2.5 piL / well of One-Gio reagent (Promega N1650). After 10 minutes incubation at room temperature, firefly luciferase expression was measured using Pherastar (BGM). Subsequently, 2.5 piL / well of Dual-Glo Stop & Gio reagents (Promega N1650) was added. After 10 minutes incubation at room temperature, nano luciferase activity was measured using Pherastar (BGM). High control wells contained hACE-2-H1299 cells + assay media H- vehicle (DMSO), and the low control and data wells had hACE-2-H1299 cells + PV + test compound or vehicle.CC50 Assay Protocol

[0056] The hACE-2-H1299 cells were seeded at 250 cells / 2 piL / well for column 4-48 of the assay plates (Aurora 2110-11020-s). For columns 1-3, 2 piL / well of assay media were dispensed. After addition of 10 nl of compounds, plates were incubated 24 hours at 37°C and 5% CO2. Following that incubation, 2 piL / well of assay media to all wells was added. The plates were further incubated for 48 hours at 37°C and 5% CO2. The plates were then removed from that atmosphere and incubated at room temperature for 10 minutes followed by the addition of 4 pj L / well of CellTiter-Glo reagents (Promega G7573). After 10 minutes incubation at room temperature, luminescence was measured using Pherastar (BGM).Counterscreen (Lassa (LASV) / Machupo (MACV)) Assay Protocol

[0057] This method is identical to the “SARS2 entry assay protocol above" but a different dilution of pseudovirus was used. The assay implemented a 1 :50 dilution of pseudovirus in media for either LASV or MACV. Either LASV or MACV was added to the 2uL of pseudovirus mix that was added to the cells as described above. This yielded a paired infection specific for these pseudouvirus, whereby the luciferase reporters were used specific for SARS2 or the counterscreen virus they measured either firefly or Nanoluc allowed the measurement of the infection level of each virus exclusively, albeit in the same well.

[0058] The range of activity for each compound in the SARS-CoV-2 entry inhibition assays are reported in the table below.

[0059] It should be appreciated that all combinations of the foregoing concepts and implementations and additional concepts and implementations discussed in greater detail below are contemplated as being part of the inventive subject matter disclosed herein and may be employed in any suitable combination to achieve the benefits as described herein. In particular, all combinations of claimed subject matter appearing at the end of this disclosure are contemplated as being part of the inventive subject matter disclosed herein

[0060] The foregoing description is given for clearness of understanding only, and no unnecessary limitations should be understood therefrom, as modifications within the scope of the invention may be apparent to those having ordinary skill in the art.

[0061] Throughout this specification and the claims which follow, unless the context requires otherwise, the word “comprise” and variations such as “comprises” and “comprising” will be understood to imply the inclusion ofa stated integer or step or group of integers or steps but not the exclusion of any other integer or step or group of integers or steps.

[0062] Throughout the specification, where compositions are described as including components or materials, it is contemplated that the compositions can also consist essentially of, or consist of, any combination of the recited components or materials, unless described otherwise. Likewise, where methods are described as including particular steps, it is contemplated that the methods can also consist essentially of, or consist of, any combination of the recited steps, unless described otherwise. The invention illustratively disclosed herein suitably may be practiced in the absence of any element or step which is not specifically disclosed herein.

[0063] The practice of a method disclosed herein, and individual steps thereof, can be performed manually and / or with the aid of or automation provided by electronic equipment. Although processes have been described with reference to particular cases, a person of ordinary skill in the art will readily appreciate that other ways of performing the acts associated with the methods may be used. For example, the order of various of the steps may be changed without departing from the scope or spirit of the method, unless described otherwise. In addition, some of the individual steps can be combined, omitted, or further subdivided into additional steps.

[0064] All patents, publications and references cited herein are hereby fully incorporated by reference. In case of conflict between the present disclosure and incorporated patents, publications and references, the present disclosure should control.

Claims

What is claimed is:1 . A method of inhibiting the angiotensin converting enzyme 2 (ACE2) receptor, cathepsin L, or transmembrane protease, serine 2 (TMPRSS2) activity comprising contacting the ACE2, cathepsin-L, or TMPRSS2 with a compound, or pharmaceutically acceptable salt thereof, having a structure according to Formula (I):R1and R1' are each independently H, OH, or Ci-ealkyl;R2and R3are each independently OH, Ci-ealkyl, Ci-ealkylhydroxy, or Ci-ealkaryl, wherein when both R2and R3are each Ci-ealkyl the alkyl substituents may be taken together to form a 5-7 membered heterocycle, wherein the heterocycle may optionally contain 1 , 2, or 3 additional heteroatoms selected from N, 0, and S;R4and R4’ are each independently H or halo; andR5is H, Ci-ealkyl, C veal kylhyrdoxy, or a 5-7 membered aryloxy.

2. The method of claim 1 , wherein R1and R1’ are each independently H or CH3.

3. The method of claim 1, wherein R1and R1’ are each independently H.

4. The method of claim 1, wherein R1and R1' are each independently CH3.

5. The method of any one of claims 1 to 4, wherein R2and R3are each independently Ci-ealkyl or Ci. ealkary I, or both R2and R3taken together form a 5-7 membered heterocycle.

6. The method of any one of claims 1 to 5, wherein R2and R3are each independently Ci-3alkyl.

7. The method of any one of claims 1 to 5, wherein the alkyl groups taken together form a 6 membered heterocycle.

8. The method of any one of claims 1 to 5, wherein one of R2and R3is a Ci-3alkyl and the other of R2and R3is a Ci-salkaryl.

9. The method of any one of claims 1 to 5, wherein the alkyl groups taken together form a 7 membered heterocycle.

10. The method of any one of claims 1 to 9, wherein R4and R4’ each independently H or halo.

11. The method of any one of claims 1 to 10, wherein R4and R4’ are each independently H12. The method of any one of claims 1 to 10, wherein R4and R4’ each independently halo, for example, Cl.

13. The method of any one of claims 1 to 12, R6is H, Ci-3alkyl, or a 6 membered aryloxy.

14. The method of any one of claims 1 to 13, wherein R6is H.

15. The method of any one of claims 1 to 11, wherein R4and R4’ are each independently H and R5is Cisalkyl.

16. The method of any one of claims 1 to 11, wherein R4and R4’ are each independently H and R5is Csalkyl.

17. The method of any one of claims 1 to 11, wherein R4and R4’ are each independently H and R5is i- propyl.

18. The method of any one of claims 1 to 11, wherein R4and R4’ are each independently H and R5is phenoxy.

19. The method of any one of claims 1 to 18, wherein the contacting occurs in vitro.

20. The method of any one of claims 1 to 19, wherein the contacting occurs in vivo.

21. A method of treating or preventing SARS-CoV-2 in a subject comprising administering to the subject a therapeutically effective amount of the compound or salt as claimed in any one of claims 1 to 18.

22. A method of inhibiting the angiotensin converting enzyme 2 (ACE2) receptor, cathepsin L, or transmembrane protease, serine 2 (TMPRSS2) activity comprising contacting the ACE2, cathepsin-L, or TMPRSS2 with a compound, or pharmaceutically acceptable salt thereof, having a structure according to Formula (II):R1is H or halo;R2is a 5-7 membered heteroaryl, wherein the heteroaryl contains 1, 2, or 3 heteroatoms selected fromN, 0, and S, and wherein the heteroaryl is optionally substituted with 1 , 2, or 3 Ra;R3is a Ci-6alkyl-N(Ra)2 or Co-ealkylene-5-7 membered heterocycle, wherein the heterocycle contains 1, 2, or 3 heteroatoms selected from N, 0, and S; and wherein the heterocycle is optionally substituted with 1 , 2, or 3 Ra;W is CH2or 0;X is CFh or 0;Y and Z are each independently CH2, 0, or NRa;Rais H.Ci-ealkyl, Ci-ealkylaryl, or (C(0))o-iCo-6alkyleneN(RN)2;R4and R4’, at each occurrence, is independently H, halo, Ci-ealkyl, Ci-ealkoxy, Ci-ealkaryl, or C(O)NHCi. ealkylene-5-7 membered heterocycle, wherein the heterocycle contains 1 , 2, or 3 heteroatoms selected from N,O, and S; andRNis H or Ci-ealkyl, or a pharmaceutically acceptable salt thereof.

23. The method of claim 22, wherein R1is H or F.

24. The method of claim 22 or 23, wherein, R1is H.

25. The method of claim 22 or 23, wherein, R1is F.

26. The method of any one of claims 22 to 25, wherein R2is 4-pyridine, 4-pyrazole, or 2-methyl-4-imidazole.

27. The method of any one of claims 22 to26, wherein R2is 4-pyridine or 4-pyrazole.

28. The method of any one of claims 22 to 27, wherein R2is 4-pyrazole.

29. The method of any one of claims 22 to 28, wherein R3is N, N-dimethyl-2-ethylene, N-benzyl-N-methyl-2- ethylene, N-methyl-3-pyrrolidine, or 1 -pyrrolidine.

30. The method of any one of claims 22 to 29, wherein R3is N, N-dimethyl-2-ethylene.

31. The method of any one of claims 22 to 30, wherein W is O.

32. The method of any one of claims 22 to 31, wherein X is CH2.

33. The method of any one of claims 22 to 32, wherein Y is CH2 or NRa.

34. The method of any one of claims 22 to 33, wherein Y is CH2.

35. The method of any one of claims 22 to 33, wherein Y is NRa.

36. The method of any one of claims 22 to 35, wherein Rais H or C2alkyl.

37. The method of any one of claims 22 to 36, wherein Rais H.

38. The method of any one of claims 22 to 37, wherein Z is CH2.

39. The method of any one of claims 22 to 37, wherein Z is O.

40. The method of any one of claims 22 to 39, wherein R4is H and R are both H.

41. The method of any one of claims 22 to 39, wherein R4is H and R4’ is F.

42. The method of any one of claims 22 to 39, wherein R4is H and R4’ is CH3.

43. The method of any one of claims 22 to 39, wherein R4is H and R4’ is OCH344. The method of any one of claims 22 to 43, wherein the contacting occurs in vitro.

45. The method of any one of claims 22 to 43, wherein the contacting occurs in vivo.

46. A method of treating or preventing SARS-CoV-2 in a subject comprising administering to the subject a therapeutically effective amount of the compound or salt of claim of any one of claims 22 to 43.

47. A method of inhibiting the angiotensin converting enzyme 2 (ACE2) receptor, cathepsin L, or transmembrane protease, serine 2 (TMPRSS2) activity comprising contacting the ACE2, cathepsin-L, or TMPRSS2 with a compound, or pharmaceutically acceptable salt thereof, having a structure according to Formula (III):whereinR1is a Co-galkylene, wherein the alkylene is optionally substituted with 1, 2, or 3 C^alkyl or NH2;R2is a 5-10 membered aryl or a 5-10 membered heterocycle, wherein the heterocycle contains 1, 2, or3 heteroatoms selected from N, 0, and S, and wherein the aryl and the heterocycle are optionally substitutedwith 1, 2, or 3 Ra; andRais halo, Co-ealkyleneCs-ioaryl, or a pharmaceutically acceptable salt thereof48. The method of claim 47, wherein R1is a Coalkylene and R2is 4-piperid I ne.

49. The method of claim 47, wherein R1is a Ci alkylene substituted with a NH2 and R2is a 4-fluorophenyl.

50. The method of any one of claims 47 to 49, wherein the contacting occurs in vitro.

51. The method of any one of claims 47 to 49, wherein the contacting occurs in vivo.

52. A method of treating or preventing SARS-CoV-2 in a subject comprising administering to the subject a therapeutically effective amount of the compound or salt of any one of claims 47 to 49.

53. A method of inhibiting the angiotensin converting enzyme 2 (ACE2) receptor, cathepsin L, or transmembrane protease, serine 2 (TMPRSS2) activity comprising contacting the ACE2, cathepsin-L, or TMPRSS2 with a compound, or pharmaceutically acceptable salt thereof, having a structure according to Formula (IV):R1is Ci-6alkylene optionally substituted with Co-ealkaryl; andR2and R2’ are each independently Ci-ealkoxy or Ci-ealkoxyaryl, or a pharmaceutically acceptable salt thereof.

54. The method of claim 53, wherein R1is Ci-2alkylene optionally substituted with a phenyl.

55. The method of claim 53, wherein R1is C2al kyl and each of R2and R2’ are H.

56. The method of any one of claims 53 to 55, wherein the contacting occurs in vitro.

57. The method of any one of claims 53 to 55, wherein the contacting occurs in vivo.

58. A method of treating or preventing SARS-CoV-2 in a subject comprising administering to the subject a therapeutically effective amount of the compound or salt of any one of claims 53 to 55.

59. A method of inhibiting the angiotensin converting enzyme 2 (ACE2) receptor, cathepsin L, or transmembrane protease, serine 2 (TMPRSS2) activity comprising contacting the ACE2, cathepsin-L, or TMPRSS2 with a compound, or pharmaceutically acceptable salt thereof, of Table A.

60. A method of treating or preventing SARS-CoV-2 in a subject comprising administering to the subject a therapeutically effective amount of the compound or salt of Table A

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