Plasma kallikrein inhibitor

Abstract

The present disclosure provides compounds of Formula (IA) and pharmaceutical compositions thereof: # imgabs0. The compounds and compositions may be used to inhibit the activity of plasma kallikrein, and they may be used in therapy and in methods of treating diseases and conditions such as ocular disorders.

Description

[0001] This application claims priority to U.S. Provisional Patent Application No. 63 / 393,427, filed Jul. 29, 2022, and No. 63 / 460,204, filed Apr. 18, 2023, which are incorporated herein by reference in their entirety as if fully set forth herein. BACKGROUND OF THE INVENTION

[0002] Plasma kallikrein is a serine protease that circulates in the blood as prekallikrein, an inactive precursor, and participates in surface-mediated defense systems through signal transduction involving the activation of factor XII and high molecular weight kininogen (HK). Elements of the kallikrein-kinin system (KKS) are involved in activities such as surface-mediated defense responses, blood flow regulation, fibrin deposition, blood pressure, smooth muscle contractility, nociception, electrolyte transport, and mediator release. See Donald H. Miller, Harry S. Margolius, Chapter 19 The kallikrein-kinin-kininogen system, edited by E. Edward Bittar, Neville Bittar, Principles of Medical Biology, Elsevier, Volume 8, 1997, pages 363-384. SUMMARY OF THE INVENTION

[0003] The present disclosure provides small molecule inhibitors of plasma kallikrein and methods of treating diseases using the inhibitors. Thus, in various embodiments, the present disclosure provides a compound of formula (IA) or a pharmaceutically acceptable salt thereof:

[0004]

[0005] In formula (IA), D 1 is N or CR 1 , D 2 is N or CR 2 , D 3 is N or CR 3 , and D 4 is N or CR 4 . Further, no more than three of D 1 , D 2 , D 3 , and D 4 are simultaneously N.

[0006] Substituent R 1 , R 2 , R 3 , and R 4 are independently selected from the group consisting of: H, C 2 -C6 -Alkenyl, C 1 -C 6 -haloalkyl, halo, NR a R b , OR a 、-NR a C(O)R b 、-C(O)R a 、-C(O)halogen、-OC(O)R a 、-OC(O)OR a 、-C(O)OR a , C 6 -C 10 -Aryl, CN, -S(O) 0-2 R a 、-S(O) 2 OR a and NO 2 .

[0007] R a and R b Each independently selected from the group consisting of: H, C 1 -C 6 -alkyl and C 1 -C 6 -haloalkyl.

[0008] Substituent R c1 , R c2 and R c3 Independently selected from the group consisting of: H, C 1 -C 6 -alkyl and C 1 -C 6 -haloalkyl.

[0009] Q 1 , Q 2 and Q 3 Independently selected from the group consisting of: CR 5 , N, O and S.

[0010] R 5 Selected from the group consisting of: H, C 1 -C 6 -alkyl, OR a 、-(C 1 -C 6 -alkyl)OR a and C 3 -C 10 -cycloalkyl.

[0011] The ring member P is C or N.

[0012] L1 -SO 2 - or -C 1 -C 8 -alkylene-.

[0013] Part is a divalent monocyclic or bicyclic part selected from the group consisting of: C 3 -C 10 -cycloalkyl, C 6 -C 10 -aryl, 3- to 10-membered heterocycloalkyl (wherein 1-4 ring members are independently selected from N, O, and S), 5- to 10-membered heteroaryl (wherein 1-4 heteroaryl members are independently selected from N, O, and S), and their fused combinations.

[0014] L 2 is selected from the group consisting of: a bond, -C 1 -C 8 -alkylene, -C 2 -C 8 -alkenylene, -C 2 -C 8 -alkynylene, and a divalent part selected from the group consisting of: C 3 -C 10 -cycloalkyl, C 3 -C 10 -cycloalkenyl, C 6 -C 10 -aryl, 5- to 10-membered heteroaryl (wherein 1-4 heteroaryl members are independently selected from N, O, and S), (C 1 -C 8 -alkyl)C 3 -C 10 -cycloalkyl, (C 1 -C 8 -alkyl)C 3 -C 10 -cycloalkenyl, (C 1 -C 8 -alkyl)C 6 -C 10 -aryl and (C 1 -C 8 -alkyl) 5- to 10-membered heteroaryl (wherein 1-4 heteroaryl members are independently selected from N, O, and S). In L 2 cases, any cycloalkyl, cycloalkenyl, aryl, and heteroaryl is monocyclic or bicyclic. L 2 is optionally substituted with 1 to 3 substituents selected from the group consisting of: C 1 -C 6 -alkyl, C 1 -C 6 -haloalkyl, and CN.

[0015] Z is selected from the group consisting of: -OR c 、-OC(O)R c 、-OC(O)NR c R d 、-S(O) 0-2 R c 、-CN、-C(O)R c 、-C(O)OR c 、-C(O)NR c R d 、-C(S)R c 、-NR c R d , =NR c 、-NR c C(O)NR c R d 、-NR c CO 2 R d 、-NR c -NO, -NO 2 、-NR c -OR d , -N=C=O, -N=C=S and -NR c -NR c R d ;and

[0016] R c and R d Each instance of is independently selected from H, C 1 -C 6 -alkyl, C 1 -C 6 -haloalkyl and C 6 -C 10 -Aryl.

[0017] In one embodiment, the compound is one of Formula I:

[0018]

[0019] In Formula I, part R is i) a cyclic hydrocarbon, bicyclic hydrocarbon or heterocycle containing up to 10 atoms consisting of C or N, or R is ii) selected from the group consisting of cyclopentyl, cyclohexyl, phenyl, naphthyl, pyridine, pyrimidine, indene, 2,3-dihydro-1H-indene or any saturated or unsaturated cyclic hydrocarbon or heterocycle thereof.

[0020] Z is selected from the group consisting of: -OH, -OR', -OC(O)H, -OC(O)R', -OC(O)NH 2, -OC(O)NHR’, -OC(O)NH(R’) 2 , -SH, -SR’, -S(O)R’, -S(O) 2 R’, -CN, -C(O)H, -C(O)R’, -C(O)OH, -C(O)OR’, -C(O)NH 2 , -C(O)NHR’, -C(O)NH(R’) 2 , -C(S)R’, -NH 2 , -NH 2 R’, -NR’ 2 , =NH, =NR’, -NHC(O)NH 2 , -NR’C(O)NH 2 , -NR’C(O)NHR’, -NR’C(O)N(R’) 2 , -NHCO 2 H, -NHCO 2 R’, -NR’CO 2 R’, -NH-NO, -NR’-NO, -NO 2 , -NH-OH, -OH, -NR’-OR’, -N=C=O, -N=C=S, -NH-NH2 and -NH-NHR’, where each R’ is independently an alkyl or alkyl halide. In some embodiments, Z is not halogen or hydrogen.

[0021] Q 1 , Q 2 and Q 3 is selected from the group consisting of C, N, O or S.

[0022] P is selected from the group consisting of C or N.

[0023] L 1 is a linking group selected from the group consisting of C 1 -C 8 alkyl linker or SO 2 .

[0024] L 2 is selected from i) a hydrocarbon that does not contain a double bond formed with O or S, ii) a hydrocarbon that does not contain heteroatoms such as O, N or S; or iii) a linking group selected from the group consisting of: C 1 -C 8 alkyl linker, which includes saturated hydrocarbons, unsaturated hydrocarbons, branched hydrocarbons, cyclic hydrocarbons and combinations thereof; cyclic hydrocarbons, which are selected from the group consisting of: cyclopentyl, cyclohexyl, phenyl, naphthyl, indene, 2,3-dihydro-1H-indene or any saturated or unsaturated cyclic hydrocarbons thereof. In some embodiments, L 2 is a hydrocarbon that contains heteroatoms such as O, N or S

[0025] In some embodiments, the composition comprises a compound of Formula I and its pharmaceutically acceptable salts. Detailed Description

[0026] The present disclosure provides compounds, compositions, and methods for inhibiting plasma kallikrein (“PKal”). In exemplary embodiments, the compounds can be used to treat inflammation and ocular conditions.

[0027] Definitions

[0028] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs.

[0029] Unless the context clearly dictates otherwise, the singular forms “a / an” and “the” as used in this specification and the appended claims include plural referents. Thus, for example, reference to “an agent” includes a plurality of such agents, and reference to “the cell” includes reference to one or more cells (or pluralities of cells) and equivalents known to those skilled in the art, and so forth.

[0030] When ranges are used herein to describe physical properties (such as molecular weight) or chemical properties (such as chemical formula), all combinations and subcombinations of the ranges and specific embodiments therein are intended to be included. The term “about” when referring to a numerical value or numerical range means that the recited numerical value or numerical range is an approximation within the experimental variability (or within the statistical experimental error), and thus, in some instances, the numerical value or numerical range will vary between 1% and 15% of the recited numerical value or numerical range.

[0031] In the present disclosure, the number of atoms of a particular element in a substituent is generally given as a range, such as an alkyl group having 1 to 4 carbon atoms or C 1-4 alkyl. Reference to such a range is intended to include specific reference to groups having each integer number of atoms within the specified range. For example, an alkyl group having 1 to 4 carbon atoms includes C 1 、C 2 、C 3 and C 4 each. C 1-12 Heteroalkyl, for example, includes 1 to 12 carbon atoms in addition to one or more heteroatoms. Other numbers of atoms and other types of atoms can be indicated in a similar manner.

[0032] The term "comprising / including" (and related terms such as "comprise", "comprises", "has", or "includes") is not intended to exclude that in certain other embodiments, for example, embodiments of any substance composition, composition, method, or process described herein "consist of the features" or "consist essentially of the features".

[0033] "Alkyl" refers to a straight-chain or branched-chain hydrocarbon group having from 1 to about 20 carbon atoms. For example, an alkyl group can have from 1 to 10 carbon atoms or from 1 to 6 carbon atoms. Exemplary alkyl groups include straight-chain alkyl groups such as methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl, etc., and also include branched isomers of straight-chain alkyl groups such as, but not limited to, -CH(CH 3 ) 2 、-CH(CH 3 )(CH 2 CH 3 )、-CH(CH 2 CH3) 2 、-C(CH 3 ) 3 、-C(CH 2 CH 3 ) 3 、-CH 2 CH(CH 3 ) 2 、-CH 2 CH(CH 3 )(CH 2 CH 3 )、-CH 2 CH(CH 2 CH 3 ) 2 、-CH 2 C(CH 3 ) 3 、-CH 2 C(CH 2 CH 3 ) 3 、-CH(CH 3 )CH(CH 3 )(CH 2 CH 3 )、-CH 2 CH 2 CH(CH 3 ) 2 、-CH 2 CH 2 CH(CH 3 )(CH 2 CH3 ) , -CH 2 CH 2 CH(CH 2 CH 3 ) 2 , -CH 2 CH 2 C(CH 3 ) 3 , -CH 2 CH 2 C(CH 2 CH 3 ) 3 , -CH(CH 3 )CH 2 CH(CH 3 ) 2 , -CH(CH 3 )CH(CH 3 )CH(CH 3 ) 2 etc. Thus, alkyl groups include primary alkyl groups, secondary alkyl groups, and tertiary alkyl groups. An alkyl group may be unsubstituted or optionally substituted with one or more substituents (such as a halogen) as described herein.

[0034] Each of the terms "halogen", "halide", and "halo group" refers to -F or fluorine, -Cl or chlorine, -Br or bromine, or -I or iodine.

[0035] The term "alkenyl" refers to a straight-chain or branched-chain hydrocarbon group having 2 to about 20 carbon atoms and having 1 - 3, 1 - 2, or at least one carbon-carbon double bond. An alkenyl group may be unsubstituted or optionally substituted with one or more substituents as described herein.

[0036] "Alkyne" or "alkynyl" refers to a straight-chain or branched-chain unsaturated hydrocarbon having a specified number of carbon atoms and at least one triple bond. (C 2 -C 8 ) Examples of alkynyl groups include, but are not limited to, acetylene, propyne, 1-butyne, 2-butyne, 1-pentyne, 2-pentyne, 1-hexyne, 2-hexyne, 3-hexyne, 1-heptyne, 2-heptyne, 3-heptyne, 1-octyne, 2-octyne, 3-octyne, and 4-octyne. An alkynyl group may be unsubstituted or optionally substituted with one or more substituents as described herein.

[0037] The term "cycloalkyl" refers to a saturated monocyclic, bicyclic, tricyclic, or polycyclic 3- to 14-membered ring system, such as C 3 -C 8 cycloalkyl. A cycloalkyl group may be attached through any atom. Representative examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl. A cycloalkyl group may be unsubstituted or optionally substituted with one or more substituents as described herein.

[0038] Embodiments of plasma kallikrein inhibitors include compounds comprising a C 1 -C 8 -alkyl linker. In an embodiment, the feature of "C 1-8 -alkyl" may be a branched or unbranched hydrocarbon group having 1 to 8 carbon atoms. C 1-8 -alkyl includes, but is not limited to, methyl, ethyl, n-propyl, isopropyl, cyclopropyl, cyclopropylmethyl, n-butyl, isobutyl, sec-butyl, tert-butyl, cyclobutyl, pentyl, and cyclopentyl.

[0039] "Aryl" (Ar), when used alone or as part of another term, means a carbocyclic aromatic group having a specified number of carbon atoms whether or not fused or, if no numerical value is specified, up to 14 carbon atoms, such as C 6 -C 10- -aryl or C 6 -C 14 -aryl-aryl. In an embodiment, the feature of Ar may be an aromatic group having a cyclic system composed of carbon atoms with conjugated π electrons (e.g., phenyl). The term includes aryl groups having 6 to 12 carbon atoms. The aryl may optionally include monocyclic, bicyclic, or tricyclic rings, where each ring has five or six members. Examples of aryl include phenyl, naphthyl, biphenyl, phenanthryl, tetracenyl, etc. (see, for example, Lang’s Handbook of Chemistry (Dean, J.A., ed.), 13th ed., Table 7-2

[1985] ). "Aryl" also encompasses aryl rings as part of a fused polycyclic system, such as aryl fused to cycloalkyl as defined herein. An exemplary aryl is phenyl. The aryl may be unsubstituted or optionally substituted with one or more substituents as described herein.

[0040] The term "heteroatom" refers to N, O, and S. Compounds of the present disclosure containing N or S atoms may optionally be oxidized to the corresponding N-oxide, sulfoxide, or sulfone compounds.

[0041] "Heteroaryl", alone or in combination with any other part described herein, is a monocyclic aromatic ring structure containing one or more (e.g., 1 - 4, 1 - 3 or 1 - 2) heteroatoms independently selected from the group consisting of O, S and N and having 5 to 10 (e.g., 5 or 6) ring atoms, or a bicyclic aromatic group having 8 to 10 atoms. Heteroaryl is also intended to include oxidized S or N, such as sulfinyl, sulfonyl and N - oxides of tertiary ring nitrogen. The carbon or heteroatom is a point of attachment of the heteroaromatic ring structure, thereby producing a stable compound. Examples of heteroaryl include, but are not limited to, pyridyl, pyridazinyl, pyrazinyl, quinoxalinyl, indolizinyl, benzo[b]thienyl, quinazolinyl, purinyl, indolyl, quinolinyl, pyrimidinyl, pyrrolyl, pyrazolyl, oxazolyl, thiazolyl, thienyl, isoxazolyl, oxadiazolyl, isothiazolyl, tetrazolyl, imidazolyl, triazolyl, furyl, benzofuryl and indolyl. Heteroaryl may be unsubstituted or optionally substituted with one or more substituents as described herein.

[0042] "Heterocycloalkyl" is a saturated or partially unsaturated non - aromatic monocyclic, bicyclic, tricyclic or polycyclic system having 3 to 14 (e.g., 3 to 6) atoms, wherein 1 to 3 carbon atoms in the ring are replaced by O, S or N heteroatoms. The ring heteroatoms may also include oxidized S or N, such as sulfinyl, sulfonyl and N - oxides of tertiary ring nitrogen. Heterocycloalkyl may be fused to another ring system, such as an aryl or heteroaryl having 5 - 6 ring members. The point of attachment of the heterocycloalkyl ring is at a carbon or heteroatom, thereby allowing for a stable ring to be retained. Examples of heterocycloalkyl include, but are not limited to, morpholino, tetrahydrofuryl, dihydropyridyl, piperidyl, pyrrolidinyl, piperazinyl, dihydrobenzofuryl and dihydroindolyl. Heterocycloalkyl may be unsubstituted or optionally substituted with one or more substituents as described herein.

[0043] The terms "nitrile" or "cyano" are used interchangeably and refer to the - CN group.

[0044] The compounds described herein may exist in various isomeric forms, including configurational isomers, geometric isomers and conformational isomers, e.g., including cis - conformation or trans - conformation. The compounds may also exist in one or more tautomeric forms, including single tautomers and mixtures of tautomers. The term "isomers" is intended to cover all isomeric forms of the compounds of the present disclosure, including the tautomeric forms of the compounds. The compounds of the present disclosure may also exist in open - chain or cyclized forms. In some cases, one or more cyclized forms may be produced due to loss of water. The specific composition of the open - chain and cyclized forms may depend on the manner of separation, storage or administration of the compound. For example, the compound may exist mainly in the open - chain form under acidic conditions, but cyclize under neutral conditions. All forms are included in the present disclosure.

[0045] Some of the compounds described herein can have chiral centers and can thus exist in different enantiomeric and diastereomeric forms. The compounds described herein can be in the form of optical isomers or diastereomers. Accordingly, the present disclosure encompasses the compounds described herein and their use in the form of their optical isomers, diastereomers, and mixtures thereof, including racemic mixtures. The optical isomers of the compounds of the present disclosure can be obtained by known techniques, such as asymmetric synthesis, chiral chromatography, simulated moving bed technology, or by chemical separation of the stereoisomers using an optically active resolving agent.

[0046] Unless otherwise indicated, the term "stereoisomer" means a stereoisomer of a compound that is substantially free of other stereoisomers of that compound. Thus, a stereoisomerically pure compound having one chiral center will be substantially free of the opposite enantiomer of that compound. A stereoisomerically pure compound having two chiral centers will be substantially free of other diastereomers of that compound. A typical stereoisomerically pure compound contains greater than about 80% by weight of one stereoisomer of the compound and less than about 20% by weight of other stereoisomers of the compound, such as greater than about 90% by weight of one stereoisomer of the compound and less than about 10% by weight of other stereoisomers of the compound, or greater than about 95% by weight of one stereoisomer of the compound and less than about 5% by weight of other stereoisomers of the compound, or greater than about 97% by weight of one stereoisomer of the compound and less than about 3% by weight of other stereoisomers of the compound, or greater than about 99% by weight of one stereoisomer of the compound and less than about 1% by weight of other stereoisomers of the compound. The above stereoisomers can be considered as compositions containing two stereoisomers, the two stereoisomers being present in their respective weight percentages as described herein.

[0047] If there is a difference between the depicted structure and the name given to that structure, the depicted structure shall prevail. In addition, if the stereochemistry of a structure or a part of a structure is not indicated, for example, by bold or dashed lines, the structure or the part of the structure shall be construed to cover all of its stereoisomers. However, in some cases, when there is more than one chiral center, the structure and the name can be represented as a single enantiomer to assist in the description of relative stereochemistry. One skilled in the art of organic synthesis will know from the method used to prepare the compound whether the compound is prepared in the form of a single enantiomer.

[0048] As used herein, and unless otherwise specified to the contrary, the term "compound" is inclusive as it encompasses a compound or its pharmaceutically acceptable salts, stereoisomers, isotopologues, and / or tautomers. Thus, for example, a compound includes a pharmaceutically acceptable salt of a tautomer of the compound. Similarly, a compound includes a pharmaceutically acceptable salt of an isotopologue of the compound.

[0049] In the present disclosure, "pharmaceutically acceptable salts" are pharmaceutically acceptable organic or inorganic acid or base salts of the compounds described herein. Representative pharmaceutically acceptable salts include, for example, alkali metal salts, alkaline earth metal salts, ammonium salts, water-soluble and water-insoluble salts such as acetate, amidotrizoate (4,4-diaminostilbene-2,2-disulfonate), benzenesulfonate, benzonatate, bicarbonate, bisulfate, bitartrate, borate, bromide, butyrate, calcium salts, calcium edetate, camphorsulfonate, carbonate, chloride, citrate, clavulanate, dihydrochloride, edetate, ethanedisulfonate, dodecyl sulfate propionate, esylate, fiunarate, glucoheptonate, gluconate, glutamate, glyceryl p-aminobenzoate, hexafluorophosphate, hexylresorcinol, hydrabamine, hydrobromide, hydrochloride, hydroxynaphthoate, iodide, isothiocyanate, lactate, lactobionate, laurate, malate, maleate, mandelate, mesylate, methyl bromide, methyl nitrate, methyl sulfate, mucate, naphthalenesulfonate, nitrate, N-methylglucamine ammonium salt, 3-hydroxy-2-naphthoate, oleate, oxalate, palmitate, pamoate (1,1'-methylene-bis-2-hydroxy-3-naphthoate, embonate), pantothenate, phosphate / diphosphate, picrate, polygalacturonate, propionate, p-toluenesulfonate, salicylate, stearate, subacetate, succinate, sulfate, sulfosalicylate, suramate, tannate, tartrate, teoclate, tosylate, triethyl iodide, and valerate. A pharmaceutically acceptable salt may have more than one charged atom in its structure. In such cases, the pharmaceutically acceptable salt may have multiple counterions. Thus, a pharmaceutically acceptable salt may have one or more charged atoms and / or one or more counterions.

[0050] The term "treat / treating / treatment" refers to ameliorating or eliminating a disease or symptoms associated with a disease. In various embodiments, the term refers to minimizing or slowing the spread, progression, or worsening of a disease by administering to a patient suffering from such a disease one or more prophylactic or therapeutic compounds described herein.

[0051] The term "prevent / preventing / prevention" refers to preventing the onset, recurrence or spread of a disease in a patient by administering a compound as described herein.

[0052] The term "effective amount" means an amount of a compound or other active ingredient as described herein sufficient to provide a therapeutic or prophylactic benefit in the treatment or prevention of a disease or sufficient to delay or minimize disease-related symptoms. Additionally, a therapeutically effective amount of a compound as described herein means an amount of a therapeutic agent, alone or in combination with other therapies, that provides a therapeutic benefit in the treatment or prevention of a disease. When used in combination with a compound as described herein, the term can encompass an amount that improves the overall therapy, reduces or avoids the symptoms or causes of a disease, or enhances the therapeutic efficacy of another therapeutic agent or has a synergistic effect with another therapeutic agent.

[0053] "Patient" or "subject" includes animals such as humans, cattle, horses, sheep, lambs, pigs, chickens, turkeys, quails, cats, dogs, mice, rats, rabbits or guinea pigs. According to some embodiments, the animal is a mammal such as a non-primate and a primate (e.g., monkeys and humans). In one embodiment, the patient is a human such as a human infant, child, adolescent or adult. In the present disclosure, the terms "patient" and "subject" can be used interchangeably.

[0054] Inhibitor of plasma kallikrein

[0055] Compound of formula IA

[0056] In some embodiments, the inhibitor is a compound of formula (IA) or a pharmaceutically acceptable salt thereof:

[0057]

[0058] In formula (IA), D 1 is N or CR 1 , D 2 is N or CR 2 , D 3 is N or CR 3 , and D 4 is N or CR 4 . Additionally, no more than three of D 1 , D 2 , D 3 and D 4 are simultaneously N.

[0059] Substituent R 1 , R 2 , R 3 and R 4 are independently selected from the group consisting of: H, C 2 -C 6 -alkenyl, C1 -C 6 -haloalkyl, halo, NR a R b , OR a 、-NR a C(O)R b 、-C(O)R a 、-C(O)halogen、-OC(O)R a 、-OC(O)OR a 、-C(O)OR a , C 6 -C 10 -Aryl, CN, -S(O) 0-2 R a 、-S(O) 2 OR a and NO 2 .

[0060] R a and R b Each independently selected from the group consisting of: H, C 1 -C 6 -alkyl and C 1 -C 6 -haloalkyl.

[0061] Substituent R c1 , R c2 and R c3 Independently selected from the group consisting of: H, C 1 -C 6 -alkyl and C 1 -C 6 -haloalkyl.

[0062] Q 1 , Q 2 and Q 3 Independently selected from the group consisting of: CR 5 , N, O and S.

[0063] R 5 Selected from the group consisting of: H, C 1 -C 6 -alkyl, OR a 、-(C 1 -C 6 -alkyl)OR a and C 3 -C 10 -cycloalkyl.

[0064] The ring member P is C or N.

[0065] L 1 For-SO2 - or -C 1 -C 8 -alkylene-.

[0066] moiety is a divalent monocyclic or bicyclic moiety selected from the group consisting of: C 3 -C 10 -cycloalkyl, C 6 -C 10 -aryl, 3- to 10-membered heteroalkyl (wherein 1-4 ring members are independently selected from N, O, and S), 5- to 10-membered heteroaryl (wherein 1-4 heteroaryl members are independently selected from N, O, and S), and their fused combinations.

[0067] L 2 is selected from the group consisting of: a bond, -C 1 -C 8 -alkylene, -C 2 -C 8 -alkenylene, -C 2 -C 8 -alkynylene, and a divalent moiety selected from the group consisting of: C 3 -C 10 -cycloalkyl, C 3 -C 10 -cycloalkenyl, C 6 -C 10 -aryl, 5- to 10-membered heteroaryl (wherein 1-4 heteroaryl members are independently selected from N, O, and S), (C 1 -C 8 -alkyl)C 3 -C 10 -cycloalkyl, (C 1 -C 8 -alkyl)C 3 -C 10 -cycloalkenyl, (C 1 -C 8 -alkyl)C 6 -C 10 -aryl and (C 1 -C 8 -alkyl) 5- to 10-membered heteroaryl (wherein 1-4 heteroaryl members are independently selected from N, O, and S). In L 2 each, any cycloalkyl, cycloalkenyl, aryl, and heteroaryl is monocyclic or bicyclic. L 2 is optionally substituted with 1 to 3 substituents selected from the group consisting of: C 1 -C 6 -alkyl, C 1 -C 6 -haloalkyl, and CN.

[0068] Z is selected from the group consisting of: -OR c 、-OC(O)R c 、-OC(O)NR c R d 、-S(O) 0-2 R c 、-CN、-C(O)R c 、-C(O)OR c 、-C(O)NR c R d 、-C(S)R c 、-NR c R d 、=NR c 、-NR c C(O)NR c R d 、-NR c CO 2 R d 、-NR c -NO、-NO 2 、-NR c -OR d 、-N=C=O、-N=C=S and -NR c -NR c R d ; and

[0069] R c and R d each instance of is independently selected from H, C 1 -C 6 -alkyl, C 1 -C 6 -haloalkyl and C 6 -C 10 -aryl.

[0070] In some embodiments, three of D 1 , D 2 , D 3 and D 4 are N. In other embodiments, two of D 1 , D 2 , D 3 and D 4 are N. In other embodiments, one of D 1 , D 2 , D 3 and D 4 is N. In other embodiments, D 1 , D 2 , D 3 and D 4At most one of them is N. In an illustrative embodiment, containing D 1 、D 2 、D 3 and D 4 The ring selected is freely from the group consisting of:

[0071]

[0072] In a specific embodiment, containing D 1 、D 2 、D 3 and D 4 The ring is:

[0073]

[0074] In various embodiments, R 1 、R 2 、R 3 and R 4 are independently selected from the group consisting of: H, OR a 、halogenated group and CN. In some embodiments, R 1 、R 2 、R 3 and R 4 are independently selected from the group consisting of: H and halogenated group. For example, in one embodiment, at least one of R 1 、R 2 、R 3 and R 4 is a halogenated group. In another embodiment, R 1 、R 2 、R 3 and R 4 are each H.

[0075] According to various embodiments, some compounds of formula (IA) are those in which R c1 、R c2 and R c3 are each H. Thus, for example, some embodiments provide compounds in which the ring containing D 1 、D 2 、D 3 and D 4 is as follows:

[0076]

[0077] In various embodiments, Q 1 is C; Q 2 and Q 3 are each independently selected from the group consisting of: CR 5, N, O, and S; and P is selected from the group consisting of C and N. In one embodiment, P is N. In other embodiments, Q 2 is CR 5 and Q 3 is N, or Q 2 is N and Q 3 is CR 5 . Exemplary embodiments include compounds in which the ring containing P, Q 1 , Q 2 and Q 3 is as follows:

[0078]

[0079] In other embodiments, L 1 is -C 1 -C 8 -alkylene-, for example -C 1 -C 3 -alkylene-. In one exemplary embodiment, L 1 is methylene.

[0080] In other embodiments, moiety is a divalent monocyclic C 6 -C 10 -aryl. In one illustrative embodiment, is

[0081] In other embodiments, the present disclosure provides compounds of formula (IA) wherein L 2 is selected from -C 1 -C 8 -alkylene, C 6 -C 10 -aryl, and -(C 1 -C 8 -alkyl)C 6 -C 10 -aryl, any of which is optionally substituted as described herein. In some embodiments, L 2 is -C 1 -C 8 -alkylene, for example -C 1 -C 3 -alkylene. As generally described herein, the alkyl (or alkylene) moiety can be straight-chain or branched-chain. In other embodiments, L 2 is C 6 -C 10 -aryl, including phenyl.

[0082] In some embodiments, moiety Z is selected from the group consisting of: -OR c, CN, -C(O)OR c and -C(O)NR c R d . In various embodiments, R c and R d are each independently H or C 1 -C 6 -alkyl. Thus, for example, Z can be selected from the group consisting of: OH, OCH 3 , -COOH, -C(O)NH 2 and -C(O)NHCH 3 . In another embodiment, Z is CN.

[0083] Compound of formula I

[0084] In some embodiments, the present disclosure provides inhibitors of PKal of formula I.

[0085]

[0086] wherein R is i) a cyclic hydrocarbon, bicyclic hydrocarbon or heterocycle containing at most 10 atoms consisting of C or N, or R is ii) selected from the group consisting of: cyclopentyl, cyclohexyl, phenyl, naphthyl, pyridine, pyrimidine, indene, 2,3-dihydro-1H-indene or any of their saturated and unsaturated cyclic hydrocarbons or heterocycles.

[0087] In some embodiments, Z is selected from the group consisting of: -OH, -OR’, -OC(O)H, -OC(O)R’, -OC(O)NH 2 , -OC(O)NHR’, -OC(O)NH(R’) 2 , -SH, -SR’, -S(O)R’, -S(O) 2 R’, -CN, -C(O)H, -C(O)R’, -C(O)OH, -C(O)OR’, -C(O)NH 2 , -C(O)NHR’, -C(O)NH(R’) 2 , -C(S)R’, -NH 2 , -NH 2 R’, -NR’ 2 , =NH, =NR’, -NHC(O)NH 2 , -NR’C(O)NH 2 , -NR’C(O)NHR’, -NR’C(O)N(R’) 2 , -NHCO 2 H, -NHCO 2 R’, -NR’CO 2 R’, -NH-NO, -NR’-NO, -NO2 , -NH-OH, -OH, -NR’-OR’, -N=C=O, -N=C=S, -NH-NH2 and -NH-NHR’, wherein each R’ is independently an alkyl or alkyl halide. In some embodiments, Z is not a halogen or hydrogen.

[0088] In some embodiments, Q 1 , Q 2 and Q 3 is selected from the group consisting of C, N, O or S.

[0089] In some embodiments, P is selected from the group consisting of C or N.

[0090] In some embodiments, L 1 is a linking group selected from the group consisting of: C 1 -C 8 alkyl linker or SO 2 .

[0091] In some embodiments, L 2 is selected from i) a hydrocarbon that does not contain a double bond formed with O or S, ii) a hydrocarbon that does not contain heteroatoms such as O, N or S; or iii) a linking group selected from the group consisting of: C 1 -C 8 alkyl linker, which includes saturated hydrocarbons, unsaturated hydrocarbons, branched hydrocarbons, cyclic hydrocarbons and combinations thereof; cyclic hydrocarbons, which are selected from the group consisting of: cyclopentyl, cyclohexyl, phenyl, naphthyl, indene, 2,3-dihydro-1H-indene or any of their saturated or unsaturated cyclic hydrocarbons. In some embodiments, L 2 is a hydrocarbon that contains heteroatoms such as O, N or S.

[0092] In some embodiments, the composition comprises a compound of formula I and its pharmaceutically acceptable salts.

[0093] In some embodiments, the compound comprises a subclass wherein Q 1 , Q 2 and Q 3 are selected from the group consisting of C and N, P is N, L 1 is CH 2 , R is para-substituted benzene, L 2 is a methylene or ethylene alkyl linker para to the R group, and Z is selected from the group consisting of CN and OH.

[0094] Other embodiments include those embodiments listed below:

[0095] ·Q 1 is C, Q 2 is C, Q 3 is N, L2 is a methylene linker and Z is CN;

[0096] ·Q 1 is C, Q 2 is C, Q 3 is N, L 2 is a methylene linker and Z is OH

[0097] ·Q 1 is C, Q 2 is C, Q 3 is N, L 2 is an ethylene linker and Z is CN;

[0098] ·Q 1 is C, Q 2 is N, Q 3 is C, L 2 is an ethylene linker and Z is CN;

[0099] ·Q 1 、Q 2 and Q 3 is selected from the group consisting of C or N, P is N, L 1 is CH2, R is phenyl, L2 is selected from the group consisting of: a branched ethyl linker and an isopropyl linker para to the R group, and Z is a nitrile group (CN);

[0100] ·Q 1 is C, Q 2 is C, Q 3 is N and L 2 is a branched ethyl linker;

[0101] ·Q 1 is C, Q 2 is N, Q 3 is C and L 2 is a branched ethyl linker;

[0102] ·Q 1 is C, Q 2 is C, Q 3 is N and L 2 is an isopropyl linker;

[0103] ·Q 1 is C, Q 2 is N, Q 3 is C and L 2 is an isopropyl linker;

[0104] ·Q 1 is C, Q 2 is selected from the group consisting of N, O or S, Q 3 is selected from the group consisting of C or N, L 1is a methylene or ethylene linker, R is phenyl, L 2 is a methylene linker para to the R group and Z is CN;

[0105] ·Q 2 is N, Q 3 is C, P is N and L 1 is a methylene linker;

[0106] ·Q 2 is O, Q 3 is N, P is C and L 1 is a methylene linker;

[0107] ·Q 2 is S, Q 3 is N, P is C and L 1 is a methylene linker;

[0108] ·Q 2 is C, Q 3 is N, P is N and L 1 is an ethylene linker;

[0109] ·Q 1 is C, Q 2 is C, Q 3 is N, P is N, L 1 is CH 2 , R is phenyl, L 2 is a phenyl linker para to the R group and Z is CN;

[0110] ·Q 1 is C, Q 2 is C, Q 3 is N, P is N, L 1 is CH 2 , R is phenyl, L 2 is a benzyl linker para to the R group and Z is CN para;

[0111] ·Q 1 is C, Q 2 is C, Q 3 is N, P is N, L 1 is CH 2 , R is phenyl, L 2 is a methylene or ethylene linker para to the R group, and Z is selected from the group consisting of: methoxy, carboxylic acid, amide, and amide substituted with C1 alkyl (methyl) bonded to N;

[0112] ·Q 1 is C, Q 2 is C, Q 3 is N, P is N, L 1 is CH 2, R is benzene, L 2 is an ethylene linker at the para position of the R group and Z is methoxy;

[0113] ·Q 1 is C, Q 2 is C, Q 3 is N, P is N, L 1 is CH2, R is benzene, L 2 is a methylene linker at the para position of the R group and Z is carboxylic acid;

[0114] ·Q 1 is C, Q 2 is C, Q 3 is N, P is N, L 1 is CH 2 , R is benzene, L 2 is a methylene linker at the para position of the R group and Z is amide;

[0115] ·Q 1 is C, Q 2 is C, Q 3 is N, P is N, L 1 is CH2, R is benzene, L 2 is a methylene linker at the para position of the R group, and Z is an amide substituted with a C1 alkyl (methyl) bonded to N;

[0116] ·Q 1 is C, Q 2 is C, Q 3 is N, P is N, L 1 is CH 2 , R is benzene, L 2 is cyclopentadiene or contains a methylene linker at the 1-position of cyclopentadiene bonded to the para position of the R group, and Z is CN bonded to the 3-position of cyclopentadiene;

[0117] ·Q 1 is C, Q 2 is C, Q 3 is N, P is N, L 1 is CH2, R is benzene, L 2 is cyclopentadiene bonded to the para position of the R group, and Z is CN bonded to the 3-position of cyclopentadiene; and

[0118] ·Q 1 is C, Q 2 is C, Q 3 is N, P is N, L 1 is CH2, R is benzene, L 2 is cyclopentadiene having a methylene linker at the 1-position bonded to the para position of the R group, and Z is CN bonded to the 3-position of cyclopentadiene.

[0119] Other embodiments include those listed below:

[0120] ·Q 1 is C, Q 2 is N, Q 3 is C, P is N, L 1 is CH2, B is 1H - indene and Z is CN; and

[0121] ·Q 1 is C, Q 2 is C, Q 3 is N, P is N, L 1 is CH2, B is 2,3 - dihydro - 1H - indene and Z is CN.

[0122] Compound of formula IB

[0123] In other embodiments, optionally in combination with any other embodiment described herein, the present disclosure provides a compound of formula (IB) or a pharmaceutically acceptable salt thereof:

[0124]

[0125] Q 1 、Q 2 and Q 3 are independently selected from the group consisting of: CR 5 、N、O and S.

[0126] R 5 is selected from the group consisting of: H, C 1 -C 6 -alkyl, OR a 、-(C 1 -C 6 -alkyl)OR a and C 3 -C 10 -cycloalkyl, wherein each R a is independently selected from the group consisting of: H, C 1 -C 6 -alkyl and C 1 -C 6 -haloalkyl.

[0127] P is C or N.

[0128] L 1 is -SO 2 - or -C 1 -C 8 -alkylene-.

[0129] is a divalent monocyclic or bicyclic moiety selected from the group consisting of: C 3 -C10 - cycloalkyl, C 6 - C 10 - aryl, 3 - to 10 - membered heteroalkyl (where 1 - 4 ring members are independently selected from N, O, and S), 5 - to 10 - membered heteroaryl (where 1 - 4 heteroaryl members are independently selected from N, O, and S), and their fused combinations.

[0130] L 2 selected from the group consisting of: a bond, - C 1 - C 8 - alkylene, - C 2 - C 8 - alkenylene, - C 2 - C 8 - alkynylene, and a divalent moiety selected from the group consisting of: C 3 - C 10 - cycloalkyl, C 3 - C 10 - cycloalkenyl, C 6 - C 10 - aryl, 5 - to 10 - membered heteroaryl (where 1 - 4 heteroaryl members are independently selected from N, O, and S), (C 1 - C 8 - alkyl)C 3 - C 10 - cycloalkyl, (C 1 - C 8 - alkyl)C 3 - C 10 - cycloalkenyl, (C 1 - C 8 - alkyl)C 6 - C 10 - aryl and (C 1 - C 8 - alkyl)5 - to 10 - membered heteroaryl (where 1 - 4 heteroaryl members are independently selected from N, O, and S), where the cycloalkyl, cycloalkenyl, aryl, and heteroaryl are monocyclic or bicyclic.

[0131] L 2 optionally substituted with 1 to 3 substituents selected from the group consisting of: C 1 - C 6 - alkyl, C 1 - C 6 - haloalkyl, and CN.

[0132] Z is selected from the group consisting of: - OR c 、- OC(O)R c 、- OC(O)NR c R d 、- S(O) 0-2 R c、-CN、-C(O)R c 、-C(O)OR c 、-C(O)NR c R d 、-C(S)R c 、-NR c R d , =NR c 、-NR c C(O)NR c R d 、-NR c CO 2 R d 、-NR c -NO, -NO 2 、-NR c -OR d , -N=C=O, -N=C=S and -NR c -NR c R d .

[0133] R c and R d Each instance of is independently selected from H, C 1 -C 6 -alkyl, C 1 -C 6 -haloalkyl and C 6 -C 10 -Aryl.

[0134] Compound of formula IB-1

[0135] In other embodiments, optionally combined with any other embodiments described herein, the present disclosure provides a compound of formula (IB-1) or a pharmaceutically acceptable salt thereof:

[0136]

[0137] Q 1 , Q 2 and Q 3 Independently selected from the group consisting of: CR 5 , N, O and S.

[0138] R 5 Selected from the group consisting of: H, C 1 -C 6 -alkyl, OR a 、-(C 1 -C 6 -alkyl)OR a and C 3 -C 10 -cycloalkyl, wherein each Ra Independently selected from the group consisting of: H, C 1 -C 6 -alkyl and C 1 -C 6 -haloalkyl.

[0139] P is C or N.

[0140] R e and R f are each independently selected from the group consisting of: H, C 1 -C 6 -alkyl and C 1 -C 6 -haloalkyl.

[0141] Compound of formula IC

[0142] In other embodiments, optionally in combination with any other embodiments described herein, the present disclosure provides a compound of formula (IC) or a pharmaceutically acceptable salt thereof:

[0143]

[0144] D 1 is N or CR 1 , D 2 is N or CR 2 , D 3 is N or CR 3 and D 4 is N or CR 4 , wherein D 1 , D 2 , D 3 and D 4 no more than three of them are simultaneously N.

[0145] R 1 , R 2 , R 3 and R 4 are independently selected from the group consisting of: H, C 2 -C 6 -alkenyl, C 1 -C 6 -haloalkyl, halo, NR a R b 、OR a 、-NR a C(O)R b 、-C(O)R a 、-C(O)halo, -OC(O)R a 、-OC(O)OR a 、-C(O)OR a 、C6 -C 10 -aryl, CN, -S(O) 0-2 R a 、-S(O) 2 OR a and NO 2 。

[0146] R a and R b each independently selected from the group consisting of: H, C 1 -C 6 -alkyl and C 1 -C 6 -haloalkyl.

[0147] R c1 、R c2 and R c3 are independently selected from the group consisting of: H, C 1 -C 6 -alkyl and C 1 -C 6 -haloalkyl.

[0148] Q 1 、Q 2 and Q 3 are independently selected from the group consisting of: CR 5 、N, O and S.

[0149] R 5 is selected from the group consisting of: H, C 1 -C 6 -alkyl, OR a 、-(C 1 -C 6 -alkyl)OR a and C 3 -C 10 -cycloalkyl.

[0150] P is C or N.

[0151] L 1 is -SO 2 - or -C 1 -C 8 -alkylene-.

[0152] is a divalent monocyclic or bicyclic moiety selected from the group consisting of: C 3 -C 10 -cycloalkyl, C 6 -C 10-aryl, 3- to 10-membered heteroalkyl (wherein 1-4 ring members are independently selected from N, O, and S), 5- to 10-membered heteroaryl (wherein 1-4 heteroaryl members are independently selected from N, O, and S), and their fused combinations.

[0153] L 2 selected from the group consisting of: a bond, -C 1 -C 8 -alkylene, -C 2 -C 8 -alkenylene, -C 2 -C 8 -alkynylene, and divalent moieties selected from the group consisting of: C 3 -C 10 -cycloalkyl, C 3 -C 10 -cycloalkenyl, C 6 -C 10 -aryl, 5- to 10-membered heteroaryl (wherein 1-4 heteroaryl members are independently selected from N, O, and S), (C 1 -C 8 -alkyl)C 3 -C 10 -cycloalkyl, (C 1 -C 8 -alkyl)C 3 -C 10 -cycloalkenyl, (C 1 -C 8 -alkyl)C 6 -C 10 -aryl and (C 1 -C 8 -alkyl) 5- to 10-membered heteroaryl (wherein 1-4 heteroaryl members are independently selected from N, O, and S), wherein the cycloalkyl, cycloalkenyl, aryl, and heteroaryl are monocyclic or bicyclic.

[0154] L 2 optionally substituted with 1 to 3 substituents selected from the group consisting of: C 1 -C 6 -alkyl, C 1 -C 6 -haloalkyl, and CN.

[0155] Z is selected from the group consisting of: -OR c 、-OC(O)R c 、-OC(O)NR c R d 、-S(O) 0-2 R c 、-CN、-C(O)R c 、-C(O)OR c, -C(O)NR c R d , -C(S)R c , -NR c R d , =NR c , -NR c C(O)NR c R d , -NR c CO 2 R d , -NR c , -NO, -NO 2 , -NR c , -OR d , -N=C=O, -N=C=S and -NR c , -NR c R d .

[0156] R c and R d Each instance of R and R is independently selected from H, C 1 -C 6 -alkyl, C 1 -C 6 -haloalkyl and C 6 -C 10 -aryl.

[0157] D 1 , D 2 , D 3 or D 4 is different from CH, or at least one of R c1 , R c2 or R c3 is different from H.

[0158] Exemplary embodiments of the present disclosure are the specific compounds presented in the tables and examples as described herein.

[0159] Pharmaceutical compositions

[0160] The present disclosure also discloses pharmaceutical compositions comprising a mixture of a therapeutically effective amount of one or more compounds as described herein or pharmaceutically acceptable salts, stereoisomers and / or tautomers thereof with a pharmaceutically acceptable carrier. In some embodiments, in accordance with accepted practices of pharmaceutical compounding, the compositions also contain one or more additional therapeutic agents, pharmaceutically acceptable excipients, diluents, adjuvants, stabilizers, emulsifiers, preservatives, colorants, buffers, flavoring agents. The pharmaceutical compositions can be administered by any suitable means such that the concentration of the compound in the subject is effective for treating a disease or disorder suitable for treatment with the compounds of the present disclosure.

[0161] In some embodiments, the compound is present in an amount of 1-95 wt% of the total weight of the composition. The "therapeutically effective amount" of the administered compound or its pharmaceutically acceptable salts, stereoisomers, and / or tautomers is governed by such considerations and is the minimum amount required to cause PKal inhibition. Such amount may be lower than the amount toxic to normal cells or the entire subject. Generally, the initial therapeutically effective amount of the compounds of the present disclosure (or its pharmaceutically acceptable salts, stereoisomers, or tautomers) administered is in the range of about 0.001 to about 200 mg / kg or about 0.1 to about 20 mg / kg of patient body weight per day, where a typical initial range is about 0.3 to about 15 mg / kg / day. Oral unit dosage forms, such as tablets and capsules, may contain from about 0.1 mg to about 1000 mg of the compounds of the present disclosure (or its pharmaceutically acceptable salts, stereoisomers, or tautomers). In another embodiment, such dosage forms contain from about 50 mg to about 500 mg of the compounds of the present disclosure (or its pharmaceutically acceptable salts, stereoisomers, or tautomers). In yet another embodiment, such dosage forms contain from about 25 mg to about 200 mg of the compounds of the present disclosure (or its pharmaceutically acceptable salts, stereoisomers, or tautomers). In yet another embodiment, such dosage forms contain from about 10 mg to about 100 mg of the compounds of the present disclosure (or its pharmaceutically acceptable salts, stereoisomers, or tautomers). In another embodiment, such dosage forms contain from about 5 mg to about 50 mg of the compounds of the present disclosure (or its pharmaceutically acceptable salts, stereoisomers, or tautomers). In other embodiments, the compound is administered to a dosage form for intraocular or periocular administration as described herein, wherein the therapeutically effective amount of the compound can be from about 0.0005 mg / kg to about 0.005 mg / kg, from about 0.0007 mg / kg to about 0.004 mg / kg, or from about 0.001 mg / kg to about 0.003 mg / kg of patient body weight. In any of the foregoing embodiments, the dosage form may be administered once a day or twice a day.

[0162] Although the attending physician will ultimately determine the appropriate amount and dosage regimen, in addition to the implementation of the regimen, a therapeutically effective amount of the compounds described herein can be, for example, in the range of from 0.0035 μg to 20 μg / kg body weight / day or from 0.010 μg to 140 μg / kg body weight / week. In some embodiments, the therapeutically effective amount is in the range of from 0.025 μg to 10 μg / kg, for example, administered at least 0.025, 0.035, 0.05, 0.075, 0.1, 0.25, 0.5, 1.0, 1.5, 2.0, 2.5, 3.0, 3.5, 4.0, 5.0, 6.0, 7.0, 8.0 or 9.0 μg / kg body weight daily, every other day, or twice a week. In some embodiments, the therapeutically effective amount can be in the range of from 0.05 μg to 20 μg / kg, for example, administered at least 0.05, 0.7, 0.15, 0.2, 1.0, 2.0, 3.0, 4.0, 5.0, 6.0, 7.0, 8.0, 10.0, 12.0, 14.0, 16.0 or 18.0 μg / kg body weight weekly, every other week, or once a month. In some embodiments, the therapeutically effective amount of the compound can be, for example, in the range of from 100 μg / m 2 to 100,000 μg / m 2 (subject body surface area), administered daily, every other day, once a week, or every other week. In some embodiments, the therapeutically effective amount is in the range of from 1000 μg / m 2 to 20,000 μg / m 2 , for example, administered at least 1000, 1500, 4000 or 14,000 μg / m 2 of the compound daily, every other day, twice a week, weekly, or every other week.

[0163] In some embodiments, for an adult, the compounds of the present disclosure are administered at a dose of about 0.01 mg to 1000 mg per day (such as 0.01, 0.05, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 2.0, 3.0, 4.0, 5.0, 6.0, 7.0, 8.0, 9.0, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 110, 120, 130, 140, 150, 200, 250, 300, 350, 400, 450, 500, 550, 600, 650, 700, 750, 800, 850, 900, 950 or 1000 mg).

[0164] In certain embodiments, the compounds or pharmaceutically acceptable salts or solvates thereof described herein are substantially pure, as they contain less than about 5%, or less than about 2%, or less than about 1%, or less than about 0.5%, or less than about 0.1% of other organic small molecules, such as unreacted intermediates or synthetic by-products generated in one or more steps of, for example, a synthetic method.

[0165] In various embodiments, the compositions are provided in dosage forms suitable for oral, parenteral (e.g., intravenous, intramuscular, subcutaneous, intra-arterial), buccal, sublingual, rectal, dermal, nasal, vaginal, intranasal, inhalational, transdermal, ophthalmic, intraosseous, otic, or intracranial routes of administration. Thus, in some embodiments, the dosage forms of the compositions are selected from tablets, capsules, pills, powders, granules, suspensions, emulsions, solutions, gels (including hydrogels), pastes, patches, ointments, creams, plasters, infusions, osmotic delivery devices, suppositories, enemas, injections, implants, sprays, and aerosols. The pharmaceutical compositions are formulated according to conventional pharmaceutical practice (see, e.g., Remington: The Science and Practice of Pharmacy, 20th Edition, 2000, edited by A.R. Gennaro, Lippincott Williams & Wilkins, Philadelphia, and Encyclopedia of Pharmaceutical Technology, edited by J. Swarbrick and J.C. Boylan, 1988 - 1999, Marcel Dekker, New York).

[0166] The pharmaceutical compositions can be formulated to release the active compound immediately upon administration or at any predetermined time or time period after administration (e.g., controlled release formulations). Examples of controlled release formulations include (i) formulations that produce a substantially constant concentration of the agents of the present disclosure in the body over an extended period of time; (ii) formulations that produce a substantially constant concentration of the agents of the present disclosure in the body over an extended period of time after a predetermined lag time; (iii) formulations that maintain the action of the agent over a predetermined time period by maintaining a relatively constant effective drug level in the body, with minimalization of adverse side effects associated with fluctuations in plasma drug levels (sawtooth kinetic pattern); (iv) formulations that localize the action of the agent, e.g., by spatially placing the controlled release composition near or within the diseased tissue or organ; (v) formulations that facilitate administration, e.g., by administering the composition once a week or once every two weeks; and (vi) formulations that target the action of the agent by delivering the compound to a specific target cell type using a carrier or chemical derivative. In some embodiments, for compounds having a narrow absorption window or a relatively short biological half-life in the gastrointestinal tract, administration in the form of a controlled release formulation is desirable.

[0167] In some embodiments, controlled release is obtained by appropriately selecting various formulation parameters and ingredients, including, for example, various types of controlled release compositions and coatings. In some embodiments, the compound is formulated with a suitable excipient into a pharmaceutical composition that releases the compound in a controlled manner after administration. Examples include single or multiple unit tablet or capsule compositions, oil solutions, suspensions, emulsions, microcapsules, molecular complexes, microspheres, nanoparticles, patches, and liposomes.

[0168] The pharmaceutical compositions containing the compounds described herein can be administered parenterally via a dosage form, formulation, by injection, infusion, or implantation (e.g., intravitreal, subcutaneous, intravenous, intramuscular, intraperitoneal), or parenterally by a suitable delivery device or implant containing conventional, non-toxic pharmaceutically acceptable carriers and adjuvants. The formulation and preparation of such compositions are well known to those skilled in the art of pharmaceutical formulations.

[0169] Suitable oral compositions described herein include, but are not limited to, tablets, pills, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, syrups, or elixirs.

[0170] In another embodiment, pharmaceutical compositions suitable for single unit dosage are also contemplated, which contain a compound of the present disclosure or a pharmaceutically acceptable stereoisomer, salt, or tautomer thereof, and a pharmaceutically acceptable carrier.

[0171] The compositions of the present disclosure suitable for oral use can be prepared according to any method known in the art for manufacturing pharmaceutical compositions. For example, liquid formulations of the compounds of the present disclosure contain one or more agents selected from the group consisting of sweetening agents, flavoring agents, coloring agents, and preservatives to provide a pharmaceutically palatable formulation of the compounds of the present disclosure.

[0172] For tablet compositions, the compounds of the present disclosure are mixed with non-toxic pharmaceutically acceptable excipients for manufacturing tablets. Examples of such excipients include, but are not limited to, inert diluents such as calcium carbonate, sodium carbonate, lactose, calcium phosphate, or sodium phosphate; granulating and disintegrating agents such as corn starch or alginic acid; binding agents such as starch, gelatin, or acacia; and lubricants such as magnesium stearate, stearic acid, or talc. Tablets can be uncoated, or they can be coated by known coating techniques to delay disintegration and absorption in the gastrointestinal tract and thus provide a sustained therapeutic effect over a desired period. For example, delayed release materials such as glyceryl monostearate or glyceryl distearate can be used.

[0173] Preparations for oral use may also be presented as hard gelatin capsules wherein the active ingredient is mixed with an inert solid diluent (such as calcium carbonate, calcium phosphate or kaolin), or as soft gelatin capsules wherein the active ingredient is mixed with water or an oil medium (such as peanut oil, liquid paraffin or olive oil).

[0174] For aqueous suspensions, the compounds of the present disclosure are mixed with excipients suitable for maintaining a stable suspension. Examples of such excipients include, but are not limited to, sodium carboxymethyl cellulose, methylcellulose, hydroxypropylmethylcellulose, sodium alginate, polyvinylpyrrolidone, tragacanth and gum arabic.

[0175] Oral suspensions may also contain dispersing or wetting agents, such as natural phospholipids, such as lecithin, or condensation products of ethylene oxide with fatty acids, such as polyoxyethylene stearate, or condensation products of ethylene oxide with long chain fatty alcohols, such as heptadecaethyleneoxycetanol, or condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol, such as polyoxyethylene sorbitan monooleate, or condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol anhydrides, such as polyethylene glycol sorbitan monooleate. Aqueous suspensions may also contain one or more preservatives (such as ethyl or n-propyl p-hydroxybenzoate), one or more coloring agents, one or more flavoring agents and one or more sweetening agents (such as sucrose or saccharin).

[0176] Oily suspensions may be formulated by suspending the compounds of the present disclosure in a vegetable oil (such as peanut oil, olive oil, sesame oil or coconut oil) or a mineral oil (such as liquid paraffin). The oily suspensions may contain thickening agents, such as beeswax, hard paraffin or cetyl alcohol.

[0177] Sweetening agents (such as those mentioned above) and flavoring agents may be added to provide a palatable oral preparation. These compositions may be preserved by the addition of an antioxidant, such as ascorbic acid.

[0178] Dispersible powders and granules suitable for the preparation of aqueous suspensions by the addition of water provide the compounds of the present disclosure mixed with a dispersing or wetting agent, a suspending agent and one or more preservatives. Suitable dispersing or wetting agents and suspending agents are exemplified by those already mentioned above. Other excipients may also be present, such as sweetening agents, flavoring agents and coloring agents.

[0179] The pharmaceutical compositions of the present disclosure may also be in the form of an oil-in-water emulsion. The oil phase may be a vegetable oil (such as olive oil or peanut oil); a mineral oil (such as liquid paraffin); or a mixture thereof. Suitable emulsifiers may be naturally occurring gums (such as gum arabic or tragacanth), naturally occurring phospholipids (such as soy, lecithin), and esters or partial esters derived from fatty acids and hexitol anhydrides (such as sorbitan monooleate), and condensation products of said partial esters with ethylene oxide (such as polyoxyethylene sorbitan monooleate). The emulsion may also contain sweetening or flavoring agents.

[0180] Syrups and elixirs may be formulated with sweetening agents such as, for example, glycerin, propylene glycol, sorbitol or sucrose. Such formulations may also contain demulcents, preservatives and flavoring and coloring agents. The pharmaceutical compositions may be in the form of a sterile injectable aqueous suspension or an oily suspension. Such suspensions may be formulated according to known techniques using those suitable dispersing or wetting agents and suspending agents which have been mentioned above. The sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally acceptable diluent or solvent, for example in the form of a solution in 1,3 - butanediol. Acceptable vehicles and solvents that may be employed are, in particular, water, Ringer's solution and isotonic sodium chloride solution. In addition, a sterile, non-volatile oil is conventionally used as a solvent or suspending medium. For this purpose, any mild non-volatile oil may be used, including synthetic mono- or diglycerides. Additionally, fatty acids such as oleic acid may be used in the preparation of injectables.

[0181] The compounds of the present disclosure may also be administered in the form of suppositories for rectal administration of drugs. These compositions may be prepared by mixing the drug with a suitable non-irritating excipient which is solid at ordinary temperature but liquid at rectal temperature and will therefore melt in the rectum to release the drug. Such materials are cocoa butter and polyethylene glycol.

[0182] Compositions for parenteral administration are administered in a sterile medium. Depending on the vehicle and the concentration of the drug in the formulation, the parenteral preparation may be a suspension or a solution containing the dissolved drug. Adjuvants such as, for example, local anesthetics, preservatives and buffering agents may also be added to the parenteral compositions.

[0183] In some embodiments, the composition is particularly suitable for administration into or around the eye. For example, the composition may be suitable for use as an eye drop, or for injection into the eye, such as by periocular injection or intravitreal injection. Such compositions should be sterile and substantially endotoxin-free, and have a pH within an acceptable range. In some embodiments, a preservative-free formulation is used. The formulation of ocular drugs is known in the art, see, for example, Ocular Therapeutics and Drug Delivery: A Multi-Disciplinary Approach, edited by Reddy (CRC Press 1995); Kaur and Kanwar, Drug Dev Ind Pharm. May 2002; 28(5):473-93; Clinical Ocular Pharmacology, Bartlett et al. (Butterworth-Heinemann; 4th Edition (March 15, 2001)); and Ophthalmic Drug Delivery Systems (Drugs and the Pharmaceutical Sciences: a Series of Textbooks and Monographs), Mitra (Marcel Dekker; 2nd Rev & Ex Edition (March 1, 2003)).

[0184] Compositions for parenteral use may be provided in unit dosage forms (e.g., single-dose ampoules), or in vials containing several doses and may include a suitable preservative (see below). The composition may be in the form of a solution, suspension, emulsion, infusion device or implantable delivery device, or it may be presented as a dry powder to be reconstituted with water or other suitable vehicle before use. In addition to the active agent, the composition may also include suitable parenterally acceptable carriers and / or excipients. The active agent may be incorporated into microspheres, microcapsules, nanoparticles, liposomes, etc. to control release. In addition, the composition may also contain suspending agents, solubilizing agents, stabilizers, pH regulators, tonicity regulators and / or dispersing agents.

[0185] In some embodiments, the pharmaceutical compositions of the present disclosure are in a form suitable for sterile injection. To prepare such compositions, the active agent is dissolved or suspended in a parenterally acceptable liquid vehicle. Acceptable vehicles and solvents that can be used are, in particular, water, water adjusted to a suitable pH by the addition of a suitable amount of hydrochloric acid, sodium hydroxide, or a suitable buffer, 1,3-butanediol, Ringer's solution, dextrose solution, and isotonic sodium chloride solution. The aqueous formulations may also contain one or more preservatives (e.g., methylparaben, ethylparaben, or propylparaben). If the compound has limited solubility in water, a solubilizing enhancer or solubilizer may be added, or the solvent may include 10 - 60% w / w propylene glycol.

[0186] The pharmaceutical composition can be administered to a subject in a single-dose or multi-dose form. For example, the compounds described herein can be administered once a week or for 2, 3, 4, 5, 6, 7, 8, 10, 15, 20 or more weeks. It should be understood that for any particular subject, the specific dosage regimen should be adjusted over time according to individual needs and the professional judgment of the health care provider who is managing or supervising the administration of the compound. For example, if a lower dose does not provide sufficient biological activity (e.g., in treating the diseases or disorders described herein), the dose of the compound can be increased. Conversely, if the disease or disorder is alleviated or eliminated, or to reduce adverse side effects, the dose of the compound can be decreased.

[0187] Methods of Use and Treatment

[0188] As an advantage of the present disclosure, the compounds described herein are effective plasma kallikrein inhibitors, that is, the compounds can reduce the activity of plasma kallikrein. In various embodiments, the compounds described herein may be characterized by an inhibition constant IC 50 (half maximal inhibitory concentration) of no more than 500 nM (e.g., less than 500, 450, 400, 350, 300, 250, 200, 150, 100, 50, 10, 1 or 0.1 nM).

[0189] In one embodiment, the present disclosure provides a method of inhibiting plasma kallikrein. In some embodiments, the method comprises contacting PKal with an amount of a compound of the present disclosure effective to inhibit the activity of PKal. In some embodiments, the compounds of the present disclosure inhibit the activity of PKal, with an IC 50The value is in the range of 0.1 to 500 nM (such as 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 2.0, 3.0, 4.0, 5.0, 6.0, 7.0, 8.0, 9.0, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 150, 200, 250, 300, 350, 400, 450 or 500 nM). It should be understood that all ranges in the present disclosure include one or both endpoints, all values accurate to one significant figure between the endpoints, and any sub-ranges between the endpoints. In some embodiments, the compounds of the present disclosure inhibit PKal activity, and their IC 50 is less than or equal to 500 nM, such as 500, 450, 400, 350, 300, 250, 200, 150, 100, 95, 90, 85, 80, 75, 70, 65, 60, 55, 50, 45, 40, 35, 30, 25, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2 or 0.1 nM.

[0190] In some embodiments, the method includes administering to a subject an effective amount of a compound of the present disclosure or a pharmaceutically acceptable salt thereof. In some embodiments, the method inhibits PKal activity in vivo, and its IC 50 (half maximal inhibitory concentration) value is in the range of 0.1 to 500 nM (such as 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 2.0, 3.0, 4.0, 5.0, 6.0, 7.0, 8.0, 9.0, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 150, 200, 250, 300, 350, 400, 450 or 500 nM). In some embodiments, the method inhibits PKal activity in vivo, and its IC 50 is greater than 100 nM, such as 110, 120, 130, 140, 150, 160, 170, 180, 190 or 200 nM.

[0191] In other embodiments, the present disclosure provides methods for treating a subject suffering from a disease or disorder. The method comprises administering to the subject an effective amount of a compound disclosed herein or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof. In some embodiments, the compound is selected from the group consisting of any compound in Table 1, combinations thereof, and pharmaceutically acceptable salts thereof.

[0192] In various embodiments, the disease or disorder is selected from the group consisting of: ischemic stroke, hemorrhagic stroke, hypertension, retinopathy, diabetic retinopathy, nephropathy, cerebral edema, pulmonary hypertension, inflammation, acute myocardial infarction, deep vein thrombosis, complications from fibrinolytic therapy, stroke, angina, angioedema, sepsis, arthritis, cardiopulmonary bypass complications, capillary leak syndrome, inflammatory bowel disease, vascular complications from diabetes, diabetic macular edema, macular degeneration, neuropathy, age-related macular degeneration, retinal vein occlusion, cerebral edema, ischemia-reperfusion injury, angiogenesis, asthma, allergic reaction, Alzheimer's disease, Parkinson's disease, multiple sclerosis, glioblastoma multiforme, complications from fibrinolytic therapy, increased albumin excretion, macroalbuminuria, pain, amyotrophic lateral sclerosis, Creutzfeldt-Jakob disease, epilepsy, traumatic brain injury, high altitude cerebral edema, cancer, disseminated intravascular coagulation, pancreatitis, inflammation, shock, hereditary angioedema (HAE), uveitis, polyangiitis, acute respiratory distress syndrome (ARDS), thrombosis, vasculitis, Crohn's disease, ulcerative colitis, enterocolitis, arteritis, glomerulonephritis, psoriasis, endometriosis, preeclampsia, malaria, arthritis, periodic and recurrent fevers, Chagas disease, Raynaud's disease, systemic sclerosis, granulomatosis with polyangiitis, small vessel vasculitis, medium vessel vasculitis, large vessel vasculitis, panvasculitis, systemic autoinflammatory diseases, renal insufficiency, cerebral malaria, Clarkson's disease (systemic vascular leak syndrome), hantavirus infection, hantavirus renal syndrome, hantavirus pulmonary syndrome, virus-associated inflammatory disorders, retinal vasculitis, uveitis, Eales' disease, Behçet's disease, sarcoidosis, whooping cough, coronavirus infection, and non-infectious posterior uveitis.

[0193] In some embodiments, the subject is an animal, such as a human or a non-human animal (e.g., a mammal), and may be used interchangeably with "patient" when the subject is receiving medical care from a healthcare provider.

[0194] Combination therapy

[0195] The present disclosure also provides a combination pharmaceutical composition, which comprises: (a) at least one compound disclosed herein or a pharmaceutically acceptable salt thereof, and (b) at least one inhibitor of inflammation, pain or edema. In some embodiments, an orally deliverable pharmaceutical composition comprising the combination pharmaceutical composition of the present disclosure is provided. The present disclosure further provides tablets, capsules, orally deliverable granules, injection suspensions and solutions, and compositions for pulmonary or nasal delivery comprising the combination pharmaceutical composition.

[0196] Kit

[0197] In another embodiment, the present disclosure provides a kit comprising a compound or a pharmaceutically acceptable salt thereof as described herein. The kit further includes instructions from a healthcare provider for administering the compound to a patient.

[0198] Examples

[0199] The following examples provide further embodiments of the present disclosure. The examples are illustrative and not restrictive. Although substantially any compositions, compounds and methods similar to those described herein can be used to practice or test the present disclosure, only illustrative compositions, compounds and methods are described.

[0200] Synthesis of Compounds

[0201] As described in the examples below, in certain illustrative embodiments, compounds are prepared according to the following general procedures. It should be understood that although the general methods describe the synthesis of certain compounds of the present invention, the following general methods, as well as other methods known to those of ordinary skill in the art, can be applied to all compounds described herein and to each subclass and species of these compounds. Other compounds of the present invention are prepared by methods known to those of skill in the art that are substantially similar to the methods described herein in the examples.

[0202] Preparation of Intermediate Compounds

[0203] Example 1A: Preparation of Ethyl 1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxylate (7_Int-5)

[0204]

[0205] Step-1. Synthesis of 2-Methyl-2-(p-tolyl)propanenitrile (7_Int-2).

[0206] At 0 °C to 5 °C under a nitrogen atmosphere, a solution of 2-(p-tolyl)acetonitrile (200 g, 1524 mmol, 1.0 eq) in N-methyl-2-pyrrolidone (1000 mL, 5V) and tetrahydrofuran (1000 mL, 5V) was added dropwise to sodium tert-butoxide (586.07 g, 6098 mmol, 4.0 eq). The reaction mixture (RM) was stirred at 0 °C to 5 °C for 30 min. At 0 °C to 5 °C, iodomethane (865.62 g, 6098 mmol, 4.0 eq) was added dropwise to the RM. The RM was stirred at 0 °C to RT for 2.5 h. After the reaction was completed, the RM was transferred to deionized water (400 mL) and the product was extracted with ethyl acetate (2000 mL). The combined organic fractions were washed with cold water (4 x 500 mL) to remove N-methyl-2-pyrrolidone, concentrated and then purified by column chromatography (2 - 5% EtOAc in hexane) to give compound 7_Int-2. 1 H NMR (400 MHz, DMSO-d6) δ 1.651 (s, 6H), 2.298 (s, 3H), 7.226 d, J = 8 Hz, 2H), 7.392 (d, J = 6.8 Hz, 2H).

[0207] Step - 2. Synthesis of 2-(4-(bromomethyl)phenyl)-2-methylpropanenitrile (7_Int-3).

[0208] A 10000 mL 4N round bottom flask (RBF) attached to a mechanical stirrer and condenser was charged with 2-methyl-2-(p-tolyl)propanenitrile (170 g, 1067 mmol, 1.0 eq) and carbon tetrachloride (3400 mL, 20V) at RT. AIBN (17.53 g, 106 mmol, 0.1 eq) was added to the RM. At RT, N-bromosuccinimide (209.04 g, 1174 mmol, 1.1 eq) was added dropwise to the RM. The resulting RM was heated to 90 °C and stirred for 2 h. Note: After the reaction was completed, the reaction mixture was cooled to RT. The RM was quenched in DM water (3400 mL) and the product was extracted with DCM (2 x 2000 mL). The combined organic fractions were concentrated and then purified by column chromatography (7% EtOAc in hexane) to give compound (7_Int-3). 1 H NMR (400 MHz, DMSO-d6) δ 1.654 (s, 6H), 4.714 (s, 2H), 7.507 (s, 4H).

[0209] Step - 3. Synthesis of ethyl 1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxylate (7_Int-5).

[0210] Charge a 3000 mL 4N RBF attached to a mechanical stirrer and condenser with ethyl 1H-pyrazole-4-carboxylate (70 g, 499 mmol, 1.0 eq), 2-(4-(bromomethyl)phenyl)-2-methylpropanenitrile (130.84 g, 549 mmol, 1.1 eq), and acetone (700 mL, 10V) at RT. Add Cs 2 CO 3 (390.8 g, 1198 mmol, 2.4 eq) to the RM. Heat the RM to 60 - 65 °C and stir for 6 h. After completion of the reaction, cool the reaction mixture to RT. Filter the RM to remove Cs 2 CO 3 ; wash with EtOAc. Concentrate the filtrate and then purify by column chromatography (ethyl acetate in hexane 15%) to give the compound (7_Int-5). 1 1H NMR (400 MHz, DMSO-d6) δ 1.656 (s, 6H), 4.204 (q, J = 6.8 Hz, 2H), 5.372 (s, 2H), 7.328 (d, J = 8 Hz, 2H), 7.498 (d, J = 8 Hz, 2H), 8.053 (s, 1H), 8.482 (s, 1H).

[0211] Example 2A: Preparation of ethyl 1-(4-(cyanomethyl)benzyl)-1H-pyrazole-4-carboxylate (1_Int-4)

[0212]

[0213] Step - 1. Synthesis of ethyl 1-(4-methylbenzyl)-1H-pyrazole-4-carboxylate (1_Int-2).

[0214] Charge a 10000 mL 4N RBF attached to a mechanical stirrer and condenser with ethyl 1H-pyrazole-4-carboxylate (250 g, 1783 mmol, 1.0 eq), 1-(bromomethyl)-4-methylbenzene (363.1 g, 1962 mmol, 1.1 eq), and acetone (6250 mL, 25V) at RT. Add Cs 2 CO 3 (390.8 g, 1198 mmol, 2.4 eq) to the RM. Heat the RM to 60 - 65 °C and stir for 16 h. After completion of the reaction, cool the reaction mixture to RT. Filter the RM to remove Cs 2 CO 3 ; wash with EtOAc; concentrate the filtrate under reduced pressure to obtain a crude product. Purify the crude product by trituration in hexane to give the compound (1_Int-2). 11H NMR (400 MHz, DMSO-d6) δ 1.227 (t, J = 6.8 Hz, 3H), 2.247 (s, 3H), 4.176 (q, J = 6.8 Hz, 2H), 5.282 (s, 2H), 7.144 - 7.136 (m, 4H), 7.828 (s, 1H), 8.390 (s, 1H).

[0215] Step - 2. Synthesis of ethyl 1-(4-(bromomethyl)benzyl)-1H-pyrazole-4-carboxylate (1_Int-3).

[0216] Charge a 10000 mL 4N RBF attached to a mechanical stirrer and condenser with ethyl 1-(4-methylbenzyl)-1H-pyrazole-4-carboxylate (300 g, 1228 mmol, 1.0 eq) and 1,2-dichloroethane (6000 mL, 20V) at RT. Add benzoyl peroxide (29.71 g, 122.8 mmol, 0.1 eq) to the RM. Add N-bromosuccinimide (240.4 g, 1350 mmol, 1.1 eq) dropwise to the RM at RT. Heat the resulting RM to 90 °C and stir for 4 h. After the reaction is complete, cool the reaction mixture to RT. Dilute the RM with DCM (2000 mL). Wash the RM with (2 x 4000 ml) saturated Na 2 SO 4 solution and concentrate under reduced pressure to obtain a crude product. Purify the crude product by column chromatography (17 - 25% EtOAc in hexane) to give the compound (1_Int-3). 1 1H NMR (400 MHz, DMSO-d6) δ 1.270 - 1.235 (m, 3H), 4.207 (q, J = 6.8 Hz, 2H), 4.682 (s, 2H), 5.364 (s, 2H), 7.243 (d, J = 8.4 Hz, 4H), 7.421 (d, J = 8.4 Hz, 2H), 7.868 (s, 1H), 8.478 (s, 1H).

[0217] Step - 3. Synthesis of ethyl 1-(4-(cyanomethyl)benzyl)-1H-pyrazole-4-carboxylate (1_Int-4).

[0218] Charge a reaction vessel with ethyl 1-(4-(bromomethyl)benzyl)-1H-pyrazole-4-carboxylate (242 g, 499748 mmol, 1.0 eq), ACN (2420 mL, 10V) and Cs 2 CO 3(488 g, 1497 mmol, 2.0 eq) was charged to a 5000 mL 4N RBF equipped with a mechanical stirrer and a condenser, and then TMSCN (334 g, 3369 mmol, 4.5 eq) was slowly added at RT. The RM was heated to 80 - 85 °C and stirred for 16 h. After the reaction was completed, the reaction mixture was cooled to RT. The RM was filtered to remove Cs 2 CO 3 ; washed with EtOAc. The solid residue was discarded and the filtrate was concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (12 - 15% EtOAc in hexane) to give the compound (1_Int - 4). 1 1H NMR (400 MHz, DMSO - d6) δ 1.282 - 1.234 (m, 3H), 4.016 (s, 2H), 4.20 (q, J = 7.2 Hz, 2H), 5.363 (s, 2H), 7.339 - 7.280 (m, 4H), 8.051 (s, 1H), 8.468 (s, 1H).

[0219] Example 3A: Preparation of Ethyl 1-(4-((3-cyano-1H-pyrrol-1-yl)methyl)benzyl)-1H-pyrazole-4-carboxylate (6_Int-1)

[0220]

[0221] Step - 1. Synthesis of Ethyl 1-(4-((3-cyano-1H-pyrrol-1-yl)methyl)benzyl)-1H-pyrazole-4-carboxylate (6_Int-1).

[0222] A stirred solution of 1H - pyrrole - 3 - carbonitrile (0.2 g, 2.16 mmol, 1.0 eq) in DMF (2 mL, 10V) was prepared and NaH (60% in mineral oil) (0.156 g, 3.24 mmol, 1.5 eq) was added at 0 °C. The RM was stirred at 0 °C for 30 min. At 0 °C, Ethyl 1-(4-(bromomethyl)benzyl)-1H-pyrazole-4-carboxylate (1_Int-3) (0.772 g, 2.39 mmol, 1.1 eq) was added to the RM. The RM was warmed to RT and stirred for 3 h. After the reaction was completed, the RM was quenched in water and extracted with ethyl acetate. The combined organic fractions were dried over Na 2 SO 4 dried, concentrated and then purified by column chromatography (20% ethyl acetate in hexane) to give the compound (6_Int-1). MS (ES): 335.30 m / z [M + H]+, LCMS purity: 95.09%, 11H NMR (400 MHz, DMSO-d6) δ 1.246 (t, J = 8 Hz, 3H), 4.194 (q, J = 7.2 Hz, 2H), 5.336 (s, 2H), 6.987 (t, J = 2.4 Hz, 1H), 7.267 - 7.208 (m, 4H), 7.699 (t, J = 2 Hz, 1H), 7.845 (s, 1H), 8.448 (s, 1H).

[0223] Example 4A: Preparation of 4-(aminomethyl)-2-fluorobenzamidine dihydrochloride (43_Int-5)

[0224]

[0225] Step - 1. Synthesis of tert-butyl (4-cyano-2-fluorobenzyl)carbamate (43_Int-2).

[0226] Prepare a stirred solution of 4-(aminomethyl)-2-fluorobenzonitrile (43_Int-1) (3 g, 19.97 mmol, 1.0 eq) in 1,4-dioxane (36 mL, 12V) and 2N NaOH (18 mL, 6V) and add di-tert-butyl dicarbonate (4.79 g, 21.97 mmol, 1.1 eq) at 0 °C. Stir the reaction mixture at RT for 3 h. After completion of the reaction, quench the RM in water and extract with ethyl acetate. Combine the organic fractions and dry over Na 2 SO 4 dry; concentrate to give the compound (43_Int-2). MS (ES): 251.00. m / z [M+H]+, LCMS purity: 100%, 1 1H NMR (400 MHz, DMSO-d6) δ 1.393 (s, 9H), 4.216 (d, J = 5.2 Hz, 2H), 7.281 (t, J = 7.5 Hz, 2H), 7.551 (s, 1H), 7.885 (t, J = 8 Hz, 1H)

[0227] Step - 2. Synthesis of tert-butyl (3-fluoro-4-(N-hydroxyformamidinyl)benzyl)carbamate (43_Int-3).

[0228] Prepare a stirred solution of tert-butyl (4-cyano-3-fluorobenzyl)carbamate (43_Int-2) (3.4 g, 13.58 mmol, 1.0 eq) and methanol (34 mL, 10 V) and add hydroxylamine hydrochloride (1.604 g, 23.09 mmol, 1.7 eq) and DIPEA (3.95 g, 23.09 mmol, 1.7 eq) at RT. Stir the RM at 70 °C for 16 h. After completion of the reaction, evaporate the RM, quench the residue in water and extract with 10% methanol in DCM. Dry the combined organic fractions over Na 2 SO 4 dry; concentrate to give compound (43_Int-3). MS (ES): 284.10 m / z [M+H]+ LCMS purity: 81.45%, 1 1H NMR (400 MHz, DMSO-d6) δ 1.393 (s, 9H), 4.133 (d, J = 5.2 Hz, 2H), 7.101 - 70.31 (m, 2H), 7.125 (s, 1H), 7.483 - 7.417 (m, 2H), 7.611 (t, J = 8 Hz, 1H), 9.593 (s, 1H).

[0229] Step - 3. Synthesis of tert-butyl (4-formamidin-3-fluorobenzyl)carbamate (43_Int-4).

[0230] Prepare a stirred solution of tert-butyl (3-fluoro-4-(N-hydroxymethylformamidine)benzyl)carbamate (43_Int-3) (3.5 g, 12.35 mmol, 1.0 eq) and methanol (35 mL, 10 V) and add ammonium formate (2.34 g, 37.06 mmol, 3.0 eq) and 10% Pd / C (0175 g, 0.065% w / w) at RT. Stir the RM in an autoclave at 70 °C under 20 kg H2 pressure for 16 h. After completion of the reaction, filter the RM through a bed of diatomaceous earth, concentrate the filtrate to give compound (43_Int-4). MS (ES): 267.80 m / z [M+H]+ LCMS purity: 96.46%, 1 1H NMR (400 MHz, DMSO-d6) δ 1.394 (s, 9H), 4.195 (d, J = 5.6 Hz, 2H), 7.244 (d, J = 8 Hz, 2H), 7.254 - 7.233 (m, 3H), 7.639 - 7.580 (m, 3H), 8.415 (s, 1H).

[0231] Step - 4. Synthesis of 4-(aminomethyl)-2-fluorobenzamidine dihydrochloride (43_Int-5).

[0232] Prepare a stirred solution of tert-butyl (4-formamido-3-fluorobenzyl)carbamate (43_Int-4) (4.8 g, 16.83 mmol, 1.0 eq) and water (52 mL, 11 V) and add concentrated HCl (16 mL, 3.3 V) at RT. Stir the RM at rt for 3 h. After completion of the reaction, concentrate the RM and triturate with methanol to give compound (43_Int-5). MS(ES): 168.13 m / z [M+H]+, 1 H NMR (400 MHz, DMSO-d6) δ 4.141 (d, J = 4 Hz, 2H), 7.658 - 7.155 (m, 3H), 7.737 - 7.686 (m, 2H), 8.772 (s, 1H), 9.564 - 9.440 (m, 2H).

[0233] Example 5A: Preparation of 4-(aminomethyl)-3-fluorobenzamidine dihydrochloride (45_Int-5)

[0234]

[0235] Step - 1. Synthesis of tert-butyl (4-cyano-2-fluorobenzyl)carbamate (45_Int-2).

[0236] Prepare a stirred solution of 4-(aminomethyl)-3-fluorobenzonitrile (45_Int-1) (1 g, 6.65 mmol, 1.0 eq) in 1,4-dioxane (12 mL, 12 V) and 2N NaOH (6 mL, 6 V) and add di-tert-butyl dicarbonate (1.59 g, 7.32 mmol, 1.1 eq) at 0 °C. Stir the reaction mixture at RT for 3 h. After completion of the reaction, quench the RM in water and extract with ethyl acetate. Dry the combined organic fractions over Na 2 SO 4 dry; concentrate to give compound (45_Int-2). 1 H NMR (400 MHz, DMSO-d6) δ 1.146 (s, 9H), 4.220 (d, J = 5.2 Hz, 2H), 7.442 (t, J = 7.5 Hz, 2H), 7.525 (s, 1H), 7.812 (t, J = 8 Hz, 2H).

[0237] Step - 2. Synthesis of tert-butyl (2-fluoro-4-(N-hydroxymethylamidine)benzyl)carbamate (45_Int-3).

[0238] Prepare a stirred solution of tert-butyl (4-cyano-2-fluorobenzyl)carbamate (45_Int-2) (5.3 g, 21.17 mmol, 1.0 eq) and methanol (53 mL, 10 V) and add hydroxylamine hydrochloride (2.5 g, 36.0 mmol, 1.7 eq) and DIPEA (4.64 g, 36.0 mmol, 1.7 eq) at RT. Stir the RM at 70 °C for 16 h. After completion of the reaction, evaporate the RM, quench the residue in water and extract by 10% methanol in DCM. Dry the combined organic fractions over Na 2 SO 4 dry; concentrate to give compound (45_Int-3). MS(ES): 284.21 m / z [M+H]+

[0239] Step - 3. Synthesis of tert-butyl (4-formamidin-2-fluorobenzyl)carbamate (45_Int-4).

[0240] Prepare a stirred solution of tert-butyl (2-fluoro-4-(N-hydroxymethylformamidine)benzyl)carbamate (45_Int-3) (2.4 g, 8.47 mmol, 1.0 eq) and methanol (24 mL, 10 V) and add ammonium formate (1.6 g, 25.41 mmol, 3.0 eq) and 10% Pd / C (0.16 g, 0.065% w / w) at RT. Stir the RM in an autoclave at 70 °C for 16 h under 20 kg H2 pressure. After completion of the reaction, filter the RM through a bed of diatomaceous earth, concentrate the filtrate to give compound (45_Int-4). MS(ES): 268.18 m / z [M+H]+

[0241] Step - 4. Synthesis of 4-(aminomethyl)-3-fluorobenzamidine dihydrochloride (45_Int-5).

[0242] Prepare a stirred solution of tert-butyl (4-formamidin-2-fluorobenzyl)carbamate (45_Int-4) (2.0 g, 7.48 mmol, 1.0 eq) and water (22 mL, 11 V) and add concentrated HCl (6.6 mL, 3.3 V) at RT. Stir the RM at RT for 3 h. After completion of the reaction, concentrate the RM and triturate with methanol to give compound (45_Int-5). MS(ES): 168.13 m / z [M+H]+, 1 1H NMR (400 MHz, DMSO-d6) δ 4.145 (s, 2H), 7.210 (s, 1H), 7.336 (s, 1H), 7.463 (s, 1H), 7.878 - 7.765 (m, 2H), 8.816 (s, 2H), 9.442 (s, 1H), 9.634 (s, 1H).

[0243] Example 6A: Preparation of Ethyl 1-(4-(3-cyanobenzyl)benzyl)-1H-pyrazole-4-carboxylate (28_Int-4)

[0244]

[0245] Step - 1. Synthesis of Ethyl 1-(4-chlorobenzyl)-1H-pyrazole-4-carboxylate (28_Int-2)

[0246] Prepare a stirred solution of ethyl 1H-pyrazole-4-carboxylate (1 g, 7.14 mmol, 1 eq) and 1-(bromomethyl)-4-chlorobenzene (28_int-1) (1.5 g, 7.85 mmol, 1.1 eq) in acetone (10 mL, 10V) and add Cs 2 CO 3 (5.53 g, 17.14 mmol, 2.4 eq). Heat the RM to 65 °C and stir for 4 h. After completion of the reaction, cool the RM to RT, quench in water and extract with ethyl acetate. Dry the combined organic fractions over Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (ethyl acetate in hexane 50%), to give the compound (28_Int-2). MS(ES): 265.30 m / z [M+], 266.30 [M+2]+, LCMS purity: 63%, 1 1H NMR (400 MHz, DMSO-d6) δ 400 MHz, DMSO-d6: δ 1.26 (t, J = 6.8 Hz, 3H), 4.28 - 4.210 (m, 2H), 5.38 (s, 2H), 7.30 (d, J = 8.40 Hz, 2H), 7.43 (d, J = 8.40 Hz, 2H), 7.88 (s, 1H), 8.49 (s, 1H).

[0247] Step - 2. Synthesis of Ethyl 1-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl)-1H-pyrazole-4-carboxylate (28_Int-3)

[0248] A stirred solution of ethyl 1-(4-chlorobenzyl)-1H-pyrazole-4-carboxylate (28_Int-2) (1.0 g, 3.78 mmol, 1.0 eq) in 1,4-dioxane (5 ml, 5V) was added with bis(pinacolato)diboron (1.14 g, 4.54 mmol, 1.2 eq), KOAc (1.11 g, 11.36 mmol, 3 eq), XPhosPdG2 (0.350 g, 0.37 mmol, 0.1 eq) and water (0.5 ml, 0.5V) at RT. The RM was heated to 100 °C and stirred for 16 h. After the reaction was completed, the RM was cooled to RT, quenched in water and extracted with ethyl acetate. The combined organic fractions were dried over Na 2 SO 4 dried; concentrated and then purified by flash column chromatography (ethyl acetate in hexane 20%), to give the compound (28_Int-3). MS (ES): 357.51 m / z [M+1]+, LCMS purity: 70%, 1 1H NMR (400 MHz, DMSO-d6) δ 1.26 (s, 12H), 1.26 (t, 3H), 4.228 - 4.210 (m, 2H), 5.38 (d, J = 12.80 Hz, 2H), 7.25 (d, J = 8.00 Hz, 2H), 7.65 (d, J = 8.00 Hz, 1H), 7.87 (s, 1H), 0.00 (s, 1H), 8.47 (s, 1H).

[0249] Step - 3. Synthesis of ethyl 1-(4-(3-cyanobenzyl)benzyl)-1H-pyrazole-4-carboxylate (28_Int-4)

[0250] A stirred solution of ethyl 1-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl)-1H-pyrazole-4-carboxylate (28_Int-3) (0.300 g, 0.84 mmol, 1.0 eq) in 1,4-dioxane (3 ml, 10V) was added with 3-(bromomethyl)benzonitrile (0.181 g, 0.92 mmol, 1.1 eq), K3PO4 (0.535 g, 3.51 mmol, 3 eq), Pd(dppf)Cl2, DCM (0.068 g, 0.11 mol, 0.1 eq) and water (1.5 ml, 5V) at RT. The RM was heated to 100 °C and stirred for 6 h. After the reaction was completed, the RM was cooled to RT, quenched in water and extracted with ethyl acetate. The combined organic fractions were dried over Na 2 SO 4Dried; concentrated and then purified by flash column chromatography (ethyl acetate in hexane 30%) to give the compound (28_Int-4). MS (ES): 346.39 m / z [M+1]+, LCMS purity: 100%, 1 H NMR (400 MHz, DMSO-d6) δ 1.22 (s, 3H), 3.98 (s, 2H), 4.20 (q, J = 6.8 Hz, 2H), 5.31 (s, 2H), 7.254 - 7.193 (m, 4H), 7.49 (t, J = 8.00 Hz, 1H), 7.56 (d, J = 8.00 Hz, 1H), 0.00 (d, J = 7.60 Hz, 1H), 7.72 (s, 1H), 7.84 (s, 1H), 8.434 (s, 1H).

[0251] Example 7A: Preparation of 4-(aminomethyl)-2,6-difluorobenzamidine dihydrochloride (44_Int-8)

[0252]

[0253] Step - 1. Synthesis of 2,6-difluoro-4-(hydroxymethyl)benzonitrile (44_Int-2)

[0254] Prepare a stirred solution of 2,6-difluoro-4-formylbenzonitrile (44_Int-1) (8 g, 0.047 mmol, 1.0 eq) in methanol (160 mL, 20 V) and add NaBH 4 (2.21 g, 0.047 mmol, 1 eq) at 0 °C. Stir the RM at RT for 1 h. After completion of the reaction, quench the RM in water and extract with DCM. Combine the organic fractions and dry over Na 2 SO 4 Dry; concentrate to give the compound (44_Int-2). 1H NMR (400 MHz, DMSO-d6) δ 4.60 (d, J = 8 Hz, 2H), 5.72 (t, J = 9.1 Hz, 1H), 7.34 (d, J = 8 Hz, 2H).

[0255] Step - 2. Synthesis of 2,6-difluoro-4-(chloromethyl)benzonitrile (44_Int-3)

[0256] Prepare a stirred solution of 2,6-difluoro-4-(hydroxymethyl)benzonitrile (44_Int-2) (7.5 g, 0.0443 mmol, 1.0 eq) in MTBE (75 mL, 10 V) and add PBr 3(14.37 g, 0.0532 mmol, 1.2 eq). Stir RM at RT for 2 h. After completion of the reaction, quench RM in cold water and extract with ethyl acetate. Combine the organic fractions and dry over Na 2 SO 4 ; concentrate to give compound (44_Int-3). 1H NMR (400 MHz, DMSO-d6) δ 4.721 (s, 2H), 7.564 (d, J = 8.8 Hz, 2H).

[0257] Step - 3. Synthesis of 4-(aminomethyl)-2,6-difluorobenzonitrile (44_Int-4)

[0258] Bubble ammonia gas through methanol at 0 °C for 30 min. Add a solution of 2,6-difluoro-4-(hydroxymethyl)benzonitrile (44_Int-3) (9.0 g, 0.06 mmol, 1.0 eq) in MeOH (40 mL, 10V) to RM and stir at 0 °C for 4 h. After completion of the reaction, concentrate RM to give compound (44_Int-4). MS (ES): 169 m / z [M+1]+, LCMS purity: 84%.

[0259] Step - 4. Synthesis of tert-butyl (4-cyano-3,5-difluorobenzyl)carbamate (44_Int-5)

[0260] Prepare a stirred solution of 4-(aminomethyl)-2,6-difluorobenzonitrile (44_Int-4) (10 g, 59.5238 mmol, 1.0 eq) in 1,4-dioxane (120 mL, 12V). Add 2N NaOH (60 ml, 6V) and Boc-anhydride (14.27 gm, 0.065 mmol, 1.1 eq) at RT and stir for 5 h. After completion of the reaction, quench RM in cold water and extract with ethyl acetate. Combine the organic fractions and dry over Na 2 SO 4 ; concentrate to give compound (44_Int-5) 1H NMR (400 MHz, DMSO-d6) δ 1.39 (s, 9H), 4.52 (s, 2H), 7.27 (d, J = 8.8 Hz, 2H).

[0261] Step - 5. Synthesis of tert-butyl (3,5-difluoro-4-(N-hydroxymethylamidinyl)benzyl)carbamate (44_Int-6)

[0262] Prepare a stirred solution of tert-butyl (4-cyano-3,5-difluorobenzyl)carbamate (44_Int-5) (6 g, 22.388 mmol, 1.0 eq) in MeOH (60 mL, 10V). Add hydroxylamine hydrochloride (2.6 g, 38.0596 mmol, 1.7 eq) and DIPEA (6.5 ml, 38.0596 mmol, 1.7 eq) at RT and stir at 70 °C for 16 h. After completion of the reaction, cool the RM to RT. Then quench the RM in water and extract with DCM. Dry the combined organic fractions over Na 2 SO 4 ; concentrate to afford the compound (44_Int-6). MS(ES): 302 m / z [M+1]+, LCMS purity: 65%.

[0263] Step - 6. Synthesis of tert-butyl (4-formamidin-3,5-difluorobenzyl)carbamate (44_Int-7)

[0264] Prepare a stirred solution of tert-butyl (3,5-difluoro-4-(N-hydroxymethylformamidine)benzyl)carbamate (44_Int-6) (6 g, 19.933 mmol, 1.0 eq) in MeOH (60 mL, 10V). Add ammonium chloride (5.38 g, 99.66 mmol, 5.0 eq) and iron (5.40 g, 99.66 mmol, 5.0 eq) at RT. Cool the RM to 0 °C and then add acetic acid (30 mL, 5V) dropwise. Then heat the RM to 70 °C and stir for 16 h. After completion of the reaction, cool the RM to RT. Concentrate the RM and quench in cold water and basify slowly to pH ~10. Separate the solid by filtration and extract with DCM. Dry the combined organic fractions over Na 2 SO 4 ; concentrate to afford the compound (44_Int-7). MS(ES): 286 m / z [M+1]+, LCMS purity: 65%.

[0265] Step - 7. Synthesis of 4-(aminomethyl)-2,6-difluorobenzamidine (44_Int-8)

[0266] Prepare a stirred solution of tert-butyl (4-formamidin-3,5-difluorobenzyl)carbamate (44_Int-7) (2.2 g, 25.473 mmol, 1.0 eq) in water (24 mL, 11V) and add concentrated HCl (7.26 mL, 3.3V) at RT and stir for 3 h. After completion of the reaction, concentrate the RM and triturate in methanol to afford the compound (44_Int-8). MS(ES): 231 m / z [M+1]+, LCMS purity: 67%.

[0267] Example 9A: Preparation of methyl 1-(4-(2-cyanopropan-2-yl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxylate (82_Int-3)

[0268]

[0269] Step - 1. Synthesis of methyl 5-(methoxymethyl)-1H-pyrazole-4-carboxylate (82_Int-2)

[0270] A solution of methyl 4-methoxy-3-oxobutanoate (82_Int-1) (15 g, 102 mmol, 1.0 eq) and dimethylformamide dimethyl acetal (12.23 g, 102 mmol, 1.0 eq) was prepared at 110 °C and stirred for 1 h. The RM was cooled to RT, and then ethanol (150 mL, 10V) and hydrazine hydrate (99%) (5.13 g, 102 mmol, 1.0 eq) were added. The RM was heated to 70 °C and stirred for 2 h. After the reaction was completed, the RM was evaporated and then purified by column chromatography (ethyl acetate in hexane 25 - 30%), to give the compound (82_Int-2). 1H NMR (400 MHz, DMSO-d6) δ 3.258 (s, 3H), 3.758 (s, 3H), 4.702 (s, 2H), 7.837 (s, 1H), 8.281 (s, 1H).

[0271] Step - 2. Synthesis of methyl 1-(4-(2-cyanopropan-2-yl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxylate (82_Int-3)

[0272] A stirred solution of methyl 5-(methoxymethyl)-1H-pyrazole-4-carboxylate (82_Int-2) (1.0 g, 5.87 mmol, 1.0 eq) in DMF (10 mL, 10V) was added with NaHMDS (1 M in THF) (5.8 mL, 5.87 mmol, 1.0 eq) at 0 °C and stirred for 30 min. Then 2-(4-(bromomethyl)phenyl)-2-methylpropanenitrile (7_Int-3) (1.39 g, 5.87 mmol, 1.0 eq) was added to the RM at 0 °C and stirred for an additional 16 h. After the reaction was completed, the RM was quenched in water and extracted with ethyl acetate. The combined organic fractions were dried over Na 2 SO 4Dry; concentrate and then purify by flash column chromatography (ethyl acetate in hexane 15%) (82_Int-3). Confirm by 2D NMR (ROE analysis). MS (ES): 328 m / z, LCMS purity: 99.11%, 1H 400 MHz, DMSO-d6: δ 1.654 (s, 6H), 3.258 (s, 3H), 3.758 (s, 3H), 4.797 (s, 2H), 5.394 (s, 2H), 7.258 (d, J = 8 Hz, 2H), 7.478 (d, J = 8 Hz, 2H), 7.903 (s, 1H).

[0273] Example 10A: Preparation of methyl 1-(4-(cyanomethyl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxylate (85_Int-3)

[0274]

[0275] Step-1. Synthesis of methyl 1-(4-(cyanomethyl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxylate (85_Int-3)

[0276] Prepare a stirred solution of methyl 5-(methoxymethyl)-1H-pyrazole-4-carboxylate (82_Int-2) (1.0 g, 5.87 mmol, 1.0 eq) in DMF (10 mL, 10V) and add NaHMDS (1 M in THF) (5.8 mL, 5.87 mmol, 1.0 eq) at 0 °C and stir for 30 min. Then add 2-(4-(bromomethyl)phenyl)acetonitrile (1.4 g, 7.05 mmol, 1.2 eq) to the RM at 0 °C and stir for an additional 16 h at RT. After completion of the reaction, quench the RM in water and extract with ethyl acetate. Combine the organic fractions and dry over Na 2 SO 4 Dry; concentrate and then purify by flash column chromatography (ethyl acetate in hexane 25 - 30%) (85_Int-3). Confirm by 2D NMR (ROE analysis). MS (ES): 300 m / z, 1H 400 MHz, DMSO-d6: δ 3.233 (s, 3H), 3.745 (s, 3H), 4.029 (s, 2H), 4.514 (s, 2H), 5.352 (s, 2H), 7.356 - 7.301 (m, 4H), 8.046 (s, 1H).

[0277] Example 11A: Preparation of ethyl 1-(4-(2-(dimethylamino)-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxylate (33_Int-3)

[0278]

[0279] Step - 1. Synthesis of 2-(4-(Bromomethyl)phenyl)-N-methylacetamide (33_Int-2)

[0280] Prepare a stirred solution of 2-(4-(bromomethyl)phenyl)acetic acid (33_Int-1) (1.0 g, 4.36 mmol, 1.0 eq) in toluene (10 mL, 10V). Add SOCl 2 (0.54 g, 4.53 mmol, 1.04 eq) and DMF (0.051 g, 0.69 mmol, 0.16 eq) at RT and stir at 85 °C for 3 h. Cool the RM to 0 °C, then add dimethylamine (2 M in THF) (2 mL, 2V). Allow the RM to reach RT and stir for 6 h. After completion of the reaction, quench the RM in saturated NaHCO3 solution and extract with ethyl acetate. Combine the organic fractions and dry over Na 2 SO 4 dry; concentrate to obtain the compound (33_Int-2). MS(ES): 256.6 m / z [M]+ 258.1. m / z [M+2]+, LCMS purity: 98.96%, 1H NMR (400 MHz, DMSO-d6) δ 2.826 (s, 3H), 2.997 (s, 3H), 3.694 (s, 2H), 4.706 (s, 3H), 7.210 (d, J = 8 Hz, 2H), 7.385 (d, J = 8 Hz, 2H).

[0281] Step - 2. Synthesis of Ethyl 1-(4-(2-(dimethylamino)-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxylate (33_Int-3).

[0282] Prepare a stirred solution of ethyl 1H-pyrazole-4-carboxylate (0.3 g, 2.14 mmol, 1.0 eq) in acetone (6 mL, 20V). Add 2-(4-(bromomethyl)phenyl)-N,N-dimethylacetamide (33_Int-2) (0.657 g, 2.56 mmol, 1.2 eq) and Cs 2 CO 3 (1.67 g, 5.13 mmol, 2.4 eq) at RT. Then heat the RM to 65 °C and stir for 16 h. After completion of the reaction, cool the RM to RT. Filter the RM to remove Cs 2 CO 3Wash with ethyl acetate. Discard the solid residue and concentrate the filtrate and then purify by column chromatography (ethyl acetate in hexane 25 - 30%) to give compound (33_Int-3) MS(ES): 316.3 m / z [M+1]+, LCMS purity: 100%, 1H NMR (400 MHz, DMSO-d6) δ 1.252 (t, J = 2.8 Hz, 3H), 2.802 (s, 3H), 2.974 (s, 3H), 3.396 - 3.295 (m, 2H), 4.188 (s, 2H), 5.324 (s, 2H), 7.189 (s, 4H), 7.852 (s, 1H), 8.466 (s, 1H).

[0283] Prepare the final compound

[0284] Example 1: Preparation of N-(4-carbamimidoylbenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxamide (7)

[0285]

[0286] Charge a 3000 mL 4N RBF attached to a mechanical stirrer and condenser with ethyl 1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxylate (7_Int5) (120 g, 403 mmol, 1.0 eq), 4-(aminomethyl)benzenecarboximidamide dihydrochloride (134.4 g, 605 mmol, 1.5 eq) and toluene (1200 mL, 10V) at RT. Add DIPEA (130.14 g, 1008 mmol, 2.5 eq) to the RM. Cool the RM to 0 °C to 5 °C and stir for 20 min. Add TMA (2 M in toluene) (605.3 mL, 121 mmol, 3.0 eq) dropwise to the RM. Heat the RM to 95 °C and stir for 16 h. After the reaction is complete, cool the reaction mixture to RT. Quench the RM slowly with DM water (151 mL) and evaporate to obtain a residue. Concentrate the filtrate under reduced pressure to obtain a crude product. Purify the crude product three times by column chromatography using silica gel of 60 - 120 mesh size. Elute the product with 10% MeOH in DCM to obtain a solid, dissolve in 4V methanol and stir at 70 °C for 2 h. Add EtOAc at 60 °C until a turbid solution is observed. Slowly cool the solution to RT and stir for 16 h until a solid precipitate is obtained, which is collected by filtration. Evaporate the filtrate and purify by column chromatography to give compound (7). MS(ES): 473.87 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 95.07% 11H NMR (400 MHz, DMSO-d6) δ 1.682 (s, 6H), 4.470 (s, 2H), 5.362 (s, 2H), 7.326 (d, J = 8.4 Hz, 2H), 7.515 - 7.419 (m, 4H), 7.772 (d, J = 8.4 Hz, 2H), 7.939 (s, 1H), 8.316 (s, 1H), 8.865 - 8.835 (m, 1H), 9.123 (s, 2H), 9.303 (s, 2H).

[0287] Example 2: N-(4-Carbamimidoylbenzyl)-1-(4-(2-cyanoethyl)benzyl)-1H-pyrazole-4-carboxamide (3)

[0288]

[0289] Step - 1. Synthesis of Ethyl 1-(4-iodobenzyl)-1H-pyrazole-4-carboxylate (3_Int-2).

[0290] Prepare a stirred solution of 1H-pyrazole-4-carboxylate (0.5 g, 3.56 mmol, 1.0 eq) in DMF (5 mL, 10V) and add NaH (60% in mineral oil) (0.213 g, 5.35 mmol, 1.5 eq) at 0 °C. Stir the RM at 0 °C for 30 minutes. Add 1-(bromomethyl)-4-iodobenzene (1.05 g, 3.56 mmol, 1.0 eq) at 0 °C and allow the RM to reach RT and stir for 2 h. After the reaction is complete, quench the RM in water and extract with ethyl acetate. Combine the organic fractions and dry over Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (ethyl acetate in hexane 12 - 15%) to give the compound (3_Int-2) MS (ES): 357.1. m / z [M + H]+, LCMS purity: 100%, 1H 400 MHz, DMSO-d6: δ 1.25 (t, J = 7.20 Hz, 3H), 4.20 (d, J = 7.20 Hz, 2H), 5.33 (s, 2H), 7.066 (d, J = 7.60 Hz, 2H), 7.715 (d, J = 8.00 Hz, 2H), 7.87 (s, 1H), 8.47 (s, 1H).

[0291] Step - 2. Synthesis of Ethyl (E)-1-(4-(2-cyanoethenyl)benzyl)-1H-pyrazole-4-carboxylate (3_Int-3).

[0292] Prepare a stirred solution of ethyl 1-(4-iodobenzyl)-1H-pyrazole-4-carboxylate (3_Int-2) (0.55 g, 1.54 mmol, 1.0 eq) and 1,4-dioxane (11 mL, 20 V) and add acrylonitrile (0.08 g, 1.54 mmol, 1 eq) and TEA (0.390 g, 3.86 mmol, 2.5 eq) at RT. Purge N2 in the RM for 10 min at RT. Add palladium acetate (0.034 g, 0.15 momol, 0.1 eq) and JohnPhos (0.092 g, 0.30 momol, 0.2 eq) and heat the RM to 110 °C and stir for 3 h. After the reaction is complete, quench the RM in water and extract with ethyl acetate. Dry the combined organic fractions over Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (22 - 25% ethyl acetate in hexane) to give compound (3_Int-3) MS (ES): 282.30. m / z [M+H]+, LCMS purity: 99.41%, 1H 400 MHz, DMSO-d6: δ 1.29 (d, J = 5.6 Hz, 3H), 2.231 (d, J = 5.6 Hz, 2H), 5.441 (d, J = 7.2 Hz, 2H), 5.92 (d, J = 12 Hz, 1H), 7.444 - 7.328 (m, Hz, 3H), 7.799 (d, J = 5.6 Hz, 2H), 7.907 (s, Hz, 1H), 8.533 (s, 1H).

[0293] Step - 3. Synthesis of ethyl 1-(4-(2-cyanoethyl)benzyl)-1H-pyrazole-4-carboxylate (3_Int-4).

[0294] Prepare a stirred solution of (E)-1-(4-(2-cyanoethenyl)benzyl)-1H-pyrazole-4-carboxylate (3_Int-3) (0.32 g, 1.13 mmol, 1.0 eq) in THF (6 mL, 20 V) and add 5% Pd / C at RT. Stir the RM under H2 and RT for 8 h. Filter the RM through a bed of diatomaceous earth and wash with ethyl acetate. Concentrate the filtrate and then purify by column chromatography (20 - 22% EtOAc in hexane) to give compound (3_Int-4). MS (ES): 284.30. m / z [M+H]+, LCMS purity: 65.00%, 1H 400 MHz, DMSO-d6: δ 1.32 (t, J = 6.8 Hz, 3H), 2.81 (d, J = 6.00 Hz, 2H), 2.86 (d, J = 6.00 Hz, 2H), 4.22 (d, J = 6.80 Hz, 2H), 5.36 (s, 2H), 7.27 (d, J = 7.60 Hz, 2H), 7.66 (d, J = 8.80 Hz, 2H), 7.87 (s, 1H), 8.50 (s, 1H).

[0295] Step - 4. Synthesis of N-(4-formamidino-benzyl)-1-(4-(2-cyanoethyl)benzyl)-1H-pyrazole-4-carboxamide (3).

[0296] A stirred solution of ethyl 1-(4-(2-cyanoethyl)benzyl)-1H-pyrazole-4-carboxylate (3_Int-4) (0.2 g, 7.05 mmol, 1.0 eq) in toluene (2 mL, 10 V) was added 4-(aminomethyl)benzimidamide dihydrochloride (0.15 g, 10.58 mmol, 1.5 eq) and DIPEA (0.23 g, 17.64 mmol, 2.5 eq) at RT. After cooling to 0 °C, TMA (2.0 M in toluene) (1 mL, 2.11 mmol, 3.0 eq) was added and the RM was heated to 100 °C and stirred for 16 h. After the reaction was complete, the RM was quenched with 1 V of water and evaporated. The residue was washed with 20% methanol in dichloromethane and filtered. The filtrate was concentrated and purified by Prep HPLC ((A) 0.1% aqueous TFA (B) 100% MeCN) to give the compound (3). MS (ES): 387.29. m / z [M+H]+, LCMS purity: 100%, HPLC purity: 100% 1H 400 MHz, DMSO-d6: δ 2.80 (d, J = 6.00 Hz, 2H), 2.84 (d, J = 6.00 Hz, 2H), 4.48 (d, J = 5.60 Hz, 2H), 5.33 (s, 2H), 7.25 (q, J = 27.20 Hz, 3H), 7.49 (d, J = 8.00 Hz, 1H), 7.74 (d, J = 8.00 Hz, 1H), 0.00 (s, 1H), 8.27 (s, 1H), 8.78 (d, J = 6.00 Hz, 1H), 8.95 (s, 2H), 9.25 (s, 2H).

[0297] Example 3: N-(4-formamidinobenzyl)-1-(4-(2-cyanoethyl)benzyl)-1H-imidazole-4-carboxamide (4)

[0298]

[0299] Step - 1. Synthesis of ethyl 1-(4-iodobenzyl)-1H-imidazole-4-carboxylate (4_Int-2).

[0300] A stirred solution of ethyl 1H-imidazole-4-carboxylate (0.5 g, 3.56 mmol, 1.0 eq) in DMF (5 mL, 10 V) was added NaHMDS (1 M in THF) (3.5 mL, 3.56 mmol, 1.0 eq) at 0 °C and stirred for 30 min. 1-(Bromomethyl)-4-iodobenzene (0.105 g, 3.56 mmol, 1.0 eq) was added to the RM at 0 °C, brought to RT and stirred for 2 h. After the reaction was complete, the RM was quenched in water and extracted with ethyl acetate. The combined organic fractions were passed through Na 2 SO4 Dried; concentrated and then purified by flash column chromatography (ethyl acetate in hexane 35 - 38%), to afford the compound (4_Int-2). MS (ES): 357.20. m / z [M+H]+, LCMS purity: 99.05%, 1H 400 MHz, DMSO-d6: δ 1.24 (t, J = 7.2 Hz, 3H), 4.19 (q, J = 7.2 Hz, 2H), 5.20 (s, 2H), 7.12 (d, J = 8.00 Hz, 2H), 7.74 (d, J = 8.00 Hz, 2H), 7.88 (s, 1H), 7.94 (s, 1H).

[0301] Step - 2. Synthesis of ethyl (E)-1-(4-(2-cyanoethenyl)benzyl)-1H-pyrazole-4-carboxylate (4_Int-3).

[0302] Prepare a stirred solution of ethyl 1-(4-iodobenzyl)-1H-imidazole-4-carboxylate (4_Int-2) (0.7 g, 1.96 mmol, 1.0 eq) and 1,4-dioxane (14 mL, 20V) and add acrylonitrile (0.104 g, 1.96 mmol, 1 eq) and TEA (0.497 g, 4.91 mmol, 2.5 eq) at RT. Purge N2 in the RM for 10 min at RT. Add palladium acetate (0.044 g, 0.19 mmol, 0.1 eq) and JohnPhos (0.117 g, 0.39 momol, 0.2 eq) to the RM. Heat the RM to 110 °C and stir for 3 h. Quench the RM in water and extract with ethyl acetate. Combine the organic fractions over Na 2 SO 4 Dried; concentrated and then purified by flash column chromatography (ethyl acetate in hexane 22 - 25%), to afford the compound (3_Int-3). MS (ES): 282.28. m / z [M+H]+, LCMS purity: 89.04%, 1H 400 MHz, DMSO-d6: δ 1.23 (t, J = 7.2 Hz, 3H), 4.21 (q, J = 5.60 Hz, 2H), 5.22 (s, 2H), 6.49 (d, J = 16.80 Hz, 1H), 7.41 (t, J = 6.8 Hz, 3H), 7.66 (d, J = 6 Hz, 1H), 7.82 (d, J = 7.60 Hz, 2H), 7.99 (s, 1H).

[0303] Step - 3. Synthesis of ethyl 1-(4-(2-cyanoethyl)benzyl)-1H-imidazole-4-carboxylate (4_Int-4).

[0304] Prepare a stirred solution of ethyl (E)-1-(4-(2-cyanoethenyl)benzyl)-1H-imidazole-4-carboxylate (4_Int-3) (0.4 g, 1.42 mmol, 1.0 eq) in THF (5 mL, 12 V) and ethanol (5 mL, 12 V) and add 5% Pd / C at RT. Stir the RM under a hydrogen atmosphere and at RT for 16 h. After the reaction is complete, filter the RM through a bed of diatomaceous earth and wash with ethyl acetate. Concentrate the filtrate and then purify by column chromatography (38 - 40% EtOAc in hexane) to obtain the compound (3_Int-4). MS (ES): 284.22. m / z [M+H]+, LCMS purity: 69.16.

[0305] Step - 4. Prepare N-(4-formamidobenzyl)-1-(4-(2-cyanoethyl)benzyl)-1H-pyrazole-4-carboxamide (4) from ethyl 1-(4-(2-cyanoethyl)benzyl)-1H-imidazole-4-carboxylate (4_Int-4) and 4-(aminomethyl)benzimidamide dihydrochloride in a similar manner to that described in Example-2(4). MS (ES): 387.30. m / z [M+H]+, LCMS purity: 100%, HPLC purity: 100% 1H 400 MHz, DMSO-d6: δ 2.80 (d, J = 6.40 Hz, 2H), 2.85 (d, J = 6.00 Hz, 2H), 4.47 (d, J = 5.60 Hz, 2H), 5.23 (s, 2H), 7.30 (s, 3H), 7.46 (d, J = 28.00 Hz, 2H), 7.74 (t, J = 8 Hz, 3H), 8.00 (s, 1H), 8.74 (s, 1H), 8.95 (s, 2H), 9.24 (s, 2H).

[0306] Example 4: Prepare N-(4-formamido-3-fluorobenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxamide (43)

[0307]

[0308] Prepare N-(4-formamido-3-fluorobenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxamide (301) from ethyl 1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxylate (7_Int-5) and 4-(aminomethyl)-2-fluorobenzimidamide dihydrochloride (43_Int-5) in a similar manner to that described in Example-2(3). MS (ES): 419.20. m / z [M+H]+, LCMS purity: 100%, HPLC purity: 10% 11H NMR (400 MHz, DMSO-d6) δ 1.667 (s, 6H), 2.504 (s, 2H), 4.771 (d, J = 4.2 Hz, 2H), 5.368 (s, 2H), 7.343 - 7.324 (m, 4H), 7.511 (d, J = 8 Hz, 2H), 7.625 (t, J = 8 Hz, 1H), 7.919 (s, 1H), 8.295 (s, 1H), 8.795 (t, J = 6 Hz, 1H), 9.184 (s, 2H), 9.369 (s, 2H).

[0309] Example 5: N-(4-Carbamimidoylbenzyl)-1-(4-(cyanomethyl)phenethyl)-1H-pyrazole-4-carboxamide (12)

[0310]

[0311] Step - 1. Synthesis of ethyl 1-(4-methylphenethyl)-1H-pyrazole-4-carboxylate (12_Int-2).

[0312] Prepare a stirred solution of ethyl 1H-pyrazole-4-carboxylate (2 g, 14.28 mmol, 1.0 eq) in acetone (20 mL, 10V) and add 1-(2-bromoethyl)-4-methylbenzene (3.04 g, 17.14 mmol, 1.2 eq) and Cs 2 CO 3 (11.14 g, 34.28 mmol, 2.4 eq) at RT. Heat the RM to 60 - 65 °C, stir for 16 h, and cool to RT. Filter the RM, wash with EtOAc and purify by column chromatography (12 - 15% ethyl acetate in hexane) to obtain the compound (12_Int-2) MS (ES): 259.23. m / z [M + H]+.

[0313] Step - 2. Synthesis of ethyl 1-(4-(bromomethyl)phenethyl)-1H-pyrazole-4-carboxylate (12_Int-3).

[0314] Prepare a stirred solution of ethyl 1-(4-methylphenethyl)-1H-pyrazole-4-carboxylate 12_Int-2) (1.7 g, 6.58 mmol, 1.0 eq) in carbon tetrachloride (34 mL, 20V) and add benzoyl peroxide (0.159 g, 0.65 mmol, 0.1 eq) and N-bromosuccinimide (1.40 g, 7.89 mmol, 1.2 eq) at RT. Stir the RM at RT for 4 h, quench in water and extract with DCM. Combine the organic fractions and pass through Na 2 SO 4Dried; concentrated and then purified by flash column chromatography (ethyl acetate in hexane 10 - 12%) to afford the compound (12_Int-3). MS (ES): 337.4. m / z [M]+ 339.4 [M+2]+, LCMS purity: 75.91%, 1 1H NMR (400 MHz, DMSO-d6) δ 1.239 (t, J = 7.2 Hz, 3H), 3.111 (t, J = 7.2 Hz, 2H), 4.179 (q, J = 7.2 Hz, 2H), 4.385 (t, J = 7.2 Hz, 2H), 4.664 (s, 2H), 7.148 - 7.127 (m, 2H), 7.350 - 7.330 (m, 2H), 7.841 (s, 1H), 8.220 (s, 1H).

[0315] Step - 3. Synthesis of ethyl 1-(4-(cyanomethyl)phenethyl)-1H-pyrazole-4-carboxylate (12_Int-4).

[0316] A stirred solution of ethyl 1-(4-(bromomethyl)phenethyl)-1H-pyrazole-4-carboxylate (12_Int-3) (0.750 g, 2.22 mmol, 1.0 eq) in MeCN (6. mL, 20 V) was prepared at RT. Tetrabutylammonium cyanide (2.38 g, 8.89 mmol, 4.0 eq) was added and the RM was stirred at RT for 16 h. After completion of the reaction, the RM was quenched in water and extracted with ethyl acetate. The combined organic fractions were dried over Na 2 SO 4 Dried; concentrated and then purified by flash column chromatography (ethyl acetate in hexane 7 - 10%) to afford the compound (12_Int-4). MS (ES): 284.28 m / z [M+H]+, LCMS purity: 93.34%, 1 1H NMR (400 MHz, DMSO-d6) δ 1.239 (t, J = 7.2 Hz, 3H), 3.112 (t, J = 7.2 Hz, 2H), 3.975 (s, 2H), 4.188 (q, J = 7.2 Hz, 2H), 4.383 (t, J = 7.2 Hz, 2H), 7.164 (d, J = 8 Hz, 2H), 7.238 (d, J = 8 Hz, 2H), 7.839 (s, 1H), 8.216 (s, 1H).

[0317] Step - 4. Synthesis of N-(4-formimidoylbenzyl)-1-(4-(cyanomethyl)phenethyl)-1H-pyrazole-4-carboxamide (12)

[0318] The title compound was prepared from ethyl 1-(4-(cyanomethyl)phenethyl)-1H-pyrazole-4-carboxylate (12_Int-4) and 4-(aminomethyl)benzimidine dihydrochloride in a similar manner as described in Example 2. MS(ES): 387.37 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 95.02% 1 H NMR (400 MHz, DMSO-d6) δ 3.110 (t, J = 7.2 Hz, 2H), 3.979 (s, 2H), 4.367 (t, J = 7.2 Hz, 2H), 4.771 (d, J = 6 Hz, 2H), 7.182 (d, J = 8 Hz, 2H), 7.244 (d, J = 8 Hz, 2H), 7.485 (d, J = 8 Hz, 2H), 7.751 (d, J = 8 Hz, 2H), 7.900 (s, 1H), 8.086 (s, 1H), 8.720 (t, J = 6 Hz, 1H), 8.974 (s, 2H), 9.248 (s, 2H).

[0319] Example 6: Preparation of 2-(4-((4-((4-carbamimidoylbenzyl)carbamoyl)-1H-pyrazol-1-yl)methyl)phenyl)acetic acid (16)

[0320]

[0321] Step - 1. Synthesis of 2-(4-((4-(ethoxycarbonyl)-1H-pyrazol-1-yl)methyl)phenyl)acetic acid (16_Int-2).

[0322] Prepare a stirred solution of 1H-pyrazole-4-carboxylate (0.5 g, 3.56 mmol, 1.0 eq) in DMF (5 mL, 10V) and add NaH (60% in mineral oil) (0.214 g, 5.35 mmol, 1.5 eq) at 0 °C. Stir the RM at 0 °C for 30 minutes. Add 2-(4-(bromomethyl)phenyl)acetic acid (0.980 g, 4.28 mmol, 1.2 eq) to the RM at 0 °C and stir at RT for 16 h. After completion of the reaction, quench the RM in water and extract with ethyl acetate. Combine the organic fractions and dry over Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (5 - 7% methanol in DCM) to give the compound (16_Int-2) MS(ES): 289.3 m / z [M+H]+, LCMS purity: 47.20%, 11H NMR (400 MHz, DMSO-d6) δ 1.252 (t, J = 7.2 Hz, 3H), 3.542 (s, 2H), 4.201 (q, J = 7.2 Hz, 2H), 5.335 (s, 2H), 7.222 (s, 4H), 7.853 (s, 1H), 8.451 (s, 1H), 12.321 (s, 1H).

[0323] Step - 2. 2-(4-((4-((4-Carbamimidoylbenzyl)carbamoyl)-1H-pyrazol-1-yl)methyl)phenyl)acetic acid (16) was prepared from 2-(4-((4-(ethoxycarbonyl)-1H-pyrazol-1-yl)methyl)phenyl)acetic acid (16_Int-2) and 4-(aminomethyl)benzimidamide dihydrochloride in a similar manner as described in Example - 2(16). MS (ES): 392.50 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 99.29% 1 1H NMR (400 MHz, DMSO-d6) δ 1.841 (s, 2H), 3.503 (s, 2H), 4.785 (s, 2H), 7.073 - 7.051 (m, 4H), 7.297 (d, J = 8 Hz, 2H), 7.504 (d, J = 8 Hz, 2H), 7.794 (s, 1H), 8.065 (s, 1H).

[0324] Example 7: Preparation of 1-(4-(2-amino-2-oxoethyl)benzyl)-N-(4-carbamimidoylbenzyl)-1H-pyrazole-4-carboxamide (18)

[0325]

[0326] Step - 1. Synthesis of 2-(4-(bromomethyl)phenyl)acetamide (18_Int-2).

[0327] A stirred solution of 2-(4-(bromomethyl)phenyl)acetic acid (0.5 g, 2.18 mmol, 1.0 eq) in toluene (5 mL, 10V) was added with SOCl2 (0.270 g, 2.26 mmol, 1.04 eq) and DMF (0.025 g, 0.34 mmol, 0.16 eq) at RT. The RM was heated to 80 - 85 °C and maintained for 3 h, followed by ammonia purge at 0 °C for 3 h. The RM was filtered and the solid was washed with EtOAc to obtain the compound (18_Int-2). MS (ES): 228.30 m / z [M]+, 230.12 m / z [M+2]+.

[0328] Step - 2. Synthesis of 1-(4-(2-amino-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxylic acid ethyl ester (18_Int-3).

[0329] Prepare a stirred solution of 1H-pyrazole-4-carboxylate (18_Int-2) (0.5 g, 3.56 mmol, 1.0 eq) in DMF (5 mL, 10V) and add NaH (60% in mineral oil) (0.214 g, 5.35 mmol, 1.5 eq) at 0 °C. Stir RM at 0 °C for 30 minutes. Add 2-(4-(bromomethyl)phenyl)acetamide (0.976 g, 4.28 mmol, 1.2 eq) to RM at 0 °C and stir at RT for 16 h. After completion of the reaction, quench RM in water and extract with ethyl acetate. Combine the organic fractions and dry over Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (ethyl acetate in hexane 40 - 42%) to give compound (18_Int-3). MS (ES): 288.23. m / z [M+H]+, LCMS purity: 97.01%, 1 1H NMR (400 MHz, DMSO-d6) δ 1.244 (t, J = 7.2 Hz, 3H), 3.231 (s, 3H), 4.201 (q, J = 7.2 Hz, 2H), 4.210 (s, 2H), 5.320 (s, 2H), 7.210 (s, 4H), 7.917 - 7.848 (m, 2H), 8.436 (s, 1H).

[0330] Step - 3. Synthesis of 1-(4-(2-amino-2-oxoethyl)benzyl)-N-(4-formamidobenzyl)-1H-pyrazole-4-carboxamide (18)

[0331] Prepare the title compound from 1-(4-(2-amino-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxylate ethyl ester (18_Int-3) and 4-(aminomethyl)benzimidamide dihydrochloride in a similar manner as described in Example 2. MS (ES): 391.32. m / z [M+H]+, LCMS purity: 99.23%, HPLC purity: 98.73% 1 1H NMR (400 MHz, DMSO-d6) δ 3.321 (d, J = 3.2 Hz, 2H), 4.479 (d, J = 6 Hz, 2H), 5.313 (s, 2H), 6.489 (s, 1H), 7.247 - 7.193 (m, 4H), 7.505 - 7.449 (m, 3H), 7.748 (d, J = 8 Hz, 1H), 7.902 (s, 1H), 8.253 (s, 1H), 8.752 (t, J = 6 Hz, 1H), 8.967 (s, 2H), 9.244 (s, 2H).

[0332] Example 8: Preparation of N-(4-formamidobenzyl)-1-(4-(2-(methylamino)-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxamide (19)

[0333]

[0334] Step - 1. Synthesis of 2-(4-(bromomethyl)phenyl)-N-methylacetamide (19_Int-2).

[0335] Prepare a stirred solution of 2-(4-(bromomethyl)phenyl)acetic acid (1.0 g, 4.36 mmol, 1.0 eq) in toluene (5 mL, 10V) at RT. Add SOCl2 (0.54 g, 4.53 mmol, 1.04 eq) and DMF (0.051 g, 0.69 mmol, 0.16 eq) at RT and then heat to 80 - 85 °C and hold for 3 h. Cool the solution to 0 °C and add methylamine (2 mL) to the RM and stir at RT for 6 h. Filter the RM and wash the solid with EtOAc to obtain the compound (19_Int-2). MS(ES): 242.1. m / z [M]+ 244.1. m / z [M+2]+, LCMS purity: 96.6%, 1 1H NMR (400 MHz, DMSO-d6) δ 3.389 (s, 2H), 4.726 (s, 2H), 7.245 (d, J = 8 Hz, 2H), 7.344 (d, J = 8 Hz, 2H), 8.019 (s, 1H).

[0336] Step - 2. Synthesis of ethyl 1-(4-(2-(methylamino)-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxylate (19_Int-3).

[0337] Prepare a stirred solution of ethyl 1H-pyrazole-4-carboxylate (0.5 g, 3.56 mmol, 1.0 eq) in acetone (10 mL, 20V) and add 2-(4-(bromomethyl)phenyl)-N-methylacetamide (19_Int-2) (1.03 g, 4.28 mmol, 1.2 eq) and s 2 CO 3 (2.78 g, 8.56 mmol, 2.4 eq) at RT. Heat the RM to 60 - 65 °C and stir for 16 h. After the reaction is complete, cool the RM to RT. Filter the RM and wash with EtOAc. Discard the solid residue and concentrate the filtrate and purify by column chromatography (12 - 15% ethyl acetate in hexane) to obtain the compound (19_Int-3) MS(ES): 302.40. m / z [M+H]+, LCMS purity: 86.48%, 11H NMR (400 MHz, DMSO-d6) δ 1.658 (s, 6H), 4.556 (s, 2H), 5.366 (s, 2H), 7.331 (d, J = 6 Hz, 2H), 7.505 (d, J = 8 Hz, 4H), 7.983 - 7.934 (m, 2H), 8.314 (s, 1H), 8.595 (s, 1H), 8.813 (s, 1H).

[0338] Step - 3. N-(4-Carbamimidoylbenzyl)-1-(4-(2-(methylamino)-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxamide (19) was prepared from ethyl 1-(4-(2-(methylamino)-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxylate (19_Int-3) and 4-(aminomethyl)benzimidamide dihydrochloride in a similar manner as described in Example 2. MS (ES): 405.55. m / z [M+H]+, LCMS purity: 100%, HPLC purity: 100% 1 1H NMR (400 MHz, DMSO-d6) δ 2.545 (d, J = 4.8 Hz, 3H), 3.361 (s, 2H), 4.476 (d, J = 6 Hz, 2H), 5.306 (s, 2H), 7.134 - 7.186 (m, 4H), 7.492 (d, J = 8 Hz, 2H), 7.743 (d, J = 8 Hz, 2H), 7.916 - 7.896 (m, 2H), 8.246 (s, 1H), 8.748 (t, J = 6 Hz, 1H), 8.896 (s, 2H), 9.234 (s, 2H).

[0339] Example 9: Preparation of N-(4-Carbamimidoylbenzyl)-1-(4-((3-cyano-1H-pyrrol-1-yl)methyl)benzyl)-1H-pyrazole-4-carboxamide (26)

[0340]

[0341] N-(4-Carbamimidoylbenzyl)-1-(4-((3-cyano-1H-pyrrol-1-yl)methyl)benzyl)-1H-pyrazole-4-carboxamide (26) was prepared from ethyl 1-(4-((3-cyano-1H-pyrrol-1-yl)methyl)benzyl)-1H-pyrazole-4-carboxylate (6_Int-1) and 4-(aminomethyl)benzimidamide dihydrochloride in a similar manner as described in the Example. MS (ES): 43841. m / z [M+H]+, LCMS purity: 100%, HPLC purity: 100% 11H NMR (400 MHz, DMSO-d6) δ 4.482 (d, J = 6 Hz, 2H), 5.156 (s, 2H), 5.335 (s, 2H), 6.467 (s, 2H), 7.001 (s, 1H), 7.254 - 7.250 (m, 4H), 7.496 (d, J = 8 Hz, 2H), 7.711 (s, 1H), 7.754 (d, J = 8 Hz, 2H), 7.909 (s, 1H), 8.270 (s, 1H), 8.772 (t, J = 6 Hz, 1H), 9.012 (s, 2H), 9.258 (s, 2H).

[0342] Example 10: Preparation of N-(4-formamidino-benzyl)-1-(4-(1-cyanoethyl)benzyl)-1H-pyrazole-4-carboxamide (5)

[0343]

[0344] Step - 1. Synthesis of ethyl 1-(4-(1-cyanoethyl)benzyl)-1H-pyrazole-4-carboxylate (5_Int-1).

[0345] Prepare a stirred solution of ethyl 1-(4-(cyanomethyl)benzyl)-1H-pyrazole-4-carboxylate (1_int-4) (0.3 g, 1.11 mmol, 1.0 eq) in THF (6 mL, 20 V) and add LiHMDS (1 M in hexanes) (1.23 mL, 1.22 mmol, 1.1 eq) at -70 °C. Stir RM at -70 °C for 30 min. Add methyl iodide (0.2 g, 1.44 mmol, 1.3 eq) to RM at -70 °C. Warm RM to RT and stir for 16 h. After completion of the reaction, quench RM in water and extract with ethyl acetate. Dry the combined organic fractions over Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (ethyl acetate in hexanes 12 - 15%) to give the compound (5_Int-1). MS (ES): 284.28 m / z [M+H]+, LCMS purity: 46.39%.

[0346] Step - 2. Prepare N-(4-formamidino-benzyl)-1-(4-(1-cyanoethyl)benzyl)-1H-pyrazole-4-carboxamide (45) from ethyl 1-(4-(1-cyanoethyl)benzyl)-1H-pyrazole-4-carboxylate (5-Int-1) in a similar manner as described in Example - 2. MS (ES): 387.33. m / z [M+H]+, LCMS purity: 94.00%.

[0347] Example 11: Preparation of N-(4-formamidin-2-fluorobenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxamide (45)

[0348]

[0349] N-(4-Formamidin-2-fluorobenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxamide (45) was prepared in a similar manner to that described in Example 2 from ethyl 1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxylate (4-Int-5) and 4-(aminomethyl)-3-fluorobenzamidine dihydrochloride (45_Int-5). MS(ES): 419.4. m / z [M+H]+, LCMS purity: 100%, HPLC purity: 100% 1 H NMR(400MHz, DMSO-d6) δ1.664(s, 6H), 4.499(d, J = 6.4Hz, 2H), 5.366(s, 2H), 7.331(d, J = 8Hz, 2H), 7.554 - 7.500(m, 3H), 7.608(d, J = 8Hz, 1H), 7.771(d, J = 8Hz, 1H), 7.925(s, 1H), 8.303(s, 1H), 8.776(t.J = 8.8Hz, 1H), 9.115(s, 2H), 9.334(s, 2H), 19 F NMR(400MHz, DMSO-d6) δ -73 - 659(1F), -116.871(0.35F).

[0350] Example 12: Preparation of N-(4-formamidinobenzyl)-1-(4-(cyanomethyl)benzyl)-1H-pyrazole-4-carboxamide (1)

[0351]

[0352] Under RT, charge a 3000 mL 4N RBF attached to a mechanical stirrer and condenser with ethyl 1-(4-(cyanomethyl)benzyl)-1H-pyrazole-4-carboxylate (1_Int4) (140 g, 517 mmol, 1.0 eq), 4-(aminomethyl)benzimidine dihydrochloride (127 g, 571 mmol, 1.1 eq), and toluene (1400 mL, 10V). Add DIPEA (266.5 mL, 1559 mmol, 3.0 eq) to the RM. Cool the RM to 0 °C to 5 °C and stir for 20 min. Add TMA (2 M in toluene) (649.8 mL, 1299 mmol, 2.5 eq) dropwise to the RM. Heat the RM to 95 °C and stir for 16 h. Cool the RM to RT, quench slowly with DM water (151 mL) and evaporate to obtain a residue. Wash the solid residue with methanol in 50% DCM (3 x 1000 mL) and discard the filtrate. Wash the solid residue again with MeOH in 20% DCM (2 x 2000 mL), then discard. Concentrate the filtrate and purify by flash column chromatography (methanol in 12 - 15% DCM) to obtain the compound (1) MS (ES): 373.35 m / z [M + H]+, LCMS purity: 96.80%, HPLC purity: 95.11% 1 H NMR (400 MHz, DMSO-d6) δ 3.805 (s, 2H), 4.455 (s, 2H), 5.314 (s, 2H), 7.304 - 7.289 (m, 4H), 7.469 (d, J = 8 Hz 2H), 7.710 (d, J = 8 Hz, 2H), 7.897 (s, 1H), 8.246 (s, 1H).

[0353] Example 13: Preparation of N-(4-formamidobenzyl)-1-(4-(hydroxymethyl)benzyl)-1H-pyrazole-4-carboxamide (2)

[0354]

[0355] Step - 1. Synthesis of ethyl 1-(4-(hydroxymethyl)benzyl)-1H-pyrazole-4-carboxylate (2_Int-2)

[0356] Prepare a stirred solution of ethyl 1H-pyrazole-4-carboxylate (2_int-1) (0.5 g, 3.56 mmol, 1.0 eq) in acetone (5 mL, 10V) and add (4-(bromomethyl)phenyl)methanol (0.86 g, 4.28 mmol, 1.2 eq) and Cs 2 CO 3(3.25 g, 9.98 mmol, 2.8 eq). Heat RM to 60 °C and stir for 3 h. After completion of the reaction, quench RM in water and extract with ethyl acetate. Combine the organic fractions, dry over Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (12 - 15% ethyl acetate in hexane) to give the compound (2_Int-2). MS (ES): 261.30 m / z [M+H]+, LCMS purity: 97.45%, 1 1H NMR (400 MHz, DMSO-d6) δ 1.255 (s, 3H), 4.203 (d, J = 6.4 Hz, 2H), 4.469 (s, 2H), 5.187 (s, 1H), 5.340 (s, 2H), 7.269 (m, 4H), 7.860 (s, 1H), 8.444 (s, 1H).

[0357] Step - 2. Prepare N-(4-formamidobenzyl)-1-(4-(hydroxymethyl)benzyl)-1H-pyrazole-4-carboxamide (2) from ethyl 1-(4-(hydroxymethyl)benzyl)-1H-pyrazole-4-carboxylate (2_Int-2) and 4-(aminomethyl)benzimidamide dihydrochloride in a similar manner as described in Example 2. MS (ES): 364.28 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 95.74% 1 1H NMR (400 MHz, DMSO-d6) δ 4.469 (s, 1H), 5.199 (s, 1H), 5.321 (s, 2H), 7.282 - 7.223 (m, 4H), 7.485 (d, J = 6.4 Hz, 2H), 7.741 (d, J = 6.4 Hz, 2H), 7.904 (s, 1H), 8.251 (s, 1H), 8.776 (s, 1H), 8.995 (s, 2H), 9.250 (s, 2H), 19 19F NMR (400 MHz, DMSO-d6) δ -73.575 (1F).

[0358] Example 14: Preparation of N-(4-formamidobenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-1H-imidazole-4-carboxamide (8)

[0359]

[0360] Step - 1. Synthesis of ethyl 1-(4-(2-cyanopropan-2-yl)benzyl)-1H-imidazole-4-carboxylate (8_Int-1)

[0361] Prepare a stirred solution of ethyl 1H-imidazole-4-carboxylate (0.45 g, 3.21 mmol, 1.0 eq) in DMF (4.5 mL, 10V) and add LiHMDS (1 M in THF) (3.2 mL, 3.21 mmol, 1.0 eq) at -78 °C and stir for 30 minutes. Add 2-(4-(bromomethyl)phenyl)-2-methylpropanenitrile (7_int-3) (0.91 g, 3.85 mmol, 1.2 eq) to the RM at -78 °C. Cool the RM to RT and stir at RT. After the reaction is complete, quench the RM in water and extract with ethyl acetate. Dry the combined organic fractions over Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (methanol in DCM at 1.2 - 1.5%), to give the compound (8_Int-1). MS (ES): 297.85 m / z [M+H]+, LCMS purity: 86.41%, 1 1H NMR (400 MHz, DMSO-d6) δ 1.246 (t, J = 5.2 Hz, 3H), 1.649 (s, 6H), 4.220 - 4.185 (m, 2H), 5.256 (s, 2H), 7.381 (d, J = 8 Hz, 2H), 7.524 (d, J = 8 Hz, 2H), 7.895 (d, J = 2 Hz, 1H), 7.952 (d, J = 2 Hz, 1H).

[0362] Step - 2. Prepare N-(4-formamidobenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-1H-imidazole-4-carboxamide (8) from 1-(4-(2-cyanopropan-2-yl)benzyl)-1H-imidazole-4-carboxylate (8_Int-1) and 4-(aminomethyl)benzimidamide dihydrochloride in a similar manner as described in Example 2. MS (ES): 401.25 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 99.63% 1 1H NMR (400 MHz, DMSO-d6) δ 1.662 (s, 6H), 4.772 (d, J = 6 Hz, 2H), 5.257 (s, 2H), 7.395 (d, J = 8 Hz, 2H), 7.536 - 7.473 (m, 4H), 7.759 - 7.717 (m, 2H), 8.717 (s, 1H), 8.873 (s, 2H), 9.231 (s, 2H).

[0363] Example 15: Preparation of N-(4-formamidobenzyl)-1-(4-(cyanomethyl)benzyl)-1H-imidazole-4-carboxamide (9)

[0364]

[0365] Step - 1. Synthesis of 2-(4-(bromomethyl)phenyl)acetonitrile (9_Int - 2)

[0366] Prepare a stirred solution of 2-(p - tolyl)acetonitrile (2 g, 15.24 mmol, 1.0 eq) and carbon tetrachloride (40 mL, 20 V) at RT. Add AIBN (0.12 g, 0.76 mmol, 0.1 eq) and N - bromosuccinimide (2.24 g, 1.67 mmol, 1.1 eq) at RT and then heat RM to 90 °C and stir for 16 h. After the reaction is complete, quench RM in water and extract with DCM.

[0367] Dry the combined organic fractions over Na 2 SO 4 dry, concentrate and then purify by flash column chromatography (7 - 10% ethyl acetate in hexane) to afford the compound (9_Int - 2). 1 1H NMR (400 MHz, DMSO - d6) δ 4.092 (s, 2H), 4.705 (s, 2H), 7.335 (d, J = 8 Hz, 2H), 7.469 (d, J = 8 Hz, 2H).

[0368] Step - 2. Synthesis of ethyl 1-(4-(cyanomethyl)benzyl)-1H - imidazole - 4 - carboxylate (9_Int - 3)

[0369] Prepare a stirred solution of ethyl 1H - imidazole - 4 - carboxylate (1.13 g, 8.09 mmol, 1.0 eq) in DMF (11.3 mL, 10 V) and add LiHMDS (1 M in THF) (8.9 mL, 8.86 mmol, 1.1 eq) at - 78 °C and stir for 30 minutes. Add 2-(4-(bromomethyl)phenyl)acetonitrile (9_Int - 2) (1.7 g, 8.09 mmol, 1.0 eq) to RM at - 78 °C. Allow RM to reach RT and stir for 16 h. After the reaction is complete, quench RM in water and extract with ethyl acetate. Dry the combined organic fractions over Na 2 SO 4 dry, concentrate and then purify by flash column chromatography (1.2 - 2.0% methanol in DCM) to afford the compound (9_Int - 3). MS (ES): 270.23 m / z [M + H]+, LCMS purity: 77.67%, 11H NMR (400 MHz, DMSO-d6) δ 1.239 (t, J = 7.2 Hz, 3H), 4.020 (s, 2H), 4.186 (q, J = 7.2 Hz, 2H), 5.240 (s, 2H), 7.348 (s, 4H), 7.877 (s, 1H), 7.929 (s, 1H).

[0370] Step - 3. N-(4-Carbamimidoylbenzyl)-1-(4-(cyanomethyl)benzyl)-1H-imidazole-4-carboxamide (9) was prepared from ethyl 1-(4-(cyanomethyl)benzyl)-1H-imidazole-4-carboxylate (9_Int-3) and 4-(aminomethyl)benzimidine dihydrochloride in a similar manner as described in Example - 2. MS (ES): 373.29 m / z [M + H]+, LCMS purity: 98.06%, HPLC purity: 98.03% 1 1H NMR (400 MHz, DMSO-d6) δ 4.023 (s, 2H), 4.470 (s, 2H), 5.250 (s, 2H), 7.354 (s, 4H), 7.481 (d, J = 6.0 Hz, 2H), 7.755 - 7.718 (m, 2H), 7.974 (s, 1H), 8.739 (s, 1H), 8.929 (s, 2H), 9.237 (s, 2H).

[0371] Example 16: Preparation of N-(4-Carbamimidoylbenzyl)-3-(4-(cyanomethyl)benzyl)isoxazole-5-carboxamide (10)

[0372]

[0373] Step - 1. Synthesis of (E)-2-(p-tolyl)acetaldoxime (10_Int-2)

[0374] A stirred solution of 2-(p-tolyl)acetaldehyde (2.2 g, 16.39 mmol, 1.0 eq) in DCM (44 mL, 20 V) was prepared at RT. Hydroxylamine hydrochloride (2.3 g, 32.79 mmol, 2.0 eq) and TEA (7 g, 68.86 mmol, 4.2 eq) were added at 0 °C. Then the RM was brought to RT and stirred for 16 h. After the reaction was complete, the reaction mixture was diluted in DCM and washed with 1N HCl in saturated NaHCO3 and brine solutions. The combined organic fractions were dried over Na 2 SO 4 dried; concentrated to give the compound (10_Int-2). MS (ES): 150.10 m / z [M + H]+

[0375] Step - 2. Synthesis of (Z)-N-hydroxy-2-(p-tolyl)iminoacetyl chloride (10_Int-3)

[0376] Prepare a stirred solution of (E)-2-(p-tolyl)acetaldoxime (10_Int-2) (4 g, 26.84 mmol, 1.0 eq) in DMF (80 mL, 20 V) and add NCS solution (3.6 g, 26.84 mmol, 1.0 eq) at 0 °C. Heat the RM to 50 °C and stir for 4 h. After completion of the reaction, evaporate the reaction mixture. Quench the remaining residue in a Na2S2O3 solution and extract with ethyl acetate. Dry the combined organic fractions over Na 2 SO 4 dry; concentrate to obtain the compound (10_Int-3). MS (ES): 184.16 m / z [M+H]+, LCMS purity: 60.43%.

[0377] Step - 3. Synthesis of ethyl 3-(4-methylbenzyl)isoxazole-5-carboxylate (10_Int-4)

[0378] Prepare a stirred solution of (Z)-N-hydroxy-2-(p-tolyl)iminoacetyl chloride (10_Int-3) (4 g, 21.78 mmol, 1.0 eq) in diethyl ether (80 mL, 20 V), and successively add TEA (3.3 g, 32.67 mmol, 1.5 eq) and ethyl propiolate (2.14 g, 21.78 mmol, 1.0 eq) at 0 °C and stir for 4 h. After completion of the reaction, filter the RM. Concentrate the filtrate and then purify by flash column chromatography (ethyl acetate in hexane 1.2 - 2.0%) to obtain the compound (10_Int-4). MS (ES): 246.24 m / z [M+H]+, LCMS purity: 100%, 1 1H NMR (400 MHz, DMSO-d6) δ 1.419 - 1.384 (m, 3H), 2.386 (s, 3H), 4.089 (s, 2H), 4.439 - 4.403 (m, 2H), 7.211 (s, 4H), 7.410 (s, 1H).

[0379] Step - 4. Synthesis of ethyl 3-(4-(bromomethyl)benzyl)isoxazole-5-carboxylate (10_Int-5).

[0380] Prepare a stirred solution of ethyl 3-(4-methylbenzyl)isoxazole-5-carboxylate (10_Int-4) (1.8 g, 7.33 mmol, 1.0 eq) and 1,2-dichloroethane (54 mL, 30 V). Add benzoyl peroxide (0.18 g, 0.73 mmol, 0.1 eq) and N-bromosuccinimide (1.3 g, 1.3 mmol, 1.0 eq) at RT. Heat the RM to 90 °C and stir for 6 h. After the reaction is complete, quench the reaction mixture in a Na2S2O3 solution and extract with DCM. Wash the combined organic fractions with Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (ethyl acetate in hexane 10 - 12%) to give the compound (10_Int-5). MS (ES): 324.2 m / z [M]+ 326.2 [M+2]+, LCMS purity: 89.94%, 1 1H NMR (400 MHz, DMSO-d6) δ 1.286 (t, J = 7.2 Hz, 3H), 4.070 (s, 2H), 4.325 (q, J = 7.2 Hz, 2H), 4.685 (s, 2H), 7.155 (s, 1H), 7.290 (d, J = 8 Hz, 2H), 7.401 (d, J = 8 Hz, 2H).

[0381] Step - 5. Synthesis of ethyl 3-(4-(cyanomethyl)benzyl)isoxazole-5-carboxylate (10_Int-6).

[0382] Prepare a stirred solution of ethyl 3-(4-(bromomethyl)benzyl)isoxazole-5-carboxylate (10_Int-5) (0.3 g, 0.92 mmol, 1.0 eq) and acetone (6 mL, 20 V), and add tetrabutylammonium cyanide (1 g, 3.7 mmol, 4 eq) under a nitrogen atmosphere and stir at RT for 16 h. After the reaction is complete, quench the RM in water and extract with ethyl acetate. Wash the combined organic fractions with Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (ethyl acetate in hexane 17 - 20%) to give the compound (10_Int-6). MS (ES): 242.42 m / z [M-42]+, LCMS purity: 100%, 1 1H NMR (400 MHz, DMSO-d6) δ 1.287 (t, J = 7.2 Hz, 3H), 4.004 (s, 2H), 4.072 (s, 2H), 4.327 (q, J = 7.2 Hz, 2H), 7.133 (s, 1H), 3.343 - 7.291 (m, 4H).

[0383] Step - 6. Prepare N-(4-formamidobenzyl)-3-(4-(cyanomethyl)benzyl)isoxazole-5-carboxamide (10) from ethyl 3-(4-(cyanomethyl)benzyl)isoxazole-5-carboxylate (10_Int-6) and 4-(aminomethyl)benzimidine dihydrochloride in a similar manner as described in Example 2. MS(ES): 374.41 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 99.64% 1 1H NMR (400 MHz, DMSO-d6) δ 3.991 (s, 2H), 4.060 (s, 2H), 4.500 (s, 2H), 6.929 (s, 1H), 7.316 (s, 4H), 7.497 (d, J = 7.2 Hz, 2H), 7.739 (d, J = 7.2 Hz, 2H).

[0384] Example 17: Preparation of N-(4-formamidobenzyl)-1-((4'-cyano-[1,1'-biphenyl]-4-yl)methyl)-1H-pyrazole-4-carboxamide (13)

[0385]

[0386] Step - 1. Synthesize ethyl 3-(4-(bromomethyl)benzyl)isoxazole-5-carboxylate (13_Int-2).

[0387] Prepare a stirred solution of 4'-methyl-[1,1'-biphenyl]-4-carbonitrile (13_Int-1) (0.1 g, 0.51 mmol, 1.0 eq) and carbon tetrachloride (2 mL, 20 V). Then add benzoyl peroxide (0.013 g, 0.05 mmol, 0.1 eq) and N-bromosuccinimide (0.1 g, 0.56 mmol, 1.1 eq) at RT. Heat the RM to 90 °C and stir for 16 h. After the reaction is complete, quench the RM in Na 2 SO 3 solution and extract with DCM. Wash the combined organic fractions with Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (2 - 5% ethyl acetate in hexane) to give the compound (13_Int-2). 1 1H NMR (400 MHz, DMSO-d6) δ 4.776 (s, 2H), 7.585 (d, J = 8 Hz, 2H), 7.845 (d, J = 8 Hz, 2H), 7.947 (s, 4H).

[0388] Step - 2. Synthesize ethyl 1-((4'-cyano-[1,1'-biphenyl]-4-yl)methyl)-1H-pyrazole-4-carboxylate (13_Int-3)

[0389] Prepare a stirred solution of ethyl 1H-imidazole-4-carboxylate (0.25 g, 1.83 mmol, 1.0 eq) in acetone (10 mL, 20V). Add 4'-(bromomethyl)-[1,1'-biphenyl]-4-carbonitrile (0.5 g, 1.89 mmol, 1.0 eq) and Cs 2 CO 3 (1.43 g, 1.40 mmol, 2.4 eq) at RT. Heat the RM to 65 °C and stir for 16 h. After the reaction is complete, cool the RM to RT, quench in water and extract with ethyl acetate. Dry the combined organic fractions over Na 2 SO 4 ; concentrate and then purify by flash column chromatography (ethyl acetate in hexane 15 - 20%) to give the compound (13_Int-3). MS (ES): 332.23 m / z [M+H]+.

[0390] Step - 3. Prepare N-(4-formamidobenzyl)-1-((4'-cyano-[1,1'-biphenyl]-4-yl)methyl)-1H-pyrazole-4-carboxamide (13) from ethyl 1-((4'-cyano-[1,1'-biphenyl]-4-yl)methyl)-1H-pyrazole-4-carboxylate (13_Int-3) and 4-(aminomethyl)benzimidine dihydrochloride in a similar manner as described in Example - 2. MS (ES): 435.37 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 99.83% 1 1H NMR (400 MHz, DMSO-d6) δ 4.485 (d, J = 6 Hz, 2H), 5.424 (s, 2H), 7.340 (d, J = 8 Hz, 2H), 7.495 (d, J = 8 Hz, 2H), 7.758 - 7.739 (m, 4H), 7.858 (s, 1H), 7.879 (s, 1H), 7.940 - 7.920 (m, 3H), 8.329 (s, 1H), 8.794 (t, J = 6 Hz, 1H), 8.984 (s, 1H), 9.248 (s, 2H).

[0391] Example 18: Preparation of N-(4-formamidobenzyl)-1-(4-(3-cyanobenzyl)benzyl)-1H-pyrazole-4-carboxamide (28)

[0392]

[0393] Step - 1. Synthesize N-(4-formamidobenzyl)-1-(4-(3-cyanobenzyl)benzyl)-1H-pyrazole-4-carboxamide (28) from ethyl 1-(4-(3-cyanobenzyl)benzyl)-1H-pyrazole-4-carboxylate (28_Int-4) and 4-(aminomethyl)benzimidamide dihydrochloride in a similar manner as described in Example - 2. MS(ES): 449.34. m / z [M+1]+, LCMS purity: 99.33%, HPLC purity: 100% 1 H NMR(400MHz, DMSO-d6) δ 3.98(s, 2H), 4.47(d, J = 6.40Hz, 2H), 5.30(s, 2H), 7.23(q, J = 8.8Hz, 4H), 7.50 - 7.47(m, 2H), 7.56(d, J = Hz, 1H), 7.66(d, J = 7.60Hz, 1H), 0.00(d, J = 8.4Hz, 2H), 7.89(s, 1H), 8.25(s, 1H), 8.75(s, 2H), 8.90(s, 2H), 9.24(s, 2H).

[0394] Example 19: Preparation of N-(4-formamidobenzyl)-1-(4-(3-cyano-1H-pyrrol-1-yl)benzyl)-1H-pyrazole-4-carboxamide (25)

[0395]

[0396] Step - 1. Synthesize ethyl 1-(4-iodobenzyl)-1H-pyrazole-4-carboxylate (25_Int-2)

[0397] Prepare a stirred solution of ethyl 1H-pyrazole-4-carboxylate (1.3g, 9.21mmol, 1eq) and 1-(bromomethyl)-4-iodobenzene (25_int-1) (3g, 10.2mmol, 1.1eq) in acetone (13mL, 10V) and add Cs 2 CO 3 (7.2g, 0.20mmol, 2.4eq). Heat the RM to 65 °C and stir for 8h. After the reaction is complete, cool the RM to RT, quench in water and extract with ethyl acetate. Combine the organic fractions and dry over Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (ethyl acetate in hexane 30%) to obtain the compound (28_Int-2). MS(ES): 357.27 m / z [M+1]+, LCMS purity: 82%.

[0398] Step - 2. Synthesis of Ethyl 1-(4-(3-cyano-1H-pyrrol-1-yl)benzyl)-1H-pyrazole-4-carboxylate (25_Int-3)

[0399] Prepare Ethyl 1-(4-iodobenzyl)-1H-pyrazole-4-carboxylate (28_Int-2). A stirred solution of (0.800 g, 2.20 mmol, 1 eq) and 1H-pyrrole-3-carbonitrile (0.426 g, 8.95 mmol, 4 eq) in DMF (8 mL, 10V) was added with K2CO3 (0.530 g, 6.78 mmol, 3 eq) and CuI (0.426 g, 2.23 mmol, 1 eq) at RT. The RM was heated to 100 °C and stirred for 8 h. After the reaction was completed, the RM was cooled at RT, quenched in water and extracted with ethyl acetate. The combined organic fractions were dried over Na 2 SO 4 dried; concentrated and then purified by flash column chromatography (ethyl acetate in hexane 30%), to give the compound (25_Int-3). MS (ES): 319.04 m / z [M+1]+, LCMS purity: 70%.

[0400] Step - 3. Synthesize N-(4-formamidobenzyl)-1-(4-(3-cyano-1H-pyrrol-1-yl)benzyl)-1H-pyrazole-4-carboxamide (25) from Ethyl 1-(4-(3-cyano-1H-pyrrol-1-yl)benzyl)-1H-pyrazole-4-carboxylate (25_Int-3) and 4-(aminomethyl)benzimidamide dihydrochloride in a similar manner as described in Example - 2. MS (ES): 424.15 m / z [M+1]+, LCMS purity: 100%, HPLC purity: 100% 1 1H NMR (400 MHz, DMSO-δ4.49 (d, J = 5.20 Hz, 2H), 5.41 (s, 2H), 6.73 (s, 1H), 7.02 (s, 1H), 7.14 (s, 2H), 7.26 (s, 1H), 7.42 (d, J = 8.00 Hz, 1H), 0.00 (t, J = 10.8 Hz, 1H), 7.65 (d, J = 8.00 Hz, 1H), 7.76 (d, J = 7.60 Hz, 1H), 7.94 (s, 1H), 8.23 (s, 1H), 8.32 (s, 1H), 8.79 (s, 1H), 9.10 (s, 2H), 9.25 (s, 2H). 19 19F NMR 400 MHz, DMSO-d6: δ - 73.478 (1F).

[0401] Example 20: Preparation of N-(4-formamidobenzyl)-1-(4-(4-cyanobenzyl)benzyl)-1H-pyrazole-4-carboxamide (14)

[0402]

[0403] Step - 1. Synthesis of 4-(4-(hydroxymethyl)benzyl)benzonitrile (14_Int-2)

[0404] Prepare a stirred solution of (4-cyanophenyl)boronic acid (1.5 g, 7.46 mmol, 1.0 eq) (14_Int-1) in 1,4-dioxane (15 ml, 10V), and add (4-(bromomethyl)phenyl)methanol (1.1 g, 8.20 mmol, 1.1 eq), Cs 2 CO 3 (7.27 g, 22.38 mmol, 3 eq), PdCl2(dppf).DCM (3.04 g, 3.73 mol, 0.5 eq) and water at RT. Heat RM to 100 °C and stir for 8 h. After the reaction is complete, cool RM to RT, quench in water and extract with ethyl acetate. Wash the combined organic fractions with Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (ethyl acetate in hexane 20%), to give the compound (14_Int-2). 1 1H NMR (400 MHz, DMSO-d6) δ 4.02 (s, 2H), 4.44 (s, 2H), 5.10 (t, J = 5.56 Hz, 1H), 7.22 (d, J = 8 Hz, 4H), 7.44 (d, J = 8.40 Hz, 2H), 7.74 (d, J = 8.40 Hz, 2H).

[0405] Step - 2. Synthesis of 4-(4-(chloromethyl)benzyl)benzonitrile (14_Int-3)

[0406] Prepare a stirred solution of ethyl 1H-pyrazole-4-carboxylate (0.300 g, 2.14 mmol, 1 eq) in DCM (10 ml, 10V) and add DIPEA (0.115 g, 8.96 mmol, 2 eq) and MsCl (0.107 g, 9.41 mmol, 2.1 eq) at 0 °C. Stir RM at RT for 6 h. After the reaction is complete, quench RM in cold water and extract with ethyl acetate. Wash the combined organic fractions with Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (ethyl acetate in hexane 4%), to give the compound (14_Int-3). 11H NMR (400 MHz, DMSO-d6) δ 400 MHz, DMSO-d6: δ 4.05 (s, 2H), 4.72 (s, 2H), 7.22 (d, J = 8 Hz, 4H), 7.44 (d, J = 8.40 Hz, 2H), 7.74 (d, J = 8.40 Hz, 2H).

[0407] Step - 3. Synthesis of ethyl 1-(4-(4-cyanobenzyl)benzyl)-1H-pyrazole-4-carboxylate (25_Int-4)

[0408] Prepare a stirred solution of ethyl 1H-pyrazole-4-carboxylate (0.300 g, 2.14 mmol, 1 eq) and 4-(4-(chloromethyl)benzyl)benzonitrile (14_Int-3) (0.568 g, 2.35 mmol, 1.1 eq) in acetone (3 mL, 10V) and add Cs 2 CO 3 (1.67 g, 5.14 mmol, 2.4 eq). Heat the RM to 65 °C and stir for 5 h. After the reaction is complete, cool the RM to RT, quench in water and extract with ethyl acetate. Combine the organic fractions and dry over Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (ethyl acetate in hexane 16%) to give the compound (28_Int-2). MS (ES): 346.3 m / z [M+1]+, LCMS purity: 88%.

[0409] Step - 4. Synthesize N-(4-formamidobenzyl)-1-(4-(4-cyanobenzyl)benzyl)-1H-pyrazole-4-carboxamide (14) from ethyl 1-(4-(4-cyanobenzyl)benzyl)-1H-pyrazole-4-carboxylate (25_Int-4) and 4-(aminomethyl)benzimidine dihydrochloride in a similar manner as described in Example - 2. MS (ES): 448 m / z [M+1]+, LCMS purity: 100%, HPLC purity: 100% 1 1H NMR (400 MHz, DMSO-d6) δ 4.02 (s, 2H), 4.47 (d, J = 6.00 Hz, 2H), 5.30 (s, 2H), 7.22 (s, 4H), 7.42 (d, J = 8.00 Hz, 2H), 7.49 (d, J = 8.40 Hz, 2H), 7.74 (d, J = 8.4 Hz, 3H), 0.00 (s, 1H), 8.25 (s, 1H), 8.76 (s, 1H), 8.92 (s, 2H), 8.95 (s, 2H).

[0410] Example 21: Preparation of N-(4-formamidobenzyl)-1-(4-(2-methoxyethyl)benzyl)-1H-pyrazole-4-carboxamide (15)

[0411]

[0412] Step - 1. Synthesis of ethyl 1-(4-(2-hydroxyethyl)benzyl)-1H-pyrazole-4-carboxylate (15_Int-2)

[0413] Prepare a stirred solution of ethyl 1H-pyrazole-4-carboxylate (1 g, 7.14 mmol, 1.0 eq) (14_Int-1) in acetone (10 ml, 10V) and add 2-(4-(bromomethyl)phenyl)ethan-1-ol (1.6 g, 7.85 mmol, 1.1 eq) and Cs 2 CO 3 (5.58 g, 17.14 mmol, 2.4 eq) at RT. Heat the RM to 65 °C and stir for 6 h. After completion of the reaction, cool the RM to RT, quench in water and extract with ethyl acetate. Dry the combined organic fractions over Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (ethyl acetate in hexane 25%), to give the compound (14_Int-2). MS (ES): 274.2 m / z [M+1]+, LCMS purity: 97%, HPLC purity: % 1 1H NMR (400 MHz, DMSO-d6) δ δ 1.22 (t, J = 7.20 Hz, 3H), 2.69 (t, J = 6.80 Hz, 2H), 3.57 (t, J = 6.80 Hz, 2H), 4.21 (t, J = 7.20 Hz, 2H), 4.63 (t, J = 5.20 Hz, 1H), 5.31 (s, 2H), 7.19 (s, 4H), 0.00 (s, 1H), 8.43 (s, 1H).

[0414] Step - 2. Synthesis of ethyl 1-(4-(2-methoxyethyl)benzyl)-1H-pyrazole-4-carboxylate (15_Int-3)

[0415] A stirred solution of ethyl 1-(4-(2-hydroxyethyl)benzyl)-1H-pyrazole-4-carboxylate (15_Int-2) (0.900 g, 5.17 mmol, 1.0 eq) in DMF (10 ml, 10V) was added with NaH (2.85 g, 10.34 mmol, 2 eq) and stirred at 0 °C for 15 min. Then MeI (0.948 g, 6.724 mmol, 1.3 eq) was added at 0 °C and the RM was stirred for 3 h. After the reaction was completed, the RM was quenched in cold water and extracted with ethyl acetate. The combined organic fractions were dried over Na 2 SO 4 dried; concentrated and then purified by flash column chromatography (ethyl acetate in hexane 12%) to give the compound (14_Int-3). MS (ES): 289.26 m / z [M+1]+, 1 1H NMR (400 MHz, DMSO-d6) δ 400 MHz, DMSO-d6: δ 1.26 (t, J = 6.80 Hz, 3H), 2.78 (t, J = 6.80 Hz, 2H), 3.27 (s, 3H), 3.51 (t, J = 6.80 Hz, 2H), 4.20 (q, J = 6.80 Hz, 2H), 5.31 (s, 2H), 7.20 (s, 2H), 0.00 (s, 2H), 8.43 (s, 2H).

[0416] Step - 3. N-(4-Carbamimidoylbenzyl)-1-(4-(2-methoxyethyl)benzyl)-1H-pyrazole-4-carboxamide (15) was synthesized from ethyl 1-(4-(2-methoxyethyl)benzyl)-1H-pyrazole-4-carboxylate (15_Int-3) and 4-(aminomethyl)benzimidamide dihydrochloride in a similar manner as described in Example - 2. MS (ES): 392.7 m / z [M+1]+, LCMS purity: 100%, HPLC purity: 100% 1 1H NMR (400 MHz, DMSO-d6) δ 2.52 (t, J = 181.60 Hz, 2H), 3.21 (s, 3H), 3.50 (t, J = 6.8 Hz, 2H), 4.48 (d, J = 5.60 Hz, 2H), 5.30 (s, 2H), 7.20 (t, J = 8.40 Hz, 4H), 7.49 (d, J = 8.40 Hz, 2H), 0.00 (d, J = 8.00 Hz, 2H), 7.90 (s, 1H), 8.26 (s, 1H), 8.77 (s, 1H), 8.97 (s, 2H), 9.25 (s, 2H).

[0417] Example 22: Preparation of N-(4-Carbamimidoyl-3-fluorobenzyl)-1-(4-(3-cyanobenzyl)benzyl)-1H-pyrazole-4-carboxamide (49)

[0418]

[0419] Step 1: N-(4-Carbamimidoyl-3-fluorobenzyl)-1-(4-(3-cyanobenzyl)benzyl)-1H-pyrazole-4-carboxamide (49) was prepared in a similar manner to that described in Example-2 from ethyl 1-(4-(3-cyanobenzyl)benzyl)-1H-pyrazole-4-carboxylate (28_Int-4) and 4-(aminomethyl)-2-fluorobenzamidine dihydrochloride (43_Int-5). MS(ES): 467.14 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 100% 1 H NMR 400 MHz, DMSO-d6: δ 3.99 (s, 2H), 4.46 (d, J = 6.00 Hz, 2H), 5.34 (s, 2H), 7.34 - 7.20 (m, 6H), 7.48 (t, J = 7.60 Hz, 1H), 7.71 - 7.56 (m, 3H), 7.89 (s, 1H), 8.25 (s, 1H), 8.76 (t, J = 6 Hz, 1H), 9.18 (s, 2H), 9.36 (s, 2H), 19 F NMR 400 MHz, DMSO-d6: δ -73.492 (1F), -113.893 (0.35F).

[0420] Example 23: N-(4-Carbamimidoyl-2-fluorobenzyl)-1-(4-(3-cyanobenzyl)benzyl)-1H-pyrazole-4-carboxamide (50)

[0421]

[0422] N-(4-Carbamimidoyl-2-fluorobenzyl)-1-(4-(3-cyanobenzyl)benzyl)-1H-pyrazole-4-carboxamide (50) was prepared in a similar manner to that described in Example-2 from ethyl 1-(4-(3-cyanobenzyl)benzyl)-1H-pyrazole-4-carboxylate (28_Int-4) and 4-(aminomethyl)-3-fluorobenzamidine dihydrochloride (45_Int5). MS(ES): 466.52 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 100% 11H NMR 400 MHz, DMSO-d6: δ 3.99 (s, 2H), 3.32 (s, 1H), 4.49 (d, J = 5.60 Hz, 1H), 5.30 (s, 1H), 7.23 (q, J = 8.00 Hz, 5H), 7.71 - 7.48 (m, 8H), 7.90 (s, 1H), 8.25 (s, 1H), 8.73 (s, 1H), 9.11 (s, 1H), 9.32 (s, 1H), 19 19F NMR 400 MHz, DMSO-d6: δ -73.536 (1F), -117.182 (0.68F).

[0423] Example 24: Preparation of N-(4-formamidobenzyl)-1-(3-(2-methoxyethyl)benzyl)-1H-pyrazole-4-carboxamide (69)

[0424]

[0425] Step 1. Synthesis of 1-bromo-3-(2-methoxyethyl)benzene (69_Int-2)

[0426] Prepare a stirred solution of 2-(3-bromophenyl)ethan-1-ol (69_Int-1) (3 g, 14.92 mmol, 1.0 eq) in DMF (30 mL, 10V) and slowly add NaH (60% in mineral oil) (0.72 g, 17.91 mmol, 1.2 eq) at 0 °C and stir for 1 h. Add methyl iodide (2.54 g, 17.91 mmol, 1.2 eq) to the RM at 0 °C and then allow to reach RT and stir for 16 h. After completion of the reaction, quench the RM slowly with cold water and extract with ethyl acetate. Dry the combined organic fractions over Na 2 SO 4 dry; concentrate and then purify by flash column chromatography (ethyl acetate in hexane 4 - 5%), to give the compound (69_Int-2) MS (ES): 162.23 m / z [M - 53]+, LCMS purity: 99.25%, 1H NMR (400 MHz, DMSO-d6) δ 2.808 (t, J = 6.4 Hz, 2H), 3.235 (s, 3H), 3.533 (t, J = 6.4 Hz, 2H), 7.250 (d, J = 5.2 Hz, 2H), 7.383 - 7.410 (m, 1H), 7.458 (s, 1H).

[0427] Step 2. Synthesis of 3-(2-methoxyethyl)benzaldehyde (69_Int-3)

[0428] Prepare a stirred solution of 1-bromo-3-(2-methoxyethyl)benzene (69_Int-2) (1.5 g, 7.0 mmol, 1.0 eq) in diethyl ether (15 mL, 10 V) and add tetramethylethylenediamine (1.75 g, 1.5 mmol, 2.16 eq) at RT. Cool the RM to -75 °C and then add n-BuLi (2.5 M in hexanes) (5.6 mL, 14.0 mmol, 2.0 eq) dropwise under N 2 2. Under N2, add n-BuLi (2.5 M in hexanes) (5.6 mL, 14.0 mmol, 2.0 eq) dropwise. Stir the RM at -75 °C for 1 h. Then warm the RM to -20 °C and stir for an additional 20 min. Cool the RM to -75 °C again and then add anhydrous DMF (7.5 mL, 5 V) dropwise and stir for 16 h. After the reaction is complete, quench the RM slowly with 1 N HCl and extract with ethyl acetate. Combine the organic fractions and dry over Na 2 SO 4 4. Dry; concentrate and then purify by flash column chromatography (ethyl acetate in hexanes 10 - 12%) to give compound (69_Int-3) MS (ES): 132.8 m / z [M - 32]+, LCMS purity: 100%, 1H NMR (400 MHz, DMSO-d6) δ 2.902 (t, J = 6.4 Hz, 2H), 3.325 (s, 3H), 3.581 (t, J = 6.4 Hz, 2H), 7.518 (t, J = 7.6 Hz, 1H), 7.588 (d, J = 8.0 Hz, 1H), 7.742 - 7.767 (m, 2H), 9.983 (s, 1H).

[0429] Step 3. Synthesis of (3-(2-methoxyethyl)phenyl)methanol (69_Int-4)

[0430] Prepare a stirred solution of 3-(2-methoxyethyl)benzaldehyde (69_Int-3) (0.23 g, 1.40 mmol, 1.0 eq) in methanol (4.6 mL, 20 V) and add sodium borohydride (0.106 g, 2.8 mmol, 2.0 eq) slowly at RT and stir for 4 h. After the reaction is complete, evaporate the RM. Quench the residue with water, adjust to pH ~5 with 2 N HCl and extract with ethyl acetate. Combine the organic fractions and dry over Na 2 SO 4Dried; concentrated and then purified by flash column chromatography (ethyl acetate in hexane 12 - 15%), to give compound (69_Int-4) MS (ES): 149.1 m / z [M-18]+, LCMS purity: 97.53%, 1H NMR (400 MHz, DMSO-d6) δ 2.794 (t, J = 6.8 Hz, 2H), 3.240 (s, 3H), 3.527 (t, J = 6.8 Hz, 2H), 4.467 (d, J = 5.6 Hz, 2H), 5.150 (t, J = 6.0 Hz, 1H), 7.092 (d, J = 7.6 Hz, 1H), 7.151 (t, J = 7.6 Hz, 2H), 7.231 (t, J = 7.6 Hz, 1H).

[0431] Step 4. Synthesis of 1-(bromomethyl)-3-(2-methoxyethyl)benzene (69_Int-5)

[0432] Prepare a stirred solution of (3-(2-methoxyethyl)phenyl)methanol (69_Int-4) (0.2 g, 1.20 mmol, 1.0 eq) in DCM (4 mL, 20 V). Then add triphenylphosphine (0.34 g, 1.32 mmol, 1.1 eq) and carbon tetrabromide (0.44 g, 1.32 mmol, 1.1 eq) at RT and stir for 16 h. After completion of the reaction, quench the RM in water and extract with DCM. Combine the organic fractions and dry over Na 2 SO 4 Dried; concentrated and then purified by flash column chromatography (ethyl acetate in hexane 8 - 10%), to give compound (69_Int-5) MS (ES): 248.2 m / z [M+18]+, LCMS purity: 100%, 1H NMR (400 MHz, DMSO-d6) δ 2.816 (t, J = 6.8 Hz, 2H), 3.343 (s, 3H), 3.538 (t, J = 6.8 Hz, 2H), 4.681 (s, 2H), 7.187 (s, 1H), 7.259 - 7.309 (m, 2H), 7.332 - 7.387 (m, 1H).

[0433] Step 5. Synthesis of ethyl 1-(3-(2-methoxyethyl)benzyl)-1H-pyrazole-4-carboxylate (69_Int-6)

[0434] A stirred solution of 1-(bromomethyl)-3-(2-methoxyethyl)benzene (69_Int-5) (0.176 g, 0.76 mmol, 1.0 eq) in acetone (3.52 mL, 20 V). Then ethyl 1H-pyrazole-4-carboxylate (0.4 g, 0.76 mmol, 1.0 eq) and cesium carbonate (0.6 g, 1.84 mmol, 2.4 eq) were added at RT. The RM was brought to 65 °C and stirred for 16 h. After the reaction was complete, the RM was filtered and the filtrate was concentrated and then purified by flash column chromatography (20 - 25% ethyl acetate in hexane) to give the compound (69_Int-6) MS (ES): 289.4 m / z [M+H]+, LCMS purity: 100%, 1H NMR (400 MHz, DMSO-d6) δ 1.25 (t, J = 8.0 Hz, 3H), 2.77 (t, J = 8.0 Hz, 2H), 3.22 (s, 3H), 3.50 (d, J = 8.0 Hz, 2H), 4.12 (q, J = 8.0 Hz, 2H), 5.33 (s, 2H), 7.08 (d, J = 8.0 Hz, 1H), 7.15 - 7.18 (m, 2H), 7.25 (t, J = 8.0 Hz, 1H), 7.87 (s, 1H), 8.45 (s, 1H).

[0435] Step 6. Synthesis of N-(4-formamidinobenzyl)-1-(3-(2-methoxyethyl)benzyl)-1H-pyrazole-4-carboxamide (69)

[0436] A stirred solution of ethyl 1-(3-(2-methoxyethyl)benzyl)-1H-pyrazole-4-carboxylate (69_Int-6) (0.1 g, 0.34 mmol, 1.0 eq) in THF (1 mL, 10 V). 4-(Aminomethyl)benzimidamide dihydrochloride (0.09 g, 0.41 mmol, 1.2 eq) and DIPEA (0.18 g, 1.38 mmol, 4.0 eq) were added at RT. Subsequently, the RM was cooled to 0 °C and then TMA (2 M in toluene) (1.0 mL, 2.08 mmol, 6.0 eq) was added. The RM was brought to 85 °C and stirred for 16 h. After completion of the reaction, the RM was quenched with 1 V of water and evaporated. The residue was washed with 20% methanol in dichloromethane and filtered. The filtrate was concentrated and then purified by PREP HPLC ((A) 0.1% aqueous TFA, (B) 100% MeCN). Finally, the pure fractions were lyophilized to give the compound (69) MS (ES): 392.6 m / z [M+H]+, LCMS purity: 97.92%, HPLC purity: 100% 1H NMR (400 MHz, DMSO-d6) δ 2.790 (t, J = 6.8 Hz, 2H), 3.226 (s, 3H), 3.522 (d, J = 6.8 Hz, 2H), 4.492 (d, J = 6.4 Hz, 2H), 5.326 (s, 2H), 7.088 (d, J = 8.0 Hz, 1H), 7.077 - 7.131 (m, 2H), 7.181 (t, J = 4.4 Hz, 1H), 7.403 (d, J = 4.4 Hz, 2H), 7.761 (d, J = 8.4 Hz, 2H), 7.924 (s, 1H), 8.274 (s, 1H), 8.776 (t, J = 6.0 Hz, 1H), 9.043 (s, 2H), 9.256 (s, 2H).

[0437] Example 25: Preparation of N-(4-formamidin-3,5-difluorobenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxamide (44)

[0438]

[0439] Step - 1. Synthesis of N-(4-formamidin-3,5-difluorobenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxamide (44)

[0440] A stirred solution of ethyl 1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxylate (7_Int-5) (0.1 g, 0.3367 mmol, 1.0 eq) in toluene (2 mL, 10 V). 4-(Aminomethyl)-2,6-difluorobenzamidine dihydrochloride (44_Int-8) (0.095 g, 0.505 mmol, 1.5 eq) and DIPEA (0.6 mL, 0.8417 mmol, 2.5 eq) were added to the RM at RT. The RM was cooled to 0 °C and TMA (2.0 M in toluene, 0.6 mL, 1.0101 mmol, 3.0 eq) was added. Then the RM was heated to 100 °C and stirred for 16 h. After the reaction was complete, the RM was quenched with 1 V of water and evaporated. The residue was washed with 20% methanol in dichloromethane and filtered. The solid residue was discarded and the filtrate was concentrated and then purified by PREP HPLC ((A) 0.1% aqueous TFA (B) 100% MeCN). Finally, the pure fractions were lyophilized to give the compound (44). MS (ES): 437.21 m / z [M+1]+, LCMS purity: 100%, HPLC purity: 94.56% 1H 400 MHz, DMSO-d6: δ 1.664 (s, 6H), 4.462 (d, J = 6.40 Hz, 2H), 5.369 (s, 2H), 7.252 (d, J = 8.8 Hz, 2H), 7.331 (d, J = 8.0 Hz, 2H), 7.511 (d, J = 8.00 Hz, 2H), 7.921 (s, 1H), 8.310 (s, 1H), 8.840 (t, J = 6.4 Hz, 1H), 9.513 (s, 2H), 9.669 (s, 2H).

[0441] Example 26: N-(4-Carbamimidoylbenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-5-cyclopropyl-1H-pyrazole-4-carboxamide (83)

[0442]

[0443] Step-1. Synthesis of tert-butyl 2-(4-(2-cyanopropan-2-yl)benzyl)hydrazine-1-carboxylate (83_Int-1)

[0444] A stirred solution of 2-(4-(bromomethyl)phenyl)-2-methylpropanenitrile (7_Int-3) (0.5 g, 2.1 mmol, 1.0 eq) in dimethylacetamide (5 mL, 10 V) was added with DIPEA (0.54 g, 4.2 mmol, 2.0 eq) and tert-butyl hydrazinecarboxylate (0.55 g, 4.2 mmol, 2.0 eq) at RT and stirred at 70 °C for 16 h. After the reaction was completed, the RM was quenched in water and extracted with ethyl acetate. The combined organic fractions were washed with brine solution, dried over Na2SO4, concentrated and purified by flash column chromatography (ethyl acetate in hexane 10 - 12%) to give the compound (83_Int-1), 1H 400 MHz, DMSO-d6: δ 1.637 (s, 6H), 1.760 (s, 9H), 4.206 (s, 2H), 7.514 (s, 4H), 7.744 (d, br, J = 8.00 Hz, 1H), 7.886 - 8.004 (d, br, J = 47.2, 1H).

[0445] Step - 2. Synthesis of 2-(4-(hydrazinomethyl)phenyl)-2-methylpropanenitrile (83_Int-2)

[0446] A stirred solution of tert-butyl 2-(4-(2-cyanopropan-2-yl)benzyl)hydrazine-1-carboxylate (83_Int-1) (2.0 g, 6.9 mmol, 1.0 eq) in methanol (10 mL, 5 V) and water (22 mL, 11 V) was added with concentrated HCl (6.6 mL, 3.3 V) at RT. The RM was heated to 80 °C and stirred for 16 h. After the reaction was completed, the RM was evaporated and triturated in ethyl acetate to give the compound (83_Int-2). MS (ES): 191.4 m / z [M + H]+, LCMS purity: 83.16%, 1H 400 MHz, DMSO-d6: δ 1.693 (s, 6H), 4.076 (s, 2H), 7.096 (s, br, 2H), 7.517 (d, J = 12.8 Hz, 4H).

[0447] Step - 3. Synthesis of ethyl 1-(4-(2-cyanopropan-2-yl)benzyl)-5-cyclopropyl-1H-pyrazole-4-carboxylate (7_Int-3)

[0448] Prepare a solution of ethyl 3-cyclopropyl-3-oxopropionate (0.22 g, 1.41 mmol, 1.0 eq) and dimethylformamide dimethyl acetal (0.185 g, 1.55 mmol, 1.1 eq) and stir at 75 °C for 90 min. Cool the RM to RT, then add ethanol (4.4 mL, 20 V), TEA (0.57 g, 5.63 mmol, 4.0 eq) and 2-(4-(hydrazinomethyl)phenyl)-2-methylpropanenitrile (83_Int-2) (0.365 g, 1.41 mmol, 1.0 eq). Heat the RM to 80 °C and stir for 16 h. After the reaction is complete, quench the RM in water and extract with ethyl acetate. Wash the combined organic fractions with brine solution, dry over Na2SO4, concentrate and purify by flash column chromatography (ethyl acetate in hexane 12 - 15%) to give the compound (83_Int-3). MS(ES): 180.88 m / z [M+H]+, LCMS purity: 89%, 1H 400 MHz, DMSO-d6: δ 1.637(s, 6H), 4.174 - 4.225(m, 2H), 5.484(s, 2H), 7.222(d, J = 8.0 Hz, 2H), 7.505(d, J = 8.0 Hz, 2H), 7.830(s, 1H).

[0449] Step - 4. Synthesis of N-(4-formamidobenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-5-cyclopropyl-1H-pyrazole-4-carboxamide (83)

[0450] Prepare the final compound (83) from ethyl 1-(4-(2-cyanopropan-2-yl)benzyl)-5-cyclopropyl-1H-pyrazole-4-carboxylate (7_Int-3) and 4-(aminomethyl)benzimidamide dihydrochloride in a similar manner as described in (Example 24). MS(ES): 441.4 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 100% 1H 400 MHz, DMSO-d6: δ 0.734 - 0.757(m, 2H), 0.919 - 0.967(m, 2H), 1.658(s, 6H), 1.753 - 1.796(m, 1H), 4.472(d, J = 6.0 Hz, 2H), 5.438(s, 2H), 7.227(d, J = 8 Hz, 2H), 7.501(t, J = 8.8 Hz, 4H), 7.762(d, J = 8.0 Hz, 2H), 7.801(s, 1H), 8.534(t, J = 6.0 Hz, 1H), 8.977(s, 2H), 9.258(s, 2H).

[0451] Example 27: N-(4-Carbamimidoylbenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-5-propyl-1H-imidazole-4-carboxamide (84)

[0452]

[0453] Step - 1. Synthesis of Ethyl 1-(4-(2-cyanopropan-2-yl)benzyl)-5-propyl-1H-imidazole-4-carboxylate (84_Int-1)

[0454] Prepare a stirred solution of ethyl 5-propyl-1H-imidazole-4-carboxylate (0.45 g, 2.47 mmol, 1.0 eq) in DMF (4.5 mL, 10V) and add NaHMDS (1 M in THF, 2.5 mL, 2.47 mmol, 1.0 eq) at 0 °C and stir for 30 min. Add 2-(4-(Bromomethyl)phenyl)-2-methylpropanenitrile (7_Int-3) (0.5 g, 3.56 mmol, 1.0 eq) to the RM at 0 °C, allow to reach RT and stir for 16 h. After completion of the reaction, quench the RM in water and extract with ethyl acetate. Dry the combined organic fractions over Na2SO4; concentrate and then purify by flash column chromatography (20 - 25% ethyl acetate in hexane). (84_Int-1). Confirm by 2D NMR (ROE analysis). MS (ES): 339.9 m / z [M+H]+, LCMS purity: 84.72%, 1H 400 MHz, DMSO-d6: δ 0.807 (t, J = 7.2 Hz, 3H), 1.251 (q, J = 8.0 Hz, 3H), 1.297 (s, 2H), 1.654 (s, 6H), 2.734 (q, J = 6.4 Hz, 2H), 4.186 (q, J = 7.2 Hz, 2H), 5.264 (s, 2H), 7.194 (d, J = 8.0 Hz, 2H), 7.512 (d, J = 8.0 Hz, 2H), 7.788 (s, 1H).

[0455] Step - 2. Synthesis of N-(4-Carbamimidoylbenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-5-propyl-1H-imidazole-4-carboxamide (84)

[0456] The final compound (84) was prepared from ethyl 1-(4-(2-cyanopropan-2-yl)benzyl)-5-propyl-1H-imidazole-4-carboxylate (84_Int-1) and 4-(aminomethyl)benzimidamide dihydrochloride in a similar manner as described in (Example-24). MS(ES): 443.35 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 99.63% 1H 400 MHz, DMSO-d6: δ 0.782 (t, J = 7.2 Hz, 3H), 1.262 - 1302 (m, 2H), 1.656 (d, J = 6.00 Hz, 2H), 2.773 (t, J = 8 Hz, 2H), 4.473 (d, J = 6.4 Hz, 2H), 5.260 (s, 2H), 7.226 (d, J = 8.4 Hz, 2H), 7.487 - 7.526 (m, 4H), 7.745 (d, J = 8.4 Hz, 2H), 7.872 (s, 1H), 8.634 (t, J = 6.0 Hz, 1H), 9.069 (s, 2H), 9.248 (s, 2H).

[0457] Example 28: Preparation of N-(4-formamidobenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-5-methyl-1H-pyrazole-4-carboxamide (81)

[0458]

[0459] Step - 1. Synthesis of ethyl 5-methyl-1H-pyrazole-4-carboxylate (81_Int-2)

[0460] A solution of ethyl 3-oxobutanoate (81_Int-1) (0.5, 3.84 mmol, 1.0 eq) and dimethylformamide dimethylacetal (0.46 g, 3.84 mmol, 1.0 eq) was prepared and stirred at 110 °C for 1 h. The RM was cooled to RT, and then ethanol (5.0 mL, 10V) and hydrazine hydrate (99%) (0.2 g, 3.84 mmol, 1.0 eq) were added. The RM was heated to 70 °C and stirred for 2 h. After the reaction was completed, the reaction mixture was evaporated to obtain the compound (81_Int-2). MS(ES): 154.96 m / z [M+H]+, LCMS purity: 100%, 1H NMR (400 MHz, DMSO-d6) δ 1.241 (t, J = 8.00 Hz, 3H), 2.446 (s, 3H), 4.178 (q, J = 4.2 Hz, 2H), 7.794 (s, 1H), 13.102 (s, 1H).

[0461] Step - 2. Synthesis of ethyl 1-(4-(2-cyanopropan-2-yl)benzyl)-5-methyl-1H-pyrazole-4-carboxylate (81_Int-3)

[0462] Prepare a stirred solution of ethyl 5-methyl-1H-pyrazole-4-carboxylate (81_Int-2) (0.25 g, 1.62 mmol, 1.0 eq) in DMF (2.5 mL, 10 V) and add NaHMDS (1 M in THF) (1.62 mL, 1.62 mmol, 1.0 eq) at 0 °C and stir for 30 min. Add 2-(4-(bromomethyl)phenyl)-2-methylpropanenitrile (7_Int-3) (0.4 g, 1.62 mmol, 1.0 eq) to the RM at 0 °C. Allow the RM to reach RT and stir for 16 h. After completion of the reaction, quench the RM in water and extract with ethyl acetate. Dry the combined organic fractions over Na2SO4; concentrate and then purify by flash column chromatography (ethyl acetate in hexane 20 - 25%), to give the compound (81_Int-3). MS (ES): 312.2 m / z [M+H]+, LCMS purity: 100%, 1H 400 MHz, DMSO-d6: δ 1.239 (t, J = 8.00 Hz, 3H), 1.656 (s, 6H), 2.299 (s, 3H), 4.171 (t, J = 8.00 Hz, 2H), 5.273 (s, 2H), 7.190 (d, J = 8.00 Hz, 2H), 7.322 (d, J = 8.00 Hz, 2H), 7.829 (s, 1H).

[0463] Step - 3. Synthesis of N-(4-carbamimidoylbenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-5-propyl-1H-imidazole-4-carboxamide (81)

[0464] Prepare the final compound (81) from ethyl 1-(4-(2-cyanopropan-2-yl)benzyl)-5-propyl-1H-imidazole-4-carboxylate (81_Int-3) and 4-(aminomethyl)benzimidamide dihydrochloride in a similar manner as described in (Example 25). MS (ES): 415.35 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 99.47% 1H 400 MHz, DMSO-d6: δ 1.655 (s, 6H), 2.457 (s, 3H), 4.777 (d, J = 6.0 Hz, 2H), 5.341 (s, 2H), 7.191 (d, J = 8 Hz, 2H), 7.515 - 7.477 (m, 4H), 7.750 (d, J = 8 Hz, 2H), 7.985 (s, 1H), 8.676 (t, J = 6 Hz, 1H), 8.886 (s, 2H), 9.247 (s, 2H).

[0465] Example 29: N-(4-Carbamimidoyl-3-fluoro-5-methoxybenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxamide (48)

[0466]

[0467] Step - 1. Synthesis of tert-Butyl (4-cyano-3-fluoro-5-methoxybenzyl)carbamate (48_Int-1)

[0468] Prepare a stirred solution of tert-Butyl (4-cyano-3,5-difluorobenzyl)carbamate (44_Int-7) (5 g, 18.36 mmol, 1.0 eq) in THF (50 mL, 10V) and MeOH (50 mL, 10V). Cool the RM to 0 °C and slowly add sodium methoxide (4.02 g, 74.6 mmol, 4.0 eq). Allow the RM to reach RT and stir for 5 h. After completion of the reaction, quench the RM in cold water and extract with ethyl acetate. Dry the combined organic fractions over Na2SO4; concentrate to afford compound (48_Int-1) 1H 400 MHz, DMSO-d6: δ 1.631 (s, 9H), 3.924 (s, 3H), 4.169 (d, J = 6.0 Hz, 2H), 6.871 (d, J = 10 Hz, 1H), 6.982 (s, 1H), 7.544 (t, J = 6.0 Hz, 1H).

[0469] Step - 2. Synthesis of tert-Butyl (3-fluoro-4-(N-hydroxycarbamimidoyl)-5-methoxybenzyl)carbamate (48_Int-2)

[0470] Prepare a stirred solution of tert-Butyl (4-cyano-3-fluoro-5-methoxybenzyl)carbamate (48_Int-1) (4.7 g, 16.78 mmol, 1.0 eq) in MeOH (50 mL, 10V). Subsequently, add hydroxylamine hydrochloride (1.96 gm, 28.536 mmol, 1.7 eq) and DIPEA (5.0 ml, 28.535 mmol, 1.7 eq) at RT. Heat the RM to 70 °C and stir for 16 h. After completion of the reaction, cool the RM to RT, quench in water and extract with DCM. Dry the combined organic fractions over Na2SO4; concentrate to afford compound (48_Int-2). MS (ES): 314.29 m / z [M+1]+, LCMS purity: 80%.

[0471] Step - 3. Synthesis of tert-Butyl (4-carbamimidoyl-3-fluoro-5-methoxybenzyl)carbamate (48_Int-3)

[0472] Prepare a stirred solution of tert-butyl (3-fluoro-4-(N-hydroxyformamidinyl)-5-methoxybenzyl)carbamate (48_Int-2) (2 g, 6.3397 mmol, 1.0 eq) in MeOH. Then add ammonium chloride (1.7 g, 31.9485 mmol, 5.0 eq) and iron (1.7 g, 31.9485 mmol, 5.0 eq) at RT (20 mL, 10V). Cool the RM to 0 °C and then add acetic acid (20 mL, 5V) dropwise. Heat the RM to 70 °C and stir for 16 h. After the reaction is complete, cool the RM to RT. Concentrate the RM, quench in cold water and basify by slowly adding saturated NaOH solution to a target pH of 10. Filter off the solid and extract the filtrate with DCM. Dry the combined organic fractions over Na2SO4; concentrate to give the compound (48_Int-3). MS (ES): 298.23 m / z [M+1]+, LCMS purity: 73.2%.

[0473] Step - 4. Synthesis of 4-(aminomethyl)-2-fluoro-6-methoxybenzimamidine (48_Int-4).

[0474] Prepare a stirred solution of tert-butyl (4-formamidinyl-3-fluoro-5-methoxybenzyl)carbamate (48_Int-3) (0.8 g, 25.473 mmol, 1.0 eq) in water (8 mL, 10V) and add concentrated HCl (2.6 mL, 3.3V) at rt and stir for 3 h. After the reaction is complete, concentrate the RM and triturate in methanol to give the compound (43_Int-4). MS (ES): 198.2 m / z [M+1]+, LCMS purity: 36%.

[0475] Step - 5. Synthesis of N-(4-formamidinyl-3-fluoro-5-methoxybenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxamide (48).

[0476] The final compound (48) was prepared from ethyl 1-(4-(2-cyanopropan-2-yl)benzyl)-1H-pyrazole-4-carboxylate (7_Int-5) and 4-(aminomethyl)-2-fluoro-6-methoxybenzimidamide dihydrochloride (48_Int-4) in a similar manner as described in (Example-25). MS(ES): 449.22 m / z [M+1]+, LCMS purity: 93.36%, HPLC purity: 93.46% 1H 400 MHz, DMSO-d6: δ1.660(s,6H), 3.843(s,3H), 4.436(d,J = 5.60Hz,2H), 5.364(s,2H), 6.878(d,J = 10Hz,2H), 6.966(s,1H), 7.324(d,J = 8Hz,2H), 7.507(d,J = 8.0Hz,2H), 7.919(s,1H), 8.299(s,1H), 8.778(t,J = 6.4Hz,1H), 9.139(s,2H), 9.383(s,2H).

[0477] Example 30: N-(4-Carbamimidoyl-3-fluorobenzyl)-1-(4-(cyanomethyl)benzyl)-1H-pyrazole-4-carboxamide (72)

[0478]

[0479] Step-1. Synthesis of N-(4-Carbamimidoyl-3-fluorobenzyl)-1-(4-(cyanomethyl)benzyl)-1H-pyrazole-4-carboxamide (72)

[0480] The final compound (72) was prepared from ethyl 1-(4-(cyanomethyl)benzyl)-1H-pyrazole-4-carboxylate (1_Int-4) and 4-(aminomethyl)-2-fluorobenzimidamide dihydrochloride (43_Int-5) in a similar manner as described in (Example-25). MS(ES): 391.22 m / z [M+1]+, LCMS purity: 100%, HPLC purity: 100% 1H 400 MHz, DMSO-d6: δ4.024(s,2H), 4.460(s,2H), 5.357(s,2H), 7.304 - 3.349(m,6H), 7.622(s,1H), 7.915(s,1H), 8.280(s,1H), 8.800(t,J = 6.4Hz,1H), 9.132(s,2H), 9.377(s,2H).

[0481] Example 31: N-(4-Carbamimidoyl-3-fluorobenzyl)-1-(4-(2-methoxyethyl)benzyl)-1H-pyrazole-4-carboxamide (73)

[0482]

[0483] Step - 1. Synthesis of N-(4-formamidin-3-fluorobenzyl)-1-(4-(2-methoxyethyl)benzyl)-1H-pyrazole-4-carboxamide (73)

[0484] The final compound (73) was prepared from ethyl 1-(4-(2-methoxyethyl)benzyl)-1H-pyrazole-4-carboxylate (15_Int-3) and 4-(aminomethyl)-2-fluorobenzamidine dihydrochloride (43_Int-5) in a similar manner as described in (Example - 25). MS(ES): 410.4 m / z [M+1]+, LCMS purity: 100%, HPLC purity: 99.67% 1H 400 MHz, DMSO-d6: δ 2.774 (t, J = 6.80 Hz, 2H), 3.212 (s, 3H), 3.500 (t, J = 6.80 Hz, 2H), 7.460 (d, J = 6.0 Hz, 2H), 5.303 (s, 2H), 7.175 - 7.224 (m, 4H), 7.319 (t, J = 7.2 Hz, 2H), 7.899 (s, 1H), 8.260 (s, 1H), 8.797 (t, J = 6.0 Hz, 1H), 9.182 (s, 2H), 9.377 (s, 2H).

[0485] Example 32: N-(4-formamidin-3-fluorobenzyl)-1-(4-(cyanomethyl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxamide (75)

[0486]

[0487] Step - 1. Synthesis of N-(4-formamidin-3-fluorobenzyl)-1-(4-(cyanomethyl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxamide (75)

[0488] The final compound (75) was prepared in a similar manner to that described in (Example-24) from methyl 1-(4-(cyanomethyl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxylate (85_Int-2) and 4-(aminomethyl)-2-fluorobenzamidine dihydrochloride (43_Int-5). MS(ES): 435.4 m / z [M+1]+, LCMS purity: 100%, HPLC purity: 100% 1H 400 MHz, DMSO-d6: δ 3.232 (s, 3H), 4.019 (s, 2H), 4.85 (d, J = 6.0 Hz, 2H), 4.812 (s, 2H), 5.365 (s, 2H), 7.242 (d, J = 7.6 Hz, 2H), 7.307 - 7.374 (M, 4H), 7.640 (t, = Hz, 1H), 8.039 (s, 1H), 8.871 (t, J = 6.0 Hz, 1H), 9.197 (s, 2H), 9.401 (s, 2H).

[0489] Example 33: N-(4-formamidinobenzyl)-1-(4-(methoxymethyl)benzyl)-1H-pyrazole-4-carboxamide (71)

[0490]

[0491] Step-1. Synthesis of ethyl 1-(4-(methoxymethyl)benzyl)-1H-pyrazole-4-carboxylate (71_Int-1)

[0492] A stirred solution of ethyl 1-(4-(hydroxymethyl)benzyl)-1H-pyrazole-4-carboxylate (36_Int-2) (2.5 g, 9.6153 mmol, 1.0 eq) in DMF (25 ml, 10V) was added with NaH (60% in mineral oil, 0.6 g, 12.4998 mmol, 2 eq) at 0 °C and stirred for 15 min. MeI (1.18 mL, 19.2306 mmol, 1.3 eq) was added dropwise to the RM at 0 °C. The RM was brought to RT and stirred for 5 h. After completion of the reaction, the RM was quenched in cold water and extracted with ethyl acetate. The combined organic fractions were dried over Na2SO4, concentrated and then purified by flash column chromatography (ethyl acetate in hexane 15%) to give the compound (71_Int-1). MS(ES): 274 m / z [M+1]+, 1H NMR (400 MHz, DMSO-d6) δ 1.267 (t, J = 7.2 Hz, 3H), 3.268 (s, 3H), 4.235 - 4.182 (m, 2H), 4.382 (s, 2H), 5.536 (s, 2H), 7.248 (t, J = 8 Hz 4H), 7.877 (s, 1H), 8.468 (s, 1H).

[0493] Step - 2. Synthesis of N-(4-formamidino benzyl)-1-(4-(methoxymethyl)benzyl)-1H-pyrazole-4-carboxamide (71).

[0494] The final compound (71) was prepared from ethyl 1-(4-(methoxymethyl)benzyl)-1H-pyrazole-4-carboxylate (71_Int-1) and 4-(aminomethyl)benzimidamide dihydrochloride in a similar manner as described in (Example - 24). MS(ES): 378.28 m / z [M+1]+, LCMS purity: 100%, HPLC purity: 98.18% 1H 400 MHz, DMSO-d6: δ 3.275 (s, 3H), 4.390 (s, 2H), 4.487 (d, J = 6.0 Hz, 2H), 5.353 (s, 2H), 7.248 - 7.318 (m, 4H), 7.50 (d, J = 8.0 Hz, 2H), 7.758 (d, J = 8.0 Hz, 2H), 7.924 (s, 1H), 8.281 (s, 1H), 8.791 (t, J = 6.0 Hz, 1H), 9.074 (s, 2H), 9.264 (s, 2H).

[0495] Example 34: N-(4-formamidino benzyl)-1-(4-(cyanomethyl)benzyl)-5-cyclopropyl-1H-pyrazole-4-carboxamide (86)

[0496]

[0497] Step - 1. Synthesis of tert-butyl 2-(4-(cyanomethyl)benzyl)hydrazine-1-carboxylate (86_Int-2)

[0498] A stirred solution of 2-(4-(bromomethyl)phenyl)acetonitrile (86_Int-1) (3 g, 14.2857 mmol, 1.0 eq) in dimethylacetamide (30 mL, 10V) was prepared at RT. Then DIPEA (3.68 g, 28.5714 mmol, 2.0 eq) and tert-butyl hydrazinecarboxylate (3.77 g, 28.5714 mmol, 2.0 eq) were added at RT. Then the RM was heated to 70 °C and stirred for 16 h. After the reaction was completed, the RM was quenched in DM water and extracted with ethyl acetate. The combined organic fractions were washed with brine solution, dried over Na2SO4; concentrated and then purified by flash column chromatography (10 - 14% ethyl acetate in hexane) to give the compound (86_Int-2) MS(ES): 261 m / z [M+1]+, LCMS purity: 70%.

[0499] Step - 2. Synthesis of 2-(4-(hydrazinomethyl)phenyl)acetonitrile (86_Int-3)

[0500] Prepare a stirred solution of tert-butyl 2-(4-(cyanomethyl)benzyl)hydrazine-1-carboxylate (86_Int-2) (1.0 g, 7.6628 mmol, 1.0 eq) in methanol (10 mL, 5V) and water (22 mL, 11V). Then add concentrated HCl (6.6 mL, 3.3V) to the RM at RT. Heat the RM to 80 °C and stir for 16 h. After the reaction is complete, evaporate the RM and triturate in ethyl acetate to obtain the compound (86_Int-3) MS (ES): 162.3 m / z [M+1]+, LCMS purity: 64%.

[0501] Step - 3. Synthesis of ethyl 1-(4-(cyanomethyl)benzyl)-5-cyclopropyl-1H-pyrazole-4-carboxylate (86_Int-4)

[0502] Prepare a solution of ethyl 3-cyclopropyl-3-oxopropanoate (1.0 g, 4.61531 mmol, 1.0 eq) and dimethylformamide dimethylacetal (0.6 g, 4.2307 mmol, 1.1 eq) and stir at 75 °C for 90 min. Cool the RM to RT, then add ethanol (12 mL, 20V), TEA (2.0 mL, 15.3844 mmol, 4.0 eq) and 2-(4-(hydrazinomethyl)phenyl)acetonitrile (86_Int-3) (0.600 g, 3.8461 mmol, 1.0 eq). Heat the RM to 80 °C and stir for 16 h. After the reaction is complete, quench the RM in water and extract with ethyl acetate. Wash the combined organic fractions with brine solution, dry over Na2SO4, concentrate and then purify by flash column chromatography (12 - 15% ethyl acetate in hexane) to obtain the compound (86_Int-4). MS (ES): 310 m / z [M+1]+, LCMS purity: 64%. 1H 400 MHz, DMSO-d6: δ 0.754 (d, J = 6.40 Hz, 2H), 0.963 (q, J = 6.4 Hz, 2H), 1.790 - 1.756 (m, 1H), 4.106 (s, 2H), 4.205 - 4.152 (m, 2H), 5.459 (s, 2H), 7.166 (d, J = 8 Hz, 2H), 7.240 (d, J = 8 Hz 2H), 7.814 (s, 1H).

[0503] Step - 4. Synthesis of N-(4-formimidoylbenzyl)-1-(4-(cyanomethyl)benzyl)-5-cyclopropyl-1H-pyrazole-4-carboxamide (86)

[0504] The final compound (86) was prepared from ethyl 1-(4-(cyanomethyl)benzyl)-5-cyclopropyl-1H-pyrazole-4-carboxylate (86_Int-4) and 4-(aminomethyl)benzimidine dihydrochloride in a similar manner as described in (Example-24). MS(ES): 412.5 m / z [M+1]+, LCMS purity: 100%, HPLC purity: 100% 1H 400 MHz, DMSO-d6: δ 0.754 (d, J = 6.40 Hz, 2H), 0.963 (q, J = 6.4 Hz, 2H), 1.790 - 1.756 (m, 1H), 4.025 (s, 2H), 4.487 (d, J = 6.4 Hz, 2H), 5.440 (s, 2H), 7.196 (d, J = 8 Hz, 2H), 7.33 (d, J = 8 Hz, 2H), 7.522 (d, J = 8 Hz 2H), 8.538 (d, J = 6.4 Hz, 2H), 8.964 (s, 2H), 9.266 (s, 2h).

[0505] Example 35: N-(4-Carbamimidoylbenzyl)-1-(4-(2-methoxyethyl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxamide (87)

[0506]

[0507] Step - 1. Synthesis of methyl 1-(4-(2-hydroxyethyl)benzyl)-3-(methoxymethyl)-1H-pyrazole-4-carboxylate (87_Int-2)

[0508] Prepare a stirred solution of methyl 5-(methoxymethyl)-1H-pyrazole-4-carboxylate (3.2 g, 1.88235 mmol, 1.0 eq) in DMF (32 mL, 10V) and add NaHMDS (1 M in THF) (18.88 mL, 1.88235 mmol, 1.0 eq) at 0 °C and stir for 30 min. Add 2-(4-(bromomethyl)phenyl)ethan-1-ol (87_Int-1) (4.0 g, 1.88235 mmol, 1.0 eq) to the RM at 0 °C. Allow the RM to reach RT and stir for 16 h. After completion of the reaction, quench the RM in water and extract with ethyl acetate. Dry the combined organic fractions over Na2SO4, concentrate and then purify by flash column chromatography (ethyl acetate in hexane 5%) (87_Int-2). Confirm by 2D NMR (ROE analysis). MS (ES): 304.9 m / z [M+1]+, LCMS purity: 99.72%, 1H 400 MHz, DMSO-d6: δ 2.679 (t, J = 4 Hz, 3H), 3.242 (s, 3H), 3.555 (t, J = 4 Hz, 2H), 3.724 (s, 3H), 4.490 (s, 2H), 5.276 (s, 2H), 7.203 - 7.184 (m, 4H), 8.416 (s, 1H).

[0509] Step - 2. Synthesis of methyl 1-(4-(2-methoxyethyl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxylate (87_Int-3)

[0510] A stirred solution of methyl 1-(4-(2-hydroxyethyl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxylate (87_Int-2) (0.4 g, 1.3157 mmol, 1.0 eq) in DMF (4 mL, 10V). NaH (60% in mineral oil) (0.126 g, 2.6314 mmol, 2 eq) was added slowly and the RM was stirred at 0 °C for 15 min, followed by the dropwise addition of MeI (0.106 mL, 1.7106 mmol, 1.3 eq). The RM was allowed to reach RT and stirred for 5 h. After completion of the reaction, the RM was quenched in cold water and extracted with ethyl acetate. The combined organic fractions were dried over Na2SO4, concentrated and then purified by flash column chromatography (ethyl acetate in hexane 15%) to give the compound (87_Int-3). MS (ES): 319 m / z [M+1]+, LCMS purity: 79%, 1H NMR (400 MHz, DMSO-d6) δ 400 MHz, DMSO-d6: δ 2.489 (t, J = 6.40 Hz, 2H), 3.211 (s, 3H), 3.225 (s, 3H), 3.518 (t, J = 6.4 Hz, 2H), 4.794 (s, 2H), 5.358 (s, 2H), 7.111 (d, J = 8 Hz, 2H), 7.200 (d, J = 8 Hz, 2H), 7.905 (s, 2H).

[0511] Step - 3. N-(4-carbamimidoylbenzyl)-1-(4-(2-methoxyethyl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxamide (87)

[0512] The final compound (87) was prepared from methyl 1-(4-(2-methoxyethyl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxylate (87_Int-3) and 4-(aminomethyl)benzimidine dihydrochloride in a similar manner as described in (Example-24). MS(ES): 435.53 m / z [M+1]+, LCMS purity: 100%, HPLC purity: 100% 1H 400 MHz, DMSO-d6: δ 2.489 (t, J = 6.40 Hz, 2H), 3.211 (s, 3H), 3.225 (s, 3H), 3.503 (t, J = 6.4 Hz, 2H), 4.492 (d, J = 6 Hz, 2H), 4.802 (d, 2H), 5.310 (d, 2H), 7.111 (d, J = 8 Hz, 2H), 7.183 (d, J = 8 Hz, 2H), 7.508 (d, J = 8 Hz, 2H), 7.752 (d, J = 8 Hz, 2H).8.012 (s, 1H), 8.817 (t, J = 6 Hz, 1H), 8.911 (s, 2H), 9.252 (s, 2H).

[0513] Example 36: Preparation of N-(4-carbamimidoylbenzyl)-1-(4-(2-(dimethylamino)-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxamide (33)

[0514]

[0515] Step-1. Synthesis of N-(4-carbamimidoylbenzyl)-1-(4-(2-(dimethylamino)-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxamide (33)

[0516] The final compound (33) was prepared from ethyl 1-(4-(2-(dimethylamino)-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxylate (33_Int-3) and 4-(aminomethyl)benzimidine dihydrochloride in a similar manner as described in (Example-24). MS(ES): 419.4 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 95.56% 1H NMR (400 MHz, DMSO-d6) δ 2.807 (s, 3H), 2.978 (s, 3H), 3.656 (s, 2H), 4.778 (d, J = 6.0 Hz, 2H), 7.183 - 7.197 (m, 5H), 7.483 - 7.504 (m, 2H), 7.734 - 7.755 (m, 1H), 7.837 (s, 1H), 7.907 (s, 1H), 8.269 (s, 1H), 8.767 (t, J = 6.0 Hz, 1H), 8.946 (s, 2H), 9.245 (s, 2H).

[0517] Example 37: Preparation of N-(4-formamidino-benzyl)-1-(2-(cyanomethyl)-benzyl)-1H-pyrazole-4-carboxamide (70)

[0518]

[0519] Step - 1. Synthesis of ethyl 1-(2-(cyanomethyl)-benzyl)-1H-pyrazole-4-carboxylate (70_Int-2)

[0520] Prepare a stirred solution of ethyl 1H-pyrazole-4-carboxylate (0.1 g, 0.71 mmol, 1.0 eq) in DMF (2 mL, 20 V) and add 2-(2-(bromomethyl)phenyl)acetonitrile (70_Int-1) (0.149 g, 0.71 mmol, 1.0 eq) at RT. Then heat the RM to 80 °C and stir for 16 h. After the reaction is complete, quench the RM in water and extract with ethyl acetate. Dry the combined organic fractions over Na2SO4, concentrate and purify by flash column chromatography (ethyl acetate in hexane 25%) to give the compound MS (ES): 270. m / z [M+1]+, LCMS purity: 98.57%, 1H 400 MHz, DMSO-d6: δ 1.244 (t, J = 6.4 Hz, 3H), 4.225 - 4.172 (m, 4H), 5.448 (s, 2H), 7.119 (d, J = 7.20 Hz, 1H), 7.454 - 7.332 (m, 3H), 7.892 (s, 1H), 8.453 (s, 1H).

[0521] Step - 2. Synthesis of N-(4-formamidino-benzyl)-1-(2-(cyanomethyl)-benzyl)-1H-pyrazole-4-carboxamide (70)

[0522] The final compound (70) was prepared from ethyl 1-(2-(cyanomethyl)benzyl)-1H-pyrazole-4-carboxylate (70_Int-2) and 4-(aminomethyl)benzimidamide dihydrochloride in a similar manner as described in (Example-24). MS(ES): 373.45 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 98.64. 1H 400 MHz, DMSO-d6: δ 4.185 (s, 2H), 4.483 (d, J = 6.0 Hz, 2H), 5.444 (s, 2H), 7.119 (s, 1H), 7.345 - 7.396 (m, 2H), 7.448 - 7.504 (m, 3H), 7.746 (d, J = 8.0 Hz, 2H), 7.943 (s, 1H), 8.259 (s, 1H), 8.784 (t, J = 6.0 Hz, 1H), 8.952 (s, 2H), 9.247 (s, 2H).

[0523] Example 38: Preparation of N-(4-formamidin-3-fluorobenzyl)-1-(4-(2-(methylamino)-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxamide (74)

[0524]

[0525] Step-1. Synthesis of N-(4-formamidin-3-fluorobenzyl)-1-(4-(2-(methylamino)-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxamide (74)

[0526] The final compound (74) was prepared from ethyl 1-(4-(2-(methylamino)-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxylate (19_Int-3) and 4-(aminomethyl)-2-fluorobenzimidamide dihydrochloride (43_Int-6) in a similar manner as described in (Example-24). MS(ES): 423.4 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 98.51% 1H NMR (400 MHz, DMSO-d6) δ 2.678 (s, 3H), 3.367 (s, 2H), 4.467 (d, J = 6 Hz, 2H), 5.317 (s, 2H), 7.196 - 7.243 (m, 4H), 7.328 (t, J = 8.4 Hz, 2H), 7.262 (t, J = 8.4 Hz, 1H), 7.906 (s, 1H), 7.961 (s, 1H), 8.268 (s, 1H), 8.807 (t, J = 6.0 Hz, 1H), 9.186 (s, 2H), 9.387 (s, 2H).

[0527] Example 39: Preparation of N-(4-carbamimidoylbenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxamide (82)

[0528]

[0529] Step-1. Synthesis of N-(4-carbamimidoylbenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxamide (82)

[0530] The final compound (70) was prepared in a similar manner as described in (Example-24) from methyl 1-(4-(2-cyanopropan-2-yl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxylate (82_Int-3) and 4-(aminomethyl)benzenecarboximidamide dihydrochloride. MS(ES): 445.4 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 100% 1H NMR (400 MHz, DMSO-d6) δ 1.675 (s, 6H), 3.246 (s, 3H), 4.514 (d, J = 6 Hz, 2H), 4.480 (s, 2H), 5.380 (s, 2H), 7.274 (d, J = 8.4 Hz, 2H), 7.486 - 7.537 (m, 4H), 7.777 (d, J = 8.4 Hz, 2H), 8.046 (s, 1H), 8.843 (t, J = 6 Hz, 1H), 9.022 (s, 2H), 9.274 (s, 2H).

[0531] Example 40: Preparation of N-(4-carbamimidoylbenzyl)-1-(4-(2-cyanopropan-2-yl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxamide (85)

[0532]

[0533] Step-1. Synthesis of N-(4-carbamimidoylbenzyl)-1-(4-(cyanomethyl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxamide (85)

[0534] The final compound (70) was prepared from methyl 1-(4-(cyanomethyl)benzyl)-5-(methoxymethyl)-1H-pyrazole-4-carboxylate (85_Int-3) and 4-(aminomethyl)benzimidine dihydrochloride in a similar manner as described in (Example-24). MS(ES): 417.4 m / z [M+H]+, LCMS purity: 100%, HPLC purity: 100% 1H NMR (400 MHz, DMSO-d6) δ 3.233 (s, 3H), 4.029 (s, 2H), 4.514 (d, J = 6 Hz, 2H), 4.829 (s, 2H), 5.375 (s, 2H), 7.232 (d, J = 8 Hz, 2H), 7.327 (d, J = 8 Hz, 2H), 7.529 (d, J = 8.0 Hz, 2H), 7.774 (d, J = 8.0 Hz, 2H), 8.046 (s, 1H), 8.839 (t, J = 6 Hz, 1H), 8.963 (s, 2H), 9.271 (s, 2H).

[0535] Example 41: Preparation of N-(4-formamidin-3-fluorobenzyl)-1-(4-(2-(dimethylamino)-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxamide (78)

[0536]

[0537] Step-1. Synthesis of 1-(4-(2-(dimethylamino)-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxylic acid (78_Int-1)

[0538] Prepare a stirred solution of ethyl 1-(4-(2-(dimethylamino)-2-oxoethyl)benzyl)-1H-pyrazole-4-carboxylate (33_Int-3) (0.5 g, 0.158 mmol, 1.0 eq) in water:methanol:THF (1:1:1) (5 mL, 10V) and add LiOH (0.3 g, 0.793 mmol, 5 eq) at RT and stir for 16 h. After completion of the reaction, quench the RM in 2N HCl (approx. pH 4) and extract with 10% methanol in DCM. Combine the organic fractions and dry over Na 2 SO 4 dry; concentrate to give compound (78_Int-1) MS(ES): 288.4. m / z [M+H]+, LCMS purity: 97.47%, 1H NMR (400 MHz, DMSO-d6) δ 2.804 (s, 3H), 2.975 (s, 3H), 5.315 (s, 2H), 7.191 - 7.219 (m, 4H), 7.795 (s, 1H), 8.357 (s, 1H), 12.301 (s, 1H).

[0539] Step - 2. Synthesis of N-(4 - formamidin - 3 - fluorobenzyl)-1-(4-(2-(dimethylamino)-2 - oxoethyl)benzyl)-1H - pyrazole - 4 - carboxamide (78)

[0540] Prepare a stirred solution of 1-(4-(2-(dimethylamino)-2 - oxoethyl)benzyl)-1H - pyrazole - 4 - carboxylic acid (78_Int - 1) (0.2 g, 0.069 mmol, 1.0 eq) in pyridine (2 mL, 10V). At RT, add -(aminomethyl)-2 - fluorobenzamidine dihydrochloride (43_Int - 5) (0.2 g, 0.083 mmol, 1.2 eq) and EDC HCl (0.67 g, 0.348 mmol, 5.0 eq) to the RM and stir for 16 h. After the reaction is complete, concentrate the RM and purify it by PREP HPLC ((A) 0.1% aqueous TFA (B) 100% MeCN). Lyophilize the pure fractions to obtain the compound (78) MS (ES): 437.36. m / z [M + H]+, LCMS purity: 98.37%, HPLC purity: 99.70% 1H NMR (400 MHz, DMSO - d6) δ 2.804 (s, 3H), 2.981 (s, 3H), 3.653 (s, 2H), 4.459 (d, J = 6.4 Hz, 2H), 5.314 (s, 2H), 7.194 (s, 4H), 7.318 (t, J = 11.6 Hz, 2H), 7.615 (t, J = 11.6 Hz, 1H), 7.900 (s, 1H), 8.266 (s, 1H), 8.790 (t, J = 6 Hz, 1H), 9.170 (s, 2H), 9.370 (s, 2H).

[0541] Biological Examples

[0542] Plasma kallikrein protease inhibition assay. Two methods are provided for determining the IC 50 of the test compound against plasma kallikrein.

[0543] In the first method, a reaction buffer consisting of 25 mM Tris - HCl (pH 8.0), 100 mM NaCl (pH 8.5), 0.01% Brij35 and 1% DMSO (final) is used. The enzyme used is plasma kallikrein (R&D Systems catalog number 2497 - SE; recombinant human plasma kallikrein, expressed in the mouse myeloma cell line, NS0 - derived Gly20 - Ala638, with a C - terminal 60 - His tag, MW = 70 kDa).

[0544] The enzyme was activated as follows: diluted to 200 μg / mL in activation buffer (100 mM Tris, 10 mM CaCl2, 150 mM NaCl, pH 7.5 (TCN)), and then combined with an equal volume of 20 μg / mL thermolysin to form a reaction buffer. Each test compound was then dissolved in DMSO and introduced into the reaction buffer. After a 20-minute pre-incubation period, the reaction was initiated by introducing a substrate solution containing 10 μM Z-FR-AMC (Enzo catalog number P-139; AMC: 7-amino-4-methylcoumarin) into the reaction wells.

[0545] Measurements were performed using EnVision (PE), with excitation and emission wavelengths of 355 nm and 460 nm, respectively. The reaction was stopped with EDTA. Enzyme activity was monitored every 5 minutes at room temperature for 120 minutes as a time course measurement of the increase in the fluorescently labeled peptide substrate signal.

[0546] Data were analyzed by obtaining the slope of the linear portion of the measurement * (signal / time). The slope was calculated using Excel, and curve fitting was performed using Prism software.

[0547] Secondly, plasma kallikrein activity in pooled human plasma was also measured. First, a 10% actin FS solution was prepared in assay buffer. Each test compound was dissolved in DMSO and sent to the reaction mixture together with the Z-FR-AMC substrate and pooled human plasma. The multi-well reaction plate was incubated at room temperature for five minutes. To initiate the reaction, the 10% actin FS solution was added to each well, and kinetic measurements were performed at Ex / Em of 355 / 460 nm. Fluorescence signals were recorded every 30 seconds for a total of 10 minutes.

[0548] Permeability assay. Permeability studies were performed using Caco-2 cells (ECACC catalog number 09042001) seeded at a density of 80,000 cells / well on cell insert plates (Millicell, catalog number PSHT010R5), and the cells were maintained in medium (1x DMEM, containing 10% FBS, 0.1 mg / mL penicillin / streptomycin) for 18 - 21 days. The medium was changed every other day. Before the experiment, the integrity of the cell monolayer was evaluated by measuring the TEER value using a volt-ohmmeter and STX100C96 electrodes. Only monolayers with a TEER value greater than 800 ohm.cm in the buffer after the first wash were used. 2 of the monolayer.

[0549] The assay buffer (HBSS containing Ca+2 and Mg+2 buffered with 10 mM HEPES and 25 mM D-glucose, pH - 7.4) was used on both the apical and basolateral sides.

[0550] Prepare an intermediate stock solution of the test compound in DMSO at a concentration of 1 mM (in DMSO). Add this stock solution to the assay buffer to achieve a target test compound concentration of 10 μM. The organic content of the final drug formulation is 1.0% v / v. The bidirectional permeability experiment is performed in singlet, and the sample analysis is performed in duplicate.

[0551] Wash the cultured cell monolayer twice with the assay buffer (add 0.4 mL and 0.8 mL to the apical side and basolateral side of the culture plate, respectively), and then discard the buffer in both compartments.

[0552] For the apical-to-basolateral (AP>BL) experiment, aliquots of 0.4 mL of the donor solution (assay buffer, pH 7.4, containing the test compound) and 0.8 mL of the receptor solution (assay buffer only, pH 7.4) are added to the apical and basolateral compartments, respectively. For the basolateral-to-apical (BL>AP) experiment, aliquots of 0.8 mL of the donor solution (pH-7.4 assay buffer containing the test compound) and 0.4 mL of the receptor solution (pH-7.4 assay buffer) are added to the basolateral and apical compartments, respectively. Then place the plate in an incubator at 37 °C for 120 minutes.

[0553] On the same experimental day, control experiments in both directions (AP>BL and BL>AP) of propranolol (high permeability), atenolol (low permeability), digoxin (high efflux - Pgp substrate), and digoxin + verapamil (Pgp inhibitor) are performed in separate wells.

[0554] After the transport experiment is completed, evaluate the integrity of the cell monolayer by measuring fluorescein yellow (LY) exclusion. For this, add 400 μL of 10 μM LY to each well of the filter plate and incubate at 37 °C for 1 hour. Subsequently, collect samples from the basolateral compartment and measure LY fluorescence using an excitation wavelength of 485 nm and an emission wavelength of 530 nm. Calculate the percentage of LY exclusion across the cell monolayer by measuring the fluorescence in the receiving plate (basolateral compartment) and comparing it with the theoretical equilibrium standard.

[0555] Analyze the samples (collected from the apical and basolateral compartments after 120 minutes of incubation) by LCMS / MS, and then calculate the Papp of the compound in the apical-to-basolateral and basolateral-to-apical directions. Papp = ([DBL] × VBL) / (A × t × [DAP]), where: [DBL] = final drug concentration in the basolateral side, VBL = volume of the basolateral compartment, A = surface area of cell culture, t = total incubation time, and [DAP] = initial drug concentration in the apical side.

[0556] Table 1 below gives the results from the above assays for representative compounds of the present disclosure. The scores for the selected compounds in each assay are shown below:

[0557]

[0558] Table 1. Plasma Kallikrein Inhibition and Cellular Permeability of Representative Compounds

[0559]

[0560]

[0561]

[0562]

[0563]

[0564]

[0565]

[0566]

[0567]

[0568]

[0569]

[0570]

[0571]

[0572]

[0573]

[0574]

[0575]

[0576]

[0577]

[0578]

[0579]

[0580]

[0581]

[0582]

[0583]

[0584]

[0585]

[0586]

[0587] It should be understood that the embodiments and implementations described herein are for illustrative purposes only, and various modifications or variations of these embodiments and implementations will occur to those skilled in the art, and such modifications or variations will be included within the spirit and scope of this application and the scope of the appended claims. All publications, patents, and patent applications cited herein are hereby incorporated by reference in their entirety for all purposes.

Claims

1. A compound of formula (IA) or a pharmaceutically acceptable salt thereof: Wherein D 1 is N or CR 1 , D 2 is N or CR 2 , D 3 is N or CR 3 , and D 4 is N or CR 4 , where D 1 , D 2 , D 3 and D 4 no more than three of them are N at the same time; R 1 , R 2 , R 3 and R 4 Independently selected from the group consisting of: H, C 2 -C 6 -Alkenyl, C 1 -C 6 -haloalkyl, halo, NR a R b , OR a 、-NR a C(O)R b 、-C(O)R a 、-C(O)halogen、-OC(O)R a 、-OC(O)OR a 、-C(O)OR a , C 6 -C 10 -Aryl, CN, -S(O) 0-2 R a 、-S(O) 2 OR a and NO 2 ; R a and R b each independently selected from the group consisting of: H, C 1 -C 6 -alkyl and C 1 -C 6 -haloalkyl; R c1 , R c2 and R c3 Independently selected from the group consisting of: H, C 1 -C 6 -alkyl and C 1 -C 6 - haloalkyl; Q 1 、 Q 2 and Q 3 are independently selected from the group consisting of: CR 5 , N, O, and S; R 5 selected from the group consisting of: H, C 1 -C 6 -alkyl, OR a 、-(C 1 -C 6 -alkyl)OR a and C 3 -C 10 -cycloalkyl; P is C or N; L 1 is -SO 2 - or -C 1 -C 8 -alkylene-; a divalent monocyclic or bicyclic moiety selected from the group consisting of: C 3 -C 10 -cycloalkyl, C 6 -C 10 -aryl, 3- to 10-membered heterocycloalkyl (wherein 1-4 ring members are independently selected from N, O, and S), 5- to 10-membered heteroaryl (wherein 1-4 heteroaryl members are independently selected from N, O, and S), and fused combinations thereof; L 2 selected from the group consisting of: -C 1 -C 8 -alkylene, -C 2 -C 8 -alkenylene, -C 2 -C 8 -alkynylene, and a divalent moiety selected from the group consisting of: C 3 -C 10 -cycloalkyl, C 3 -C 10 -cycloalkenyl, C 6 -C 10 -aryl, 5- to 10-membered heteroaryl (wherein 1-4 heteroaryl members are independently selected from N, O, and S), (C 1 -C 8 -alkyl)C 3 -C 10 -cycloalkyl, (C 1 -C 8 -alkyl)C 3 -C 10 -cycloalkenyl, (C 1 -C 8 -alkyl)C 6 -C 10 -aryl and (C 1 -C 8 -alkyl) 5- to 10-membered heteroaryl (wherein 1-4 heteroaryl members are independently selected from N, O, and S), wherein the cycloalkyl, cycloalkenyl, aryl, and heteroaryl are monocyclic or bicyclic; wherein L 2 is optionally substituted with 1 to 3 substituents selected from the group consisting of C 1 -C 6 -alkyl, C 1 -C 6 -haloalkyl and CN; Z is selected from the group consisting of: -OR c 、-OC(O)R c 、-OC(O)NR c R d 、-S(O) 0-2 R c 、-CN, -C(O)R c 、-C(O)OR c 、-C(O)NR c R d 、-C(S)R c 、-NR c R d 、=NR c 、-NR c C(O)NR c R d 、-NR c CO 2 R d 、-NR c -NO, -NO 2 、-NR c -OR d 、-N=C=O, -N=C=S and -NR c -NR c R d ; and R c and R d each instance of which is independently selected from H, C 1 -C 6 -alkyl, C 1 -C 6 -haloalkyl, and C 6 -C 10 -aryl.

2. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein no more than one of D 1 , D 2 , D 3 and D 4 is N.

3. The compound according to claim 1 or 2, or a pharmaceutically acceptable salt thereof, which contains D 1 , D 2 , D 3 and D 4 The rings are selected from the group consisting of:

4. The compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof, wherein the ring containing D 1 , D 2 , D 3 and D 4 is 5. A compound according to any one of claims 1 to 4 or a pharmaceutically acceptable salt thereof, wherein R 1 , R 2 , R 3 and R 4 are independently selected from the group consisting of: H, OR a , a halogenated group, and CN.

6. The compound according to any one of claims 1 to 5 or a pharmaceutically acceptable salt thereof, wherein R 1 , R 2 , R 3 and R 4 are independently selected from the group consisting of: H and halo groups.

7. A compound according to any one of claims 1 to 6 or a pharmaceutically acceptable salt thereof, wherein R 1 , R 2 , R 3 and R 4 is at least one halo group.

8. A compound according to any one of claims 1 to 6 or a pharmaceutically acceptable salt thereof, wherein R 1 , R 2 , R 3 and R 4 are each H.

9. The compound according to any one of claims 1 to 8 or a pharmaceutically acceptable salt thereof, wherein R c1 , R c2 and R c3 are each H.

10. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 9, wherein Q 1 is C; Q 2 and Q 3 each independently is selected from the group consisting of: CR 5 , N, O, and S; and P is selected from the group consisting of C and N.

11. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 10, wherein P is N.

12. The compound according to any one of claims 1 to 11 or a pharmaceutically acceptable salt thereof, wherein Q 2 is CR 5 and Q 3 is N.

13. The compound according to any one of claims 1 to 11 or a pharmaceutically acceptable salt thereof, wherein Q 2 is N and Q 3 is CR 5 .

14. The compound according to any one of claims 1 to 13 or a pharmaceutically acceptable salt thereof, wherein L 1 is -C 1 -C 8 -alkylene-.

15. The compound according to any one of claims 1 to 14 or a pharmaceutically acceptable salt thereof, wherein L 1 is -C 1 -C 3 -alkylene-.

16. The compound according to any one of claims 1 to 15 or a pharmaceutically acceptable salt thereof, wherein is a divalent monocyclic C 6 -C 10 -aryl.

17. The compound according to any one of claims 1 to 16 or a pharmaceutically acceptable salt thereof, wherein is 18. A compound according to any one of claims 1 to 17 or a pharmaceutically acceptable salt thereof, wherein L 2 is selected from -C 1 -C 8 -alkylene, C 6 -C 10 -aryl and (C 1 -C 8 -alkyl)C 6 -C 10 -aryl.

19. The compound according to any one of claims 1 to 18 or a pharmaceutically acceptable salt thereof, wherein L 2 is -C 1 -C 8 -alkylene.

20. The compound according to any one of claims 1 to 19 or a pharmaceutically acceptable salt thereof, wherein L 2 is C 1 -C 3 -alkylene.

21. The compound according to any one of claims 1 to 18 or a pharmaceutically acceptable salt thereof, wherein L 2 is C 6 -C 10 -aryl.

22. The compound according to any one of claims 1 to 21 or a pharmaceutically acceptable salt thereof, wherein L 2 is phenyl.

23. The compound according to any one of claims 1 to 22 or a pharmaceutically acceptable salt thereof, wherein Z is selected from the group consisting of: -OR c , CN, -C(O)OR c and -C(O)NR c R d .

24. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 23, wherein Z is CN.

25. The compound according to any one of claims 1 to 23 or a pharmaceutically acceptable salt thereof, wherein R c and R d are each independently H or C 1 -C 6 -alkyl.

26. The compound according to any one of claims 1 to 24 or a pharmaceutically acceptable salt thereof, wherein Z is selected from the group consisting of: OH, OCH 3 , -COOH, -C(O)NH 2 and -C(O)NHCH 3 .

27. The compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein the compound has the formula (I): Wherein Q 1 , Q 2 and Q 3 is selected from the group consisting of C, N, O, and S; P is selected from the group consisting of C and N; L 1 is a linking group selected from the group consisting of: C 1 -C 8 alkyl linker and SO 2 ; For i) a cyclic hydrocarbon, bicyclic hydrocarbon or heterocycle containing at most 10 atoms consisting of C or N; or ii) selected from the group consisting of: cyclopentyl, cyclohexyl, phenyl, naphthyl, pyridine, pyrimidine, indene, 2,3-dihydro-1H-indene or any of their saturated and unsaturated cyclic hydrocarbons or heterocycles; L 2 For i) a hydrocarbon without a double bond formed with O or S, ii) a hydrocarbon and without heteroatoms such as O, N or S; or iii) A linking group selected from the group consisting of: C 1 -C 8 an alkyl linker comprising a saturated hydrocarbon, an unsaturated hydrocarbon, a branched hydrocarbon, a cyclic hydrocarbon, and combinations thereof; a cyclic hydrocarbon selected from the group consisting of cyclopentyl, cyclohexyl, phenyl, naphthyl, indene, and 2,3-dihydro-1H-indenene, or any saturated or unsaturated cyclic hydrocarbon thereof; Z is i) selected from the group consisting of: -OH, -OR’, -OC(O)H, -OC(O)R’, -OC(O)NH 2 , -OC(O)NHR’, -OC(O)NH(R’) 2 , -SH, -SR’, -S(O)R’, -S(O) 2 R’, -CN, -C(O)H, -C(O)R’, -C(O)OH, -C(O)OR’, -C(O)NH 2 , -C(O)NHR’, -C(O)NH(R’) 2 , -C(S)R’, -NH 2 , -NH 2 R’, -NR’ 2 , =NH, =NR’, -NHC(O)NH 2 , -NR’C(O)NH 2 , -NR’C(O)NHR’, -NR’C(O)N(R’) 2 , -NHCO 2 H, -NHCO 2 R’, -NR’CO 2 R’, -NH-NO, -NR’-NO, -NO 2 , -NH-OH, -OH, -NR’-OR’, -N=C=O, -N=C=S, -NH-NH2 and -NH-NHR’, where each R’ is independently an alkyl or alkyl halide, or ii) not a halogen or hydrogen.

28. The compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein the compound is selected from the following table:

29. A pharmaceutical composition comprising the compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 28 and a pharmaceutically acceptable carrier.

30. A method for treating a subject suffering from a disease or disorder, which comprises administering to the subject a compound according to any one of claims 1 to 28 or a pharmaceutically acceptable salt thereof, wherein the disease or disorder is selected from the group consisting of: ischemic stroke, hemorrhagic stroke, hypertension, retinopathy, diabetic retinopathy, nephropathy, cerebral edema, pulmonary hypertension, inflammation, acute myocardial infarction, deep vein thrombosis, complications from fibrinolytic therapy, stroke, angina, angioedema, sepsis, arthritis, cardiopulmonary bypass complications, capillary leak syndrome, inflammatory bowel disease, vascular complications from diabetes, diabetic macular edema, macular degeneration, neuropathy, age-related macular degeneration, retinal vein occlusion, cerebral edema, ischemia-reperfusion injury, angiogenesis, asthma, allergic reaction, Alzheimer's disease, Parkinson's disease, multiple sclerosis, glioblastoma multiforme, complications from fibrinolytic therapy, increased albumin excretion, macroalbuminuria, pain, amyotrophic lateral sclerosis, Creutzfeldt-Jakob disease, epilepsy, traumatic brain injury, high altitude cerebral edema, cancer, disseminated intravascular coagulation, pancreatitis, inflammation, shock, hereditary angioedema (HAE), uveitis, polyangiitis, acute respiratory distress syndrome (ARDS), thrombosis, vasculitis, Crohn's disease, ulcerative colitis, enterocolitis, arteritis, glomerulonephritis, psoriasis, endometriosis, preeclampsia, malaria, arthritis, periodic and recurrent fevers, Chagas disease, Raynaud's disease, systemic sclerosis, granulomatosis with polyangiitis, small vessel vasculitis, medium vessel vasculitis, large vessel vasculitis, panvasculitis, systemic autoinflammatory diseases, renal insufficiency, cerebral malaria, Clarkson's disease (systemic vascular leak syndrome), hantavirus infection, hantavirus renal syndrome, hantavirus pulmonary syndrome, virus-related inflammatory disorders, retinal vasculitis, uveitis, Eales' disease, Behçet's disease, sarcoidosis, whooping cough, coronavirus infection and non-infectious posterior uveitis.

31. The method according to claim 30, wherein the disease or disorder is selected from the group consisting of: diabetic macular edema, diabetic retinopathy and uveitis.

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