Plasma kallikrein inhibitors

Abstract

The present invention provides compounds of formula (I), pharmaceutical compositions comprising one or more of said compounds, and methods of using said compounds to treat or prevent one or more disease states in which inhibition of plasma kallikrein is believed to be beneficial, such as hereditary angioedema, uveitis, posterior uveitis, wet age-related macular edema, diabetic macular edema, diabetic retinopathy, and retinal vein occlusion. The compounds are selective inhibitors of plasma kallikrein. [Formula 1] TIFF2023551150000099.tif37138

Description

[Technical Field]

[0001] Plasma kallikrein is a trypsin-like serine protease zymogen present in plasma. Its gene structure is similar to that of factor XI. Overall, the amino acid sequence of plasma kallikrein shares 58% homology with factor XI. Proteolytic activation by factor XIIa at the internal I389-R390 bond generates a heavy chain (371 amino acids) and a light chain (248 amino acids). The active site of plasma kallikrein is contained in the light chain. The light chain of plasma kallikrein reacts with protease inhibitors such as α2-macroglobulin and Cl- inhibitors. Interestingly, heparin significantly accelerates the inhibition of plasma kallikrein by antithrombin III in the presence of high molecular weight kininogen (HMWK). In blood, the majority of plasma kallikrein circulates in a complex with HMWK. Plasma kallikrein cleaves HMWK, releasing bradykinin. The release of bradykinin leads to increased vascular permeability and vasodilation (for reviews, see Coleman, R., "Contact Activation Pathway", Hemostasis and Thrombosis, pp. 103-122, Lippincott Williams & Wilkins (2001); Schmaier AH, "Contact Activation", Thrombosis and Hemorrhage, pp. 105-128 (1998)).

[0002] Patients with a genetic deficiency in C1-esterase inhibitor suffer from hereditary angioedema (HAE), a lifelong condition that causes intermittent swelling throughout the body, including the hands, feet, face, throat, genitals, and gastrointestinal tract. Analysis of blisters resulting from acute episodes has shown that they contain high levels of plasma kallikrein, and treatment with ecallantide (Carbitol), a protein-based reversible plasma kallikrein inhibitor, has been approved by the FDA for the treatment of acute attacks of HAE (Schneider, L, et al., J. Allergy Clin. Immunol., 120; p. 416 (2007)).

[0003] Furthermore, the plasma kallikrein-kinin system is abnormally abundant in patients diagnosed with advanced diabetic macular edema (DME). Recent publications have shown that plasma kallikrein contributes to the retinal vascular leakage and dysfunction observed in diabetic rodent models (A. Clermont, et al., Diabetes, 60:1590(2011)), and that treatment with a small molecule plasma kallikrein inhibitor improved the observed retinal vascular permeability and other abnormalities related to retinal blood flow. [Prior art documents] [Non-patent literature]

[0004] [Non-Patent Document 1] Coleman, R., ″Contact Activation Pathway″, Hemostasis and Thrombosis, pp.103-122, Lippincott Williams & Wilkins (2001) [Non-patent document 2] Schmaier AH, ″Contact Activation″, Thrombosis and Hemorrhage, pp.105-128(1998) [Non-patent document 3] Schneider, L, et al., J. Allergy Clin. Immunol., 120; p.416(2007) [Non-patent document 4] A. Clermont, et al., Diabetes, 60:1590(2011) Summary of the Invention [Problem to be solved by the invention]

[0005] It would be desirable in the art to develop plasma kallikrein inhibitors that have utility in treating a wide range of disorders, including hereditary angioedema, diabetic macular edema, and diabetic retinopathy. [Means for solving the problem]

[0006] The present invention relates to compounds of formula I: [ka] and pharmaceutically acceptable salts thereof. The compounds of Formula I are inhibitors of plasma kallikrein and may therefore be useful in the treatment, inhibition, or amelioration of one or more disease states that would benefit from inhibition of plasma kallikrein, such as hereditary angioedema, uveitis, posterior uveitis, wet age-related macular edema, diabetic macular edema, diabetic retinopathy, and retinal vein occlusion. The compounds of the present invention may also be used in combination with other therapeutically effective agents, including, but not limited to, other agents useful in the treatment of hereditary angioedema, uveitis, posterior uveitis, wet age-related macular edema, diabetic macular edema, diabetic retinopathy, and retinal vein occlusion. The present invention further relates to methods for preparing the compounds of Formula I, as well as pharmaceutical compositions comprising the compounds of Formula I and pharmaceutically acceptable salts thereof. DETAILED DESCRIPTION OF THE INVENTION

[0007] The present invention relates to compounds of formula I below or a pharmaceutically acceptable salt thereof: [ka] During the ceremony, X is N or CH; R 1 is hydrogen, halo, hydroxy and C 1-6 selected from the group consisting of alkyl; R 2 is hydrogen, halo, hydroxy and C 1-6 selected from the group consisting of alkyl; R 3 is hydrogen, halo, hydroxy and C 1-6 alkyl, wherein said alkyl groups are independently selected from the group consisting of halo, cyano, and OR x may be substituted with 1 to 4 substituents selected from the group consisting of: R 4 is hydrogen, halo, hydroxy and C 1-6 alkyl; said alkyl groups are independently selected from the group consisting of halo, cyano, and OR x may be substituted with 1 to 4 substituents selected from the group consisting of: R 5 is NR 9 R 10 OR x and; Each R 6 are independently hydrogen, halo, hydroxy and C 1-6 alkyl, wherein the alkyl group is optionally substituted with 1 to 3 halo; Each R 7 is hydrogen, halo, hydroxy and C 1-6 alkyl, wherein the alkyl group is optionally substituted with 1 to 3 halo; or R 6 and R 7 may be taken together with the carbon atom to which they are attached to form a 3- to 6-membered cycloalkyl group optionally substituted with 1 or 2 halo; R 8 is hydrogen; halo; hydroxy; R x ;OR x ;Phenyl;Indan;OR yheteroaryl, which may be monocyclic or bicyclic; heterocycle; and C, which may be monocyclic or bicyclic 3-6 cycloalkyl; the phenyl and heteroaryl groups are independently selected from the group consisting of oxo, halo, R x , OR x , N.R. 9 R 10 , N.R. 9 (C=O)R x , N.R. 9 (C=O)OR x , (C=O)OR x , (C=O)NR 9 , R y and OR y and wherein the cycloalkyl and heterocyclic groups are independently substituted with 1 to 3 substituents selected from the group consisting of oxo, halo, R x and OR x may be substituted with 1 to 3 substituents selected from the group consisting of: R 9 is hydrogen or C 1-3 is alkyl; R 10 is hydrogen or C 1-3 is alkyl; R x is hydrogen or C optionally substituted with 1 to 3 substituents selected from the group consisting of halo and hydroxy 1-6 is alkyl, R y is phenyl, heterocycle or C 3-6 cycloalkyl, wherein the phenyl group is optionally substituted with 1 to 3 halo, the heterocyclic group is optionally substituted with 1 or 2 oxo, and the cycloalkyl group is C 1-6 optionally substituted with alkyl; n is an integer of 0 to 2.

[0008] In one embodiment of the invention, X is CH. In another embodiment of the invention, X is N.

[0009] In one embodiment of the present invention, R 1 is a halo. In one class of embodiments, R1 is chloro.

[0010] In one embodiment of the present invention, R 2 is halo. In one class of the invention, R 2 is fluoro.

[0011] In one embodiment of the present invention, R 3 is hydrogen. In another embodiment of the present invention, R 3 is methyl.

[0012] In one embodiment of the present invention, R 4 is hydrogen. In another embodiment of the present invention, R 3 is methyl.

[0013] In one embodiment of the present invention, R 5 is NH2. In another embodiment of the present invention, R 5 is OH.

[0014] In one embodiment of the present invention, R 6 is hydrogen.

[0015] In one embodiment of the present invention, R 7 is hydrogen.

[0016] In one embodiment of the present invention, R 8 is phenyl; said phenyl is independently halo, R x , OR x , N.R. 9 R 10 , N.R. 9 (C=O)R x , N.R. 9 (C=O)OR x , (C=O)NR 9 , (C=O)OR x , R y and OR y In one class of the embodiment, R 8 is phenyl; said phenyl is independently halo, Rx , OR x , R y and OR y It may be substituted with 1 to 3 substituents selected from the group consisting of:

[0017] In one embodiment of the invention, n is 0. In another embodiment of the invention, n is 1. In another embodiment of the invention, n is 2.

[0018] Reference to the above preferred classes and subclasses is meant to include all combinations of particular and preferred groups unless otherwise specified.

[0019] Specific embodiments of the present invention include, but are not limited to, the compounds identified herein as Examples 1-133, or pharmaceutically acceptable salts thereof.

[0020] Also included within the scope of the present invention is a pharmaceutical composition comprising a compound of Formula I above and a pharmaceutically acceptable carrier. The present invention is also intended to encompass pharmaceutical compositions containing a pharmaceutically acceptable carrier and any of the compounds specifically disclosed herein. These and other aspects of the present invention will be apparent from the teachings contained herein.

[0021] The present invention includes compositions for treating diseases or conditions in which plasma kallikrein activity is implicated. Accordingly, the present invention includes compositions for treating visual activity disorders, diabetic retinopathy, diabetic macular edema, retinal vein occlusion, hereditary angioedema, diabetes, pancreatitis, cerebral hemorrhage, nephropathy, cardiomyopathy, neuropathy, inflammatory bowel disease, arthritis, inflammation, septic shock, hypotension, cancer, adult respiratory distress syndrome, disseminated intravascular coagulation, blood clotting during cardiopulmonary bypass surgery, and bleeding from postoperative surgery in mammals, comprising a compound of the present invention in a pharmaceutically acceptable carrier. One class of the present invention includes compositions for treating hereditary angioedema, uveitis, posterior uveitis, wet age-related macular edema, diabetic macular edema, diabetic retinopathy, and retinal vein occlusion. These compositions may optionally contain anti-inflammatory agents, anti-VEGF agents, immunosuppressants, anticoagulants, antiplatelet agents, and thrombolytic agents. The compositions can be added to blood, blood products, or mammalian organs to exert the desired inhibition.

[0022] The present invention also includes a composition for preventing or treating retinal vascular permeability associated with diabetic retinopathy and diabetic macular edema in a mammal, the composition comprising a compound of the present invention in a pharmaceutically acceptable carrier.

[0023] These compositions may optionally include anti-inflammatory agents, anti-VEGF agents, immunosuppressants, anticoagulants, antiplatelet agents, and thrombolytic agents.

[0024] The present invention also includes compositions for treating ocular inflammatory conditions, including, but not limited to, uveitis, posterior uveitis, macular edema, acute macular degeneration, wet age-related macular edema, retinal detachment, retinal vein occlusion, ocular tumors, fungal infections, viral infections, multifocal choroiditis, diabetic uveitis, diabetic macular edema, diabetic retinopathy, proliferative vitreoretinopathy, sympathetic ophthalmia, Vogt-Koyanagi-Harada syndrome, histoplasmosis, and uveal ulcers. These compositions may optionally include anti-inflammatory agents, anti-VEGF agents, immunosuppressants, anticoagulants, antiplatelet agents, and thrombolytic agents.

[0025] The present invention also includes compositions for treating posterior ocular diseases such as, but not limited to, uveitis, posterior uveitis, wet age-related macular edema, diabetic macular edema, diabetic retinopathy, and retinal vein occlusion. These compositions may optionally include anti-inflammatory agents, anti-VEGF agents, immunosuppressants, anticoagulants, antiplatelet agents, and thrombolytic agents.

[0026] It will be understood that the present invention relates to compounds of structural formula I as described herein, as well as pharmaceutically acceptable salts of compounds of structural formula I, and also to salts that are not pharmaceutically acceptable when used as precursors to the free compounds or pharmaceutically acceptable salts thereof or in other synthetic procedures.

[0027] The compounds of the present invention can be administered in the form of pharmaceutically acceptable salts. The term "pharmaceutically acceptable salts" refers to salts prepared from pharmaceutically acceptable non-toxic bases or acids, including inorganic or organic bases and inorganic or organic acids. Salts of basic compounds encompassed by the term "pharmaceutically acceptable salts" generally refer to non-toxic salts of the compounds of the present invention, prepared by reacting the free base with a suitable organic or inorganic acid. Representative salts of basic compounds of the present invention include acetate, ascorbate, adipate, alginate, aspirate, benzenesulfonate, benzoate, bicarbonate, bisulfate, bitartrate, borate, bromide, butyrate, camphorate, camphorsulfonate, camsylate, carbonate, chloride, clavulanate, citrate, cyclopentanepropionate, diethylacetate, digluconate, dihydrochloride, dodecylsulfate, and the like. Dodecylsulfanate, edetate, edisylate, estolate, esylate, ethanesulfonate, formate, fumarate, gluceptate, glucoheptanoate, gluconate, glutamate, glycerophosphate, glycollylarsanilate, hemisulfate, heptanoate, hexanoate, hexylresorcinol, hydrabamine, hydrobromide, hydrochloride, 2-hydroxybenzoate Ethanesulfonate, hydroxynaphthoate, iodide, isonicotinic acid, isothionate, lactate, lactobionate, laurate, malate, maleate, mandelate, mesylate, methyl bromide, methyl nitrate, methyl sulfate, methanesulfonate, mucate, 2-naphthalenesulfonate, napsylate, nicotinate, nitrate, N-methylglucamine ammonium salt, oleate, oxalate, pamoate (embonate), palmitate Acid salts include, but are not limited to, acetate, pantothenate, pectinate, persulfate, phosphate / diphosphate, pimelate, phenylpropionate, polygalacturonate, propionate, salicylate, stearate, sulfate, subacetate, succinate, tannate, tartrate, teoclate, thiocyanate, tosylate, triethiodide, trifluoroacetate, undeconate, valerate, and the like.Additionally, when a compound of the present invention possesses an acidic moiety, suitable pharmaceutically acceptable salts thereof include, but are not limited to, salts derived from inorganic bases, including aluminum, ammonium, calcium, copper, ferric, ferrous, lithium, magnesium, manganic, manganous, potassium, sodium, zinc, and the like. Salts derived from pharmaceutically acceptable organic non-toxic bases include salts of primary, secondary, and tertiary amines, cyclic amines, dicyclohexylamines, and basic ion exchange resins, such as arginine, betaine, caffeine, choline, N,N-dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine, ethylamine, ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucamine, glucosamine, histidine, hydrabamine, isopropylamine, lysine, methylglucamine, morpholine, piperazine, piperidine, polyamine resins, procaine, purines, theobromine, triethylamine, trimethylamine, tripropylamine, tromethamine, and the like. Basic nitrogen-containing groups may be quaternized with agents such as lower alkyl halides, e.g., methyl, ethyl, propyl, and butyl chlorides, bromides, and iodides; dialkyls such as dimethyl, diethyl, dibutyl sulfates; and diamyl sulfates; long chain halides, e.g., decyl, lauryl, myristyl, and stearyl chlorides, bromides, and iodides; aralkyl halides such as benzyl and phenethyl bromide; and others.

[0028] These salts can be obtained by known methods, for example, by mixing the compound of the present invention with a solution containing an equivalent amount of the desired acid, base, etc., and then filtering the salt or distilling off the solvent to recover the desired salt. The compound of the present invention and its salt may form a solvate with a solvent such as water, ethanol, or glycerol. The compound of the present invention can simultaneously form an acid addition salt and a salt with a base, depending on the type of substituent on the side chain.

[0029] When the compounds of formula I contain both acidic and basic groups in the molecule, the present invention includes, in addition to the salt forms described above, also inner salts or betaines (zwitterions).

[0030] The present invention encompasses all stereoisomeric forms of the compounds of Formula I. Unless a specific stereochemistry is indicated, the present invention is meant to encompass all such isomers of these compounds. All asymmetric centers present in compounds of Formula I may, independently of one another, have either the (R) or (S) configuration. When a bond to a chiral carbon is depicted as a straight line in a structural formula of the present invention, it is understood that both the (R) and (S) configurations of that chiral carbon, and thus both individual enantiomers and mixtures thereof, are encompassed by the formula. When a specific configuration is depicted, that enantiomer (either (R) or (S) at that center) is intended. Similarly, when a compound name is given without a chiral designation of a chiral carbon, it is understood that both the (R) and (S) configurations of that chiral carbon, and thus individual enantiomers and mixtures thereof, are encompassed by the name. The preparation of a specific stereoisomer or mixture thereof may be specified in the examples in which such stereoisomer or mixture was obtained, but this in no way limits the inclusion of all stereoisomers and mixtures thereof within the scope of the present invention.

[0031] Unless a specific enantiomer or diastereomer is indicated, the present invention includes all possible enantiomers and diastereomers, as well as mixtures of two or more stereoisomers, e.g., mixtures of enantiomers and / or diastereomers, in all ratios. Accordingly, enantiomers are the subject of the present invention in enantiomerically pure form, both as levorotatory and dextrorotatory antipodes, in the form of racemates, and in the form of mixtures of the two enantiomers in all ratios. In the case of cis / trans isomerism, the present invention includes both the cis and trans forms, as well as mixtures of these forms in all ratios. The preparation of individual stereoisomers can be carried out, if desired, by conventional methods, such as separation of mixtures by chromatography or crystallization, the use of stereochemically uniform starting materials for synthesis, or stereoselective synthesis. Optionally, derivatization can be carried out before the separation of stereoisomers. Separation of stereoisomeric mixtures can be carried out at an intermediate stage during the synthesis of compounds of formula I or on the final racemate. Absolute stereochemistry can be determined by X-ray crystallography of crystalline products or crystalline intermediates which are derivatized, if necessary, with a reagent containing a stereocenter of known configuration. Where compounds of the invention can tautomerize, all individual tautomers and mixtures thereof are included within the scope of the invention. The invention includes all such isomers, as well as salts, solvates (including hydrates), and solvated salts of such racemates, enantiomers, diastereomers, and tautomers, and mixtures thereof.

[0032] In the compounds of the present invention, atoms may exhibit natural isotopic abundance, or one or more atoms may be artificially enriched in a particular isotope having the same atomic number but an atomic mass or mass number different from that predominantly found in nature. The present invention is intended to include all suitable isotopic variations of the specifically and generically described compounds. For example, different isotopic forms of hydrogen (H) include protium (1 H ) and deuterium (2 H) is the predominant hydrogen isotope found in nature. Enrichment with deuterium may confer certain therapeutic advantages, such as increased in vivo half-life or reduced dosage requirements, or may provide compounds useful as standards for characterization of biological samples. Isotopically enriched compounds can be prepared without undue experimentation by conventional techniques known to those of skill in the art, or by processes similar to those described in the general schemes and examples herein using appropriate isotopically enriching reagents and / or intermediates.

[0033] In any component, any variable (e.g., R x When a group (such as aryl, aryl, aryl) occurs more than once, its definition at each occurrence is independent at all other occurrences. Also, combinations of substituents and variables are permissible only if such combinations result in stable compounds. Lines drawn from substituents into ring systems indicate that the indicated bond may be attached to any of the substitutable ring atoms. When the ring system is bicyclic, it is intended that the bond be attached to any of the suitable atoms on either ring of the bicyclic moiety.

[0034] It is clear to those skilled in the art that by incorporating one or more silicon (Si) atoms in place of one or more carbon atoms into the compounds of the present invention, it is possible to obtain chemically stable compounds that can be easily synthesized from readily available raw materials using techniques known in the art. Carbon and silicon have different covalent bond radii, which result in differences in bond distances and steric configurations compared to the analogous C-element and Si-element bonds. These differences result in slight variations in the size and shape of silicon-containing compounds compared to carbon. Those skilled in the art will understand that differences in size and shape can lead to minor or significant changes in potency, solubility, lack of targeted activity, packaging characteristics, etc. (Diass, JO et al. Organometallics (2006) 5:1188-1198; Showell, GA et al. Bioorganic & Medicinal Chemistry Letters (2006) 16:2555-2558).

[0035] It will be apparent that one skilled in the art can select substituents and substitution patterns on the compounds of the present invention to provide compounds that are chemically stable and can be readily synthesized from readily available starting materials by techniques known in the art and the methods described below. It will be apparent that when a substituent is itself substituted with multiple groups, those multiple groups can be on the same carbon or on different carbons, so long as a stable structure results. It will also be understood that the phrase "optionally substituted" (with one or more substituents) means that the group in question may be unsubstituted or substituted with one or more substituents.

[0036] Furthermore, compounds of the present invention may exist in amorphous form and / or one or more crystalline forms, and all such amorphous and crystalline forms of the compounds of Formula I, as well as mixtures thereof, are intended to be included within the scope of the present invention. Furthermore, some compounds of the present invention may form solvates with water (i.e., hydrates) or common organic solvents. Such solvates and hydrates, particularly pharmaceutically acceptable solvates and hydrates of the compounds of the present invention, are included within the scope of the present invention, as are unsolvated and anhydrous forms.

[0037] Furthermore, when a carboxylic acid (—COOH) or alcohol group is present in a compound of the invention, pharmaceutically acceptable esters of the carboxylic acid derivative, such as methyl, ethyl, or pivaloyloxymethyl, or acyl derivatives of the alcohol, such as O-acetyl, O-pivaloyl, O-benzoyl, and O-aminoacyl, can also be used. These include ester and acyl groups known in the art for altering solubility or hydrolysis characteristics for use as sustained-release or prodrug formulations.

[0038] Pharmaceutically acceptable prodrug modifications of the compounds of the invention, which are converted in vivo to compounds within the scope of the invention, are also within the scope of the invention. For example, esters may be formed by esterification of available carboxylic acid groups or by ester formation at available hydroxy groups in the compounds. Similarly, unstable amides may be formed. Pharmaceutically acceptable esters or amides of the compounds of the invention may be hydrolyzed in vivo to give acid (or -COO, depending on the pH of the fluid or tissue where the conversion occurs). - ) or hydroxy form and are therefore within the scope of the present invention. Examples of pharmaceutically acceptable prodrug modifications include -C 1-6 -C substituted with alkyl esters and phenyl esters 1-6 Examples include, but are not limited to, alkyl.

[0039] Thus, compounds within the scope of the general structural formulae, embodiments and specific compounds described and claimed herein include, unless otherwise indicated, salts, all possible stereoisomers and tautomers, physical forms (e.g., amorphous and crystalline), solvates and hydrate forms, and any combination thereof, as well as salts thereof, prodrugs thereof, and salts of the prodrugs thereof, where such forms are possible.

[0040] Except as otherwise noted herein, the terms "alkyl" and "alkylene" are intended to include both branched and straight-chain saturated aliphatic hydrocarbon groups having the specified number of carbon atoms. Commonly used abbreviations for alkyl groups are used throughout this specification, e.g., methyl is represented by conventional abbreviations such as "Me" or CH3, or by symbols that are extended bonds without a defined end group, e.g., [ka] Ethyl can be represented by "Et" or CH2CH3, propyl can be represented by "Pr" or CH2CH2CH3, butyl can be represented by "Bu" or CH2CH2CH2CH3, etc. For example, "C 1-4 "Alkyl" (or "C1-C4 alkyl") means a straight or branched chain alkyl group having the specified number of carbon atoms, including all isomers. For example, the structure: [ka] have equivalent meanings. C 1-4 Alkyl includes n-, iso-, sec- and t-butyl, n- and isopropyl, ethyl and methyl, etc. If no number is specified, 1 to 4 carbon atoms are intended for straight or branched chain alkyl groups.

[0041] Unless otherwise noted, the term "cycloalkyl" means a mono- or bicyclic saturated aliphatic hydrocarbon group having the specified number of carbon atoms. For example, "cycloalkyl" includes cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and the like.

[0042] Unless otherwise noted, the term "aryl," as used herein, refers to a stable monocyclic or bicyclic ring system of up to 10 carbon atoms in each ring, where at least one ring is aromatic. Bicyclic aryl ring systems include fused ring systems in which two rings share two atoms, and spiro ring systems in which two rings share one atom. Aryl groups within this definition include, but are not limited to, phenyl, indene, isoindene, naphthalene, and tetralin.

[0043] As used herein, except where otherwise indicated, the term "heteroaryl" refers to a stable monocyclic or bicyclic ring system of up to 10 atoms in each ring, in which at least one ring is aromatic and at least one ring contains 1 to 4 heteroatoms selected from the group consisting of O, N, and S. Bicyclic heterocyclic ring systems include fused ring systems in which two rings share two atoms, and spiro ring systems in which two rings share one atom. Heteroaryl groups within the scope of this definition include azaindolyl, benzimidazolyl, benzisoxazolyl, benzofuranyl, benzofurazanyl, benzopyrazolyl, benzotriazolyl, benzothiophenyl, benzoxazolyl, carbazolyl, carbolinyl, cinnolinyl, dihydroindenyl, furanyl, indolinyl, indolyl, indolazinyl, indazolyl, isobenzofuranyl, isoindolyl, isoquinolyl, isothiazolyl, isoxazolyl, naphthalenyl, naphthopyridinyl, oxadiazolyl, oxazolyl, oxazoline, isoxazoline, pyranyl, pyrazinyl, pyrazolyl, pyrazolopyrimidinyl, pyridazinyl, pyridopyridinyl, pyridyl, pyrimidinyl, pyrrolyl, quinazoline, and the like. Heteroaryls include, but are not limited to, linyl, quinolyl, quinoxalinyl, tetrazolyl, tetrazolopyridyl, thiadiazolyl, thiazolyl, thienyl, triazolyl, dihydrobenzimidazolyl, dihydrobenzofuranyl, dihydrobenzothiophenyl, dihydrobenzoxazolyl, dihydroindolyl, dihydroquinolinyl, dihydrobenzodioxinyl, dihydropyrazolooxazinyl, dihydropyrazoliothiazinedioxidyl, methylenedioxybenzene, benzothiazolyl, benzothienyl, quinolinyl, isoquinolinyl, oxazolyl, tetrahydroquinoline, sulfonyl, 1,3-benzodioxolyl, and 3-oxo-3,4-dihydro-2N-benzo[b][1,4]thiazine. Where heteroaryl contains a nitrogen atom, it is understood that the corresponding N-oxide thereof is also encompassed by this definition.

[0044] The term "heterocycle" or "heterocyclyl," as used herein, unless otherwise specified, is intended to mean a stable non-aromatic monocyclic or bicyclic ring system having up to 10 atoms in each ring, including one to four heteroatoms selected from the group consisting of O, N, S, SO, or SO. Bicyclic heterocyclic ring systems include fused ring systems in which two rings share two atoms, and spiro ring systems in which two rings share one atom. Thus, "heterocyclyl" includes, but is not limited to, the following: azaspirononanyl, azaspirooctanyl, azetidinyl, dioxanyl, oxadiazaspirodecenyl, oxaspirooctanyl, oxazolidinonyl, piperazinyl, piperidinyl, pyrrolidinyl, morpholinyl, thiomorpholinyl, tetrahydrofurnayl, tetrahydropyranyl, dihydropiperidinyl, tetrahydrothiophenyl, and the like. If the heterocycle contains a nitrogen, it is understood that the corresponding N-oxide thereof is also encompassed by this definition.

[0045] Except where otherwise noted, the term "halogen" or "halo" means fluorine, chlorine, bromine, or iodine.

[0046] "Celite®" (Fluka) diatomaceous earth is diatomaceous earth and can be referred to as "celite."

[0047] Except as noted herein, structures containing substituent variables such as the variable "R" below: [ka] (which is depicted as not being attached to a specific bicyclic ring carbon atom) represents a structure in which that variable can be optionally attached to any bicyclic ring carbon atom. For example, the variable R shown in the structure above can be attached to any one of the six bicyclic ring carbon atoms i, ii, iii, iv, v, or vi.

[0048] Except as otherwise stated herein, bicyclic ring systems include fused ring systems in which two rings share two atoms, and spiro ring systems in which two rings share one atom.

[0049] The present invention also relates to medicinal products comprising at least one compound of formula I and / or a pharmaceutically acceptable salt of a compound of formula I and / or any stereoisomeric form of a compound of formula I or of a pharmaceutically acceptable salt of a compound of formula I, together with pharmaceutically suitable and pharmaceutically acceptable vehicles, additives and / or other active substances and adjuvants.

[0050] As used herein, the term "patient" is intended to mean mammals such as primates, humans, sheep, horses, cows, pigs, dogs, cats, rats, and mice.

[0051] The pharmaceutical preparations according to the invention can be administered orally, by inhalation, rectally or transdermally, or by subcutaneous, intraarticular, intraperitoneal or intravenous injection. Oral administration is preferred. Coating of stents and other surfaces in the body that come into contact with blood with the compounds of formula I is possible.

[0052] The present invention also relates to a method for producing a medicament, which comprises bringing at least one compound of formula I into a suitable administration form using a pharmaceutically suitable and pharmaceutically acceptable carrier and optionally further suitable active substances, additives or auxiliaries.

[0053] Suitable solid or galenical preparations include, for example, granules, powders, coated tablets, tablets, (micro)capsules, suppositories, syrups, juices, suspensions, emulsions, drops or injectable solutions and preparations with extended release of active substances, in the preparation of which common excipients such as vehicles, disintegrants, binders, coating agents, swelling agents, propellants and lubricants, flavorings, sweeteners and solubilizers are used. Frequently used adjuvants that may be mentioned are magnesium carbonate, titanium dioxide, lactose, mannitol and other sugars, talc, lactose, gelatin, starch, cellulose and its derivatives, animal and vegetable oils such as cod liver oil, sunflower oil, peanut oil or sesame oil, polyethylene glycol and solvents such as sterile water and mono- or polyhydric alcohols such as glycerin.

[0054] The administration regimen using a plasma kallikrein inhibitor is selected depending on a variety of factors, such as the type of patient, species, age, weight, sex, and medical condition of the patient; the severity of the condition being treated; the route of administration; the renal and hepatic function of the patient; and the particular compound or salt thereof being used. A physician or veterinarian of ordinary skill can readily determine and prescribe the effective amount of the drug required to prevent, eliminate, or arrest the progress of the condition.

[0055] When used for the indicated effects, oral dosages of plasma kallikrein inhibitors range from about 0.01 mg / kg body weight per day (mg / kg / day) to about 30 mg / kg / day, preferably 0.025 to 7.5 mg / kg / day, more preferably 0.1 to 2.5 mg / kg / day, and most preferably 0.1 to 0.5 mg / kg / day (amounts of active ingredient are on a free base basis unless otherwise specified). For example, an 80 kg patient would receive between about 0.8 mg / day and 2.4 g / day, preferably 2 to 600 mg / day, more preferably 8 to 200 mg / day, and most preferably 8 to 40 mg / kg / day. Thus, a pharmaceutical preparation suitable for once-daily administration would contain between 0.8 mg and 2.4 g, preferably 2 mg to 600 mg, more preferably 8 mg to 200 mg, and most preferably 8 mg to 40 mg, e.g., 8 mg, 10 mg, 20 mg, and 40 mg. Advantageously, the plasma kallikrein inhibitor may be administered in divided doses two, three or four times daily. For twice daily administration, a suitable prepared medicament would contain from 0.4 mg to 4 g, preferably from 1 mg to 300 mg, more preferably from 4 mg to 100 mg, most preferably from 4 mg to 20 mg, for example, 4 mg, 5 mg, 10 mg and 20 mg.

[0056] For intravenous administration, a patient will receive an amount of active ingredient sufficient to deliver 0.025-7.5 mg / kg / day, preferably 0.1-2.5 mg / kg / day, and more preferably 0.1-0.5 mg / kg / day. Such amounts can be administered in a number of suitable ways, e.g., a single, long-term, or several times daily dose of a low-concentration active ingredient, or a short-term, e.g., once daily, dose of a high-concentration active ingredient. Typically, conventional intravenous formulations containing active ingredient at concentrations of about 0.01-1.0 mg / mL, e.g., 0.1 mg / mL, 0.3 mg / mL, and 0.6 mg / mL, can be prepared and administered at daily doses of 0.01 mL / kg and 10.0 mL / kg patient body weight, e.g., 0.1 mL / kg, 0.2 mL / kg, and 0.5 mL / kg. In one example, an 80 kg patient receiving an 8 mL twice-daily intravenous formulation having an active ingredient concentration of 0.5 mg / mL would receive 8 mg of active ingredient per day. Glucuronic acid, L-lactic acid, acetic acid, citric acid, or any pharmaceutically acceptable acid / conjugate base with adequate buffering capacity in the pH range acceptable for intravenous administration can be used as a buffer. Selection of an appropriate buffer and formulation pH is within the skill of one in the art, depending on the solubility of the drug being administered.

[0057] The compounds of Formula I can be administered both as monotherapy and in combination with other therapeutic agents, including, but not limited to, anti-inflammatory agents, anti-VEGF agents, immunosuppressants, anticoagulants, antiplatelet agents, and thrombolytic agents.

[0058] An "anti-inflammatory agent" is an agent that, when administered at therapeutically effective levels, is effective in reducing inflammation, either directly or indirectly. "Anti-inflammatory agents" include, but are not limited to, steroidal anti-inflammatory agents and glucocorticoids. Suitable anti-inflammatory agents include, but are not limited to, cortisone, dexamethasone, hydrocortisone, methylprednisolone, prednisolone, prednisone, and triamcinolone.

[0059] An "anti-VEGF agent" is an agent that is effective directly or indirectly in inhibiting the activity of VEGF (vascular endothelial growth factor). Suitable anti-VEGF agents include, but are not limited to, bevacizumab, ranibizumab, and aflibercept.

[0060] An "immunosuppressant" is a drug that has the effect of directly or indirectly suppressing or reducing the strength of the body's immune system. Suitable immunosuppressants include, but are not limited to, corticosteroids (e.g., prednisone, budesonide, prednisolone), Janus kinase inhibitors (e.g., tofacitinib), calcineurin inhibitors (e.g., cyclosporine, tacrolimus), mTOR inhibitors (e.g., sirolimus, everolimus), IMDH inhibitors (e.g., azathioprine, leflunomide, mycophenolate), biologics (e.g., abatacept, adalimumab, anakinra, certolizumab, etanercept, golimumab, infliximab, ixekizumab, natalizumab, rituximab, secukinumab, tocilizumab, ustekinumab, vedolizumab), and monoclonal antibodies (e.g., basiliximab, daclizumab).

[0061] Suitable anticoagulants include, but are not limited to, factor XIa inhibitors, thrombin inhibitors, thrombin receptor antagonists, factor VIIa inhibitors, factor Xa inhibitors, factor IXa inhibitors, factor XIIa inhibitors, adenosine diphosphate antiplatelet agents (e.g., P2Y12 antagonists), fibrinogen receptor antagonists (e.g., for treating or preventing unstable angina or preventing re-occlusion after angioplasty and restenosis), other anticoagulants such as aspirin, and thrombolytic agents such as plasminogen activators or streptokinase for synergistic effects in the treatment of various vascular diseases. Examples of such anticoagulants include apixaban, dabigatran, cangrelor, ticagrelor, vorapaxar, clopidogrel, edoxaban, mipomersen, prasugrel, rivaroxaban, and semuloparin. For example, patients suffering from coronary artery disease and those who have undergone angioplasty may benefit from the co-administration of a fibrinogen receptor antagonist and a thrombin inhibitor.

[0062] In certain embodiments, the anti-inflammatory agents, anti-VEGF agents, immunosuppressants, anticoagulants, antiplatelet agents, and thrombolytic agents described herein are used within their conventional dosage ranges and administration regimens as reported in the art, e.g., the dosages set forth in the Physicians' Desk Reference, e.g., 70th Edition (2016) and earlier editions. In other embodiments, the anti-inflammatory agents, anti-VEGF agents, immunosuppressants, anticoagulants, antiplatelet agents, and thrombolytic agents described herein are used at doses lower than their conventional dosage ranges.

[0063] Alternatively or additionally, one or more other pharmacologically active agents can be co-administered with the compounds of the present invention. The term "other active agent" refers to a pharmaceutically active agent (or agent) that is different from the compounds of the present invention and is active in the body, such as a prodrug that is converted to a pharmaceutically active form after administration, and includes the free acid, free base, and pharmaceutically acceptable salts of the other active agent, if such formulations are commercially available or otherwise chemically feasible. Generally, any suitable other active substance (or active substances), such as, but not limited to, an antihypertensive drug, another diuretic, an antiatherosclerotic drug, a lipid-modifying agent, an antidiabetic drug, and / or an antiobesity drug, can be used in any combination with the compounds of the present invention in a single formulation (fixed-dose combination), or can be administered to a patient in one or more separate formulations that allow for simultaneous or sequential administration of the active substances (co-administration of separate active substances). Examples of other active substances that can be used include angiotensin-converting enzyme inhibitors (e.g., alacepril, benazepril, captopril, ceronapril, cilazapril, delapril, enalapril, enalaprilat, fosinopril, imidapril, lisinopril, moveltipril, perindopril, quinapril, ramipril, spirapril, temocapril or trandolapril);angiotensin II receptor antagonists, also known as angiotensin receptor blockers or ARBs, which can be in free base, free acid, salt, or prodrug form, such as azilsartan, e.g., azilsartan medoxomil potassium (EDARBI®), candesartan, e.g., candesartan cilexetil (ATACAND®), eprosartan, e.g., eprosartan mesylate (TEVETAN®), irbesartan (AVAPRO®), losartan, e.g., losartan potassium (COZAAR®), olmesartan, e.g., olmesartan medoximil (BENICAR®), telmesartan (MICARDIS®), valsartan (DIOVAN®), and hydrochlorothiazide (e.g., HYZAAR®, DIOVAN HCT®, ATACAND®). Any of these agents used in combination with a thiazide-like diuretic, such as HCTZ®; potassium-sparing diuretics, such as amiloride HCl, spironolactone, epleranone, and triamterene, each in combination with or without HCTZ; neutral endopeptidase inhibitors (e.g., thiorphan and phosphoramidon); aldosterone antagonists; aldosterone synthase inhibitors; renin inhibitors; enalclein; RO 42-5892; A 65317; CP 80794; ES 1005; ES 8891; SQ 34017; aliskiren (2(S),4(S),5(S),7(S)-N-(2-carbamoyl-2-methylpropyl)-5-amino-4-hydroxy-2,7-diisopropyl-8-[4-methoxy-3-(3-methoxypropoxy)-phenyl]-octanamide hemifumarate) SPP600, SPP630, and SPP635; endothelin receptor antagonists, vasodilators (e.g., nitroprusside); calcium channel blockers (e.g., amlodipine, nifedipine, verapamil, diltiazem, felodipine, gallopamil, niludipine, nimodipine, nicardipine); potassium channel activators (e.g., nicorandil, pinacidil, cromakalim, minoxidil, aprilkalim, loprazolam); sympatholytics;beta-adrenergic blocking agents (e.g., acebutolol, atenolol, betaxolol, bisoprolol, carvedilol, metoprolol, metoprolol tartrate, nadolol, propranolol, sotalol, timolol); alpha-adrenergic blocking agents (e.g., doxazosin, prazosin, or alpha-methyldopa); central alpha-adrenergic agonists; peripheral vasodilators (e.g., hydralazine); lipid-lowering agents, such as HMG-CoA reductase inhibitors, e.g., simvastatin and lovastatin (which are sold in lactone prodrug form as ZOCOR® and MEVACOR®, and function as inhibitors after administration), as well as atorvastatin (particularly the calcium salt sold under the trademark LIPITOR®), rosuvastatin (particularly the calcium salt sold under the trademark CRESTOR®), pharmaceutically acceptable salts of dihydroxy open-acid HMG-CoA reductase inhibitors such as prazolam (especially the calcium salt sold under the trademark PRAVACHOL®), pravastatin (especially the sodium salt sold under the trademark LESCOL®), and fluvastatin (especially the sodium salt sold under the trademark LESCOL®); cholesterol absorption inhibitors, for example ezetimibe (ZETIA®), and any other lipid-lowering agent such as the HMG-CoA reductase inhibitors mentioned above, in particular ezetimibe in combination with simvastatin (VYTORIN®) or atorvastatin calcium; immediate-release or sustained-release niacin, in particular niacin in combination with a DP antagonist and / or HMG-CoA reductase inhibitor, such as laropiprant; niacin receptor agonists, for example acipimox and acifran, and niacin receptor partial agonists;Insulin sensitizers and related compounds for the treatment of diabetes, such as biguanides (e.g., metformin), meglitinides (e.g., repaglinide, nateglinide), sulfonylureas (e.g., chlorpropamide, glimepiride, glipizide, glyburide, tolazamide, tolbutamide), thiazolidinediones, also known as glitazones (e.g., pioglitazone, rosiglitazone), alpha-glucosidase inhibitors (e.g., acarbose, miglitol), dipeptidyl peptidase inhibitors (e.g., sitagliptin (JANUVIA®), alogliptin, vildagliptin, saxagliptin, linagliptin, dutogliptin, gemigliptin), metabolic modifiers including ergot alkaloids (e.g., bromocriptine), JANUMET® (including metformin), inhibitors of glucose uptake, for example, sodium-glucose transporter (SGLT) inhibitors and their various isoforms, such as SGLT-1, SGLT-2 (e.g., ASP-1941, TS-071, BI-10773, tofogliflozin, LX-4211, canagliflozin, dapagliflozin, ertugliflozin, ipragliflozin, remogliflozin, and sotagliflozin), and SGLT-3; or other drugs useful in the prevention or treatment of the above diseases, such as, but not limited to, diazoxide; and chemically possible free acid, free base, and pharmaceutically acceptable salt forms, prodrug forms, such as esters, and prodrug salts, of the above pharmaceuticals, but not limited to these. The trade names of the above pharmaceutical agents are provided as examples of the marketed forms of the active agents, and it is contemplated that such agents may be used in separate formulations for simultaneous or sequential administration with a compound of the invention, or the active agents in the formulations may be used in fixed-dose pharmaceutical combinations with a compound of the invention;

[0064] A typical dose of a plasma kallikrein inhibitor of the present invention in combination with other suitable agents may be the same as the dose of the plasma kallikrein inhibitor administered without co-administration of additional agents, or may be significantly less than the dose of the plasma kallikrein inhibitor administered without co-administration of additional agents, depending on the therapeutic needs of the patient.

[0065] The compound is administered to a mammal in a therapeutically effective amount. By "therapeutically effective amount" is meant an amount of a compound of the invention that, when administered alone or in combination with an additional therapeutic agent to a mammal, is effective to treat (i.e., prevent, inhibit, or ameliorate) a disease state or treat the progression of a disease in the host.

[0066] The compounds of the present invention are preferably administered alone to mammals in a therapeutically effective amount. However, the compounds of the present invention can also be administered to mammals in a therapeutically effective amount in combination with an additional therapeutic agent, as defined below. When administered in combination, the combination of compounds is preferably, but not necessarily, a synergistic combination. Synergy occurs when the effect of the compounds when administered in combination (in this case, inhibition of the desired target) is greater than the additive effect of each compound when administered individually as a single agent, as described, for example, by Chou and Talalay, Adv. Enzyme Regul. 1984, 22, 27-55. Generally, synergy is most clearly demonstrated at suboptimal concentrations of the compounds. Synergy can be in terms of lower cytotoxicity, increased anticoagulant effect of the combination compared to the individual components, or some other beneficial effect.

[0067] "Combined administration" or "combined therapy" means that a compound of the present invention and one or more additional therapeutic agents are administered simultaneously to a mammal being treated. When administered concurrently, each component may be administered simultaneously or sequentially at different times and in any order. Thus, each component may be administered separately but sufficiently close in time to provide the desired therapeutic effect. Administration of each component need not be by the same route of administration; for example, one component can be administered orally and another component can be administered intravitreally.

[0068] The present invention is not to be limited in scope by the specific embodiments disclosed in the examples which are intended as illustrative of some aspects of the invention, and all embodiments which are functionally equivalent are intended to be within the scope of the invention. Indeed, various modifications of the invention in addition to those shown and described herein will become apparent to those skilled in the relevant arts and are intended to be encompassed within the scope of the appended claims.

[0069] general law The compounds of the invention can be made using conventional techniques or according to the methods outlined in the following general synthetic schemes. Those skilled in the art can vary the procedures and reagents shown to obtain analogous intermediates and / or final compounds.

[0070] NMR spectra were measured using a VARIAN or Bruker NMR system (400, 500, or 600 MHz). Chemical shifts are reported in ppm downfield and upfield from tetramethylsilane (TMS) and are relative to internal TMS or solvent resonances ( 1 H NMR: δ 7.27 for CDCl3, δ 2.50 for (CD3)(CHD2)SO, and 13C NMR: δ 77.02 for CDCl3 and δ 39.51 for (CD3)2SO. Coupling constants (J) are expressed in hertz (Hz), and spin multiplicities are indicated as s (singlet), d (doublet), dd (double doublet), t (triplet), m (multiplet), and br (broad). Chiral separations were performed on a Waters Thar 80 SFC or Berger MG II preparative SFC system. LC-MS data were recorded on a SHIMADAZU LC-MS-2020, SHIMADAZU LC-MS-2010, Agilent 1100 series LC-MS, Agilent Prime-1260, or Waters Acquity LC-MS instrument using a C18 column with a MeCN / water gradient containing 0.02 to 0.1% TFA. UV detection was performed at 220 nm and 254 nm, and ESI ionization was used for MS detection.

[0071] When chiral resolution was performed by chromatography using a chiral column, the chiral columns used for the SFC chiral resolution are listed in the table. Some of the chiral columns used were CHIRALPAK AD, CHIRALCEL OJ, CHIRALPAK AS, CHIRALPAK AY, CHIRALPAK IA, CHIRALPAK AD-H, and CHIRALPAK AS-H. They will be referred to hereafter by their two- or three-letter abbreviations. By convention, the fast-eluting isomer from a chiral resolution is always listed first in the table, followed immediately by the slower-eluting isomer from the same resolution. When more than two isomers are separated, they are always listed in the order of elution in the table: Peak 1, then Peak 2, Peak 3, etc. Near the chiral center in the structure * The symbol indicates that the chiral center was resolved by chiral resolution without its stereochemical configuration being unambiguously determined.

[0072] Also, TLC is thin layer chromatography; UV is ultraviolet; W is watts; wt. is weight percent; xg is times gravity; α Dis the specific rotation of polarized light at 589 nm; °C is degrees Celsius; %w / v is the percentage of the weight of the former agent to the volume of the latter agent; Hz is hertz; cpm is counts per minute; δ H is a chemical shift; d is a doublet; dd is a doublet of a doublet; MHz is megahertz; MS is a mass spectrum; a mass spectrum obtained by ES-MS may be referred to herein as "LC-MS"; m / z is mass / charge ratio; n is normal; N is normality; nm is nanometer; and nM is nanomolar concentration.

[0073] For purposes of this specification, the following abbreviations have the indicated meanings: [Mes-Acr-Me] + is 9-mesityl-10-methylacridinium tetrafluoroborate; and X-PHOS Pd G2 is chloro(2-dicyclohexylphosphino-2′,4′,6′-triisopropyl-1,1′-biphenyl)[2-(2′-amino-1,1′-biphenyl)]palladium(II).

[0074] Ac: Acetyl ACN: acetonitrile AcOH acetic acid aq.:Aqueous solution Boc or BOC: tert-butoxycarbonyl br:Wide Bu or n Bu: Butyl (normal) Bz: benzoyl ℃: Celsius calcd.: Calculated value δ: chemical shift d: double line DAST: (Diethylamino) sulfur trifluoride DBU: 1,8-diazabicyclo[5.4.0]undec-7-ene DCM: dichloromethane dd: Double line of double lines DIEA, DIPEA: N,N-diisopropylethylamine or Hunig's base DMF: dimethylformamide DMSO: dimethyl sulfoxide DPPA: Diphenylphosphoryl azide dqd: double line quad line double line DTT: dithiothreitol EDC: 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide EDTA: Ethylenediaminetetraacetic acid equiv: equivalent amount ESI: electrospray ionization Et: Ethyl Et2O: Diethyl ether EtOH: ethanol EtOAc: ethyl acetate g: grams GST: glutathione S-transferase h: time HATU:N,N,N',N'-Tetramethyl-O-(7-azabenzotriazol-1-yl)uronium hexafluorophosphate HPLC: High-performance liquid chromatography Hz: Hertz IPA: Isopropanol i Pr: Isopropyl J: Coupling constant LC: liquid chromatography LCMS: Liquid Chromatography Mass Spectrometry LED: Light-emitting diode m: multiplet M: Molar concentration Me: Methyl MeOH: Methanol mg: milligram MHz: Megahertz min:minutes μL: microliter mL: milliliter mmol: millimolar MS: Mass spectrometry MTBE: Methyl tert-butyl ether N: Nitrogen substituted nm: nanometer nM: nanomolar concentration NMP: 1-methylpyrrolidinone NMR: nuclear magnetic resonance spectroscopy OAc: acetate Ph: Phenyl Pr: Propyl q:Quarter rac: racemic RT or rt: room temperature (ambient temperature, approximately 25°C) s: Single line satd.:saturated SFC: Supercritical Fluid Chromatography t: Mie line T3P: Propylphosphonic anhydride TBAF: tert-butylammonium fluoride TBS or TBDMS: tert-butyldimethylsilyl TBSCl: tert-butyldimethylsilyl chloride t Bu: tert-butyl t BuOH: tert-butyl alcohol TCFH: tetramethylchloroformamidinium hexafluorophosphate TEA: Triethylamine (Et3N) tert: tertiary TFA: Trifluoroacetic acid THF: tetrahydrofuran TLC: Thin Layer Chromatography TMS: Trimethylsilyl TMSCl: chlorotrimethylsilane Tris: Tris(hydroxymethyl)aminomethane Ts: toluenesulfonyl (tolyl) tt: Triple line of triple lines X-phos or X-PHOS: 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl general law Starting materials used were obtained from commercial sources or prepared as described elsewhere unless otherwise noted. The methods used to prepare the compounds of the present invention are illustrated by the following schemes. Unless otherwise noted, all raw materials used are commercially available.

[0075] Chiral Separation Methods General preparative conditions for separating diastereomeric or enantiomeric mixtures of compounds using chiral SFC are as follows: [Table 1] General diagram Diagram A [ka] Scheme A depicts a synthetic procedure for the preparation of substituted spirocarbamates such as A6 from Boc-protected anilines A1 and ketones such as A2. Directed lithiation of aniline A1 and addition to heterocyclic ketone A2 is carried out in the presence of a Lewis acid (e.g., LaCl).

[0076] In situ cyclization of a tertiary alcohol onto the carbamate affords a spirocarbamate derivative such as A3, which can be subjected to chiral separation, preferably using supercritical fluid chromatography (SFC), to afford the enantiomers A4 and A5. Deprotection of either enantiomer (e.g., A4) affords the secondary amine A6.

[0077] Diagram B [ka] Scheme B depicts a synthetic procedure for the preparation of alkylhydrazines such as B4 from carbonyl derivatives such as B1. Condensation of the carbonyl B1 with a benzohydrazide gives intermediate B2, which is reduced to the protected hydrazine B3. Deprotection under acidic conditions gives the alkylhydrazine B4.

[0078] Diagram C [ka] Scheme C depicts a synthetic procedure for the preparation of CF3-ethyl hydrazine derivatives such as C4 from carbonyl derivatives such as C1. Condensation of carbonyl C1 with a benzohydrazide gives intermediate C2. Addition of CF3 to C2 with TMSCF3 gives the CF3-ethyl intermediate C3, which is deprotected under acidic conditions to give hydrazine derivative C4.

[0079] Diagram D [ka] Scheme D depicts the synthesis of alkylhydrazine derivatives such as D2 from alkyl halides such as D1. Alkylation of hydrazine with alkyl halide D1 gives alkylhydrazine D2.

[0080] Diagram E [ka] Scheme E depicts a synthetic procedure for the preparation of alkylhydrazine derivatives such as E3 from carboxylic acid derivatives such as E1. Photoredox decarboxylative hydrazide conversion of carboxylic acid E1 provides the protected hydrazine intermediate E2. Deprotection provides the alkylhydrazine derivative E3.

[0081] Diagram F [ka] Scheme F depicts a synthetic procedure for the preparation of alkylhydrazine derivatives such as F4 from alkylcarboxylic acid derivatives such as F1. Curtius rearrangement of alkylcarboxylic acid F1 gives protected amine F2. Oxidation of F2 gives N-nitroso intermediate F3, which is reduced and deprotected to give alkylhydrazine derivative F4.

[0082] Diagram G [ka] Scheme G depicts the synthesis of aryl hydrazines such as G2 from aryl bromides such as G1.

[0083] Aryl hydrazine G2 is prepared by palladium catalyzed cross-coupling reaction of an aryl bromide such as G1 with hydrazine in the presence of a suitable base.

[0084] Diagram H [ka] Scheme H depicts a synthetic procedure for the preparation of aminopyrazole derivatives such as H4 from hydrazine derivatives such as H2 and cyanoalkoxyacrylates such as H1. Condensation of substituted hydrazine H2 with cyanoethoxyacrylate H1 gives ester aminopyrazole derivatives such as H3. Saponification of H3 gives carboxylic acid H4.

[0085] Scheme I [ka] Scheme I depicts a synthetic procedure for the preparation of hydroxypyrazole derivatives, such as I4, from hydrazine derivatives, such as I2, and alkoxymethylene malonates, I1. Condensation of substituted hydrazine I2 with malonates, I1, affords ester hydroxypyrazole derivatives, such as I3. Saponification of I3 affords carboxylic acids, I4.

[0086] Diagram J [ka] Scheme J depicts a synthetic procedure for the preparation of N-substituted aminopyrazole derivatives, such as J4, from unsubstituted aminopyrazole derivatives, such as J1. Alkylation of aminopyrazole J1 provides N-substituted aminopyrazole J3. Saponification of J3 provides the N-substituted aminopyrazole carboxylic acid J4.

[0087] Diagram K [ka] Scheme K depicts the preparation of spirocarbamate pyrazole or triazole derivatives such as K1 from spirocarbamate piperidine derivatives such as K1 and carboxylic acid derivatives such as K2. Coupling of carboxylic acid K2 with spirocarbamate K1 using a peptide coupling agent such as TCFH, EDC, HATU, or T3P provides amide K3.

[0088] Diagram L [ka] Scheme L depicts a synthetic procedure for the preparation of spirocarbamate aminopyrazole derivatives such as L5 from spirocarbamate piperidine derivatives such as L1. Coupling of spirocarbamate L1 with cyanoacetic acid gives intermediate L2, which undergoes condensation to give cyanoacrylate derivatives such as L3. Condensation of L3 with various hydrazines L4 under basic conditions gives aminopyrazoles such as L5.

[0089] Intermediate A2-1 [ka] tert-Butyl 3-cyclopropyl-5-oxopiperidine-1-carboxylate Cyclopropylmagnesium bromide (30.0 mL, 15.2 mmol, 0.5 M solution in THF) was added to a suspension of CuI (1.45 g, 7.61 mmol) in THF (20 mL) at −78 °C under a N atmosphere. The resulting mixture was warmed to 0 °C and stirred for an additional 1 h to provide the organocuprate reagent. The solution was cooled to −78 °C, followed by the addition of a solution of tert-butyl 3-oxo-3,6-dihydropyridine-1(2H)-carboxylate (1.0 g, 5.1 mmol) and TMSCl (1.30 mL, 10.1 mmol) in THF (5 mL). The reaction mixture was stirred at −78 °C for 1 h before being quenched with MeOH. The mixture was diluted with EtOAc and saturated aqueous NH4Cl, and the layers were separated. The organic layer was dried over MgSO4, filtered, and concentrated under reduced pressure. The crude residue was purified by silica gel chromatography to provide the title compound. 1 H NMR (600 MHz, CDCl3) δ 4.12 - 3.74 (m, 3H), 3.18 (br s, 1H), 2.66 (dd, J = 16.3, 4.6 Hz, 1H), 2.33 (dd, J = 16.1, 10.5 Hz, 1H), 1.49 (s, 9H), 1.32 - 1.24 (m, 1H), 0.63 (tt, J = 8.7, 4.7 Hz, 1H), 0.55 (dd, J = 12.9, 5.3 Hz, 2H), 0.25 (s, 1H), 0.18 - 0.11 (m, 1H). Intermediate A2-2 [ka] tert-Butyl 3-(difluoromethyl)-5-oxopiperidine-1-carboxylate To a solution of tert-butyl 3-oxo-3,6-dihydropyridine-1(2H)-carboxylate (1.1 g, 5.6 mmol) and zinc difluoromethanesulfinate (2.5 g, 8.4 mmol) in trifluorotoluene (22 mL) and HO (9 mL) was added tert-butyl peroxide (1.70 mL, 12.6 mmol, 70% w / v solution in HO) dropwise at room temperature. The reaction mixture was heated to 35 °C and stirred for 12 h. The flask was then cooled to room temperature, diluted with HO and DCM, and the layers were separated. The combined organic layers were dried over MgSO, filtered, and concentrated under reduced pressure. The crude residue was purified by silica gel chromatography (EtOAc:hexanes) to provide the title compound. 1 H NMR (600 MHz, CDCl3) δ 5.80 (t, J = 55.5 Hz, 1H), 4.12 - 3.72 (m, 3H), 3.50 (br s, 1H), 2.63 (dd, J = 15.7, 5.3 Hz, 1H), 2.59 - 2.53 (m, 1H), 2.50 (dd, J = 15.6, 8.7 Hz, 1H), 1.47 (s, 9H). Intermediate A2-3 [ka] tert-Butyl 3-oxo-5-(trifluoromethyl)piperidine-1-carboxylate To a solution of tert-butyl 3-hydroxy-5-(trifluoromethyl)piperidine-1-carboxylate (8.0 g, 29.7 mmol) in DCM (50 mL) was added NaHCO (7.5 g, 89 mmol) and Dess-Martin periodinane (15.1 g, 35.7 mmol) at room temperature. The reaction mixture was stirred at room temperature for 1 h. It was then quenched with H2O and diluted with DCM. The layers were separated, and the aqueous phase was extracted with DCM. The combined organic layers were dried over MgSO, filtered, and concentrated under reduced pressure. The crude residue was purified by silica gel chromatography (EtOAc:hexanes) to provide the title compound. 1H NMR (600 MHz, CDCl3) δ 4.32 - 3.77 (m, 3H), 3.72 - 3.20 (m, 1H), 2.83 (d, J = 6.9 Hz, 1H), 2.73 (dd, J = 16.7, 6.1 Hz, 1H), 2.57 (dd, J = 16.6, 9.4 Hz, 1H), 1.47 (s, 9H). Intermediate A6-1 [ka] 6-chloro-5-fluoro-5',5'-dimethylspiro[benzo[d]1,3]oxazine-4,3'-piperidin]-2(1H)-one Step 1: tert-butyl 6-chloro-5-fluoro-5′,5′-dimethyl-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidine]-1′-carboxylate: To a round-bottom flask containing tert-butyl (4-chloro-3-fluorophenyl)carbamate (2.21 g, 9.0 mmol) under a N atmosphere was added THF (55 mL) and the solution was cooled to −78° C. To the stirred solution was added n BuLi (11.2 mL, 27.9 mmol, 2.5 M solution in hexane) was added over 40 min. The reaction mixture was stirred at -78 °C for an additional 45 min, at which point a solution of LaCl3·2LiCl (22.5 mL, 13.5 mmol, 0.6 M solution in THF) and tert-butyl 3,3-dimethyl-5-oxopiperidine-1-carboxylate (3.10 g, 13.5 mmol) was added to the reaction mixture over 40 min at -78 °C. The reaction mixture was allowed to warm to room temperature and stirred for 16 h. KO t Bu (5.3 mL, 9.0 mmol, 1.7 M solution in THF) was added to the reaction mixture and the solution was heated to 60° C. for an additional 3 h. The reaction was cooled to room temperature, quenched with 1 M HCl, and diluted with EtOAc. The layers were separated and the aqueous phase was extracted with EtOAc. The combined organic layers were dried over MgSO4, filtered, and concentrated under reduced pressure. The crude residue was purified by silica gel chromatography (EtOAc:Hexanes) to give the title compound. LCMS [M+Na] + =421.1 (calculated value 421.1).

[0090] Step 2: 6-chloro-5-fluoro-5′,5′-dimethylspiro[benzo[d][1,3]oxazine-4,3′-piperidin]-2(1H)-oneHCl (25.0 mL, 100 mmol, 4 M solution in dioxane) was added to a round-bottom flask containing a suspension of tert-butyl 6-chloro-5-fluoro-5',5'-dimethyl-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3'-piperidine]-1'-carboxylate (7.98 g, 20.0 mmol) in 1,4-dioxane (30 mL). The reaction mixture was heated to 90°C and stirred vigorously for 12 hours. The reaction was cooled to room temperature and concentrated to give the crude title compound, which was used in the next step without further purification. LCMS [M+H] + =299.1 (calculated value 299.1).

[0091] Table A. Using the appropriate starting materials, the following compounds were prepared using procedures similar to those described for intermediate A6-1. [Table 2] TIFF0007787890000024.tif194170TIFF0007787890000025.tif126170 Intermediate A6-15 [ka] (rac)-tert-butyl (4R or S,5'R or S)-6-chloro-5-fluoro-5'-hydroxy-2-oxo-1,2-dihydrospiro[benzo[1,3]oxazine-4,3'-piperidine]-1'-carboxylate Step 1: tert-butyl 3-((tert-butyldimethylsilyl)oxy)-5-oxopiperidine-1-carboxylate A flask containing a solution of tert-butyl 3-hydroxy-5-oxopiperidine-1-carboxylate (22.4 g, 104 mmol) in DMF (224 mL) was cooled to 0 °C. Imidazole (21.2 g, 312 mmol) and TBSCl (18.8 g, 125 mmol) were added, and the reaction mixture was warmed to room temperature and stirred for 16 h. The reaction was quenched with H2O and extracted with MTBE. The layers were separated, and the combined organic layer was washed with brine. The organic layer was dried over Na2SO4, filtered, and concentrated to give a crude residue that was purified by silica gel chromatography (EtOAc:petroleum ether) to give the title compound. LCMS [M-55] + =274.3 (calculated value 274.2).

[0092] Step 2: (rac)-tert-butyl (4R or 5,5′R or S)-5′-((tert-butyldimethylsilyl)oxy)-6-chloro-5-fluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidine]-1′-carboxylate and butyl (4S or R,5′S or R)-5′-((tert-butyldimethylsilyl)oxy)-6-chloro-5-fluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidine]-1′-carboxylateTo a round-bottom flask containing tert-butyl (4-chloro-3-fluorophenyl)carbamate (12.5 g, 50.9 mmol) was added THF (188 mL) and the mixture was cooled to -78°C. n BuLi (63.1 mL, 158 mmol, 2.5 M solution in hexanes) was added over 1 h, followed by the addition of a solution of LaCl3·2LiCl (× 102 mL, 61.1 mmol, 0.6 M solution in THF) and tert-butyl 3-((tert-butyldimethylsilyl)oxy)-5-oxopiperidine-1-carboxylate (20.1 g, 61.1 mmol) over 5 min at −78 °C. The reaction mixture was stirred at −78 °C for 1 h and then warmed to room temperature for an additional 12 h. The reaction was quenched with saturated aqueous NH4Cl, poured into a flask containing ice, and stirred for 15 min. The mixture was diluted with EtOAc, the layers separated, and the combined organic layers were dried over MgSO4, filtered, and concentrated to give a crude residue that was purified by silica gel chromatography (EtOAc:petroleum ether) to give the title compound as a mixture of diastereomers. The diastereomers were separated by preparative reverse phase HPLC (ACN / water + 10 mM NH4HCO3). The first eluting diastereomer of the title compound was obtained: LCMS [M+Na] + = 523.3 (calculated 523.2). The later eluting diastereomer of the title compound was obtained: LCMS [M+Na] + =523.3 (calculated value 523.2).

[0093] Step 3: (rac)-(4R or S,5′R or S)-6-chloro-5-fluoro-5′-hydroxyspiro[benzo[d][1,3]oxazine-4,3′-piperidin]-2(1H)-one:A flask containing (rac)-tert-butyl (4S or R, 5′S or R)-5′-((tert-butyldimethylsilyl)oxy)-6-chloro-5-fluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidine]-1′-carboxylate (later-eluting peak from the previous step, 1.0 g, 2.0 mmol) in THF (20 mL) was cooled to 0 °C. TBAF (6.0 mL, 6.0 mmol) was added, and the reaction mixture was warmed to 40 °C for 12 h. The reaction was quenched with ice water and stirred for 15 min. The mixture was diluted with EtOAc, the layers were separated, and the combined organic layer was washed with brine. The organic layer was dried over MgSO, filtered, and concentrated under reduced pressure to give the crude product, which was used in the next step without further purification. HCl (0.9 mL, 3.5 mmol, 4 M solution in dioxane) was added to a vial containing a suspension of the crude product (34 mg, 0.090 mmol) in 1,4-dioxane (0.8 mL). The reaction mixture was vigorously stirred at room temperature for 12 hours and concentrated to give the crude title compound. The crude product was used in the next step without further purification. LCMS [M+H] + =287.0 (calculated value 287.1).

[0094] Intermediate A6-16 [ka] (rac)-(4R or S,5′R or S)-6-chloro-5,5′-difluorospiro[benzo[d][1,3]oxazine-4,3′-piperidin]-2(1H)-one Step 1: (rac)-tert-butyl (4R or S, 5R or S)-6-chloro-5,5′-difluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidine]-1′-carboxylate:A flask containing (rac)-tert-butyl (4S or R, 5′S or R)-5′-((tert-butyldimethylsilyl)oxy)-6-chloro-5-fluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidine]-1′-carboxylate (later eluting peak from Step 1 of Intermediate A6-15, 1.0 g, 2.0 mmol) in THF (20 mL) was cooled to 0 °C. TBAF (6.0 mL, 6.0 mmol, 1 M solution in THF) was added to the flask, and the reaction mixture was warmed to 40 °C for 12 h. The reaction was quenched with ice water and stirred for 15 min. The mixture was diluted with EtOAc, the layers were separated, and the combined organic layer was washed with brine. The organic layer was dried over MgSO, filtered, and concentrated under reduced pressure to give the crude product, which was carried forward without further purification. The crude alcohol (100 mg, 0.260 mmol) in DCM (5 mL) was cooled to −78° C. under a N atmosphere. DAST (625 mg, 0.390 mmol) in DCM (2 mL) was added dropwise and the reaction mixture was stirred at −78° C. for 1 h. The reaction mixture was directly purified by silica gel chromatography (EtOAc:DCM) to give the title compound. LCMS [M+H] + =411.3 (calculated value 411.1).

[0095] Step 2: (rac)-(4R or S,5′R or S)-6-chloro-5,5′-difluorospiro[benzo[d][1,3]oxazine-4,3′-piperidin]-2-one: HCl (0.6 mL, 2.3 mmol, 4 M solution in dioxane) was added to a vial containing (rac)-tert-butyl (4R or S,5'R or S)-6-chloro-5,5'-difluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3'-piperidine]-1'-carboxylate (45 mg, 0.12 mmol) in 1,4-dioxane (0.6 mL), and the reaction mixture was stirred at room temperature for 3 hours. The solvent was removed under reduced pressure to give the crude product, which was used in the next step without further purification. LCMS [M+H] + =289.1 (calculated value 289.1).

[0096] Intermediate A6-17 [ka] (4R and S,6'S)-6-chloro-5-fluoro-6'-methylspiro[benzo[d][1,3]oxazine-4,3'-piperidin]-2(1H)-one Step 1: Benzyl (4R and S,6′S)-6-chloro-5-fluoro-6′-methyl-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidine]-1′-carboxylate: THF (15 mL) was added to tert-butyl (4-chloro-3-fluorophenyl)carbamate (600 mg, 2.44 mmol) and cooled to -78°C. n BuLi (3.0 mL, 7.57 mmol, 2.5 M solution in hexane) was added over 40 min, and the resulting mixture was stirred at -78 °C for an additional 45 min. A solution of LaCl3·2LiCl (6.1 mL, 3.66 mmol, 0.6 M solution in THF) and benzyl (S)-2-methyl-5-oxopiperidine-1-carboxylate (900 mg, 3.66 mmol) was added over 40 min at -78 °C, and the reaction mixture was allowed to warm to room temperature and stirred for 16 h. t Bu (1.40 mL, 2.44 mmol, 1.7 M in THF) was added and the reaction was heated to 60° C. for an additional 3 h. The reaction was cooled to room temperature, quenched with 1 M HCl, and diluted with EtOAc. The layers were separated and the aqueous layer was extracted with EtOAc. The combined organic layers were dried over MgSO4, filtered, and concentrated to give a crude residue that was purified by silica gel chromatography (EtOAc:hexanes) to give the title compound as a mixture of diastereomers. LCMS [M+H] + =419.0 (calculated value 419.1).

[0097] Step 2: (4R and S,6′S)-6-chloro-5-fluoro-6′-methylspiro[benzo[d][1,3]oxazine-4,3′-piperidin]-2-one: HBr (3.9 mL, 71.6 mmol, 33 wt% solution in AcOH) was added to a vial containing benzyl (4R and S,6'S)-6-chloro-5-fluoro-6'-methyl-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3'-piperidine]-1'-carboxylate (600 mg, 1.43 mmol). The reaction mixture was heated to 80°C for 12 hours. The reaction was cooled to room temperature and concentrated to give the crude title compound, which was used in the next step without further purification. LCMS [M+H] + =285.1 (calculated value 285.1).

[0098] Intermediate B4-1 [ka] rac-(1,1,1-trifluorobutan-2-yl)hydrazine Step 1: N'-(1,1,1-trifluorobutan-2-ylidene)benzohydrazide: To a solution of 1,1,1-trifluorobutan-2-one (1.39 g, 11.0 mmol) in toluene (10 mL) was added benzohydrazide (1.50 g, 11.0 mmol) and the reaction mixture was heated to 110° C. for 18 hours. The reaction was cooled to room temperature, poured into water, and filtered. The solid was washed with water and dried to give the crude desired title compound. LCMS [M+H] + =245.4 (calculated value 245.1).

[0099] Step 2: N'-(1,1,1-trifluorobutan-2-yl)benzohydrazide A solution of N'-(1,1,1-trifluorobutan-2-ylidene)benzohydrazide (500 mg, 2.05 mmol) in THF (6 mL) was cooled to 0 °C and BH3·THF (4.09 mL, 4.09 mmol, 1.0 M in THF) was added dropwise. The reaction was warmed to room temperature and stirred for 14 h. The reaction was cooled again to 0 °C and quenched with MeOH. The solvent was removed under reduced pressure and dichloromethane was added. The resulting slurry was filtered to remove insoluble material, and the organic layer was washed with saturated aqueous NH4Cl, dried over MgSO4, filtered, and concentrated to give the title compound. LCMS [M+H] + =246.7 (calculated value 247.1).

[0100] Step 3: (1,1,1-trifluorobutan-2-yl)hydrazine To a solution of N'-(1,1,1-trifluorobutan-2-yl)benzohydrazide (274 mg, 1.11 mmol) in MeOH (3 mL) was added hydrogen chloride (1.48 mL, 17.8 mmol, 37% aqueous solution), and the resulting mixture was heated to 80°C for 16 h. The reaction was cooled to room temperature and concentrated under reduced pressure. EtOAc was added, and the precipitate was filtered and washed with EtOAc to give the title compound. 1 H NMR (400 MHz, CD3OD) δ 3.34-3.49 (m, 1H), 1.80 (dqd, J=14.7, 7.5, 4.4 Hz, 1H), 1.46-1.65 (m, 1H), 0.97-1.27 (m, 3H). Table B. Using appropriate starting materials and procedures similar to those described in Intermediate B4-1, the following compounds were prepared. [Table 3] Intermediate C4-1 [ka] (2,2,2-trifluoro-1-(1-fluorocyclopropyl)ethyl)hydrazine Step 1: N'-((1-fluorocyclopropyl)methylene)benzohydrazide: 1-Fluorocyclopropane-1-carbaldehyde (176 mg, 2.0 mmol) was added to a solution of benzhydrazide (272 mg, 2.0 mmol) in toluene (4 mL) and the reaction mixture was heated to 60° C. for 1 h. The reaction was cooled to room temperature and concentrated to give the crude product which was used in the next step without any purification. LCMS [M+H] + =207.1 (calculated value 207.1).

[0101] Step 2: N'-(2,2,2-trifluoro-1-(1-fluorocyclopropyl)ethyl)benzohydrazide Allyltrimethylsilane (0.48 mL, 3.0 mmol) and BF3·Et2O (0.37 mL, 3.0 mmol) were added, in that order, to a suspension of N'-((1-fluorocyclopropyl)methylene)benzohydrazide (412 mg, 2.0 mmol) in 1,2-dichloroethane (4.0 mL), and the mixture was heated to reflux for 5 min. The solvent was removed under reduced pressure, and the resulting residue was dissolved in DMF (4 mL). TMSCF3 (0.60 mL, 4.0 mmol) and NaOAc (660 mg, 8.0 mmol) were added, and the mixture was heated to 55 °C for 3 h. The reaction was cooled to room temperature, quenched with saturated aqueous Na2CO3, and stirred for an additional 5 min. The mixture was diluted with H2O and extracted with EtOAc. The combined organic layers were dried over MgSO4, filtered, and concentrated to give a crude residue which was purified by silica gel chromatography (EtOAc:hexanes) to give the title compound. LCMS [M+H] + =277.1 (calculated value 277.1).

[0102] Step 3: (2,2,2-trifluoro-1-(1-fluorocyclopropyl)ethyl)hydrazineTo a solution of N'-(2,2,2-trifluoro-1-(1-fluorocyclopropyl)ethyl)benzohydrazide (140 mg, 0.51 mmol) in MeOH (0.75 mL) was added HCl (0.7 mL, 8.1 mmol, 37% aqueous solution), and the resulting mixture was heated to 80°C for 16 h. The reaction was cooled to room temperature and concentrated to give the crude product, which was azeotroped with toluene to give the title compound, which was carried forward without purification. LCMS [M+H] + =173.1 (calculated value 173.1).

[0103] Table C. Using appropriate starting materials and procedures similar to those described in Intermediate C4-1, the following compounds were prepared. [Table 4] TIFF0007787890000033.tif53168 Intermediate D2-1 [ka] ((1-fluorocyclopropyl)methyl)hydrazine Hydrazine (0.33 mL, 0.33 mmol, 1.0 M solution in THF) was added to a vial containing 1-(bromomethyl)-1-fluorocyclopropane (50 mg, 0.33 mmol) in EtOH (0.3 mL), and the resulting mixture was heated to 70° C. for 16 h. The reaction was cooled to room temperature and concentrated to give the crude product, which was azeotroped with toluene to give the title compound, which was carried forward without purification. LCMS [M+H] + =105.1 (calculated value 105.1).

[0104] Table D. Using appropriate starting materials and procedures similar to those described in Intermediate D2-1, the following compounds were prepared. [Table 5] Intermediate E3-1 [ka] (1-(4-fluorophenyl)cyclopropyl)hydrazine Step 1: Di-tert-butyl 1-(1-(4-fluorophenyl)cyclopropyl)hydrazine-1,2-dicarboxylate : Acetonitrile (20 mL) was dissolved in 1-(4-fluorophenyl)cyclopropane-1-carboxylic acid (360 mg, 2.0 mmol) and [Mes-Acr-Me] + This was added to a vial containing the photocatalyst (16 mg, 0.04 mmol). The solution was degassed with N2 for 5 minutes. After degassing, DBU (0.06 mL, 0.4 mmol) and di-tert-butyl azodicarboxylate (576 mg, 2.5 mmol) were added in rapid succession. The vial was placed in front of a 450 nm blue LED (Merck photoreactor) and stirred for 12 hours. The solvent was removed under reduced pressure, and the crude mixture was purified by silica gel chromatography (EtOAc:hexanes) to give the title compound. LCMS [M-155] + =211.1 (calculated value 211.2).

[0105] Step 2: (1-(4-fluorophenyl)cyclopropyl)hydrazine HCl (6.6 mL, 26.2 mmol, 4.0 M solution in dioxane) was added to a vial containing di-tert-butyl 1-(1-(4-fluorophenyl)cyclopropyl)hydrazine-1,2-dicarboxylate (640 mg, 1.75 mmol) and the reaction mixture was stirred at room temperature for 16 hours. The solvent was removed under reduced pressure and the mixture was azeotroped with toluene to give the title compound, which was carried forward without purification. LCMS [M+H] + =167.0 (calculated value 167.1).

[0106] Table E. Using appropriate starting materials and procedures similar to those described in Intermediate E3-1, the following compounds were prepared. [Table 6] Intermediate F4-1 [ka] (1-(trifluoromethyl)cyclopropyl)hydrazine Step 1: Butyl (1-(trifluoromethyl)cyclopropyl)carbamate 1-(trifluoromethyl)cyclopropanecarboxylic acid (5.00 g, 32.4 mmol) tTo a solution in BuOH (5 mL) was added TEA (5.00 mL, 35.7 mmol) and diphenylphosphinyl azide (11.8 g, 48.7 mmol), and the resulting mixture was stirred at room temperature for 0.5 h and then heated to 100 °C for 15 h. The reaction was diluted with EtOAc and washed with 5% citric acid, saturated aqueous NaHCO, and brine. The organic layer was dried over NaSO, filtered, and concentrated to give the crude product, which was purified by silica gel chromatography (EtOAc:petroleum ether) to give the title compound. 1 H NMR (400 MHz, CDCl3) δ 4.99-5.12 (m, 1H), 1.45 (s, 9H), 1.26 (br s, 2H), 1.11 (br s, 2H). Step 2: Butyl nitroso(1-(trifluoromethyl)cyclopropyl)carbamate Nitrosyl tetrafluoroborate (78 mg, 0.67 mmol) was added portionwise to a solution of tert-butyl (1-(trifluoromethyl)cyclopropyl)carbamate (100 mg, 0.440 mmol) in pyridine (0.2 mL) and acetonitrile (2 mL) at −30° C. The solution was stirred at −30° C. for 30 min and then warmed to 0° C. for 2 h. The reaction was concentrated under reduced pressure to give the crude product, which was purified by preparative TLC (EtOAc:petroleum ether) to give the title compound. 1 H NMR (400 MHz, CDCl3) δ 1.69 (s, 9H), 1.57-1.62 (m, 4H). Step 3: (1-(trifluoromethyl)cyclopropyl)hydrazine tert-Butyl nitroso(1-(trifluoromethyl)cyclopropyl)carbamate (100 mg, 0.390 mmol) in MeOH (2 mL) was stirred at −78° C. for 30 minutes. HCl (0.32 mL, 3.9 mmol, 37% aqueous solution) and zinc (257 mg, 3.93 mmol) were added at −78° C., and the resulting mixture was stirred for 2 hours. The reaction was allowed to warm to room temperature, filtered, and concentrated to give the title compound, which was carried forward without purification. LCMS [M+H] + =141.0 (calculated value 141.1).

[0107] Intermediate G2-1 [ka] (2-(difluoromethoxy)phenyl)hydrazine THF (1 mL) and NaO t To a mixture of Bu (129 mg, 1.35 mmol) and X-PHOS Pd G2 (10.6 mg, 0.0130 mmol) was added. 1-Bromo-2-(difluoromethoxy)benzene (300 mg, 1.35 mmol) was added, and the mixture was stirred at room temperature for 10 minutes. Hydrazine (42 μL, 1.3 mmol) was added in one portion, and the vial was heated to 90 °C (preheated bath) and stirred for 12 hours. The reaction was cooled to room temperature, diluted with MeOH, and filtered. The filtrate was concentrated to dryness to give a crude mixture that was purified by reverse-phase preparative HPLC (C18 stationary phase, ACN / water with 0.1% TFA) to give the title compound. LCMS [M+H] + =175.0 (calculated value 175.1).

[0108] Table G. Using appropriate starting materials and procedures similar to those described in Intermediate G2-1, the following compounds were prepared. [Table 7] TIFF0007787890000041.tif30165 Intermediate H4-1 [ka] 5-Amino-1-((tetrahydro-2H-pyran-4-yl)methyl)-1H-pyrazole-4-carboxylic acid Step 1: Ethyl 5-amino-1-((tetrahydro-2H-pyran-4-yl)methyl)-1H-pyrazole-4-carboxylate : To a stirring solution of ethyl 5-amino-1H-pyrazole-4-carboxylate (100 mg, 0.650 mmol) in ACN (1 mL) was added sodium hydride (30.9 mg, 1.29 mmol) at 0 °C. After 1 h, 4-(bromomethyl)tetrahydro-2H-pyran (173 mg, 0.970 mmol) was added, and the resulting mixture was heated to 80 °C for 12 h. The reaction was cooled to room temperature and quenched with saturated aqueous NH4Cl. The aqueous phase was extracted with EtOAc, and the combined organic layers were dried over Na2SO4, filtered, and concentrated under reduced pressure. The crude product was purified by preparative reverse-phase HPLC (C18 stationary phase, ACN / water with 0.04% NH4OH) to give the title compound. LCMS [M+H] + =254.2 anhydrous (calculated 254.1).

[0109] Step 2: 5-amino-1-((tetrahydro-2H-pyran-4-yl)methyl)-1H-pyrazole-4-carboxylic acid To a solution of ethyl 5-amino-1-((tetrahydro-2H-pyran-4-yl)methyl)-1H-pyrazole-4-carboxylate (100 mg, 0.400 mmol) in EtOH (1 mL) and HO (0.2 mL) was added LiOH·HO (20 mg, 0.47 mmol), and the resulting mixture was heated to 60° C. for 12 h. The mixture was concentrated to give the crude product, which was used in the next step without purification. LCMS [M+H] + =226.0 (calculated value 226.1).

[0110] Table H. Using appropriate starting materials and procedures similar to those described in Intermediate H4-1, the following compounds were prepared. [Table 8] Intermediate I4-1 [ka] 5-Amino-1-(1-(trifluoromethyl)cyclopropyl)-1H-pyrazole-4-carboxylic acid Step 1: Ethyl 5-amino-1-(1-(trifluoromethyl)cyclopropyl)-1H-pyrazole-4-carboxylateA solution of (1-(trifluoromethyl)cyclopropyl)hydrazine (50 mg, 0.36 mmol), methyl 2-cyano-3-ethoxyacrylate (56 mg, 0.36 mmol), and DIEA (0.31 mL, 1.8 mmol) in EtOH (1 mL) was heated to 80° C. for 12 h. The reaction was cooled to room temperature and concentrated under reduced pressure to give the crude product, which was purified by preparative TLC (EtOAc:petroleum ether) to give the title compound. LCMS [M+H] + =264.1 (calculated value 264.1).

[0111] Step 2: 5-amino-1-(1-(trifluoromethyl)cyclopropyl)-1H-pyrazole-4-carboxylic acid To a solution of ethyl 5-amino-1-(1-(trifluoromethyl)cyclopropyl)-1H-pyrazole-4-carboxylate (50 mg, 0.20 mmol) in MeOH (2 mL) and HO (0.5 mL) was added LiOH·HO (42.1 mg, 1.0 mmol). The resulting mixture was heated to 70 °C for 5 h. The reaction was cooled to room temperature, concentrated, acidified to pH 2 with 1 M HCl, and extracted with EtOAc. The combined organic layers were dried over NaSO, filtered, and concentrated under reduced pressure to give the crude product, which was carried forward without further purification. LCMS [M+H] + =235.9 (calculated value 236.1).

[0112] Intermediate I4-2 [ka] Intermediate I4-2 was prepared following a procedure similar to that described above for intermediate I4-1. LCMS [M+H] + =210.1 (calculated value 210.0).

[0113] Intermediate L3-1 [ka] (R,E / Z)-2-(6-chloro-5-fluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidine]-1′-carbonyl)-3-ethoxyacrylonitrile Step 1: 3-(6-chloro-5-fluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidine]-1′-yl)-3-oxopropanenitrileDIEA (7.06 mL, 40.4 mmol) was added to a solution of 6-chloro-5-fluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidine]-1′-ium chloride (4.14 g, 13.5 mmol) and 2-cyanoacetic acid (1.26 g, 14.8 mmol) in EtOAc (41 mL) and DMF (6.2 mL). 1-Propaphonic anhydride (9.60 mL, 16.2 mmol) was added, and the resulting mixture was stirred at room temperature overnight. The reaction was quenched with H2O and extracted with EtOAc. The combined organic layers were washed with brine, dried over MgSO4, and concentrated to give the crude residue, which was purified by silica gel chromatography (((3:1) EtOH:EtOAc):hexanes) to give the title compound. LCMS [M+H] + =337.9 (calculated value 338.1).

[0114] Step 2: (R,E / Z)-2-(6-chloro-5-fluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidine]-1′-carbonyl)-3-ethoxyacrylonitrile ZnCl (anhydrous, 303 mg, 2.21 mmol) was added to 3-(6-chloro-5-fluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidine]-1′-yl)-3-oxopropanenitrile (2.50 g, 7.40 mmol) in triethyl orthoformate (18.5 mL, 111 mmol) and NMP (0.8 mL), and the reaction mixture was heated to 130° C. for 4 h. The reaction was cooled to room temperature, quenched with saturated aqueous NaHCO and extracted with EtOAc. The combined organic layers were dried over MgSO, filtered, and concentrated to give a crude residue that was purified by silica gel chromatography (EtOAc:hexanes) to give the title compound. LCMS [M+H] + =394.0 (calculated value 394.1).

[0115] Intermediate L3-2 [ka] (R)-2-(6-chloro-5-fluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidine]-1′-carbonyl)-3-hydroxyacrylonitrile THF (15 mL) was added to a mixture of (R)-3-(6-chloro-5-fluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidin]-1′-yl)-3-oxopropanenitrile (1.82 g, 4.43 mmol) and methyl formate (3.99 g, 66.5 mmol). The reaction mixture was sonicated to dissolve the solids, and then KO t A solution of Bu (14.2 mL, 14.2 mmol, 1M in THF) was added dropwise and the resulting mixture was stirred at room temperature for 12 h. The reaction was diluted with H2O, neutralized to pH 6 with 1M HCl, and extracted with EtOAc. The combined organic layers were dried over MgSO4, filtered, and concentrated to give the crude title compound, which was used in the next step without purification. LCMS [M+H] + =366.4 (calculated value 366.1).

[0116] Example 1 [ka] (R)-1'-(5-amino-1-(2,2,2-trifluoroethyl)-1H-pyrazole-4-carbonyl)-6-chloro-5-fluorospiro[benzo[d][1,3]oxazine-4,3'-piperidin]-2(1H)-one (R,E / Z)-2-(6-chloro-5-fluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidine]-1′-carbonyl)-3-ethoxyacrylonitrile (0.25 mL, 0.10 mmol, 0.4 M stock solution in EtOH) was added to a vial containing (2,2,2-trifluoroethyl)hydrazine (17 mg, 0.15 mmol). TEA (42 μL, 0.30 mmol) was added to the vial, and the mixture was heated to 70° C. for 12 h. The reaction was cooled to room temperature and directly purified by preparative reverse-phase HPLC (C18 stationary phase, ACN / water with 0.05% TFA) to provide the title compound. 1H NMR (500 MHz, CD3OD) δ 7.55 (s, 1H), 7.43 (t, J = 8.1 Hz, 1H), 6.74 (d, J = 8.6 Hz, 1H), 4.76 - 4.66 (m, 3H), 4.46 (d, J = 13.3 Hz, 1H), 3.53 (s, 1H), 3.24 - 3.16 (m, 1H), 2.52 (t, J = 11.5 Hz, 1H), 2.30 (d, J = 13.6 Hz, 1H), 2.17 (q, J = 10.4 Hz, 1H), 1.73 (d, J = 12.7 Hz, 1H).LCMS [M+H] + = 462.3 (calculated value 462.1). Example 2 [ka] (R)-1'-(5-amino-1-(2,2,2-trifluoroethyl)-1H-pyrazole-4-carbonyl)-6-chloro-5-fluoro-5',5'-dimethylspiro[benzo[d][1,3]oxazine-4,3'-piperidin]-2(1H)-one A 3 mL vial was charged with (R)-6-chloro-5-fluoro-5′,5′-dimethylspiro[benzo[d][1,3]oxazine-4,3′-piperidin]-2(1H)-one (67 mg, 0.20 mmol) and 5-amino-1-(2,2,2-trifluoroethyl)-1H-pyrazole-4-carboxylic acid (84 mg, 0.40 mmol) in DMF (0.2 mL). To this mixture was added TEA (0.14 mL, 1.0 mmol), followed by T3P (0.18 μL, 0.6 mmol, 50% w / v solution in DMF), and the resulting mixture was stirred at room temperature for 2 h. The reaction mixture was directly purified by preparative reverse-phase HPLC (C18 stationary phase, ACN / water + 0.05% HCOH) to provide the title compound. 1H NMR (500 MHz, CD3OD) δ 7.62 (s, 1H), 7.44 - 7.40 (m, 1H), 6.74 (dd, J = 8.7, 1.1 Hz, 1H), 4.79 -4.70 (m, 3H), 4.27 (d, J = 12.9 Hz, 1H), 3.73 (s, 1H), 2.97 (d, J = 12.4 Hz, 1H), 2.17 - 2.03 (m, 2H), 1.24 (s, 3H), 1.03 (s, 3H). LCMS [M+H] + = 490.4 (calculated value 490.1). Examples 3 and 4 [ka] (R)-1'-(5-amino-1-((R)-1,1,1-trifluorobutan-2-yl)-1H-pyrazole-4-carbonyl)-6-chloro-5-fluorospiro[benzo[d][1,3]oxazine-4,3'-piperidine]-2(1H)-one and (R)-1'-(5-amino-1-((S)-1,1,1-trifluorobutan-2-yl)-1H-pyrazole-4-carbonyl)-6-chloro-5-fluorospiro[benzo[d][1,3]oxazine-4,3'-piperidine]-2(1H)-one (R)-2-(6-chloro-5-fluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidine]-1′-carbonyl)-3-hydroxyacrylonitrile (0.25 mL, 0.01 mmol, 0.4 M stock solution in AcOH) was added to a vial containing R- and S-(1,1,1-trifluorobutan-2-yl)hydrazine (31 mg, 0.15 mmol), and the resulting mixture was heated to 80° C. for 12 hours. The crude mixture was purified directly by preparative reverse-phase HPLC (C18 stationary phase, ACN / water with 0.05% TFA) to afford a mixture of diastereomers of the title compound. The title compound was resolved by preparative chiral SFC using Method A. The faster-eluting isomer of the title compound was obtained (Example 3): 1H NMR (400 MHz, CD3OD) δ 7.57 (s, 1H), 7.41 (dd, J = 8.6, 7.8 Hz, 1H), 6.72 (dd, J = 8.7, 1.3 Hz, 1H), 4.74-4.81 (m, 1H), 4.68 (br d, J = 13.4 Hz, 1H), 4.45 (br d, J = 12.2 Hz, 1H), 3.43-3.58 (m, 1H), 3.05-3.25 (m, 1H), 2.42-2.58 (m, 1H), 2.24-2.34 (m, 2H), 2.11-2.22 (m, 1H), 1.95-2.07 (m, 1H), 1.72 (dt, J = 11.3, 2.2 Hz, 1H), 0.82 (t, J = 7.3 Hz, 3H). LCMS [M+H] + = 490.1 (calculated 490.1). The later eluting isomer of the title compound was obtained (Example 4). 1 H NMR (400 MHz, CD3OD) δ 7.57 (s, 1H), 7.41 (dd, J = 8.7, 7.7 Hz, 1H), 6.73 (dd, J = 8.7, 1.3 Hz, 1H), 4.76-4.80 (m, 1H), 4.68 (br d, J = 14.9 Hz, 1H), 4.46 (br d, J = 11.5 Hz, 1H), 3.41-3.62 (m, 1H), 3.02-3.25 (m, 1H), 2.45-2.60 (m, 1H), 2.22-2.36 (m, 2H), 2.10-2.22 (m, 1H), 2.00 (dqd, J = 14.1, 7.3, 3.9 Hz, 1H), 1.65-1.77 (m, 1H), 0.80-0.88 (m, 3H). LCMS [M+H] + = 490.2 (calculated value 490.1). Table 1. Following procedures similar to those described for Examples 1-4, the following compounds were prepared using the appropriate starting materials. [Table 9] TIFF0007787890000052.tif210165TIFF0007787890000053.tif210165TIFF0007787890000054.tif209164TIFF0007787890000055. tif198164TIFF0007787890000056.tif210164TIFF0007787890000057.tif211165TIFF0007787890000058.tif210164TIFF00077878 90000059.tif209164TIFF0007787890000060.tif210164TIFF0007787890000061.tif211165TIFF0007787890000062.tif210164TIF F0007787890000063.tif210165TIFF0007787890000064.tif210164TIFF0007787890000065.tif210165TIFF0007787890000066.tif 209165TIFF0007787890000067.tif187166TIFF0007787890000068.tif210165TIFF0007787890000069.tif210165TIFF00077878900 00070.tif188164TIFF0007787890000071.tif209165TIFF0007787890000072.tif210165TIFF0007787890000073.tif209165TIFF00 07787890000074.tif210166TIFF0007787890000075.tif210165TIFF0007787890000076.tif210165TIFF0007787890000077.tif189165TIFF0007787890000078.tif209164TIFF0007787890000079.tif48166Table 2. The following compounds were prepared using appropriate starting materials following procedures similar to those described for Examples 3 and 4. The diastereomeric products were separated using the chiral SFC method specified in the table. For diastereomeric pairs, the faster-eluting isomer is listed first. [Table 10] TIFF0007787890000081.tif189167TIFF0007787890000082.tif187167TIFF0007787890000083.tif199166TIFF0007787890000084.tif199166TIFF0007787890000085.tif199167TIFF0007787890000086.tif199166Example 132 [ka] (R)-6-chloro-5-fluoro-1'-(1-(4-fluorobenzyl)-5-hydroxy-1H-pyrazole-4-carbonyl)spiro[benzo[d][1,3]oxazine-4,3'-piperidin]-2(1H)-one Step 1: Ethyl 1-(4-fluorobenzyl)-5-hydroxy-1H-pyrazole-4-carboxylate: To a solution of (4-fluorobenzyl)hydrazine hydrochloride (650 mg, 3.68 mmol) and diethyl 2-(ethoxymethylene)malonate (875 mg, 4.05 mmol) in HO (3 mL) was added KCO (1.27 g, 9.20 mmol), and the resulting mixture was heated to 100 °C for 3 h. The reaction was cooled to room temperature, and the mixture was washed with EtOAc. The aqueous phase was acidified to pH 2 with 1 M HCl and extracted with EtOAc. The organic layer was dried over NaSO, filtered, and concentrated under reduced pressure to give the crude title compound, which was used without further purification. LCMS [M+H] + =265.0 (calculated value 265.1).

[0117] Step 2: 1-(4-fluorobenzyl)-5-hydroxy-1H-pyrazole-4-carboxylic acid To a solution of ethyl 1-(4-fluorobenzyl)-5-hydroxy-1H-pyrazole-4-carboxylate (25 mg, 0.095 mmol) in EtOH (1 mL) and HO (0.2 mL) was added NaOH (38 mg, 0.95 mmol), and the resulting mixture was heated to 90 °C for 4 h. The reaction was cooled to room temperature and concentrated to give a crude residue that was acidified to pH 2 with 1 M HCl and extracted with EtOAc. The organic layer was dried over NaSO, filtered, and concentrated under reduced pressure to give the crude title compound, which was used in the next step without further purification. LCMS [M+H] + =237.0 (calculated value 237.1).

[0118] Step 3: (R)-6-chloro-5-fluoro-1′-(1-(4-fluorobenzyl)-5-hydroxy-1H-pyrazole-4-carbonyl)spiro[benzo[d][1,3]oxazine-4,3′-piperidin]-2(1H)-one To a solution of 1-(4-fluorobenzyl)-5-hydroxy-1H-pyrazole-4-carboxylic acid (20 mg, 0.085 mmol) and (R)-6-chloro-5-fluorospiro[benzo[d][1,3]oxazine-4,3′-piperidin]-2(1H)-one (31 mg, 0.085 mmol) in ACN (2 mL) was added TCFH (26 mg, 0.093 mmol) and 1-methylimidazole (21 mg, 0.25 mmol), and the resulting mixture was stirred at room temperature for 12 h. The reaction was concentrated to give a crude residue, which was purified by preparative reverse-phase HPLC (C18 stationary phase, ACN / water with 0.1% TFA) to give a mixture of diastereomers of the title compound. 1 H NMR (500 MHz, CD3OD) δ 7.91 (br s, 1H), 7.46 (t, J = 8.2 Hz, 1H), 7.32 (br s, 2H), 7.09 (br t, J = 8.5 Hz, 2H), 6.76 (br d, J = 8.4 Hz, 1H), 5.13 (br s, 2H), 4.36 (s, 2H), 3.19 (br s, 2H), 2.52 (br s, 1H), 2.31 (br d, J = 13.7 Hz, 1H), 2.17 (br d, J = 13.1 Hz, 1H), 1.75 (br d, J = 14.5 Hz, 1H).LCMS [M+H] + = 489.1 (calculated value 489.1). Example 133 [ka] (R)-1'-(5-amino-1-benzyl-1H-1,2,3-triazole-4-carbonyl)-6-chloro-5-fluorospiro[benzo[d][1,3]oxazine-4,3'-piperidin]-2(1H)-one A 4 mL vial was charged with (R)-6-chloro-5-fluoro-2-oxo-1,2-dihydrospiro[benzo[d][1,3]oxazine-4,3′-piperidin]-1-ium 2,2,2-trifluoroacetate (50 mg, 0.13 mmol) and 5-amino-1-benzyl-1H-1,2,3-triazole-4-carboxylic acid (28 mg, 0.13 mmol) in DMF (5 mL). DIEA (23 μL, 0.13 mmol) was added to the mixture, followed by HATU (49 mg, 0.13 mmol) in one portion, and the resulting mixture was stirred at room temperature for 2 h. The reaction mixture was directly purified by preparative reverse-phase HPLC (C18 stationary phase, ACN / water with 0.05% TFA) to provide the title compound. 1 H NMR (500 MHz, CD3OD) δ 7.91 (br s, 1H), 7.46 (t, J = 8.2 Hz, 1H), 7.32 (br s, 2H), 7.09 (br t, J = 8.5 Hz, 2H), 6.76 (br d, J = 8.4 Hz, 1H), 5.13 (br s, 2H), 4.36 (s, 2H), 3.19 (br s, 2H), 2.52 (br s, 1H), 2.31 (br d, J = 13.7 Hz, 1H), 2.17 (br d, J = 13.1 Hz, 1H), 1.75 (br d, J = 14.5 Hz, 1H).LCMS [M+H] + = 489.1 (calculated value 489.1). Factor XIa assay The efficacy of the compounds of the present invention as inhibitors of coagulation factor XIa can be determined using the relevant purified serine protease and an appropriate synthetic substrate. The rate of hydrolysis of a chromogenic or fluorogenic substrate by the relevant serine protease was measured both in the absence and presence of the compounds of the present invention. Assays were performed at room temperature or 37°C. Hydrolysis of the substrate resulted in the release of aminotrifluoromethylcoumarin (AFC), which was monitored spectrofluorimetrically by measuring the increase in emission at 510 nm upon excitation at 405 nm. A decrease in the rate of fluorescence change in the presence of an inhibitor indicates enzyme inhibition. Such methods are known to those skilled in the art. The results of this assay are reported as the half-maximal inhibitory concentration (IC 50 ), or the inhibition constant Ki.

[0119] Compounds were preincubated with human factor XIa (0.04 nM) in 50 mM HEPES buffer containing 150 mM sodium chloride, 5 mM calcium chloride, 0.1% PEG 8000, pH 7.4, for 30 minutes at 25°C. Factor XIa enzymatic activity was determined by adding the substrate glycine-proline-arginine-7-amido-4-trifluoromethylcoumarin (GPR-AFC) and measuring fluorescence at 400 / 505 nm after 60 minutes of incubation at 25°C. The percent inhibition of each data point was calculated from the data and analyzed using a four-parameter equation of log(inhibitor) versus response to determine the half-maximal inhibitory concentration (IC). 50 ) was calculated. IC 50 was converted to an equilibrium inhibition constant (Ki) using the Cheng-Prusoff equation.

[0120] Activity demonstrated by this assay indicates that the compounds of the invention may be therapeutically useful in the treatment or prevention of a variety of cardiovascular and / or cerebrovascular thromboembolic conditions in patients with unstable angina, acute coronary syndromes, refractory angina, myocardial infarction, transient ischemic attacks, atrial fibrillation, stroke such as thrombotic or embolic stroke, venous thrombosis, coronary and cerebral artery thrombosis, cerebral and pulmonary embolism, arteriosclerosis, deep vein thrombosis, disseminated intravascular coagulation, and reocclusion or restenosis of recanalized vessels.

[0121] Plasma kallikrein assay The efficacy of the compounds of the present invention as inhibitors of plasma kallikrein can be determined using the relevant purified serine protease and an appropriate synthetic substrate. The rate of hydrolysis of a chromogenic or fluorogenic substrate by the relevant serine protease was measured both in the absence and presence of the compounds of the present invention. Assays were performed at room temperature or 37°C. Hydrolysis of the substrate resulted in the release of aminotrifluoromethylcoumarin (AFC), which was monitored spectrofluorimetrically by measuring the increase in emission at 510 nm upon excitation at 405 nm. A decrease in the rate of fluorescence change in the presence of an inhibitor indicates enzyme inhibition. Such methods are known to those skilled in the art. The results of this assay are reported as half-maximal inhibitory concentrations (IC 50 ), or the inhibition constant Ki.

[0122] Plasma kallikrein measurements were performed in 50 mM HEPES buffer, pH 7.4, containing 150 mM NaCl, 5 mM CaCl, and 0.1% PEG 8000 (polyethylene glycol; Fisher Scientific). Measurements were performed using purified human plasma kallikrein (Enzyme Research Laboratories) at a final concentration of 0.5 nM and the synthetic substrate, acetyl-KPR-AFC (Sigma #C6608), at a concentration of 100 mM.

[0123] Activity measurements were performed by diluting the substrate stock solution at least 10-fold to a final concentration ≤0.2 Km and injecting it into a solution containing enzyme or enzyme equilibrated with inhibitor. The time required for equilibration between enzyme and inhibitor was determined in control experiments. Reactions were run under linear progress curve conditions, and the increase in fluorescence was measured at 405Ex / 510Em nm. Values ​​were converted to percent inhibition of the control reaction (after subtracting the 100% inhibition value). IC 50 was determined by the inflection point from a four-parameter logistic curve fit. Ki was calculated using the Cheng-Prusoff equation: Ki=IC 50 Calculated using / (1+([S] / Km)).

[0124] Activity demonstrated by this assay indicates that the compounds of the invention may be therapeutically useful for the treatment or prevention of a variety of ophthalmic, cardiovascular and / or cerebrovascular thromboembolic conditions in patients with unstable angina, acute coronary syndromes, refractory angina, myocardial infarction, transient ischemic attack, atrial fibrillation, stroke such as thrombotic or embolic stroke, venous thrombosis, coronary and cerebral artery thrombosis, cerebral and pulmonary embolism, arteriosclerosis, deep vein thrombosis, disseminated intravascular coagulation, and reocclusion or restenosis of recanalized vessels, hereditary angioedema, uveitis, posterior uveitis, wet age-related macular edema, diabetic macular edema, diabetic retinopathy and retinal vein occlusion.

[0125] Plasma kallikrein IC for selected compounds 50 (nM) and FXIa IC 50 (nM) is as follows: [Table 11] TIFF0007787890000090.tif21268TIFF0007787890000091.tif21267TIFF0007787890000092.tif21267TIFF0007787890 000093.tif21266TIFF0007787890000094.tif21266TIFF0007787890000095.tif21367TIFF0007787890000096.tif19572

Claims

1. A compound of the formula: or a pharmaceutically acceptable salt of said compound: 【Chemistry 1】 [In the formula, X is N or CH; R １ is a halo; R ２ is a halo; R ３ is selected from the group consisting of hydrogen and methyl; R ４ is selected from the group consisting of hydrogen and methyl; R ５ is NH2; Each R ６ are independently hydrogen, halo, hydroxy and C １－６ alkyl, said alkyl group being optionally substituted with 1 to 3 halo; Each R ７ are independently hydrogen, halo, hydroxy and C １－６ alkyl, said alkyl group being optionally substituted with 1 to 3 halo; or R ６ and R ７ may be taken together with the carbon atoms to which they are attached to form a 3- to 6-membered cycloalkyl group optionally substituted with 1 or 2 halo; R ８ is hydrogen; halo; hydroxy; R ｘ ; OR ｘ ; phenyl; indan; OR ｙ heteroaryl, which can be monocyclic or bicyclic; heterocycle; and C, which can be monocyclic or bicyclic ３－６ cycloalkyl; the phenyl and heteroaryl groups are independently selected from the group consisting of oxo, halo, R ｘ , OR ｘ , N.R. ９ R １０ , N.R. ９ (C=O)R ｘ , N.R. ９ (C=O)OR ｘ , (C=O)OR ｘ , (C═O)NR ９ , R ｙ and OR ｙ and wherein the cycloalkyl and heterocyclic groups are independently substituted with 1 to 3 substituents selected from the group consisting of oxo, halo, R ｘ and OR ｘ optionally substituted with 1 to 3 substituents selected from the group consisting of: R ９ is hydrogen or C １－３ is alkyl; R １０ is hydrogen or C １－３ is alkyl; R ｘ is hydrogen or C optionally substituted with 1 to 3 substituents selected from the group consisting of halo and hydroxy １－６ is alkyl, R ｙ is phenyl, a heterocycle or C ３－６ cycloalkyl, wherein the phenyl group is optionally substituted with 1 to 3 halo, the heterocyclic group is optionally substituted with 1 or 2 oxo, and the cycloalkyl group is C １－６ may be substituted with alkyl; n is an integer of 0 to 2.

2. 2. The compound of claim 1, wherein n is 0 or 1; or a pharmaceutically acceptable salt of said compound.

3. R ８ is phenyl; and said phenyl is independently halo, R ｘ , OR ｘ , N.R. ９ R １０ , N.R. ９ (C=O)R ｘ , N.R. ９ (C=O)OR ｘ , (C═O)NR ９ , (C=O)OR ｘ , R ｙ and OR ｙ The compound according to claim 1 or 2, which is optionally substituted with 1 to 3 substituents selected from the group consisting of:

4. R ８ is phenyl; and said phenyl is independently halo, R ｘ , OR ｘ , R ｙ and OR ｙ The compound according to any one of claims 1 to 3, which is optionally substituted with 1 to 3 substituents selected from the group consisting of: or a pharmaceutically acceptable salt of said compound.

5. n is 1; R ６ is hydrogen; R ７ The compound of any one of claims 1 to 4, wherein is hydrogen; or a pharmaceutically acceptable salt of said compound.

6. Compounds 1-133 2. The compound according to claim 1, selected from any one of the following:

7. A pharmaceutical composition comprising a compound according to any one of claims 1 to 6 or a pharmaceutically acceptable salt of said compound and a pharmaceutically acceptable carrier.

8. 10. Use of the composition of claim 7 in the manufacture of a medicament for a method of treating visual activity disorder, diabetic retinopathy, diabetic macular edema, retinal vein occlusion, hereditary angioedema, diabetes, pancreatitis, cerebral hemorrhage, nephropathy, cardiomyopathy, neuropathy, inflammatory bowel disease, arthritis, inflammation, septic shock, hypotension, cancer, adult respiratory distress syndrome, disseminated intravascular coagulation, blood clots during cardiopulmonary bypass surgery, or bleeding from post-operative surgery in a mammal, said method of treatment comprising administering the composition of claim 7 to a mammal in need of treatment.

9. Use of the composition of claim 7 in the manufacture of a medicament for a method of treating uveitis, posterior uveitis, wet age-related macular edema, diabetic macular edema, diabetic retinopathy or retinal vein occlusion in a mammal, the method of treatment comprising administering the composition of claim 7 to a mammal in need of treatment.

10. Use of the composition of claim 7 in the manufacture of a medicament for a method for treating diabetic retinopathy or diabetic macular edema in a mammal, the method comprising administering the composition of claim 7 to a mammal in need of treatment.

11. 10. Use of the composition of claim 7 in the manufacture of a medicament for a method of treating retinal vein occlusion in a mammal, the method of treatment comprising administering the composition of claim 7 to a mammal in need of treatment.

12. 10. The compound of any one of claims 1 to 6, or a pharmaceutically acceptable salt thereof, for use in the manufacture of a medicament for treating uveitis, posterior uveitis, wet age-related macular edema, diabetic macular edema, diabetic retinopathy or retinal vein occlusion in a mammal in need thereof.

13. 10. A compound according to any one of claims 1 to 6, or a pharmaceutically acceptable salt thereof, for use in therapy.

14. 8. The composition of claim 7, further comprising another agent selected from the group consisting of an anti-inflammatory agent, an anti-VEGF agent, an immunosuppressant, an anticoagulant, an antiplatelet agent, and a thrombolytic agent.

15. The use of claim 8, wherein the treatment method further comprises administering another drug selected from the group consisting of anti-inflammatory agents, anti-VEGF agents, immunosuppressants, anticoagulants, antiplatelet agents, and thrombolytic agents.

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