Patent Application
20220402950
PAD4 is a member of the peptidylarginine deiminase (PAD) family of enzymes capable of catalysing the citrullination of arginine into citrulline within peptide sequences. PAD4 is responsible for the deimination or citrullination of a variety of proteins in vitro and in vivo, with consequences of diverse functional responses in a variety of diseases (Jones J. E. et al, Curr. Opin. Drug Discov. Devel., 12(5), (2009), 616-627). Examples of exemplar diseases include rheumatoid arthritis, diseases with neutrophilic contributions to pathogenesis (for example vasculitis, systemic lupus erythematosus, ulcerative colitis) in addition to oncology indications. PAD4 inhibitors also have wider applicability as tools and therapeutics for human disease through epigenetic mechanisms.
Inhibitors of PAD4 have utility against Rheumatoid Arthritis (RA). RA is an auto-immune disease affecting approximately 1% of the population (Wegner N. et al, Immunol. Rev., 233(1) (2010), 34-54). It is characterized by inflammation of articular joints leading to debilitating destruction of bone and cartilage. A weak genetic association between PAD4 polymorphisms and susceptibility to RA has been suggested, albeit inconsistently, in a number of population studies (Kochi Y. et al, Ann. Rheum. Dis., 70, (2011), 512-515). PAD4 (along with family member PAD2) has been detected in synovial tissue where it is responsible for the deimination of a variety of joint proteins. This process is presumed to lead to a break of tolerance to, and initiation of immune responses to, citrullinated substrates such as fibrinogen, vimentin and collagen in RA joints. These anti-citrullinated protein antibodies (ACPA) contribute to disease pathogenesis and may also be used as a diagnostic test for RA (e.g. the commercially available CCP2 or cyclic citrullinated protein 2 test). In addition, increased citrullination may also offer additional direct contributions to disease pathogenesis through its ability to affect directly the function of several joint and inflammatory mediators (e.g. fibrinogen, anti-thrombin, multiple chemokines). In a smaller subset of RA patients, anti-PAD4 antibodies can be measured and may correlate with a more erosive form of the disease.
PAD4 inhibitors are also useful for the reduction of pathological neutrophil activity in a variety of diseases. Studies suggest that the process of Neutrophil Extracellular Trap (NET) formation, an innate defense mechanism by which neutrophils are able to immobilize and kill pathogens, is associated with histone citrullination and is deficient in PAD4 knockout mice (Neeli I. et at, J. Immunol., 180, (2008), 1895-1902 and Li P. et al, J. Exp. Med., 207(9), (2010), 1853-1862). PAD4 inhibitors may therefore have applicability for diseases where NET formation in tissues contributes to local injury and disease pathology. Such diseases include, but are not limited to, small vessel vasculitis (Kessenbrock K. et al, Nat. Med., 15(6), (2009), 623-625), systemic lupus erythematosus (Hakkim A. et al, Proc. Natl. Acad. Sci. USA, 107(21), (2010), 9813-9818 and Villanueva E. et al, J. Immunol., 187(1), (2011), 538-52), ulcerative colitis (Savchenko A. et al, Pathol. Int., 61(5), (2011), 290-7), cystic fibrosis, asthma (Dworski R. et al, 0.1. Allergy Clin. Immunol., 127(5), (2011), 1260-6), deep vein thrombosis (Fuchs T, et at, Proc. Natl. Acad. Sci. USA, 107(36), (2010), 15880-5), periodontitis (Vilkov L. et al, Ultrastructural Pathol., 34(1), (2010), 25-30), sepsis (Clark, S. R. et al. Nat. Med., 13(4), (2007), 463-9), appendicitis (Brinkmann V. et al, Science, 303, (2004), 1532-5), and stroke. In addition, there is evidence that NETs may contribute to pathology in diseases affecting the skin, e.g., in cutaneous lupus erythematosis (Villanueva E. et al, J. Immunol., 187(1), (2011), 538-52) and psoriasis (Lin A. M. et al., J. Immunol., 187(1), (2011), 490-500), so a PAD4 inhibitor may show benefit to tackle NET skin diseases, when administered by a systemic or cutaneous route. PAD4 inhibitors may affect additional functions within neutrophils and have wider applicability to neutrophilic diseases.
Studies have demonstrated efficacy of tool PAD inhibitors (for example chloro-amidine) in a number of animal models of disease, including collagen-induced arthritis (Willis VC. et al, J. Immunol., 186(7), (2011), 4396-4404), dextran sulfate sodium (DSS)-induced experimental colitis (Chumanevich A. A. et al, Am. J. Physiol. Gastrointest. Liver Physiol., 300(6), (2011), G929-G938), spinal cord repair (Lange S. et al, Dev. Biol., 355(2), (2011), 205-14), and experimental autoimmune encephalomyelitis (EAE). The DSS colitis report also demonstrates that chloro-amidine drives apoptosis of inflammatory cells both in vitro and in vivo, suggesting that PAD4 inhibitors may be effective more generally in widespread inflammatory diseases.
PAD4 inhibitors are also useful in the treatment of cancers (Slack J. L. et al, Cell. Mol. Life Sci., 68(4), (2011), 709-720). Over-expression of PAD4 has been demonstrated in numerous cancers (Chang X et al, BMC Cancer, 9, (2009), 40). An antiproliferative role has been suggested for PAD4 inhibitors from the observation that PAD4 citrullinates arginine residues in histones at the promoters of p53-target genes such as p21, which are involved in cell cycle arrest and induction of apoptosis (Li P. et al, Mol. Cell Biol., 28(15), (2008), 4745-4758).
The aforementioned role of PAD4 in deiminating arginine residues in histones may be indicative of a role for PAD4 in epigenetic regulation of gene expression. PAD4 is the primary PAD family member observed to be resident in the nucleus as well as the cytoplasm. Early evidence that PAD4 may act as a histone demethyliminase as well as a deiminase is inconsistent and unproven. However, it may reduce histone arginine methylation (and hence epigenetic regulation associated with this mark) indirectly via depletion of available arginine residues by conversion to citrulline. PAD4 inhibitors are useful as epigenetic tools or therapeutics for affecting expression of varied target genes in additional disease settings. Through such mechanisms, PAD4 inhibitors may also be effective in controlling citrullination levels in stem cells and may therefore therapeutically affect the pluripotency status and differentiation potential of diverse stem cells including, but not limited to, embryonic stem cells, neural stem cells, haematopoietic stem cells and cancer stem cells. Accordingly, there remains an unmet need to identify and develop PAD4 inhibitors for the treatment of PAD4-mediated disorders.
It has now been found that compounds of Formula (I) are useful as inhibitors of PAD4:
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L, R1, R2, R3, R4, R7, and R8, along with other variables is as defined herein.
In some embodiments, a provided compound demonstrates selectivity for PAD4 with respect to PAD2. The present invention also provides pharmaceutically acceptable compositions comprising a provided compound. Provided compounds are useful in treatment of various disorders associated with PAD4. Such disorders are described in detail, herein, and include, for example rheumatoid arthritis, vasculitis, systemic lupus erythematosus, ulcerative colitis, cancer, cystic fibrosis, asthma, cutaneous lupus erythematosis, and psoriasis.
In some embodiments, such compounds include those of the formulae described herein, or a pharmaceutically acceptable salt thereof, wherein each variable is as defined herein and described in embodiments. Such compounds have the structure of Formula (I):
or a pharmaceutically acceptable salt thereof, wherein:
cycloalkyl, benzo[d][1,2,3]triazol-5-yl, 6-acetamido-n-hex-1-yl, and (3-(hydroxymethyl)oxetan-3-yl)methyl.
Throughout the specification and the appended claims, a given chemical formula or name shall encompass all stereo and optical isomers and racemates thereof where such isomers exist. Unless otherwise indicated, all chiral (enantiomeric and diastereomeric) and racemic forms are within the scope of the invention. Many geometric isomers of C═C double bonds, C═N double bonds, ring systems, and the like can also be present in the compounds, and all such stable isomers are contemplated in the present invention. Cis- and trans- (or E- and Z-) geometric isomers of the compounds of the present invention are described and may be isolated as a mixture of isomers or as separated isomeric forms. The present compounds can be isolated in optically active or racemic forms. Optically active forms may be prepared by resolution of racemic forms or by synthesis from optically active starting materials. All processes used to prepare compounds of the present invention and intermediates made therein are considered to be part of the present invention. When enantiomeric or diastereomeric products are prepared, they may be separated by conventional methods, for example, by chromatography or fractional crystallization. Depending on the process conditions the end products of the present invention are obtained either in free (neutral) or salt form. Both the free form and the salts of these end products are within the scope of the invention. If so desired, one form of a compound may be converted into another form. A free base or acid may be converted into a salt; a salt may be converted into the free compound or another salt; a mixture of isomeric compounds of the present invention may be separated into the individual isomers. Compounds of the present invention, free form and salts thereof, may exist in multiple tautomeric forms, in which hydrogen atoms are transposed to other parts of the molecules and the chemical bonds between the atoms of the molecules are consequently rearranged. It should be understood that all tautomeric forms, insofar as they may exist, are included within the invention.
As used herein, the term “alkyl” or “alkylene” is intended to include both branched and straight-chain saturated aliphatic hydrocarbon groups having the specified number of carbon atoms. For examples, “C1 to C12 alkyl” or “C1-12 alkyl” (or alkylene), is intended to include C1, C2, C3, C4, C5, C6, C7, C8, C9, C10, C11 and C12 alkyl groups; “C4 to C18 alkyl” or “C4-18 alkyl” (or alkylene), is intended to include C4, C5, C6, C7, C8, C9, C10, C11, C12, C13, C14, C15, C16, C17, and C18 alkyl groups. Additionally, for example, “C1 to C6 alkyl” or “C1-6 alkyl” denotes alkyl having 1 to 6 carbon atoms. Alkyl group can be unsubstituted or substituted with at least one hydrogen being replaced by another chemical group. Example alkyl groups include, but are not limited to, methyl (Me), ethyl (Et), propyl (e.g., n-propyl and isopropyl), butyl (e.g., n-butyl, isobutyl, i-butyl), and pentyl (e.g., n-pentyl, isopentyl, neopentyl). When “C0 alkyl” or “C0 alkylene” is used, it is intended to denote a direct bond.
“Alkenyl” or “alkenylene” is intended to include hydrocarbon chains of either straight or branched configuration having the specified number of carbon atoms and one or more, preferably one to two, carbon-carbon double bonds that may occur in any stable point along the chain. For example, “C2 to C6 alkenyl” or “C2-6 alkenyl” (or alkenylene), is intended to include C2, C3, C4, C5, and C6 alkenyl groups. Examples of alkenyl include, but are not limited to, ethenyl, 1-propenyl, 2-propenyl, 2-butenyl, 3-butenyl, 2-pentenyl, 3, pentenyl, 4-pentenyl, 2-hexenyl, 3-hexenyl, 4-hexenyl, 5-hexenyl, 2-methyl-2-propenyl, and 4-methyl-3-pentenyl.
“Alkynyl” or “alkynylene” is intended to include hydrocarbon chains of either straight or branched configuration having one or more, preferably one to three, carbon-carbon triple bonds that may occur in any stable point along the chain. For example, “C2 to C6 alkynyl” or “C2-6 alkynyl” (or alkynylene), is intended to include C2, C3, C4, C5, and C6 alkynyl groups; such as ethynyl, propynyl, butynyl, pentynyl, and hexynyl.
The term “alkoxy” or “alkyloxy” refers to an —O-alkyl group. For example, “C1 to C6 alkoxy” or “C1-6 alkoxy” (or alkyloxy), is intended to include C1, C2, C3, C4, C5, and C6 alkoxy groups. Example alkoxy groups include, but are not limited to, methoxy, ethoxy, propoxy (e.g., n-propoxy and isopropoxy), and t-butoxy. Similarly, “alkylthio” or “thioalkoxy” represents an alkyl group as defined above with the indicated number of carbon atoms attached through a sulphur bridge; for example methyl-S— and ethyl-S—.
“Halo” or “halogen” includes fluoro, chloro, bromo, and iodo. “Haloalkyl” is intended to include both branched and straight-chain saturated aliphatic hydrocarbon groups having the specified number of carbon atoms, substituted with 1 or more halogens. Examples of haloalkyl include, but are not limited to, fluoromethyl, difluoromethyl, trifluoromethyl, trichloromethyl, pentafluoroethyl, pentachloroethyl, 2,2,2-trifluoroethyl, heptafluoropropyl, and heptachloropropyl. Examples of haloalkyl also include “fluoroalkyl” that is intended to include both branched and straight-chain saturated aliphatic hydrocarbon groups having the specified number of carbon atoms, substituted with 1 or more fluorine atoms.
The term “cycloalkyl” refers to cyclized alkyl groups, including mono-, bi- or poly-cyclic ring systems. For example, “C3 to C6 cycloalkyl” or “C3-6 cycloalkyl” is intended to include C3, C4, C5, and C6 cycloalkyl groups. Example cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and norbornyl. Branched cycloalkyl groups such as 1-methylcyclopropyl and 2-methylcyclopropyl are included in the definition of “cycloalkyl”. The term “cycloalkenyl” refers to cyclized alkenyl groups. C4-6 cycloalkenyl is intended to include C4, C5, and C6 cycloalkenyl groups. Example cycloalkenyl groups include, but are not limited to, cyclobutenyl, cyclopentenyl, and cyclohexenyl.
As used herein, “carbocycle”, “carbocyclyl”, or “carbocyclic residue” is intended to mean any stable 3-, 4-, 5-, 6-, 7-, or 8-membered monocyclic or bicyclic or 7-, 8-, 9-, 10-, 11-, 12-, or 13-membered bicyclic or tricyclic hydrocarbon ring, any of which may be saturated, partially unsaturated, unsaturated or aromatic. Examples of such carbocycles include, but are not limited to, cyclopropyl, cyclobutyl, cyclobutenyl, cyclopentyl, cyclopentenyl, cyclohexyl, cycloheptenyl, cycloheptyl, cycloheptenyl, adamantyl, cyclooctyl, cyclooctenyl, cyclooctadienyl, [3.3.0]bicyclooctane, [4.3.0]bicyclononane, [4.4.0]bicyclodecane (decalin), [2.2.2]bicyclooctane, fluorenyl, phenyl, naphthyl, indanyl, adamantyl, anthracenyl, and tetrahydronaphthyl (tetralin). As shown above, bridged rings are also included in the definition of carbocycle (e.g., [2.2.2]bicyclooctane). Preferred carbocycles, unless otherwise specified, are cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, phenyl, indanyl, and tetrahydronaphthyl. When the term “carbocycle” is used, it is intended to include “aryl.” A bridged ring occurs when one or more, preferably one to three, carbon atoms link two non-adjacent carbon atoms. Preferred bridges are one or two carbon atoms. It is noted that a bridge always converts a monocyclic ring into a tricyclic ring. When a ring is bridged, the substituents recited for the ring may also be present on the bridge.
As used herein, the term “bicyclic carbocycle” or “bicyclic carbocyclic group” is intended to mean a stable 9- or 10-membered carbocyclic ring system that contains two fused rings and consists of carbon atoms. Of the two fused rings, one ring is a benzo ring fused to a second ring; and the second ring is a 5- or 6-membered carbon ring which is saturated, partially unsaturated, or unsaturated. The bicyclic carbocyclic group may be attached to its pendant group at any carbon atom which results in a stable structure. The bicyclic carbocyclic group described herein may be substituted on any carbon if the resulting compound is stable. Examples of a bicyclic carbocyclic group are, but not limited to, naphthyl, 1,2-dihydronaphthyl, 1,2,3,4-tetrahydronaphthyl, and indanyl. “Carbocycle”, “carbocyclyl”, or “carbocyclic residue” can also refer to spiro compounds, for example, a spiro[3.3]heptane.
“Aryl” groups refer to monocyclic or bicyclic aromatic hydrocarbons, including, for example, phenyl, and naphthyl. Aryl moieties are well known and described, for example, in Lewis, R. J., ed., Hawley's Condensed Chemical Dictionary, 15th Edition, John Wiley & Sons, Inc., New York (2007). “C6-10 aryl” refers to phenyl and naphthyl.
As used herein, the term “heterocycle”, “heterocyclyl”, or “heterocyclic group” is intended to mean a stable 3-, 4-, 5-, 6-, or 7-membered monocyclic or bicyclic or 7-, 8-, 9-, 10-, 11-, 12-, 13-, or 14-membered polycyclic heterocyclic ring that is saturated, partially unsaturated, or fully unsaturated, and that contains carbon atoms and 1, 2, 3 or 4 heteroatoms independently selected from the group consisting of N, O and S; and including any polycyclic group in which any of the above-defined heterocyclic rings is fused to a benzene ring. The nitrogen and sulfur heteroatoms may optionally be oxidized (i.e., N→O and S(O)p, wherein p is 0, 1 or 2). The nitrogen atom may be substituted or unsubstituted (i.e., N or NR wherein R is H or another substituent, if defined). The heterocyclic ring may be attached to its pendant group at any heteroatom or carbon atom that results in a stable structure. The heterocyclic rings described herein may be substituted on carbon or on a nitrogen atom if the resulting compound is stable. A nitrogen in the heterocycle may optionally be quaternized. It is preferred that when the total number of S and O atoms in the heterocycle exceeds 1, then these heteroatoms are not adjacent to one another. It is preferred that the total number of S and O atoms in the heterocycle is not more than 1. When the term “heterocycle” is used, it is intended to include heteroaryl.
Examples of heterocycles include, but are not limited to, acridinyl, azetidinyl, azocinyl, benzimidazolyl, benzofuranyl, benzothiofuranyl, benzothiophenyl, benzoxazolyl, benzoxazolinyl, benzthiazolyl, benztriazolyl, benztetrazolyl, benzisoxazolyl, benzisothiazolyl, benzimidazolinyl, carbazolyl, 4aH-carbazolyl, carbolinyl, chromanyl, chromenyl, cinnolinyl, decahydroquinolinyl, 2H,6H-1,5,2-dithiazinyl, dihydrofuro[2,3-b]tetrahydrofuran, furanyl, furazanyl, imidazolidinyl, imidazolinyl, imidazolyl, 1H-indazolyl, imidazolopyridinyl, indolenyl, indolinyl, indolizinyl, indolyl, 3H-indolyl, isatinoyl, isobenzofuranyl, isochromanyl, isoindazolyl, isoindolinyl, isoindolyl, isoquinolinyl, isothiazolyl, isothiazolopyridinyl, isoxazolyl, isoxazolopyridinyl, methylenedioxyphenyl, morpholinyl, naphthyridinyl, octahydroisoquinolinyl, oxadiazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl, oxazolidinyl, oxazolyl, oxazolopyridinyl, oxazolidinylperimidinyl, oxindolyl, pyrimidinyl, phenanthridinyl, phenanthrolinyl, phenazinyl, phenothiazinyl, phenoxathiinyl, phenoxazinyl, phthalazinyl, piperazinyl, piperidinyl, piperidonyl, 4-piperidonyl, piperonyl, pteridinyl, purinyl, pyranyl, pyrazinyl, pyrazolidinyl, pyrazolinyl, pyrazolopyridinyl, pyrazolyl, pyridazinyl, pyridooxazolyl, pyridoimidazolyl, pyridothiazolyl, pyridinyl, pyrimidinyl, pyrrolidinyl, pyrrolinyl, 2-pyrrolidonyl, 2H-pyrrolyl, pyrrolyl, quinazolinyl, quinolinyl, 4H-quinolizinyl, quinoxalinyl, quinuclidinyl, tetrazolyl, tetrahydrofuranyl, tetrahydroisoquinolinyl, tetrahydroquinolinyl, 6H-1,2,5-thiadiazinyl, 1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl, 1,2,5-thiadiazolyl, 1,3,4-thiadiazolyl, thianthrenyl, thiazolyl, thienyl, thiazolopyridinyl, thienothiazolyl, thienooxazolyl, thienoimidazolyl, thiophenyl, triazinyl, 1,2,3-triazolyl, 1,2,4-triazolyl, 1,2,5-triazolyl, 1,3,4-triazolyl, and xanthenyl. Also included are fused ring and spiro compounds containing, for example, the above heterocycles.
Examples of 5- to 10-membered heterocycles include, but are not limited to, pyridinyl, furanyl, thienyl, pyrrolyl, pyrazolyl, pyrazinyl, piperazinyl, piperidinyl, imidazolyl, imidazolidinyl, indolyl, tetrazolyl, isoxazolyl, morpholinyl, oxazolyl, oxadiazolyl, oxazolidinyl, tetrahydrofuranyl, thiadiazinyl, thiadiazolyl, thiazolyl, triazinyl, triazolyl, benzimidazolyl, 1H-indazolyl, benzofuranyl, benzothiofuranyl, benztetrazolyl, benzotriazolyl, benzisoxazolyl, benzoxazolyl, oxindolyl, benzoxazolinyl, benzthiazolyl, benzisothiazolyl, isatinoyl, isoquinolinyl, octahydroisoquinolinyl, tetrahydroisoquinolinyl, tetrahydroquinolinyl, isoxazolopyridinyl, quinazolinyl, quinolinyl, isothiazolopyridinyl, thiazolopyridinyl, oxazolopyridinyl, imidazolopyridinyl, and pyrazolopyridinyl.
Examples of 5- to 6-membered heterocycles include, but are not limited to, pyridinyl, furanyl, thienyl, pyrrolyl, pyrazolyl, pyrazinyl, piperazinyl, piperidinyl, imidazolyl, imidazolidinyl, indolyl, tetrazolyl, isoxazolyl, morpholinyl, oxazolyl, oxadiazolyl, oxazolidinyl, tetrahydrofuranyl, thiadiazinyl, thiadiazolyl, thiazolyl, triazinyl, and triazolyl. Also included are fused ring and spiro compounds containing, for example, the above heterocycles.
As used herein, the term “bicyclic heterocycle” or “bicyclic heterocyclic group” is intended to mean a stable 9- or 10-membered heterocyclic ring system which contains two fused rings and consists of carbon atoms and 1, 2, 3, or 4 heteroatoms independently selected from the group consisting of N, O and S. Of the two fused rings, one ring is a 5- or 6-membered monocyclic aromatic ring comprising a 5-membered heteroaryl ring, a 6-membered heteroaryl ring or a benzo ring, each fused to a second ring. The second ring is a 5- or 6-membered monocyclic ring which is saturated, partially unsaturated, or unsaturated, and comprises a 5-membered heterocycle, a 6-membered heterocycle or a carbocycle (provided the first ring is not benzo when the second ring is a carbocycle).
The bicyclic heterocyclic group may be attached to its pendant group at any heteroatom or carbon atom which results in a stable structure. The bicyclic heterocyclic group described herein may be substituted on carbon or on a nitrogen atom if the resulting compound is stable. It is preferred that when the total number of S and O atoms in the heterocycle exceeds 1, then these heteroatoms are not adjacent to one another. It is preferred that the total number of S and O atoms in the heterocycle is not more than 1.
Examples of a bicyclic heterocyclic group are, but not limited to, quinolinyl, isoquinolinyl, phthalazinyl, quinazolinyl, indolyl, isoindolyl, indolinyl, 1H-indazolyl, benzimidazolyl, 1,2,3,4-tetrahydroquinolinyl, 1,2,3,4-tetrahydroisoquinolinyl, 5,6,7,8-tetrahydro-quinolinyl, 2,3-dihydro-benzofuranyl, chromanyl, 1,2,3,4-tetrahydro-quinoxalinyl, and 1,2,3,4-tetrahydro-quinazolinyl.
As used herein, the term “aromatic heterocyclic group” or “heteroaryl” is intended to mean stable monocyclic and polycyclic aromatic hydrocarbons that include at least one heteroatom ring member such as sulfur, oxygen, or nitrogen. Heteroaryl groups include, without limitation, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, triazinyl, furyl, quinolyl, isoquinolyl, thienyl, imidazolyl, thiazolyl, indolyl, pyrroyl, oxazolyl, benzofuryl, benzothienyl, benzthiazolyl, isoxazolyl, pyrazolyl, triazolyl, tetrazolyl, indazolyl, 1,2,4-thiadiazolyl, isothiazolyl, purinyl, carbazolyl, benzimidazolyl, indolinyl, benzodioxolanyl, and benzodioxane. Heteroaryl groups are substituted or unsubstituted. The nitrogen atom is substituted or unsubstituted (i.e., N or NR wherein R is H or another substituent, if defined). The nitrogen and sulfur heteroatoms may optionally be oxidized (i.e., N→O and S(O)p, wherein p is 0, 1 or 2).
Examples of 5- to 6-membered heteroaryls include, but are not limited to, pyridinyl, furanyl, thienyl, pyrrolyl, pyrazolyl, pyrazinyl, imidazolyl, imidazolidinyl, tetrazolyl, isoxazolyl, oxazolyl, oxadiazolyl, oxazolidinyl, thiadiazinyl, thiadiazolyl, thiazolyl, triazinyl, and triazolyl.
Bridged rings are also included in the definition of heterocycle. A bridged ring occurs when one or more, preferably one to three, atoms (i.e., C, O, N, or S) link two non-adjacent carbon or nitrogen atoms. Examples of bridged rings include, but are not limited to, one carbon atom, two carbon atoms, one nitrogen atom, two nitrogen atoms, and a carbon-nitrogen group. It is noted that a bridge always converts a monocyclic ring into a tricyclic ring. When a ring is bridged, the substituents recited for the ring may also be present on the bridge.
The term “counter ion” is used to represent a negatively charged species such as chloride, bromide, hydroxide, acetate, and sulfate or a positively charged species such as sodium (Na+), potassium (K+), ammonium (RnNHm+ where n=0-4 and m=0-4) and the like.
When a dotted ring is used within a ring structure, this indicates that the ring structure may be saturated, partially saturated or unsaturated.
As used herein, the term “amine protecting group” means any group known in the art of organic synthesis for the protection of amine groups which is stable to an ester reducing agent, a disubstituted hydrazine, R4-M and R7-M, a nucleophile, a hydrazine reducing agent, an activator, a strong base, a hindered amine base and a cyclizing agent. Such amine protecting groups fitting these criteria include those listed in Wuts, P. G. M. et al., Protecting Groups in Organic Synthesis, 4th Edition, Wiley (2007) and The Peptides: Analysis, Synthesis, Biology, Vol. 3, Academic Press, New York (1981), the disclosure of which is hereby incorporated by reference. Examples of amine protecting groups include, but are not limited to, the following: (1) acyl types such as formyl, trifluoroacetyl, phthalyl, and p-toluenesulfonyl; (2) aromatic carbamate types such as benzyloxycarbonyl (Cbz) and substituted benzyloxycarbonyls, 1-(p-biphenyl)-1-methylethoxycarbonyl, and 9-fluorenylmethyloxycarbonyl (Fmoc); (3) aliphatic carbamate types such as tert-butyloxycarbonyl (Boc), ethoxycarbonyl, diisopropylmethoxycarbonyl, and allyloxycarbonyl; (4) cyclic alkyl carbamate types such as cyclopentyloxycarbonyl and adamantyloxycarbonyl; (5) alkyl types such as triphenylmethyl and benzyl; (6) trialkylsilane such as trimethylsilane; (7) thiol containing types such as phenylthiocarbonyl and dithiasuccinoyl; and (8) alkyl types such as triphenylmethyl, methyl, and benzyl; and substituted alkyl types such as 2,2,2-trichloroethyl, 2-phenylethyl, and t-butyl; and trialkylsilane types such as trimethylsilane.
As referred to herein, the term “substituted” means that at least one hydrogen atom is replaced with a non-hydrogen group, provided that normal valencies are maintained and that the substitution results in a stable compound. Ring double bonds, as used herein, are double bonds that are formed between two adjacent ring atoms (e.g., C═C, C═N, or N═N).
In cases wherein there are nitrogen atoms (e.g., amines) on compounds of the present invention, these may be converted to N-oxides by treatment with an oxidizing agent (e.g., mCPBA and/or hydrogen peroxides) to afford other compounds of this invention. Thus, shown and claimed nitrogen atoms are considered to cover both the shown nitrogen and its N-oxide (N→O) derivative.
When any variable occurs more than one time in any constituent or formula for a compound, its definition at each occurrence is independent of its definition at every other occurrence. Thus, for example, if a group is shown to be substituted with 0-3 R, then said group may optionally be substituted with up to three R groups, and at each occurrence R is selected independently from the definition of R.
When a bond to a substituent is shown to cross a bond connecting two atoms in a ring, then such substituent may be bonded to any atom on the ring. When a substituent is listed without indicating the atom in which such substituent is bonded to the rest of the compound of a given formula, then such substituent may be bonded via any atom in such substituent.
Combinations of substituents and/or variables are permissible only if such combinations result in stable compounds.
The phrase “pharmaceutically acceptable” is employed herein to refer to those compounds, materials, compositions, and/or dosage forms that are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, and/or other problem or complication, commensurate with a reasonable benefit/risk ratio.
As used herein, “pharmaceutically acceptable salts” refer to derivatives of the disclosed compounds wherein the parent compound is modified by making acid or base salts thereof. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic groups such as amines; and alkali or organic salts of acidic groups such as carboxylic acids. The pharmaceutically acceptable salts include the conventional non-toxic salts or the quaternary ammonium salts of the parent compound formed, for example, from non-toxic inorganic or organic acids. Examples of pharmaceutically acceptable, nontoxic acid addition salts are salts of an amino group formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid or with organic acids such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid or by using other methods used in the art such as ion exchange. Other pharmaceutically acceptable salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate salts, and the like.
Salts derived from appropriate bases include alkali metal, alkaline earth metal, ammonium and N+(C1-4alkyl)4 salts. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like. Further pharmaceutically acceptable salts include, when appropriate, nontoxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulfate, phosphate, nitrate, loweralkyl sulfonate and aryl sulfonate.
The pharmaceutically acceptable salts of the present invention can be synthesized from the parent compound that contains a basic or acidic moiety by conventional chemical methods. Generally, such salts can be prepared by reacting the free acid or base forms of these compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent, or in a mixture of the two; generally, nonaqueous media like ether, ethyl acetate, ethanol, isopropanol, or acetonitrile are preferred. Lists of suitable salts are found in Allen, Jr., L. V., ed., Remington: The Science and Practice of Pharmacy, 22nd Edition, Pharmaceutical Press, London, UK (2012), the disclosure of which is hereby incorporated by reference.
In addition, compounds of formula I may have prodrug forms. Any compound that will be converted in vivo to provide the bioactive agent (i.e., a compound of formula I) is a prodrug within the scope and spirit of the invention. Various forms of prodrugs are well known in the art. For examples of such prodrug derivatives, see:
Compounds containing a carboxy group can form physiologically hydrolyzable esters that serve as prodrugs by being hydrolyzed in the body to yield formula I compounds per se. Such prodrugs are preferably administered orally since hydrolysis in many instances occurs principally under the influence of the digestive enzymes. Parenteral administration may be used where the ester per se is active, or in those instances where hydrolysis occurs in the blood. Examples of physiologically hydrolyzable esters of compounds of formula I include C1-6alkyl, C1-6alkylbenzyl, 4-methoxybenzyl, indanyl, phthalyl, methoxymethyl, C1-6 alkanoyloxy-C1-6alkyl (e.g., acetoxymethyl, pivaloyloxymethyl or propionyloxymethyl), C1-6alkoxycarbonyloxy-C1-6alkyl (e.g., methoxycarbonyl-oxymethyl or ethoxycarbonyloxymethyl, glycyloxymethyl, phenylglycyloxymethyl, (5-methyl-2-oxo-1,3-dioxolen-4-yl)-methyl), and other well known physiologically hydrolyzable esters used, for example, in the penicillin and cephalosporin arts. Such esters may be prepared by conventional techniques known in the art.
Preparation of prodrugs is well known in the art and described in, for example, King, F. D., ed., Medicinal Chemistry: Principles and Practice, The Royal Society of Chemistry, Cambridge, UK (2nd Edition, reproduced (2006)); Testa, B. et al., Hydrolysis in Drug and Prodrug Metabolism. Chemistry, Biochemistry and Enzymology, VCHA and Wiley-VCH, Zurich, Switzerland (2003); Wermuth, C. G., ed., The Practice of Medicinal Chemistry, 3rd Edition, Academic Press, San Diego, Calif. (2008).
The present invention is intended to include all isotopes of atoms occurring in the present compounds. Isotopes include those atoms having the same atomic number but different mass numbers. By way of general example and without limitation, isotopes of hydrogen include deuterium (symbol D or 2H) and tritium (symbol T or 3H). For example, a methyl group may be represented by CH3 or CD3. Isotopes of carbon include 13C and 14C. Isotopically-labeled compounds of the invention can generally be prepared by conventional techniques known to those skilled in the art or by processes analogous to those described herein, using an appropriate isotopically-labeled reagent in place of the non-labeled reagent otherwise employed.
The term “solvate” means a physical association of a compound of this invention with one or more solvent molecules, whether organic or inorganic. This physical association includes hydrogen bonding. In certain instances the solvate will be capable of isolation, for example when one or more solvent molecules are incorporated in the crystal lattice of the crystalline solid. The solvent molecules in the solvate may be present in a regular arrangement and/or a non-ordered arrangement. The solvate may comprise either a stoichiometric or nonstoichiometric amount of the solvent molecules. “Solvate” encompasses both solution-phase and isolable solvates. Exemplary solvates include, but are not limited to, hydrates, ethanolates, methanolates, and isopropanolates. Methods of solvation are generally known in the art.
The terms “measurable affinity” and “measurably inhibit,” as used herein, means a measurable change in PAD4 activity between a sample comprising a compound of the present invention, or composition thereof, and PAD4, and an equivalent sample comprising PAD4 in the absence of said compound, or composition thereof.
Abbreviations as used herein, are defined as follows: “1×” for once, “2×” for twice, “3×” for thrice, “° C.” for degrees Celsius, “eq” for equivalent or equivalents, “g” for gram or grams, “mg” for milligram or milligrams, “L” for liter or liters, “mL” for milliliter or milliliters, “μL” for microliter or microliters, “N” for normal, “M” for molar, “mmol” for millimole or millimoles, “min” for minute or min, “h” for hour or h, “rt” for room temperature, “RT” for retention time, “atm” for atmosphere, “psi” for pounds per square inch, “conc.” for concentrate, “aq” for “aqueous”, “sat” or “sat'd” for saturated, “MW” for molecular weight, “mp” for melting point, “MS” or “Mass Spec” for mass spectrometry, “ESI” for electrospray ionization mass spectroscopy, “HR” for high resolution, “HRMS” for high resolution mass spectrometry, “LCMS” for liquid chromatography mass spectrometry, “HPLC” for high pressure liquid chromatography, “RP HPLC” for reverse phase HPLC, “TLC” or “tlc” for thin layer chromatography, “NMR” for nuclear magnetic resonance spectroscopy, “nOe” for nuclear Overhauser effect spectroscopy, “1H” for proton, “S” for delta, “s” for singlet, “d” for doublet, “t” for triplet, “q” for quartet, “in” for multiplet, “br” for broad, “Hz” for hertz, and “α”, “β”, “R”, “S”, “E”, “Z” and “ee” are stereochemical designations familiar to one skilled in the art. As used herein, the term “pharmaceutically acceptable salt” refers to those salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio.
In a first aspect, the present invention provides a compound of Formula (I):
or a pharmaceutically acceptable salt thereof, wherein:
cycloalkyl, benzo[d][1,2,3]triazol-5-yl, 6-acetamido-n-hex-1-yl, and (3-(hydroxymethyl)oxetan-3-yl)methyl.
In a second aspect, the present invention provides a compound of Formula
or a pharmaceutically acceptable salt thereof, within the scope of the first aspect, wherein:
In a third aspect, the present invention provides a compound of Formula (III):
or a pharmaceutically acceptable salt thereof, within the scope of the first and second aspects, wherein:
In a fourth aspect, the present invention provides a compound or a pharmaceutically acceptable salt thereof, within the scope of the first, second, and third aspects, wherein:
In a fifth aspect, the present invention provides a compound or a pharmaceutically acceptable salt thereof, within the scope of the first, second, and third aspects, wherein:
In a sixth aspect, the present invention provides a compound or a pharmaceutically acceptable salt thereof, within the scope of the fifth aspect, wherein:
In a seventh aspect, the present invention provides a compound or a pharmaceutically acceptable salt thereof, within the scope of the first, second, and third aspects, wherein:
In an eighth aspect, the present invention provides a compound or a pharmaceutically acceptable salt thereof, within the scope of the first, second, and third aspects, wherein:
In a ninth aspect, the present invention provides a compound or a pharmaceutically acceptable salt thereof, within the scope of the eighth aspect, wherein: R2 is selected from —CH3 and —CH2-cyclopropyl substituted with 0-2 F and Cl;
R5, at each occurrence, is independently selected from H, F, Cl, Br, C1-4alkyl, aryl substituted with 0-4 Re, and heterocyclyl, substituted with 0-4 Re;
In a tenth aspect, the present invention provides a compound or a pharmaceutically acceptable salt thereof, within the scope of the first, second, and third aspects, wherein:
In an eleventh aspect, the present invention provides a compound of Formula (IV):
or a pharmaceutically acceptable salt thereof, within the scope of the first aspect, wherein:
In a twelfth aspect, the present invention provides a compound or a pharmaceutically acceptable salt thereof, within the scope of the first aspect, wherein:
In a thirteenth aspect, the present invention provides a compound of Formula (I):
or a pharmaceutically acceptable salt thereof, wherein:
and
2) when L is NRa-L-R4 is not
In a fourteenth aspect, the present invention provides a compound of Formula (III):
or a pharmaceutically acceptable salt thereof, within the scope of the thirteenth aspect, wherein:
is selected from
R5, at each occurrence, is independently selected from H, F, Cl, Br, C1-4alkyl substituted with 0-5 Re, C2-4alkenyl, C2-4alkynyl, nitro, —(CH2)rORb, —CN, —NRaRa, —(CH2)rNRaC(═O)Rb, —NRaC(═O)NRaRa, —C(═O)ORb, —C(═O)Rb, —OC(═O)Rb, —C(═O)NRaRa, —P(═O)(C1-4alkyl)2, —S(O)pRc, —S(O)pNRaRa, —NRaS(O)pRc, —C3-6 cycloalkyl substituted with 0-4 Re, aryl substituted with 0-4 Re, and heterocyclyl substituted with 0-4 Re;
In a fifteenth aspect, the present invention provides a compound of Formula (III), or a pharmaceutically acceptable salt thereof, within the scope of the fourteenth aspect, wherein:
is selected from
In a sixteenth aspect, the present invention provides a compound of Formula (III), or a pharmaceutically acceptable salt thereof, within the scope of the fourteenth aspect, wherein:
In a seventeenth aspect, the present invention provides a compound of Formula (III), or a pharmaceutically acceptable salt thereof, within the scope of the fourteenth aspect, wherein:
In an eighteenth aspect, the present invention provides a compound of Formula (III), or a pharmaceutically acceptable salt thereof, within the scope of the seventeenth aspect, wherein:
In a nineteenth aspect, the present invention provides a compound of Formula (I), or a pharmaceutically acceptable salt thereof, within the scope of the fourteenth aspect, wherein:
In a twentieth aspect, the present invention provides a compound of Formula (III), or a pharmaceutically acceptable salt thereof, within the scope of the fourteenth aspect, wherein:
In a twenty first aspect, the present invention provides a compound of Formula (III), or a pharmaceutically acceptable salt thereof, within the scope of the twentieth aspect, wherein:
In a twenty second aspect, the present invention provides a compound of Formula (III), or a pharmaceutically acceptable salt thereof, within the scope of the fourteenth aspect, wherein:
R5, at each occurrence, is independently selected from H, F, Cl, Br, and C1-4alkyl substituted with 0-5 Re, ORb, —CN, —C(═O)Rb, —C(═O)ORb, —OC(═O)Rb, —C(═O)NRaRa;
In a twenty third aspect, the present invention provides a compound of Formula (III), or a pharmaceutically acceptable salt thereof, within the scope of the fourteenth aspect, wherein:
is selected from
In a twenty fourth aspect, the present invention provides a compound of Formula (III), or a pharmaceutically acceptable salt thereof, within the scope of the fourteenth aspect, wherein:
is selected from
In a twenty fifth aspect, the present invention provides a compound of Formula (V):
or a pharmaceutically acceptable salt thereof, within the scope of the fourteenth aspect, wherein:
is selected from
In a twenty sixth aspect, the present invention provides a compound of Formula VI:
or a pharmaceutically acceptable salt thereof, wherein:
and
In a twenty seventh aspect, the present invention provides a compound of Formula (VII):
or a pharmaceutically acceptable salt thereof, wherein:
is selected from
In a twenty eighth aspect, the present invention provides a compound of Formula (VIII):
or a pharmaceutically acceptable salt thereof, wherein:
is selected from
In a twenty ninth aspect, the present invention provides a compound of Formula (IX):
or a pharmaceutically acceptable salt thereof, wherein:
is selected from
R5b is selected from OH, —C(═O)NRaRa, —C(═O)ORb, NHC(═O)Rb, and NH2, Ra, at each occurrence, is independently selected from H, CH3, and CD3; or Ra and Ra together with the nitrogen atom to which they are both attached form a heterocyclic ring substituted with 0-2 OH; and
In a thirtieth aspect, the present invention provides a compound of Formula (X):
or a pharmaceutically acceptable salt thereof, wherein:
is selected from
In a thirty first aspect, the present invention provides a compound of Formula
or a pharmaceutically acceptable salt thereof, wherein:
is selected from
and
As defined above and described herein, R1 is selected from —CH3 —CD3, and —CH2-5-6 membered heterocyclyl comprising carbon atoms and 1-3 heteroatoms selected from N, NH, and NC1-3alkyl. In some embodiments, R1 is —CH3. In some embodiments, R1 is —CD3. In some embodiments, R1 is
As defined above and described herein, R2 is hydrogen, C1-3 alkyl substituted with 0-5 Re, or C3-6 cycloalkyl substituted with 0-5 Re. In some embodiments, R2 is hydrogen. In some embodiments, R2 is C1-2 alkyl substituted with C3-6 cycloalkyl. In some embodiments, R2 is C3-6 cycloalkyl. In some embodiments, R2 is methyl. In some embodiments, R2 is ethyl. In some embodiments, R2 is cyclopropyl. In some embodiments, R2 is cyclobutyl. In some embodiments, R2 is cyclopentyl. In some embodiments, R2 is cyclohexyl. In some embodiments, R2 is cyclopropylmethyl. In some embodiments, R2 is cyclobutylmethyl. In some embodiments, R2 is cyclopentylmethyl. In some embodiments, R2 is cyclohexylmethyl. In some embodiments, R2 is cyclopropylethyl. In some embodiments, R2 is cyclobutylethyl. In some embodiments, R2 is cyclopentylethyl. In some embodiments, R2 is cyclohexylethyl. In some embodiments, R2 is —CH2-cyclopropyl or —CH2-cyclobutyl. In some embodiments, R2 is —CH2-cyclobutyl optionally substituted with methyl and —OH. In certain embodiments, R2 is selected from those functional groups depicted in the examples below.
As defined above and described herein, R3 is selected from H, F, Cl, Br, —ORb, and C1-3 alkyl substituted with 0-5 Re. In some embodiments, R3 is H, F, Cl, Br. In some embodiments, R3 is F. In some embodiments, R3 is H. In some embodiments, R3 is C1-3 alkyl. In some embodiments, R3 is methyl. In some embodiments, R3 is ethyl. In some embodiments, R3 is propyl. In some embodiments, R3 is ORb. In some embodiments, R3 is —CH3. In some embodiments, R3 is —CH2CH3. In some embodiments, R3 is —OCH2CH2CH3. In certain embodiments, R3 is —OCH(F)2. In certain embodiments, R3 is selected from those functional groups depicted in the examples below.
As defined above and described herein, L is absent, —NRa—, —O—, —C(═O)NRd—, or —S(O)p—, In some embodiments, L is absent. In some embodiments, L is —NRa—, Ra is H or C1-3alkyl. In some embodiments, L is —O—. In some embodiments, L is —C(═O)NH—. In some embodiments, L is —S(O)2—. In some embodiments, L is —S—. In certain embodiments, L is selected from those functional groups depicted in the examples below.
As defined above and described herein, each R4 is
In some embodiments, R4 is
In some embodiments, R4 is
In some embodiments, R4 is
In some embodiments, R4 is
In some embodiments, R4 is
In some embodiments, R4 is
In some embodiments, R4 is
In some embodiments, R4 is
In some embodiments, R4 is
In some embodiments, R4 is
In some embodiments, R4 is
In some embodiments, R4 is
In some embodiments, R4 is
In certain embodiments, R4 is selected from those functional groups depicted in the examples below.
As defined above and described herein, R1 is H, F, Cl, Br, CN, C1-4alkyl substituted with 0-5 Re, C2-4alkenyl, C2-4alkynyl, nitro, —S(O)pRc, —S(O)pNRaRa, —NRaS(O)pRc, —(CHRd)rORb, —(CH2)rNRaRa, —NRaC(═O)Rb, NRaC(═O)ORb —NRaC(═O)NRaRa, —C(═O)Rb, —C(═O)ORb, C(═O)NRaRa, —OC(═O)Rb, C3-6cycloalkyl substituted with 0-4 Re, aryl substituted with 0-4 Re, and heterocyclyl substituted with 0-4 Re
In some embodiments, R5 is H, F, Cl, CN, C1-4alkyl, C1-4alkyl (substituted with OH, NH2, and COOH), SC1-4alkyl, S(O)2C1-4alkyl, S(O)2NH-cyclopropyl, —(CH2)0-1NHS(O)2C1-4alkyl, N(Rd)S(O)2C2-4alkenyl, —(CH2)0-1OH, OC1-4alkyl, —(CH2)0-1NH2, —(CH2)0-1NHC(═O)C1-4alkyl, —NRdC(═O)C2-4alkenyl, —NHC(═O)C2-4alkynyl, —(CH2)0-1C(═O)OH, —C(═O)OC1-4alkyl, —NHC(═O)OC1-4alkyl, —NHC(═O)O(CH2)2OC1-4alkyl, —NHC(═O)OCH2-cyclopropyl, —NHC(═O)NH2, C(═O)NHC1-4alkyl, CONH(CH2)1-2C(═O)OH, —(CH2)0-1C(═O)NH2, —(CH2)0-1C(═O)NHC1-4alkyl, C(═O)NH-pyridine, —C(═O)NH(CH2)2N(C1-4alkyl)2, —C(═O)NH(CH2)2OH, —C(═O)NH(CH2)2S(O)2C1-4alkyl, and —OC(═O)C1-4alkyl.
In some embodiments, R5 is
In some embodiments, R5 is F. In some embodiments, R5 C1-4alkyl. In some embodiments, R5 is —OH or —OC1-3alkyl. In some embodiments, R5 is —NHS(O)2C2-4alkenyl. In certain embodiments, R5 is selected from those functional groups depicted in the examples below.
As defined above and described herein, R6 is H, C1-3alkyl substituted with 0-4 Re, —S(O)pRc, —C(═O)Rb, —(CH2)r—C(═O)NRaRa, —C(═O)(CH2)rNRaC(═O)Rb, —C(═O)ORb, —S(O)pNRaRa, aryl substituted with 0-4 Re, or heterocyclyl substituted with 0-4 Re.
In some embodiments, R6 is H. In some embodiments, R6 is methyl or isopropyl. In some embodiments, R6 is —(CH2)2C(═O)NH2. In some embodiments, R6 is —(CH2)2OH. In some embodiments, R6 is C(═O)C1-4alkyl. In certain embodiments, R6 is selected from those functional groups depicted in the examples below.
As defined above and described herein, R7 is H, F, Cl, C1-3alkyl, —NRaRa, or —NRaC(═O)ORb. In some embodiments, R7 is NH2. In some embodiments, R7 is F.
As defined above and described herein, R8 is H, F, Cl, Br, or C1-4alkyl substituted with 0-5 Re. In some embodiments, R is H. In some embodiments, R8 is C1-3alkyl.
As defined above, Ring A including its substituent R7 is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In some embodiments, Ring A is
In certain embodiments, R4 is selected from those functional groups depicted in the examples below.
As defined above and described herein, r is 0-4. In some embodiments, r is 0. In some embodiments, r is 1. In some embodiments, r is 2. In some embodiments, r is 3. In some embodiments, r is 4.
In some embodiments, Ring A is
R1 is —CH3, R2 is cyclopropylmethyl, R3 is H, F, or —OCH3, R4 is
and R5 is H, F, Cl, CN, C1-4alkyl, C1-4alkyl substituted with OH, NH2, and COOH, SC1-4alkyl, S(O)2C1-4alkyl, S(O)2NH-cyclopropyl, —(CH2)0-1NHS(O)2C1-4alkyl, N(Ra)S(O)2C2-4alkenyl, —(CH2)0-1OH, OC1-4alkyl, —(CH2)0-1NH2, —(CH2)0-1NHC(═O)C1-4alkyl, —NRdC(═O)C2-4alkenyl, —NHC(═O)C2-4alkynyl, —(CH2)0-1C(═O)OH, —C(═O)OC1-4alkyl, —NHC(═O)OC1-4alkyl, —NHC(═O)O(CH2)2OC1-4alkyl, —NHC(═O)OCH2-cyclopropyl, —NHC(═O)NH2, C(═O)NHC1-4alkyl, CONH(CH2)1-2C(═O)OH, —(CH2)0-1C(═O)NH2, —(CH2)0-1C(═O)NHC1-4alkyl, C(═O)NH-pyridine, —C(═O)NH(CH2)2N(C1-4alkyl)2, —C(═O)NH(CH2)2OH, —C(═O)NH(CH2)2S(O)2C1-4alkyl, and —OC(═O)C1-4alkyl,
In some embodiments, Ring A is
R1 is —CH3, R2 is cyclopropylmethyl, R3 is H, F, or —OCH3, R5 is
and R5 is H, F, Cl, CN, C1-4alkyl, C1-4alkyl substituted with OH, NH2, and COOH, SC1-4alkyl, S(O)2C1-4alkyl, S(O)2NH-cyclopropyl, —(CH2)0-1NHS(O)2C1-4alkyl, N(Rd)S(O)2C2-4alkenyl, —(CH2)0-1OH, OC1-4alkyl, —(CH2)0-1NH2, —(CH2)0-1NHC(═O)C1-4alkyl, —NRdC(═O)C2-4alkenyl, —NHC(═O)C2-4alkynyl, —(CH2)0-1C(═O)OH, —C(═O)OC1-4alkyl, —NHC(═O)OC1-4alkyl, —NHC(═O)O(CH2)2OC1-4alkyl, —NHC(═O)OCH2-cyclopropyl, —NHC(═O)NH2, C(═O)NHC1-4alkyl, CONH(CH2)1-2C(═O)OH, —(CH2)0-1C(═O)NH2, —(CH2)0-1C(═O)NHC1-4alkyl, C(═O)NH-pyridine, —C(═O)NH(CH2)2N(C1-4alkyl)2, —C(═O)NH(CH2)2OH, —C(═O)NH(CH2)2S(O)2C1-4alkyl, and —OC(═O)C1-4alkyl,
In some embodiments, Ring A is
R1 is —CH3, R2 is cyclopropylmethyl, R3 is H, F or —OCH3, R4 is
In some embodiments, Ring A is
R1 is —CH3, R2 is cyclopropylmethyl, R3 is H, F or —OCH3, R4 is
In some embodiments, Ring A is
R1 is —CH3, R2 is cyclopropylmethyl, R3 is H, F or —OCH3, R4 is
In some embodiments, Ring A is
R1 is —CH3, R2 is cyclopropylmethyl, R3 is H, F or —CH3, R4 is
In some embodiments, Ring A is
R1 is —CH3, R2 is cyclopropylmethyl, R3 is H, F or —OCH3, R4 is
In some embodiments, Ring A is
R1 is —CH3, R2 is cyclopropylmethyl, R3 is H, F or —OCH3, R4 is
and R5 H, F, R5 C1-4alkyl, —OH, —OC1-3alkyl and is —NHS(O)2C2-4alkenyl.
In some embodiments, the compound of Formula (I) is selected from examples depicted below. In certain embodiments, the present invention provides any compound described above and herein, or a pharmaceutically acceptable salt thereof. In some embodiments, the present invention provides any compound described above and herein in isolated form.
According to another embodiment, the invention provides a composition comprising a compound of this invention or a pharmaceutically acceptable derivative thereof and a pharmaceutically acceptable carrier, adjuvant, or vehicle. The amount of compound in compositions of this invention is such that is effective to measurably inhibit PAD4, in a biological sample or in a patient. In certain embodiments, the amount of compound in compositions of this invention is such that is effective to measurably inhibit PAD4, in a biological sample or in a patient. In certain embodiments, a composition of this invention is formulated for administration to a patient in need of such composition. In some embodiments, a composition of this invention is formulated for oral administration to a patient.
The term “subject,” as used herein, is used interchangeably with the term “patient” and means an animal, preferably a mammal. In some embodiments, a subject or patient is a human. In other embodiments, a subject (or patient) is a veterinary subject (or patient). In some embodiments, a veterinary subject (or patient) is a canine, a feline, or an equine subject.
The term “pharmaceutically acceptable carrier, adjuvant, or vehicle” refers to a non-toxic carrier, adjuvant, or vehicle that does not destroy the pharmacological activity of the compound with which it is formulated. Pharmaceutically acceptable carriers, adjuvants or vehicles that may be used in the compositions of this invention include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, serum proteins, such as human serum albumin, buffer substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone, cellulose-based substances, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, polyethylene glycol and wool fat.
Compositions of the present invention may be administered orally, parenterally, by inhalation spray, topically, rectally, nasally, buccally, vaginally or via an implanted reservoir. The term “parenteral” as used herein includes subcutaneous, intravenous, intramuscular, intra-articular, intra-synovial, intrasternal, intrathecal, intrahepatic, intralesional and intracranial injection or infusion techniques. Preferably, the compositions are administered orally, intraperitoneally or intravenously. Sterile injectable forms of the compositions of this invention may be aqueous or oleaginous suspension. These suspensions may be formulated according to techniques known in the art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally acceptable diluent or solvent, for example as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium.
For this purpose, any bland fixed oil may be employed including synthetic mono- or di-glycerides. Fatty acids, such as oleic acid and its glyceride derivatives are useful in the preparation of injectables, as are natural pharmaceutically-acceptable oils, such as olive oil or castor oil, especially in their polyoxyethylated versions. These oil solutions or suspensions may also contain a long-chain alcohol diluent or dispersant, such as carboxymethyl cellulose or similar dispersing agents that are commonly used in the formulation of pharmaceutically acceptable dosage forms including emulsions and suspensions. Other commonly used surfactants, such as Tweens, Spans and other emulsifying agents or bioavailability enhancers which are commonly used in the manufacture of pharmaceutically acceptable solid, liquid, or other dosage forms may also be used for the purposes of formulation.
Pharmaceutically acceptable compositions of this invention may be orally administered in any orally acceptable dosage form including, but not limited to, capsules, tablets, aqueous suspensions or solutions. In the case of tablets for oral use, carriers commonly used include lactose and corn starch. Lubricating agents, such as magnesium stearate, are also typically added. For oral administration in a capsule form, useful diluents include lactose and dried cornstarch. When aqueous suspensions are required for oral use, the active ingredient is combined with emulsifying and suspending agents. If desired, certain sweetening, flavoring or coloring agents may also be added.
Alternatively, pharmaceutically acceptable compositions of this invention may be administered in the form of suppositories for rectal administration. These can be prepared by mixing the agent with a suitable non-irritating excipient that is solid at room temperature but liquid at rectal temperature and therefore will melt in the rectum to release the drug. Such materials include cocoa butter, beeswax and polyethylene glycols.
Pharmaceutically acceptable compositions of this invention may also be administered topically, especially when the target of treatment includes areas or organs readily accessible by topical application, including diseases of the eye, the skin, or the lower intestinal tract. Suitable topical formulations are readily prepared for each of these areas or organs.
Topical application for the lower intestinal tract can be effected in a rectal suppository formulation (see above) or in a suitable enema formulation. Topically-transdermal patches may also be used.
For topical applications, provided pharmaceutically acceptable compositions may be formulated in a suitable ointment containing the active component suspended or dissolved in one or more carriers. Carriers for topical administration of compounds of this invention include, but are not limited to, mineral oil, liquid petrolatum, white petrolatum, propylene glycol, polyoxyethylene, polyoxypropylene compound, emulsifying wax and water. Alternatively, provided pharmaceutically acceptable compositions can be formulated in a suitable lotion or cream containing the active components suspended or dissolved in one or more pharmaceutically acceptable carriers. Suitable carriers include, but are not limited to, mineral oil, sorbitan monostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol and water.
For ophthalmic use, provided pharmaceutically acceptable compositions may be formulated as micronized suspensions in isotonic, pH adjusted sterile saline, or, preferably, as solutions in isotonic, pH adjusted sterile saline, either with or without a preservative such as benzylalkonium chloride. Alternatively, for ophthalmic uses, the pharmaceutically acceptable compositions may be formulated in an ointment such as petrolatum.
Pharmaceutically acceptable compositions of this invention may also be administered by nasal aerosol or inhalation. Such compositions are prepared according to techniques well-known in the art of pharmaceutical formulation and may be prepared as solutions in saline, employing benzyl alcohol or other suitable preservatives, absorption promoters to enhance bioavailability, fluorocarbons, and/or other conventional solubilizing or dispersing agents.
Most preferably, pharmaceutically acceptable compositions of this invention are formulated for oral administration. Such formulations may be administered with or without food. In some embodiments, pharmaceutically acceptable compositions of this invention are administered without food. In other embodiments, pharmaceutically acceptable compositions of this invention are administered with food.
Pharmaceutically acceptable compositions of this invention can be administered to humans and other animals orally, rectally, parenterally, intracisternally, intravaginally, intraperitoneally, topically (as by powders, ointments, or drops), bucally, as an oral or nasal spray, or the like, depending on the severity of the infection being treated. In certain embodiments, the compounds of the invention may be administered orally or parenterally at dosage levels of about 0.01 mg/kg to about 50 mg/kg and preferably from about 1 mg/kg to about 25 mg/kg, of subject body weight per day, one or more times a day, to obtain the desired therapeutic effect.
Liquid dosage forms for oral administration include, but are not limited to, pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to the active compounds, the liquid dosage forms may contain inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof. Besides inert diluents, the oral compositions can also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.
Injectable preparations, for example, sterile injectable aqueous or oleaginous suspensions may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution, suspension or emulsion in a nontoxic parenterally acceptable diluent or solvent, for example, as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution, U.S.P. and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil can be employed including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid are used in the preparation of injectables.
Injectable formulations can be sterilized, for example, by filtration through a bacterial-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable medium prior to use.
In order to prolong the effect of a compound of the present invention, it is often desirable to slow the absorption of the compound from subcutaneous or intramuscular injection. This may be accomplished by the use of a liquid suspension of crystalline or amorphous material with poor water solubility. The rate of absorption of the compound then depends upon its rate of dissolution that, in turn, may depend upon crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered compound form is accomplished by dissolving or suspending the compound in an oil vehicle. Injectable depot forms are made by forming microencapsule matrices of the compound in biodegradable polymers such as polylactide-polyglycolide. Depending upon the ratio of compound to polymer and the nature of the particular polymer employed, the rate of compound release can be controlled. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations are also prepared by entrapping the compound in liposomes or microemulsions that are compatible with body tissues.
Compositions for rectal or vaginal administration are preferably suppositories which can be prepared by mixing the compounds of this invention with suitable non-irritating excipients or carriers such as cocoa butter, polyethylene glycol or a suppository wax which are solid at ambient temperature but liquid at body temperature and therefore melt in the rectum or vaginal cavity and release the active compound.
Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active compound is mixed with at least one inert, pharmaceutically acceptable excipient or carrier such as sodium citrate or dicalcium phosphate and/or a) fillers or extenders such as starches, lactose, sucrose, glucose, mannitol, and silicic acid, b) binders such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidinone, sucrose, and acacia, c) humectants such as glycerol, d) disintegrating agents such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate, e) solution retarding agents such as paraffin, f) absorption accelerators such as quaternary ammonium compounds, g) wetting agents such as, for example, cetyl alcohol and glycerol monostearate, h) absorbents such as kaolin and bentonite clay, and i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets and pills, the dosage form may also comprise buffering agents.
Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols and the like. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulating art. They may optionally contain opacifying agents and can also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes. Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polethylene glycols and the like.
The active compounds can also be in micro-encapsulated form with one or more excipients as noted above. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings, release controlling coatings and other coatings well known in the pharmaceutical formulating art. In such solid dosage forms the active compound may be admixed with at least one inert diluent such as sucrose, lactose or starch. Such dosage forms may also comprise, as is normal practice, additional substances other than inert diluents, e.g., tableting lubricants and other tableting aids such a magnesium stearate and microcrystalline cellulose. In the case of capsules, tablets and pills, the dosage forms may also comprise buffering agents. They may optionally contain opacifying agents and can also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes.
Dosage forms for topical or transdermal administration of a compound of this invention include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants or patches. The active component is admixed under sterile conditions with a pharmaceutically acceptable carrier and any needed preservatives or buffers as may be required. Ophthalmic formulation, ear drops, and eye drops are also contemplated as being within the scope of this invention. Additionally, the present invention contemplates the use of transdermal patches, which have the added advantage of providing controlled delivery of a compound to the body. Such dosage forms can be made by dissolving or dispensing the compound in the proper medium. Absorption enhancers can also be used to increase the flux of the compound across the skin. The rate can be controlled by either providing a rate controlling membrane or by dispersing the compound in a polymer matrix or gel.
The amount of compounds of the present invention that may be combined with the carrier materials to produce a composition in a single dosage form will vary depending upon the host treated, the particular mode of administration. Preferably, provided compositions should be formulated so that a dosage of between 0.01-100 mg/kg body weight/day of the inhibitor can be administered to a patient receiving these compositions.
A compound of the current invention can be administered alone or in combination with one or more other therapeutic compounds, possible combination therapy taking the form of fixed combinations or the administration of a compound of the invention and one or more other therapeutic compounds being staggered or given independently of one another, or the combined administration of fixed combinations and one or more other therapeutic compounds. Exemplary of such other therapeutic agents include corticosteroids, rolipram, calphostin, cytokine-suppressive anti-inflammatory drugs (CSAIDs), Interleukin-10, glucocorticoids, salicylates, nitric oxide, and other immunosuppressants; nuclear translocation inhibitors, such as deoxyspergualin (DSG); non-steroidal antiinflammatory drugs (NSAIDs) such as ibuprofen, celecoxib and rofecoxib; steroids such as prednisone or dexamethasone; antiviral agents such as abacavir; antiproliferative agents such as methotrexate, leflunomide, FK506 (tacrolimus, Prograf); cytotoxic drugs such as azathiprine and cyclophosphamide; TNF-α inhibitors such as tenidap, anti-TNF antibodies or soluble TNF receptor, and rapamycin (sirolimus or Rapamune) or derivatives thereof. A compound of the current invention can besides or in addition be administered especially for tumor therapy in combination with chemotherapy, radiotherapy, immunotherapy, phototherapy, surgical intervention, or a combination of these. Long-term therapy is equally possible as is adjuvant therapy in the context of other treatment strategies, as described above. Other possible treatments are therapy to maintain the patient's status after tumor regression, or even chemopreventive therapy, for example in patients at risk.
Those additional agents may be administered separately from an inventive compound-containing composition, as part of a multiple dosage regimen. Alternatively, those agents may be part of a single dosage form, mixed together with a compound of this invention in a single composition. If administered as part of a multiple dosage regime, the two active agents may be submitted simultaneously, sequentially or within a period of time from one another normally within five hours from one another.
As used herein, the term “combination,” “combined,” and related terms refers to the simultaneous or sequential administration of therapeutic agents in accordance with this invention. For example, a compound of the present invention may be administered with another therapeutic agent simultaneously or sequentially in separate unit dosage forms or together in a single unit dosage form. Accordingly, the present invention provides a single unit dosage form comprising a compound of the current invention, an additional therapeutic agent, and a pharmaceutically acceptable carrier, adjuvant, or vehicle.
The amount of both an inventive compound and additional therapeutic agent (in those compositions which comprise an additional therapeutic agent as described above) that may be combined with the carrier materials to produce a single dosage form will vary depending upon the host treated and the particular mode of administration. Preferably, compositions of this invention should be formulated so that a dosage of between 0.01-100 mg/kg body weight/day of an inventive compound can be administered.
In those compositions which comprise an additional therapeutic agent, that additional therapeutic agent and the compound of this invention may act synergistically. Therefore, the amount of additional therapeutic agent in such compositions will be less than that required in a monotherapy utilizing only that therapeutic agent.
The amount of additional therapeutic agent present in the compositions of this invention will be no more than the amount that would normally be administered in a composition comprising that therapeutic agent as the only active agent. Preferably the amount of additional therapeutic agent in the presently disclosed compositions will range from about 50% to 100% of the amount normally present in a composition comprising that agent as the only therapeutically active agent.
It should also be understood that a specific dosage and treatment regimen for any particular patient will depend upon a variety of factors, including the activity of the specific compound employed, the age, body weight, general health, sex, diet, time of administration, rate of excretion, drug combination, and the judgment of the treating physician and the severity of the particular disease being treated. The amount of a compound of the present invention in the composition will also depend upon the particular compound in the composition.
Compounds and compositions described herein are generally useful for the inhibition of PAD4.
The activity of a compound utilized in this invention as an inhibitor of PAD4, may be assayed in vitro, in vivo or in a cell line. In vitro assays include assays that determine the inhibition of PAD4. Detailed conditions for assaying a compound utilized in this invention as an inhibitor of PAD4 are set forth in the Examples below. In some embodiments, a provided compound inhibits PAD4 selectively as compared to PAD2.
As used herein, the terms “treatment,” “treat,” and “treating” refer to reversing, alleviating, delaying the onset of, or inhibiting the progress of a disease or disorder, or one or more symptoms thereof, as described herein. In some embodiments, treatment may be administered after one or more symptoms have developed. In other embodiments, treatment may be administered in the absence of symptoms. For example, treatment may be administered to a susceptible individual prior to the onset of symptoms (e.g., in light of a history of symptoms and/or in light of genetic or other susceptibility factors). Treatment may also be continued after symptoms have resolved, for example to prevent or delay their recurrence.
Provided compounds are inhibitors of PAD4 and are therefore useful for treating one or more disorders associated with activity of PAD4. Thus, in certain embodiments, the present invention provides a method for treating a PAD4-mediated disorder comprising the step of administering to a patient in need thereof a compound of the present invention, or pharmaceutically acceptable composition thereof.
In one embodiment, a PAD4-mediated disorder is a disease, condition, or disorder mediated by inappropriate PAD4 activity. In some embodiments, a PAD4-mediated disorder is selected from the group consisting of rheumatoid arthritis, vasculitis, systemic lupus erythematosus, ulcerative colitis, cancer, cystic fibrosis, asthma, cutaneous lupus erythematosis, and psoriasis. In a further embodiment, the disorder mediated by inappropriate PAD4 activity is rheumatoid arthritis. In a further embodiment, the disorder mediated by inappropriate PAD4 activity is systemic lupus. In a further embodiment, the disorder mediated by inappropriate PAD4 activity is vasculitis. In a further embodiment, the disorder mediated by inappropriate PAD4 activity is cutaneous lupus erythematosis. In a further embodiment, the disorder mediated by inappropriate PAD4 activity is psoriasis.
In one embodiment there is provided a method of treatment of rheumatoid arthritis, vasculitis, systemic lupus erythematosus, ulcerative colitis, cancer, cystic fibrosis, asthma, cutaneous lupus erythematosis, or psoriasis, which method comprises administering to a human subject in need thereof, a therapeutically effective amount of a provided compound or a pharmaceutically acceptable salt thereof.
In one embodiment there is provided a method of treatment of rheumatoid arthritis, which method comprises administering to a human subject in need thereof, a therapeutically effective amount of a provided compound, or a pharmaceutically acceptable salt thereof. In one embodiment there is provided a method of treatment of systemic lupus, which method comprises administering to a human subject in need thereof, a therapeutically effective amount of a provided compound, or a pharmaceutically acceptable salt thereof. In one embodiment there is provided a method of treatment of vasculitis, which method comprises administering to a human subject in need thereof, a therapeutically effective amount of a provided compound, or a pharmaceutically acceptable salt thereof. In one embodiment there is provided a method of treatment of cutaneous lupus erythematosis, which method comprises administering to a human subject in need thereof, a therapeutically effective amount of a provided compound, or a pharmaceutically acceptable salt thereof. In one embodiment there is provided a method of treatment of psoriasis, which method comprises administering to a human subject in need thereof, a therapeutically effective amount of a provided compound, or a pharmaceutically acceptable salt thereof.
In some embodiments, a PAD4-mediated disorder is selected from the group consisting of acid-induced lung injury, acne (PAPA), acute lymphocytic leukemia, acute, respiratory distress syndrome, Addison's disease, adrenal hyperplasia, adrenocortical insufficiency, ageing, AIDS, alcoholic hepatitis, alcoholic hepatitis, alcoholic liver disease, allergen induced asthma, allergic bronchopulmonary, aspergillosis, allergic conjunctivitis, alopecia, Alzheimer's disease, amyloidosis, amyotropic lateral sclerosis, and weight loss, angina pectoris, angioedema, anhidrotic ecodermal dysplasia-ID, ankylosing spondylitis, anterior segment, inflammation, antiphospholipid syndrome, aphthous stomatitis, appendicitis, arthritis, asthma, atherosclerosis, atopic dermatitis, autoimmune diseases, autoimmune hepatitis, bee sting-induced inflammation, behcet's disease, Behcet's syndrome, Bells Palsey, berylliosis, Blau syndrome, bone pain, bronchiolitis, burns, bursitis, cancer, cardiac hypertrophy, carpal tunnel syndrome, catabolic disorders, cataracts, cerebral aneurysm, chemical irritant-induced inflammation, chorioretinitis, chronic heart failure, chronic lung disease of prematurity, chronic lymphocytic leukemia, chronic obstructive pulmonary disease, colitis, complex regional pain syndrome, connective tissue disease, corneal ulcer, crohn's disease, cryopyrin-associated periodic syndromes, cyrptococcosis, cystic fibrosis, deficiency of the interleukin-1-receptor antagonist (DIRA), dermatitis, dermatitis endotoxemia, dermatomyositis, diffuse intrinsic pontine glioma, endometriosis, endotoxemia, epicondylitis, erythroblastopenia, familial amyloidotic polyneuropathy, familial cold urticarial, familial Mediterranean fever, fetal growth retardation, glaucoma, glomerular disease, glomerular nephritis, gout, gouty arthritis, graft-versus-host disease, gut diseases, head injury, headache, hearing loss, heart disease, hemolytic anemia, Henoch-Scholein purpura, hepatitis, hereditary periodic fever syndrome, herpes zoster and simplex, HIV-1, Hodgkin's disease, Huntington's disease, hyaline membrane disease, hyperammonemia, hypercalcemia, hypercholesterolemia, hyperimmunoglobulinemia D with recurrent fever (HIDS), hypoplastic and other anemias, hypoplastic anemia, idiopathic thrombocytopenic purpura, incontinentia pigmenti, infectious mononucleosis, inflammatory bowel disease, inflammatory lung disease, inflammatory neuropathy, inflammatory pain, insect bite-induced inflammation, iritis, irritant-induced inflammation, ischemia/reperfusion, juvenile rheumatoid arthritis, keratitis, kidney disease, kidney injury caused by parasitic infections, kidney injury caused by parasitic infections, kidney transplant rejection prophylaxis, leptospiriosis, leukemia, Loeffler's syndrome, lung injury, lung injury, lupus, lupus, lupus nephritis, lymphoma, meningitis, mesothelioma, mixed connective tissue disease, Muckle-Wells syndrome (urticaria deafness amyloidosis), multiple sclerosis, muscle wasting, muscular dystrophy, myasthenia gravis, myocarditis, mycosis fungiodes, mycosis fungoides, myelodysplastic syndrome, myositis, nasal sinusitis, necrotizing enterocolitis, neonatal onset multisystem inflammatory disease (NOMID), nephrotic syndrome, neuritis, neuropathological diseases, non-allergen induced asthma, obesity, ocular allergy, optic neuritis, organ transplant, osterarthritis, otitis media, paget's disease, pain, pancreatitis, Parkinson's disease, pemphigus, pericarditis, periodic fever, periodontitis, peritoneal endometriosis, pertussis, pharyngitis and adenitis (PFAPA syndrome), plant irritant-induced inflammation, pneumonia, pneumonitis, pneumosysts infection, poison ivy/urushiol oil-induced inflammation, polyarteritis nodosa, polychondritis, polycystic kidney disease, polymyositis, psoriasis, psoriasis, psoriasis, psoriasis, psychosocial stress diseases, pulmonary disease, pulmonary hypertension, pulmonayr fibrosis, pyoderma gangrenosum, pyogenic sterile arthritis, renal disease, retinal disease, rheumatic carditis, rheumatic disease, rheumatoid arthritis, sarcoidosis, seborrhea, sepsis, severe pain, sickle cell, sickle cell anemia, silica-induced disease, Sjogren's syndrome, skin diseases, sleep apnea, solid tumors, spinal cord injury, Stevens-Johnson syndrome, stroke, subarachnoid hemorrhage, sunburn, temporal arteritis, tenosynovitis, thrombocytopenia, thyroiditis, tissue transplant, TNF receptor associated periodic syndrome (TRAPS), toxoplasmosis, transplant, traumatic brain injury, tuberculosis, type 1 diabetes, type 2 diabetes, ulcerative colitis, urticarial, uveitis, and Wegener's granulomatosis.
In one embodiment, the invention provides a provided compound, or a pharmaceutically acceptable salt thereof, for use in therapy. In another embodiment, the invention provides a provided compound, or a pharmaceutically acceptable salt thereof, for use in the treatment of a disorder mediated by inappropriate PAD4 activity. In another embodiment, the invention provides a provided compound, or a pharmaceutically acceptable salt thereof, for use in the treatment of rheumatoid arthritis, vasculitis, systemic lupus erythematosus, ulcerative colitis, cancer, cystic fibrosis, asthma, cutaneous lupus erythematosis, or psoriasis. In another embodiment, the invention provides a provided compound, or a pharmaceutically acceptable salt thereof, for use in the treatment of rheumatoid arthritis. In another embodiment, the invention provides a provided compound, or a pharmaceutically acceptable salt thereof, for use in the treatment of systemic lupus. In another embodiment, the invention provides a provided compound, or a pharmaceutically acceptable salt thereof, for use in the treatment of vasculitis. In another embodiment, the invention provides a provided compound, or a pharmaceutically acceptable salt thereof, for use in the treatment of cutaneous lupus erythematosis. In another embodiment, the invention provides a provided compound, or a pharmaceutically acceptable salt thereof, for use in the treatment of psoriasis. In another embodiment, the invention provides the use of a provided compound, or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for use in the treatment of a disorder mediated by inappropriate PAD4 activity. In another embodiment, the invention provides the use of a provided compound, or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for use in the treatment of rheumatoid arthritis, vasculitis, systemic lupus erythematosus, ulcerative colitis, cancer, cystic fibrosis, asthma, cutaneous lupus erythematosis, or psoriasis. In another embodiment, the invention provides the use of a provided compound, or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for use in the treatment of rheumatoid arthritis. In another embodiment, the invention provides the use of a provided compound, or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for use in the treatment of systemic lupus. In another embodiment, the invention provides the use of a provided compound, or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for use in the treatment of vasculitis. In another embodiment, the invention provides the use of a provided compound, or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for use in the treatment of cutaneous lupus erythematosis. In another embodiment, the invention provides the use of a provided compound, or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for use in the treatment of psoriasis. In a further embodiment, the invention provides a pharmaceutical composition for the treatment or prophylaxis of a disorder mediated by inappropriate PAD4 activity comprising a provided compound, or a pharmaceutically acceptable salt thereof. In a further embodiment, the invention provides a pharmaceutical composition for the treatment or prophylaxis of rheumatoid arthritis, vasculitis, systemic lupus erythematosus, ulcerative colitis, cancer, cystic fibrosis, asthma, cutaneous lupus erythematosis, or psoriasis, comprising a provided compound, or a pharmaceutically acceptable salt thereof. In a further embodiment, the invention provides a pharmaceutical composition for the treatment or prophylaxis of rheumatoid arthritis comprising a provided compound, or a pharmaceutically acceptable salt thereof. In a further embodiment, the invention provides a pharmaceutical composition for the treatment or prophylaxis of systemic lupus comprising a provided compound, or a pharmaceutically acceptable salt thereof. In a further embodiment, the invention provides a pharmaceutical composition for the treatment or prophylaxis of vasculitis comprising a provided compound, or a pharmaceutically acceptable salt thereof. In a further embodiment, the invention provides a pharmaceutical composition for the treatment or prophylaxis of cutaneous lupus erythematosis comprising a provided compound, or a pharmaceutically acceptable salt thereof. In a further embodiment, the invention provides a pharmaceutical composition for the treatment or prophylaxis of psoriasis comprising a provided compound, or a pharmaceutically acceptable salt thereof
All features of each of the aspects of the invention apply to all other aspects mutatis mutandis.
In order that the invention described herein may be more fully understood, the following examples are set forth. It should be understood that these examples are for illustrative purposes only and are not to be construed as limiting this invention in any manner.
As depicted in the Examples below, in certain exemplary embodiments, compounds are prepared according to the following general procedures. It will be appreciated that, although the general methods depict the synthesis of certain compounds of the present invention, the following general methods, and other methods known to one of ordinary skill in the art, can be applied to all compounds and subclasses and species of each of these compounds, as described herein.
The following schemes elucidate the synthesis of example compounds embodied in this application.
For the synthesis of compounds involving photoredox chemistry, see Scheme 6.
For the synthesis of compounds involving photoredox chemistry of cyclic, bicyclic, or tricyclic protected amines followed by deprotection and subsequent coupling see Scheme 7. Note that the cyclic amine used in the photoredox step can also be in fully elaborated form requiring no subsequent deprotection and coupling steps.
For the synthesis of compounds involving photoredox chemistry of cyclic, bicyclic, or tricyclic protected carboxylic acids (for example, esters) followed by deprotection and subsequent coupling see Scheme 8. Note that the cyclic carboxylic acid used in the photoredox step can also be in fully derivatized form requiring no subsequent deprotection and coupling steps.
For the synthesis of compounds involving the Suzuki, Stille or other aromatic cross-coupling reactions, see Scheme 9.
For the synthesis of 1,2,3-triazoles, see Scheme 10.
For the synthesis of Buchwald reaction-type coupled products, see Scheme 11.
And by analogy:
For the synthesis of nitrile coupled products, see Scheme 12. The nitrile can subsequently be converted into many other different functional groups familiar to one skilled in the art.
The following Examples are offered as illustrative, as a partial scope and particular embodiments of the invention and are not meant to be limiting of the scope of the invention. Abbreviations and chemical symbols have their usual and customary meanings unless otherwise indicated. Unless otherwise indicated, the compounds described herein have been prepared, isolated and characterized using the schemes and other methods disclosed herein or may be prepared using the same.
Method 1: Column: Waters XBridge C18, 2.1 mm×50 mm, 1.7 μm particles; Mobile Phase A: 5:95 acetonitrile:water with 10 mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile:water with 10 mM ammonium acetate; Temperature: 50° C.; Gradient: 0% B to 100% B over 3 min, then a 0.75 min hold at 100% B; Flow: 1 mL/min; Detection: MS and UV (220 nm).
Method 2: Column: Waters XBridge C18, 2.1 mm×50 mm, 1.7 μm particles; Mobile Phase A: 5:95 acetonitrile:water with 0.1% c trifluoroacetic acid; Mobile Phase B: 95:5 acetonitrile:water with 0.1% trifluoroacetic acid; Temperature: 50° C.; Gradient: 0% B to 100% B over 3 min, then a 0.75 min hold at 100% B; Flow: 1 mL/min; Detection: MS and UV (220 nm).
Method A: Column: Waters Acquity UPLC BEH C18, 2.1×50 mm, 1.7 μm particles; Mobile Phase A: 5:95 ACN:water with 10 mM NH4OAc; Mobile Phase B: 95:5 ACN:water with 10 mM NH4OAc; Temperature: 50° C.; Gradient: 0-100% B over 3 minutes, then a 0.75 minute hold at 100% B; Flow: 1.11 mL/min; Detection: UV at 220 nm.
Method B: Column: Waters Acquity UPLC BEH C18, 2.1×50 mm, 1.7 μm particles; mobile Phase A: 5:95 ACN:water with 0.1% TFA; Mobile Phase B: 95:5 ACN:water with 0.1% TFA; Temperature: 50° C.; Gradient: 0-100% B over 3 minutes, then a 0.75 minute hold at 100% B; Flow: 1.11 mL/min; Detection: UV at 220 nm.
Method C: Column: PHENOMENEX® Luna 3 μm C18 (2.0×30 mm); mobile Phase A: 10:90 MeOH:water with 0.1% TFA; Mobile Phase B: 90:10 MeOH:water with 0.1% TFA; Gradient: 0-100% B over 2 minutes, then a 1 minute hold at 100% B; Flow: 1 mL/min; Detection: UV at 220 nm.
Method D: Waters Acquity UPLC BEH C18, 2.1×50 mm, 1.7 μm particles; Mobile Phase A: water with 0.05% TFA; Mobile Phase B: ACN with 0.05% TFA; Gradient: 2-98% B over 1 minute, then a 0.5 minute hold at 98% B; Flow: 0.8 mL/min; Detection: UV at 220 or 254 nm.
A stirring mixture of ethyl 7-bromo-1H-indole-2-carboxylate (2.8 g, 10.44 mmol) and potassium carbonate (4.33 g, 31.3 mmol) in DMF (20 mL) was treated with (bromomethyl)cyclopropane (2.026 mL, 20.89 mmol). The reaction was heated to 60° C. and stirred under a nitrogen atmosphere for 4 hours, at which point it was judged to be complete by LCMS. The reaction mixture poured into ethyl acetate (200 mL), and the turbid solution was washed 3× with 100% lithium chloride and once with brine, then dried over sodium sulfate and concentrated in vacuo. The residue was dried under high vacuum to yield ethyl 7-bromo-1-(cyclopropylmethyl)-1H-indole-2-carboxylate (3.36 g, 10.43 mmol, 100% yield). The material was used in the next step without further purification. 1H NMR (499 MHz, chloroform-d) δ 7.65 (dd, J=7.9, 1.0 Hz, 1H), 7.54 (dd, J=7.6, 1.0 Hz, 1H), 7.35 (s, 1H), 7.00 (t, J=7.7 Hz, 1H), 5.12 (d, J=6.9 Hz, 2H), 4.39 (q, J=7.2 Hz, 2H), 1.43 (t, J=7.2 Hz, 3H), 1.38-1.26 (m, 1H), 0.42 (d, 0.1=6.6 Hz, 4H). MS ESI m/z=322 (M+H).
A stirring solution of ethyl 7-bromo-1-(cyclopropylmethyl)-1H-indole-2-carboxylate (5.2 g, 16.14 mmol) in methanol/THF (2:1) (75 mL) was treated with 1M sodium hydroxide (48.4 mL, 48.4 mmol). The reaction was stirred at 50° C. for 18 hours, at which point it was judged to be complete by LCMS. The organic solvents were evaporated, and the remaining aqueous solution was washed twice with diethyl ether, then treated with 1M HCl (50 mL). The turbid mixture was extracted 4× with ethyl acetate, then the combined organic phases were washed once with brine, dried over sodium sulfate, and concentrated in vacuo to yield 7-bromo-1-(cyclopropylmethyl)-1H-indole-2-carboxylic acid (4.64 g, 15.77 mmol, 98% yield) as a colorless solid, which was used in the next step without further purification. 1H NMR (499 MHz, DMSO-d6) δ 13.17 (br s, 1H), 7.74 (dd, J=7.9, 1.0 Hz, 1H), 7.57 (dd, J=7.5, 1.0 Hz, 1H), 7.35 (s, 1H), 7.05 (t, J=7.7 Hz, 1H), 5.03 (d, J=7.0 Hz, 2H), 1.29-1.11 (m, 1H), 0.45-0.35 (m, 2H), 0.34-0.30 (m, 2H). MS ESI m/z=294 (M+H).
A stirring solution of methyl 3-amino-5-methoxy-4-(methylamino)benzoate hydrochloride (4.0 g, 16.21 mmol), 7-bromo-1-(cyclopropylmethyl)-1H-indole-2-carboxylic acid (4.64 g, 15.77 mmol), and Hunig's Base (6.89 mL, 39.4 mmol) in DMF (20 mL) was treated with HATU (7.20 g, 18.93 mmol). The reaction was stirred under a nitrogen atmosphere at room temperature for 18 hours, at which point it was judged to be complete by LCMS based on the disappearance of starting material, and the formation of a single product peak with m/z=487 (M+H for intermediate carboximide). The reaction mixture was concentrated in vacuo, and the residue was taken up in ethyl acetate (350 mL) and water (100 mL). The turbid mixture was stirred for 15 minutes, and the resulting solids were collected by filtration, rinsed thoroughly with ethyl acetate and water, and dried under vacuum to yield 2.7 g of a colorless solid. LCMS of this material detects a single peak with m/z=487. The combined filtrate and rinsings were transferred to a separatory funnel, the layers were separated, and the ethyl acetate phase was washed twice with 10% lithium chloride and once with brine, then dried over sodium sulfate and concentrated in vacuo. The residue and solids from the filtration were suspended in acetic acid (30 mL). The mixture was heated to 75° C., resulting in a homogeneous solution, and the reaction was stirred for 90 minutes, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo, and the residue was taken up in ethyl acetate (200 mL) and water (50 mL). A magnetic stir bar was added to the flask, and the stirring mixture was carefully treated with half-saturated sodium carbonate until gas evolution ceased. The mixture was stirred for another 10 minutes, during which time a colorless solid precipitated. The solids were collected by filtration, rinsed thoroughly with water, ethyl acetate, and methanol, and then dried under vacuum to yield 4.42 g of the title compound as a colorless solid. The combined filtrate and rinsings were transferred to a separatory funnel, and the layers were separated. The aqueous phase was extracted twice with ethyl acetate (75 mL), then the organic phases were combined, washed once with saturated sodium carbonate, once with water, and once with brine, then dried over sodium sulfate, and concentrated in vacuo. The residue was taken up in boiling methanol (100 mL), and the mixture was stirred for 10 minutes, then allowed to cool to room temperature. The resulting solids were collected by filtration, rinsed 3× with methanol, and dried under vacuum to yield 1.50 g of the title compound as a slightly amber solid. Combined, the two crops yielded methyl 2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylate (5.9 g, 12.60 mmol, 80% yield). 1H NMR (499 MHz, DMSO-d6) δ 7.98 (d, 0.1=1.2 Hz, 1H), 7.75 (dd, 0.1=7.9, 0.8 Hz, 1H), 7.55 (dd, J=7.5, 1.0 Hz, 1H), 7.41 (d, J=1.2 Hz, 1H), 7.17 (s, 1H), 7.09 (t, J=7.7 Hz, 1H), 4.76 (d, J=7.0 Hz, 2H), 4.09 (s, 3H), 4.04 (s, 3H), 3.90 (s, 3H), 1.09-0.99 (m, 1H), 0.30-0.22 (m, 2H), −0.14-−0.21 (m, 2H). MS ESI m/z=468 (M+H).
A stirring solution of methyl 2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylate (1.47 g, 3.14 mmol) in 1:1 methanol/THF (10 mL) was treated with 1 M sodium hydroxide (9.42 mL, 9.42 mmol). The reaction was stirred at 50° C. for three hours, at which point it was judged to be complete by LCMS. The organic solvents were removed on the rotary evaporator, and the remaining aqueous suspension was adjusted to pH 5 with 1M HCl. The heterogeneous mixture was vigorously stirred for 20 minutes, then extracted 3× with ethyl acetate (100 mL). The combined organic phases were washed once with brine, dried over sodium sulfate and concentrated in vacuo to yield 2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylic acid (1.34 g, 2.95 mmol, 94% yield) as an off-white solid, which was used in the next step without further purification. 1H NMR (499 MHz, CHLOROFORM-d) δ 7.94 (d, J=1.2 Hz, 1H), 7.74 (dd, J=7.9, 0.8 Hz, 1H), 7.54 (dd, J=7.6, 0.9 Hz, 1H), 7.40 (d, J=1.1 Hz, 1H), 7.15 (s, 1H), 7.08 (t, J=7.7 Hz, 1H), 4.75 (d, J=6.9 Hz, 2H), 4.08 (s, 3H), 4.02 (s, 3H), 1.07-0.98 (m, 1H), 0.29-0.21 (m, 2H), −0.16-−0.22 (m, 2H). MS ESI m/z=454 (M+H).
A stirring solution of tert-butyl ((1R,4R,7R)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (0.689 g, 3.24 mmol), 2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylic acid (1.34 g, 2.95 mmol), and triethylamine (1.233 mL, 8.85 mmol) in DMF (20 mL) was treated with BOP (1.370 g, 3.10 mmol). The reaction was stirred at room temperature for 4 hours, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo, and the residue was taken up in ethyl acetate (150 mL). The turbid solution was washed 3× with water, 3× with 1 M sodium hydroxide, and once with brine, then dried over sodium sulfate and concentrated in vacuo. The residue was chromatographed via MPLC over a 120 g silica gel column, eluting at 85 mL/min with a 0% to 10% methanol/dichloromethane gradient over 15 column volumes, holding at 4% methanol as the product eluted. Fractions containing the desired product were pooled and concentrated in vacuo to yield tert-butyl ((1R,4R,7R)-2-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (1.86 g, 2.87 mmol, 97% yield) as a white solid. 1H NMR (499 MHz, chloroform-d) δ 7.70-7.62 (m, 1H), 7.57-7.47 (m, 2H), 7.08-6.99 (m, 2H), 6.88-6.80 (m, 1H), 4.87-4.70 (m, 2H), 4.68-4.46 (m, 1H), 4.36 (br s, 1H), 4.16-4.09 (m, 3H), 4.04 (s, 3H), 3.90-3.72 (m, 2H), 3.31-3.20 (m, 1H), 2.54 (br s, 1H), 2.11-1.83 (m, 3H), 1.77-1.68 (m, 1H), 1.52-1.35 (m, 9H), 1.20-1.07 (m, 1H), 0.37-0.23 (m, 2H), −0.02-−0.25 (m, 2H). MS ESI m/z=648 (M+H).
A stirring solution of tert-butyl ((1R,4R,7R)-2-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (1.6 g, 2.467 mmol) in dichloromethane (10 mL) was treated with 4M HCl in dioxane (10 mL). The reaction was stirred at room temperature for 1 hour, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo, and the residue was taken up in water (15 mL). The solution was treated carefully with saturated sodium carbonate until further addition failed to produce gas evolution. The turbid solution was extracted 3× with ethyl acetate, then the combined organic phases were washed once with brine, dried over sodium sulfate, and concentrated in vacuo to yield ((1R,4R,7R)-7-amino-2-azabicyclo[2.2.1]heptan-2-yl)(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-5-yl)methanone (1.23 g, 2.243 mmol, 91% yield) as a colorless solid. 1H NMR (499 MHz, chloroform-d) δ 7.70-7.62 (m, 1H), 7.58-7.49 (m, 2H), 7.08-7.00 (m, 2H), 6.88-6.80 (m, 1H), 4.87-4.71 (m, 2H), 4.17-4.10 (m, 3H), 4.08-4.00 (m, 4H), 3.81-3.67 (m, 1H), 3.52-3.34 (m, 1H), 3.33-3.17 (m, 1H), 2.39-2.24 (m, 1H), 2.16-1.90 (m, 3H), 1.69-1.59 (m, 1H), 1.20-1.09 (m, 1H), 0.35-0.24 (m, 2H), −0.08-−0.21 (m, 2H). MS ESI m/z 548 (M+H). Anal. HPLC Retention time: 0.82 minutes, Method D.
In a 2-dram vial, a stirring mixture of 3-bromo-1-propanol (10.05 μl, 0.116 mmol), tert-butyl (2-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (50 mg, 0.077 mmol), tris(trimethylsilyl)silane (0.036 mL, 0.116 mmol), Ir(dF(CF3)ppy)2(dtbbpy)PF6 (4.32 mg, 3.85 μmol), and sodium carbonate (32.7 mg, 0.308 mmol) in 1,4-Dioxane (2 mL) was degassed with bubbling nitrogen for 10 minutes. In a second vial, a stirring mixture of nickel(II) chloride ethylene glycol dimethyl ether complex (4.23 mg, 0.019 mmol), and 4,4′-di-tert-butyl-2,2′-bipyridine (6.21 mg, 0.023 mmol) in 1,4-Dioxane (1 mL) was degassed with nitrogen for 20 minutes. The nickel complex was transferred via syringe to the first vial, and the mixture was degassed with bubbling nitrogen for an additional 5 minutes. The vial was sealed, and the reaction was stirred at room temperature under two blue Kessil lamps for 18 hours, at which point it was judged to be complete by LCMS based on the disappearance of starting material. The reaction mixture was diluted with dichloromethane (3 mL). Solids were removed by filtration and rinsed with dichloromethane, and the combined filtrates and rinsings were treated with 4 M HCl in dioxane (2 mL). The reaction was stirred at room temperature for 1 hour, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo. The crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 20% B, 20-60% B over 20 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS and UV signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield ((1R,4R,7R)-7-amino-2-azabicyclo [2.2.1]heptan-2-yl)(2-(1-(cyclopropylmethyl)-7-(3-hydroxypropyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-5-yl)methanone (19.9 mg, 0.036 mmol, 47.0% yield). 1H NMR (500 MHz, DMSO-d6) δ 7.34 (br d, J=7.0 Hz, 1H), 7.26-7.12 (m, 1H), 6.92-6.85 (m, 2H), 6.82 (s, 1H), 6.77-6.68 (m, 1H), 4.26 (br d, J=5.8 Hz, 2H), 3.92-3.84 (m, 3H), 3.78 (s, 3H), 3.59-3.45 (m, 3H), 3.34 (br t, J=6.0 Hz, 2H), 2.94-2.82 (m, 3H), 2.15-1.97 (m, 1H), 1.87-1.44 (m, 6H), 1.39-1.14 (m, 1H), 0.71-0.52 (m, 1H), 0.01 (br d, J=7.6 Hz, 2H), −0.43-−0.69 (m, 2H). MS ESI m/z 528 (M+H). Anal. HPLC Retention time: 1.56 minutes, Method 1.
The following compounds in Table 1 can be made by the procedures described in Example 2, substituting the appropriate alkyl halide for 3-bromo-1-propanol. For examples where the substituent at the indole 7-position contains a basic amine, the appropriate Boc-protected aminoalkyl halide was used, and the Boc-group was cleaved during the final deprotection step.
In a 1-dram vial, a stirring mixture of nickel(II) chloride ethylene glycol dimethyl ether complex (10.31 mg, 0.047 mmol) and 4,4′-di-tert-butyl-2,2′-bipyridine (15.11 mg, 0.056 mmol) in 1,4-Dioxane (2 mL) was degassed with bubbling nitrogen for 20 minutes. In a separate 20 mL scintillation vial, a stirring mixture of methyl 2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylate (293 mg, 0.626 mmol), tert-butyl 3-iodoazetidine-1-carboxylate (354 mg, 1.251 mmol), Ir(dF(CF3)ppy)2(dtbbpy)PF6 (10.53 mg, 9.38 μmol), tris(trimethylsilyl)silane (0.290 mL, 0.938 mmol), and sodium carbonate (265 mg, 2.502 mmol) in 1,4-Dioxane (10 mL) was degassed with nitrogen for 10 minutes. The nickel complex was transferred to the vial containing the reaction mixture, the vial was sealed, and the reaction was stirred at room temperature under a blue Kessil lamp for 60 hours, at which point it was judged to be complete by LCMS based on the disappearance of starting material. The reaction mixture was diluted with dichloromethane (20 mL). Solids were removed by filtration and rinsed with dichloromethane, and the combined filtrates and rinsings were concentrated in vacuo. The residue was chromatographed via MPLC over a 40 g silica gel column, eluting at 40 mL/min with a 0% to 5% methanol/dichloromethane gradient over 20 column volumes. Fractions containing the major peak were pooled and concentrated in vacuo to yield methyl 2-(7-(1-(tert-butoxycarbonyl)azetidin-3-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylate (278 mg, 0.510 mmol, 82% yield) as a colorless solid. 1H NMR (499 MHz, chloroform-d) δ 8.18 (d, J=1.2 Hz, 11H), 7.65 (dd, J=7.8, 0.8 Hz, 1H), 7.51-7.43 (m, 2H), 7.27 (t, J=7.7 Hz, 1H), 6.90 (s, 1H), 4.68-4.56 (m, 1H), 4.50-4.41 (m, 4H), 4.27 (br s, 2H), 4.17 (s, 3H), 4.07 (s, 3H), 3.98 (s, 3H), 3.73 (s, 2H), 1.50 (s, 9H), 1.47-1.43 (m, 2H), 0.88-0.79 (m, 1H), 0.30-0.24 (m, 2H), −0.26 (br s, 2H). MS ESI m/z=545 (M+H).
A stirring solution of methyl 2-(7-(1-(tert-butoxycarbonyl)azetidin-3-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylate (0.39 g, 0.716 mmol) in dichloromethane (4 mL) was treated with TFA (0.4 mL, 5.19 mmol). The reaction was stirred at room temperature for 1 hour, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo, and the residue was taken up in dichloromethane. The mixture was treated with triethylamine (0.399 mL, 2.86 mmol) and stirred for 5 minutes, then treated with acetic anhydride (0.068 mL, 0.716 mmol). The reaction was stirred at room temperature for 1 hours, at which point it was judged to be complete by LCMS. The mixture was treated with methanol and concentrated in vacuo, and residue was chromatographed via MPLC over a 40 g silica gel column, eluting at 40 mL/min with a 0% to 2% methanol/dichloromethane gradient over 5 column volumes, then 2% methanol/dichloromethane for 7 column volumes as impurities eluted, then a 2-3.5% methanol/dichloromethane gradient over 3 column volumes, then 3.5% methanol/dichloromethane to fully elute the desired product. Fractions containing the desired product were pooled and concentrated in vacuo to yield methyl 2-(7-(1-acetylazetidin-3-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylate (270 mg, 0.555 mmol, 77% yield) as a colorless solid. 1H NMR (499 MHz, chloroform-d) δ 8.18 (d, J=1.2 Hz, 1H), 7.67 (dd, J=7.8, 0.8 Hz, 1H), 7.48 (d, J=1.2 Hz, 1H), 7.44 (d, J=7.3 Hz, 1H), 7.28-7.26 (m, 1H), 6.91 (s, 1H), 4.74-4.63 (m, 2H), 4.61-4.31 (m, 5H), 4.18 (s, 3H), 4.07 (s, 3H), 3.99 (s, 3H), 1.97 (s, 3H), 0.88-0.79 (m, 1H), 0.36-0.21 (m, 2H), −0.16 (dq, J=9.7, 4.9 Hz, 1H), −0.35 (dq, J=9.7, 4.9 Hz, 1H). MS ESI m/z=487 (M+H).
A stirring solution of methyl 2-(7-(1-acetylazetidin-3-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylate (105 mg, 0.216 mmol) in methanol (2 mL) was treated with 1M sodium hydroxide (0.259 mL, 0.259 mmol), and the reaction was stirred at room temperature for 18 hours, at which point it was judged to be complete by LCMS. The methanol was evaporated, and the remaining aqueous mixture was diluted with water (4 mL) and treated with 1 M HCl (0.26 mL). The heterogeneous mixture was extracted 5× with dichloromethane, and the combined organic phases were dried over sodium sulfate and concentrated in vacuo to yield 2-(7-(1-acetylazetidin-3-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylic acid (70 mg, 0.148 mmol, 68.6% yield) as a colorless solid. 1H NMR (499 MHz, DMSO-d6) δ 13.12-12.21 (m, 1H), 7.95-7.92 (m, 1H), 7.66-7.61 (m, 1H), 7.48 (d, J=7.0 Hz, 1H), 7.41 (d, J=1.2 Hz, 1H), 7.22 (t, J=7.6 Hz, 1H), 7.11 (s, 1H), 4.72-4.60 (m, 2H), 4.50-4.33 (m, 4H), 4.15-4.06 (m, 4H), 4.03 (s, 3H), 1.85 (s, 3H), 0.86-0.73 (m, 1H), 0.28-0.16 (m, 2H), −0.24-−0.41 (m, 2H). MS ESI m/z=473 (M+H).
A stirring solution of tert-butyl ((1R,4R,7R)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (47.2 mg, 0.222 mmol), 2-(7-(1-acetylazetidin-3-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylic acid (70 mg, 0.148 mmol), and triethylamine (0.062 mL, 0.444 mmol) in dichloromethane (2 mL) was treated with BOP (79 mg, 0.178 mmol). The reaction was stirred at room temperature for 18 hours, at which point it was judged to be complete by LCMS. The mixture was diluted with dichloromethane (10 mL), and the solution was washed twice with 1M HCl, twice with 1M NaOH, and once with brine, then dried over sodium sulfate and concentrated in vacuo. The residue was chromatographed via MPLC over a 24 g silica gel column, eluting at 40 mL/min with a 0% to 7% methanol/dichloromethane gradient over 13 column volumes. Fractions containing the desired product were pooled and concentrated in vacuo to yield tert-butyl ((1R,4R,7R)-2-(2-(7-(1-acetylazetidin-3-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate. 1H NMR (499 MHz, chloroform-d) δ 7.67 (d, J=7.7 Hz, 1H), 7.54-7.47 (m, 1H), 7.44 (d, J=7.4 Hz, 1H), 7.28-7.23 (m, 1H), 7.06-6.98 (m, 1H), 6.93-6.87 (m, 1H), 4.75-4.62 (m, 2H), 4.59-4.29 (m, 7H), 4.16 (s, 3H), 4.04 (d, J=1.0 Hz, 3H), 3.92-3.72 (m, 2H), 3.28 (dd, J=11.3, 1.9 Hz, 1H), 2.54 (br s, 1H), 1.97 (d, J=2.6 Hz, 5H), 1.61-1.35 (m, 10H), 0.96-0.76 (m, 2H), 0.38-0.18 (m, 2H), −0.06-−0.46 (m, 2H). MS ESI m/z=667 (M+H).
A stirring solution of tert-butyl ((1R,4R,7R)-2-(2-(7-(1-acetylazetidin-3-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (79 mg, 0.118 mmol) in dichloromethane (2 mL) was treated with TFA (0.5 mL). The reaction was stirred at room temperature for 3 hours, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo, and the residue was taken up in water (3 mL). The solution was treated with saturated sodium bicarbonate (5 mL), and the homogeneous solution was stirred for 10 minutes. The mixture was extracted 4× with ethyl acetate (5 mL) (failed to fully extract the product), then 3× with 9:1 dichloromethane/methanol (completely extracted the product). All organic phases were combined and dried over sodium sulfate, then concentrated in vacuo. The residue was chromatographed via MPLC over a 24 g silica gel column, eluting at 40 mL/min with a 3% to 10% methanol/dichloromethane gradient over 10 column volumes, then with 10% methanol/dichloromethane to completely elute the product. Fractions containing the desired product were pooled and concentrated in vacuo. The residue was taken up in 5:1 dichloromethane/methanol, and treated with Si-pyridine resin. The mixture was shaken for 4 hours so the resin could remove any trace metal contaminants, then the resin was removed by filtration and rinsed with 5:1 dichloromethane/methanol. The combined filtrate and rinsings were concentrated in vacuo, and the residue was taken up in 2:1 acetonitrile/water. The solution was freeze dried to yield 1-(3-(2-(5-((1R,4R,7R)-7-amino-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)azetidin-1-yl)ethan-1-one (51 mg, 0.088 mmol, 74.4% yield) as a colorless solid. 1H NMR (499 MHz, chloroform-d) δ 7.67 (d, J=7.7 Hz, 1H), 7.54-7.50 (m, 1H), 7.44 (d, 0.1=7.4 Hz, 1H), 7.28-7.25 (m, 1H), 7.04-7.00 (m, 1H), 6.92-6.87 (m, 1H), 4.74-4.62 (m, 2H), 4.54 (br t, J=8.8 Hz, 1H), 4.50-4.33 (m, 4H), 4.19-4.15 (m, 3H), 4.04 (s, 4H), 3.79-3.66 (m, 1H), 3.52-3.34 (m, 1H), 3.31-3.16 (m, 1H), 2.41-2.25 (m, 1H), 2.14-1.89 (m, 6H), 1.76-1.62 (m, 2H), 1.31-0.76 (m, 2H), 0.37-0.20 (m, 2H), −0.11-−0.24 (m, 1H), −0.28-−0.44 (m, 1H). MS ESI m/z=567.2 (M+H). HPLC retention time 0.66 minutes, Method D.
The following compounds in Table 2 can be made by the procedures described in Example 37, substituting tert-butyl 3-iodopiperidine-1-carboxylate for tert-butyl 3-iodoazetidine-1-carboxylate in step 1, and the appropriate anhydride or isocyanate for acetic anhydride in step 2.
The title compound was prepared via the same procedures used to prepare Example 37, substituting methyl chloroformate for acetic anhydride in step 2. 1H NMR (499 MHz, chloroform-d) δ 7.65 (d, J=7.7 Hz, 1H), 7.54-7.50 (m, 1H), 7.46 (d, J=7.4 Hz, 1H), 7.27-7.24 (m, 1H), 7.05-6.99 (m, 1H), 6.90-6.85 (m, 1H), 4.68 (quin, J=7.4 Hz, 1H), 4.52 (t, J=8.5 Hz, 2H), 4.46-4.40 (m, 2H), 4.32 (br d, J=4.4 Hz, 2H), 4.18-4.13 (m, 3H), 4.06-3.98 (m, 4H), 3.79-3.65 (m, 4H), 3.50-3.34 (m, 1H), 3.31-3.16 (m, 1H), 2.40-2.23 (m, 1H), 2.14-1.88 (m, 3H), 1.70-1.60 (m, 2H), 0.92-0.77 (m, 1H), 0.33-0.22 (m, 2H), −0.17-−0.37 (m, 2H). MS ESI m/z=583.6 (M+H). HPLC retention time 0.80 minutes, Method D.
The title compound was prepared via the same procedures used to prepare Example 37, substituting tert-butyl ((3R, 5R)-5-fluoropiperidin-3-yl)carbamate for ((1R,4R,7R)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate in step 4. 1H NMR (500 MHz, DMSO-d6) δ 7.39 (d, J=7.7 Hz, 1H), 7.21 (d, J=7.3 Hz, 1H), 7.07 (s, 1H), 6.98 (t, J=7.6 Hz, 1H), 6.81 (s, 1H), 6.61 (s, 1H), 4.77-4.57 (m, 1H), 4.49-4.35 (m, 2H), 4.23-4.06 (m, 4H), 3.84 (s, 4H), 3.74 (s, 3H), 3.17 (br s, 6H), 2.80 (br t, 0.1=10.8 Hz, 1H), 1.95 (br t, J=11.1 Hz, 1H), 1.61 (s, 3H), 1.42-1.21 (m, 1H), 0.62-0.50 (m, 1H), −0.01 (br d, J=8.1 Hz, 2H), −0.51 (br dd, J=8.8, 4.7 Hz, 2H). MS ESI m/z=572.9 (M+H). HPLC retention time 1.48 minutes, Method 1.
The title compound was prepared via the same procedures used to prepare Example 37, substituting tert-butyl ((3R,6S)-6-methylpiperidin-3-yl)carbamate for ((1R,4R,7R)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate in step 4. 1H NMR (500 MHz, DMSO-d6) δ 7.43 (d, J=7.6 Hz, 1H), 7.25 (br d, 0.1=7.6 Hz, 1H), 7.06 (s, 1H), 7.01 (t, J=7.6 Hz, 1H), 6.86 (s, 1H), 6.61 (s, 1H), 4.50-4.38 (m, 2H), 4.25-4.10 (m, 4H), 3.87 (s, 4H), 3.78 (s, 3H), 3.37-3.21 (m, 1H), 2.63-2.43 (m, 2H), 1.68 (s, 3H), 1.64 (s, 3H), 1.58-1.41 (m, 2H), 1.32 (br d, J=8.2 Hz, 2H), 1.00 (br d, J=6.7 Hz, 3H), 0.57 (br d, J=5.8 Hz, 1H), 0.01 (br d, J=7.9 Hz, 2H), −0.44-−0.59 (m, 2H). MS ESI m/z=569.1 (M+H). HPLC retention time 1.49 minutes, Method 1.
In a 40 mL scintillation vial, a stirring solution of methyl 3-hydroxycyclobutane-1-carboxylate (834 mg, 6.41 mmol), Ts-Cl (1833 mg, 9.61 mmol), and triethylamine (1.786 mL, 12.82 mmol) in dichloromethane (10 mL) was treated with DMAP (78 mg, 0.641 mmol). The vial was sealed, and the reaction was stirred at room temperature for 18 hours. TLC (50% EtOAc/hexane, UV, KMnO4) indicated that the reaction was complete. The reaction mixture was concentrated onto celite and chromatographed via MPLC over an 80 g silica gel column, eluting at 60 mL/min with a 0% to 50% acetone/hexanes gradient over 15 column volumes. Fractions containing the desired product were pooled and concentrated in vacuo to yield methyl 3-(tosyloxy) cyclobutane-1-carboxylate (1.55 g, 5.45 mmol, 85% yield) as a colorless oil. 1H NMR (499 MHz, chloroform-d) δ 7.83-7.75 (m, 2H), 7.36 (d, J=8.0 Hz, 2H), 4.81-4.72 (m, 1H), 3.68 (s, 3H), 2.69-2.59 (m, 1H), 2.56-2.39 (m, 7H).
A mixture of methyl 3-(tosyloxy)cyclobutane-1-carboxylate (1.1 g, 3.87 mmol) and sodium iodide (2.320 g, 15.48 mmol) in 2-butanone (4 mL) was stirred at 100° C. for 18 hours. The mixture was allowed to come to room temperature and diluted with dichloromethane (25 mL), and the resulting solids were removed by filtration. The residue was chromatographed via MPLC over a 24 g silica gel column, eluting at 40 mL/min with a 0% to 70% ethyl acetate/hexanes gradient over 15 column volumes. The cis- and trans-isomers were partially separated on the column, but all fractions were combined to yield methyl 3-iodocyclobutane-1-carboxylate (805 mg, 3.35 mmol, 87% yield) as a colorless oil. 1H NMR (499 MHz, chloroform-d) δ 4.73-4.34 (m, 1H), 3.76-3.65 (m, 3H), 3.49-3.10 (m, 1H), 3.02-2.89 (m, 31H), 2.89-2.73 (m, 1H).
The title compound was prepared from methyl 3-iodocyclobutane-1-carboxylate and tert-butyl ((1R,4R,7R)-2-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate using the procedure described in Example 37, step 1. MS ESI m/z=682.3 (M+H). HPLC retention time 0.94 and 0.95 minutes, Method D.
In a 2-dram vial, a solution of methyl 3-(2-(5-((1R,4R,7R)-7-((tert-butoxycarbonyl)amino)-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)cyclobutane-1-carboxylate (213 mg, 0.312 mmol) was treated with 1 M sodium hydroxide (0.937 mL, 0.937 mmol). The vial was sealed, and the reaction was stirred at 60° C. for 18 hours, at which point it was judged to be complete by LCMS. The mixture was diluted with water (3 mL), and the methanol was evaporated. The remaining, cloudy solution was washed 3× with ethyl acetate (the layers separated very slowly, so this took two days). LCMS of the combined washings detected some of the desired product. The washings were extracted 3× with 1M NaOH (2 mL) (a yellow, amorphous material settled to the bottom of the vial during the first extraction—this was removed and combined with the combined aqueous phases from the initial workup and these combined sodium hydroxide extractions. All aqueous phases were combined and the mixture was acidified to pH 3 with 1M HCl. The mixture was treated with pH 7 buffer (5 mL), and extracted 5× with ethyl acetate. The combined organic phases were dried over sodium sulfate and concentrated in vacuo to yield 3-(2-(5-((1R,4R,7R)-7-((tert-butoxycarbonyl)amino)-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)cyclobutane-1-carboxylic acid (103 mg, 0.154 mmol, 49.4% yield) as an amber solid, which was used in the next step without further purification. MS ESI m/z=668.3 (M+H). HPLC retention time 0.86 and 0.87 minutes, Method D.
A stirring mixture of 3-(2-(5-((1R,4R,7R)-7-((tert-butoxycarbonyl)amino)-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)cyclobutane-1-carboxylic acid (30 mg, 0.045 mmol), methanamine hydrochloride (6.07 mg, 0.090 mmol), and triethylamine (0.025 mL, 0.180 mmol) in DMF (2 mL) was treated with BOP (23.84 mg, 0.054 mmol). The reaction was stirred at room temperature for 18 hours, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo, and the residue was taken up in dichloromethane (2 mL). The mixture was treated with 4M HCl in dioxane (2 mL), and the reaction was stirred at room temperature for 1 hour, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo, and the crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 19% B, 19-45% B over 35 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS and UV signals. Two isomers were separated, and each was handled separately for the remainder of the process. Fractions were dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield:
First Eluting: 3-(2-(5-((1R,4R,7R)-7-amino-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)-N-methylcyclobutane-1-carboxamide, ISOMER 1 (8.2 mg, 0.014 mmol, 31.4% yield). 1H NMR (500 MHz, DMSO-d6) δ 7.60 (br d, J=4.3 Hz, 1H), 7.37 (d, J=7.6 Hz, 1H), 7.29-7.13 (m, 1H), 7.07 (d, J=7.3 Hz, 1H), 7.02-6.94 (m, 1H), 6.86 (s, 1H), 6.79-6.71 (m, 1H), 4.29 (br d, J=6.4 Hz, 2H), 3.97-3.55 (m, 7H), 3.31-2.95 (m, 1H), 2.93-2.79 (m, 2H), 2.41 (d, J=4.3 Hz, 3H), 2.36 (s, 4H), 2.27-2.16 (m, 2H), 2.06-1.92 (m, 1H), 1.86-1.72 (m, 2H), 1.60-1.44 (m, 1H), 1.30-1.11 (m, 11H), 0.65-0.50 (m, 1H), 0.01 (br d, J=7.6 Hz, 2H), −0.48-−0.63 (m, 2H). MS ESI m/z=581.1 (M+H). HPLC retention time 1.31 minutes, Method 2.
Second Eluting: 3-(2-(5-((1R,4R,7R)-7-amino-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)-N-methylcyclobutane-1-carboxamide, ISOMER 2 (5.9 mg, 9.84 μmol, 21.91% yield). 1H NMR (500 MHz, DMSO-d6) δ 7.63-7.53 (m, 1H), 7.39 (d, J=7.6 Hz, 1H), 7.30-7.14 (m, 2H), 6.99 (t, J=7.6 Hz, 1H), 6.87 (s, 1H), 6.81-6.72 (m, 1H), 4.23 (br d, J=6.7 Hz, 2H), 4.16-4.06 (m, 1H), 3.96-3.87 (m, 3H), 3.83-3.69 (m, 3H), 3.62-3.31 (m, 1H), 3.04-2.82 (m, 2H), 2.51-2.41 (m, 4H), 2.36 (s, 3H), 2.29-2.17 (m, 2H), 2.08-1.93 (m, 1H), 1.87-1.73 (m, 2H), 1.65-1.51 (m, 1H), 1.33-1.14 (m, 1H), 0.64-0.47 (m, 1H), 0.01 (br d, J=7.6 Hz, 2H), −0.45-−0.59 (m, 2H). MS ESI m/z=581.0 (M+H). HPLC retention time 1.34 minutes, Method 2.
The following compounds in Table 3 can be made by the procedures described in Example 43, substituting the appropriate amine for methanamine hydrochloride in step 2. Examples 55 and 56 resulted from unreacted starting material from step 5 in the reaction which gave Examples 45 and 46 respectively carried through the subsequent deprotection step.
A stirring mixture of 3-aminocyclobutan-1-ol (170 mg, 1.951 mmol) and triethylamine (0.816 mL, 5.85 mmol) in dichloromethane (5 mL) was cooled to 0° C. and treated with acetic anhydride (0.184 mL, 1.951 mmol). The reaction was allowed to come to room temperature and stirred for 7 days. The mixture was concentrated in vacuo, and the residue was taken up in dichloromethane (5 mL). The turbid solution was treated with tosyl-Cl (409 mg, 2.145 mmol), and triethylamine (0.544 mL, 3.90 mmol), followed by DMAP (11.91 mg, 0.098 mmol). The reaction was stirred at room temperature for 18 hours. The reaction mixture was injected onto a 24 g silica gel column, and chromatographed via MPLC eluting at 40 mL/min with a 0% to 10% methanol/dichloromethane gradient over 14 column volumes. Fractions containing the desired product were pooled and concentrated in vacuo to yield 3-acetamidocyclobutyl 4-methylbenzenesulfonate (226 mg, 0.798 mmol, 40.9% yield) as a colorless solid, which was used in the next step without further purification. 1H NMR (499 MHz, chloroform-d) δ 7.83-7.76 (m, 2H), 7.39-7.34 (m, 2H), 5.71 (br d, J=4.9 Hz, 1H), 4.51 (quin, J=7.2 Hz, 1H), 4.07-3.97 (m, 1H), 2.81-2.69 (m, 2H), 2.47 (s, 3H), 2.12-2.02 (m, 2H), 1.95 (s, 3H). MS ESI m/z=284.0 (M+H). HPLC retention time 0.74 minutes, Method D.
The title compound was prepared via the procedure used in Example 43, step 2, substituting 3-acetamidocyclobutyl 4-methylbenzenesulfonate for methyl 3-(tosyloxy)cyclobutane-1-carboxylate. 1H NMR (500 MHz, CHLOROFORM-d) (NMR indicates a mixture of isomers) δ 6.00-5.77 (m, 0.6H), 5.76-5.59 (m, 0.4H), 4.87-4.76 (m, 0.3H), 4.56-4.32 (m, 1H), 4.11 (tt, J=9.2, 7.3 Hz, 0.7H), 3.27-3.07 (m, 1.3H), 2.94-2.75 (m, 0.7H), 2.71-2.58 (m, 0.7H), 2.56-2.40 (m, 1.3H), 1.99-1.97 (m, 3H).
The title compound was prepared from N-(3-iodocyclobutyl)acetamide and ((1R,4R,7R)-7-amino-2-azabicyclo[2.2.1]heptan-2-yl)(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-5-yl)methanone via the procedure described in Example 2. 1H NMR (500 MHz, DMSO-d6) (Proton count is low due to the water suppression algorithm used during data processing) S 8.11-7.86 (m, 1H), 7.35 (t, J=8.0 Hz, 1H), 7.18-7.12 (m, 1H), 7.08-6.92 (m, 1H), 6.81 (d, J=5.0 Hz, 1H), 6.71 (br s, 1H), 4.26 (br d, J=6.3 Hz, 1H), 4.18 (br d, J=6.1 Hz, 1H), 4.15-4.05 (m, 1H), 3.96 (br dd, J=8.8, 4.1 Hz, 1H), 3.86 (s, 3H), 3.78 (s, 2H), 3.63-3.51 (m, 1H), 3.44-3.27 (m, 2H), 2.77 (br d, J=6.3 Hz, 1H), 2.54 (br d, J=7.9 Hz, 1H), 2.36 (br d, J=6.1 Hz, 1H), 2.05-1.90 (m, 2H), 1.85-1.71 (m, 2H), 1.65-1.48 (m, 4H), 1.30-1.10 (m, 1H), 0.62-0.44 (m, 1H), 0.06-−0.09 (m, 2H), −0.46-−0.62 (m, 2H). MS ESI m/z=581.2 (M+H). HPLC retention time 1.42 minutes, Method 2.
Examples 58 and 59: The two isomers of Example 57 were resolved using the following conditions:
Isomer 1 (first-eluting): 1H NMR (500 MHz, DMSO-d6) δ 8.00 (br d, J=7.6 Hz, 1H), 7.37 (br d, J=7.6 Hz, 1H), 7.30-7.15 (m, 1H), 7.06 (br d, J=7.3 Hz, 1H), 6.98 (br t, J=7.3 Hz, 1H), 6.86 (s, 1H), 6.80-6.72 (m, 1H), 4.31 (br d, J=6.1 Hz, 2H), 4.23-4.10 (m, 1H), 3.90 (br s, 3H), 3.81 (s, 3H), 3.67-3.57 (m, 1H), 3.53-3.15 (m, 1H), 3.04-2.85 (m, 1H), 2.81 (s, 1H), 2.55 (br d, J=6.7 Hz, 2H), 2.12-1.92 (m, 3H), 1.87-1.74 (m, 2H), 1.62 (s, 3H), 1.28 (br s, 1H), 1.05 (br s, 3H), 0.72-0.51 (m, 2H), 0.01 (br d, 0.1=7.6 Hz, 2H), −0.42-−0.70 (m, 2H). MS ESI m/z=581.2 (M+H). HPLC retention time 1.41 minutes, Method 2.
Isomer 2 (second-eluting): 1H NMR (500 MHz, DMSO-d6) δ 8.15 (br d, J=7.0 Hz, 1H), 7.40 (br d, J=7.6 Hz, 1H), 7.30-7.15 (m, 2H), 6.99 (br t, J=7.5 Hz, 1H), 6.88 (s, 1H), 6.81-6.72 (m, 1H), 4.22 (br d, J=6.1 Hz, 2H), 4.17-4.09 (m, 1H), 4.07-3.95 (m, 1H), 3.91 (br s, 3H), 3.81 (s, 3H), 3.69-3.48 (m, 1H), 3.42-3.31 (m, 1H), 2.90 (br d, J=10.7 Hz, 1H), 2.82 (s, 1H), 2.37-2.24 (m, 1H), 2.14-2.04 (m, 1H), 1.89-1.74 (m, 2H), 1.68-1.54 (m, 4H), 1.29 (br d, J=7.6 Hz, 1H), 1.06 (br s, 4H), 0.73-0.52 (m, 2H), 0.01 (br d, J=7.6 Hz, 2H), −0.56 (br d, J=4.0 Hz, 2H). MS ESI m/z=581.2 (M+H). HPLC retention time 1.41 minutes, Method 2.
The title compound was prepared from tert-butyl ((1R,4R,7R)-2-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate and methyl 4-bromobutanoate using the procedure described in Example 37, step 1. MS ESI m/z=670.4 (M+H). HPLC retention time 0.95 minutes, Method D.
A stirring solution of methyl 4-(2-(5-((1R,4R,7R)-7-((tert-butoxycarbonyl)amino)-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)butanoate (100 mg, 0.15 mmol) in methanol (2 mL) was treated with 1 M sodium hydroxide (0.750 ml, 0.750 mmol). The reaction was stirred at 60° C. for 3 hours, at which point it was judged to be complete by LCMS. The methanol was allowed to evaporate, and the remaining aqueous mixture was washed twice with diethyl ether. The aqueous phase was adjusted to pH 4 with 1M HCl, and extracted 5× with ethyl acetate. The combined organic phases were washed once with brine, dried over sodium sulfate and concentrated in vacuo. The residue was taken up in dichloromethane, and the solution was treated with 4 M HCl in dioxane (2 ml, 8.00 mmol). The reaction was stirred at room temperature for 18 hours, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo, and The crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 9% B, 9-49% B over 20 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS and UV signals. Fractions containing the desired product were combined and dried via centrifugal evaporation.
The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield the title compound (26.4 mg, 0.048 mmol, 31.7% yield). 1H NMR (500 MHz, DMSO-d6) δ 7.33 (br t, J=4.5 Hz, 1H), 7.23-7.10 (m, 1H), 6.86 (br d, J=4.5 Hz, 2H), 6.78 (s, 1H), 6.70 (br s, 1H), 4.26 (br d, J=5.7 Hz, 2H), 3.85 (s, 3H), 3.76 (s, 3H), 3.31-3.22 (m, 5H), 2.93-2.80 (m, 3H), 2.14 (br t, J=6.7 Hz, 2H), 2.09-1.94 (m, 1H), 1.84-1.65 (m, 5H), 1.61-1.47 (m, 1H), 1.29-1.08 (m, 1H), 0.60 (br s, 1H), −0.01 (br d, 0.1=7.8 Hz, 2H), −0.52 (br s, 2H). MS ESI m/z=556.4 (M+H). HPLC retention time 1.30 minutes, Method 1.
The title compound was prepared from tert-butyl 6-hydroxy-2-azaspiro[3.3]heptane-2-carboxylate using the procedures described in Example 43, steps 1 and 2. 1H NMR (499 MHz, chloroform-d) δ 4.31 (quin, J=7.8 Hz, 1H), 3.96 (d, J=15.1 Hz, 4H), 2.99-2.89 (m, 2H), 2.77-2.68 (m, 2H), 1.45 (s, 9H).
The title compound was prepared from tert-butyl 6-iodo-2-azaspiro[3.3]heptane-2-carboxylate and tert-butyl ((1R,4R,7R)-2-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate using the procedure described in Example 37, step 1.
tert-butyl 6-(2-(5-((1R,4R,7R)-7-((tert-butoxycarbonyl)amino)-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)-2-azaspiro[3.3]heptane-2-carboxylate (27 mg, 0.035 mmol) was dissolved in 10% TFA/dichloromethane (1 mL), and the reaction was stirred at room temperature for 3 hours, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo, and the residue was concentrated 3× from dichloromethane to remove residual TFA. The crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 0.1% trifluoroacetic acid; Mobile Phase B: 95:5 acetonitrile: water with 0.1% trifluoroacetic acid; Gradient: a 0-minute hold at 7% B, 7-47% B over 20 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield the title compound, (19.3 mg, 0.034 mmol, 97% yield). 1H NMR (500 MHz, DMSO-de) δ 7.35 (d, J=7.7 Hz, 1H), 7.31-7.15 (m, 1H), 7.04 (d, J=7.3 Hz, 1H), 6.98-6.91 (m, 1H), 6.83 (s, 1H), 6.76 (br s, 1H), 4.23 (br d, J=6.4 Hz, 2H), 3.99 (s, 2H), 3.88 (s, 3H), 3.87-3.82 (m, 1H), 3.80 (s, 3H), 3.75 (s, 2H), 3.54-3.23 (m, 2H), 2.59 (br t, J=10.4 Hz, 2H), 2.53-2.38 (m, 1H), 1.74 (br s, 3H), 1.45 (br s, 1H), 1.15-0.68 (m, 1H), 0.61-0.46 (m, 1H), −0.01 (br d, J=7.9 Hz, 2H), −0.52 (br s, 2H). (Proton count is low due to the water suppression algorithm used during data processing). MS ESI m/z=565.4 (M+H). HPLC retention time 1.02 minutes, Method 2.
tert-Butyl 6-(2-(5-((1R,4R,7R)-7-((tert-butoxycarbonyl)amino)-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)-2-azaspiro[3.3]heptane-2-carboxylate (54 mg, 0.071 mmol) was dissolved in 10% TFA/dichloromethane (1 mL), and the reaction was stirred at room temperature for 3 hours, at which point it was judged to be complete by LCMS. The mixture was diluted with dichloromethane (20 mL) and concentrated in vacuo, and the residue was taken up in dichloromethane (3 mL). The mixture was treated with 1.5 M potassium phosphate (dibasic) (3 mL), and vigorously shaken for 10 minutes. The layers were separated, and the aqueous phase was extracted 3× with dichloromethane (2 mL). The combined organic phases were dried over sodium sulfate and concentrated in vacuo, and the residue was taken up in dichloromethane (2 mL). Triethylamine (0.013 mL, 0.096 mmol) was added, and the solution was cooled to 0° C. and treated with acetic anhydride (3.38 μl, 0.036 mmol). The reaction was allowed to slowly come to room temperature and stirred for 1 hour, at which point it was judged to be complete by LCMS. The mixture was diluted with methanol to quench any residual acetic anhydride, then concentrated in vacuo. The crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 18% B, 18-58% B over 20 minutes, then a 4-minute hold at 100° % B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS and UV signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield the title compound, (22.9 mg, 0.038 mmol, 79% N yield). 1H NMR (500 MHz, DMSO-d6) δ 7.38 (br d, J=7.9 Hz, 1H), 7.28-7.14 (m, 1H), 7.07 (br d, J=7.3 Hz, 1H), 6.96 (br t, J=7.5 Hz, 1H), 6.87 (br s, 1H), 6.79-6.71 (m, 1H), 4.27 (br d, J=4.9 Hz, 2H), 4.14 (s, 1H), 3.95-3.77 (m, 9H), 3.69-3.52 (m, 2H), 3.52-3.30 (m, 1H), 3.02-2.80 (m, 1H), 2.53 (br d, J=8.9 Hz, 2H), 2.05-1.91 (m, 1H), 1.87-1.51 (m, 6H), 1.30-1.12 (m, 1H), 0.56 (br s, 1H), 0.01 (br d, J=7.9 Hz, 2H), −0.54 (br d, J=3.7 Hz, 2H). (Proton count is low due to the water suppression algorithm used during data processing). MS ESI m/z=607.0 (M+H). HPLC retention time 1.69 minutes, Method 1.
A stirring solution of ((1R,4R,7R)-7-amino-2-azabicyclo[2.2.1]heptan-2-yl)(2-(7-(azetidin-3-ylmethyl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-5-yl)methanone (Example 26) (68 mg, 0.126 mmol) and triethylamine (0.070 mL, 0.505 mmol) in dichloromethane (2 mL) was cooled to 0° C. and treated with acetic anhydride (9.53 μl, 0.101 mmol). The reaction was allowed to slowly come to room temperature and stirred for 1 hour, at which point it was judged to be complete by LCMS. The mixture was diluted with methanol to quench any residual acetic anhydride, then concentrated in vacuo. The crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 0.1% trifluoroacetic acid; Mobile Phase B: 95:5 acetonitrile: water with 0.1% trifluoroacetic acid; Gradient: a 0-minute hold at 12% B, 12-52% B over 20 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS and UV signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield the title compound, (43 mg, 0.072 mmol, 57.3% yield). 1H NMR (500 MHz, DMSO-d6) δ 7.34 (br d, J=7.8 Hz, 1H), 7.26-7.11 (m, 1H), 6.85 (br d, J=7.6 Hz, 1H), 6.81 (br d, J=7.0 Hz, 1H), 6.78 (s, 1H), 6.73 (br s, 1H), 4.23 (br d, J=6.3 Hz, 2H), 4.00 (br t, J=7.6 Hz, 2H), 3.84 (s, 3H), 3.81-3.61 (m, 5H), 3.40 (br dd, J=9.0, 5.8 Hz, 1H), 3.27-3.07 (m, 2H), 2.99-2.91 (m, 1H), 2.89-2.76 (m, 1H), 2.52-2.38 (m, 1H), 1.70 (br s, 3H), 1.51 (s, 3H), 1.46-1.37 (m, 1H), 0.63 (br d, J=5.0 Hz, 1H), −0.01 (br d, J=7.9 Hz, 2H), −0.53 (br s, 2H). (Proton count is low due to the water suppression algorithm used during data processing). MS ESI m/z=581.0 (M+H). HPLC retention time 1.29 minutes, Method 2.
In a 40 mL scintillation vial, a stirring mixture of benzyl 3-iodoazetidine-1-carboxylate (0.782 g, 2.467 mmol), tert-butyl ((1R,4R,7R)-2-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (0.8 g, 1.233 mmol), tris(trimethylsilyl)silane (0.571 mL, 1.850 mmol), Ir(dF(CF3)ppy)2(dtbbpy)PF6 (0.042 g, 0.037 mmol), and sodium carbonate (0.523 g, 4.93 mmol) in 1,4-Dioxane (15 mL) was degassed with bubbling nitrogen for 10 minutes. In a separate 2-dram vial, a stirring mixture of nickel(II) chloride ethylene glycol dimethyl ether complex (0.041 g, 0.185 mmol), and 4,4′-di-tert-butyl-2,2′-bipyridine (0.056 g, 0.210 mmol) in 1,4-dioxane (5 mL) was degassed with nitrogen for 20 minutes. The nickel complex was transferred to the containing the other mixture, the vial was sealed, and the reaction was stirred at room temperature under a blue Kessil lamp for 18 hours, at which point it was judged to be complete by LCMS. The mixture was diluted with ethyl acetate (15 mL) and solids were removed by filtration and rinsed thoroughly with ethyl acetate. The combined filtrate and rinsings were concentrated in vacuo, and the residue was concentrated 3× from methanol to completely remove the other solvents. The crude material was used in the next step without further purification. MS ESI m/z=759.5 (M+H). HPLC retention time 0.98 minutes, Method D.
A stirring mixture of benzyl 3-(2-(5-((1R,4R,7R)-7-((tert-butoxycarbonyl)amino)-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)azetidine-1-carboxylate (0.936 g, 1.233 mmol) and 10% Pd—C, Degussa type (1 g, 0.940 mmol) in methanol (15 mL) was degassed 3× with nitrogen/vacuum. The reaction was hydrogenated at atmospheric pressure for 48 hours, at which point it was judged to be complete by LCMS. The catalyst was removed by filtration through 3 layers of Whatman GF/A filter paper and rinsed thoroughly with methanol and ethyl acetate, and the combined filtrate and rinsings were concentrated in vacuo. The residue was chromatographed via MPLC over a 40 g silica gel column, eluting at 40 mL/min with a 0% to 10% (7 M ammonia in methanol)/dichloromethane gradient over 15 column volumes. Fractions containing the desired product were pooled and concentrated in vacuo to yield the title compound (407 mg, 0.651 mmol, 52.8% yield) as a yellow solid. 1H NMR (499 MHz, chloroform-d) δ 7.62 (d, J=7.7 Hz, 1H), 7.55-7.47 (m, 1H), 7.35 (d, J=7.4 Hz, 1H), 7.23 (t, 0.1=7.6 Hz, 1H), 7.06-6.97 (m, 1H), 6.91-6.84 (m, 1H), 4.77 (quin, J=8.0 Hz, 1H), 4.60-4.42 (m, 3H), 4.35 (br s, 1H), 4.19-4.05 (m, 5H), 4.05-3.97 (m, 5H), 3.90-3.72 (m, 2H), 3.32-3.19 (m, 1H), 2.54 (br s, 1H), 2.12-1.85 (m, 3H), 1.75-1.67 (m, 1H), 1.60-1.31 (m, 10H), 0.89-0.78 (m, 1H), 0.32-0.18 (m, 2H), −0.17-−0.41 (m, 2H). MS ESI m/z=625.3 (M+H). HPLC retention time 0.75 minutes, Method D.
In a 2-dram vial, a stirring solution of tert-butyl ((1R,4R,7R)-2-(2-(7-(azetidin-3-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (15 mg, 0.024 mmol) and triethylamine (5.02 μl, 0.036 mmol) in dichloromethane (1 mL) was treated with methanesulfonyl chloride (1.777 μl, 0.023 mmol). The vial was sealed, and the reaction was stirred at room temperature for 2 hours, at which point it was judged to be complete by LCMS. The mixture was treated with TFA (100 μl, 1.298 mmol), and the reaction was stirred at room temperature for 2 hours, at which point it was judged to be complete by LCMS. The mixture was diluted with dichloromethane (20 mL), and concentrated in vacuo, and the residue was concentrated twice from DCM to remove residual TFA. The crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 17% B, 17-57% B over 20 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS and UV signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield the title compound (7.2 mg, 0.011 mmol, 47.3% yield). 1H NMR (500 MHz, DMSO-d6) δ 7.45 (br d, J=7.6 Hz, 1H), 7.28 (br d, J=7.3 Hz, 1H), 7.25-7.12 (m, 1H), 7.03 (br t, J=7.5 Hz, 1H), 6.87 (br s, 1H), 6.77-6.68 (m, 1H), 4.48-4.37 (m, 1H), 4.21-4.11 (m, 4H), 3.94 (br s, 2H), 3.87 (br s, 3H), 3.78 (br s, 3H), 3.61-3.40 (m, 2H), 3.00-2.83 (m, 5H), 2.72-2.48 (m, 1H), 2.10-1.94 (m, 1H), 1.87-1.67 (m, 2H), 1.61-1.47 (m, 1H), 1.33-1.16 (m, 1H), 1.02 (br s, 1H), 0.57 (br s, 1H), 0.01 (br d, J=7.6 Hz, 2H), −0.56 (br d, J=4.0 Hz, 2H). MS ESI m/z=603.1 (M+H). HPLC retention time 1.44 minutes, Method 2.
In a 2-dram vial, a stirring solution of tert-butyl ((1R,4R,7R)-2-(2-(7-(azetidin-3-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (15 mg, 0.024 mmol), (tert-butoxycarbonyl)glycine (4.63 mg, 0.026 mmol), and triethylamine (8.37 μl, 0.060 mmol) in dichloromethane (1 mL) was treated with BOP (13 mg, 0.030 mmol). The vial was sealed, and the reaction was stirred at room temperature for 2 hours, at which point it was judged to be complete by LCMS. The mixture was treated with TFA (100 μl, 1.298 mmol), and the reaction was stirred at room temperature for 2 hours, at which point it was judged to be complete by LCMS. The mixture was diluted with dichloromethane (20 mL), and concentrated in vacuo, and the residue was concentrated twice from DCM to remove residual TFA. The crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate, Gradient: a 0-minute hold at 7% B, 7-47% B over 20 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The material was further purified via preparative LC/MS with the following conditions: Column: XBridge C18, 150 mm×30 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 8% B, 8-48% B over 20 minutes, then a 2-minute hold at 100% B; Flow Rate: 40 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS and UV signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield the title compound (3.8 mg, 5.25 μmol, 21.85% yield). 1H NMR (500 MHz, DMSO-d6) δ 7.40 (br d, J=7.8 Hz, 1H), 7.28-7.18 (m, 1H), 7.12 (br s, 1H), 6.98 (br t, J=7.6 Hz, 1H), 6.81 (s, 1H), 6.70 (br s, 1H), 4.45 (br s, 2H), 4.30-4.07 (m, 3H), 3.95-3.81 (m, 3H), 3.78-3.48 (m, 3H), 3.36-2.91 (m, 1H), 2.09-1.87 (m, 1H), 1.83-1.58 (m, 5H), 1.56-1.39 (m, 1H), 1.30-1.15 (m, 1H), 1.10-0.95 (m, 2H), 0.89-0.69 (m, 1H), 0.56 (br s, 1H), −0.01 (br d, J=7.6 Hz, 2H), −0.12-−0.35 (m, 1H), −0.51 (br s, 2H). (Proton count is low due to the water suppression algorithm used during data processing). MS ESI m/z=582.6 (M+H). HPLC retention time 1.10 minutes, Method 1.
The following compounds in Table 4 can be made using the procedures described in Examples 64 and 65, substituting the appropriate acid chloride, isocyanate, anhydride, or sulfonyl chloride for methanesulfonyl chloride in Example 64 step 2, or the appropriate carboxylic acid for (tert-butoxycarbonyl)glycine in Example 65.
In a 2-dram vial, a stirring mixture of tert-butyl ((1R,4R,7R)-2-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (50 mg, 0.077 mmol), tert-butyl 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3,6-dihydropyridine-1(2H)-carboxylate (28.6 mg, 0.093 mmol), and 2 M potassium phosphate, tribasic (0.116 mL, 0.231 mmol) in dioxane (2 mL) was degassed with bubbling nitrogen for 5 minutes. The mixture was treated with 2nd generation XPhos precatalyst (2.96 mg, 3.85 μmol) and degassed for another five minutes, then the vial was sealed. The reaction was heated at 50° C. for 4 hours, at which point it was judged to be complete by LCMS. Most of the dioxane was evaporated, and the residue was taken up in ethyl acetate (5 mL). The turbid solution was washed twice with water and once with brine, then dried over sodium sulfate and concentrated in vacuo. The residue was chromatographed via MPLC over a 12 g silica gel column, eluting at 30 mL/min with a 0% to 6% methanol/dichloromethane gradient over 40 column volumes. Fractions containing the desired product were pooled and concentrated in vacuo to yield tert-butyl 4-(2-(5-((1R,4R,7R)-7-((tert-butoxycarbonyl)amino)-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)-3,6-dihydropyridine-1(2H)-carboxylate (46 mg, 0.061 mmol, 79% yield) as a colorless solid. MS ESI m/z=751.5 (M+H). HPLC retention time 1.03 minutes, Method D. The material was taken up in dichloromethane (2 mL) and treated with 4M HCl in dioxane (2 ml, 8.00 mmol). The reaction was stirred at room temperature for 2 hours, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo, and the crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 10% B, 10-50% B over 20 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield the title compound,
(3.7 mg, 5.85 μmol, 24.39% yield). 1H NMR (500 MHz, DMSO-d6) δ 7.59-7.46 (m, 1H), 7.36-7.20 (m, 1H), 7.10-6.92 (m, 3H), 6.90-6.74 (m, 2H), 5.80-5.15 (m, 1H), 3.99 (br d, J=6.7 Hz, 2H), 3.87 (br d, J=3.4 Hz, 2H), 3.29-2.83 (m, 4H), 2.80-2.55 (m, 3H), 2.22-2.05 (m, 1H), 1.99-1.80 (m, 2H), 1.71-1.29 (m, 2H), 1.24-1.08 (m, 3H), 1.03-0.71 (m, 2H), 0.49 (br s, 1H), 0.11-−0.22 (m, 4H), −0.32-−0.60 (m, 1H). (Proton count is low due to the water suppression algorithm used during data processing). MS ESI m/z=551.4 (M+H). HPLC retention time 1.23 minutes, Method 1.
A stirring solution of tert-butyl ((1R,4R,7R)-2-(2-(7-(3-cyanocyclohexyl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.]heptan-7-yl)carbamate (33 mg, 0.049 mmol) (prepared from 3-iodocyclohexane-1-carbonitrile and tert-butyl ((1R,4R,7R)-2-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate using the conditions described in Example 37, step 1) in 1:1 DMSO/water (1 mL) was treated with potassium carbonate (13.48 mg, 0.098 mmol) and hydrogen peroxide (4.98 μl, 0.049 mmol). The reaction was stirred at room temperature for 18 hours. LCMS indicated that the reaction had not gone to completion. The mixture was treated with potassium carbonate (13.48 mg, 0.098 mmol) and hydrogen peroxide (4.98 μl, 0.049 mmol), and the reaction was stirred at 50° C. for 18 hours. The above additions/heating at 50° C. was repeated each day for five days, until the reaction was judged to be essentially complete by LCMS. The mixture was diluted with ethyl acetate (15 mL), and the turbid solution was washed 3× with water and once with brine. The organic phase was dried over sodium sulfate and concentrated in vacuo. The residue was taken up in dichloromethane (2 mL), and the solution was treated with 4 M HCl (2 mL). The reaction was stirred for 1 hour, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo, and the crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 30% B, 30-60% B over 30 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS signals. The cis- and trans-isomers were resolved. The two isomers were handled separately for the remainder of the process. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation.
Example 85, ISOMER 1 (First eluting), (2.0 mg, 3.34 μmol, 6.85% yield). 1H NMR (500 MHz, DMSO-d6) δ 7.29 (d, J=7.7 Hz, 1H), 7.14 (br s, 1H), 7.01 (d, J=7.4 Hz, 1H), 6.90-6.86 (m, 1H), 6.76-6.67 (m, 2H), 4.61-4.49 (m, 1H), 4.20-4.11 (m, 1H), 3.89-3.80 (m, 3H), 3.77 (s, 3H), 3.70-3.55 (m, 2H), 3.39-3.06 (m, 4H), 2.95-2.76 (m, 1H), 2.47 (br s, 1H), 2.03 (br d, J=10.4 Hz, 2H), 1.88-1.69 (m, 5H), 1.60-1.20 (m, 8H), 0.70-0.60 (m, 2H), 0.04-−0.06 (m, 2H), −0.36-−0.55 (m, 2H). MS ESI m/z=595.0 (M+H). HPLC retention time 1.75 minutes, Method 1.
Example 86, ISOMER 2 (Second eluting), (4.3 mg, 6.15 μmol, 12.60% yield). 1H NMR (500 MHz, DMSO-d6) δ 7.75 (d, J=7.7 Hz, 1H), 7.58 (br s, 1H), 7.44 (br d, J=7.4 Hz, 1H), 7.39-7.31 (m, 2H), 7.24-7.18 (m, 1H), 7.15 (br s, 1H), 6.81-6.70 (m, 1H), 4.78-4.57 (m, 2H), 4.28 (s, 3H), 4.21 (s, 3H), 4.03-3.90 (m, 1H), 3.86-3.72 (m, 2H), 3.70-3.56 (m, 1H), 3.36-3.23 (m, 1H), 2.68-2.58 (m, 1H), 2.50-2.36 (m, 1H), 2.32-2.15 (m, 5H), 2.11 (br d, J=2.9 Hz, 1H), 2.01-1.61 (m, 6H), 1.53-1.43 (m, 2H), 1.19-1.10 (m, 1H), 0.50 (br d, J=8.1 Hz, 2H), −0.01 (br s, 2H). MS ESI m/z=595.4 (M+H). HPLC retention time 1.37 minutes, Method 2.
In a 2-dram vial, a stirring mixture of 2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenol (15 mg, 0.068 mmol), ((1R,4R,7R)-7-amino-2-azabicyclo[2.2.1]heptan-2-yl)(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-5-yl)methanone (32 mg, 0.058 mmol), PdCl2(dppf) (4.27 mg, 5.83 μmol), and 2 M potassium phosphate, tribasic (0.117 mL, 0.233 mmol) (previously degassed) in 1,4-Dioxane (2 mL) was degassed with bubbling nitrogen for 10 minutes. The vial was sealed, and the reaction was stirred at 80° C. for 18 hours, at which point it was judged to be complete by LCMS. The solvents were evaporated, and the residue was taken up in DMF (2 mL). The crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 0.1% trifluoroacetic acid; Mobile Phase B: 95:5 acetonitrile: water with 0.1% trifluoroacetic acid; Gradient: a 0-minute hold at 16% B, 16-56% B over 20 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS and UV signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The material was further purified via preparative LC/MS with the following conditions: Column: XBridge Shield RP18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 35% B, 35-57% B over 25 minutes, then a 2-minute hold at 100% B: Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield the title compound (3.2 mg, 5.70 μmol, 9.76% yield). 1H NMR (500 MHz, DMSO-d6) δ 8.03 (br s, 1H), 7.75 (br d, J=7.0 Hz, 1H), 7.51-7.21 (m, 4H), 7.16 (br s, 1H), 7.11-6.93 (m, 3H), 4.16 (br s, 3H), 4.10-3.92 (m, 4H), 3.85-3.43 (m, 2H), 3.28-3.05 (m, 1H), 2.32-2.15 (m, 1H), 2.13-1.92 (m, 3H), 1.87-1.69 (m, 1H), 1.51 (br d, J=9.8 Hz, 1H), 0.93 (br s, 1H), 0.52 (br s, 1H), 0.27-0.06 (m, 2H), −0.38-−0.75 (m, 2H). MS ESI m/z=562.2 (M+H). HPLC retention time 1.86 minutes, Method 1.
The following compounds in Table 5 can be made by the procedures described in Example 87, substituting the appropriate boronic acid or boronic ester for 2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenol.
In a 40 mL vial, a mixture of tert-butyl ((1R,4R,7R)-2-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (100 mg, 0.154 mmol), diethyl phosphite (0.070 mL, 0.540 mmol), triethylamine (0.071 mL, 0.509 mmol), and PdCl2(dppf) (6 mg, 8.20 μmol) in toluene (2 mL) was placed in a sonicator and degassed with bubbling nitrogen for 1 minute. The vial was sealed, and the reaction was stirred at 100° C. for 18 hours. LCMS detected mostly starting material, but did detect a trace of the desired product. The mixture was treated with diethyl phosphite (0.070 mL, 0.540 mmol), triethylamine (0.071 mL, 0.509 mmol), and PdCl2(dppf) (6 mg, 8.20 μmol), and degassed with bubbling nitrogen for 10 minutes. The vial was sealed, and the reaction was stirred at 120° C. for 18 hours, at which point it was judged to be complete by LCMS. The mixture was filtered and concentrated in vacuo, and the residue was taken up in dichloromethane (2 mL). The solution was treated with TFA (1 mL), and the reaction was stirred at room temperature for 3 hours, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo, and the crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 19% B, 19-59/6 B over 20 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS and UV signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The material was further purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 25% B, 25-46% B over 25 minutes, then a 2-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield the title compound, (12 mg, 0.019 mmol, 12.14% yield). 1H NMR (500 MHz, DMSO-d6) δ 7.87 (br d, J=7.6 Hz, 1H), 7.72 (br dd, J=16.2, 7.3 Hz, 1H), 7.54-7.40 (m, 1H), 7.36-7.21 (m, 1H), 7.16 (br t, J=6.7 Hz, 1H), 7.10 (s, 1H), 6.88-6.77 (m, 1H), 4.76-4.59 (m, 2H), 4.02 (br t, J6.7 Hz, 4H), 3.98 (br s, 3H), 3.87 (s, 3H), 3.69-3.35 (m, 1H), 3.19 (br s, 1H), 3.07-2.88 (m, 1H), 2.17-2.00 (m, 1H), 1.93-1.70 (m, 2H), 1.68-1.52 (m, 1H), 1.39-1.06 (m, 8H), 0.61 (br s, 1H), 0.01 (br d, J=7.3 Hz, 2H), −0.44-−0.63 (m, 2H). MS ESI m/z=606.0 (M+H). HPLC retention time 1.49 minutes, Method 2.
In a 2 mL microwave vial, a solution of tert-butyl (2-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (265 mg, 0.409 mmol) and copper(I) iodide (9 mg, 0.047 mmol) in DMF (1.5 mL) was degassed with bubbling nitrogen for 10 minutes. The mixture was treated with bis(triphenylphosphine)palladium(II) dichloride (31.5 mg, 0.045 mmol) and triethylamine (1.139 mL, 8.17 mmol), and degassed for 5 minutes. trimethylsilylacetylene (0.113 mL, 0.817 mmol) was added, the vial was sealed, and the reaction was heated at 120° C. via microwave for 25 minutes, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo, and the residue was taken up in ethyl acetate (35 mL). The solution was filtered, then washed twice with 10% lithium chloride and once with brine, dried over sodium sulfate, and concentrated in vacuo. The residue was chromatographed via MPLC over a 40 g silica gel column, eluting at 40 mL/min with a 20% to 100% ethyl acetate/hexanes gradient over 25 column volumes. Fractions containing the desired product were pooled and concentrated in vacuo to yield 256 mg of an amber solid, which was used without further purification in the next step. MS ESI m/z=666.7 (M+H). HPLC retention time 1.27 minutes, Method D.
A stirring solution of tert-butyl (2-(2-(1-(cyclopropylmethyl)-7-((trimethylsilyl)ethynyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (256 mg, 0.384 mmol) in methanol (5 mL) was treated with potassium carbonate (26.6 mg, 0.192 mmol). The reaction was stirred at room temperature for 2 hours, at which point it was judged to be complete by LCMS. Most of the methanol was evaporated, and the residue was taken up in ethyl acetate. The turbid solution was washed 3× with water, dried over sodium sulfate, and concentrated in vacuo. The residue was chromatographed via MPLC over a 40 g silica gel column, eluting at 40 mL/min with a 0% to 5% methanol/dichloromethane gradient over 12 column volumes. Fractions containing the desired product were pooled and concentrated in vacuo to yield the title compound as an amber solid (80 mg, 0.135 mmol, 35.0% yield). MS ESI m/z=594.6 (M+H). HPLC retention time 1.07 minutes, Method D.
In a 2-dram vial, a mixture of tert-butyl (2-(2-(1-(cyclopropylmethyl)-7-ethynyl-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (13 mg, 0.022 mmol), 2-azidoethan-1-ol (10% in ethanol) (38.1 mg, 0.044 mmol), and sodium ascorbate (0.51 M in water) (0.013 mL, 6.57 μmol) in 5:1 THF/water (0.2 mL) was treated with copper(II) sulfate (0.62 M in water) (3.53 μl, 2.190 μmol). The vial was sealed, and the reaction was stirred at 50° C. for 18 hours. The mixture was treated with 2-azidoethan-1-ol (10% in ethanol) (38.1 mg, 0.044 mmol), and the reaction was stirred at 50° C. for 18 hours, at which point it was judged to be essentially complete by LCMS. The mixture was diluted with ethyl acetate (5 mL) and filtered, and the filtrate was washed once with brine. The organic phase was dried over sodium sulfate and concentrated in vacuo. The residue was taken up in dichloromethane (3 mL), and the solution was treated with 4 M HCl in dioxane (2 mL, 8.00 mmol). The reaction was stirred at room temperature for 2 hours, at which point it was judged to be complete by LCMS. The reaction mixture was concentrated in vacuo, and the crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 19×200 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: 10-50% B over 20 minutes, then a 4-minute hold at 100% B; Flow: 20 mL/min. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield the title compound, (5.5 mg, 8.90 μmol, 40.5% yield). 1H NMR (500 MHz, DMSO-d6) δ 8.30 (s, 1H), 7.79 (br d, J=6.8 Hz, 1H), 7.45-7.31 (m, 1H), 7.26-7.19 (m, 2H), 7.11 (s, 1H), 6.98-6.88 (m, 1H), 4.53 (t, J=5.4 Hz, 2H), 4.14-4.04 (m, 5H), 3.98 (s, 3H), 3.91-3.85 (m, 2H), 3.15-2.97 (m, 1H), 2.94-2.66 (m, 1H), 2.28-2.12 (m, 1H), 2.05-1.84 (m, 5H), 1.79-1.64 (m, 1H), 1.55-1.34 (m, 1H), 1.09-0.78 (m, 1H), 0.42 (br s, 1H), −0.01 (br d, J=7.8 Hz, 2H), −0.50 (br s, 2H). MS ESI m/z=581.4 (M+H). HPLC retention time 1.28 minutes, Method 1.
In a scintillation vial, a mixture of tert-butyl ((1r,4r)-4-ethynylcyclohexyl)carbamate (31.0 mg, 0.139 mmol), tert-butyl ((1r,4r)-4-ethynylcyclohexyl)carbamate (31.0 mg, 0.139 mmol), trans-n,n′-dimethylcyclohexane-1,2-diamine (0.036 mL, 0.231 mmol), and sodium azide (7.89 mg, 0.121 mmol) in DMSO (0.4 mL) and water (0.1 mL) was treated with copper(I) iodide (24.22 mg, 0.127 mmol) and sodium ascorbate (25.2 mg, 0.127 mmol). The vial was sealed, and the reaction was stirred at 70° C. for 18 hours. LCMS indicated that the reaction had not gone to completion. The mixture was allowed to come to room temperature and treated with tert-butyl ((1r,4r)-4-ethynylcyclohexyl)carbamate (31.0 mg, 0.139 mmol), sodium azide (7.89 mg, 0.121 mmol), copper(I) iodide (24.22 mg, 0.127 mmol), and sodium ascorbate (25.2 mg, 0.127 mmol). The vial was sealed, and the reaction was stirred at 70° C. for 7 hours, then at room temperature for 2 days. The mixture was cooled to room temperature and diluted with ethyl acetate. The turbid solution was washed 3× with water, then dried over sodium sulfate and concentrated in vacuo. The residue was taken up in dichloromethane (5 mL), and the solution was treated with 4M HCl in dioxane (7.05 μl, 0.232 mmol). The reaction was stirred at room temperature for 2 hours, at which point it was judged to be complete by LCMS. The reaction mixture was concentrated in vacuo, and the crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 7% B, 7-47% B over 20 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS and UV signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield the title compound, (17.7 mg, 0.028 mmol, 24.07% yield). 1H NMR (500 MHz, DMSO-d6) δ 8.35 (s, 11H), 7.95-7.87 (m, 1H), 7.41-7.24 (m, 3H), 7.18 (s, 11H), 6.94-6.84 (m, 1H), 4.03 (s, 3H), 3.94 (s, 3H), 3.47 (br d, J=10.9 Hz, 2H), 3.20-2.91 (m, 1H), 2.80-2.70 (m, 2H), 2.21-2.00 (m, 3H), 1.91 (br d, J=13.0 Hz, 4H), 1.72-1.60 (m, 1H), 1.56-1.42 (m, 2H), 1.40-1.16 (m, 3H), 0.41 (br d, J=5.4 Hz, 1H), −0.01 (br d, J=7.8 Hz, 2H), −0.57 (br s, 2H). (Proton count is low due to the water suppression algorithm used during data processing). MS ESI m/z=634.2 (M+H). HPLC retention time 1.42 minutes, Method 1.
The title compound was prepared via the procedure described in Example 109. MS ESI m/z=592.5 (M+H). HPLC retention time 1.10 minutes, Method 2.
The title compound was prepared from 2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylic acid and tert-butyl ((3R, 5R)-5-fluoropiperidin-3-yl)carbamate using the conditions described in Example 1, Step 5. 1H NMR (499 MHz, chloroform-d) δ 7.66 (dd, J=7.9, 1.0 Hz, 1H), 7.54-7.49 (m, 2H), 7.05 (t, J=7.7 Hz, 1H), 6.94 (d, J=1.0 Hz, 1H), 6.84 (s, 1H), 5.02-4.30 (m, 5H), 4.12 (s, 3H), 4.09-4.00 (m, 4H), 3.45-3.29 (m, 1H), 3.23-3.02 (m, 1H), 2.43-2.31 (m, 1H), 2.02-1.74 (m, 1H), 1.56-1.24 (m, 9H), 1.19-1.07 (m, 1H), 0.29 (br dd, J=8.0, 0.8 Hz, 2H), −0.14 (br d, J=4.6 Hz, 2H). MS ESI m/z=654.4 (M+H). HPLC retention time 1.00 minutes, Method D.
The title compound was prepared from tert-butyl ((3R,5R)-1-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-5-fluoropiperidin-3-yl)carbamate and methyl 3-iodocyclobutane-1-carboxylate using the conditions described in Examples 43 and 44, Steps 3 and 4. MS ESI m/z=674.6 (M+H). HPLC retention time 0.85, 0.86 minutes, Method D.
The title compounds were prepared from 3-(2-(5-((3R,5R)-3-((tert-butoxycarbonyl)amino)-5-fluoropiperidine-1-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)cyclobutane-1-carboxylic acid (cis/trans mixture) and methanamine hydrochloride using the procedure described in Examples 43 and 44, Step 5.
Example 111, First Eluting: 3-(2-(5-((3R,5R)-3-amino-5-fluoropiperidine-1-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)-N-methylcyclobutane-1-carboxamide, ISOMER 1. 1H NMR (500 MHz, DMSO-d6) δ 7.59 (br d, J=4.3 Hz, 1H), 7.35 (d, J=7.8 Hz, 1H), 7.09 (s, 1H), 7.05 (br d, J=7.0 Hz, 1H), 7.00-6.92 (m, 1H), 6.84 (s, 1H), 6.63 (s, 1H), 5.01-4.48 (m, 1H), 4.26 (br d, J=5.1 Hz, 2H), 3.88 (s, 3H), 3.85-3.79 (m, 1H), 3.79-3.74 (m, 3H), 3.31 (br d, J=1.6 Hz, 2H), 2.91-2.76 (m, 2H), 2.38 (d, J=4.5 Hz, 3H), 2.35-2.32 (m, 4H), 2.24-2.12 (m, 2H), 2.06-1.89 (m, 1H), 1.44-1.23 (m, 1H), 0.64-0.50 (m, 1H), −0.01 (br d, J=7.9 Hz, 2H), −0.49-−0.64 (m, 2H). MS ESI m/z=587.3 (M+H). HPLC retention time 1.22 minutes, Method 2.
Example 112, Second Eluting: 3-(2-(5-((3R,5R)-3-amino-5-fluoropiperidine-1-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)-N-methylcyclobutane-1-carboxamide, ISOMER 2. 1H NMR (500 MHz, DMSO-d6) δ 7.59 (br d, J=4.4 Hz, 1H), 7.37 (d, J=7.8 Hz, 1H), 7.15 (d, J=7.2 Hz, 1H), 7.10 (s, 1H), 6.97 (t, J=7.6 Hz, 1H), 6.85 (s, 1H), 6.64 (s, 1H), 4.99-4.51 (m, J H), 4.20 (br d, J=6.4 Hz, 2H), 4.12-4.02 (m, 11H), 3.88 (s, 3H), 3.77 (s, 3H), 3.56-3.39 (m, 11H), 2.90-2.76 (m, 2H), 2.49-2.41 (m, 5H), 2.38-2.33 (m, 5H), 2.27-2.18 (m, 2H), 2.06-1.91 (m, 11H), 1.44-1.23 (m, 11H), 0.64-0.48 (m, 1H), −0.01 (br d, J=−8.2 Hz, 2H), −0.55 (br d, J=2.6 Hz, 2H). MS ESI m/z=587.3 (M+H). HPLC retention time 1.27 minutes, Method 2.
The following compounds in Table 6 can be made by the procedures described in Examples 11, and 112, substituting the appropriate amine for methanamine hydrochlroide in step 3.
In a 2-dram vial, a stirring mixture of methyl 2-hydroxy-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzoate (129 mg, 0.463 mmol), tert-butyl ((1R,4R,7R)-2-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (200 mg, 0.308 mmol), PdCl2(dppf) (22.56 mg, 0.031 mmol), and 2 M potassium phosphate, tribasic (0.694 mL, 1.388 mmol) (previously degassed) in DMF (2 mL) was degassed with bubbling nitrogen for 10 minutes. The vial was sealed, and the reaction was stirred at 80° C. for hour, at which point it was judged to be complete by LCMS. The solution was allowed to come to room temperature, diluted with ethyl acetate (10 mL) and water (5 mL), and filtered. The layers were separated, and the organic phase was washed twice with 10% lithium chloride and once with brine, then dried over sodium sulfate and concentrated in vacuo. The residue was chromatographed via MPLC over a 24 g silica gel column, eluting at 40 mL/min with a 0% to 100% ethyl acetate/hexanes gradient over 14 column volumes, then with ethyl acetate to completely elute the product. Fractions containing the desired product were pooled and concentrated in vacuo to yield the title compound, (77 mg, 0.107 mmol, 34.7% yield). 1H NMR (500 MHz, chloroform-d) δ 10.88 (s, 1H), 8.01 (d, J=2.1 Hz, 1H), 7.74 (dd, J=7.9, 1.0 Hz, 1H), 7.64 (dd, J=8.5, 2.3 Hz, 1H), 7.47 (s, 1H), 7.24 (t, J=7.5 Hz, 1H), 7.14-7.07 (m, 2H), 7.04-6.99 (m, 1H), 6.95 (s, 1H), 4.68-4.44 (m, 1H), 4.42-4.29 (m, 1H), 4.22-4.11 (m, 3H), 4.03 (s, 3H), 3.98 (s, 5H), 3.89-3.71 (m, 2H), 3.30-3.18 (m, 1H), 2.54 (br s, 1H), 2.06-1.82 (m, 3H), 1.76-1.59 (m, 4H), 1.46-1.27 (m, 9H), 0.51-0.39 (m, 1H), −0.37-−0.73 (m, 2H). MS ESI m/z=720.5 (M+H). HPLC retention time 1.03 minutes, Method D.
A stirring solution of methyl 5-(2-(5-((1R,4R,7R)-7-((tert-butoxycarbonyl)amino)-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)-2-hydroxybenzoate (38 mg, 0.053 mmol) in anhydrous THF (1.5 mL) was cooled to 0 C and treated with lithium borohydride (2 M in THF) (0.053 mL, 0.106 mmol). The reaction was allowed to come to room temperature and stirred for 18 hours, at which point it was judged to be complete by LCMS. The reaction was quenched with methanol (2 mL) and stirred for 30 minutes, then the mixture was concentrated in vacuo. The residue was taken up in dichloromethane (2 mL), and the solution was treated with 4M HCl in dioxane (2 mL). The reaction was stirred at room temperature for 1 hour, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo. The crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 17% B, 17-57% B over 20 minutes, then a 4-minute hold at 100% B: Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield the title compound. 1H NMR (500 MHz, DMSO-d6) δ 9.56 (br s, 1H), 7.71-7.63 (m, 1H), 7.44-7.32 (m, 2H), 7.23-7.17 (m, 2H), 7.15-7.11 (m, 1H), 7.07-7.00 (m, 1H), 6.97-6.88 (m, 2H), 5.05 (br s, 1H), 4.57 (br d, J=3.5 Hz, 2H), 4.18-4.08 (m, 3H), 3.99 (s, 3H), 3.95 (br d, J=6.9 Hz, 1H), 3.75 (s, 1H), 3.67-3.47 (m, 1H), 3.16 (s, 1H), 3.10-3.00 (m, 1H), 2.25-2.10 (m, 1H), 2.07-1.92 (m, 2H), 1.81-1.63 (m, 1H), 1.51-1.32 (m, 1H), 0.46-0.34 (m, 1H), −0.05 (br d, J=8.1 Hz, 2H), −0.45-−0.73 (m, 2H). MS ESI m/z=592.2 (M+H). HPLC retention time 1.30 minutes, Method 2.
In a 2 dram vial, a stirring solution of methyl 5-(2-(5-((1R,4R,7R)-7-((tert-butoxycarbonyl)amino)-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)-2-hydroxybenzoate (38 mg, 0.053 mmol) in methanol (1 mL) was treated with 1M sodium hydroxide (0.158 mL, 0.158 mmol). The vial was sealed, and the reaction was stirred at 70 C for 18 hours, at which point it was judge to be complete by LCMS. The mixture was diluted with water (1 mL) and the methanol was allowed to evaporate. The aqueous mixture was acidified to pH 5 with 1M HCl, and the product was extracted into dichloromethane. The combined organic phases were dried over sodium sulfate and concentrated in vacuo to yield the title compound, which was used without further purification in the next step. MS ESI m/z=706.3 (M+H). HPLC retention time 0.91 minutes, Method D.
A stirring mixture of 5-(2-(5-((1R,4R,7R)-7-((tert-butoxycarbonyl)amino)-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)-2-hydroxybenzoic acid (30 mg, 0.043 mmol), ammonium hydroxide (0.017 mL, 0.425 mmol), and triethylamine (0.024 mL, 0.170 mmol) in DMF (2 mL) was treated with BOP (22.56 mg, 0.051 mmol). The reaction was stirred at room temperature for 18 hours, at which point LCMS detected the desired product and unreacted starting carboxylic acid. The mixture was concentrated in vacuo, and the residue was taken up in dichloromethane (2 mL). The mixture was treated with 4M HCl in dioxane (2 mL), and the reaction was stirred at room temperature for 1 hour, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo. The crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 0.1% trifluoroacetic acid; Mobile Phase B: 95:5 acetonitrile: water with 0.1% trifluoroacetic acid; Gradient: a 0-minute hold at 17% B, 17-57% B over 24 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by UV signals. Fractions containing the Example 120 were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield the title compound Example 120, (9.1 mg, 0.014 mmol, 33.7% yield). 1H NMR (500 MHz, DMSO-d6) δ 8.49 (br s, 1H), 8.11 (s, 1H), 7.98 (br s, 1H), 7.78 (d, J=7.6 Hz, 1H), 7.66-7.57 (m, 1H), 7.55-7.37 (m, 1H), 7.27 (br t, J=7.6 Hz, 1H), 7.20 (s, 1H), 7.14 (br d, J=7.3 Hz, 1H), 7.07 (d, J=8.5 Hz, 1H), 7.03-6.96 (m, 1H), 4.61-4.20 (m, 1H), 4.15 (br s, 3H), 4.03 (br s, 3H), 3.97-3.76 (m, 2H), 3.71-3.59 (m, 1H), 3.57-3.38 (m, 1H), 3.30-3.17 (m, 1H), 2.97 (br d, J=5.2 Hz, 4H), 2.08-1.85 (m, 3H), 1.79-1.58 (m, 1H), 1.40-1.28 (m, 1H), 0.60-0.40 (m, 1H), 0.07-−0.09 (m, 2H), −0.57 (br d, J=0.9 Hz, 2H). MS ESI m/z=605.3 (M+H). HPLC retention time 1.41 minutes, Method 2.
Fractions containing the Example 121 were combined and dried via centrifugal evaporation. The material was further purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 11% B, 11-51% B over 25 minutes, then a 6-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation to yield the title compound Example 121, (4.2 mg, 6.64 μmol, 15.63% yield). 1H NMR (500 MHz, DMSO-d6) δ 7.83 (s, 1H), 7.72 (br d, J=7.5 Hz, 1H), 7.62-7.44 (m, 1H), 7.37 (br d, J=8.4 Hz, 1H), 7.25 (br t, J=7.6 Hz, 1H), 7.20 (s, 1H), 7.10 (br d, J=7.2 Hz, 1H), 7.04-6.96 (m, 1H), 6.84 (d, J=8.2 Hz, 1H), 4.24-4.13 (m, 3H), 4.10-3.98 (m, 4H), 3.81-3.52 (m, 3H), 3.29-3.11 (m, 3H), 2.17-1.83 (m, 4H), 1.80-1.44 (m, 2H), 1.40-1.20 (m, 1H), 0.58-0.35 (m, 1H), −0.01 (br d, J=7.6 Hz, 2H), −0.45-−0.69 (m, 2H). MS ESI m/z=606.1 (M+H). HPLC retention time 1.48 minutes, Method 1.
A vial was charged with tert-butyl-((1R,4R,7R)-2-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (30 mg, 0.046 mmol), 1-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole (9.62 mg, 0.046 mmol), Pd(dppf)Cl2 (3.38 mg, 4.63 μmol) and dioxane (2 mL). The slurry was sparged with argon for 5 min. 2M K2HPO4 (0.060 mL, 0.120 mmol) was added and the vial was capped and heated at 85° C. for 4 hours. LC/MS detects (M+H)+=650.60 for titled compound. The reaction was filtered and concentrated. The residue was dissolved in dioxane (2 mL) and 4N HCl in dioxane (0.116 mL, 0.463 mmol) was then added thereto. The reaction was stirred for 4 h until LC/MS detected no starting material. The reaction was concentrated 3 times from methylene chloride to remove traces of HCl to yield an amber oil. The crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 19×200 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: 16-56% B over 25 minutes, then a 4-minute hold at 100% B; Flow: 20 mL/min. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation. The yield of the titled compound Example 122 was 7.0 mg (26%), and its estimated purity by analytical LC/MS analysis was 97.30/a (Method 1) and 98.30a (Method 2). H NMR (500 MHz, DMSO-d6) δ 7.90 (s, 1H), 7.63 (d, J=7.6 Hz, 1H), 7.60-7.56 (m, 1H), 7.41-7.24 (m, 1H), 7.16-7.08 (m, 1H), 7.08-7.04 (m, 1H), 7.03-6.98 (m, 1H), 6.92-6.82 (m, 1H), 4.12-4.00 (m, 5H), 3.95-3.90 (m, 3H), 3.90-3.87 (m, 3H), 3.74-3.43 (m, 1H), 3.12-3.05 (m, 1H), 3.07-2.88 (m, 1H), 2.26-2.13 (m, 1H), 2.00-1.80 (m, 3H), 1.78-1.58 (m, 1H), 1.47-1.23 (m, 1H), 1.23-1.08 (m, 1H), 0.43 (br d, J=4.9 Hz, 1H), 0.02-−0.13 (m, 2H), −0.51-−0.66 (m, 2H). MS ESI m/z 550.10 (M+H). Analytical LC/MS retention time: 1.51 min (Method 1).
The following compounds in Table 7 can be made by the procedures described in Example 122 using the appropriate starting materials.
The following compounds in Table 8 can be synthesized by the methods discussed in Examples 1 and/or 122 using the appropriate starting materials.
To a solution of tert-butyl ((1R,4R,7R)-2-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (30 mg, 0.046 mmol) in dioxane (1.5 mL) were added tert-butyl 4-aminopiperidine-1-carboxylate (9.26 mg, 0.046 mmol), Cs2CO3 (45.2 mg, 0.139 mmol), and Xantphos (8.03 mg, 0.014 mmol) at rt. The reaction was purged with nitrogen for 5 minutes. Pd2(dba)3 (8.47 mg, 9.25 μmol) was added and the reaction purged again with nitrogen for 5 minutes. The reaction vial under nitrogen was capped and stirred at 100° C. for 16 hours. LC/MS detects (M+H)+=768.40 for intermediate product. The reaction was filtered and concentrated. The residue was dissolved in dioxane (2 mL) then 4N HCl in dioxane (0.116 mL, 0.463 mmol) added thereto. The reaction was stirred at rt for 20 hours. LC/MS detects (M+H)=568.40 for product. The reaction was concentrated 5 times from methylene chloride to remove traces of HCl to yield an amber oil. The crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 11% B, 11-51% B over 20 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25° C. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation. The yield of the product was 1.9 mg (7%), and its estimated purity by analytical LC/MS analysis was 96.4% (Method 1) and 97.7% (Method 2). 1H NMR (500 MHz, DMSO-D6) δ 7.40-7.37 (m, 1H), 7.31 (s, 1H), 7.13 (br d, J=7.6 Hz, 1H), 6.99-6.93 (m, 1H), 6.91-6.87 (m, 1H), 6.86-6.83 (m, 1H), 6.71-6.65 (m, 1H), 5.34-4.81 (m, 1H), 4.43-4.28 (m, 1H), 3.58-3.40 (m, 2H), 3.32-3.19 (m, 2H), 3.02-2.92 (m, 2H), 2.77-2.59 (m, 2H), 2.16-2.03 (m, 2H), 2.00-1.92 (m, 1H), 1.70-1.58 (m, 2H), 1.25-1.04 (m, 6H), 1.43-1.04 (m, 6H), 0.97-0.82 (m, 1H), 0.79-0.66 (m, 2H), 0.12-0.04 (m, 2H), −0.28 (br d, J=4.6 Hz, 2H). MS ESI m/z 568.2 (M+H). Analytical LC/MS retention time: 1.27 (Method 1).
The following compounds in Table 9 can be made by the procedures described in Example 158 using the appropriate starting materials.
The following compounds in Table 10 can be synthesized by the methods discussed in Examples 1, 122, and/or 158 using the appropriate starting materials.
To a solution of ethyl 2-(2-(5-((1R,4R,7R)-7-amino-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)benzoate (20 mg, 0.032 mmol) in anhydrous THF (3 mL) at 0° C. was added 2N lithium borohydride in THF (0.016 mL, 0.032 mmol). The reaction was stirred for 16 hours at room temperature. LC/MS detects incomplete reaction and therefore more 2N lithium borohydride in THF (0.016 mL, 0.032 mmol) as above. After 1 hour, LC/MS showed reaction had progressed about 60%. At this point, the reaction was quenched under nitrogen with the slow addition of a few drops of saturated aqueous NH4Cl solution. Workup entailed extracting the aqueous 3 times from methylene chloride, drying over Na2SO4 and evaporating to obtain an amber oil. The crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 19×200 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: 24-64% B over 20 minutes, then a 4-minute hold at 100% B; Flow: 20 mL/min. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation. The yield of the product was 2.3 mg, and its estimated purity by LCMS analysis was 88%. The yield of the product was 2.3 mg (11%), and its estimated purity by analytical LC/MS analysis was 95.1% (Method 1) and 87.6% (Method 2). 1H NMR (500 MHz, DMSO-d6) δ 7.69 (br d, J=7.9 Hz, 1H), 7.60 (br d, J=7.6 Hz, 1H), 7.45 (br t, J=7.2 Hz, 1H), 7.35-7.25 (m, 3H), 7.17 (br t, J=7.5 Hz, 1H), 7.11-7.06 (m, 1H), 6.98-6.93 (m, 1H), 6.91-6.85 (m, 1H), 4.28-4.21 (m, 1H), 4.15-4.07 (m, 1H), 4.07-4.00 (m, 31H), 3.93 (s, 3H), 3.80-3.68 (m, 1H), 3.61-3.51 (m, 1H), 3.54 (br s, 1H), 3.06-2.94 (m, 1H), 2.87-2.65 (m, 2H), 2.23-2.10 (m, 1H), 1.96-1.84 (m, 2H), 1.75-1.56 (m, 1H), 1.47-1.35 (m, 1H), 1.02-0.86 (m, 1H), 0.83-0.75 (m, 1H), 0.33 (br s, 1H), −0.10 (br d, J=6.1 Hz, 3H), −0.55-−0.69 (m, 1H), −0.90 (br s, 1H). MS ESI m/z 576.4 (M+H). Analytical LC/MS retention time: 1.91 (Method 1).
The following compounds in Table 11 can be synthesized by the methods discussed in Examples 158 and 181 using the appropriate starting materials.
tert-Butyl ((1R,4R,7R)-2-(2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (50 mg, 0.077 mmol) was dissolved in dioxane (2 mL), and to this mixture was added dicyanozinc (9.05 mg, 0.077 mmol), zinc (5.04 mg, 0.077 mmol) and PdCl2(dppf) (56.4 mg, 0.077 mmol). The mixture was heated to 100° C. overnight. LC/MS shows reaction to be essentially complete. The reaction was filtered and concentrated. The residue was dissolved in dioxane (2 mL) and 4N HCl in dioxane (0.109 mL, 0.435 mmol) was then added. The contents were stirred at room temperature for several hours after which LC/MS showed reaction to be complete. The reaction was concentrated 5 times from methylene chloride to get rid of traces of HCl, yielding an amber oil.
The crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 19×200 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: 24-64% B over 20 minutes, then a 4-minute hold at 100% B; Flow: 20 mL/min. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation. The yield of the product was 10.6 mg (27%), and its estimated purity by analytical LC/MS analysis was 98.5% (Method 1) and 98.9% (Method 2). 1H NMR (500 MHz, DMSO-d6) δ 8.05 (br d, J=7.9 Hz, 1H), 7.78 (d, J=7.3 Hz, 2H), 7.44-7.32 (m, 1H), 7.30-7.24 (m, 1H), 7.22-7.18 (m, 1H), 6.94-6.87 (m, 1H), 4.65 (br d, J=7.0 Hz, 2H), 4.12-3.98 (m, 3H), 3.97-3.80 (m, 3H), 3.48 (br s, 1H), 3.06-2.94 (m, 1H), 2.18 (br s, 1H), 1.99-1.80 (m, 2H), 1.75-1.58 (m, 1H), 1.44-1.27 (m, 1H), 1.26-1.09 (m, 1H), 1.09-1.01 (m, 1H), 0.27 (br d, J=7.6 Hz, 2H), −0.08-−0.19 (m, 2H). MS ESI m/z 495.37 (M+H). Analytical LC/MS retention time: 1.59 (Method 1).
2-(5-((1R,4R,7R)-7-Amino-2-azabicyclo[2.2.1]heptane-2-carbonyl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indole-7-carbonitrile (8 mg, 0.016 mmol) was dissolved in DMSO (1 mL) at 25° C. and then 5M KOH (0.016 ml, 0.081 mmol) was added followed by 33% aqueous hydrogen peroxide (15 uL, 0.162 mmol). Some gas evolution was observed. The reaction was stirred at room temperature for 1 hour after which LC/MS showed reaction to be essentially finished. The reaction was filtered, and the filtrate was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 10% B, 10-50% B over 20 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS and UV signals. Fractions containing the desired product were combined and dried via centrifugal evaporation.
The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation.
The yield of the product was 4.9 mg (59%), and its estimated purity by analytical LC/MS analysis was 100.0% (Method 1) and 100% (Method 2). 1H NMR (500 MHz, DMSO-d6) δ 8.11 (br s, 1H), 7.74 (br d, J=7.6 Hz, 1H), 7.63 (br s, 1H), 7.35-7.25 (m, 2H), 7.16-7.10 (m, 1H), 7.08-7.02 (m, 1H), 6.81 (br d, J=10.1 Hz, 1H), 4.41 (br d, J=7.0 Hz, 2H), 4.06-3.98 (m, 3H), 3.94 (s, 3H), 3.52-3.26 (m, 1H), 3.07-2.95 (m, 1H), 2.24-2.10 (m, 1H), 2.02-1.88 (m, 2H), 1.77-1.61 (m, 1H), 1.46-1.28 (m, 1H), 1.03-0.78 (m, 2H), 0.16-0.08 (m, 2H), 0.07-−0.13 (m, 1H), −0.28 (br s, 2H). MS ESI m/z 513.43 (M+H). Analytical LC/MS retention time: 1.17 (Method 1).
A mixture of 7-methoxy-1H-indole-2-carboxylic acid (120 mg, 0.628 mmol), potassium carbonate (260 mg, 1.883 mmol) and (bromomethyl)cyclopropane (254 mg, 1.883 mmol) in DMF (5 mL) was stirred for 2 hours at 70° C. and then overnight at room temperature. The reaction was complete by LC/MS. The reaction mixture was diluted with EtOAc and washed with 10% LiCl (2×) followed by brine. The combined aqueous layers were extracted with EtOAc. The combined organic layers were dried over anhydrous sodium sulfate and evaporated to yield an amber oil. The crude product was dissolved in MeOH (10 mL) and 1.0N sodium hydroxide (1.255 mL, 1.255 mmol) was added thereto. The reaction was heated at 50° C. for 2 hours when LC/MS showed reaction to be complete. The basic aqueous mixture was acidified to pH=3 with 1N HCl. Precipitate formed which was dissolved and extracted (2×) with methylene chloride. The organic layers were combined, dried (sodium sulfate) and concentrated to yield 1-(cyclopropylmethyl)-7-methoxy-1H-indole-2-carboxylic acid (120 mg, 0.440 mmol, 70% yield) of an amber oil as product. The material was used without further purification in the next step.
1-(Cyclopropylmethyl)-7-methoxy-1H-indole-2-carboxylic acid (120 mg, 0.489 mmol), methyl 3-amino-5-methoxy-4-(methylamino)benzoate (123 mg, 0.587 mmol) (WO2017/100594) and Hunig's Base (0.214 mL, 1.223 mmol) and lastly added HATU (223 mg, 0.587 mmol) were added to DMF (10 mL) at room temperature. The reaction was stirred for 60 hours after which LC/MS showed that reaction was complete. The reaction mixture was diluted with EtOAc and washed with 10% LiCl (2×) followed by brine. The combined aqueous layers were extracted with EtOAc. The combined organic layers were dried over anhydrous sodium sulfate and evaporated to yield an amber oil. The crude material was then taken dissolved in acetic acid (10.00 mL) and the solution stirred at 70° C. for 2 h when LC/MS showed reaction to be complete. The reaction was concentrated to yield a tan solid (250 mg, 0.447 mmol). The isolated intermediate was used without further purification in the next step.
Methyl 2-(1-(cyclopropylmethyl)-7-methoxy-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylate (250 mg, 0.447 mmol) was dissolved in methanol (10 mL and then 1N NaOH (0.894 mL, 0.894 mmol) was added thereto. The mixture was heated at 50° C. for 2 hours when LC/MS showed the reaction to be complete. The organic solvent was evaporated and the residual basic aqueous was acidified to pH=3 with 1N HCl. Precipitate formed which was dissolved and extracted (2×) with methylene chloride. The organic layers were combined, dried (sodium sulfate) and concentrated to yield 2-(1-(cyclopropylmethyl)-7-methoxy-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylic acid (130 mg, 0.192 mmol) as a tan solid. The isolated intermediate was used without further purification in the next step.
To a mixture of 2-(1-(cyclopropylmethyl)-7-methoxy-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylic acid (25 mg, 0.062 mmol), triethylamine (8.59 μl, 0.062 mmol), and BOP (27.3 mg, 0.062 mmol) in 1,4-dioxane (2 mL) was added tert-butyl ((3R,5R)-5-fluoropiperidin-3-yl)carbamate (13.46 mg, 0.062 mmol). The mixture was stirred at room temperature for 16 hours at which LC/MS showed reaction to be complete. Ethyl acetate was added and the mixture washed with 10% LiCl (3×). The organic layer was dried over sodium sulfate and concentrated to yield an amber solid as product. This BOC-protected intermediate was dissolved in 1,4-dioxane (2 mL) at 25° C. and to this solution was added 4N HCl in dioxane (0.018 mL, 0.073 mmol). The reaction was stirred for 2 hours after which LC/MS showed reaction to be complete. The reaction was evaporated 5 times from methylene chloride to yield an amber oil. The crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 23% B, 23-63% B over 20 minutes, then a 4-minute hold at 1000% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS and UV signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation. The yield of the product was 6.4 mg (91%), and its estimated purity by analytical LC/MS was 97.0% (Method 1) and 91.4% (Method 2); 1H NMR (500 MHz, DMSO-d6) δ 7.36-7.28 (m, 1H), 7.26-7.21 (m, 1H), 7.09-7.02 (m, 1H), 6.96-6.91 (m, 1H), 6.86-6.78 (m, 2H), 4.52 (br d, J=7.0 Hz, 2H), 4.24-3.99 (m, 3H), 3.98-3.90 (m, 6H), 3.61-3.48 (m, 4H), 3.09-2.93 (m, 1H), 2.26-2.09 (m, 1H), 1.66-1.42 (m, 1H), 0.95 (br s, 2H), 0.21 (br d, J=7.9 Hz, 2H), −0.12 (br d, J=4.0 Hz, 2H). MS ESI m/z 506.09 (M+H). Analytical LC/MS retention time: 1.79 min. (Method 1).
The following compounds in Table 12 can be synthesized by the methods discussed heretofore using the appropriate starting materials.
A stirring solution of tert-butyl ((7R)-2-(2-(1-(cyclopropylmethyl)-7-(piperidin-4-yl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (15 mg, 0.023 mmol) in dichloromethane (1 mL) was treated with phenyl isocyanate (2.51 μl, 0.023 mmol). The reaction was stirred at room temperature for 2 hours, at which point it was judged to be complete by LCMS. The mixture was treated with 4M HCl in dioxane (1 mL), and the reaction was stirred at room temperature for 1 hour, at which point it was judged to be complete by LCMS. The mixture was concentrated in vacuo, and the crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 0.05% trifluoroacetic acid; Mobile Phase B: 95:5 acetonitrile: water with 0.05% trifluoroacetic acid; Gradient: a 0-minute hold at 20% B, 20-60% B over 23 minutes, then a 0-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Fraction collection was triggered by MS signals. Fractions containing the desired product were combined and dried via centrifugal evaporation. The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation.
The yield of the product was 13.6 mg, and its estimated purity by LCMS analysis was 100%. Analytical LC/MS was used to determine the final purity.
Proton NMR was acquired in deuterated DMSO and shows multiple conformers. 1H NMR (500 MHz, DMSO-86) δ 8.57-8.52 (m, 1H), 7.53-7.33 (m, 4H), 7.27-7.13 (m, 4H), 7.12-7.06 (m, 1H), 7.06-6.96 (m, 1H), 6.96-6.87 (m, 2H), 4.55-3.87 (m, 10H), 3.62-3.39 (m, 1H), 3.23-3.07 (m, 1H), 3.02-2.90 (m, 2H), 2.66-2.52 (m, 1H), 2.03-1.52 (m, 9H), 1.24-1.11 (m, 1H), 0.95-0.82 (m, 1H), 0.27-0.15 (m, 2H), −0.29-−0.45 (m, 2H). MS ESI m/z=670.2 (M+H). HPLC retention time 1.91 minutes, Method 1.
A stirring solution of tert-butyl ((7R)-2-(2-(1-(cyclopropylmethyl)-7-(piperidin-4-yl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-2-azabicyclo[2.2.1]heptan-7-yl)carbamate (20 mg, 0.031 mmol) in dichloromethane (1 mL) was treated with triethylamine (4.3 μL, 0.031 mmol) and dimethylcarbamic chloride (2.8 μL, 0.031 mmol) at rt. After 10 minutes, LCMS had detected product. 4N HCl in dioxane was added thereto (153 μL, 0.61 mmol) and the mixture stirred at rt. After 30 minutes, the reaction was judged to be complete by LCMS. The mixture was evaporated, and the crude material was purified via preparative LC/MS with the following conditions: Column: XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 0.05% trifluoroacetic acid; Mobile Phase B: 95:5 acetonitrile: water with 0.05% trifluoroacetic acid; Gradient: a 0-minute hold at 15% B, 15-55% B over 20 minutes, then a 0-minute hold at 100% B; Flow Rate: 20 mL/min: Column Temperature: 25 C. Fraction collection was triggered by MS and UV signals. Fractions containing the desired product were combined and dried via centrifugal evaporation.
The purified material was then diluted with DMF, treated with Si-Pyridine and shaken for a minimum of 2 h. The resulting mixture was filtered and dried via centrifugal evaporation.
The yield of the product was 12.1 mg, and its estimated purity by LCMS analysis was 95%. Analytical LC/MS was used to determine the final purity.
Proton NMR was acquired in deuterated DMSO and shows multiple conformers. 1H NMR (500 MHz, DMSO-d6) δ 7.57-7.47 (m, 1H), 7.47-7.32 (m, 1H), 7.19-7.15 (m, 1H), 7.12-7.06 (m, 1H), 7.00-6.97 (m, 1H), 6.95-6.88 (m, 1H), 4.55-3.86 (m, 8H), 3.78-3.64 (m, 1H), 3.55-3.41 (m, 1H), 3.16-3.07 (m, 1H), 2.91-2.80 (m, 2H), 2.79-2.65 (m, 7H), 2.64-2.52 (m, 1H), 2.02-1.51 (m, 9H), 1.28-1.11 (m, 1H), 0.93-0.83 (m, 1H), 0.26-0.15 (m, 2H), −0.27-−0.44 (m, 2H). MS ESI m/z=624.0 (M+H). HPLC retention time 1.88 minutes, Method 1.
To a solution of 7-bromo-1-(cyclopropylmethyl)-1H-indole-2-carbaldehyde (350 mg, 1.258 mmol) and methyl 3-methoxy-4-(((1-methyl-1H-pyrazol-4-yl)methyl)amino)-5-nitrobenzoate (403 mg, 1.258 mmol) in EtOH (8.0 mL) was added a solution of sodium dithionite (657 mg, 3.77 mmol) in water (4.00 mL), the mixture was stirred at 80° C. for 6 hour. The mixture was diluted with EtOAc (25 mL) and was washed with a solution of aqueous saturated sodium bicarbonate (2×25 mL). The ethyl acetate layer was dried over sodium sulfate and concentrated. The crude product was subjected to ISCO flash chromatography (silica gel/hexane-EtOAc 100:0 to 0:100 gradient). Yield methyl 2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-((1-methyl-1H-pyrazol-4-yl)methyl)-1H-benzo[d]imidazole-5-carboxylate (409 mg, 0.708 mmol, 56.3% yield) off-white foam. 1H NMR (499 MHz, CHLOROFORM-d) δ 8.22 (d, J=1.2 Hz, 1H), 7.66 (dd, J=7.9, 1.0 Hz, 1H), 7.55-7.53 (m, 2H), 7.28 (s, 1H), 7.08-6.99 (m, 2H), 6.84 (s, 1H), 5.56 (s, 2H), 4.63 (d, J=6.9 Hz, 2H), 4.09 (s, 3H), 3.99 (s, 3H), 3.81 (s, 3H), 1.15-1.03 (m, 1H), 0.24-0.17 (m, 2H), −0.07-−0.11 (m, 2H). LC/MS (M+H): 549; LC retention time: 1.07 min (analytical HPLC Method B).
A stirring mixture of methyl 2-(7-bromo-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-((1-methyl-1H-pyrazol-4-yl)methyl)-1H-benzo[d]imidazole-5-carboxylate (49.0 mg, 0.089 mmol), tris(trimethylsilyl)silane (0.055 mL, 0.179 mmol), Ir(dF(CF3)ppy)2(dtbbpy)PF6 (3.01 mg, 2.68 μmol), tert-butyl 4-bromopiperidine-1-carboxylate (47.2 mg, 0.179 mmol) and sodium carbonate (37.9 mg, 0.357 mmol) in 1,4-dioxane (2 mL) was degassed with bubbling nitrogen for 10 minutes. In a separate vial, a stirring mixture of nickel (II) chloride ethylene glycol dimethyl ether complex (2.94 mg, 0.013 mmol), and 4,4′-di-tert-butyl-2,2′-bipyridine (4.08 mg, 0.015 mmol) in 1,4-dioxane (1 mL) was degassed with nitrogen for 15 minutes. The nickel complex was transferred to the vial containing the other mixture and the reaction was stirred at room temperature under a blue Kessil lamp for 18 hours. The mixture was diluted with EtOAc (10 mL) and was washed with a solution of aqueous saturated sodium bicarbonate (2×10 mL). The ethyl acetate layer was dried over sodium sulfate and concentrated to give crude methyl 2-(7-(1-(tert-butoxycarbonyl)piperidin-4-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-((1-methyl-1H-pyrazol-4-yl)methyl)-1H-benzo[d]imidazole-5-carboxylate. The crude methyl 2-(7-(1-(tert-butoxycarbonyl)piperidin-4-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-((1-methyl-1H-pyrazol-4-yl)methyl)-1H-benzo[d]imidazole-5-carboxylate in DCM (1.0 mL) and TFA (1.0 mL) was stirred at RT for 30 min. The mixture was concentrated. The crude product was purified by prep-HPLC (Phenomenex, Luna 5 micron 30×250 mm, flow rate=30 ml/min., gradient=20% A to 100% B in 30 min., A=H2O/ACN/TFA (90:10:0.1), B=H2O/ACN/TFA (10:90:0.1)). Yield methyl 2-(1-(cyclopropylmethyl)-7-(piperidin-4-yl)-1H-indol-2-yl)-7-methoxy-1-((1-methyl-1H-pyrazol-4-yl)methyl)-1H-benzo[d]imidazole-5-carboxylate (12 mg, 0.021 mmol, 23.09% yield) as clear gum. LC/MS (M+H): 553; LC retention time: 0.74 min (analytical HPLC Method B).
To a solution of methyl 2-(1-(cyclopropylmethyl)-7-(piperidin-4-yl)-1H-indol-2-yl)-7-methoxy-1-((1-methyl-1H-pyrazol-4-yl)methyl)-1H-benzo[d]imidazole-5-carboxylate (12 mg, 0.022 mmol) and TEA (15.13 μl, 0.109 mmol) in THF (1.0 mL) was added acetic anhydride (2.049 μl, 0.022 mmol), the mixture was stirred at RT for 1 hours. The mixture was diluted with EtOAc (5 mL) and was washed with a solution of aqueous saturated sodium bicarbonate (2×5 mL). The ethyl acetate layer was dried over sodium sulfate and concentrated to give crude methyl 2-(7-(1-acetylpiperidin-4-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-((1-methyl-1H-pyrazol-4-yl)methyl)-1H-benzo[d]imidazole-5-carboxylate.
A mixture of methyl 2-(7-(1-acetylpiperidin-4-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-((1-methyl-1H-pyrazol-4-yl)methyl)-1H-benzo[d]imidazole-5-carboxylate and 1.0 M aqueous sodium hydroxide (109 μl, 0.109 mmol) in MeOH (2.0 mL) was stirred at 50° C. for 3 hours. The mixture was cooled to RT. A solution of 1.0 M aqueous HCl (0.10 mL) was added and the mixture was concentrated to give crude 2-(7-(1-acetylpiperidin-4-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-((1-methyl-1H-pyrazol-4-yl)methyl)-1H-benzo[d]imidazole-5-carboxylic acid (14 mg, 0.019 mmol, 89% yield) as white solid. LC/MS (M+H): 581; LC retention time: 0.78 min (analytical HPLC Method B).
A mixture of 2-(7-(1-acetylpiperidin-4-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-((1-methyl-1H-pyrazol-4-yl)methyl)-1H-benzo[d]imidazole-5-carboxylic acid (14 mg, 0.024 mmol), tert-butyl ((3R,5R)-5-fluoropiperidin-3-yl)carbamate (5.26 mg, 0.024 mmol), BOP (10.66 mg, 0.024 mmol) and TEA (16.80 μl, 0.121 mmol) in DMF (1.0 mL) was stirred at RT for 2 hours. The mixture was diluted with EtOAc (5 mL) and was washed with a solution of aqueous saturated sodium bicarbonate (2×5 mL). The ethyl acetate layer was dried over sodium sulfate and concentrated to give tert-butyl ((3R,5R)-1-(2-(7-(1-acetylpiperidin-4-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-((1-methyl-1H-pyrazol-4-yl)methyl)-1H-benzo[d]imidazole-5-carbonyl)-5-fluoropiperidin-3-yl)carbamate. LC/MS (M+H): 781; LC retention time: 0.84 min (analytical HPLC Method B).
A mixture of tert-butyl ((3R,5R)-1-(2-(7-(1-acetylpiperidin-4-yl)-1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-((1-methyl-1H-pyrazol-4-yl)methyl)-1H-benzo[d]imidazole-5-carbonyl)-5-fluoropiperidin-3-yl)carbamate in DCM (1.0 mL) and TFA (1.0 mL) was stirred at RT for 30 min. The mixture was concentrated. The crude product was purified by XBridge C18, 200 mm×19 mm, 5-μm particles; Mobile Phase A: 5:95 acetonitrile: water with 10-mM ammonium acetate; Mobile Phase B: 95:5 acetonitrile: water with 10-mM ammonium acetate; Gradient: a 0-minute hold at 14% B, 14-54% B over 20 minutes, then a 4-minute hold at 100% B; Flow Rate: 20 mL/min; Column Temperature: 25 C. Yield 1-(4-(2-(5-((3R,5R)-3-amino-5-fluoropiperidine-1-carbonyl)-7-methoxy-1-((1-methyl-1H-pyrazol-4-yl)methyl)-1H-benzo[d]imidazol-2-yl)-1-(cyclopropylmethyl)-1H-indol-7-yl)piperidin-1-yl)ethan-1-one (6.10 mg, 8.84 μmol, 36.7% yield). 1H NMR (500 MHz, DMSO-d6) δ 7.81-7.75 (m, 11H), 7.64-7.58 (m, 1H), 7.55-7.50 (m, 1H), 7.45-7.39 (m, 1H), 7.38-7.31 (m, 1H), 7.28-7.23 (m, 1H), 7.22-7.17 (m, 1H), 7.13-7.08 (m, 1H), 5.75-5.68 (m, 2H), 5.25-4.99 (m, 1H), 4.86-4.78 (m, 1H), 4.56-4.48 (m, 2H), 4.25-4.15 (m, 3H), 3.95-3.89 (m, 2H), 3.86-3.76 (m, 1H), 3.49-3.37 (m, 1H), 3.28-3.18 (m, 1H), 3.15-2.92 (m, 1H), 2.93-2.84 (m, 1H), 2.45-2.32 (m, 1H), 2.29-2.14 (m, 5H), 2.14-2.07 (m, 1H), 2.07-1.93 (m, 2H), 1.90-1.62 (m, 3H), 1.53-1.40 (m, 1H), 1.26-1.03 (m, 2H), 0.42-0.32 (m, 2H), 0.06-−0.05 (m, 2H). LC/MS (M+H): 681; LC retention time: 1.19 min (analytical HPLC Method 2).
The following compounds in Table 13 can be synthesized by the methods discussed heretofore using the appropriate starting materials.
Compounds of the present invention were assayed as inhibitors of PAD4 using the assay protocol described below.
Compounds were solubilized in 100% DMSO to achieve a 10 mM compound concentration. Compound stock solutions were stored at RT. A series of dilutions were prepared in DMSO and mixed 8 times with 20 μL mixing volume. Final top concentration of compound in the assay is 50 μM. Final assay conditions were as follows:
0.13 μL of compound solution was added to 13 μL of 10 nM PAD4 in assay buffer. After 30 min 13 μl of 500 μM of BAEE was added in 25 mM hepes, pH 7.5, 5 mM NaCl, 1 mM DTT, 0.2 mg/ml BSA, 0.01% CHAPS, 50 μM Calcium, 5 μM TPEN was added and the reaction incubated for 90 min at 37° C. The enzymatic reaction was quenched by addition of 15 μl of 6.1N TCA, 100% Final Concentration is 20%, 35 μl of 8.5 mM phenyl glyoxal (final concentration 4 mM) is then added and the reaction is incubated for 30 min at 37° C.
After 30 minutes the plates are spun down to remove all precipitate. The enzyme reaction was quenched with an equal volume of methanol containing internal standard (modified citrulline). Samples were loaded onto the Rapid Fire RF300 system (Agilent) wherein they were first sipped for 1000 ms and then directly loaded to a C18 separations cartridge using a mixture of acetonitrile containing 0.01% formic acid for 3000 ms desalting. The flow rate of the mobile phase was 1.5 ml/min. Once the samples were eluted from the cartridge, a mobile phase of acetonitrile containing 0.01% formic acid was used to move the samples into the mass spectrometer for 4000 ms at a flow rate of 1.25 ml/min/Sciex API5500 triple quadrupole mass spectrometer (Applied Biosystems) equipped with ESI was used to analyze the peptidyl citrulline and internal standard ions.
MRM transition of product and internal standard were monitored at m/z 424.5 to 350.4 and m/z 293 to 247 respectively. The dwell time for each transition was set at 200 ms, and the ESI voltage was used at 5500 with a source temperature of 400° C. Extracted ion peaks for each transition were integrated using the Rapid Fire Integrator software. Peak area of analyte was normalized with internal standard.).
For a given compound example, the Table below shows the human PAD4 (hPAD4) IC50 in the rapid-fire mass spectrum (RFMS) assay.
This application claims the benefit of U.S. Provisional Application Ser. No. 62/715,858 filed Aug. 8, 2018 which is incorporated herein in its entirety.
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/US2019/045424 | 8/7/2019 | WO |
| Number | Date | Country | |
|---|---|---|---|
| 62715858 | Aug 2018 | US |
