Discovery of pyrrolo[2,1-f][1,2,4]triazine C6-ketones as potent, orally active p38α MAP kinase inhibitors.

Pyrrolo[2,1-f][1,2,4]triazine based inhibitors of p38α have been prepared exploring functional group modifications at the C6 position. Incorporation of aryl and heteroaryl ketones at this position led to potent inhibitors with efficacy in in vivo models of acute and chronic inflammation.

Imidazo[4,5-d]thiazolo[5,4-b]pyridine based inhibitors of IKK2: synthesis, SAR, PK/PD and activity in a preclinical model of rheumatoid arthritis.

The synthesis, structure-activity relationships (SAR) and biological evaluation of thiazole based tricyclic inhibitors of IKK2 are described. Compound 9 was determined to be orally efficacious in a murine model of rheumatoid arthritis.

Discovery of BMS-986202: A Clinical Tyk2 Inhibitor that Binds to Tyk2 JH2.

A search for structurally diversified Tyk2 JH2 ligands from 6 (BMS-986165), a pyridazine carboxamide-derived Tyk2 JH2 ligand as a clinical Tyk2 inhibitor currently in late development for the treatment of psoriasis, began with a survey of six-membered heteroaryl groups in place of the N-methyl triazolyl moiety in 6. The X-ray co-crystal structure of an early lead (12) revealed a potential new binding pocket. Exploration of the new pocket resulted in two frontrunners for a clinical candidate. The potential hydrogen bonding interaction with Thr599 in the pocket was achieved with a tertiary amide moiety, confirmed by the X-ray co-crystal structure of 29. When the diversity search was extended to nicotinamides, a single fluorine atom addition was found to significantly enhance the permeability, which directly led to the discovery of 7 (BMS-986202) as a clinical Tyk2 inhibitor that binds to Tyk2 JH2. The preclinical studies of 7, including efficacy studies in mouse models of IL-23-driven acanthosis, anti-CD40-induced colitis, and spontaneous lupus, will also be presented.

Driving Potency with Rotationally Stable Atropisomers: Discovery of Pyridopyrimidinedione-Carbazole Inhibitors of BTK.

Bruton's tyrosine kinase (BTK) has been shown to play a key role in the pathogenesis of autoimmunity. Therefore, the inhibition of the kinase activity of BTK with a small molecule inhibitor could offer a breakthrough in the clinical treatment of many autoimmune diseases. This Letter describes the discovery of BMS-986143 through systematic structure-activity relationship (SAR) development. This compound benefits from defined chirality derived from two rotationally stable atropisomeric axes, providing a potent and selective single atropisomer with desirable efficacy and tolerability profiles.

Periodic, partial inhibition of IkappaB Kinase beta-mediated signaling yields therapeutic benefit in preclinical models of rheumatoid arthritis.

We have previously shown that inhibitors of IkappaB kinase beta (IKKbeta), including 4(2'-aminoethyl)amino-1,8-dimethylimidazo(1,2-a)quinoxaline (BMS-345541), are efficacious against experimental arthritis in rodents. In our efforts to identify an analog as a clinical candidate for the treatment of autoimmune and inflammatory disorders, we have discovered the potent and highly selective IKKbeta inhibitor 2-methoxy-N-((6-(1-methyl-4-(methylamino)-1,6-dihydroimidazo[4,5-d]pyrrolo[2,3-b]pyridin-7-yl)pyridin-2-yl)methyl)acetamide (BMS-066). Investigations of its pharmacology in rodent models of experimental arthritis showed that BMS-066 at doses of 5 and 10 mg/kg once daily was effective at protecting rats against adjuvant-induced arthritis, despite showing only weak inhibition at 10 mg/kg against a pharmacodymanic model of tumor necrosis factor alpha production in rats challenged with lipopolysaccharide. The duration of exposure in rats indicated that just 6 to 9 h of coverage per day of the concentration necessary to inhibit IKKbeta by 50% in vivo was necessary for protection against arthritis. Similar findings were observed in the mouse collagen-induced arthritis model, with efficacy observed at a dose providing only 6 h of coverage per day of the concentration necessary to inhibit IKKbeta by 50%. This finding probably results from the cumulative effect on multiple cellular mechanisms that contribute to autoimmunity and joint destruction, because BMS-066 was shown to inhibit a broad spectrum of activities such as T cell proliferation, B cell function, cytokine and interleukin secretion from monocytes, T(H)17 cell function and regulation, and osteoclastogenesis. Thus, only partial and transient inhibition of IKKbeta is sufficient to yield dramatic benefit in vivo, and this understanding will be important in the clinical development of IKKbeta inhibitors.

Synthesis, initial SAR and biological evaluation of 1,6-dihydroimidazo[4,5-d]pyrrolo[2,3-b]pyridin-4-amine derived inhibitors of IkappaB kinase.

A new series of tricyclic-based inhibitors of IKK have been derived from an earlier lead compound. The synthesis and structure-activity relationships (SAR) are described. Compound 4k inhibited TNF production in rats stimulated with LPS.

Autoimmune pathways in mice and humans are blocked by pharmacological stabilization of the TYK2 pseudokinase domain.

TYK2 is a nonreceptor tyrosine kinase involved in adaptive and innate immune responses. A deactivating coding variant has previously been shown to prevent receptor-stimulated activation of this kinase and provides high protection from several common autoimmune diseases but without immunodeficiency. An agent that recapitulates the phenotype of this deactivating coding variant may therefore represent an important advancement in the treatment of autoimmunity. BMS-986165 is a potent oral agent that similarly blocks receptor-stimulated activation of TYK2 allosterically and with high selectivity and potency afforded through optimized binding to a regulatory domain of the protein. Signaling and functional responses in human TH17, TH1, B cells, and myeloid cells integral to autoimmunity were blocked by BMS-986165, both in vitro and in vivo in a phase 1 clinical trial. BMS-986165 demonstrated robust efficacy, consistent with blockade of multiple autoimmune pathways, in murine models of lupus nephritis and inflammatory bowel disease, supporting its therapeutic potential for multiple immune-mediated diseases.

Discovery of Branebrutinib (BMS-986195): A Strategy for Identifying a Highly Potent and Selective Covalent Inhibitor Providing Rapid in Vivo Inactivation of Bruton's Tyrosine Kinase (BTK).

Bruton's tyrosine kinase (BTK), a non-receptor tyrosine kinase, is a member of the Tec family of kinases and is essential for B cell receptor (BCR) mediated signaling. BTK also plays a critical role in the downstream signaling pathways for the Fcγ receptor in monocytes, the Fcε receptor in granulocytes, and the RANK receptor in osteoclasts. As a result, pharmacological inhibition of BTK is anticipated to provide an effective strategy for the clinical treatment of autoimmune diseases such as rheumatoid arthritis and lupus. This article will outline the evolution of our strategy to identify a covalent, irreversible inhibitor of BTK that has the intrinsic potency, selectivity, and pharmacokinetic properties necessary to provide a rapid rate of inactivation systemically following a very low dose. With excellent in vivo efficacy and a very desirable tolerability profile, 5a (branebrutinib, BMS-986195) has advanced into clinical studies.

Design, synthesis, and anti-inflammatory properties of orally active 4-(phenylamino)-pyrrolo[2,1-f][1,2,4]triazine p38alpha mitogen-activated protein kinase inhibitors.

A novel structural class of p38 mitogen-activated protein (MAP) kinase inhibitors consisting of substituted 4-(phenylamino)-pyrrolo[2,1- f][1,2,4]triazines has been discovered. An initial subdeck screen revealed that the oxindole-pyrrolo[2,1- f][1,2,4]triazine lead 2a displayed potent enzyme inhibition (IC 50 60 nM) and was active in a cell-based TNFalpha biosynthesis inhibition assay (IC 50 210 nM). Replacement of the C4 oxindole with 2-methyl-5- N-methoxybenzamide aniline 9 gave a compound with superior p38 kinase inhibition (IC 50 10 nM) and moderately improved functional inhibition in THP-1 cells. Further replacement of the C6 ester of the pyrrolo[2,1- f][1,2,4]triazine with amides afforded compounds with increased potency, excellent oral bioavailability, and robust efficacy in a murine model of acute inflammation (murine LPS-TNFalpha). In rodent disease models of chronic inflammation, multiple compounds demonstrated significant inhibition of disease progression leading to the advancement of 2 compounds 11b and 11j into further preclinical and toxicological studies.

Synthesis and SAR of new pyrrolo[2,1-f][1,2,4]triazines as potent p38 alpha MAP kinase inhibitors.

A novel series of compounds based on the pyrrolo[2,1-f][1,2,4]triazine ring system have been identified as potent p38 alpha MAP kinase inhibitors. The synthesis, structure-activity relationships (SAR), and in vivo activity of selected analogs from this class of inhibitors are reported. Additional studies based on X-ray co-crystallography have revealed that one of the potent inhibitors from this series binds to the DFG-out conformation of the p38 alpha enzyme.

The discovery of (R)-2-(sec-butylamino)-N-(2-methyl-5-(methylcarbamoyl)phenyl) thiazole-5-carboxamide (BMS-640994)-A potent and efficacious p38alpha MAP kinase inhibitor.

A novel structural class of p38alpha MAP kinase inhibitors has been identified via iterative SAR studies of a focused deck screen hit. Optimization of the lead series generated 6e, BMS-640994, a potent and selective p38alpha inhibitor that is orally efficacious in rodent models of acute and chronic inflammation.

A selective small molecule agonist of the melanocortin-1 receptor inhibits lipopolysaccharide-induced cytokine accumulation and leukocyte infiltration in mice.

It is well established that melanocortins are peptides that have potent anti-inflammatory activity. Recent research has focused on understanding which of the known melanocortin receptors mediates the anti-inflammatory actions of the melanocortins. The aim of this study was to assess the anti-inflammatory activity of a synthetic MC-1R agonist. BMS-470539 is a potent, selective, full agonist of human and murine MC-1R with EC(50) values in a cAMP accumulation assay of 16.8 and 11.6 nM, respectively. BMS-470539 dose-dependently inhibited TNF-alpha-induced activation of a NF-kappaB transcriptional reporter in human melanoma cells, which endogenously express MC-1R. In vivo studies with BMS-470539 demonstrated that subcutaneous administration of BMS-470539 resulted in a dose-dependent inhibition of LPS-induced TNF-alpha production in BALB/c mice. In this model, the compound had an ED(50) of approximately 10 micromol/kg and a pharmacodynamic half-life of approximately 8 h. Pharmacokinetic analysis of the compound indicated that the compound had a t(1/2) of 1.7 h. In a model of lung inflammation, administration of 15 micromol/kg BMS-470539 resulted in a 45% reduction in LPS-induced leukocyte infiltration (an infiltrate comprised primarily of neutrophils). The compound was also effective in a model of delayed-type hypersensitivity, reducing paw swelling by 59%, comparable with that seen with 5 mg/kg dexamethasone. These studies demonstrate that a selective small molecule agonist of the melanocortin-1 receptor is a potent anti-inflammatory agent in vivo and provides compelling evidence for the involvement of this receptor in the modulation of inflammation.

Bruton's tyrosine kinase inhibitor BMS-986142 in experimental models of rheumatoid arthritis enhances efficacy of agents representing clinical standard-of-care.

Bruton's tyrosine kinase (BTK) regulates critical signal transduction pathways involved in the pathobiology of rheumatoid arthritis (RA) and other autoimmune disorders. BMS-986142 is a potent and highly selective reversible small molecule inhibitor of BTK currently being investigated in clinical trials for the treatment of both RA and primary Sjögren's syndrome. In the present report, we detail the in vitro and in vivo pharmacology of BMS-986142 and show this agent provides potent and selective inhibition of BTK (IC50 = 0.5 nM), blocks antigen receptor-dependent signaling and functional endpoints (cytokine production, co-stimulatory molecule expression, and proliferation) in human B cells (IC50 ≤ 5 nM), inhibits Fcγ receptor-dependent cytokine production from peripheral blood mononuclear cells, and blocks RANK-L-induced osteoclastogenesis. Through the benefits of impacting these important drivers of autoimmunity, BMS-986142 demonstrated robust efficacy in murine models of rheumatoid arthritis (RA), including collagen-induced arthritis (CIA) and collagen antibody-induced arthritis (CAIA). In both models, robust efficacy was observed without continuous, complete inhibition of BTK. When a suboptimal dose of BMS-986142 was combined with other agents representing the current standard of care for RA (e.g., methotrexate, the TNFα antagonist etanercept, or the murine form of CTLA4-Ig) in the CIA model, improved efficacy compared to either agent alone was observed. The results suggest BMS-986142 represents a potential therapeutic for clinical investigation in RA, as monotherapy or co-administered with agents with complementary mechanisms of action.

Identification of a Potent, Selective, and Efficacious Phosphatidylinositol 3-Kinase δ (PI3Kδ) Inhibitor for the Treatment of Immunological Disorders.

PI3Kδ plays an important role controlling immune cell function and has therefore been identified as a potential target for the treatment of immunological disorders. This article highlights our work toward the identification of a potent, selective, and efficacious PI3Kδ inhibitor. Through careful SAR, the successful replacement of a polar pyrazole group by a simple chloro or trifluoromethyl group led to improved Caco-2 permeability, reduced Caco-2 efflux, reduced hERG PC activity, and increased selectivity profile while maintaining potency in the CD69 hWB assay. The optimization of the aryl substitution then identified a 4'-CN group that improved the human/rodent correlation in microsomal metabolic stability. Our lead molecule is very potent in PK/PD assays and highly efficacious in a mouse collagen-induced arthritis model.

2-aminothiazole as a novel kinase inhibitor template. Structure-activity relationship studies toward the discovery of N-(2-chloro-6-methylphenyl)-2-[[6-[4-(2-hydroxyethyl)-1- piperazinyl)]-2-methyl-4-pyrimidinyl]amino)]-1,3-thiazole-5-carboxamide (dasatinib, BMS-354825) as a potent pan-Src kinase inhibitor.

2-aminothiazole (1) was discovered as a novel Src family kinase inhibitor template through screening of our internal compound collection. Optimization through successive structure-activity relationship iterations identified analogs 2 (Dasatinib, BMS-354825) and 12m as pan-Src inhibitors with nanomolar to subnanomolar potencies in biochemical and cellular assays. Molecular modeling was used to construct a putative binding model for Lck inhibition by this class of compounds. The framework of key hydrogen-bond interactions proposed by this model was in agreement with the subsequent, published crystal structure of 2 bound to structurally similar Abl kinase. The oral efficacy of this class of inhibitors was demonstrated with 12m in inhibiting the proinflammatory cytokine IL-2 ex vivo in mice (ED50 approximately 5 mg/kg) and in reducing TNF levels in an acute murine model of inflammation (90% inhibition in LPS-induced TNFalpha production when dosed orally at 60 mg/kg, 2 h prior to LPS administration). The oral efficacy of 12m was further demonstrated in a chronic model of adjuvant arthritis in rats with established disease when administered orally at 0.3 and 3 mg/kg twice daily. Dasatinib (2) is currently in clinical trials for the treatment of chronic myelogenous leukemia.

Discovery of 6-Fluoro-5-(R)-(3-(S)-(8-fluoro-1-methyl-2,4-dioxo-1,2-dihydroquinazolin-3(4H)-yl)-2-methylphenyl)-2-(S)-(2-hydroxypropan-2-yl)-2,3,4,9-tetrahydro-1H-carbazole-8-carboxamide (BMS-986142): A Reversible Inhibitor of Bruton's Tyrosine Kinase (BTK) Conformationally Constrained by Two Locked Atropisomers.

Bruton's tyrosine kinase (BTK), a nonreceptor tyrosine kinase, is a member of the Tec family of kinases. BTK plays an essential role in B cell receptor (BCR)-mediated signaling as well as Fcγ receptor signaling in monocytes and Fcε receptor signaling in mast cells and basophils, all of which have been implicated in the pathophysiology of autoimmune disease. As a result, inhibition of BTK is anticipated to provide an effective strategy for the clinical treatment of autoimmune diseases such as lupus and rheumatoid arthritis. This article details the structure-activity relationships (SAR) leading to a novel series of highly potent and selective carbazole and tetrahydrocarbazole based, reversible inhibitors of BTK. Of particular interest is that two atropisomeric centers were rotationally locked to provide a single, stable atropisomer, resulting in enhanced potency and selectivity as well as a reduction in safety liabilities. With significantly enhanced potency and selectivity, excellent in vivo properties and efficacy, and a very desirable tolerability and safety profile, 14f (BMS-986142) was advanced into clinical studies.

5-Cyanopyrimidine derivatives as a novel class of potent, selective, and orally active inhibitors of p38alpha MAP kinase.

A novel class of 5-cyanopyrimidine-based inhibitors of p38alpha MAP kinase has been investigated. Analogues optimized through SAR iterations display low nanomolar enzymatic and cellular activity. The in vivo efficacy of this class of p38 inhibitors was demonstrated by 3a and 3b (>50% reduction in TNF levels when orally dosed at 5 mg/kg, 5 h prior to LPS administration in an acute murine model of inflammation). For 3a and 3b, the previously identified N-methoxybenzamide moiety (1) was replaced with N-(isoxazol-3-yl)benzamide, thereby providing increased metabolic stability. Cyanopyrimidine 3a demonstrated 100% oral bioavailability in mouse. High p38 kinase selectivity versus over 20 kinases was observed for analogue 3b. Direct hydrogen bonding of the cyano nitrogen of the 5-cyanopyrimidine core to the backbone NH of Met109 was confirmed by X-ray crystallographic analysis of 3a bound to p38alpha.

Rational design and synthesis of an orally active indolopyridone as a novel conformationally constrained cannabinoid ligand possessing antiinflammatory properties.

A series of unique indazoles and pyridoindolones have been rationally designed and synthesized as novel classes of cannabinoid ligands based on a proposed bioactive amide conformation. This has led to the discovery of the novel indolopyridone 3a as a conformationally constrained cannabinoid ligand that displays high affinity for the CB2 receptor (K(i)(CB2) = 1.0 nM) and possesses antiinflammatory properties when administered orally in an in vivo murine inflammation model.

The discovery of orally active triaminotriazine aniline amides as inhibitors of p38 MAP kinase.

A new structural class of triaminotriazine aniline amides possessing potent p38 enzyme activity has been discovered. The initial hit (compound 1a) was identified through screening the Pharmacopeia ECLiPS compound collection. SAR modification led to the identification of a short acting triaminotriazine aniline methoxyamide (compound 1m) possessing in vitro and in vivo oral activity in animal models of acute and chronic inflammatory disease. An X-ray crystal structure of compound 1m in this class, cocrystallized with unactivated p38 alpha protein, indicates that these compounds bind to the ATP binding pocket and possess key H-bonding interactions within a deeper cleft. Hydrogen bonding between one of the triazine nitrogens and the backbone NH of the Met109 residue occurs through a water molecule. The methoxyamide NH and carbonyl oxygen are within H-bonding distance of Glu71 and Asp168.

Discovery of 4-(5-(cyclopropylcarbamoyl)-2-methylphenylamino)-5-methyl-N-propylpyrrolo[1,2-f][1,2,4]triazine-6-carboxamide (BMS-582949), a clinical p38α MAP kinase inhibitor for the treatment of inflammatory diseases.

The discovery and characterization of 7k (BMS-582949), a highly selective p38α MAP kinase inhibitor that is currently in phase II clinical trials for the treatment of rheumatoid arthritis, is described. A key to the discovery was the rational substitution of N-cyclopropyl for N-methoxy in 1a, a previously reported clinical candidate p38α inhibitor. Unlike alkyl and other cycloalkyls, the sp(2) character of the cyclopropyl group can confer improved H-bonding characteristics to the directly substituted amide NH. Inhibitor 7k is slightly less active than 1a in the p38α enzymatic assay but displays a superior pharmacokinetic profile and, as such, was more effective in both the acute murine model of inflammation and pseudoestablished rat AA model. The binding mode of 7k with p38α was confirmed by X-ray crystallographic analysis.