ABSTRACT
[This corrects the article DOI: 10.1021/acsmedchemlett.8b00344.].
ABSTRACT
RIP2 kinase has been identified as a key signal transduction partner in the NOD2 pathway contributing to a variety of human pathologies, including immune-mediated inflammatory diseases. Small-molecule inhibitors of RIP2 kinase or its signaling partners on the NOD2 pathway that are suitable for advancement into the clinic have yet to be described. Herein, we report our discovery and profile of the prodrug clinical compound, inhibitor 3, currently in phase 1 clinical studies. Compound 3 potently binds to RIP2 kinase with good kinase specificity and has excellent activity in blocking many proinflammatory cytokine responses in vivo and in human IBD explant samples. The highly favorable physicochemical and ADMET properties of 3 combined with high potency led to a predicted low oral dose in humans.
Subject(s)
Benzothiazoles/pharmacology , Phosphates/pharmacology , Protein Kinase Inhibitors/pharmacology , Quinazolines/pharmacology , Receptor-Interacting Protein Serine-Threonine Kinase 2/antagonists & inhibitors , Animals , Benzothiazoles/chemistry , Benzothiazoles/pharmacokinetics , Benzothiazoles/therapeutic use , Colitis/drug therapy , Dogs , Drug Discovery , Humans , Male , Mice , Molecular Docking Simulation , Phosphates/chemistry , Phosphates/pharmacokinetics , Phosphates/therapeutic use , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/pharmacokinetics , Protein Kinase Inhibitors/therapeutic use , Quinazolines/chemistry , Quinazolines/pharmacokinetics , Quinazolines/therapeutic use , Rats, Sprague-Dawley , Receptor-Interacting Protein Serine-Threonine Kinase 2/metabolism , Swine , Swine, MiniatureABSTRACT
RIP1 kinase regulates necroptosis and inflammation and may play an important role in contributing to a variety of human pathologies, including inflammatory and neurological diseases. Currently, RIP1 kinase inhibitors have advanced into early clinical trials for evaluation in inflammatory diseases such as psoriasis, rheumatoid arthritis, and ulcerative colitis and neurological diseases such as amyotrophic lateral sclerosis and Alzheimer's disease. In this paper, we report on the design of potent and highly selective dihydropyrazole (DHP) RIP1 kinase inhibitors starting from a high-throughput screen and the lead-optimization of this series from a lead with minimal rat oral exposure to the identification of dihydropyrazole 77 with good pharmacokinetic profiles in multiple species. Additionally, we identified a potent murine RIP1 kinase inhibitor 76 as a valuable in vivo tool molecule suitable for evaluating the role of RIP1 kinase in chronic models of disease. DHP 76 showed efficacy in mouse models of both multiple sclerosis and human retinitis pigmentosa.
Subject(s)
Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/pharmacology , Nuclear Pore Complex Proteins/antagonists & inhibitors , Pyrazoles/chemical synthesis , Pyrazoles/pharmacology , RNA-Binding Proteins/antagonists & inhibitors , Animals , Biological Availability , Cell Line , Chronic Disease , Drug Design , Encephalomyelitis, Autoimmune, Experimental/drug therapy , Enzyme Inhibitors/pharmacokinetics , Haplorhini , High-Throughput Screening Assays , Humans , Mice , Mice, Inbred C57BL , Models, Molecular , Multiple Sclerosis/drug therapy , Pyrazoles/pharmacokinetics , Rats , Retinitis Pigmentosa/drug therapy , Structure-Activity RelationshipABSTRACT
RIP2 kinase was recently identified as a therapeutic target for a variety of autoimmune diseases. We have reported previously a selective 4-aminoquinoline-based RIP2 inhibitor GSK583 and demonstrated its effectiveness in blocking downstream NOD2 signaling in cellular models, rodent in vivo models, and human ex vivo disease models. While this tool compound was valuable in validating the biological pathway, it suffered from activity at the hERG ion channel and a poor PK/PD profile thereby limiting progression of this analog. Herein, we detail our efforts to improve both this off-target liability as well as the PK/PD profile of this series of inhibitors through modulation of lipophilicity and strengthening hinge binding ability. These efforts have led to inhibitor 7, which possesses high binding affinity for the ATP pocket of RIP2 (IC50 = 1 nM) and inhibition of downstream cytokine production in human whole blood (IC50 = 10 nM) with reduced hERG activity (14 µM).
ABSTRACT
RIP1 regulates necroptosis and inflammation and may play an important role in contributing to a variety of human pathologies, including immune-mediated inflammatory diseases. Small-molecule inhibitors of RIP1 kinase that are suitable for advancement into the clinic have yet to be described. Herein, we report our lead optimization of a benzoxazepinone hit from a DNA-encoded library and the discovery and profile of clinical candidate GSK2982772 (compound 5), currently in phase 2a clinical studies for psoriasis, rheumatoid arthritis, and ulcerative colitis. Compound 5 potently binds to RIP1 with exquisite kinase specificity and has excellent activity in blocking many TNF-dependent cellular responses. Highlighting its potential as a novel anti-inflammatory agent, the inhibitor was also able to reduce spontaneous production of cytokines from human ulcerative colitis explants. The highly favorable physicochemical and ADMET properties of 5, combined with high potency, led to a predicted low oral dose in humans.
Subject(s)
Anti-Inflammatory Agents/chemistry , Anti-Inflammatory Agents/pharmacology , Colitis, Ulcerative/drug therapy , Inflammation/drug therapy , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/pharmacology , Receptor-Interacting Protein Serine-Threonine Kinases/antagonists & inhibitors , Animals , Benzazepines/chemistry , Benzazepines/pharmacology , Colitis, Ulcerative/immunology , Cytokines/immunology , Dogs , Haplorhini , Humans , Inflammation/immunology , Mice , Molecular Docking Simulation , Rabbits , Rats , Receptor-Interacting Protein Serine-Threonine Kinases/immunology , Swine , Swine, Miniature , Tumor Necrosis Factor-alpha/immunologyABSTRACT
RIP2 kinase is a central component of the innate immune system and enables downstream signaling following activation of the pattern recognition receptors NOD1 and NOD2, leading to the production of inflammatory cytokines. Recently, several inhibitors of RIP2 kinase have been disclosed that have contributed to the fundamental understanding of the role of RIP2 in this pathway. However, because they lack either broad kinase selectivity or strong affinity for RIP2, these tools have only limited utility to assess the role of RIP2 in complex environments. We present, herein, the discovery and pharmacological characterization of GSK583, a next-generation RIP2 inhibitor possessing exquisite selectivity and potency. Having demonstrated the pharmacological precision of this tool compound, we report its use in elucidating the role of RIP2 kinase in a variety of in vitro, in vivo, and ex vivo experiments, further clarifying our understanding of the role of RIP2 in NOD1 and NOD2 mediated disease pathogenesis.
Subject(s)
Aminoquinolines/pharmacology , Protein Kinase Inhibitors/pharmacology , Receptor-Interacting Protein Serine-Threonine Kinase 2/antagonists & inhibitors , Sulfones/pharmacology , Aminoquinolines/blood , Aminoquinolines/chemistry , Animals , Dose-Response Relationship, Drug , Female , Humans , Male , Mice , Mice, Inbred C57BL , Models, Molecular , Molecular Structure , Protein Kinase Inhibitors/blood , Protein Kinase Inhibitors/chemistry , Rats , Rats, Sprague-Dawley , Receptor-Interacting Protein Serine-Threonine Kinase 2/metabolism , Structure-Activity Relationship , Sulfones/blood , Sulfones/chemistryABSTRACT
The recent discovery of the role of receptor interacting protein 1 (RIP1) kinase in tumor necrosis factor (TNF)-mediated inflammation has led to its emergence as a highly promising target for the treatment of multiple inflammatory diseases. We screened RIP1 against GSK's DNA-encoded small-molecule libraries and identified a novel highly potent benzoxazepinone inhibitor series. We demonstrate that this template possesses complete monokinase selectivity for RIP1 plus unique species selectivity for primate versus nonprimate RIP1. We elucidate the conformation of RIP1 bound to this benzoxazepinone inhibitor driving its high kinase selectivity and design specific mutations in murine RIP1 to restore potency to levels similar to primate RIP1. This series differentiates itself from known RIP1 inhibitors in combining high potency and kinase selectivity with good pharmacokinetic profiles in rodents. The favorable developability profile of this benzoxazepinone template, as exemplified by compound 14 (GSK'481), makes it an excellent starting point for further optimization into a RIP1 clinical candidate.
Subject(s)
DNA/chemistry , Isoxazoles/pharmacology , Oxazepines/pharmacology , Protein Kinase Inhibitors/pharmacology , Receptor-Interacting Protein Serine-Threonine Kinases/antagonists & inhibitors , Small Molecule Libraries/pharmacology , Animals , Cell Line, Tumor , Crystallography, X-Ray , Dose-Response Relationship, Drug , HT29 Cells , Humans , Isoxazoles/chemical synthesis , Isoxazoles/chemistry , Mice , Models, Molecular , Molecular Structure , Oxazepines/chemical synthesis , Oxazepines/chemistry , Protein Kinase Inhibitors/chemical synthesis , Protein Kinase Inhibitors/chemistry , Receptor-Interacting Protein Serine-Threonine Kinases/metabolism , Small Molecule Libraries/chemical synthesis , Small Molecule Libraries/chemistry , Structure-Activity Relationship , U937 CellsABSTRACT
Potent inhibitors of RIP1 kinase from three distinct series, 1-aminoisoquinolines, pyrrolo[2,3-b]pyridines, and furo[2,3-d]pyrimidines, all of the type II class recognizing a DLG-out inactive conformation, were identified from screening of our in-house kinase focused sets. An exemplar from the furo[2,3-d]pyrimidine series showed a dose proportional response in protection from hypothermia in a mouse model of TNFα induced lethal shock.
