ABSTRACT
Guided by co-crystal structural information obtained from a different series we were exploring, a scaffold morphing and SBDD approach led to the discovery of the 1,4-disubstituted indazole series as a novel class of GKAs that potently activate GK in enzyme and cell assays. anti-diabetic OGTT efficacy was demonstrated with 29 in a rodent models of type 2 diabetes.
Subject(s)
Diabetes Mellitus, Experimental/drug therapy , Diabetes Mellitus, Type 2/drug therapy , Drug Discovery , Enzyme Activators/pharmacology , Glucokinase/metabolism , Indazoles/pharmacology , Administration, Oral , Allosteric Regulation/drug effects , Animals , Diabetes Mellitus, Experimental/metabolism , Diabetes Mellitus, Type 2/metabolism , Dose-Response Relationship, Drug , Enzyme Activators/administration & dosage , Enzyme Activators/chemistry , Ether-A-Go-Go Potassium Channels/antagonists & inhibitors , Ether-A-Go-Go Potassium Channels/metabolism , Glucose Tolerance Test , Humans , Indazoles/administration & dosage , Indazoles/chemistry , Mice , Mice, Inbred C57BL , Mice, Obese , Models, Molecular , Molecular Structure , Potassium Channel Blockers/pharmacology , Structure-Activity RelationshipABSTRACT
The MAPK signaling cascade, comprised of several linear and intersecting pathways, propagates signaling into the nucleus resulting in cytokine and chemokine release. The Map Kinase Kinase isoforms 3 and 6 (MKK3 and MKK6) are responsible for the phosphorylation and activation of p38, and are hypothesized to play a key role in regulating this pathway without the redundancy seen in downstream effectors. Using FBDD, we have discovered efficient and selective inhibitors of MKK3 and MKK6 that can serve as tool molecules to help further understand the role of these kinases in MAPK signaling, and the potential impact of inhibiting kinases upstream of p38.
Subject(s)
Drug Design , MAP Kinase Kinase 3/antagonists & inhibitors , MAP Kinase Kinase 6/antagonists & inhibitors , Protein Kinase Inhibitors/chemistry , Binding Sites , Humans , MAP Kinase Kinase 3/metabolism , MAP Kinase Kinase 6/metabolism , MAP Kinase Signaling System/drug effects , Molecular Docking Simulation , Protein Kinase Inhibitors/metabolism , Protein Kinase Inhibitors/pharmacology , Protein Structure, Tertiary , U937 Cells , p38 Mitogen-Activated Protein Kinases/metabolismABSTRACT
Spleen Tyrosine Kinase (SYK) is a non-receptor cytoplasmic tyrosine kinase that is primarily expressed in hematopoietic cells. SYK is a key mediator for a variety of inflammatory cells, including B cells, mast cells, macrophages and neutrophils and therefore, an attractive approach for treatment of both inflammatory diseases and oncology indications. Using in house co-crystal structure information, and structure-based drug design, we designed and optimized a novel series of heteroaromatic pyrrolidinone SYK inhibitors resulting in the selection of the development candidate TAK-659. TAK-659 is currently undergoing Phase I clinical trials for advanced solid tumor and lymphoma malignancies, a Phase Ib study in advanced solid tumors in combination with nivolumab, and PhIb/II trials for relapsed/refractory AML.
Subject(s)
Drug Discovery , Protein Kinase Inhibitors/pharmacology , Pyrimidines/pharmacology , Pyrrolidinones/pharmacology , Syk Kinase/antagonists & inhibitors , Animals , Antineoplastic Agents , Cell Line, Tumor , Cell Proliferation/drug effects , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Humans , Mice , Mice, Inbred C57BL , Molecular Structure , Neoplasms, Experimental/drug therapy , Neoplasms, Experimental/pathology , Protein Kinase Inhibitors/administration & dosage , Protein Kinase Inhibitors/chemistry , Pyrimidines/administration & dosage , Pyrimidines/chemistry , Pyrrolidinones/administration & dosage , Pyrrolidinones/chemistry , Structure-Activity Relationship , Syk Kinase/metabolismABSTRACT
The RAS/RAF/MEK/ERK signal transduction cascade plays an important role in the regulation of critical cellular processes such as cell proliferation, migration, and differentiation. The up-regulation of this pathway can negatively affect cell homeostasis and is responsible for the development of various forms of cancer and inflammation processes. Therefore, there is a strong interest in pursuing drug programs targeting some of the enzymes involved in this pathway. In addition to the determination of Ki, Kd, IC50, and/or EC50, a more thorough kinetic analysis can provide useful information for the selection of the best lead series during the early stage of the drug discovery process. This study describes a medium-throughput fluorescent probe displacement assay for the rapid determination of the k(off) constant, residence time, and kinetic efficiency for ERK (extracellular signal-regulated kinase) inhibitors. Using this method, we have identified several inhibitors that we have subjected to further kinetic analysis by comparing k(off) constants determined for these time-dependent inhibitors using either the active or inactive form of ERK2.
Subject(s)
Drug Evaluation, Preclinical/methods , High-Throughput Screening Assays/methods , Mitogen-Activated Protein Kinase 1/antagonists & inhibitors , Protein Kinase Inhibitors/pharmacology , Fluorescent Dyes/chemistry , Kinetics , Mitogen-Activated Protein Kinase 1/chemistryABSTRACT
Structure based drug design of a series of novel 1,4-benzoxazin-3-one derived PARP-1 inhibitors are described. The synthesis, enzymatic & cellular activities and pharmacodynamic effects are described. Optimized analogs demonstrated inhibition of poly-ADP-ribosylation in SW620 tumor bearing nude mice through 24h following a single dose.
Subject(s)
Benzoxazines/chemistry , Benzoxazines/pharmacology , Drug Design , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/pharmacology , Poly(ADP-ribose) Polymerase Inhibitors , Animals , Cell Line, Tumor , Enzyme Activation/drug effects , Enzyme Inhibitors/chemistry , Humans , Mice , Mice, Nude , Molecular StructureSubject(s)
Biomedical Research/methods , Information Dissemination/ethics , Molecular Probes/chemistry , Small Molecule Libraries/chemistry , Biomedical Research/instrumentation , Humans , Intellectual Property , Internet , Molecular Probes/pharmacology , Molecular Weight , Sensitivity and Specificity , Small Molecule Libraries/pharmacologyABSTRACT
Two different signaling pathways lead to the activation of the transcription factor NF-κB, initiating distinct biological responses: The canonical NF-κB pathway activation has been implicated in host immunity and inflammatory responses, whereas the noncanonical pathway activation has been involved in lymphoid organ development and B-cell maturation, as well as in the development of chronic inflammatory diseases and some hematologic cancers. The NF-κB-inducing kinase (NIK) is a cytoplasmic Ser/Thr kinase and is a key regulator of the noncanonical pathway. NIK activation results in the processing of the p100 subunit to p52, leading to the formation of the RelB/p52 complex and noncanonical pathway activation. Because of its role in the development of lymphoid malignancies, this kinase has always been considered as an attractive target for the treatment of certain types of cancers and immune diseases. We at Takeda have pursued a drug discovery program to identify small-molecule inhibitors against NIK. This report provides an overview of the data generated from our screening campaign using a small fragment library. Most importantly, we also provide a kinetic analysis of published compounds and chemical series developed at Takeda that are associated with a slow tight-binding mechanism and excellent cellular potency.
Subject(s)
Drug Discovery/methods , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/pharmacology , Protein Serine-Threonine Kinases/antagonists & inhibitors , Protein Serine-Threonine Kinases/chemistry , Drug Screening Assays, Antitumor/methods , Humans , Protein Binding , Signal Transduction/drug effects , Small Molecule Libraries , NF-kappaB-Inducing KinaseABSTRACT
A series of N-(2-amino-5-substituted phenyl)benzamides (3-21) were designed, synthesized and evaluated for their inhibition of HDAC2 and their cytotoxicity in HCT116 cancer cells. Multiple compounds from this series demonstrated time-dependent binding kinetics that is rationalized using a co-complex crystal structure of HDAC2 and N-(4-aminobiphenyl-3-yl)benzamide (6).
Subject(s)
Benzamides/chemistry , Histone Deacetylase 2/antagonists & inhibitors , Histone Deacetylase Inhibitors/chemical synthesis , Benzamides/chemical synthesis , Benzamides/toxicity , Binding Sites , Catalytic Domain , Crystallography, X-Ray , HCT116 Cells , Histone Deacetylase 2/metabolism , Histone Deacetylase Inhibitors/chemistry , Histone Deacetylase Inhibitors/toxicity , Humans , Kinetics , Structure-Activity RelationshipABSTRACT
Protein arginine methyltransferase (PRMT) 4 (also known as coactivator-associated arginine methyltransferase 1; CARM1) is involved in a variety of biological processes and is considered as a candidate oncogene owing to its overexpression in several types of cancer. Selective PRMT4 inhibitors are useful tools for clarifying the molecular events regulated by PRMT4 and for validating PRMT4 as a therapeutic target. Here, we report the discovery of TP-064, a potent, selective, and cell-active chemical probe of human PRMT4 and its co-crystal structure with PRMT4. TP-064 inhibited the methyltransferase activity of PRMT4 with high potency (half-maximal inhibitory concentration, IC50 < 10 nM) and selectivity over other PRMT family proteins, and reduced arginine dimethylation of the PRMT4 substrates BRG1-associated factor 155 (BAF155; IC50= 340 ± 30 nM) and Mediator complex subunit 12 (MED12; IC50 = 43 ± 10 nM). TP-064 treatment inhibited the proliferation of a subset of multiple myeloma cell lines, with affected cells arrested in G1 phase of the cell cycle. TP-064 and its negative control (TP-064N) will be valuable tools to further investigate the biology of PRMT4 and the therapeutic potential of PRMT4 inhibition.
ABSTRACT
Potent, selective and broadly characterized small molecule modulators of protein function (chemical probes) are powerful research reagents. The pharmaceutical industry has generated many high-quality chemical probes and several of these have been made available to academia. However, probe-associated data and control compounds, such as inactive structurally related molecules and their associated data, are generally not accessible. The lack of data and guidance makes it difficult for researchers to decide which chemical tools to choose. Several pharmaceutical companies (AbbVie, Bayer, Boehringer Ingelheim, Janssen, MSD, Pfizer, and Takeda) have therefore entered into a pre-competitive collaboration to make available a large number of innovative high-quality probes, including all probe-associated data, control compounds and recommendations on use (
Subject(s)
Molecular Probes/metabolism , Pharmacology/methods , Proteins/metabolism , Technology, Pharmaceutical/methodsABSTRACT
Prolyl hydroxylases (PHDs) down-regulate the level of hypoxia-inducible factors (HIFs) by hydroxylating key proline residues that trigger the degradation of the protein and affect the cell and its ability to respond to hypoxic stress. Several small molecule PHD inhibitors are now in various preclinical and clinical stages for the treatment of anemia. The present study provides a detail kinetic analysis for some of these inhibitors. The data generated in the present study suggest that these compounds are reversible and compete directly with the co-substrate, 2-oxoglutarate (2-OG) for binding at the enzyme active site. Most of these compounds are pan PHD inhibitors and exhibit a time-dependent inhibition (TDI) mechanism due to an extremely slow dissociation rate constant, koff, and a long residence time.
Subject(s)
Enzyme Inhibitors/pharmacology , Hypoxia-Inducible Factor-Proline Dioxygenases/antagonists & inhibitors , Small Molecule Libraries/pharmacology , Catalytic Domain , Enzyme Inhibitors/chemistry , Humans , Hypoxia-Inducible Factor-Proline Dioxygenases/chemistry , Hypoxia-Inducible Factor-Proline Dioxygenases/metabolism , Ketoglutaric Acids/metabolism , Kinetics , Protein Binding , Small Molecule Libraries/chemistryABSTRACT
Trelagliptin (SYR-472), a novel dipeptidyl peptidase-4 inhibitor, shows sustained efficacy by once-weekly dosing in type 2 diabetes patients. In this study, we characterized in vitro properties of trelagliptin, which exhibited approximately 4- and 12-fold more potent inhibition against human dipeptidyl peptidase-4 than alogliptin and sitagliptin, respectively, and >10,000-fold selectivity over related proteases including dipeptidyl peptidase-8 and dipeptidyl peptidase-9. Kinetic analysis revealed reversible, competitive and slow-binding inhibition of dipeptidyl peptidase-4 by trelagliptin (t1/2 for dissociation ≈ 30 minutes). X-ray diffraction data indicated a non-covalent interaction between dipeptidyl peptidase and trelagliptin. Taken together, potent dipeptidyl peptidase inhibition may partially contribute to sustained efficacy of trelagliptin.