ABSTRACT
Cytidine triphosphate synthase 1 (CTPS1) is necessary for an effective immune response, as revealed by severe immunodeficiency in CTPS1-deficient individuals [E. Martin et al], [Nature] [510], [288-292] ([2014]). CTPS1 expression is up-regulated in activated lymphocytes to expand CTP pools [E. Martin et al], [Nature] [510], [288-292] ([2014]), satisfying increased demand for nucleic acid and lipid synthesis [L. D. Fairbanks, M. Bofill, K. Ruckemann, H. A. Simmonds], [J. Biol. Chem. ] [270], [29682-29689] ([1995]). Demand for CTP in other tissues is met by the CTPS2 isoform and nucleoside salvage pathways [E. Martin et al], [Nature] [510], [288-292] ([2014]). Selective inhibition of the proliferative CTPS1 isoform is therefore desirable in the treatment of immune disorders and lymphocyte cancers, but little is known about differences in regulation of the isoforms or mechanisms of known inhibitors. We show that CTP regulates both isoforms by binding in two sites that clash with substrates. CTPS1 is less sensitive to CTP feedback inhibition, consistent with its role in increasing CTP levels in proliferation. We also characterize recently reported small-molecule inhibitors, both CTPS1 selective and nonselective. Cryo-electron microscopy (cryo-EM) structures reveal these inhibitors mimic CTP binding in one inhibitory site, where a single amino acid substitution explains selectivity for CTPS1. The inhibitors bind to CTPS assembled into large-scale filaments, which for CTPS1 normally represents a hyperactive form of the enzyme [E. M. Lynch et al], [Nat. Struct. Mol. Biol.] [24], [507-514] ([2017]). This highlights the utility of cryo-EM in drug discovery, particularly for cases in which targets form large multimeric assemblies not amenable to structure determination by other techniques. Both inhibitors also inhibit the proliferation of human primary T cells. The mechanisms of selective inhibition of CTPS1 lay the foundation for the design of immunosuppressive therapies.
Subject(s)
Carbon-Nitrogen Ligases/metabolism , Protein Isoforms/metabolism , Cell Proliferation/physiology , Humans , Immunologic Deficiency Syndromes/metabolism , T-Lymphocytes/metabolismABSTRACT
The derivatization of pharmaceuticals is a core activity in the discovery and development of new medicines. Late-stage functionalization via modern CH functionalization chemistry has emerged as a powerful technique with which to diversify advanced pharmaceutical intermediates. We report herein a case study in late-stage functionalization towards the development of a new class of indazole-based mineralocorticoid receptor antagonists (MRA). An effort to modify the electronics of the core indazole heterocycle inspired the use of modern CH borylation chemistry. New reactivity patterns were revealed and studied computationally. Ultimately, a de novo synthesis delivered a key 6-fluoroindazole compound 26, a potent MRA with excellent metabolic stability.
Subject(s)
Drug Development/methods , Indazoles/chemistry , Mineralocorticoid Receptor Antagonists/chemistry , Molecular StructureABSTRACT
The parallel medicinal chemistry (PMC) was effectively applied to accelerate the optimization of diacylglycerol O-acyltransferase I (DGAT-1) inhibitors. Through a highly collaborative and iterative library design, synthesis and testing, a benzimidazole lead was rapidly and systematically advanced to a highly potent, selective and bioavailable DGAT1 inhibitor with the potential for further development.
Subject(s)
Benzimidazoles/pharmacology , Diacylglycerol O-Acyltransferase/antagonists & inhibitors , Drug Discovery , Enzyme Inhibitors/pharmacology , Benzimidazoles/chemical synthesis , Benzimidazoles/chemistry , Chemistry, Pharmaceutical , Diacylglycerol O-Acyltransferase/metabolism , Dose-Response Relationship, Drug , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/chemistry , Humans , Molecular Structure , Structure-Activity RelationshipABSTRACT
Previously disclosed benzimidazole-based DGAT1 inhibitors containing a cyclohexane carboxylic acid moiety suffer from isomerization at the alpha position of the carboxylic acid group, generating active metabolites which exhibit DGAT1 inhibition comparable to the corresponding parent compounds. In this report, we describe the design, synthesis and profiling of benzimidazole-based DGAT1 inhibitors with a [3.1.0] bicyclohexane carboxylic acid moiety. Our results show that single isomer 3A maintains in vitro and in vivo inhibition against DGAT1. In contrast to previous lead compounds, 3A does not undergo isomerization during in vitro hepatocyte incubation study or in vivo mouse study.
Subject(s)
Benzimidazoles/chemistry , Carboxylic Acids/chemistry , Diacylglycerol O-Acyltransferase/antagonists & inhibitors , Enzyme Inhibitors/chemistry , Animals , Benzimidazoles/metabolism , Carboxylic Acids/metabolism , Chromatography, High Pressure Liquid , Cyclohexanones/chemistry , Diacylglycerol O-Acyltransferase/metabolism , Enzyme Inhibitors/analysis , Enzyme Inhibitors/metabolism , Hepatocytes/chemistry , Hepatocytes/metabolism , Humans , Inhibitory Concentration 50 , Isomerism , Mass Spectrometry , Mice , RatsABSTRACT
We describe the performance of multiple pose prediction methods for the D3R 2016 Grand Challenge. The pose prediction challenge includes 36 ligands, which represent 4 chemotypes and some miscellaneous structures against the FXR ligand binding domain. In this study we use a mix of fully automated methods as well as human-guided methods with considerations of both the challenge data and publicly available data. The methods include ensemble docking, colony entropy pose prediction, target selection by molecular similarity, molecular dynamics guided pose refinement, and pose selection by visual inspection. We evaluated the success of our predictions by method, chemotype, and relevance of publicly available data. For the overall data set, ensemble docking, visual inspection, and molecular dynamics guided pose prediction performed the best with overall mean RMSDs of 2.4, 2.2, and 2.2 Å respectively. For several individual challenge molecules, the best performing method is evaluated in light of that particular ligand. We also describe the protein, ligand, and public information data preparations that are typical of our binding mode prediction workflow.
Subject(s)
Computer-Aided Design , Drug Design , Drug Discovery , Molecular Docking Simulation , Receptors, Cytoplasmic and Nuclear/metabolism , Binding Sites , Crystallography, X-Ray , Databases, Protein , Humans , Ligands , Molecular Dynamics Simulation , Protein Binding , Protein Conformation , Receptors, Cytoplasmic and Nuclear/chemistry , ThermodynamicsABSTRACT
The 2016 D3R Grand Challenge 2 includes both pose and affinity or ranking predictions. This article is focused exclusively on affinity predictions submitted to the D3R challenge from a collaborative effort of the modeling and informatics group. Our submissions include ranking of 102 ligands covering 4 different chemotypes against the FXR ligand binding domain structure, and the relative binding affinity predictions of the two designated free energy subsets of 15 and 18 compounds. Using all the complex structures prepared in the same way allowed us to cover many types of workflows and compare their performances effectively. We evaluated typical workflows used in our daily structure-based design modeling support, which include docking scores, force field-based scores, QM/MM, MMGBSA, MD-MMGBSA, and MacroModel interaction energy estimations. The best performing methods for the two free energy subsets are discussed. Our results suggest that affinity ranking still remains very challenging; that the knowledge of more structural information does not necessarily yield more accurate predictions; and that visual inspection and human intervention are considerably important for ranking. Knowledge of the mode of action and protein flexibility along with visualization tools that depict polar and hydrophobic maps are very useful for visual inspection. QM/MM-based workflows were found to be powerful in affinity ranking and are encouraged to be applied more often. The standardized input and output enable systematic analysis and support methodology development and improvement for high level blinded predictions.
Subject(s)
Drug Discovery , Molecular Docking Simulation , Receptors, Cytoplasmic and Nuclear/metabolism , Thermodynamics , Workflow , Binding Sites , Computer-Aided Design , Databases, Protein , Drug Design , Humans , Ligands , Protein Binding , Protein Conformation , Receptors, Cytoplasmic and Nuclear/agonists , Receptors, Cytoplasmic and Nuclear/antagonists & inhibitors , Receptors, Cytoplasmic and Nuclear/chemistry , Spiro Compounds/chemistry , Spiro Compounds/pharmacology , Sulfonamides/chemistry , Sulfonamides/pharmacologyABSTRACT
We report the discovery and hit-to-lead optimization of a structurally novel indazole series of CYP11B2 inhibitors. Benchmark compound 34 from this series displays potent inhibition of CYP11B2, high selectivity versus related steroidal and hepatic CYP targets, and lead-like physical and pharmacokinetic properties. On the basis of these and other data, the indazole series was progressed to lead optimization for further refinement.
Subject(s)
Antihypertensive Agents/pharmacology , Cytochrome P-450 CYP11B2/antagonists & inhibitors , Hypertension/drug therapy , Indazoles/pharmacology , Animals , Antihypertensive Agents/chemical synthesis , Antihypertensive Agents/pharmacokinetics , Aromatase Inhibitors/chemical synthesis , Aromatase Inhibitors/pharmacokinetics , Aromatase Inhibitors/pharmacology , Cell Line , Cricetulus , Cytochrome P-450 CYP2D6 Inhibitors/chemical synthesis , Cytochrome P-450 CYP2D6 Inhibitors/pharmacokinetics , Cytochrome P-450 CYP2D6 Inhibitors/pharmacology , Humans , Indazoles/chemical synthesis , Indazoles/pharmacokinetics , Macaca mulatta , Male , Rats, Sprague-Dawley , Stereoisomerism , Steroid 11-beta-Hydroxylase/antagonists & inhibitorsABSTRACT
Multitask deep learning has emerged as a powerful tool for computational drug discovery. However, despite a number of preliminary studies, multitask deep networks have yet to be widely deployed in the pharmaceutical and biotech industries. This lack of acceptance stems from both software difficulties and lack of understanding of the robustness of multitask deep networks. Our work aims to resolve both of these barriers to adoption. We introduce a high-quality open-source implementation of multitask deep networks as part of the DeepChem open-source platform. Our implementation enables simple python scripts to construct, fit, and evaluate sophisticated deep models. We use our implementation to analyze the performance of multitask deep networks and related deep models on four collections of pharmaceutical data (three of which have not previously been analyzed in the literature). We split these data sets into train/valid/test using time and neighbor splits to test multitask deep learning performance under challenging conditions. Our results demonstrate that multitask deep networks are surprisingly robust and can offer strong improvement over random forests. Our analysis and open-source implementation in DeepChem provide an argument that multitask deep networks are ready for widespread use in commercial drug discovery.
Subject(s)
Drug Discovery/methods , Machine Learning , Absorption, Radiation , Inhibitory Concentration 50 , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/pharmacology , Serine Proteinase Inhibitors/chemistry , Serine Proteinase Inhibitors/pharmacology , Software , Ultraviolet RaysABSTRACT
The development of new antimalarial therapies is essential, and lowering the barrier of entry for the screening and discovery of new lead compound classes can spur drug development at organizations that may not have large compound screening libraries or resources to conduct high-throughput screens. Machine learning models have been long established to be more robust and have a larger domain of applicability with larger training sets. Screens over multiple data sets to find compounds with potential malaria blood stage inhibitory activity have been used to generate multiple Bayesian models. Here we describe a method by which Bayesian quantitative structure-activity relationship models, which contain information on thousands to millions of proprietary compounds, can be shared between collaborators at both for-profit and not-for-profit institutions. This model-sharing paradigm allows for the development of consensus models that have increased predictive power over any single model and yet does not reveal the identity of any compounds in the training sets.
Subject(s)
Antimalarials/pharmacology , Machine Learning , Malaria/drug therapy , Models, Theoretical , Quantitative Structure-Activity Relationship , Antimalarials/therapeutic use , Bayes Theorem , Drug Discovery , Malaria/blood , ROC Curve , TemperatureABSTRACT
Bioisosteres are integral components of modern pharmaceutical research that allow structural optimization to maximize in vivo efficacy and minimize adverse effects by selectively modifying pharmacodynamic, pharmacokinetic and physicochemical properties. A recent medicinal chemistry campaign focused on identifying small molecule inhibitors of prolylcarboxypeptidase (PrCP) initiated an investigation into the use of pyrazoles as bioisosteres for amides. The results indicate that pyrazoles are suitable bioisosteric replacements of amide functional groups. The study is an example of managing bioisosteric replacement by incorporating subsequent structural modifications to maintain potency against the selected target. A heuristic model for an embedded pharmacophore is also described.
Subject(s)
Carboxypeptidases/antagonists & inhibitors , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Pyrazoles/pharmacology , Animals , Carboxypeptidases/metabolism , Dose-Response Relationship, Drug , Enzyme Inhibitors/chemical synthesis , Humans , Mice , Molecular Structure , Pyrazoles/chemical synthesis , Pyrazoles/chemistry , Structure-Activity RelationshipABSTRACT
The synthesis, SAR, binding affinities and pharmacokinetic profiles are described for a series of cyclohexane-based prolylcarboxypeptidase (PrCP) inhibitors discovered by high throughput screening. Compounds show high levels of ex vivo target engagement in mouse plasma 20 h post oral dose.
Subject(s)
Carboxypeptidases/antagonists & inhibitors , Cyclohexanes/chemistry , Cyclohexanes/pharmacology , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Animals , Cyclohexanes/pharmacokinetics , Drug Discovery , Male , Mice , Mice, Inbred C57BL , Molecular Structure , Obesity/drug therapy , Structure-Activity RelationshipABSTRACT
Efforts were dedicated to develop potent and brain penetrant prolylcarboxypeptidase (PrCP) inhibitors by replacing the amide group of original leads 1 and 2 with heterocycles. Aminopyrimidines including compound 32a were identified to display good PrCP inhibitory activity (32a, IC(50)=43 nM) and impressive ability to penetrate brain in mice (brain/plasma ratio: 1.4).
Subject(s)
Amines/chemical synthesis , Brain/drug effects , Carboxypeptidases/antagonists & inhibitors , Drug Discovery , Enzyme Inhibitors , Heterocyclic Compounds/chemical synthesis , Heterocyclic Compounds/pharmacology , Amides/chemistry , Amines/chemistry , Amines/pharmacology , Animals , Brain/enzymology , Enzyme Activation/drug effects , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/pharmacology , Heterocyclic Compounds/chemistry , Inhibitory Concentration 50 , Mice , Molecular Structure , Structure-Activity RelationshipABSTRACT
Novel prolylcarboxypeptidase (PrCP) inhibitors with nanomolar IC(50) values were prepared by replacing the previously described dichlorobenzimidazole-substituted pyrrolidine amides with a variety of substituted benzylamine amides. In contrast to prior series, the compounds demonstrated minimal inhibition shift in whole serum and minimal recognition by P-glycoprotein (P-gp) efflux transporters. The compounds were also cell permeable and demonstrated in vivo brain exposure. The in vivo effect of compound (S)-6e on weight loss in an established diet-induced obesity (eDIO) mouse model was studied.
Subject(s)
Benzimidazoles/pharmacology , Brain/metabolism , Carboxypeptidases/antagonists & inhibitors , ATP Binding Cassette Transporter, Subfamily B/genetics , ATP Binding Cassette Transporter, Subfamily B/metabolism , Amides/chemistry , Animals , Biological Transport , Body Weight , Brain/drug effects , Disease Models, Animal , Humans , Inhibitory Concentration 50 , Mice , Models, Chemical , Obesity/drug therapy , Pyrrolidines/chemistry , Time FactorsABSTRACT
A series of benzodihydroisofurans were discovered as novel, potent, bioavailable and brain-penetrant prolylcarboxypeptidase (PrCP) inhibitors. The structure-activity relationship (SAR) is focused on improving PrCP activity and metabolic stability, and reducing plasma protein binding. In the established diet-induced obese (eDIO) mouse model, compound ent-3a displayed target engagement both in plasma and in brain. However, this compound failed to induce significant body weight loss in eDIO mice in a five-day study.
Subject(s)
Carboxypeptidases/antagonists & inhibitors , Drug Discovery , Enzyme Inhibitors/pharmacology , Furans/chemistry , Furans/pharmacology , Animals , Cells, Cultured , Disease Models, Animal , Drug Stability , Enzyme Activation/drug effects , Furans/chemical synthesis , Humans , Mice , Mice, Obese , Molecular Structure , Structure-Activity RelationshipABSTRACT
A new structural class of potent prolylcarboxypeptidase (PrCP) inhibitors was discovered by high-throughput screening. The series possesses a tractable SAR profile with sub-nanomolar in vitro IC(50) values. Compared to prior inhibitors, the new series demonstrated minimal activity shifts in pure plasma and complete ex vivo plasma target engagement in mouse plasma at the 20 h post-dose time point (po). In addition, the in vivo level of CNS and non-CNS drug exposure was measured.
Subject(s)
Carboxypeptidases/antagonists & inhibitors , Drug Discovery , Enzyme Inhibitors , Animals , Butanols/chemical synthesis , Butanols/chemistry , Butanols/pharmacology , Enzyme Activation/drug effects , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Inhibitory Concentration 50 , Male , Mice , Mice, Inbred C57BL , Molecular Structure , Obesity/drug therapy , Pyrrolidines/chemical synthesis , Pyrrolidines/chemistry , Pyrrolidines/pharmacologyABSTRACT
A series of potent inhibitors of prolylcarboxypeptidase (PrCP) was developed by modifying a lead structure that was discovered by high-throughput screening. The tert-butyl pyrrolidine was replaced by an aminocyclopentane to reduce the metabolic liabilities of the original lead. The compounds demonstrated sub-nanomolar in vitro IC(50) values, minimal activity shifts in pure plasma and improved pharmacokinetics. Complete ex vivo plasma target engagement was achieved with low brain exposure at the 20 h time point following p.o. dosing in a mouse. The results indicate that the aminocyclopentanes are useful tools for studying the therapeutic potential of peripheral (non-CNS) PrCP inhibition.
Subject(s)
Amines/pharmacology , Carboxypeptidases/antagonists & inhibitors , Cyclopentanes/pharmacology , Drug Discovery , Enzyme Inhibitors , Amines/chemical synthesis , Amines/chemistry , Animals , Cyclization , Cyclopentanes/chemical synthesis , Cyclopentanes/chemistry , Enzyme Activation/drug effects , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Inhibitory Concentration 50 , Male , Mice , Mice, Inbred C57BL , Molecular Structure , Obesity/drug therapyABSTRACT
Efforts to modify the central proline portion of lead compound 4 lead to the discovery of novel prolylcarboxypeptidase (PrCP) inhibitors. Especially, replacement with alanine afforded compound 19 displaying more potent human and mouse PrCP inhibitory activity than 4 and an overall comparable profile.
Subject(s)
Alanine/chemistry , Carboxypeptidases/antagonists & inhibitors , Drug Discovery , Alanine/pharmacology , Animals , Enzyme Activation/drug effects , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Humans , Inhibitory Concentration 50 , Mice , Molecular Structure , Structure-Activity RelationshipABSTRACT
Activation of PKG1α is a compelling strategy for the treatment of cardiovascular diseases. As the main effector of cyclic guanosine monophosphate (cGMP), activation of PKG1α induces smooth muscle relaxation in blood vessels, lowers pulmonary blood pressure, prevents platelet aggregation, and protects against cardiac stress. The development of activators has been mostly limited to cGMP mimetics and synthetic peptides. Described herein is the optimization of a piperidine series of small molecules to yield activators that demonstrate in vitro phosphorylation of vasodilator-stimulated phosphoprotein as well as antiproliferative effects in human pulmonary arterial smooth muscle cells. Hydrogen/deuterium exchange mass spectrometry experiments with the small molecule activators revealed a mechanism of action consistent with cGMP-induced activation, and an X-ray co-crystal structure with a construct encompassing the regulatory domains illustrated a binding mode in an allosteric pocket proximal to the low-affinity cyclic nucleotide-binding domain.
Subject(s)
Cyclic GMP-Dependent Protein Kinase Type I , Cyclic GMP , Cyclic GMP/metabolism , Cyclic GMP-Dependent Protein Kinase Type I/genetics , Cyclic GMP-Dependent Protein Kinase Type I/metabolism , Humans , Myocytes, Smooth Muscle , Phosphorylation , Protein Processing, Post-TranslationalABSTRACT
A series of benzimidazole pyrrolidinyl amides containing a piperidinyl group were discovered as novel prolylcarboxypeptidase (PrCP) inhibitors. Low-nanomolar IC(50)'s were achieved for several analogs, of which compound 9b displayed modest ex vivo target engagement in eDIO mouse plasma. Compound 9b was also studied in vivo for its effect on weight loss and food intake in an eDIO mouse model and the results will be discussed.
Subject(s)
Amides , Benzimidazoles , Carboxypeptidases/antagonists & inhibitors , Drug Discovery , Enzyme Inhibitors , Pyrrolidines , Amides/chemistry , Amides/pharmacology , Animals , Benzimidazoles/chemistry , Benzimidazoles/pharmacology , Disease Models, Animal , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Humans , Inhibitory Concentration 50 , Mice , Molecular Structure , Pyrrolidines/chemistry , Pyrrolidines/pharmacology , Structure-Activity RelationshipABSTRACT
PKG1α is a central node in cGMP signaling. Current therapeutics that look to activate this pathway rely on elevation of cGMP levels and subsequent activation of PKG1α. Direct activation of PKG1α could potentially drive additional efficacy without associated side effects of blanket cGMP elevation. We undertook a high-throughput screen to identify novel activators. After triaging through numerous false positive hits, attributed to compound mediated oxidation and activation of PKG1α, a piperidine series of compounds was validated. The hit 1 was a weak activator with EC50 = 47 µM. The activity could be improved to single digit micromolar, as seen in compounds 21 and 25 (7.0 and 3.7 µM, respectively). Several compounds were tested in a pVASP cell-based assay, and for compounds with moderate permeability, good agreement was observed between the biochemical and functional assays. These compounds will function as efficient tools to further interrogate PKG1α biology.