ABSTRACT
Lipids are used as cellular building blocks and condensed energy stores and also act as signaling molecules. The glycerolipid/ fatty acid cycle, encompassing lipolysis and lipogenesis, generates many lipid signals. Reliable procedures are not available for measuring activities of several lipolytic enzymes for the purposes of drug screening, and this resulted in questionable selectivity of various known lipase inhibitors. We now describe simple assays for lipolytic enzymes, including adipose triglyceride lipase (ATGL), hormone sensitive lipase (HSL), sn-1-diacylglycerol lipase (DAGL), monoacylglycerol lipase, α/ß-hydrolase domain 6, and carboxylesterase 1 (CES1) using recombinant human and mouse enzymes either in cell extracts or using purified enzymes. We observed that many of the reported inhibitors lack specificity. Thus, Cay10499 (HSL inhibitor) and RHC20867 (DAGL inhibitor) also inhibit other lipases. Marked differences in the inhibitor sensitivities of human ATGL and HSL compared with the corresponding mouse enzymes was noticed. Thus, ATGListatin inhibited mouse ATGL but not human ATGL, and the HSL inhibitors WWL11 and Compound 13f were effective against mouse enzyme but much less potent against human enzyme. Many of these lipase inhibitors also inhibited human CES1. Results describe reliable assays for measuring lipase activities that are amenable for drug screening and also caution about the specificity of the many earlier described lipase inhibitors.
Subject(s)
Drug Discovery/methods , Enzyme Inhibitors/pharmacology , Lipolysis/drug effects , Adipose Tissue/metabolism , Animals , Carboxylic Ester Hydrolases/antagonists & inhibitors , Carboxylic Ester Hydrolases/metabolism , Fatty Acids/metabolism , Humans , Lipase/antagonists & inhibitors , Lipase/metabolism , Lipogenesis/physiology , Lipolysis/physiology , Lipoprotein Lipase/antagonists & inhibitors , Lipoprotein Lipase/metabolism , Mice , Monoacylglycerol Lipases/metabolism , Sterol Esterase/antagonists & inhibitors , Sterol Esterase/metabolism , Triglycerides/metabolismABSTRACT
ATR is a key kinase in the DNA-damage response (DDR) that is synthetic lethal with several other DDR proteins, making it an attractive target for the treatment of genetically selected solid tumors. Herein we describe the discovery of a novel ATR inhibitor guided by a pharmacophore model to position a key hydrogen bond. Optimization was driven by potency and selectivity over the related kinase mTOR, resulting in the identification of camonsertib (RP-3500) with high potency and excellent ADME properties. Preclinical evaluation focused on the impact of camonsertib on myelosuppression, and an exploration of intermittent dosing schedules to allow recovery of the erythroid compartment and mitigate anemia. Camonsertib is currently undergoing clinical evaluation both as a single agent and in combination with talazoparib, olaparib, niraparib, lunresertib, or gemcitabine (NCT04497116, NCT04972110, NCT04855656). A preliminary recommended phase 2 dose for monotherapy was identified as 160 mg QD given 3 days/week.
Subject(s)
Neoplasms , Humans , Ataxia Telangiectasia Mutated Proteins , Neoplasms/drug therapy , Protein Kinase Inhibitors/pharmacology , Protein Kinase Inhibitors/therapeutic use , GemcitabineABSTRACT
The trifluoroethylamine group found in cathepsin K inhibitors like odanacatib can be replaced by a difluoroethylamine group. This change increased the basicity of the nitrogen which positively impacted the log D. This translated into an improved oral bioavailability in pre-clinical species. Difluoroethylamine compounds exhibit a similar potency against cathepsin K and selectivity profile against other cathepsins when compared to trifluoroethylamine analogs.
Subject(s)
Cathepsin K/antagonists & inhibitors , Ethylamines/chemistry , Protease Inhibitors/chemistry , Administration, Oral , Amides/chemistry , Animals , Biphenyl Compounds/chemistry , Biphenyl Compounds/pharmacology , Cathepsin K/metabolism , Dogs , Ethylamines/chemical synthesis , Ethylamines/pharmacokinetics , Protease Inhibitors/chemical synthesis , Protease Inhibitors/pharmacokinetics , RatsABSTRACT
A weak antagonist of the pyrimidinergic receptor P2Y(14) containing a dihydropyridopyrimidine core was identified through high-throughput screening. Subsequent optimization led to potent, non-UTP competitive antagonists and represent the first reported non-nucleotide antagonists of this receptor. Compound 18q was identified as a 10 nM P2Y(14) antagonist with good oral bioavailability and provided sufficient exposure in mice to be used as a tool for future in vivo studies.
Subject(s)
Purinergic P2 Receptor Antagonists/chemical synthesis , Pyrimidines/chemical synthesis , Receptors, Purinergic P2/chemistry , Administration, Oral , Animals , Biological Availability , Mice , Molecular Structure , Pan troglodytes , Purinergic P2 Receptor Antagonists/chemistry , Pyrimidines/administration & dosage , Pyrimidines/chemistry , Receptors, Purinergic P2Y , Structure-Activity RelationshipABSTRACT
In this manuscript we wish to report the discovery of MK-7246 (4), a potent and selective CRTH2 (DP2) antagonist. SAR studies leading to MK-7246 along with two synthetic sequences enabling the preparation of this novel class of CRTH2 antagonist are reported. Finally, the pharmacokinetic and metabolic profile of MK-7246 is disclosed.
Subject(s)
Carbolines/chemistry , Lung Diseases/drug therapy , Receptors, Immunologic/antagonists & inhibitors , Receptors, Prostaglandin/antagonists & inhibitors , Animals , Carbolines/pharmacokinetics , Carbolines/therapeutic use , Humans , Macaca mulatta , Microsomes, Liver/metabolism , Rats , Rats, Sprague-Dawley , Receptors, Immunologic/metabolism , Receptors, Prostaglandin/metabolism , Structure-Activity RelationshipABSTRACT
MK-0674 is a potent and selective cathepsin K inhibitor from the same structural class as odanacatib with a comparable inhibitory potency profile against Cat K. It is orally bioavailable and exhibits long half-life in pre-clinical species. In vivo studies using deuterated MK-0674 show stereoselective epimerization of the alcohol stereocenter via an oxidation/reduction cycle. From in vitro incubations, two metabolites could be identified: the hydroxyleucine and the glucuronide conjugate which were confirmed using authentic synthetic standards.
Subject(s)
Biphenyl Compounds/administration & dosage , Biphenyl Compounds/pharmacokinetics , Cathepsin K/antagonists & inhibitors , Cysteine Proteinase Inhibitors/administration & dosage , Cysteine Proteinase Inhibitors/pharmacokinetics , Drug Discovery/methods , Administration, Oral , Animals , Biological Availability , Biphenyl Compounds/chemistry , Cathepsin K/metabolism , Cysteine Proteinase Inhibitors/chemistry , Dogs , Hepatocytes/drug effects , Hepatocytes/metabolism , Humans , Macaca mulatta , Rabbits , RatsABSTRACT
Odanacatib is a potent, selective, and neutral cathepsin K inhibitor which was developed to address the metabolic liabilities of the Cat K inhibitor L-873724. Substituting P1 and modifying the P2 side chain led to a metabolically robust inhibitor with a long half-life in preclinical species. Odanacatib was more selective in whole cell assays than the published Cat K inhibitors balicatib and relacatib. Evaluation in dermal fibroblast culture showed minimal intracellular collagen accumulation relative to less selective Cat K inhibitors.
Subject(s)
Biphenyl Compounds/pharmacology , Cathepsins/antagonists & inhibitors , Cysteine Proteinase Inhibitors/pharmacology , Animals , Azepines/chemistry , Azepines/pharmacology , Cathepsin K , Collagen/drug effects , Collagen/immunology , Dogs , Fibroblasts/drug effects , Humans , Models, Biological , Molecular Structure , Osteoporosis, Postmenopausal/drug therapy , Skin/cytology , Structure-Activity Relationship , Sulfones/chemistry , Sulfones/pharmacologyABSTRACT
Identification, characterization and structure elucidation of human metabolites of drug candidates is crucial for the pharmaceutical industry to assess their activity against the therapeutic target of interest and potential toxicological effects. It often requires in vitro synthesis of microgram quantities of metabolites of interest with enzymatic preparations, pre-concentration of the reaction mixture by solid phase extraction (SPE), metabolite isolation using HPLC systems coupled to fraction collectors prior to nuclear magnetic resonance characterization. The method reported herein is a rapid and simple technique using solely off-line mixed phase anionic exchange lipophilic SPE cartridges to selectively isolate glucuronide and sulfate metabolites from their parent compound. This approach capitalizes on the pKa differences between the parent compound, devoided of acidic moieties, and the negatively charged glucuronide and/or sulfate metabolites. Once loaded on the SPE cartridge, the incubation mixture is washed successively with a basic aqueous solution, methanol to elute the non-anionic parent compounds, and then with an acidic methanolic solution to protonate and recover the phase II conjugates. Over 100 microg (>95% purity) of 17 alpha-ethynylestradiol-3-glucuronide and 6-gingerol-4'-glucuronide were successfully isolated using this technique, as well as glucuronide and a sulfate conjugates of 1-{4'-[(1R)-2,2-difluoro-1-hydroxyethyl]biphenyl-4-yl}cyclopropanecarboxamide (DHBC) synthesized in-house. Their structures were confirmed by Ultra Performance Liquid Chromatography coupled to Quadrupole-Time of flight (UPLC-QTof) and nuclear magnetic resonance analysis.
Subject(s)
Anions/chemistry , Chromatography, Ion Exchange/instrumentation , Metabolic Detoxication, Phase II , Solid Phase Extraction/methods , Spectrometry, Mass, Electrospray Ionization/methods , Animals , Chromatography, High Pressure Liquid/instrumentation , Chromatography, High Pressure Liquid/methods , Chromatography, Ion Exchange/methods , Glucuronides/chemistry , Humans , Hydrophobic and Hydrophilic Interactions , Intestinal Mucosa/metabolism , Intestines/chemistry , Magnetic Resonance Spectroscopy/methods , Microsomes, Liver/chemistry , Microsomes, Liver/metabolism , Rats , Rats, Sprague-Dawley , Sensitivity and Specificity , Solid Phase Extraction/instrumentation , Solvents/chemistry , Spectrometry, Mass, Electrospray Ionization/instrumentation , Tandem Mass Spectrometry/methodsABSTRACT
Condensation of N-tert-butanesulfinamide (S)-1 with trifluoroacetaldehyde hydrate 2a afforded 2-methyl-N-(2,2,2-trifluoroethylidene)propane-2-sulfinamide 3. Without isolation and purification, imine 3 was added to various aryllithium reagents to give highly diastereomerically enriched adducts 5a-g. Acidic methanolysis of 5a-g provided the desired 1-aryl-2,2,2-trifluoroethylamine hydrochloride compounds 6a-g. [reaction: see text].
Subject(s)
Ethylamines/chemical synthesis , Hydrocarbons, Fluorinated/chemical synthesis , Sulfonium Compounds/chemistry , Chromatography, High Pressure Liquid , Hydrocarbons, Fluorinated/chemistry , Imines/chemical synthesis , Indicators and Reagents , Organometallic Compounds/chemistry , StereoisomerismABSTRACT
A novel HIV protease inhibitor was designed using a morpholine core as the aspartate binding group. Analysis of the crystal structure of the initial lead bound to HIV protease enabled optimization of enzyme potency and antiviral activity. This afforded a series of potent orally bioavailable inhibitors of which MK-8718 was identified as a compound with a favorable overall profile.
ABSTRACT
[reaction: see text] The use of anionic polycyclization (AP) in constructing the steroidal backbone of cardenolides was investigated. The reaction of 2-carbomethoxy-2-cyclohexenone I with the enolate of Nazarov reagent II gave, after decarboxylation and aldol condensation, steroid III with control of stereochemistry.
Subject(s)
Cardiotonic Agents/chemical synthesis , Steroids/chemical synthesis , Cardenolides/chemical synthesis , Cyclization , Ouabain/chemical synthesis , StereoisomerismABSTRACT
Highly potent, selective, and bioavailable inhibitors of human, mouse, or rat cathepsin S are described. The key structural features combine a sulfonyl moiety attached to a large group in P2 and a small substituent in P3.
Subject(s)
Cathepsins/antagonists & inhibitors , Chemistry, Pharmaceutical/methods , Animals , Cysteine Proteinase Inhibitors/chemistry , Drug Design , Drug Evaluation, Preclinical , Humans , Inhibitory Concentration 50 , Mice , Models, Chemical , Models, Molecular , Molecular Conformation , Molecular Structure , Protein Binding , Rats , Structure-Activity RelationshipABSTRACT
Based on our previous study with trifluoroethylamine as a P2-P3 amide isostere of cathepsin K inhibitor, further optimization led to identification of compound 22 (L-873724) as a potent and selective non-basic cathepsin K inhibitor. This compound showed excellent pharmacokinetics and efficacy in an ovariectomized (OVX) rhesus monkey model. The volumes of distribution close to unity were consistent with this compound not being lysosomotropic, which is a characteristic of basic cathepsin K inhibitors.