ABSTRACT
We describe here our attempts to optimise the human fatty acid amide hydrolase (FAAH) inhibition and physicochemical properties of our previously reported tetrasubstituted azetidine urea FAAH inhibitor, VER-156084. We describe the SAR of a series of analogues and conclude with the demonstration of in vivo dose-dependant FAAH inhibition in an anandamide-loading study in rats.
Subject(s)
Amidohydrolases/antagonists & inhibitors , Azetidines/pharmacology , Enzyme Inhibitors/pharmacology , Urea/pharmacology , Amidohydrolases/metabolism , Animals , Azetidines/chemical synthesis , Azetidines/chemistry , Cytochrome P-450 Enzyme Inhibitors , Cytochrome P-450 Enzyme System/metabolism , Dose-Response Relationship, Drug , ERG1 Potassium Channel , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/chemistry , Ether-A-Go-Go Potassium Channels/antagonists & inhibitors , Humans , Models, Molecular , Molecular Structure , Rats , Stereoisomerism , Structure-Activity Relationship , Urea/chemical synthesis , Urea/chemistryABSTRACT
We herein report the discovery of a novel class of antagonists of the human adenosine A2B receptor. This low molecular weight scaffold has been optimized to offer derivatives with potential utility for the alleviation of conditions associated with this receptor subtype, such as nociception, diabetes, asthma and COPD. Furthermore, preliminary pharmacokinetic analysis has revealed compounds with profiles suitable for either inhaled or systemic routes of administration.
Subject(s)
Adenosine A2 Receptor Antagonists , Pyrimidines/chemistry , Administration, Inhalation , Animals , Asthma/drug therapy , Drug Design , Humans , Pulmonary Disease, Chronic Obstructive/drug therapy , Pyrimidines/chemical synthesis , Pyrimidines/pharmacokinetics , Rats , Receptor, Adenosine A2A/metabolism , Receptor, Adenosine A2B/metabolismABSTRACT
We report the discovery of a novel, chiral azetidine urea inhibitor of Fatty Acid Amide Hydrolase (FAAH,) and describe the surprising species selectivity of VER-156084 versus rat and human FAAH and also hCB1.
Subject(s)
Amidohydrolases/antagonists & inhibitors , Amidohydrolases/chemistry , Azetidines/chemistry , Azetidines/chemical synthesis , Enzyme Inhibitors/chemical synthesis , Piperidines/chemical synthesis , Urea/chemistry , Animals , Azetidines/pharmacology , Catalysis , Chemistry, Pharmaceutical/methods , Drug Design , Enzyme Inhibitors/pharmacology , Humans , Mice , Mice, Knockout , Models, Chemical , Piperidines/pharmacology , Rats , Receptor, Cannabinoid, CB1/chemistry , StereoisomerismABSTRACT
Alkyl quinolone molecules 2-heptyl-4-quinolone (HHQ) and 2-heptyl-3-hydroxy-4(1H)-quinolone (PQS) are important quorum sensing signals, which play a mediatory role in the pathogenesis of acute and chronic Pseudomonas aeruginosa infection. A targeted approach inhibiting the bacterial 'multiple virulence factor regulon' (MvfR) protein complex, offers the possibility to block the synthesis of MvfR-dependant signal molecules. Here, a high throughput bioanalytical method was developed using LC-MS/MS detection for the selective determination of HHQ and PQS in mouse tissue homogenate, over a sensitive range of 1-5000 and 10-5000pg/mL, respectively. Chromatographic peak distortion of the iron chelator PQS was overcome with the applied use of a bidentate chelator mobile phase additive 2-Picolinic acid at 0.2mM concentration, giving an improved separation and response for the analyte, whilst maintaining overall MS system robustness. Following thigh infection with P. aeruginosa strain 2-PA14 in mice, the concentration and time course of HHQ and PQS (4-hydroxy-2-alkyl-quinolone (HAQ) biomarkers) residing in the biophase were evaluated, and exhibited a low level combined with a substantial inter-individual variability. Quantifiable levels could be obtained from approximately 15h post infection, to the study termination at 21-22h. A dose dependant reduction in HAQ tissue concentrations at selected time points were obtained following MvfR inhibitor administration versus drug vehicle (p<0.01, Kruskal-Wallis-one way ANOVA) and meta -analyses of several studies enabled an inhibitory concentration (IC50) of 80nM free drug to be determined. However, due to the experimental limitations a defined time profile for in-vivo HAQ production could not be characterised. Microsomal stability measurements demonstrated a rapid metabolic clearance of both alkyl quinolone biomarkers in the bacterial host, with a hepatic extraction ratio greater than 0.96 (the measurable assay limit). High clearance underpinned the low concentrations present in the well-perfused thigh tissue. Along with method development and validation details, this paper considers the kinetics of in-vivo HAQ bio-synthesis during Pseudomonas infection; and risks of biomarker over-estimation from samples which contain an exogenous population of bacteria.