Benzothiazole-based compounds as potent endothelial lipase inhibitors.

A series of benzothiazoles with a cyano group was synthesized and evaluated as endothelial lipase (EL) inhibitors for the potential treatment of cardiovascular diseases. Efforts to reduce molecular weight and polarity in the series led to improved physicochemical properties of these compounds, as well as selectivity for EL over hepatic lipase (HL). As a benchmark compound, 8i demonstrated potent EL activity, an acceptable absorption, distribution, metabolism and elimination (ADME) profile and pharmacokinetic (PK) exposure which allowed further evaluation in preclinical animal efficacy studies.

Characterization of monoacylglycerol acyltransferase 2 inhibitors by a novel probe in binding assays.

Monoacylglycerol acyltransferase 2 (MGAT2) is a membrane-bound lipid acyltransferase that catalyzes the formation of diacylglycerol using monoacylglycerol and fatty acyl CoA as substrates. MGAT2 is important for intestinal lipid absorption and is an emerging target for the treatment of metabolic diseases. In the current study, we identified and characterized four classes of novel MGAT2 inhibitors. We established both steady state and kinetic binding assay protocols using a novel radioligand, [(3)H]compound A. Diverse chemotypes of MGAT2 inhibitors were found to compete binding of [(3)H]compound A to MGAT2, indicating the broad utility of [(3)H]compound A for testing various classes of MGAT2 inhibitors. In the dynamic binding assays, the kinetic values of MGAT2 inhibitors such as Kon, Koff, and T1/2 were systematically defined. Of particular value, the residence times of inhibitors on MGAT2 enzyme were derived. We believe that the identification of novel classes of MGAT2 inhibitors and the detailed kinetic characterization provide valuable information for the identification of superior candidates for in vivo animal and clinical studies. The current work using a chemical probe to define inhibitory kinetics can be broadly applied to other membrane-bound acyltransferases.

Cell-based assay of MGAT2-driven diacylglycerol synthesis for profiling inhibitors: use of a stable isotope-labeled substrate and high-resolution LC/MS.

To demonstrate monoacylglycerol acyltransferase 2 (MGAT2)-mediated enzyme activity in a cellular context, cells of the murine secretin tumor cell-1 line of enteroendocrine origin were used to construct human MGAT2-expressing recombinant cell lines. Low throughput and utilization of radiolabeled substrate in a traditional TLC technique were circumvented by development of a high-resolution LC/MS platform. Monitoring incorporation of stable isotope-labeled D31-palmitate into diacylglycerol (DAG) allowed selective tracing of the cellular DAG synthesis activity. This assay format dramatically reduced background interference and increased the sensitivity and the signal window compared with the TLC method. Using this assay, several MGAT2 inhibitors from different chemotypes were characterized. The described cell-based assay adds a new methodology for the development and evaluation of MGAT2 inhibitors for the treatment of obesity and type 2 diabetes.