Funding will be used to screen existing compound hbraries for newer chemical classes for selective LDHA inhibitors. In Task Element One, NCATS will adopt for HTS an existing assay to monitor the activity of LDHA by measuring the change in fluorescence of the NADH substrate as a function of reaction time. Under this Task, we will also validate the? assay by running a pilot screen using both oftline/standalone and online/robotic equipment. For Technical Task Element Two, NCATS will perform a high-throughput screen of the NCGC's compound collection using a fully-integrated Kalypsys robotic system. Compounds will be screened in concentration-response fonnat. Hits from this screen wi11 first be selected after confirmatory assays and further examined in a series of assays for selectivity. Under Technical Task Element Three, NCATS will develop counterscrecn assays including pyruvate kinase and unrelated dehydrogenase GAPDH. Working in parallel, Technical Task Element Four will develop and propose desired characteristics of compounds that may act as LOH-A inhibitors, perform in silico screening of a virtual library for desired pharmacophores, and identify new lead scaffolds. Under Technical Task Element Five, NCATS and the Southern Research Institute will establish structure activity relationships (SARs), ensuring that emerging groups oflead compounds exert biological activities through the intended mechanism of action, and evaluate the lead compounds for physicochemical properties.
