Abstract 2914: Creation of a novel biochemical and biophysical assay suite to enable the identification of inhibitors targeting inactive kinases

Abstract

Abstract We have recently reported the implementation of a structural biology-based drug design platform for the identification of protein kinase inhibitors which utilize hydrophobic clusters to stabilize the inactive state of a kinase. These clusters are not formed in active kinases and result in the conversion of the ATP binding cleft into a non-polar environment which is sterically and electrostatically incompatible with ATP binding. We have industrialized this approach, designated the ArQule Kinase Inhibitor Platform (AKIPTM), and synthesized a library of more than 15,000 small molecules through the application of an in silico guided process. This has allowed us to rapidly generate leads to a variety of kinases, including receptor tyrosine kinases, non-receptor tyrosine kinases, and serine-threonine kinases. By virtue of their unique binding modes, many of these inhibitors would not be identified in standard assays using highly activated kinases. We have therefore deliberately re-engineered standard biochemical kinase assays using unphosphorylated inactive kinases to aid in the optimization of these inhibitors. In addition to these biochemical assays, we have also implemented a variety of well-established technologies not commonly used early in the hit generation process, including a thermal shift assay (TSA), affinity mass spectrometry, endogenous tryptophan fluorescence detection, an ATP-exclusion assay using a non-hydrolyzable ATP analogue, classical kinetic analysis to assess ATP-dependence and mechanism-of-inhibition, X-ray crystallography, and finally, cross-competition experiments with known inhibitors. Using the cumulative knowledge gained from these technologies throughout the hit generation, hit-to-lead, and lead optimization stages has enabled us to make informed decisions and resulted in identification of many potent ATP-independent inhibitors. The AKIP technology to date has produced at least one clinical candidate, ARQ 092, which potently inhibits AKT with a high degree of selectivity amongst the human kinome. Further examples of the application of these various technologies will be provided for a diverse range of kinases, including c-Met, FGFR, Ack, and TNIK. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2914. doi:1538-7445.AM2012-2914