This NCDDG application proposes the design and development of novel nonpeptidic, small-molecule inhibitors of Bcl-2 and Bcl-xL proteins as a new class of anti-cancer drugs through a contemporary, multidisciplinary and integrated drug discovery approach. The first primary goal of Laboratory Program #1 is the design and synthesis of highly potent and optimized small-molecule inhibitors of Bcl-2/Bcl-xL through a structure-based design approach. The second primary goal is the characterization of their binding affinities to Bcl-2 and Bcl-xL proteins in biochemical binding assays. The third primary goal is the characterization of their activity in cancer cells with high levels of Bcl-2 and/or Bcl-xL protein and their specificity to normal cells. The fourth primary goal is the elucidation of the molecular mechanisms of action by which they induce apoptosis using isogenic model cell systems. We will perform the following specific Aims: (1). Performance of computational structure-based design for lead optimization based upon the experimental structures of small-molecule inhibitors in complex with Bcl-2 and Bcl-xL, as determined in Laboratory Program #2. (2). Chemical synthesis of new compounds designed in Aim 1. (3). Biochemical characterization for their binding affinities to Bcl-2/xL proteins and their specificity to other proteins using our established fluorescence-polarization (FP)-based binding assays. This is followed by a conclusive confirmation by the NMR Heteronuclear Single Quantum Coherence (HSQC) experiments outlined in Laboratory Program #2 (Aim 1) to rule out any potential false positives. (4). Determination of the activity, specificity and molecular mechanisms by which small molecule inhibitors of Bcl-2/Bcl-xL induce apoptosis. Laboratory Program #1 forms the foundation of this NCDDG grant and provides highly potent and optimized small-molecule inhibitors of Bcl-2/Bcl-xL with a clear understanding of their activity, specificity, and molecular mechanism of action to be studied in Laboratory Program #2 and #3.
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