_ , ...... =5 -;??, . Core A will focus on biological evaluation of extracts, chromatograpnic'fractions,'and pure isolates prepared by all of the projects within the program in order to help prioritize subsequent biological and chemical work. This strategy of using bioassay-guided isolation will afford us the opportunity to pursue biological activity within complex mixtures. Extracts found active by Core A will be further purified by Projects 1-3 (OSU, UIC, and RTI, respectively) and the pure isolates returned to us for further analysis. Promising pure compounds will ultimately be further tested in animal models to assess efficacy and toxicity. Mechanism of action studies will be conduced to study the molecular pharmacology of pure compounds with the most favorable therapeutic indices. The Leader of Core A has well-established collaborations with all of the Project Leaders of this Program Project and has previously employed a variety of assays for natural product drug discovery. Our ongoing working relationships with other Projects and Cores and our ability to develop new biological assays as needed will assure that Core A is fully integrated in the program project and serves all projects therein.
The Specific Aims for Core A are to (1) conduct molecular-target based assays including the histone deacetylase assay (HDAC) and inhibition of the 20S proteasome. These assays will be conducted initially on extract of materials Our second specific aim is to follow active leads identified by initial analysis using cell based assays such as HL60, which is known to overexpress the 20S proteasome, or HeLa cells that are the source of the HDAC used in the molecular target assays. Our third specific aim is to assess the activity of our most promising active pure compounds in vivo. These experiments will include initial dose ranging studies followed by efficacy studies using the hollow fiber assay or tumor xenografts studies in immunodeficient mice. Finally, studies will be performed on those pure compounds that prove to be effective anticancer agents with limited toxicity. in mice. These studies will focus on determining the mechanism of action and may include studies such as pathway of apoptosis induction or mechanism of cell cycle arrest. The biological assays outlined in this Core, used in concert with Cores B through D, will provide a solid foundation of support for all the Projects within the program.
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