LC-MS/MS Assessments of Apoptosis/DNA Damage/Repair

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DESCRIPTION (provided by applicant): It is proposed that Liquid Chromatography-tandem Mass Spectrometry (LC-MS/MS) be utilized to characterize the response of tumor cells to chemotherapy. Modern LC-MS/MS technology, employing an electrospray interface and triple quadrapole detection, provides unprecedented sensitivity, selectivity and the capability for measuring multiple products. It is proposed that LCMS/ MS technology be adapted to the measurement of DNA modifications induced by anticancer drugs including direct and/or indirect damage, various types of strand breaks and the products of apoptosis. Using a method of sample preparation that is highly effective in eliminating background it is feasible to measure simultaneously this variety of DNA lesions and modifications in a single sample. The feasibility of this approach is demonstrated in the application using LC-MS/MS measurements on cells exposed to cisplatin as an example. It is proposed that LC-MS/MS assay can be applied to cells obtained at surgery suing the culture methods presently employed to measure drug resisitivity. Differences in cell function revealed by LC-MS/MS assay will be complemented by array studies of gene expression. The first clinical application will be ovarian cancer tissue.