Cisplatin is a platinum-based chemotherapy drug used in the treatment of testicular, ovarian, cervical, lung, and breast cancer. Despite its clinical utility, development of resistance and toxic side effects remain big challenges associated with cisplatin therapy. Potential mechanisms of resistance include changes in drug import and efflux, cisplatin metabolism/activation, and repair rates of cisplatin-induced DNA damage. All three resistance mechanisms influence the abundance of DNA cross-links, including 1,2- and 1,3-intrastrand and interstrand cross-links. Therefore, we hypothesize that a patient’s response to cisplatin can be accurately predicted by measuring the formation and repair of these cisplatin DNA cross-links in tumors. To address this goal, we have developed an isotope dilution UPLC-MS assay to quantify both cisplatin-induced intra- and interstrand crosslinks. The DNA digestion and monomer enrichment methodology was first optimized using synthetic DNA oligos and plasmids containing specific cross-links, followed by validating the assay in cisplatin-treated cell lines. We are currently investigating the formation and repair kinetics of each lesion in cell deficient in nucleotide excision repair (addressing intrastrand crosslinks) and ICL-repair (addressing interstrand crosslinks) to investigate synergism between repair pathways in counteracting cisplatin therapy. Successful application of this assay in cells is expected to be useful in clinical trials and practice to predict the outcome of cisplatin therapy.

Development of an ultra performance liquid chromatography-mass spectrometry (UPLC-MS) assay for the quantification of cisplatin-Induced DNA intra- and interstrand cross-links

2019 Fall National Meeting

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