Modeling the Responsiveness of Sensitive Populations to Genotoxic Agents Using DNA Repair Inhibitors

Project Summary It is well recognized that conventional toxicology assays exhibit two crucial deficiencies. First, the use of limited numbers of cell lines and laboratory animal models/strains does not provide information about the degree to which a particular chemical may exhibit highly variable potencies across exposed humans. Although studying very large numbers of cell lines and laboratory rodent models could theoretically address this issue, it is not a practical answer. There are simply too many legacy chemicals and new chemical substances being developed for this to represent a routine testing scheme. A second issue with the current schema is that it does not typically generate much information regarding the molecular mechanism(s) responsible for a toxic effect. Fortunately, in the area of genetic toxicology, a solution exists for both of these issues. Our innovative approach takes into account the fact that inter-subject differences in sensitivity to genotoxic agents is often driven by variable DNA repair capacity. Our solution is based on a single p53-competent human cell line (TK6). Whereas test chemical-exposed TK6 cells will be used to generate reference responses, we will model sensitive subpopulation responses by co-exposing cells to each test chemical in combination with a panel of small molecule DNA repair inhibitors. These inhibitors will be chosen to cover each of the major DNA repair pathways. This platform will simultaneously allow us to identify for further testing those chemicals that are most likely to exhibit the greatest variation in genotoxic potency across exposed humans, and also provide insights into genotoxic mechanism. Innovative aspects of our project include our use of an automated, multiplexed DNA damage assay to cover multiple biomarkers of genotoxicity and cytotoxicity, and state-of-the-art benchmark dose software to characterize the family of dose-response relationships that will be generated for each chemical studied. Once reduced to practice, we will make the assay available through fee-for-service chemical testing, with an eventual goal of selling reagent-containing kits.