Cell-Based Assays for Identification of Novel Double-Strand Break-Inducing Agents

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Abstract
Background:We are developing cell-based assays to identify anticancer agents that are selectively toxic to cells with defined mutations. As a test, we used a three-stage strategy to screen compounds from the National Cancer Institute's repository for agents that are selectively toxic to double-strand break repair-deficient yeast cells. Methods:Compounds identified in the screen were further analyzed by use of yeast and vertebrate cell-based and in vitroassays to distinguish between topoisomerase I and II poisons. Results:Of the more than 85 000 compounds screened, 126 were selectively toxic to yeast deficient in DNA double-strand break repair. Eighty-seven of these 126 compounds were structurally related to known topoisomerase poisons, and 39 were not. Twenty-eight of the 39 were characterized, and we present data for eight of the compounds. Among these eight compounds, we identified two novel topoisomerase II poisons (NSC 327929 and NSC 638432) that were equipotent to etoposide in biochemical tests and in cells, five (NSC 63599, NSC 65601, NSC 380271, NSC 651646, and NSC 668370) with topoisomerase I-dependent toxicity in yeast that induced DNA damage and toxicity in mammalian cells, and one (NSC 610898) that directly bound to DNA and induced strand breaks. Conclusions:Cell-based assays can be used to identify molecules that are selectively toxic to cells with a predetermined genetic background, including mutations in genes involved in the cell cycle and its checkpoints, for which there are currently no selectively toxic compounds.