Topoisomerase II is a well-established anti-cancer drug target. Drugs such as etoposide and doxorubicin cause an increase in Top2:DNA covalent complexes in cells, resulting in the formation of protein linked DNA double strand breaks. We have taken advantage of newly developed yeast genomic tools to identify genes encoding DNA repair or DNA damage tolerance functions that play key roles in sensitivity to drugs targeting topoisomerases. We have used a set of yeast strains that are deleted for all the non-essential open reading frames of yeast to identify genes that contribute to the survival of cells exposed to Top2 targeting agents. In order to increase the sensitivity of this screen, we used vectors that overexpress yTop2, or that express a top2 mutant hypersensitive to the Top2 targeting agent mAMSA. As a first step, we examined the mAMSA sensitivity of strains carrying gene deletions that were previously shown to confer hypersensitivity to ionizing radiation or MMS. We identified over 40 genes from these repair deficient strains that were also hypersensitive to mAMSA. Loss-of-function in genes involved in DNA repair and recombination, such as HPR5 and MUS81, chromosome stability such as CTF4, transcription such as SPT10, chromatin assembly such as ASF1, as well as other genes of unknown biological function all conferred hypersensitivity to mAMSA. We identified several gene deletions, such as deletion of the RTF1 gene, which confer hypersensitivity to mAMSA, but have minimal effects on sensitivity to camptothecin, a drug targeting Top1. RTF1 encodes a protein that is part of the Paf1 complex, which is plays an important but not essential role in transcriptional elongation by RNA polymerase II. As a confirmation step, we introduced the deletions into strains with enhanced drug accumulation, and tested sensitivity etoposide, a clinically important Top2 targeting drug. All deletions that have been subjected to this confirmation step confer hypersensitivity to etoposide, even when Top2 is not overexpressed. These results indicate the utility of genomic approaches using model organisms to identifying pathways for sensitivity to chemotherapeutic agents. Our results may also provide important clues to the differences in cellular responses to drugs that target topoisomerase I and drugs that target topoisomerase II. (Supported by grants for NIH (CA 82313) and the American Lebanese Syrian Associated Charities).
[Proc Amer Assoc Cancer Res, Volume 46, 2005]