Abstract 570: Role of NAD(P)H:quinone oxidoreductase 1 (NQO1) in the cytotoxicity of the Hsp90 inhibitor 17AAG in human pancreatic cancer cells

Abstract

Abstract Previous work has demonstrated that NQO1 can reduce the benzoquinone ansamycin Hsp90 inhibitor 17-allyamino-demethoxy geldanamycin (17AAG) to the corresponding hydroquinone (17AAGH2). The formation of 17AAGH2 by NQO1 results in a more water soluble molecule and that binds with greater affinity to Hsp90 when compared to the parent quinone 17AAG. Experiments were conducted with the human pancreatic cancer cell line Panc1 which is homozygous for the NQO1*2 polymorphism and contains only trace levels of NQO1 protein and no NQO1 catalytic activity. An isogenic cell line expressing high levels of NQO1 (Panc1-C5) was generated by stable transfection of wild type NQO1 in Panc1 cells. 17AAG was more cytotoxic to Panc1-C5 cells with high levels of NQO1 when compared to NQO1-null parental Panc1 cells. Pretreatment of Panc1-C5 cells with the NQO1 mechanism-based inhibitor ES936 reduced the cytotoxicity of 17AAG to the level observed in Panc1 cells. No difference in the cytotoxicity of the non-quinone Hsp90 inhibitor radicicol was observed between the two cell lines Biomarkers of Hsp90 inhibition were measured in both Panc1 and Panc1-C5 cells following treatment with 17AAG. Higher protein levels of Hsp70 and lower amounts of cRaf, phosphorylated ERK1/2 and phosphylated MEK were observed in Panc1-C5 cells compared to Panc1 cells. In addition, cell cycle analysis revealed a greater number of Panc1-C5 cells arrested in G2M following 17AAG treatment compared to Panc1 cells. Cytotoxicity studies were also performed in additional human pancreatic cancer cell lines (BxPC3 and MiaPaCa2) that contain high levels of catalytically active NQO1*1 protein and in agreement with the studies described above the cytotoxicity of 17AAG could be reduced significantly following pretreatment with the NQO1 inhibitor ES936. The intracellular concentrations of 17AAG and 17AAH2 were measured in human pancreatic cancer cell lines and it was observed that greater amounts of both 17AAG and 17AAGH2 could be detected in cell lines with catalytically active NQO1 when compared to cells lacking NQO1 activity or cells pretreated with ES936. These data demonstrate that in addition to generating a more potent Hsp90 inhibitor (17AAGH2) the reduction of 17AAG to 17AAGH2 by NQO1 leads to significantly greater intracellular concentrations of 17AAG and 17AAGH2. These data confirm the role of NQO1 in sensitivity to 17AAG and suggest that genotyping for the NQO1*2 polymorphism should be considered in patients undergoing 17AAG treatment for pancreatic cancer or other solid tumors that are known to contain high levels of NQO1 activity (Supported by CA51210). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 570.