Abstract 2622: C282Y HFE polymorphism imparts temozolomide and 17-DMAG resistance via p16INK4A

Abstract

Abstract Cancer cells have a robust requirement for iron. The HFE protein interacts with transferrin receptor and limits the cellular iron uptake. HFE polymorphisms - the most common genetic variants in Caucasians - have been associated with a variety of cancers. Therefore we hypothesized that HFE polymorphisms would impact the phenotype of cancer cells. Using stably transfected human neuroblastoma cell lines and a number of human glioma cell lines we determined the effect of HFE polymorphisms on the therapy resistance. In the HFE variants stably transfected neuroblastoma cells, cells carrying the C282Y allelic variant were significantly more resistant to the chemotherapeutic agent temozolomide (currently the standard of care for brain tumors) than the same cell line carrying wildtype or H63D variant of HFE. The resistance to temozolomide was also found in commercially available human astrocytoma cell lines with the C282Y allele. The mechanism of greater resistance in the cells with the C282Y allele did not appear to involve O6-methylguanine-DNA methyltransferase traditionally associated with temozolomide resistance. In addition to temozolomide, the C282Y allele expressing glioma cells were also resistant to geldanamycin and its derivatives, 17-allylaminogeldanamycin (17-AAG) and 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG). These ansamycin compounds inhibit heat shock protein 90 (Hsp90) and thus have a different mechanism of action on the cancer cells than temozolomide. To discover the mechanism by which C282Y HFE variant cells were resistant to different therapeutic agents, we performed functional gene arrays in HFE stably transfected neuroblastoma cells. The expression of p16INK4A (cyclin dependent kinase inhibitor 2A) gene was found significantly increased in the C282Y cells compared to the wildtype cells. Consistent with the gene expression, p16INK4A protein expression was elevated in association with the C282Y allele relative to wildtype HFE. C282Y expressing astrocytoma cells also expressed high levels of p16INK4A protein. Therefore, we further determined the role of p16INK4A in drug resistance using p16INK4A siRNA, and found that p16INK4A knockdown sensitized C282Y expressing cells to temozolomide and 17-DMAG. Furthermore, siRNA of HFE decreased p16INK4A expression in C282Y cells. Taking together, these data suggest that C282Y HFE polymorphism impacts the cancer phenotype such as drug resistance, and the mechanism appears to be via increased expression of p16INK4A. Given the frequency of the HFE variants in the general population, these data provide compelling evidence that C282Y HFE polymorphism should be a pharmacogenetic consideration for evaluating treatment efficacy in cancer cells. 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 2622.