Abstract
Cisplatin, a commonly used chemotherapeutic, is associated with ototoxicity, renal toxicity and neurotoxicity, thus identifying means to increase the therapeutic index of cisplatin may allow for improved outcomes. A SNP (rs4343077) within EPS8, discovered through a genome wide association study of cisplatin-induced cytotoxicity and apoptosis in lymphoblastoid cell lines (LCLs), provided impetus to further study this gene. The purpose of this work was to evaluate the role of EPS8 in cellular susceptibility to cisplatin in cancerous and non-cancerous cells. We used EPS8 RNA interference to determine the effect of decreased EPS8 expression on LCL and A549 lung cancer cell sensitivity to cisplatin. EPS8 knockdown in LCLs resulted in a 7.9% increase in cisplatin-induced survival (P = 1.98×10−7) and an 8.7% decrease in apoptosis (P = 0.004) compared to control. In contrast, reduced EPS8 expression in lung cancer cells resulted in a 20.6% decrease in cisplatin-induced survival (P = 5.08×10−5). We then investigated an EPS8 inhibitor, mithramycin A, as a potential agent to increase the therapeutic index of cisplatin. Mithramycin A decreased EPS8 expression in LCLs resulting in decreased cellular sensitivity to cisplatin as evidenced by lower caspase 3/7 activation following cisplatin treatment (42.7%±6.8% relative to control P = 0.0002). In 5 non-small-cell lung carcinoma (NSCLC) cell lines, mithramycin A also resulted in decreased EPS8 expression. Adding mithramycin to 4 NSCLC cell lines and a bladder cancer cell line, resulted in increased sensitivity to cisplatin that was significantly more pronounced in tumor cell lines than in LCL lines (p<0.0001). An EGFR mutant NSCLC cell line (H1975) showed no significant change in sensitivity to cisplatin with the addition of mithramycin treatment. Therefore, an inhibitor of EPS8, such as mithramycin A, could improve cisplatin treatment by increasing sensitivity of tumor relative to normal cells.
Highlights
Cisplatin is a platinum agent used for the treatment of head and neck, ovarian, cervical, testicular, and lung cancers; severe toxicities and intrinsic/acquired resistance interfere with its efficacy [1]
These results reveal that lower levels of epidermal growth factor receptor pathway substrate 8 (EPS8) in lymphoblastoid cell lines (LCLs) decrease cellular sensitivity to cisplatin as evidenced by increased cell survival and reduced apoptotic activity when treated with cisplatin
EPS8 was chosen based on previous preclinical GWAS results using multiple cellular phenotypes: cisplatin-induced cytotoxicity and cisplatin-induced apoptosis as measured by caspase 3/7 activation
Summary
Cisplatin is a platinum agent used for the treatment of head and neck, ovarian, cervical, testicular, and lung cancers; severe toxicities and intrinsic/acquired resistance interfere with its efficacy [1]. One of the common SNPs, rs4343077, in which the minor allele had lower cisplatin induced apoptosis (P = 0.0007) and higher survivial (P = 0.0007) is an expression quantitative trait locus (eQTL) associated with the baseline gene expression levels of 28 genes at P#1024 [12]. This SNP is in an intron of epidermal growth factor receptor pathway substrate 8 (EPS8)
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