Abstract 5562: Screening of in vitro cytotoxicity of nucleoside analogs in HapMap cell lines to identify predictive markers of drug sensitivity
Authors: Ghosh Taraswi Mitra , Feldberg Tanya J. , Bhise Neha , Lamba Jatinder K. 
Published: May 2015

Nucleoside analogues (NAs) are synthetic molecules that resemble naturally occurring nucleosides. These analogs are themselves inactive and require activation by multiple phosphorylation steps to their triphosphate forms which depending on the particular agent either inhibits DNA/RNA (cytarabine, clofarabine), or inhibits, ribonucleotide reductases (clofarabine, gemcitabine) or DNA methyl transferases (decitabine). Resistance to nucleoside analogue-based chemotherapy is a major problem in the treatment of cancers. In the current study, we determined the in vitro cytotoxicity of the nucleoside analogs clofarabine, fludarabine and decitabine in 60 unrelated Epstein Barr virus (EBV)-transformed B-lymphoblastoid HapMap cell lines with European ancestry [Centre d'Etude du Polymorphisme Humain (CEU) samples] were obtained from the Coriell Institute for Medical Research. The cell lines were maintained at 37°C in RPMI1640 medium supplemented with 2mM Glutamine. In vitro cytotoxicity was determined by plating LCLs in a 96 well plate at density of 250,000cells/ml. Cell survival was determined using MTT ((3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay 48hr after exposure to varying concentrations of each nucleoside analog. Gen 5 software was used to calculate IC50 values for each cell line and agent. Our results showed wide inter-individual variation in cytotoxicity to nucleoside analogs among HapMap cell lines. Significant correlation was observed between the IC50 values of fludarabine vs. clofarabine (r = 0.732; P = 0.00001). However no significant correlation was observed with the IC50 values of decitabine. The reason for the observed relationship might be the fact that clofarabine is a 2nd generation purine nucleoside and structurally related to fludarabine. Additionally, the mechanism of action of decitabine is different than clofarabine and fludarabine with it being a hypomethylating agent. Since genome-wide genotype and gene expression data on these cell lines is available publicly, currently we are analyzing the data for identification of genomics markers predictive of cell survival and response to these nucleoside analogs. Our preliminary analysis showed that polymorphisms in candidate genes involved in drug metabolism pathway may be involved in cancer drug resistance. For example, a promoter SNP in the gene DCK was associated with both clofarabine IC50 and fludarabine IC50 while another promoter SNP in NT5C2 was associated with fludarabine IC50.

These pharmacogenomic factors may then serve as predictive markers for improvement of cancer chemotherapy.

Citation Format: Taraswi Mitra Ghosh, Tanya J. Feldberg, Neha Bhise, Jatinder K. Lamba. Screening of in vitro cytotoxicity of nucleoside analogs in HapMap cell lines to identify predictive markers of drug sensitivity. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5562. doi:10.1158/1538-7445.AM2014-5562