Abstract 2214: Genome-wide association analysis identified genetic markers associated with triptolide cellular sensitivity using HapMap LCLs as model system.

Abstract

Abstract Triptolide is a biological diterpenoid derived from the Chinese herb Tripterygium wilfordii Hook f. It has shown to have immunosuppression and anti-inflammation, and antineoplastic effect [Pan et al, Cancer Lett, 20102;3,5]. The anti-tumor effects of triptolide have been shown in wide range of solid tumor and leukemic cell lines in vitro. Present study focused on identifying the genetic markers using EBV transformed (lymphoblast cell lines as model systems). International HapMap Project LCLs derived from subjects with European ancestry (CEU; n=60 unrelated) were obtained from the Coriell Institute for Medical Research. Genotype data and gene expression data on these cell lines is available publically. The cell lines were maintained at 370C in RPMI1640 medium supplemented with 2mM Glutamine. In vitro cytotoxicity was determined by plating LCLs in a 96 well plate at the density of 250,000 cells/mL followed by treatment with six different concentration of triptolide (500, 50, 20, 10, 6.67, and 1.3 nM) 24hr after plating. Cell viability was determined 48 hr post drug treatment by MTT assays. Genome-wide gene expression and genotype data was obtained from publically available source and was evaluated for identification of SNPs and/or gene expression signatures predictive of triptolide cellular sensitivity. Interestingly we observed “clumping” of small p-values in a region on chromosome 2 consisting of SNPs with smallest p-value. SNPs within this region on chromosome 2 spanned across many biologically and functionally important genes such as: genes of importance in apoptosis caspase 8, caspase 10 and CFLR (caspase 8 and FADD like apoptosis regulator). The most significant SNP rs6759004 occurred in the 3’UTR of CFLAR gene and was associated with triptolide cytotoxicity (p=2.5E-07). SNPs in genes involved in DNA replication and repair as ORC2 (origin of replication complex subunit2); in formation of reactive oxygen species as AOX1 and AOX2P (aldehyde oxidase); proteins involved in transcription, RNA processing and splicing as protein kinase CLK1 (CDC-kine linase-1), NIF3L1 etc were also significantly associated with triptolide chemosensitivity. The two most interesting candidate markers identified by association of genome-wide expression with triptolide cytotoxicity includes JAK1, (Janus kinase 1) which is involved in JAK/STAT signaling and plays significant role in cell differentiation, growth, transformation, apoptosis, proliferation etc. and DTX1, which plays role in NOTCH1 signaling. Overall our results identified biomarkers of biological and clinical significance that predict response to triptolide and would be of clinical relevance to patients being treated by triptolide Citation Format: Jatinder K. lamba, Tanya Feldberg, Taraswi Mitra Ghosh, Neha Bhise, Brooke Fridley. Genome-wide association analysis identified genetic markers associated with triptolide cellular sensitivity using HapMap LCLs as model system. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2214. doi:10.1158/1538-7445.AM2013-2214