Gemcitabine and Cytosine Arabinoside Cytotoxicity: Association with Lymphoblastoid Cell Expression

Abstract

Two cytidine analogues, gemcitabine (dFdC) and 1-B-Darabinofuranosylcytosine (AraC), show significant therapeutic effect in a variety of cancers. However, response to these drugs varies widely. Evidence from tumor biopsy samples shows that expression levels for genes involved in the cytidine transport, metabolism, and bioactivation pathway contribute to this variation in response. In the present study, we set out to test the hypothesis that variation in gene expression both within and outside of this ''pathway'' might influence sensitivity to gemcitabine and AraC. Specifically, Affymetrix U133 Plus 2.0 GeneChip and cytotoxicity assays were performed to obtain basal mRNA expression and IC 50 values for both drugs in 197 ethnically defined Human Variation Panel lymphoblastoid cell lines. Genes with a high degree of association with IC 50 values were involved mainly in cell death, cancer, cell cycle, and nucleic acid metabolism pathways. We validated selected significant genes by performing real-time quantitative reverse transcription-PCR and selected two representative candidates, NT5C3 (within the pathway) and FKBP5 (outside of the pathway), for functional validation. Those studies showed that down-regulation of NT5C3 and FKBP5 altered tumor cell sensitivity to both drugs. Our results suggest that cell-based model system studies, when combined with complementary functional characterization, may help to identify biomarkers for response to chemotherapy with these cytidine analogues.

DNA methylation contributes to natural human variation

“…Many genome-wide association studies (GWAS) have attempted to establish genetic associations with differences between distinct populations (Li et al 2008a;Lachance et al 2012), diseases (Kamatani et al 2009), and the response to external stimuli (Li et al 2008b;Niu et al 2010). However, fewer associations were observed than expected, and direct genotype-phenotype relations were not easily explicable since the majority of variant sites are located in noncoding loci (Kilpinen and Dermitzakis 2012).…”

“…We determined differences in DNA methylation between three populations extensively characterized in the Human Variation Panel in terms of single nucleotide polymorphisms (SNPs) and gene expression (Li et al 2008b;Niu et al 2010). The DNA methylation profile was assessed for 288 B-cell lymphoblastoid cell lines (LCL) representing 96 Caucasian-American (CA), 96 AfricanAmerican (AF), and 96 Han Chinese-American (AS) individuals.…”

Identification of genes associated to 2′,2′-difluorodeoxycytidine resistance in HeLa cells with a lentiviral short-hairpin RNA library

“…Li and co-workers have performed a genome-wide association study with multiple cell lines to analyse single nucleotide polymorphisms and mRNA expression levels to identify genes that affected dFdC sensitivity [26,27]. Using these techniques they have identified genes and genetic variations associated to nucleoside analog resistance.…”

Leveraging Lymphoblastoid Cell Lines for Drug Response Modeling

“…Several studies have used LCL models to functionally validate findings using knock-down experiments [15][16][17] and the advantages offered by these techniques are described in greater detail below.…”