DNA Methylation-Independent Reversion of Gemcitabine Resistance by Hydralazine in Cervical Cancer Cells

Abstract

Background Down regulation of genes coding for nucleoside transporters and drug metabolism responsible for uptake and metabolic activation of the nucleoside gemcitabine is related with acquired tumor resistance against this agent. Hydralazine has been shown to reverse doxorubicin resistance in a model of breast cancer. Here we wanted to investigate whether epigenetic mechanisms are responsible for acquiring resistance to gemcitabine and if hydralazine could restore gemcitabine sensitivity in cervical cancer cells. Methodology/Principal Findings The cervical cancer cell line CaLo cell line was cultured in the presence of increasing concentrations of gemcitabine. Down-regulation of hENT1 & dCK genes was observed in the resistant cells (CaLoGR) which was not associated with promoter methylation. Treatment with hydralazine reversed gemcitabine resistance and led to hENT1 and dCK gene reactivation in a DNA promoter methylation-independent manner. No changes in HDAC total activity nor in H3 and H4 acetylation at these promoters were observed. ChIP analysis showed H3K9m2 at hENT1 and dCK gene promoters which correlated with hyper-expression of G9A histone methyltransferase at RNA and protein level in the resistant cells. Hydralazine inhibited G9A methyltransferase activity in vitro and depletion of the G9A gene by iRNA restored gemcitabine sensitivity. Conclusions/Significance Our results demonstrate that acquired gemcitabine resistance is associated with DNA promoter methylation-independent hENT1 and dCK gene down-regulation and hyper-expression of G9A methyltransferase. Hydralazine reverts gemcitabine resistance in cervical cancer cells via inhibition of G9A histone methyltransferase.

Association between hypermethylation of DNA repetitive elements in white blood cell DNA and early-onset colorectal cancer

“…Chemotherapy is an important factor to consider when investigating DNA methylation changes, as some studies suggest that chemotherapy can result in increased DNA methylation. [30][31][32] Apart from the Alu repetitive element, our results show no significant difference in the methylation levels of cases who underwent treatment with 5FU and those who did not, suggesting that 5FU in the MUTYH gene as previously described. 49 Those cases carrying a pathogenic mutation in a mismatch repair gene (Lynch Syndrome) or biallelic or compound heterozygous mutations in MUTYH were identified for the stratified analysis.…”

Expression Levels of Human Equilibrative Nucleoside Transporter 1 and Deoxycytidine Kinase Enzyme as Prognostic Factors in Patients with Acute Myeloid Leukemia Treated with Cytarabine

“…Other metabolic steps are also implicated in chemotherapy-resistant leukemic cells including kinase activities, such as inactivation of the dCK enzyme [21,22]. This modification was also documented in clinical conditions, including pancreatic, esophageal and biliary tract cancer [23,24].…”

PharmGKB summary

“…More recently, due to its epigenetic effects, one group has investigated its use in combination with valproic acid in clinical trials with the hypothesis of reducing tumor resistance and increasing anti-cancer chemotherapy efficacy [121–123]. Its beneficial epigenetic effects in cancer cells are thought to be as an inhibitor of DNA methyltransferase (DNMT) enzymes in order to reactivate tumor suppressor genes silenced by DNA methylation [124], and may also inhibit histone methyltransferase activity [123] and histone acetyltransferases [65]. Hydralazine is thought to be metabolized by two pathways, both of which involve acetylation [125].…”