Abstract
BackgroundDown 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 FindingsThe 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/SignificanceOur 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.
Highlights
Gemcitabine (29,29-difluoro 29deoxycytidine, dFdC) is an analog of cytosine arabinoside that possesses distinctive pharmacological properties and wide antitumor-spectrum activity
Cervical cancer cell lines were examined for the basal expression of hENT1, dCK, RRM1, RRM2, Cytidine deaminase (CDA) genes and their sensitivity to gemcitabine was evaluated
To investigate whether the induction of gemcitabine resistance is related to changes in the expression of hENT1, dCK, RRM1, RRM2, CDA genes, CaLo cells were exposed to increasing concentrations of gemcitabine and once cells became resistant they were treated with hydralazine at different times and concentrations
Summary
Gemcitabine (29,29-difluoro 29deoxycytidine, dFdC) is an analog of cytosine arabinoside that possesses distinctive pharmacological properties and wide antitumor-spectrum activity. Sensitivity to nucleoside analogs including gemcitabine in vitro and in the clinical setting has been shown to correlate with expression of this transporter whereas hENT1-deficient cells are highly resistant to this nucleoside [5,6,7,8]. Cytidine deaminase (CDA) catalyses the deamination of cytidine, dexoycytidine, and their analogs such as gemcitabine its role mediating gemcitabine resistance is controversial [18] These data clearly suggest that a diminished or lack of expression of dCK and hENT1 are crucial for gemcitabine resistance, the mechanisms leading to their transcriptional silencing are yet to be defined. 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. 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
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
