Epigenetic Regulation of hTERT Expression in Breast Cancer Cells: Analysis of Methylation Status, Expression Levels, and Apoptosis with HDACi and Calpeptin Combination Therapy

• Further investigation, such as by CHIP analysis to look at histone acetylation and methylation, is required to better understand the way by which HDACi alter telomerase expression. Figure 4A: Methylation Levels at CpG islands in MDA-231 (blue) and MCF-7 (green) Graphs on the right are from -634bp to -524bp and on the left are 1233bp to 1386bp. The hTERT gene encodes the catalytic subunit of human telomerase reverse transcriptase and is required for telomerase activity. Telomerase is turned off in differentiated somatic cells but is re-expressed in the vast majority of cancer cells, allowing for sustained cellular division and growth. Because there are two CpG islands upstream of hTERT, suggesting a role for methylation in the control of this gene’s transcription, we looked at the correlation between expression levels and methylation status of the hTERT gene in breast cancer cells. Further, we used a combination drug treatment of HDAC inhibitors (HDACi) and calpeptin to determine the treatment’s effects on methylation status and expression of hTERT. Though the drug treatment caused decreased expression of the hTERT gene, the methylation status remained unaffected, indicating other mechanisms of control in the expression of telomerase. Because telomerase inhibition has been shown to cause apoptosis in cancer cells and expression of telomerase in somatic cells leads to apoptotic resistance, we looked at the effect of HDACi on apoptosis. We found increased expression of both intrinsic and extrinsic pro-apoptotic genes with this combination treatment.