Epigenetic Regulation of CFTR in Salivary Gland Cancer

The cystic fibrosis transmembrane conductance regulator (CFTR) is a ligand (ATP)-gated chloride channel mostly regulated by cAMP/ PKA signaling. CFTR plays a central role in electrolyte and fluid homeostasis in epithelia including airways, intestine, and pancreas and salivary gland ducts. Whereas mutation or lack of CFTR is widely associated with diseases including cystic fibrosis, cholera, autoimmune disease and cancer development. Despite extensive studies of the properties of luminal CFTR in salivary glands, the epigenetic modification responsible for regulating functional expression of CFTR in salivary gland cancer has been unidentified.

Here, we provide evidence that mRNA and protein expression level of CFTR were downregulated in A253, human submandibular carcinoma cancer cells, compare to HSG, human submandibular ducts, using western blot assay and RT-PCR and immunocytochemistry. To determine DNA methylation of CFTR, bisulfite sequencing and methylation-specific PCR revealed that the silencing was caused by hypermethylation of CG pairs in the CFTR CpG island and thus rectified by 5-Aza 2-deoxycytidine (5-Aza-CdR) treatment as demethylation agent. We also found similar results in human cancer tissue compared to normal tissue. we further estimated functional expression of CFTR in A253 in the presence or absence of 5-Aza-CdR by whole cell patch clamp recording that CFTR mediated inward current was observed in HSG cells but not in A253; however, CFTR was reactivated in A253 after 5-Aza-CdR treatment.

To conclude, epigenetics regulates functional expression of CFTR in salivary gland cancer cells that the CFTR is silenced by hypermethylation; therefore, we suggest that demethylation of CFTR is a strong candidate for therapeutic strategy for salivary gland cancer.