– (-) Menthol Induces Reversal of Promoter Hypermethylation and Associated Up-Regulation of the FANCF Gene in the SiHa Cell Line

OBJECTIVE: To identify natural bioactive molecules with potential to inhibit DNA methyltransferase 1 (DNMT1) and cause reactivation of genes silenced due to promoter hypermethylation. METHODS AND RESULTS: -(-) Menthol and epigallocatechin-3-gallate (EGCG) (reference molecule) were investigated using an in vitro methylation assay, which indicated potential of -(-) menthol as an epigenetic modulator with the ability to directly inhibit M.SssI (an analogue of DNMT1) activity at 100µM. Methylation specific PCR and bisulphite sequencing revealed complete hypomethylation of 15 CpG sites in the Fanconi anemia, complementation group F (FANCF) gene between +280 and + 432 nucleotides relative to the transcription start site, which resulted in significant (P<0.001) up-regulation of FANCF gene expression by 2.1 and 2.5 fold respectively after treatment with menthol (80µM) and EGCG (80µM) for 4 days in the SiHa cell line as analyzed by qRT PCR. CONCLUSION: The present work highlighted the potential of -(-) menthol, a naturally occurring cyclic monoterpene, as an epigenetic modulator causing promoter hypomethylation induced reactivation of the FANCF gene mediated by possible inhibition of DNMT1 activity in the SiHa cell line.