Kaempferol Modulates DNA Methylation and Downregulates DNMT3B in Bladder Cancer

Background: Genomic DNA methylation plays an important role in both the occurrence and development of bladder cancer. Kaempferol (Kae), a natural flavonoid that is present in many fruits and vegetables, exhibits potent anti-cancer effects in bladder cancer. Similar to other flavonoids, Kae possesses a flavan nucleus in its structure. This structure was reported to inhibit DNA methylation by suppressing DNA methyltransferases (DNMTs). However, whether Kae can inhibit DNA methylation remains unclear. Methods: Nude mice bearing bladder cancer were treated with Kae for 31 days. The genomic DNA was extracted from xenografts and the methylation changes was determined using an Illumina Infinium HumanMethylation 450 BeadChip Array. The ubiquitination was detected using immuno-precipitation assay. Results: Our data indicated that Kae modulated DNA methylation in bladder cancer, inducing 103 differential DNA methylation positions (dDMPs) associated with genes (50 hyper-methylated and 53 hypo-methylated). DNA methylation is mostly relied on the levels of DNMTs. We observed that Kae specifically inhibited the protein levels of DNMT3B without altering the expression of DNMT1 or DNMT3A. However, Kae did not downregulate the transcription of DNMT3B. Interestingly, we observed that Kae induced a premature degradation of DNMT3B by inhibiting protein synthesis with cycloheximide (CHX). By blocking proteasome with MG132, we observed that Kae induced an increased ubiquitination of DNMT3B. These results suggested that Kae could induce the degradation of DNMT3B through ubiquitin-proteasome pathway. Conclusion: Our data indicated that Kae is a novel DNMT3B inhibitor, which may promote the degradation of DNMT3B in bladder cancer.