Mechanistic Insights into Harmine-Mediated Inhibition of Human DNA Methyltransferases and Prostate Cancer Cell Growth

摘要

Mammalian DNA methyltransferases (DNMTs), including DNMT1,DNMT3A,and DNMT3B, are key DNA methylation enzymes and play important rolesin gene expression regulation. Dysregulation of DNMTs is linked tovarious diseases and carcinogenesis, and therefore except for thetwo approved anticancer azanucleoside drugs, various non-nucleosideDNMT inhibitors have been identified and reported. However, the underlyingmechanisms for the inhibitory activity of these non-nucleoside inhibitorsstill remain largely unknown. Here, we systematically tested and comparedthe inhibition activities of five non-nucleoside inhibitors towardthe three human DNMTs. We found that harmine and nanaomycin A blockedthe methyltransferase activity of DNMT3A and DNMT3B more efficientlythan resveratrol, EGCG, and RG108. We further determined the crystalstructure of harmine in complex with the catalytic domain of the DNMT3B-DNMT3Ltetramer revealing that harmine binds at the adenine cavity of theSAM-binding pocket in DNMT3B. Our kinetics assays confirm that harminecompetes with SAM to competitively inhibit DNMT3B-3L activity witha K (i) of 6.6 mu M. Cell-based studiesfurther show that harmine treatment inhibits castration-resistantprostate cancer cell (CRPC) proliferation with an IC50 of similar to 14 mu M. The CPRC cells treated with harmine resultedin reactivating silenced hypermethylated genes compared to the untreatedcells, and harmine cooperated with an androgen antagonist, bicalutamide,to effectively inhibit the proliferation of CRPC cells. Our studythus reveals, for the first time, the inhibitory mechanism of harmineon DNMTs and highlights new strategies for developing novel DNMT inhibitorsfor cancer treatment.