Protein arginine methyltransferase 5 (PRMT5) is the main type II enzyme that catalyzes symmetric dimethylarginine of histone proteins to induce gene silencing by generating repressive histone marks, including H2AR3me2s, H3R8me2s, and H4R3me2s. PRMT5 can also methylate nonhistone proteins such as the transcription factors p53, E2F1 and p65[2]. HLCL-61 hydrochloride is a first-in-class inhibitor of PRMT5. HLCL-61 showed effective inhibition of symmetric arginine dimethylation (me2) of histones H3 and H4 in AML samples, starting at 12 hours post-treatment and persisting after 48 hours. PRMT5 inhibition by HLCL-61 and PRMT5 overexpression led to a decrease or increase, respectively, of both H4R3me2 and the presence of PRMT5 on the miR-29b regulatory element in MV4–11 cells. HLCL-61 led not only to the decrease of the PRMT5 downstream target Sp1 but also to a substantial decrease of FLT3 (both WT and ITD) levels in cell lines and blasts. A significant decrease in Sp1 enrichment on the FLT3 promoter was associated with downregulation of FLT3 expression in MV4–11.Similarly, knock-down of PRMT5 by shRNA decreased FLT3 mRNA and protein expression. Conversely, forced PRMT5 expression resulted in increased levels of FLT3 mRNA and protein in vitro and in vivo. HLCL-61–treated AML cell lines and patient samples exhibited induction of differentiation as suggested by dose-dependent increases in the expression of CD11b. HLCL-61 treatment for 6 hours decreased the luciferase activity (4-fold) compared to DMSO-treated control. In conclusion, treatment with this selective PRMT5 inhibitor significantly increased miR-29b expression, downregulated FLT3 levels, and affected cell survival and proliferation of AML blasts[3].
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