UNC0646

UNC0646 is a novel G9a inhibitor with excellent potency in a variety of cell lines and excellent separation of functional potency versus cell toxicity. UNC0646 has high in vitro potency versus G9a and improved lipophilicity, are highly potent (IC50 < 0.06 μM) in reducing H3K9me2 levels in MDA-MB-231 cells and has low cell toxicity.^[1]

Cells are seeded at 5–10 thousand cells in black walled 96 well plates and exposed to various inhibitor concentrations for 48h. The media is removed and 2% Formaldehyde in PBS for fixation is added for 15 min. After five washes with 0.1% Triton X100 in PBS, cells are blocked for 1h with 1% BSA in PBS. Three out of four replicates are exposed to the primary H3K9me2 antibody for 2 h. One replicate is reserved for the background control. The wells are washed five times with 0.1% Tween 20 in PBS, then secondary IR800 conjugated antibody and nucleic acid-intercalating dye, DRAQ5 added for 1 h. After 5 washes with 0.1% Tween 20 PBS, the plates are read on an Odyssey scanner at 800 nm (H3K9me2 signal) and 700 nm (DRAQ5 signal).

Due to its water insolubility, the in vivo efficacy of UNC0646 is not satisfactory. Nanodiamonds (NDs) are utilized as a drug delivery platform to improve in vivo delivery of this small-molecule inhibitor. ND-UNC0646 complexes can be rapidly synthesized by physical adsorption, meanwhile possessing favorable drug delivery properties and is able to improve the dispersibility of UNC0646 in water, therefore making it amenable for intravenous administration. The release profile of UNC0646 from NDUNC0646 is demonstrated to be pH-responsive. Moreover, ND-UNC0646 maintains the biological functionality of UNC0646, with higher efficacy in reducing H3K9 methylation as well as enhanced invasion suppressive effects. Most importantly, increased in vivo efficacy is demonstrated using an orthotopic Hepatocellular carcinoma (HCC) mouse model, which paves the way of translating this small-molecule inhibitor toward HCC treatment.^[2]