EZM2302 是选择性口服精氨酸甲基转移酶 CARM1 抑制剂，IC50值为 6 nM。

EZM2302 is a selective, and orally available arginine methyltransferase CARM1 inhibitor (IC50: 6 nM).

EZM2302 (GSK3359088) is an inhibitor of CARM1 enzymatic activity in biochemical assays (IC50?=?6?nM) with broad selectivity against other histone methyltransferases. Ninety-six-hour EZM2302 treatment led to a concentration-dependent decrease in methylation of PABP1 (IC50?=?0.038?μM) and SmB (increased levels of SmBme0, EC50?=?0.018?μM), as well as in multiple aDMA bands (IC50?=?0.009?μM). Similar results were also observed in the NCI-H929 and U266B1 MM cell lines.

Tumor growth inhibition studies were performed in SCID mice bearing subcutaneous RPMI-8226 xenografts with twice daily (BID) oral dosing on four dose groups: 37.5, 75, 150 and 300?mg per kilogram (mg/kg). EZM2302 showed dose-dependent exposure and tumor growth inhibition (TGI) after 21 days in the RPMI-8226 xenograft model. Tumors in all EZM2302 dose groups measured on day 21 showed significant decreases in tumor growth compared to vehicle. Tumor growth inhibition ranged from 45% in the 37.5?mg/kg dose group to 63% in the 300?mg/kg dose group.

CARM1 activity was measured as previously reported for the histone methyltransferases PRMT1/6/865 and PRMT558. Briefly, CARM1 was preincubated with compounds for 30?minutes at room temperature before reactions were initiated. Final assay conditions were 0.25?nM CARM1, 30?nM 3H-S-adenosyl-methionine (SAM), and 250?nM biotinylated peptide in buffer containing 20?mM bicine, 1?mM tris(2-carboxyethyl)phosphine, 0.005% bovine skin gelatin and 0.002% Tween-20, pH 7.5. The assays were quenched by the addition of 300?μM unlabeled SAM. The quantity of 3H-labeled peptide produced was measured by Flashplate.

Cultured cells in linear/log phase growth were split to a seeding density of 2e5 cells/mL in 2–20mLs of media, depending on the yield required at the end of the growth period. The compound was diluted in DMSO and added to each culture vessel with a final DMSO concentration of 0.2%. Cells were allowed to grow for 96?hours. At the conclusion of each treatment period, cells were harvested by centrifugation (5?minutes, 1200?rpm), and cell pellets were rinsed once with PBS before being frozen on dry ice pending further processing.

For the in vivo efficacy studies, there were 8 mice per dose group and each mouse was inoculated subcutaneously at the right flank. All cells were suspended in a 0.2?mL mixture of base media and Matrigel at 1:1 for tumor development. RPMI-8226 cells were inoculated at 5?×?10^6 cells/mouse and treatment began when the mean tumor sizes reached 120?mm3 (28 days post-inoculation). CB-17 SCID Mice were assigned into groups using a randomized block design. EZM2302 or vehicle (0.5% methylcellulose in water) was administered orally BID at a dose volume of 37.5, 75, 150, or 300?mg/kg for 21 days. Body weights were measured twice a week for the duration of the study. Tumor size was measured twice weekly in two dimensions using a caliper, and the volume was expressed in cubic millimeters. Animals were euthanized 3?hours post-final dose, with blood and tissues collected for analysis.

1628830-21-6

C29H37ClN6O5

585.1

GSK3359088

DMSO:100 mg/mL (170.91 mM)
Powder: -20°C for 3 years
In solvent: -80°C for 2 years