USP7/USP47 inhibitor (USP7/47 inhibitor-1) is a selective ubiquitin-specific protease 7/47 (USP7/USP47) inhibitor, with EC50s of 0.42 μM and 1.0 μM, respectively.

USP47:1.0 μM(EC50), USP7:0.42 Μm(EC50)

USP7/USP47抑制剂不抑制caspase 3、calpain 1、20S蛋白酶体以及一组代表性的USPs（USP2、USP5、USP8、USP21和USP28；EC50 > 31.6 μM）。USP7/USP47抑制剂能够抑制HCT-116细胞的生长，其EC50为7.6 μM。

The cloning, expression and purification of USP21 from BL21 (DE3) bacteria are performed using standard molecular biology techniques. USP2, USP5, USP7, USP8, USP28, USP47, Ub-PLA2 (Ub-CHOP) and Ub-EKL (Ub-CHOP2) are generated. Caspase 3 and the caspase 3 substrate DEVD-Rh110 are used. Deubiquitylating enzyme, cathepsin B and 20S proteasome chymotrypsin like protease activities are measured. Caspase 3 activity is determined using a similar protocol. Briefly, dose ranges of compound (including USP7/USP47 inhibitor) are incubated with caspase 3 for 30 minutes before the addition of DEVD-Rh110 and reading on a fluorometric plate reader using excitation and emission maxima of 485 nm and 531 nm respectively. The final concentrations of caspase 3 and DEVD-Rh110 are 2 nM and 100 nM respectively