DI-591 is an effective, high-affinity and cell-permeability inhibitor for DCN1-UBC12 interaction. DI-591 binds to DCN1 and DCN2 with Ki values of 12 and nM and 10.4 nM, respectively, and has no obvious binding to DCN3, DCN4 and DCN5 proteins.

DI-591 binds to DCN1 and DCN2 with Ki values of 12 nM and 10.4 nM, respectively, and has no obvious binding to DCN3, DCN4 and DCN5 proteins. Therefore, DI-591 has a high binding affinity for recombinant human DCN1 and DCN2 proteins, and its selectivity is more than 1000 times that of recombinant human DCN3-5 protein [1]. DI-591 (0-10μM; 1 hour; KYSE70 cells) binds to the DCN1 and DCN2 proteins of the cells and disrupts the binding of the DCN1 and UBC12 proteins. DI-591 (10 μM; 24 hours; THLE2 cells) treatment can strongly increase the mRNA levels of NQO1 and HO1, resulting in the up-regulation of HO1 protein in the cells. It is worth noting that DI-591 has no effect on the mRNA level of NRF2. DI-591's selective inhibition of cullin 3's dienylation leads to the accumulation of NRF2 protein and its transcriptional activation. Knockdown experiments show that DCN1, but not DCN2, plays a key role in the regulation of dimerization of cullin 3 but not other cullins. DI-591 is an excellent probe compound for studying the role of cullin 3 CRL (Cullin-RING E3 ubiquitin ligase) in biological processes and human diseases.