DI-591 is a potent, high-affinity and cell-permeable inhibitor of the DCN1-UBC12 interaction. DI-591 binds to DCN1 and DCN2 with K i values of 12 nM and 10.4 nM, respectively and has no appreciable binding to DCN3, DCN4, and DCN5 proteins. DI-591 selectively inhibits neddylation of cullin 3 but has no or minimal effect on neddylation of other cullin family members
In Vitro
DI-591 (Compound 44) binds to DCN1 and DCN2 with K i values of 12 nM and 10.4 nM, respectively and has no appreciable binding to DCN3, DCN4, and DCN5 proteins. Hence, DI-591 displays a very-high binding affinity to recombinant human DCN1 and DCN2 proteins and >1000-fold selectivity over recombinant human DCN3-5 proteins. DI-591 (Compound 44; 0-10 μM; 1 hour; KYSE70 cells) binds to both cellular DCN1 and DCN2 proteins and disrupts the association of cellular DCN1 and UBC12 proteins. DI-591 (Compound 44; 10 μM; 24 hours; THLE2 cells) treatment robustly increases the mRNA levels of NQO1 and HO1, leading to upregulation of HO1 protein in the cells. Significantly, DI-591 has no effect on the mRNA level of NRF2. The selective inhibition of neddylation of cullin 3 by DI-591 leads to accumulation NRF2 protein and its transcriptional activation. Knockdown experiments indicate that DCN1, but not DCN2, plays a key role in regulation of neddylation of cullin 3 but not of other cullins. DI-591 is an excellent probe compound to investigate the role of the cullin 3 CRL (Cullin-RING E3 ubiquitin ligase) in biological processes and human diseases. MCE has not independently confirmed the accuracy of these methods. They are for reference only. Western Blot AnalysisCell Line: KYSE70 cells Concentration: 0 μM, 1 μM, 3 Μm or 10 μM Incubation Time: 1 hour Result: Potently bound to cellular DCN1 and DCN2 proteins. Enhanced the stability of DCN1 and DCN2 protein in a dose-dependent manner. RT-PCRCell Line: THLE2 cells Concentration: 10 μM Incubation Time: 24 hours Result: Robustly increases the mRNA levels of NQO1 and HO1.
