Discovery of new 7-azaindole derivatives as CSN5 inhibitors with in vitro antitumor activity against triple-negative breast cancer.

COP9 signalosome subunit 5 (CSN5) exerts critical deneddylase activity and is frequently overexpressed in triple-negative breast cancer (TNBC), representing a compelling antitumour target. However, to date, only a limited number of inhibitors targeting CSN5 have been reported, and none of them have advanced into clinical studies. Based on lead compound 1, we designed and synthesized two series of 7-azaindole derivatives functionalized with piperazinylethylamine (2-7) and propylenediamine (8-19) side-chains. Combined enzymatic fluorescence polarization assays and cellular NEDD8-Cul1-based biomarker screening identified CSN5-6 and CSN5-14 as key hits. Both compounds induce the accumulation of NEDD8-Cul1 in TNBC cells without altering the protein level of CSN5. Molecular docking rationalized their binding interactions inside the CSN5 catalytic pocket. CSN5-6 and CSN5-14 inhibited TNBC cell proliferation and clonogenic survival while inducing cell-cycle redistribution and apoptosis, with CSN5-14 showing superior efficacy. Mechanistically, high-level CSN5 inhibition provoked ROS production and G2/M-phase arrest. Furthermore, CSN5-14 displayed rapid absorption, favorable systemic exposure and moderate elimination upon intraperitoneal dosing in SD rats, showing promising preliminary pharmacokinetic properties. This study broadens the chemical space for CSN5 inhibitors, and CSN5-14 represents a promising lead compound for further anti-TNBC therapeutic development.