A bis(benzimidazolyl)-based anion transporter triggers chloride-initiated pyroptosis of MDA-MB-468 and 4T1 triple-negative breast cancer cells in vitro and in vivo.

Abstract

As a highly heterogeneous subtype of breast cancers, triple-negative breast cancer (TNBC) poses significant therapeutic challenges. This study aims to systematically investigate the activity and mechanism of action of a benzimidazole-based anion transporter (i.e., compound 1) in treating TNBC both in vivo and in vitro. The results showed that compound 1 markedly inhibited the proliferation of TNBC cells with high selectivity. Mechanistic studies demonstrated that compound 1 increased the levels of intracellular Cl- and induced osmotic stress, mitochondrial dysfunction, and lysosomal alkalinization, leading to reactive oxygen species (ROS) accumulation, DNA damage, and reduced deoxyribonuclease II (DNase II) and cathepsin B (CTSB) levels. These alterations further activated the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING)-TANK-binding kinase 1 (TBK1)-interferon regulatory factor 3 (IRF3)/nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway, promoted NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome activation and gasdermin D (GSDMD) cleavage, and ultimately induced pyroptosis in MDA-MB-468 and 4T1 cells. In MDA-MB-468 and 4T1 breast cancer nude mouse models, compound 1 exhibited significant antitumor activity without obvious toxicity. In vivo analyses further confirmed the activation of pyroptosis-related signaling in tumor tissues. These results demonstrate that benzimidazole-based anion transporters drive inflammatory signaling and pyroptosis in TNBC cells.