Salvianolic acid B suppresses non-small cell lung cancer growth by promoting Nrf2 proteasomal degradation and triggering autophagy-dependent ferroptosis.

Abstract

<p>Non-small cell lung cancer (NSCLC) constitutes the predominant lung cancer subtype and remains a leading cause of cancer-related mortality worldwide, underscoring an imperative need for mechanistically informed therapeutic strategies. Salvianolic acid B (SalB), a bioactive polyphenolic compound isolated from Salvia miltiorrhiza, has demonstrated antitumor potential across several malignancies; however, the precise molecular mechanisms governing its activity in NSCLC remain largely undefined. The anticancer activity of SalB was evaluated in human NSCLC cell lines (A549, H460 and PC-9) and in an A549 xenograft mouse model. Apoptosis, proliferation, autophagy, oxidative stress, ferroptosis-associated phenotypes, and mitochondrial function were assessed using flow cytometry, clonogenic assays, immunofluorescence, transmission electron microscopy, and Western blotting. Nuclear factor erythroid 2-related factor 2 (Nrf2) protein stability and ubiquitination were examined using proteasome inhibition and immunoprecipitation assays. The functional relevance of Nrf2 was further validated by gain-of-function approaches both in vitro and in vivo. SalB potently induced apoptosis and abrogated the proliferative capacity of NSCLC cells. Mechanistically, SalB engaged autophagic machinery and elicited hallmark features of ferroptosis, including excessive reactive oxygen species generation, labile iron accumulation, glutathione depletion, elevated malondialdehyde (MDA) accumulation, mitochondrial depolarization, and coordinate downregulation of critical ferroptosis defense proteins-glutathione peroxidase 4 (GPX4), ferritin heavy chain 1 (FTH1), and the cystine/glutamate antiporter xCT (solute carrier family 7 member 11, SLC7A11). Pharmacological blockade of autophagic flux partially attenuated SalB-induced oxidative stress and iron dysregulation, indicating that autophagy contributes to SalB-induced ferroptosis-associated phenotypes. At the molecular level, SalB accelerated proteasome-dependent degradation of Nrf2 by selectively potentiating K48-linked polyubiquitination. Enforced Nrf2 overexpression partially attenuated SalB-induced autophagy activation, ferroptosis-associated molecular alterations, apoptotic cell death, and proliferative suppression. Concordantly, Nrf2 overexpression substantially abrogated the antitumor efficacy of SalB in vivo. These findings establish that SalB suppresses NSCLC progression by promoting Nrf2 proteasomal degradation, thereby enhancing autophagy-associated ferroptotic vulnerability. Pharmacological targeting of the Nrf2-autophagy-ferroptosis regulatory axis may provide a potential therapeutic avenue for NSCLC.</p>