Abstract

Bladder cancer (BC) is an aggressive and treatment-resistant malignancy with a high recurrence rate. Current first-line therapies only provide limited relief due to chemoresistance and toxicity. Natural products are emerging as promising chemotherapeutic agents due to their multi-target mechanisms and excellent safety profile. In this study, we evaluated the anti-BC effects of echinocystic acid (EA), a pentacyclic triterpenoid, on BC cells and investigated the underlying mechanism. We found that EA effectively inhibited proliferation, triggered G1-phase cell cycle arrest, and simultaneously suppressed cellular invasion and migration capabilities in vitro . Although EA did not obviously induce apoptosis in BC cells at the doses that were effective in suppressing their growth, it successfully triggered ferroptosis within these cells. Moreover, EA suppressed xenograft tumor growth of BC cells in vivo and demonstrated a favorable safety profile. Mechanistically, EA interacted with AKT1, reducing its phosphorylation and thereby inhibiting the AKT1/GSK3β/β-Catenin signaling pathway. Furthermore, EA exhibited synergistic effects with gemcitabine in inhibiting BC cells in vitro and in vivo . Overall, our results suggest that EA may exert its anti-BC effects, at least in part, by inhibiting the AKT1/GSK3β/β-Catenin signaling pathway. When combined with the clinically approved anti-tumor drug gemcitabine, EA may produce a synergistic anti-BC effect. Most importantly, this study identifies EA as a novel inducer of ferroptosis in BC and underscores its potential for clinical application.