Targeting the HSPA8‐CMA‐ATP6V1A Axis Triggers Lysosomal Hyperacidification and Catastrophic Vacuolation in Prostate Cancer

Prostate cancer (PCa) ranks among the most common and deadly malignancies worldwide. The clinical treatment of advanced prostate cancer is particularly challenging due to acquired drug resistance. Autophagy and lysosome-related pathways are key drivers of this resistance. Targeting the lysosome represents a potential therapeutic strategy for PCa. In this study, we identified Heat Shock Protein Family A Member 8 (HSPA8) as a critical functional node of Aloperine (ALO). ALO suppresses autophagic flux, disrupts lysosomal homeostasis, and induces lysosomal vacuolation in cancer cells by inhibiting the function of HSPA8, impairing chaperone-mediated autophagy (CMA)-mediated ATP6V1A degradation. The resulting pathological accumulation and enhanced V1-V0 association of the V-ATPase complex drive pronounced lysosomal hyperacidification and severe osmotic swelling. This biochemical and physical stress is associated with lysosomal membrane permeabilization (LMP) and downstream loss of lysosomal integrity. Furthermore, we reveal that ALO-induced vacuolation triggers a compensatory upregulation of cholesterol biosynthesis to buffer membrane expansion; preemptively disrupting this adaptive response with the DHCR7 inhibitor AY9944 yields significant synergistic lethality. Collectively, our findings reveal the specific cytotoxic mechanism of ALO and demonstrate that pharmacological targeting of the HSPA8-CMA-ATP6V1A axis is a valuable strategy for inducing lethal lysosomal vacuolation in advanced PCa.