Targeting the HSP60/p53 Axis with Extracellular Vesicle-Delivered siRNA Reprograms Glycolysis in Prostate Cancer

Prostate cancer (PCa), a most prevalent urologic malignancy in men, remains a therapeutic challenge due to limited targeted strategies. This study investigates heat shock protein 60 (HSP60) (<i>HSPD1</i>-encoded), employing multi-dimensional approaches to decipher its oncogenic role and develop siRNA-loaded extracellular vesicles (siRNA@EVs) for PCa targeted therapy. Bioinformatics screening identified <i>HSPD1</i> overexpression in PCa, which was validated via qPCR/Western blot in clinical tissues and cell lines. Metabolomic-transcriptomic integration and molecular biology experiments revealed HSP60-mediated glycolytic reprogramming. EVs were harvested from UV-irradiated PCa cells via high-speed centrifugation. siRNA@EVs were constructed via electroporation and evaluated <i>in vitro</i> (glycolysis phenotyping: glucose consumption, lactate/pyruvate production, hexokinase activity, and ATP production) and <i>in vivo</i> using xenograft models. Data were analyzed using R 4.3.1 and GraphPad Prism 9.0 (two-tailed t-test, <i>P</i> < 0.05). Multiple bioinformatics analyses (DepMap/TCGA/HPA) confirmed that HSP60 is specifically overexpressed and associated with advanced PCa progression and poor prognosis. <i>HSPD1</i> knockdown and pharmacological HSP60 inhibition suppressed proliferation, metastasis, and subcutaneous tumor growth, while overexpression exacerbated oncogenicity. Multi-omics integration revealed HSP60 enhances glycolysis via p53 suppression, driving metabolic reprogramming. siRNA@EVs achieved significant <i>HSPD1</i> silencing, effectively inhibiting the proliferation and metastasis of PCa cells, and blocking xenografts tumor growth in nude mice with safety. siRNA@EVs targeting <i>HSPD1</i> demonstrate precision therapeutic potential with robust efficacy and safety, offering a novel approach for targeted therapy in PCa.