Currently chemotherapy drugs are usually used as first-line treatments for castration-resistant prostate cancer (CRPC), but they are ineffective and accompanied by serious side effects. MicroRNA-34a (miR-34a) simultaneously targets multiple genes related to the cell apoptosis in CRPC cells without obvious side effects. It has shown great potential in the treatment of CRPC. Previous studies focused on miR-34a increasing the sensitivity of chemotherapy drugs to chemoresistant prostate cancer cells. There are few researches on miR-34a alone in the treatment of CRPC. But the macromolecular miR-34a is difficult to enter the cell and is easily degraded by nuclease. Therefore, we constructed methoxy polyethylene glycol-polylacticco-glycolic acid-polylysine (mPEG-PLGA-PLL) nanoparticles to encapsulate miR-34a (miR-34a/NP). The results showed that miR-34a/NP protects miR-34a from degradation by nucleases and can be phagocytized by PC-3 CRPC cells. Ultrasound induces microbubble cavitation (UIMC) improves cell membrane permeability and capillary gaps, and further promotes miR-34a/NP to enter cells PC-3 and prostate cancer xenografts. The miR-34a/NP that enters the cell and tumor tissue releases miR-34a, which suppressed CRPC cells PC-3 proliferation, promoted its apoptosis, and inhibited the growth of CRPC xenografts. Our research verified that miR-34a/NP, especially combined with UIMC, has a significant anti-tumor effect on CRPC.