AIM: To explore the effect of microRNA-34a (miR-34a) on androgen receptor gene (AR) targeting regulation and proliferation and apoptosis of prostate cancer (PCa) LNCaP cells. METHODS: Thirty-six patients with PCa confirmed by prostate biopsy in urology department of our hospital were collected from October 2016 to September 2019, and 41 cases of benign prostatic hyperplasia (BPH) tissue samples were taken and operated at the same time. LNCaP cells were cultured in vitro and transfected with miR-34a mimics (mimics group), miR-34a mimic NC (NC group) respectively, and normal growth cells were set as blank control group (BC group). In addition, on the basis of mimics group, the AR over-expression vector (AR over-expression group) and its control group (AR control group) were transfected. The cell activity was detected by CCK-8 kit, the apoptosis rate was detected by Annexin V-FITC/PI double staining flow cytometry test kit, the expression of miR-34a and AR mRNA was detected by real-time quantitative PCR (RT-qPCR), the expression of AR protein, CyclinD1, c-Myc and apoptosis-related proteins (Bcl-2, Bax, capase-3) was detected by Western blotting. RESULTS: Compared with BPH tissues, the expression of miR-34a in PCa tissues decreased significantly (P<0.05), and the expression of AR mRNA and protein increased significantly (P<0.05). Compared with BC and NC group, miR-34a expression level, apoptosis rate and Bax protein expression level of LNCaP cells in mimics group increased significantly (P<0.05), the expression levels of AR, Cyclin D1, c-Myc and Bcl-2 decreased significantly (P<0.05), the activity of LNCaP cells decreased significantly at the same time (P<0.05). The dual-luciferase assay showed that miR-34a may have a direct targeting relationship with AR. Over-expression of AR gene could reverse the inhibition of miR-34a mimics on LNCaP cell proliferation and promote apoptosis. CONCLUSION: miR-34a may inhibit the proliferation and promote the apoptosis of prostate cancer LNCaP cells by regulating AR gene.