The present study investigated the effect of signal transducer and activator of transcription 3 (Stat3) interference on RM1 prostate cancer cell viability in vitro, using plasmid‑based Stat3 specific short hairpin RNA (sh‑Stat3) delivered by hydroxyapatite nanoparticles (HAP). HAP carrying sh‑Stat3 plasmids were transfected into tumor cells. MTT assays were used to measure RM1 cell viability 24 and 48 h following transfection, and the apoptosis rate and cell cycle phase distribution were determined by flow cytometry. Stat3 mRNA expression levels were measured by reverse transcription‑quantitative polymerase chain reaction and Stat3, Cyclin D1, B cell lymphoma 2 apoptosis regulator (Bcl‑2), vascular endothelial growth factor (VEGF), Bcl‑2 associated X apoptosis regulator (Bax) and cleaved‑caspase‑3 protein expression levels were detected using western blot analysis. The results demonstrated that HAP‑delivered sh‑Stat3 significantly decreased RM1 cell viability through the promotion of cell cycle arrest and apoptosis. Stat3 mRNA and protein expression levels were significantly downregulated in RM1 cells. Bcl‑2, VEGF and Cyclin D1 were also significantly downregulated, but cleaved‑caspase‑3 and Bax mRNA and protein expression levels were significantly upregulated. HAP‑delivered sh‑Stat3 decreased RM1 cell viability in vitro, and HAP assisted plasmid‑based delivery of shRNA into tumor cells. The present results suggest that HAP may be a useful method for successful shRNA delivery into tumors.