To explore the in vitro antitumor effect of interleukin-24 (IL-24) gene regulated by ovarian-specific promoter-1 (OSP-1) on human ovarian cancer cell line SKOV3. An expression vector (pcDNA3.0-OSP-1-IL-24) containing IL-24 gene under ovarian-specific promoter-1 was constructed by molecular biological methods and then transfected into human ovarian cancer cell SKOV3, human hepatoma cell HepG2 and human fibroblast BJ by cationic liposome. Reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) were used to detect the IL-24 gene in cells and supernatants. The antitumor effects of IL-24 gene were investigated by direct cell count, flow cytometry and methyl thiazolyl tetrazolium (MTT) assay. After transfections with pcDNA3.0-OSP-1-IL-24, the expression of IL-24 gene was detected only in SKOV3 cells. And only the growth of SKOV3 cells was inhibited significantly while those of HepG2 and BJ cells were not affected. The culture supernatant of SKOV3 cells transfected with pcDNA3.0-OSP-1-IL-24 could significantly inhibit the growth of SKOV3 cells. But the antitumor efficiency in SKOV3 cells of OSP-1 system was significantly lower than that of CMV promoter system. With a high specificity, IL-24 gene therapy under the control of ovarian-specific promoter-1 is a novel target for the treatment of ovarian cancer.