Ovarian cancer is the most frequent cause of gynecological cancer-related mortality as a majority of patients are diagnosed at an advanced stage with intraperitoneal dissemination because of the absence of initial symptoms. Granulocyte-macrophage colony-stimulating factor (GM-CSF) plays an important role in the maturation of specialized antigen-presenting cells. In this study, we utilized a herpes simplex virus (HSV) amplicon expressing murine GM-CSF combined with HF10 (mGM-CSF amplicon), a highly attenuated HSV type 1 strain functioning as a helper virus to strengthen anti-tumor immune response, for the treatment of ovarian cancer with intraperitoneal dissemination. A mouse ovarian cancer cell line, OV2944-HM-1 (HM-1), was intraperitoneally injected, following which HF10 only or the mGM-CSF amplicon was injected intraperitoneally three times. HF10 injection prolonged survival and decreased intraperitoneal dissemination, but to a lesser extent than the mGM-CSF amplicon. Although HF10 replication was not observed in HM-1 cells, expression of VP5, a late gene coding the major capsid protein of HSV, was detected. Moreover, mGM-CSF production was detected in transfected HM-1 cells. Immunohistochemical staining revealed the infiltration of CD4- and CD8-positive cells into the peritoneal tumor(s). A significantly increased CD4+ T cell concentration was observed in the spleen. Murine splenic cells after each treatment were stimulated with HM-1 cells, and the strongest immune response was observed in the mice that received mGM-CSF amplicon injections. These results suggested that the mGM-CSF amplicon is a promising agent for the treatment of advanced ovarian cancer with intraperitoneal dissemination.