Ascofuranone (AF) showed an antitumor protective effect on L-1210 leukemia when AF was administered once 7 days before tumor challenge. However, effect was not elicited when host mice were treated with AF simultaneously with tumor challenge. AF pretreatment on day 7, 5 and 3 before tumor challenge protected the host from the ascites form of S-180. AF also retarded tumor growth when administered once daily for 5 consecutive days 24 hours after transplantation, but antitumor effect was not seen with combined treatments before and after the transplantation. Similar results were noted with Ehrlich ascites carcinoma. AF treatment of normal mice enlarged the solid lymphoid organs without affecting body weight gain. The splenocytes derived from AF-treated mice lowered mitogenic response to phytohemagglutinin, while the mitogenic response to concanavalin A and lipopolysaccharide was unaffected.