Currently, there is no long-term effective treatment for unresectable hepatic malignancies. Salmonella species are known to naturally track to the liver during active infection. To develop a biological vector for delivery of interleukin-2 (IL-2) to the liver for antitumor purposes, the thi 4550 attenuated strain of Salmonella typhimurium was used as a vector for IL-2. The gene for human IL-2 was cloned into plasmid pYA292 and inserted into the attenuated S typhimurium and renamed (thi 4550(pIL-2)]. MCA-38 murine adenocarcinoma cells were injected intrasplenically into C57BL/6 mice to produce hepatic metastases that were subsequently enumerated after 12 days. We previously have demonstrated that the thi 4550(pIL-2) produces biologically active IL-2 and that a single gavage feeding of 10(7) thi 4550(pIL-2) significantly reduced the number of hepatic metastases when compared with animals fed salmonella lacking the IL-2 gene or nontreated controls. The aims of the current studies were to determine which host effector cell populations were responsible for the antitumor effect seen with thi 4550(pIL-2) by depletion of natural killer (NK), cytotoxic T lymphocytes (CD8+), T helper (CD4+) cells, and Kupffer cells. Multiple experiments were conducted for each host effector cell population depleted. We found a consistent reduction in the mean number of hepatic metastases in animals fed thi 4550(pIL-2) (55.6 metastases; n = 54) when compared with controls (162.3 metastases; n = 53) (P < .0001). Depletion of NK cells and CD8+ T cells significantly inhibited the antitumor effect of thi 4550(pIL-2) (analysis of variance [ANOVA], P < .01). Elimination of CD4+ T cells and Kupffer cells had no significant impact on the antitumor effect of thi 4550(pIL-2) (ANOVA, P value was not significant). Salmonella IL-2 may represent a novel form of in vivo biotherapy for unresectable hepatic malignancies that employs the oral route of administration. Furthermore, both NK cells or CD8+ cells are required for the antitumor effect seen while CD4+ T cells and Kupffer cells do not appear to be as essential.