Interaction of SR-4233 with Hyperthermia and Radiation in the FSallC Murine Fibrosarcoma Tumor System in Vitro and in Vivo

The effects of SR-4233 (3-amino-l, 2, 4-benzotriazine-l, 4-dioxide), a hypoxic cell cytotoxic agent, were assayed against the FSallC murine fibrosarcoma in vitro and in vivo alone and in conjunction with hyperthermia and radiation. In vitro, a concentration of 500 pM of SR-4233 upon exposure of the cells for 1 h decreased the survival of hypoxic cells by about 1 log more than euoxic cells at 37°C and pH 7.40. At the same concentration at pH 6.45, this difference in cytotoxicity increased to about 3 logs. In conjunction with 42 or 43°C hyperthermia at pH 7.40, the killing of both euoxic and hypoxic cells was markedly increased (hypoxic >oxic), and the effect of hyperthermia on SR-4233 cytotoxicity was further increased at pH 6.45. SR-4233 proved to be an effective radiosensitizer of hypoxic cells in vitro, producing an enhancement ratio of 2.6 ± 0.2 at pH 7.40 and 2.7 ± 0.2 at pH 6.45. In vivo, however, SR-4233 (50 mg/kg) used with single dose radiation (10, 20, or 30 Gy) did not alter the slope of the radiation dose-dependent tumor growth delay curve but did produce a significant additive increase in tumor growth delay. Local hyperthermia (43°C, 30 min) plus SR-4233 (30 mg/kg) produced a tumor growth delay of 9.1 ± 2.2 days, whereas SR-4233 alone caused a tumor growth delay of only 1.7 ± 0.9 days and the hyperthermia, only 1.4 ± 0.7 days. The tumor growth delay increased to 28.2 ± 4.4 days with the addition of daily radiation (3 Gy for 5 days) to SR-4233 and hyperthermia given on treatment day 1 only. Hoechst 33342 dye-selected tumor subpopulation analysis at 24 h following treatment demonstrated that SR-4233 (30 mg/kg) was more toxic to dim (presumably hypoxic) cells by about 1.8-fold. The addition of hyperthermia to treatment with SR-4233 increased the killing of dim cells by about 5-fold but of bright cells by only 2-fold. Trimodality treatment with SR-4233, hyperthermia, and radiation increased the killing of bright cells by about 6.5-fold and of dim cells by about 16.5-fold as compared with radiation alone. These results indicate that SR-4233 might be used quite effectively with radiation and/or hyperthermia to treat tumors with significant hypoxic subpopulations.