OBJECTIVE To determine if a maximally radiosensitizing but non-toxic moderate hyperthermia treatment enhances radiation-induced mitotic catastrophe. METHODS HeLa S3 cells were given a non-lethal heat treatment (41.5 degrees C, 4 h) and irradiated with 5 Gy X-irradiation. Alterations in cell-cycle distribution, intracellular cyclin B1 levels, and the yield of mitotic catastrophe were then measured and compared to heat-only and radiation-only groups, as a function of time following treatment. RESULTS A greater accumulation of cells in S and G2 phases of the first cell-cycle post-treatment was observed in the combined heat and radiation groups, when compared with that observed following treatment with heat or radiation alone. Similarly, intracellular levels of cyclin B1 and the incidence of mitotic catastrophe were found to be greater in the combined treatment groups. CONCLUSIONS This study provides further evidence that delays late in the cell cycle are implicated in increases in intracellular cyclin B1 levels and the subsequent development of mitotic catastrophe. Further, these data suggest a role for mitotic catastrophe occurring as a result of G2/M checkpoint abrogation in the process of thermal radiosensitization.