Complementary killing effect of tirapazamine in combination with radiation therapy on cells with high aldehyde dehydrogenase activity in SAS cell line

Tumor cells with high aldehyde dehydrogenase activity (ALDHhigh cells) are induced by an intratumoral hypoxic condition and lead to radioresistance, tumor recurrence, and metastasis. Therefore, to enhance the anti-tumor effect of radiotherapy, it is reasonable to efficiently control ALDHhigh cells by targeting them. In this study, we evaluated the effect of tirapazamine, a hypoxic toxin, combined with irradiation on ALDHhigh cells.

Human tongue squamous cell carcinoma SAS cells were used in this study. Spheroids were irradiated with 6 Gy following treatment with 40 μM tirapazamine. After 24, 48, and 72 hours, the populations of ALDHhigh cells were analyzed. The frozen sections of spheroids were prepared, and hypoxia-inducible factor-1α-positive areas and ALDH1-positive areas were detected.

Compared with the cells grown in monolayer culture, the SAS cells grown in spheroids exhibited radioresistance. Furthermore, the proportion of ALDHhigh cells was significantly higher in spheroids than in monolayer culture. The ALDHhigh cells were sustained in a hypoxic fraction localized at the center of the spheroids after irradiation. Tirapazamine effectively reduced these ALDHhigh cells. The combination of tirapazamine with irradiation showed an additive cytotoxic effect in spheroids, but not in parental cells, which was consistent with a preferential killing effect of tirapazamine on ALDHhigh cells.

Combined tirapazamine and radiotherapy administration appeared to be a reasonable approach to control ALDHhigh cells in hypoxic regions that may be involved in recurrence and metastasis.