In vitro and in vivo radiosensitization induced by the DNA methylating agent temozolomide.
OBJECTIVE
Temozolomide, a DNA methylating agent, is currently undergoing clinical evaluation for cancer therapy. Because temozolomide has been shown to increase survival rates of patients with malignant gliomas when given combined with radiation, and there is conflicting preclinical data concerning the radiosensitizing effects of temozolomide, we further investigated the possible temozolomide-induced enhancement of radiosensitivity.


METHODS
The effects of temozolomide on the in vitro radiosensitivity of U251 (a human glioma) and MDA-MB231BR (a brain-seeking variant of a human breast tumor) cell lines was evaluated using clonogenic assay. DNA damage and repair were evaluated using phosphorylated histone H2AX (gammaH2AX), and mitotic catastrophe was measured using nuclear fragmentation. Growth delay was used to evaluate the effects of temozolomide on in vivo (U251) tumor radiosensitivity.


RESULTS
Exposure of each cell line to temozolomide for 1 h before irradiation resulted in an increase in radiosensitivity with dose enhancement factors at a surviving fraction of 0.1 ranging from 1.30 to 1.32. Temozolomide had no effect on radiation-induced apoptosis or on the activation of the G(2) cell cycle checkpoint. As a measure of DNA double strand breaks, gammaH2AX foci were determined as a function of time after the temozolomide + irradiation combination. The number of gammaH2AX foci per cell was significantly greater at 24 h after the combined modality compared with the individual treatments. Mitotic catastrophe, measured at 72 h, was also significantly increased in cells receiving the temozolomide + irradiation combination compared with the single treatments. In vivo studies revealed that temozolomide administration to mice bearing U251 tumor xenografts resulted in a greater than additive increase in radiation-induced tumor growth delay with a dose enhancement factor of 2.8.


CONCLUSIONS
These results indicate that temozolomide can enhance tumor cell radiosensitivity in vitro and in vivo and suggest that this effect involves an inhibition of DNA repair leading to an increase in mitotic catastrophe.
Kil
Whoon Jong
WJ
Radiation Oncology Branch, National Cancer Institute, Bethesda, Maryland 20892, USA.
Cerna
David
D
Burgan
William E
WE
Beam
Katie
K
Carter
Donna
D
Steeg
Patricia S
PS
Tofilon
Philip J
PJ
Camphausen
Kevin
K
D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D001932:Brain Neoplasms; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D019175:DNA Methylation; D004261:DNA Replication / Q000187:drug effects; D003606:Dacarbazine / Q000031:analogs & derivatives* / Q000494:pharmacology; D005909:Glioblastoma; D006801:Humans; D008938:Mitosis / Q000187:drug effects; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology*; D000077204:Temozolomide; D014410:Tumor Stem Cell Assay
D018906:Antineoplastic Agents, Alkylating; D011838:Radiation-Sensitizing Agents; D003606:Dacarbazine; D000077204:Temozolomide