Alkylator-induced DNA excision repair in human leukemia CCRF-CEM cells in vitro, measured using the single-cell gel electrophoresis (comet) assay.
The capacity to repair DNA damage is an important factor that affects the therapeutic outcome in cancer treatment. To clarify the cellular repair response, we investigated the kinetics of DNA excision repair initiated by 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) in human leukemia CCRF-CEM cells at an exponential growth phase in vitro. Using the alkaline single-cell gel electrophoresis (comet) assay, we quantitated the repair kinetics as the amount of DNA single-strand breaks that were generated from the incision and were diminished by the rejoining in the repair process. CEM cells could initiate DNA excision repair in response to BCNU by starting an incision reaction. However, the incision capacity came to a plateau at a concentration of 80 to 100 microM or after an incubation time of 90 to 120 minutes. When the cells were pulsed with 40 microM BCNU, the maximal incision occurred at the end of the incubation period, and the repair process was completed within 4 hours When cells were treated with 100 microM BCNU, the incised DNA was not rejoined at 4 hours, suggesting that the repair was not completed. Higher concentrations might surpass the cellular capacity for repair and would be associated with increased cell death. Evaluation of the repair process may provide a clue for therapeutic strategies to improve clinical efficacy if accelerated DNA repair is responsible for the drug resistance.
Yamauchi
Takahiro
T
First Department of Internal Medicine, Fukui Medical University, Matsuoka, Fukui, Japan. tyamauch@fmsrsa.fukui-med.ac.jp
Kawai
Yasukazu
Y
Ueda
Takanori
T
D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology*; D002330:Carmustine / Q000494:pharmacology; D016923:Cell Death / Q000187:drug effects; D020552:Comet Assay; D004260:DNA Repair / Q000187:drug effects*; D006801:Humans; D007700:Kinetics; D007938:Leukemia / Q000188:drug therapy / Q000235:genetics* / Q000473:pathology; D014407:Tumor Cells, Cultured
D018906:Antineoplastic Agents, Alkylating; D002330:Carmustine