S-phase cell-specific modification by gemcitabine of PFGE-analyzed radiation-induced DNA fragmentation and rejoining.
OBJECTIVE
To assess the cell cycle-dependent influence of gemcitabine on ionizing radiation-induced DNA double-strand breakage (DSB) and rejoining measured by pulsed-field gel electrophoresis (PFGE).


METHODS
WIDR cells (human colon carcinoma) were synchronized by serum starvation/stimulation providing populations with 7% (G1) or 50% S-phase cells, respectively. Following drug treatment (0.5 microg/ml for 2 hours) cells were irradiated (up to 90 Gy) or incubated for repair (up to 6 h after 40 Gy). Cell cycle changes were monitored by flow cytometry, DNA fragmentation was assessed by PFGE as fraction of electrophoretically mobile DNA.


RESULTS
Without drug treatment, irradiated S-phase cells exhibited lower PFGE signals than the G1 cells due to the well known electrophoretic immobility of replicative DNA fragments, but DSB rejoining was not different. Gemcitabine pretreatment increased the apparent initial radiation-induced DNA fragmentation specifically for S-phase cells. This effect was rapidly reversed (1 h) during incubation for repair.


CONCLUSIONS
The data indicate that gemcitabine causes the formation of additional radiation-induced DSB in S-phase cells or destabilizes the replicative structures that otherwise prevent DNA fragment migration during PFGE. The latter would be rapidly restituted superimposing DSB rejoining. This is discussed in relation to the recently proposed role of mismatch repair in gemcitabine radiosensitization.
Jensen
Alexandra
A
Department of Radiooncology, University of Heidelberg, Germany.
Debus
Jürgen
J
Weber
Klaus-Josef
KJ
D000970:Antineoplastic Agents / Q000494:pharmacology*; D045744:Cell Line, Tumor; D004247:DNA / Q000235:genetics; D053938:DNA Fragmentation / Q000187:drug effects* / Q000528:radiation effects*; D004260:DNA Repair / Q000187:drug effects*; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D016521:Electrophoresis, Gel, Pulsed-Field; D006801:Humans; D016196:S Phase / Q000187:drug effects* / Q000528:radiation effects*; D012680:Sensitivity and Specificity; D000093542:Gemcitabine
D000970:Antineoplastic Agents; D003841:Deoxycytidine; D004247:DNA; D000093542:Gemcitabine