Dependence of Fluorodeoxyuridine-mediated Radiosensitization on S Phase Progression

Abstract

Recent evidence casts doubt on the hypotheses that fluoropyrimidine-mediated radiosensitization is related to cytotoxicity or to cell cycle redistribution into the G1/S boundary. We hypothesized that cells that are capable of progressing into S phase in the presence of fluorodeoxyuridine may also be more susceptible to radiation-induced damage. To test this hypothesis, fluorodeoxyuridine (FdUrd)-treated HT29 human colon cancer cells were separated by centrifugal elutriation into four fractions (1-4) containing a range of cells from those at the G1/S boundary (fraction 1) to those which had progressed approximately 11% into S phase (fraction 4). We found that fraction 4 cells showed significantly greater radiosensitization than fraction 1 cells. We also compared the effects of fluorodeoxyuridine on HT29 and SW620 human colon cancer cells. We found that, in contrast with HT29 cells, SW620 cells arrested at the G1/S boundary and were minimally radiosensitized. Finally, we found that an increase in sensitivity was correlated with a decrease in the rate of repair of DNA double-strand and single-strand breaks (assessed by asymmetric field inversion gel electrophoresis and alkaline elution respectively). These findings are consistent with the hypothesis that fluorodeoxyuridine-mediated radiosensitization depends on S phase progression and a decreased ability to repair radiation-induced DNA damage.

5-Fluoro-2′-Deoxyuridine-Induced cdc25A Accumulation Correlates with Premature Mitotic Entry and Clonogenic Death in Human Colon Cancer Cells

“…This result is consistent with the incomplete drug-induced S-phase arrest previously observed in HT29 cells (24,42,44). The genomic cdc25A sequence from both SW620 and HT29 cells matches that reported for wild-type human cdc25A in the National Center for Biotechnology Information database (accession no.…”

“…2A). This decrease correlates temporally with the previously described druginduced early S-phase arrest observed in these cells (24,42,44). In addition, the FdUrd-induced decrease in cdc25A was prevented by cotreatment with the proteasome inhibitor LLnL, supporting the hypothesis that FdUrd stimulated degradation of cdc25A (Fig.…”

MLH1 Deficiency Enhances Radiosensitization with 5-Fluorodeoxyuridine by Increasing DNA Mismatches

“…Because dTTP depletion is necessary for radiosensitization with FdUrd (Davis et al, 1995), we wanted to determine whether the inability of the MLH1 proficient HCT116 1-2 cells to undergo radiosensitization by FdUrd was due to lesser depletion of dTTP compared with the MLH1 inactivated HCT116 0-1 cells. Equitoxic concentrations (IC 50 ) of FdUrd produced similar changes in dTTP and other dNTPs in each cell line at 4 h, with ϳ40% reduction in dTTP and Ͼ50% reduction in dGTP with a concomitant Ն35% increase in dATP (Table 2).…”

“…Previous studies have determined that the radiosensitizing effect of FdUrd correlates with dTTP depletion but is not dependent on cytotoxicity (Davis et al, 1995). The ability of FdUrd to radiosensitize under minimally or noncytotoxic conditions was demonstrated to be sequence-dependent, occurring only when cells were exposed to drug before radiation (Bruso et al, 1990) and to correlate with the accumulation of cells in early to mid-S phase (Miller and Kinsella, 1992;McGinn et al, 1994).…”

“…Until recently, the most commonly proposed models for radiosensitization by antimetabolites included either an increase in DNA dsbs before or with radiation compared with radiation alone, or the inhibition of the repair of DNA dsbs after irradiation. Although FdUrd has been demonstrated to enhance cytotoxicity when administered with radiation by inhibiting the repair of radiation-induced DNA damage, radiosensitization can also occur in the absence of detectable DNA dsbs, suggesting another mechanism exists to explain the radiosensitizing effect of these drugs (Bruso et al, 1990;Davis et al, 1995). Our recent studies with another antimetabolite radiosensitizer, gemcitabine (dFdCyd), demonstrated that, at radiosensitizing concentrations, the dFdCyd-mediated depletion in dATP pools via inhibition of ribonucleotide reductase produced DNA mismatches that, if left unrepaired, resulted in radiosensitization (Flanagan et al, 2007).…”