Fas-dependent and -independent mechanisms of cell death following DNA damage in human colon carcinoma cells

Istvan Petak, David M. Tillman, Franklin G. Harwood, Rudolf Mihalik, Janet A. Houghton

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38 Citations (Scopus)


In thymidylate synthase-deficient (TS-) colon carcinoma cells, thymineless death is mediated via Fas/Fas ligand (FasL) interactions after thymidine deprivation and inhibited by the Fas-inhibitory monoclonal antibody NOK-1. The objective of the study was to elucidate whether other modes of DNA damage induced by doxorubicin, topotecan, and etoposide (VP-16) could elicit a similar cytotoxic response in TS- cells by signaling via the Fas death receptor. After a 72-h drug exposure, a loss in clonogenic survival that was not prevented by NOK-1 was induced by each agent in the absence of acute apoptosis, yielding IC50 values of 5 (doxorubicin), 10 (topotecan), and 150 nm (VP-16). Furthermore, TS- cell clones selected for resistance to Fas- mediated apoptosis (CH-11) were cross-resistant to the induction of thymineless death after thymidine deprivation but were not cross-resistant to doxorubicin, topotecan, or VP-16. A close correlation was found between acute induction of apoptosis (24 h) and up-regulated expression of FasL at high concentrations of each of the three agents (0.3-3 μM doxorubicin, 0.3-3 μM topotecan, and 10-90 μM VP-16), which was caspase dependent but Fas independent. At all drug concentrations, cell cycle distribution analyses demonstrated marked accumulation of cells in the G2-M phase. At nanomolar drug concentrations, prolonged arrest of TS cells in G2-M phase resulted in the upregulation of FasL expression and the delayed appearance of apoptotic cells (6 days), which could also be inhibited by the general caspase inhibitor Z-VAD-FMK, but not by NOK-1 or Fas-Fc. In clonogenic assays, Z-VAD- FMK did not rescue cells treated with VP-16 in contrast to treatment with CH- 11 or thymineless stress, suggesting an irreversible commitment to cell death in G2-M phase. Expression of FasL at all drug concentrations appeared to be unrelated to the mechanism of drug-induced apoptosis. This was in contrast to the Fas-dependent regulation of thymineless death, which could be inhibited by blocking Fas/FasL interactions.

Original languageEnglish
Pages (from-to)2643-2650
Number of pages8
JournalCancer Research
Issue number10
Publication statusPublished - May 15 2000

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

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