Vol. 53, Issue 2, 252-261, February 1998

Modulation of Fluorouracil Cytotoxicity by Interferon- and -

A.-S.A. Ismail,¹ C. J. Van Groeningen,² A. Hardcastle, Q.-F. Ren, G. W. Aherne, F. Geoffroy, C. J. Allegra, and J. L. Grem.

National Cancer Institute-Navy Medical Oncology Branch (A.-S.A.I., C.J.V.G., Q.-F.R., F.G., C.J.A., J.L.G.), National Naval Medical Center, Bethesda, Maryland 20889, and CRC Centre for Cancer Therapeutics at The Institute of Cancer Research (A.H., G.W.A.), Belmont, Sutton, Surrey SM2 5NG, UK

Because interferons (IFN)- and - individually have increased fluorouracil (FUra) cytotoxicity in several in vitro models, we studied the effects of FUra combined with IFN- +  in HT29 colon cancer cells. A 96-hr exposure to IFN- (500 units/ml) plus IFN- (10 units/ml) and a 72-hr exposure to 0.25-1 µM FUra (hr 24-96) inhibited cell growth and colony formation in an additive or more-than-additive fashion. When cells were exposed to IFN- +  and FUra, free FdUMP levels became detectable, whereas [³H]FUra-RNA incorporation decreased. Exposure to IFN- + , FUra, or the combination decreased dTTP pools to 58%, 43%, and 17% of control, respectively. A marked increase in the dATP to dTTP ratio was seen with FUra with or without IFN- + . Thymidylate synthase catalytic activity was reduced to 28% and 24% of control with FUra with or without IFN- + , suggesting that the enhanced dTTP depletion must be due to another mechanism. FUra-mediated thymidylate synthase inhibition was accompanied by a 124-fold increase in total deoxyuridylate immunoreactivity and a 31-fold increase in dUTP pools, but the addition of IFN- +  attenuated the accumulation. Treatment with IFN- +  and FUra individually interfered with nascent DNA chain elongation, whereas the three-drug combination produced the most striking effects. IFN- +  plus FUra produced the greatest amount of single-strand breaks in nascent DNA and dramatically decreased net DNA synthesis. IFN- +  with or without FUra produced double-strand breaks in parental DNA. These results suggest that dTTP depletion, dATP/dTTP imbalance, pronounced inhibition of DNA synthesis, and damage to nascent and parental DNA contribute to the enhanced cytotoxicity with the triple combination.