Genetic Variation in Thymidylate Synthase Confers Resistance to 5-Fluorodeoxyuridine
The human colorectal tumor cell line HCT 116 was resistant to the cytotoxic effects of 5-fluorodeoxyuridine (FdUrd). The response to FdUrd was increased only slightly by the presence of 10 μM folinic acid (CF). HCT 116 formed FdUMP and CH2H4PteGlu polyglutamates after exposure to FdUrd and CF. The sensitivity to FdUrd correlated well with the extent of TS inhibition. The role of TS in the resistance of the cells to FdUrd was examined. HCT 116 expresses two TS enzymes, which differ in pI. The more basic TS has been detected in only HCT 116 cells. The other TS is identical in pI to the enzymes detected in other human cells. The variant TS differs from the common by His replacement of Tyr at residue 33. The variant TS exhibited a 3-fold lower affinity for FdUMP than the common TS. The enzymes co-expressed in HCT 116 exhibited an FdUMP binding constant similar to that of the variant TS. TS-deficient cells were transfected with cDNAs encoding the two TS polypeptides. Transfectants expressing the variant TS were more resistant to FdUrd cytotoxicity than cells expressing the common TS. Thus, the structural variation in TS reduced enzyme affinity for FdUMP and conferred resistance to FdUrd.
KeywordsOncol Methotrexate Folate Charcoal Lactobacillus
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- 7.Peters, G.J., van Groeningen, C.J., van der Wilt, C.L., Meijer, S., Smid, K., Laurensse, E., and Pinedo, H.M. Time course of inhibition of thymidylate synthase in patients treated with fluorouracil and leucovorin. Sem. Oncol. 19: 26–35 (1992).Google Scholar
- 13.Danenberg, P.V. and Bapat, A.R. Thymidylate synthases isolated from human colon tumors have varied affinity towards 5-fluoro-2′-deoxyuridylate. Reg. Cancer Treat. 2: 1–4 (1989).Google Scholar
- 16.Nussbaum, R.L., Walmsley, R.M., Lesko, J.G., Airhart, S.D., and Ledbetter, D.H. Thymidylate synthase-deficient Chinese hamster cells: a selection system for human chromosome 18 and experimental system for the study of thymidylate synthase regulation and fragile X expression. Am. J. Hum. Genet. 37: 1192–1205 (1985).PubMedGoogle Scholar
- 18.Moran, R.G., Spears, C.P., and Heidelberger, C. Biochemical determinants of tumor sensitivity to 5-fluorouracil: ultrasensitive methods for the determination of 5-fluoro-2′-deoxyuridylate, 2′-deoxyuridylate, and thymidylate synthetase. Proc. Natl. Acad. Sci. USA 76: 1456–1460 (1979).PubMedCrossRefGoogle Scholar
- 21.Banerjee, C.K., Bennett, L.L., Jr., Brockman, R.W., Sani, B.P., and Temple, C., Jr. A convenient procedure for purification of thymidylate synthase from L1210 cells. Anal. Biochem. 72: 248–254 (1982).Google Scholar
- 22.Spears, C.P., Shahinian, A.H., Moran, R.G., Heidelberger, C., and Corbett, T.H. In vivo kinetics of thymidylate synthetase inhibition in 5-fluorouracil-sensitive and — resistant murine colon adenocarcinomas. Cancer Res. 42: 450–456 (1982).Google Scholar
- 27.Berger, S.H. Biochemical factors which influence the action of 5-fluoropyrimidines on thymidylate synthetase. Ph.D. Thesis, State University of New York at Buffalo (1982).Google Scholar