Skip to main content
Log in

Comparative cytotoxicity of folate-based inhibitors of thymidylate synthase and 5-fluorouracil ± leucovorin in MGH-U1 cells

  • Original Articles
  • 5-Fluorouracil, Inhibitors, Thymidylate Synthase
  • Published:
Cancer Chemotherapy and Pharmacology Aims and scope Submit manuscript

Abstract

Thymidylate synthase (TS) is a critical enzyme in the synthesis of DNA and an important target for cancer chemotherapy. 5-Fluorouracil (5FU) combined with leucovorin (LV) has been used to inhibit TS, and inhibition is dependent on the formation of a ternary complex between a folate cofactor, TS, and 5-fluorodeoxyuridine monophosphate (FdUMP), a metabolite of FU. The folate-based TS inhibitors CB3717, its analogs, and BW1843U89 have been synthesized as specific inhibitors of TS that do not require activation or the presence of a cofactor. We have compared the cytotoxicity of 5FU ± LV with that of these folate-based TS inhibitors in human bladder cancer MGH-U1 cells using a colony-forming assay. After a 6-h exposure, FU+LV, CB3717, dCB3717, or C2 methyl dideazafolate analogs demonstrated similar cytotoxic potency that was 0.96 to 2.9 times that of 5FU alone. A 24-h exposure did not increase the potency of 5FU+LV relative to 5FU alone, but there was a marked increase in the cytotoxicity of the dideazafolates as compared with 5FU+LV. Similarly, BW1843U89 was more cytotoxic than 5FU+LV. This was reflected in a 3.2- to 1333-fold decrease in the 50% inhibitory concentration (IC50). Simultaneous exposure to LV and thymidine (TdR) protected MGH-U1 cells from the cytotoxicity of CB3717, its analogs, and BW1843U89. We conclude that (a) the folate-based TS inhibitors are more potent than 5FU+LV after a 24-h exposure, (b) protection by LV and TdR indicates that TS inhibition is the primary site of action, and (c) BW1843U89 is more potent than D1694 in MGH-U1 cells.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Bisset GMF, Pawelczak K, Jackman AL, Calvert AH, Hughes LR (1992) Syntheses and thymidylate synthase inhibitory activity of the poly-gamma-glutamyl conjugates ofN-<5-<N-(3,4-dihydro-2-methyl-4-oxoquinazolin-6-ylmethyl)-N-methylamino>-2-thenoyl>-l-glutamic acid (ICI D1694) and other quinazoline antifolates. J Med Chem 35: 859

    Google Scholar 

  2. Chen T-L, Erlichman C (1992) Biochemical modulation of 5-fluorouracil with or without leucovorin by low dose of brequinar in MGH-U1 cells. Cancer Chemother Pharmacol 30: 370

    Google Scholar 

  3. Duch DS, Banks S, Inderjit KD, Dickerson SH, Ferone R, Heath LS, Humphreys J, Knick V, Pendergast W, Singer S, Smith GK, Waters K, Wilson HR (1993) Biochemical and cellular pharmacology of 1843U89, a novel benzoquinazoline inhibitor of thymidylate synthase. Cancer Res 53: 810

    Google Scholar 

  4. Erlichman C, Wu A (1991) Effects of 5-fluorouracil and leucovorin in spheroids — a model for solid tumours. Anticancer Res 11: 671

    Google Scholar 

  5. Erlichman C, Fine S, Wong A, Elhakim T (1988) A randomized trial of fluorouracil and folinic acid in patients with metastatic colorectal carcinoma. J Clin Oncol 6: 469

    Google Scholar 

  6. Evans RM, Laskin JD, Hakala MT (1981) Effect of excess folates and deoxyinosine on the activity site of action of 5-fluorouracil. Cancer Res 41: 3288

    Google Scholar 

  7. Houghton JA, Maroda SJ, Phillips JO, Houghton PJ (1981) Biochemical determinants of responsiveness to 5-fluorouracil and its derivatives in xenografts of human colorectal adenocarcinomas in mice. Cancer Res 41: 144

    Google Scholar 

  8. Jackman AL, Taylor GA, Moran RG, Bishop JAM, Bisset G, Pawelczak K, Balmanno K, Hughes LR, Calvert AH (1988) Biologic properties of 2-desamino-2-substituted-5,8-deazafolates that inhibit thymidylate synthase (TS). Proc Am Assoc Cancer Res 29: 287

    Google Scholar 

  9. Jackman AL, Taylor GA, Gibson W, Kimbell R, Brown M, Calvert AH, Judson IR, Hughes LR (1991) ICI-D1694, a quinazoline antifolate thymidylate synthase inhibitor that is a potent inhibitor of L1210 tumor cell growth in vitro and in vivo — a new agent for clinical study. Cancer Res 51: 5579

    Google Scholar 

  10. Jansen G, Schornagel JH, Westerhof GR, Rijksen G, Newell DR, Jackman AL (1990) Multiple membrane transport systems for the uptake of folate-based thymidylate synthase inhibitors. Cancer Res 50: 7544

    Google Scholar 

  11. Jones TR, Calvert AH, Jackman AL, Brown SJ, Jones M, Harrap KR (1981) A potent antitumor quinazoline inhibitor of thymidylate synthetase. Synthesis, biologic properties and therapeutic results in mice. Eur J Cancer 17: 11

    Google Scholar 

  12. Jones TR, Calvert AH, Jackman AL, Eakin MA, Smithers MJ, Beteridge RF, Newell DR, Hayter AJ, Stocker A, Harland SJ, Davies LC, Harrap KR (1985) Quinazoline antifolates inhibiting thymidylate synthase: variations of the N10 substituent. J Med Chem 28: 1468

    Google Scholar 

  13. Jones TR, Smithers MJ, Betteridge RF, Taylor MA, Jackman AL, Calvert AH, Davies LC, Harrap KR (1986) Quinazoline antifolates inhibiting thymidylate synthase: variation of the amino acid. J Med Chem 29: 1114

    Google Scholar 

  14. Jones TR, Betteridge RF, Neidle S, Jackman AL, Calvert AH (1989) Quinazoline antifolates inhibiting thymidylate synthase: computer modelling of the N10 substituent. Anticancer Drug Des 3: 243

    Google Scholar 

  15. Jones TR, Thornton TJ, Flinn A, Jackman AL, Newell DR, Calvert AH (1989) Quinazoline antifolates inhibiting thymidylate synthase: 2-desamino derivatives with enhanced solubility and potency. J Med Chem 32: 847

    Google Scholar 

  16. Keyomarsi K, Moran RG (1988) Mechanism of the cytotoxic synergism of fluoropyrimidines and folinic acid in mouse leukemic cells. J Biol Chem 263: 14402

    Google Scholar 

  17. Marsham PR, Hughes LR, Hayter AJ et al (1990) Quinazoline antifolate thymidylate synthase inhibitors: potent cytotoxic agent containing heterocyclic isosteres of the para-amino benzoate unit. In: Curtius H-C, Ghisla S, Blau N, (ed) Chemistry and biology of pteridines 1989. Walter de Gruyter, New York, pp 1048–1051

    Google Scholar 

  18. Marsham PR, Hughes LR, Jackman AL, Hayter AJ, Oldfield J, Wardleworth JM, Bishop JA, O'Connor BM, Calvert AH (1991) Quinazoline antifolate thymidylate synthase inhibitors: heterocyclic benzoyl ring modifications. J Med Chem 34: 1594

    Google Scholar 

  19. Pawelczak K, Jones TR, Kempny M, Jackman AL, Newell DR, Krzyzanowski L, Rzeszotarska B (1989) Quinazoline antifolates inhibiting thymidylate synthase: synthesis of four oligo(l-gamma-glutamyl) conjugates ofN10-propargyl-5,8-dideazafolic acid and their enzyme inhibition. J Med Chem 32: 160

    Google Scholar 

  20. Pendergast W, Dickerson SH, Inderjit K, Ferone R, Duch DS, Smith GK (1993) Benzoquinazoline inhibitors of thymidylate synthase: effect on cytotoxicity and thymidylate synthase activity of variation of sidechain structure and C3 substitution. Proc Am Assoc Cancer Res 33: 407

    Google Scholar 

  21. Piedbois P, Buyse M, Rustum Y, Machover D, Erlichman C, Carlson RW, Valone F, Labianca R, Doroshow JH, Petrelli N (1992) Modulation of fluorouracil by leucovorin in patients with advanced colorectal cancer — evidence in terms of response rate. J Clin Oncol 10: 896

    Google Scholar 

  22. Pinedo HM, Peters GFJ (1988) Fluorouracil: biochemistry and pharmacology. J Clin Oncol 6: 1653

    Google Scholar 

  23. Poon MA, O'Connell MJ, Wieand HS, Krook JE, Gerstner JB, Tschetter LK, Levitt R, Kardinal CG, Mailliard JA (1991) Biochemical modulation of fluorouracil with leucovorin — confirmatory evidence of improved therapeutic efficacy in advanced colorectal cancer. J Clin Oncol 9: 1967

    Google Scholar 

  24. Sikora E, Jackman AL, Newell DR, Calvert AH (1988) Formation and retention and biological activity ofN10-propargyl-5,8-dideazafolic acid (CB3717) polyglutamates in L1210 cells in vitro. Biochem Pharmacol 37: 4047

    Google Scholar 

  25. Spears CP, Gustavsson BG, Berne M, Frosing R, Bernstein L, Hayes AA (1988) Mechanisms of innate resistance to thymidylate synthase inhibition after 5-fluorouracil. Cancer Res 48: 5894

    Google Scholar 

  26. Westerhof GR, Jansen G, Vanemmerik N, Kathmann I, Rijksen G, Jackman AL, Schornagel JH (1991) Membrane transport of natural folates and antifolate compounds in murine L1210 leukemia cells — role of carrier-mediated and receptor-mediated transport systems. Cancer Res 51: 5507

    Google Scholar 

  27. Yin MB, Zakrezewski F, Hakala MT (1983) Relationship of cellular folate cofactor pools to the activity of 5-fluorouracil. Mol Pharmacol 23: 190

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Additional information

This study was supported by a grant from the National Cancer Institute of Canada

Rights and permissions

Reprints and permissions

About this article

Cite this article

Erlichman, C., Mitrovski, B. Comparative cytotoxicity of folate-based inhibitors of thymidylate synthase and 5-fluorouracil ± leucovorin in MGH-U1 cells. Cancer Chemother. Pharmacol. 34, 51–56 (1994). https://doi.org/10.1007/BF00686111

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00686111

Keywords

Navigation