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Lookup NU author(s): Professor Alan Calvert
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The properties are described of a mutant L1210 cell line (L1210:C15) with acquired resistance (>200-fold) to the thymidylate synthase (TS) inhibitor N10-propargyl-5,8-dideazafolic acid. TS was overproduced 45-fold and was accompanied by a small increase in the activity of dihydrofolate reductase (2.6-fold). Both the level of resistance and enzyme activities were maintained in drug-free medium (>300 generations). Failure of N10-propargyl-5,8-dideazafolic acid to suppress the [3H]-2'-deoxyuridine incorporation into the acid-precipitable material of the resistant line supported the evidence that TS overproduction was the mechanism of resistance; consequently the L1210:C15 cells were largely cross-resistant to another (but weaker) TS inhibitor, 5,8-dideazafolic acid. Minimal cross-resistance was observed to the dihydrofolate reductase inhibitors methotrexate and 5-methyl-5,8-dideazaaminopterin (5- and 2-fold, respectively). L1210 and L1210:C15 cells were, however, equally sensitive to 5-fluorodeoxyuridine (FdUrd), an unexpected finding since a metabolite, 5-fluorodeoxyuridine monophosphate, is a potent TS inhibitor; however, this cytotoxicity against the L1210:C15 cells was antagonized by coincubation with 5 µM folinic acid although folinic acid potentiated the cytotoxicity of FdUrd to the N10-propargyl-5,8-dideazafolic acid-sensitive L1210 line. Thymidine was much less effective as a FdUrd protecting agent in the L1210:C15 when compared with the L1210 cells; however, a combination of thymidine plus hypoxanthine was without any additional effect (compared with thymidine alone) against the sensitive line but effectively protected L1210:C15 cells such that the concentration of FdUrd necessary to reduce the cell count to 50% of control at 48 h was increased >11,000-fold. We propose that the elevated TS levels result in sequestration of the reduced-folate pool (as N5,10-methylene tetrahydrofolic acid) into the TS ternary complex with 5-fluoro-2'-deoxyuridine 5'-monophosphate. Despite "free" TS, the de novo synthesis of thymidylate and purines is inhibited by substrate depletion. The fact that folinic acid is able to reverse the inhibition of [3H]-2'-deoxyuridine incorporation by FdUrd into the resistant cells supports this hypothesis.
Author(s): Jackman AL, Alison DL, Calvert AH, Harrap KR
Publication type: Article
Publication status: Published
Journal: Cancer Research
Year: 1986
Volume: 46
Issue: 6
Pages: 2810-2815
Print publication date: 01/06/1986
ISSN (print): 0008-5472
ISSN (electronic): 1538-7445
URL: http://cancerres.aacrjournals.org/cgi/content/abstract/46/6/2810
