The modes of action of 5-fluoro-2'deoxyuridine(FdUrd) and 5-fluoro-2'-deoxycytidine(FdCyd) were studied in PHA- stimulated lymphocytes from normal volunteer donors and a fragile X patient. In both cell types, FdUrd and FdCyd inhibited cell proliferation at concentrations of 3x10-8M. Thymidylate synthetase was identified as the decisive target for the action of both FdUrd and FdCyd, as judged from the following observations: First, addition of thymidine to the culture medium was able to counteract both FdUrd and FdCyd toxicities, whereas addition of dCyd had no observable effect. Second, inhibition of the in situ thymidylate synthetase activity measured as an increase in the level of [3H]-dUrd into newly synthesized DNA coincided with the inhibition of the cell proliferation. The effects of FdUrd and FdCyd on the in vitro expression of the fragile site Xq27 of the fragile X chromosomes was shown to the based on the depletion of the intracellular pool of thymidine-5'- monophosphate(dTMP), as judged from the following observations: First, both the FdUrd- and FdCyd- dependent induction of Xq27 coincided with the antiproliferative effects of the respective fluoropyrimidines. Second, addition of thymidine (dThd) to the culture medium both prevented the expression of site Xq27 and neutralized the cytotoxicity of FdUrd and of FdCyd. On the basis of these findings we provide further evidence for the concept that fragile X site is located in an AT-rich region.
|Number of pages||6|
|Publication status||Published - 20 Jan 1988|
- fragile X chromosome