Uracil misincorporation, DNA strand breaks, and gene amplification are associated with tumorigenic cell transformation in folate deficient/repleted Chinese hamster ovary cells

Clinical and experimental evidence has linked nutritional folic acid status to both anti- and procarcinogenic activity. Folate supplementation of norm al cells appears to have a protective effect; however, folate supplementati on of initiated cells may promote neoplastic progression. Given these consi derations, the present series of experiments examines alterations in DNA me tabolism and cumulative DNA lesions using an in vitro model of folate depri vation and repletion. DNA repair-deficient CHO-UV5 cells were cultured in H am's F-12 medium or in custom-prepared Ham's F-12 medium lacking in folic a cid, thymidine and hypoxanthine for a period of 18 days without cell passag e. The results indicated that progressive folate and nucleotide depletion l eads to a significant increase in the ratio of dUTP/dTTP and to the misinco rporation of uracil into DNA, These alterations were accompanied by growth inhibition, DNA strand breaks, abasic sites and phenotypic abnormalities. A fter 14 days in culture, there was significant increase in gene amplificati on potential in the chronically folate-deficient cells, but no significant increase in anchorage-independent growth or in neoplastic transformation. A cute folate repletion of the deficient cells was used as a proliferative st imulus under conditions of dNTP pool imbalance and multiple lesions in DNA. A further increase in gene amplification was accompanied by anchorage-inde pendent growth and neoplastic cell transformation as evidenced by aggressiv e tumor growth in Balb/c nu/nu mice. Using a sensitive in vitro model syste m, these results emphasize the essentiality of folic acid for de novo nucle otide synthesis and the integrity of the DNA. However, the in vivo relevanc e, especially in terms of tumorigenic potential, is not clear. (C) 1999 Pub lished by Elsevier Science Ireland Ltd. All rights reserved.