Patients with malignant diseases often have associated chromosomal abnormalities, but the factors which initiate or foster these alterations are poorly understood. The goal of this project is to investigate the possibility that folate deficiency, which is relatively common in this patient population, may play an important role in the development or progression of these diseases by promoting chromosomal aberrations including breaks, gaps, despiralization and increased sister chromatid exchanges (SCE). The first specific aim of this proposal is to characterize the chromosomal alterations induced by nutritional folate deficiency and their modulation by metabolites. Chinese hamster ovary cells and murine B16 melanoma cells will be grown in folate replete medium, folate deficient medium, or the latter medium supplemented with thymidine, hypoxanthine, or both nucleotides. The extent of folate deficiency will be evaluated by bioassay, the deoxyuridine (dU) suppression test, morphology and flow cytometry. Chromosomal alterations will be assessed with 5 assays: cytogenetics or flow cytometry for gain, loss or unequal exchange of DNA; detection of DNA strand breaks by alkaline filter elution and nick translation analysis; and mutation induction at specific loci (hprt and diphtheria toxin resistance). Our second specific aim is to measure the effects of folate deficiency on the mutagenicity of chemotherapeutic agents. Nutrient lack and drug effects might combine synergistically to contribute to the development of second malignancies in cancer patients. Cells will be incubated in control or folate deficient medium containing selected chemotherapeutic drugs. The effects of the drugs in control and deficient medium will be compared using the assays listed above, to look for additive or synergistic effects. The third specific aim is to determine the effect of folate status on the frequency of chemotherapy induced chromosomal alterations in patients with breast cancer. The incidence of chromosomal changes before and after chemotherapy will be determined and correlated with patient folate status as evaluated by a nutritional questionnaire, morphology, and blood folate levels. The frequency of chromosomal alterations will be assessed by karyotypic analysis, measurement of SCE, and quantification of somatic mutants at the hprt locus in lymphocytes. These studies will provide information about the incidence of folate deficiency, the degree of chromosomal instability, and the extent of synergy between folate deficiency and chemotherapeutic drugs in causing chromosomal damage.
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