Lymphocytes of BRCA1 and BRCA2 germ-line mutation carriers, with or without breast cancer, are not abnormally sensitive to the chromosome damaging effect of moderate folate deficiency

Abstract

Mutations in BRCA1 and BRCA2 genes may cause defective DNA repair and increase the risk for breast cancer. Folate deficiency is associated with increased breast cancer risk and induces chromosome abnormalities. We hypothesized that BRCA1 and BRCA2 germline mutation carriers are more sensitive to the genome damaging effect of folate deficiency compared with healthy non-carrier controls and that this sensitivity is further increased in those carriers who develop breast cancer. We tested these hypotheses in lymphocytes cultured in a medium containing 12 or 120 nM folic acid (FA) for 9 days and measured proliferative capacity and chromosomal instability using the cytokinesis-block micronucleus assay. BRCA1 and BRCA2 mutation carriers with or without breast cancer were not abnormally sensitive to FA deficiency-induced chromosome instability; however, BRCA2 mutation carriers had significantly reduced cell proliferation. FA deficiency reduced cell proliferation and increased micronucleus formation significantly, accounting for 45-59% and 70-75% of the variance in these parameters compared with 0.3-8.5% and 0.2-0.3% contributed by BRCA1 or BRCA2 mutation carrier status, respectively. The results of this study suggest that moderate folate deficiency has a stronger effect on chromosomal instability than BRCA1 or BRCA2 mutations found in breast cancer families.

Methionine-Dependence Phenotype in the de novo Pathway in BRCA1 and BRCA2 Mutation Carriers with and without Breast Cancer

“…The description and distribution of the BRCA1 and BRCA2 mutations among carriers with and without breast cancer have been published previously (23). Before presymptomatic testing, individuals not affected by breast cancer were at 25% to 50% risk of having the pathogenic mutation, which had been documented in another relative.…”

Cytokinesis-block micronucleus cytome assay

“…1-5) [3][4][5][6][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22] . This method is now also used to measure NPBs, a biomarker of dicentric chromosomes resulting from telomere end-fusions or DNA misrepair, and to measure NBUDs, a biomarker of gene amplification [20][21][22][23][24][25][26][27][28][29][30][31][32][33][34][35][36][37][38][39] . The significance of these developments and the concept of the CBMN assay as a ''cytome'' assay of chromosomal instability are explained in the following sections.…”

“…We have proposed that NPBs between nuclei in BN cells should be scored in the CBMN assay because they provide a measure of chromosome rearrangement, which is otherwise not achievable in this assay if only MNi are scored 3,22,23,[35][36][37][38][39] . NPBs occur when centromeres of dicentric chromosomes are pulled to opposite poles of the cell at anaphase.…”

“…In a series of studies on folic acid deficiency in long-term primary human lymphocyte cultures, we carefully quantified the interrelationship between MNi, NPBs and NBUDs in an attempt to validate the use of these biomarkers and to determine more comprehensively the impact of folic acid deficiency, within the physiological range, on various aspects of genomic stability [35][36][37][38] . Folic acid concentration correlated significantly (Po0.0001) and negatively (r ¼ À0.…”

“…Furthermore, determining nuclear division index (NDI) and proportion of cells undergoing necrosis and apoptosis provides important information on cytostatic and cytotoxic properties of the agent being examined that is relevant to the toxicity assessment. In human lymphocytes, the NDI also provides a measure of mitogen response, which is a useful biomarker of immune response in nutrition studies and may also be related to genotoxic exposure 3,23,[36][37][38]56 . The cytome approach in the CBMN Cyt assay is important because it allows genotoxic (MNi, NPBs and NBUDs in BN cells), cytotoxic (proportion of necrotic and apoptotic cells) and cytostatic (proportion and ratios of mono-, bi-and multinucleated cells, NDI) events to be captured within one assay.…”