Heterogenic Loss of the Wild-Type BRCA Allele in Human Breast Tumorigenesis
ABSTRACT For individuals genetically predisposed to breast and ovarian cancer through inheritance of a mutant BRCA allele, somatic loss of heterozygosity affecting the wild-type allele is considered obligatory for cancer initiation and/or progression. However, several lines of evidence suggest that phenotypic effects may result from BRCA haploinsufficiency.
Archival fixed and embedded tissue specimens from women with germ line deleterious mutations in BRCA1 or BRCA2 were identified. After pathologic review, focal areas of normal breast epithelium, atypical ductal hyperplasia, ductal carcinoma-in-situ, and invasive ductal carcinoma were identified from 14 BRCA1-linked and 9 BRCA2-linked breast cancers. Ten BRCA-linked prophylactic mastectomy specimens and 12 BRCA-linked invasive ovarian carcinomas were also studied. Laser catapult microdissection was used to isolate cells from the various pathologic lesions and corresponding normal tissues. After DNA isolation, real-time polymerase chain reaction assays were used to quantitate the proportion of wild-type to mutant BRCA alleles in each tissue sample.
Quantitative allelotyping of microdissected cells revealed a high level of heterogeneity in loss of heterozygosity within and between preinvasive lesions and invasive cancers from BRCA1 and BRCA2 heterozygotes with breast cancer. In contrast, all BRCA-associated ovarian cancers displayed complete loss of the wild-type BRCA allele.
These data suggest that loss of the wild-type BRCA allele is not required for BRCA-linked breast tumorigenesis, which would have important implications for the genetic mechanism of BRCA tumor suppression and for the clinical management of this patient population.
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- "Initial studies of LOH in the breast tumours of BRCA1 carriers reported a rate of 75% (75/101) of LOH of the wild type allele -. More recently, quantitative allelotyping has demonstrated a significant degree of variability in the extent and direction of LOH in breast tumours . In some cases, normal tissue shows LOH of the wild type BRCA1 allele, and tumor tissue shows loss of the mutant allele. "
ABSTRACT: The assessment of BRCA1 and BRCA2 coding sequences to identify pathogenic mutations associated with inherited breast/ovarian cancer syndrome has provided a method to identify high-risk individuals, allowing them to seek preventative treatments and strategies. However, the current test is expensive, and cannot differentiate between pathogenic variants and those that may be benign. Focusing only on one of the two BRCA partners, we have developed a biological assay for haploinsufficiency of BRCA1. Using a series of EBV-transformed cell lines, we explored gene expression patterns in cells that were BRCA1 wildtype compared to those that carried (heterozygous) BRCA1 pathogenic mutations. We identified a subset of 43 genes whose combined expression pattern is a sensitive predictor of BRCA1 status. The gene set was disproportionately made up of genes involved in cellular differentiation, lending credence to the hypothesis that single copy loss of BRCA1 function may impact differentiation, rendering cells more susceptible to undergoing malignant processes.PLoS ONE 06/2014; 9(6):e100068. DOI:10.1371/journal.pone.0100068 · 3.23 Impact Factor
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- "There is mounting evidence that heterozygosity for BRCA2 is sufficient for human carcinogenesis in tissues other than the pancreas, with the attendant implications for therapy using PARP1i. Lack of LOH has been documented using different methods in up to ∼30% of breast cancers (King et al., 2007), by NGS in ∼25% high-grade serous ovarian carcinomas (Cancer Genome Atlas, 2011), and in up to 55% of prostatic cancers associated with BRCA2 mutation carriers (Willems-Jones et al., 2012). Thus, it will be essential in future clinical trial designs to test for LOH in tumour samples to help stratify likely responders to PARP1i or similar targeted therapies. "
ABSTRACT: Cancer is unique amongst human diseases in that its cellular manifestations arise and evolve through the acquisition of somatic alterations in the genome. In particular, instability in the number and structure of chromosomes is a near-universal feature of the genomic alterations associated with epithelial cancers, and is triggered by the inactivation of tumour suppressor mechanisms that preserve chromosome integrity in normal cells. The nature of these mechanisms, and how their inactivation promotes carcinogenesis, remains enigmatic. I will review recent work from our laboratory on the tumour suppressor BRCA2 that addresses these issues, focusing on new insights into cancer pathogenesis and therapy that are emerging from improved understanding of the molecular basis of chromosomal instability in BRCA2-deficient cancer cells.Moleculer Cells 02/2014; 37(2):95-9. DOI:10.14348/molcells.2014.2346 · 2.09 Impact Factor
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- "However, loss of the mutant allele is not an unprecedented finding even for a bona fide breast cancer predisposition gene. In a recent study performed by King et al., 23 BRCA-linked breast tumors and 10 BRCA-linked prophylactic mastectomy (PM) specimens were analyzed for loss of the wild-type allele . No loss of heterozygosity (LOH) or LOH involving the mutant allele was observed in a substantial fraction of pre-invasive and invasive breast carcinomas. "
ABSTRACT: SLX4 encodes a DNA repair protein that regulates three structure-specific endonucleases and is necessary for resistance to DNA crosslinking agents, topoisomerase I and poly (ADP-ribose) polymerase (PARP) inhibitors. Recent studies have reported mutations in SLX4 in a new subtype of Fanconi anemia (FA), FA-P. Monoallelic defects in several FA genes are known to confer susceptibility to breast and ovarian cancers. To determine if SLX4 is involved in breast cancer susceptibility, we sequenced the entire SLX4 coding region in 738 (270 Jewish and 468 non-Jewish) breast cancer patients with 2 or more family members affected by breast cancer and no known BRCA1 or BRCA2 mutations. We found a novel nonsense (c.2469G>A, p.W823*) mutation in one patient. In addition, we also found 51 missense variants [13 novel, 23 rare (MAF<0.1%), and 15 common (MAF>1%)], of which 22 (5 novel and 17 rare) were predicted to be damaging by Polyphen2 (score = 0.65-1). We performed functional complementation studies using p.W823* and 5 SLX4 variants (4 novel and 1 rare) cDNAs in a human SLX4-null fibroblast cell line, RA3331. While wild type SLX4 and all the other variants fully rescued the sensitivity to mitomycin C (MMC), campthothecin (CPT), and PARP inhibitor (Olaparib) the p.W823* SLX4 mutant failed to do so. Loss-of-function mutations in SLX4 may contribute to the development of breast cancer in very rare cases.PLoS ONE 06/2013; 8(6):e66961. DOI:10.1371/journal.pone.0066961 · 3.23 Impact Factor