Quantification of BRCA1 protein in sporadic breast carcinoma with or without loss of heterozygosity of the BRCA1 gene

Abstract

In sporadic breast cancer, no mutations of the BRCA1 gene have been reported so far, whereas BRCA1 mRNA is markedly decreased in invasive breast cancer. To elucidate the contribution of the BRCA1 gene in sporadic breast cancer, we quantified the BRCA1 protein, using [ 125 I] labeling of wholecell proteins, lentil-lectin affinity chromatography, immunoprecitation by anti-BRCA1 antibodies (C-20, D-20, I-20 and K-18), purification of the immune complex by protein A affinity chromatography and chromato-focusing. As loss of 1 allele may lead to a decreased expression of the gene, 10 tumors were previously checked for loss of heterozygosity (LOH) of the BRCA1 gene, using 3 intragenic microsatellite markers. Our results indicated that the BRCA1 gene product was decreased in the 4 tumors with LOH compared with matched normal breast tissues. Reduced amounts of BRCA1 protein were also detected in 3 of 6 tumors without LOH. Our quantitative method allowed us to demonstrate that the BRCA1 protein level was decreased in sporadic invasive breast carcinomas with or without LOH of the BRCA1 gene, implying multiple mechanisms of BRCA1 expression downregulation in these tumors. Our data suggest that the amount of BRCA1 protein present may play an important role in human sporadic breast carcinoma. Int. J. Cancer 80:823-826, 1999. Wiley-Liss, Inc.While familial breast cancer or familial breast/ovarian cancer often accompanies germ-line mutations of the BRCA1 gene (Miki et al., 1994), there is no evidence of BRCA1 mutations in sporadic breast cancer and BRCA1 mutations in sporadic ovarian cancer are rare (Futreal et al., 1994, Merajver et al., 1995. However BRCA1 gene transcription is markedly reduced in invasive breast tumors relative to normal breast epithelium, ductal carcinoma in situ (Thompson et al., 1995) and breast cancer cell lines (Ribieras et al., 1997), suggesting a role for BRCA1 in sporadic tumors. So far, BRCA1 gene expression in tumor tissues and cell lines has been monitored mainly at the mRNA level, by either Northern blot (Marquis et al., 1995), ribonuclease protection assay (Thompson et al., 1995) or competitive RT-PCR (Ribieras et al., 1997). In this study, we evaluated the relative amount of BRCA1 protein in ductal invasive carcinomas, using a strategy of biochemical purification previously devised in our laboratory for measureing HLA-DR antigen amounts (Bernard et al., 1984(Bernard et al., , 1985 and adapted for the BRCA1 protein (Bernard-Gallon et al., 1997).As loss of 1 allele might lead to reduced expression of the corresponding gene, we searched for loss of heterozygosity (LOH) of the BRCA1 gene, using 3 intragenic microsatellite markers. The BRCA1 protein was then quantified in the 2 groups of tumors, with and without LOH, and compared in tumors and normal matched tissues. Our results show that the amount of BRCA1 protein was decreased in all tumors with LOH as well as in a majority of ductal invasive carcinomas without LOH. MATERIAL AND METHODS Patient samplesTen sporadic invasive ductal breast carcinomas...

MTA1-mediated transcriptional repression of BRCA1 tumor suppressor gene

“…On the other hand, the remaining 90% of breast cancers occur sporadically in the absence of any apparent genetic predisposition. In these nonfamilial cases, tumorigenesis is likely not due to mutation but rather to a drastic decrease of BRCA1 mRNA or protein levels (Magdinier et al, 1998;Rio et al, 1999). Multiple studies have shown a correlation between loss of BRCA1 expression and increased metastatic disease.…”

BRCA1 in hormonal carcinogenesis: basic and clinical research

“…Recent studies indicate that BRCA1 mRNA and protein are absent or significantly reduced in 30-40% of sporadic breast cancer cases (Ozcelik et al 1998, Taylor et al 1998, Rio et al 1999, Wilson et al 1999. Decreased BRCA1 expression may be due to hypermethylation of the BRCA1 promoter on CpG islands and/or loss of one of the BRCA1 alleles (Rice et al 1998, Esteller et al 2000, Staff et al 2003.…”

Genetic instability in the RAD51 and BRCA1 regions in breast cancer

“…Recent studies indicated that allelic loss in the aforementioned regions might be associated with clinicopathological features of breast cancer. Statistically significant differences between breast tumors with and without LOH in the RAD51 and BRCA1 regions have been found with respect to estrogen receptor content, progesterone receptor content, higher grade, and stage [56,65,66]. Johnson et al [62] detected allelic loss of BRCA1 with higher frequency in women under 36 compared to postmenopausal patients.…”