Haploinsufficiency for BRCA1 is associated with normal levels of DNA nucleotide excision repair in breast tissue and blood lymphocytes

Abstract

Background: Screening mammography has had a positive impact on breast cancer mortality but cannot detect all breast tumors. In a small study, we confirmed that low power magnetic resonance imaging (MRI) could identify mammographically undetectable tumors by applying it to a high risk population. Tumors detected by this new technology could have unique etiologies and/or presentations, and may represent an increasing proportion of clinical practice as new screening methods are validated and applied A very important aspect of this etiology is genomic instability, which is associated with the loss of activity of the breast cancer-predisposing genes BRCA1 and BRCA2. In sporadic breast cancer, however, there is evidence for the involvement of a different pathway of DNA repair, nucleotide excision repair (NER), which remediates lesions that cause a distortion of the DNA helix, including DNA cross-links.

Case presentation: We describe a breast cancer patient with a mammographically undetectable stage I tumor identified in our MRI screening study. She was originally considered to be at high risk due to the familial occurrence of breast and other types of cancer, and after diagnosis was confirmed as a carrier of a Q1200X mutation in the BRCA1 gene. In vitro analysis of her normal breast tissue showed no differences in growth rate or differentiation potential from disease-free controls. Analysis of cultured blood lymphocyte and breast epithelial cell samples with the unscheduled DNA synthesis assay (UDS) revealed no deficiency in nucleotide excision repair (NER).

Conclusion: As new breast cancer screening methods become available and cost effective, patients such as this one will constitute an increasing proportion of the incident population, so it is important to determine whether they differ from current patients in any clinically important ways. Despite her status as a BRCA1 mutation carrier, and her mammographically dense breast tissue, we did not find increased cell proliferation or deficient differentiation potential in her breast epithelial cells, which might have contributed to her cancer susceptibility. Although NER deficiency has been demonstrated repeatedly in blood samples from sporadic breast cancer patients, analysis of blood cultured lymphocytes and breast epithelial cells for this patient proves definitively that heterozygosity for inactivation of BRCA1 does not intrinsically confer this type of genetic instability. These data suggest that the mechanism of genomic instability driving the carcinogenic process may be fundamentally different in hereditary and sporadic breast cancer, resulting in different genotoxic susceptibilities, oncogene mutations, and a different molecular pathogenesis.

Figure 1

Pedigree of the patient (indicated by arrow). She, one maternal aunt and one maternal cousin had breast cancer diagnosed at 36, 44 and 41 years old, respectively, as indicated by the half-filled symbols, and her aunt died of the disease. Her cousin underwent lumpectomy followed by chemotherapy, radiotherapy and is presently on tamoxifen. Her mother had breast cancer in both breasts, diagnosed at ages 41 and 42, as indicated by the completely filled symbol. She underwent bilateral mastectomy and hysterectomy followed by chemotherapy and radiotherapy and died of the disease at age 44. A second maternal aunt was diagnosed with colon cancer at age 52 (light half-filled symbol) and breast cancer at age 55 (dark half-filled symbol). Based on this pattern of familial cancer the patient was considered to be at high risk of developing breast cancer and was entered into the low power MRI screening validation and feasibility study. Following her diagnosis, she was confirmed as carrying a Q1200X mutation in the BRCA1 gene.

Figure 2

Ultrasound of the MRI-detected lesion. Following MRI, the patient was scheduled for ultrasound to identify the questionable lesions seen on MRI for possible core biopsy. Under ultrasound the lesion of concern was identified and biopsied at the 1:00 location in the left breast. Additionally, one lesion seen by MRI in the right breast at the 4:00 location was identified and biopsied.

Figure 3

Micrographs of the non-diseased primary human mammary epithelial cultures (HMEC) from the BRCA1 mutation carrier. A) Contralateral breast – A cluster of epithelial cells called a mammosphere is shown on the left center of the image sitting on a field of fibroblasts. B) Ipsilateral breast – The original fresh tissue block from which this culture was derived was located 4 cm from the infiltrating ductal carcinoma. The structure shown is a cluster of rounded epithelial cells manifesting a column configuration called "pre-ductal linearization". Both images were captured under Differential Interference Contrast (DIC) optics on a Zeiss Axiovert 100 microscope at a total of 140x magnification.

Figure 4

Comparison of the NER capacity of a PBL sample from our BRCA1 mutation carrier patient with those of a population of disease-free controls. The dark horizontal line indicates the average for the normal population, while the dotted lines indicate upper limits for residual NER activity in patients with the hereditary NER deficiency disease XP (0.50) and the cut-off established in our breast tissue study that identified tumors with high sensitivity and specificity (0.70).

Figure 5

Comparison of the NER capacities of two samples of normal breast epithelium from our BRCA1 mutation carrier patient with those of a population of disease-free controls who underwent breast reduction mammoplasty. The dark horizontal line indicates the average for the normal population of breast reduction epithelium (BRE), while the dotted lines indicate upper limits for residual NER activity in patients with the hereditary NER deficiency disease XP (0.50) and the cut-off established in our breast tissue study that identified tumors with high sensitivity and specificity (0.70). The patient sample on the left was derived from the ipsilateral (left) breast, while the sample on the right was from the contralateral (right) breast.