Sporadic breast cancer in young women: prevalence of loss of heterozygosity at p53, BRCA1 and BRCA2

Previous studies have shown that breast cancers have more aggressive pathologic features in young women. In order to examine genetic alterations associated with early-onset breast cancer, 31 patients with no known family history, aged 26-35 years at diagnosis, were examined for loss of heterozygosity (LOH) at 3 key chromosomal intervals: 17p (p53), 17q 21 (BRCA1) and 13q12-13 (BRCA2) using polymerase chain reaction analysis of polymorphic microsatellite markers. These were compared with 31 patients aged 55-72 years that were matched for size, type and grade. All young breast cancer cases exhibited LOH for at least 1 marker and 20 cases (64.5%) exhibited LOH at 1 or more markers from each interval. The frequency of LOH detected for each of the markers was as follows 17p: p534N (33.3%), D17S796 (36.7%), D17S799 (63.3%) and D17S513 (59.3%); 17q: D17S855 (64.5%), THRA1 (46.7%) and D17S579 (33.3%); and 13q: D13S260 (74.2%), D13S171 (47.6%) and D13S267 (40.0%). These frequencies are higher than those observed at the 3 markers studied in the matched postmenopausal patients: D17S799 (41.4%), D17S855 (35.5%), D13S260 (30.0%). These differences in frequency of LOH were statistically significant for the D17S855 and D13S260 markers (p < 0.025 and p < 0.001 respectively). Although there were more grade III carcinomas (21 of 31 cases), there was no correlation between number of alterations and high grade in younger cases. These data suggest that LOH at these regions could be related to early-onset sporadic breast cancer.     

Association between loss of heterozygosity of BRCA1 and BRCA2 and morphological attributes of sporadic breast cancer

Germline mutations in the breast cancer-associated genes BRCA1 and BRCA2 confer a lifetime risk of malignancy. Distinctive morphological features have been attributed to these familial tumours; however, in sporadic breast cancer, the inter-relationship between loss of heterozygosity (LOH) of these loci and tumour morphology remains to be fully elucidated. We studied a series of 120 sporadic breast carcinomas using microsatellite markers to identify LOH of BRCA1, BRCA2, p53 and PTEN. The associations between loss at each of the loci were examined and related to tumour morphology. LOH of the 4 loci did not occur independently; there were highly significant associations between LOH of BRCA1 and both BRCA2 (p < 0.001) and p53 (p < 0.001). LOH at all 4 loci was significantly associated with a high degree of nuclear pleomorphism. Tumours with LOH of BRCA1 also had high mitotic indices, few tubules and a paucity of DCIS, all of which are morphological features similar to those described for familial cases. Following Bonferroni's correction for multiple tests, we found that the tumours with LOH of BRCA1 were still significantly associated with a high mitotic index (p = 0.0006) and a high degree of nuclear pleomorphism (p = 0.001).     

Detection of loss of heterozygosity at RAD51, RAD52, RAD54 and BRCA1 and BRCA2 loci in breast cancer: pathological correlations.

Loss of heterozygosity (LOH) in loci of the 15q15.1, 12p13, 1p32, 17q21 and 13q12-13 regions may collaborate in the inactivation of RAD51, RAD52, RAD54, BRCA1, BRCA2 and possibly other genes implicated in the repair of double-stranded DNA and in DNA recombination. We investigate allelic losses in microsatellites of the RAD51, RAD52, RAD54, BRCA1 and BRCA2 regions, and their correlations with nine pathologic parameters in 127 breast carcinomas. The LOH analysis was performed by amplifying DNA by PCR, using 15 markers of the 15q15.1, 12p13.3, 1p32, 17q21 and 13q12-13 regions. LOH was found in the RAD51 region in 32% of tumours, in the RAD52 region in 16%, in RAD54 in 20% and in the BRCA1 and BRCA2 regions in 49% and 44% respectively. Significant correlations between one or more regions with concomitant LOH and pathologic parameters were observed with respect to age (P = 0.008), oestrogen receptor content (P = 0.03), progesterone receptors (P = 0.003), higher grade (P = 0.001), more advanced stage (P = 0.004) and peritumoural vessel involvement (P < 0.0001). The number of cases in which LOH was observed simultaneously in two or more regions was always higher than expected on the basis of their statistical probability, and curiously, the three patients with LOH at five regions concomitantly were under the age of 30 years. These results suggest that LOH at these regions could be related to breast cancer, and probably to a poor tumour prognosis.     

Familial breast cancer: double heterozygosity for BRCA1 and BRCA2 mutations with differing phenotypes

The co-existence of mutations in the BRCA1 and BRCA2 genes is unusual, and to date almost all cases reported have had at least one of the Ashkenazi founder mutations. We report on a family in whom individuals are double heterozygotes for a mutation in BRCA1 and a novel splice site mutation in BRCA2. The phenotypes are discordant, where one sister has had multiple cancers in the BRCA spectrum, while the other is unaffected at 65 years of age. The utility of testing is discussed, and the completion of diagnostic testing despite the finding of a potentially causal mutation is validated.     

Clinical impact of detection of loss of heterozygosity of BRCA1 and BRCA2 markers in sporadic breast cancer.

The development of familial and sporadic breast cancer is based on genetic alterations of tumour-suppressor genes, for which loss of heterozygosity (LOH) is one mechanism of gene inactivation. To investigate LOH of BRCA1 (17q21) and BRCA2 (13-q12-13) in sporadic breast cancer, polymerase chain reaction (PCR)-based fluorescent DNA technology for detection of microsatellite polymorphisms was applied. A total of 137 breast cancer and 15 benign breast specimens with matched normal tissue were examined. Fluorescent-labelled PCR products were analysed in an automated DNA sequencer (ALFTM Pharmacia). Losses at both loci were correlated with different histological types, age, tumour size, lymph node status, grading and steroid hormone receptor expression, [SHR: oestrogen receptor (ER), progesterone receptor (PgR)]. For BRCA1 (D17S855, THRA1, D17S579) losses could be detected in invasive ductal carcinoma (IDC; n = 108) in 32-38%, invasive lobular carcinoma (ILC; n = 19) in 21-42% depending on the marker applied, but not in benign breast tumours (n = 15). Losses of BRCA1 markers correlated with larger tumour size, higher grade, and PgR expression. For BRCA2 (D13S260, D13S267, D13S171) losses could be detected in 108 IDCs in 30-38%, in 19 ILCs in 17-39% depending on the marker applied, but not in benign breast tumours. Losses of BRCA2 markers correlated only with higher grade. Microsatellite analyses combined with detection of fluorescent-labelled PCR products by an automated laser DNA sequencer can be used for routine determination of LOH. In sporadic breast cancer, LOH of BRCA1 of BRCA2 does not add decisive prognostic value as stated for familial breast cancer.     

Breast cancer in patients irradiated for Hodgkin's disease: a clinical and pathologic analysis of 45 events in 37 patients.

PURPOSE: To characterize the clinical and pathologic features of patients who developed breast cancer (BC) after treatment for Hodgkin's disease (HD). Recent epidemiologic studies have shown that women who are cured of HD have an increased risk of developing BC. PATIENTS AND METHODS: The clinical data, mammograms, and pathologic specimens of 37 women who developed 45 BCs (eight bilateral events), and had a prior history of treatment for HD were analyzed. RESULTS: The median age at diagnosis of HD was 27 years (range, 11 to 60). All patients received radiotherapy (RT) to the upper part of their body, and 10 also had chemotherapy for HD. The median interval from the treatment of HD to the diagnosis of BC was 15 years (range, 8 to 34). The median age at diagnosis of BC was 43 years (range, 27 to 75), 41% of patients were 39 years old or younger. Most mammograms (81%) showed abnormal findings of mass and/or microcalcifications. Of the eight patients (22%) with bilateral tumors, four were synchronous and four were metachronous. Involvement of the medial half of the breast occurred more frequently than in patients with primary BC (39% and 21%, respectively; P < .002). But, the histologic types, grades, presence of lymphocytic reaction, and lymphatic invasion were similar to those observed in 935 primary BC patients who were previously analyzed at our center. The 6-year actuarial relapse-free survival (RFS) for node-negative BC after HD was 85%. Node-positive patients had a significantly lower RFS of 33% (P = .002). CONCLUSIONS: In comparison to patients with primary BC, patients who develop BC after HD are more likely to be younger, have bilateral disease, and have their tumors more frequently involve the medial half of the breast. Pathologic characteristics, nodal involvement, and prognosis are similar to those of primary BC. BC in women who were treated for HD is becoming an increasing problem, as more patients cured of HD reach a follow-up time of 10 to 15 years. Breast examination and mammography at an early age should be part of the follow-up program for women who are cured of HD.     

Ovarian carcinoma in situ with germline BRCA1 mutation and loss of heterozygosity at BRCA1 and TP53

BACKGROUND: The two-hit hypothesis for the genesis of cancer predicts that cancer can develop when the wild-type allele of a tumor suppressor gene is lost in an individual with a germline mutation in that gene. Neither loss of heterozygosity (LOH) for BRCA1 nor mutations of the TP53 (also known as p53) gene have been documented prior to invasion in ovarian cancers arising in women with germline BRCA1 mutations. Such documentation is difficult because lesions are rarely identified in ovarian epithelium. We, therefore, looked for LOH at microsatellite polymorphisms linked to the BRCA1 and TP53 tumor suppressor loci in an incidental carcinoma in situ of the ovary removed prophylactically from a woman with a germline BRCA1 mutation. METHODS: By use of laser-capture microdissection, we obtained pure populations of atypical ovarian epithelial cells and normal stromal cells. DNA was extracted, amplified with primers flanking polymorphic microsatellites linked to BRCA1 (D17S855 and D17S579) and TP53 (TP53 and D17S786), and analyzed for LOH at these microsatellites. We also tested for p53 expression in the abnormal epithelium by immunohistochemistry. RESULTS: Both of the markers linked to TP53 showed LOH, as did an intragenic BRCA1-linked marker (D17S855). The other microsatellite marker for BRCA1 was uninformative. Immunohistochemical staining with an antibody to p53 showed strong immunoreactivity confined to the atypical epithelium. CONCLUSIONS: BRCA1, as well as TP53, can undergo LOH prior to stromal invasion in BRCA1-associated ovarian cancer. Strong immunoreactivity for p53 suggests the presence of mutated p53 in these cells as well. These findings suggest that loss of function of these two tumor suppressor genes occurs early in ovarian carcinogenesis in BRCA1 mutation carriers.     

Double heterozygosity for mutations in BRCA1 and BRCA2 in German breast cancer patients: implications on test strategies and clinical management

Double heterozygosity for disease-causing BRCA1 and BRCA2 mutations is a very rare condition in most populations. Here we describe genetic and clinical data of eight female double heterozygotes (DH) for BRCA1 and BRCA2 mutations found in a cohort of 8162 German breast/ovarian cancer families and compare it with the data of their single heterozygous relatives and of the index patients of the German Consortium for Hereditary Breast and Ovarian Cancer. Furthermore, we analyze the phenotypic features of these patients with respect to age at onset of first cancer, first breast/ovarian cancer and the number of disease manifestations and compare them to that of published Caucasian female DHs and their single heterozygous female relatives. German DHs were not significantly younger at diagnosis of first breast cancer than the single heterozygous index patients of the German Consortium. However, if the data of our study were pooled with that of the literature, DHs were substantially younger at onset of first cancer (mean age 40.4 years, 95 % CI = 36.6-44.1) than their single heterozygous female relatives (mean age 51.9 years, 95 % CI = 46.8-57.0). The two groups also differed concerning the onset of first breast cancer (mean age 40.6 years, 95 % CI = 36.6-44.5 vs. 52.6, 95 % CI = 47.5-57.6). In addition, DHs had a more severe disease than their female relatives carrying a single BRCA mutation (1.4 vs. 0.6 manifestations per person). In contrast to Ashkenazi Jewish females, Caucasian DH females might develop breast cancer at an earlier age and have a more severe disease than single heterozygous BRCA mutation carriers. Therefore, DHs may benefit from more intensive surveillance programs/follow-up care and prophylactic surgery.     

Characteristics of double heterozygosity for BRCA1 and BRCA2 germline mutations in Korean breast cancer patients

To investigate clinical, pathological, and familial characteristics of Korean patients with double heterozygosity for BRCA1 and BRCA2 mutations, six breast tumors of five patients who carried deleterious mutations in both of the genes were included. Medical records of the patients were reviewed and genetic testing by direct sequencing was undertaken to detect mutations in BRCA1 and BRCA2. Seven frameshift and three nonsense mutations were identified, and four mutations are novel in the Breast Cancer Information Core. There were no Ashkenazi founder mutations detected. The mean age at diagnosis for breast cancer was 33 years. All six tumors were infiltrating ductal carcinoma and poorly differentiated. Pathologic stage was I or II, and immunohistochemistry showed negative immunoreactivity for estrogen receptor and Her-2/neu in all tumors. Positive immunoreactivity for progesterone receptor was found only in one tumor. Three patients had familial history of breast, ovarian or other cancers. One patient who was diagnosed for breast cancer at the age of 26 had two maternal family members of metachronous bilateral breast cancer. Another patient who experienced metachronous bilateral breast cancer had maternal history of ovarian and esophageal cancer. In summary, Korean patients with double heterozygosity for BRCA1 and BRCA2 were young at diagnosis of breast cancer. Tumors were early stage, high grade, and almost triple-negative phenotype. All familial history of breast, ovary or other cancer was maternal. Close surveillance and accurate risk assessment should be provided for the patients with mutations in the both of the genes.     

BRCA1, BRCA2, AR and IGF-I expression in prostate cancer: correlation between RT-qPCR and immunohistochemical detection

Identification and characterization of biomarkers in prostate cancer are important for improving the diagnosis. The aim of this study was to determine differences in the expression of 4 genes according to the stage of malignancy in prostate cancer. We analyzed BRCA1, BRCA2, androgen receptor (AR) and IGF-I gene expression in a cohort of 98 prostate biopsies. We used TaqMan RT-qPCR for mRNA detection, and correlation with proteins was performed using immunohistochemistry. Among the 98 studied prostate biopsies, high heterogeneity in the expression of the 4 genes was detected among the different histological types. However, down-regulation of BRCA1 and BRCA2 mRNA was detected, particularly in the normal tissues. The expression of AR was dependent on the stage of the tumor. The IGF-I gene was specifically expressed in the tumor tissues. Upon comparison between protein and mRNA expression for BRCA1, BRCA2 and AR, we obtained a trend; however, this did not achieve statistical significance. Regarding IGF-I, a correlation between mRNA expression and staining intensity of the protein was found to be significant (p<0.012). The AR biomarker was found to be slightly correlated with the prostate cancer diagnosis (p=0.013). AR was found to be decreased in the tumors with a 43% sensitivity and 90% specificity. The relative risk of 2.05 (1.13-3.69) indicated a 2?fold higher chance of cancer occurrence when AR was ≤0.206.     

Estrogen receptor positive breast cancers in BRCA1 mutation carriers: clinical risk factors and pathologic features

Introduction

Recent, international declines in breast cancer incidence are unprecedented, and the causes remain controversial. Few data sources can address breast cancer incidence trends according to pertinent characteristics like hormone therapy use history.

Methods

We used the prospective California Teachers Study to evaluate changes in self-reported use of menopausal hormone therapy (HT) between 1995 to 1996 and 2005 to 2006 and age-adjusted breast cancer incidence among 74,647 participants aged 50 years or older. Breast cancer occurrence was determined by linkage with the California Cancer Registry.

Results

During 517,286 woman years of follow up, 565 in situ and 2,668 invasive breast cancers were diagnosed. In situ breast cancer incidence rates in this population did not change significantly from 2000 to 2002 to 2003 to 2005, whereas rates of invasive breast cancer declined significantly by 26.0% from 528.0 (95% confidence intervals (CI) = 491.1, 564.9) per 100,000 women in 2000 to 2002 to 390.6 (95% CI = 355.6, 425.7) in 2003 to 2005. The decline in invasive breast cancer incidence rates was restricted to estrogen receptor-positive tumors. In 1996 to 1999 and 2000 to 2002 invasive breast cancer incidence was higher for women who reported current HT use especially estrogen-progestin (EP) use at baseline than for never or past users; but by 2003 to 2005 rates were comparable between these groups. For women who were taking EP in 2001 to 2002,75% of whom had stopped use by 2005 to 2006, incidence had declined 30.6% by 2003 to 2005 (P = 0.001); whereas incidence did not change significantly for those who never took HT (P = 0.33).

Conclusions

Few data resources can examine prospectively individual HT use and breast cancer diagnosis. Stable in situ breast cancer rates imply consistent levels of screening and suggest recent declines in invasive breast cancer to be explained predominantly by changes in HT use.     

Selective loss of heterozygosity in multiple breast cancers from a carrier of mutations in both BRCA1 and BRCA2

Carriers of one mutant allele of either BRCA1 or BRCA2 are at risk for somatic loss of the second wild-type allele, leading to the initiation of breast tumorigenesis. We identified a patient of Ashkenazi Jewish heritage with germ-line heterozygous mutations in both BRCA1 (5382insC) and BRCA2 (6174delT), who had developed three independent breast cancers by age 47. Two breast cancers demonstrated inactivation of both BRCA2 alleles but retention of the wild-type BRCA1 allele, and the third showed loss of heterozygosity for BRCA1 but not BRCA2. The observation that breast tumors arising in a double heterozygote show biallelic inactivation of either BRCA1 or BRCA2, but not both, suggests that these genetic events are functionally equivalent in initiating tumorigenesis. The distinct histopathological features of these tumors may reflect the acquisition of subsequent genetic events.     

BRCA1 gene mutation and loss of heterozygosity on chromosome 17q21 in primary prostate cancer

The tumor suppressor gene BRCA1 on chromosome 17q21 has been characterized and shown to be mutated in patients with familial breast and ovarian cancer. Several studies examined the relatives of women with breast cancer and noted an association with ovarian and prostate cancer. This study investigated 24 human prostate cancer specimens for BRCA1 gene mutations and loss of heterozygosity (LOH) on chromosome 17q21 assessed by the polymerase chain reaction. LOH was identified using 7 highly polymorphic tandem repeat markers on chromosome 17q21, in addition to an analysis of the whole coding region of the BRCA1 gene. Four of the 24 prostate cancer specimens showed LOH at one or more loci, all of which were histologically poorly differentiated (4 of 11) and stage D (4 of 15). One of the 24 cases showed a germ‐line mutation of the BRCA1 gene, and a sister of this patient died of ovarian cancer. It appears that the BRCA1 gene is not frequently involved in the development of primary prostate cancer. Int. J. Cancer (Pred. Oncol.) 84:19–23, 1999. © 1999 Wiley‐Liss, Inc.