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Purpose: Mammographic density phenotypes, adjusted for age and body mass index (BMI), are strong predictors of breast cancer risk. BMI is associated with mammographic density measures, but the role of circulating sex hormone concentrations is less clear. We investigated the relationship between BMI, circulating sex hormone concentrations, and mammographic density phenotypes using Mendelian randomization (MR)., Methods: We applied two-sample MR approaches to assess the association between genetically predicted circulating concentrations of sex hormones [estradiol, testosterone, sex hormone-binding globulin (SHBG)], BMI, and mammographic density phenotypes (dense and non-dense area). We created instrumental variables from large European ancestry-based genome-wide association studies and applied estimates to mammographic density phenotypes in up to 14,000 women of European ancestry. We performed analyses overall and by menopausal status., Results: Genetically predicted BMI was positively associated with non-dense area (IVW: β = 1.79; 95% CI = 1.58, 2.00; p = 9.57 × 10 -63 ) and inversely associated with dense area (IVW: β = - 0.37; 95% CI = - 0.51,- 0.23; p = 4.7 × 10 -7 ). We observed weak evidence for an association of circulating sex hormone concentrations with mammographic density phenotypes, specifically inverse associations between genetically predicted testosterone concentration and dense area (β = - 0.22; 95% CI = - 0.38, - 0.053; p = 0.009) and between genetically predicted estradiol concentration and non-dense area (β =  - 3.32; 95% CI = - 5.83, - 0.82; p = 0.009), although results were not consistent across a range of MR approaches., Conclusion: Our findings support a positive causal association between BMI and mammographic non-dense area and an inverse association between BMI and dense area. Evidence was weaker and inconsistent for a causal effect of circulating sex hormone concentrations on mammographic density phenotypes. Based on our findings, associations between circulating sex hormone concentrations and mammographic density phenotypes are weak at best., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Purpose: The American Cancer Society (ACS) published an updated Guideline for Cancer Prevention (ACS Guideline) in 2020. Research suggests that adherence to the 2012 ACS Guideline might lower breast cancer risk, but there is limited evidence that this applies to women at increased familial and genetic risk of breast cancer., Methods: Using the Breast Cancer Family Registry (BCFR), a cohort enriched for increased familial and genetic risk of breast cancer, we examined adherence to three 2020 ACS Guideline recommendations (weight management (body mass index), physical activity, and alcohol consumption) with breast cancer risk in 9615 women. We used Cox proportional hazard regression modeling to calculate hazard ratios (HRs) and 95% confidence intervals (CI) overall and stratified by BRCA1 and BRCA2 pathogenic variant status, family history of breast cancer, menopausal status, and estrogen receptor-positive (ER +) breast cancer., Results: We observed 618 incident invasive or in situ breast cancers over a median 12.9 years. Compared with being adherent to none (n = 55 cancers), being adherent to any ACS recommendation (n = 563 cancers) was associated with a 27% lower breast cancer risk (HR = 0.73, 95% CI: 0.55-0.97). This was evident for women with a first-degree family history of breast cancer (HR = 0.68, 95% CI: 0.50-0.93), women without BRCA1 or BRCA2 pathogenic variants (HR = 0.71, 95% CI: 0.53-0.95), postmenopausal women (HR = 0.63, 95% CI: 0.44-0.89), and for risk of ER+ breast cancer (HR = 0.63, 95% CI: 0.40-0.98)., Discussion: Adherence to the 2020 ACS Guideline recommendations for BMI, physical activity, and alcohol consumption could reduce breast cancer risk for postmenopausal women and women at increased familial risk., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Purpose: The main aim of this study was to screen epigenetic modifier genes and known breast cancer driver genes for germline mutations in non-BRCA1/2 (BRCAx) breast cancer families in order to identify novel susceptibility genes of moderate-high penetrance., Methods: We screened 264 candidate susceptibility genes in 656 index cases from non-BRCA1/2 families. Potentially pathogenic candidate mutations were then genotyped in all available family members for the assessment of co-segregation of the variant with disease in the family in order to estimate the breast cancer risks associated with these mutations. For 11 of the candidate susceptibility genes, we screened an additional 800 non-BRCA1/2 breast cancer cases and 787 controls., Results: Only two genes, CHD8 and USH2A showed any evidence of an increased risk of breast cancer (RR = 2.40 (95% CI 1.0-7.32) and 2.48 (95% CI 1.11-6.67), respectively)., Conclusions: We found no convincing evidence that epigenetic modifier and known breast cancer driver genes carry germline mutations that increase breast cancer risk. USH2A is no longer regarded as a breast cancer driver gene and seems an implausible candidate given its association with Usher syndrome. However, somatic mutations in CHD8 have been recently reported, making it an even more promising candidate, but further analysis of CHD8 in very large cohorts of families or case-control studies would be required to determine if it is a moderate-risk breast cancer susceptibility gene.

Purpose: Women diagnosed with breast cancer have heterogeneous survival outcomes that cannot be fully explained by known prognostic factors, and germline variation is a plausible but unconfirmed risk factor., Methods: We used three approaches to test the hypothesis that germline variation drives some differences in survival: mortality loci identification, tumor aggressiveness loci identification, and whole-genome prediction. The 2954 study participants were women diagnosed with breast cancer before age 50, with a median follow-up of 15 years who were genotyped on an exome array. We first searched for loci in gene regions that were associated with all-cause mortality. We next searched for loci in gene regions associated with five histopathological characteristics related to tumor aggressiveness. Last, we also predicted 10-year all-cause mortality on a subset of 1903 participants (3,245,343 variants after imputation) using whole-genome prediction methods., Results: No risk loci for mortality or tumor aggressiveness were identified. This null result persisted when restricting to women with estrogen receptor-positive tumors, when examining suggestive loci in an independent study, and when restricting to previously published risk loci. Additionally, the whole-genome prediction model also found no evidence to support an association., Conclusion: Despite multiple complementary approaches, our study found no evidence that mortality in women with early onset breast cancer is influenced by germline variation.

Purpose: Cis-acting regulatory SNPs resulting in differential allelic expression (DAE) may, in part, explain the underlying phenotypic variation associated with many complex diseases. To investigate whether common variants associated with DAE were involved in breast cancer susceptibility among BRCA1 and BRCA2 mutation carriers, a list of 175 genes was developed based of their involvement in cancer-related pathways., Methods: Using data from a genome-wide map of SNPs associated with allelic expression, we assessed the association of ~320 SNPs located in the vicinity of these genes with breast and ovarian cancer risks in 15,252 BRCA1 and 8211 BRCA2 mutation carriers ascertained from 54 studies participating in the Consortium of Investigators of Modifiers of BRCA1/2., Results: We identified a region on 11q22.3 that is significantly associated with breast cancer risk in BRCA1 mutation carriers (most significant SNP rs228595 p = 7 × 10 -6 ). This association was absent in BRCA2 carriers (p = 0.57). The 11q22.3 region notably encompasses genes such as ACAT1, NPAT, and ATM. Expression quantitative trait loci associations were observed in both normal breast and tumors across this region, namely for ACAT1, ATM, and other genes. In silico analysis revealed some overlap between top risk-associated SNPs and relevant biological features in mammary cell data, which suggests potential functional significance., Conclusion: We identified 11q22.3 as a new modifier locus in BRCA1 carriers. Replication in larger studies using estrogen receptor (ER)-negative or triple-negative (i.e., ER-, progesterone receptor-, and HER2-negative) cases could therefore be helpful to confirm the association of this locus with breast cancer risk., Competing Interests: The authors declare that they have no conflict of interest. Ethical approval Study participants were recruited through the CIMBA Initiative, following the approval of the corresponding protocol by the Institutional Review Board or Ethics Committee at each participating center. Written informed consent was obtained from all study participants.

BReast CAncer genes 1 and 2 (BRCA1 and BRCA2) mutation carriers diagnosed with breast cancer are at increased risk of developing a second primary breast cancer. Data from high-risk clinics may be subject to different biases which can cause both over and underestimation of this risk. Using data from a large multi-institutional family registry we estimated the 10-year cumulative risk of second primary breast cancer including more complete testing information on family members. We prospectively followed 800 women diagnosed with breast cancer from the Breast Cancer Family Registry (BCFR) who were carriers of a BRCA1 or BRCA2 pathogenic mutation or a variant of unknown clinical significance. In order to limit survival and ascertainment bias, cases were limited to those diagnosed with a first primary breast cancer from 1994 to 2001 and enrolled in the BCFR within 3 years after their cancer diagnosis. We excluded women enrolled after being diagnosed with a second breast cancer. We calculated 10-year incidence of second primary breast cancers. The 10-year incidence of a second primary breast cancer was highest in BRCA1 mutation carriers (17 %; 95 % CI 11-25 %), with even higher estimates in those first diagnosed under the age of 40 (21 %; 95 % CI 13-34 %). Lower rates were found in BRCA2 mutation carriers (7 %; 95 % CI 3-15 %) and women with a variant of unknown clinical significance (6 %; 95 % CI 4-9 %). Whereas the cumulative 10-year incidence of second primary breast cancer is high in BRCA1 mutation carriers, the estimates in BRCA2 mutation carriers and women with variants of unknown clinical significance are similar to those reported in women with sporadic breast cancer.

Though the role of Estrogen Receptor (ER)α in breast cancer has been studied extensively, there is little consensus about the role of alternative ER isoform ERβ in breast cancer biology. ERβ has significant sequence homology to ERα but is located on a different chromosome and maintains both overlapping and unique functional attributes. Five variants exist, resulting from alternative splicing of the C-terminal region of ERβ. The relevance of ERβ variants in breast cancer outcomes and response to therapy is difficult to assess because of conflicting reports in the literature, likely due to variable methods used to assess ERβ in patient tumors. Here, we quantitatively assess expression of ERβ splice variants on over 2,000 breast cancer patient samples. Antibodies against ERβ variants were validated for staining specificity in cell lines by siRNA knockdown of ESR2 and staining reproducibility on formalin-fixed paraffin-embedded tissue by quantitative immunofluorescence (QIF) using AQUA technology. We found antibodies against splice variants ERβ1 and ERβ5, but not ERβ2/cx, which were sensitive, specific, and reproducible. QIF staining of validated antibodies showed both ERβ1 and ERβ5 QIF scores, which have a normal (bell shaped) distribution on most cohorts assessed, and their expression is significantly associated with each other. Extensive survival analyses show that ERβ1 is not a prognostic or predictive biomarker for breast cancer. ERβ5 appears to be a context-dependent marker of worse outcome in HER2-positive and triple-negative patients, suggesting an unknown biological function in the absence of ERα.

Human breast cancer cells with a CD44(+)/CD24(-/low) or ALDH1+ phenotype have been demonstrated to be enriched for cancer stem cells (CSCs) using in vitro and in vivo techniques. The aim of this study was to determine the association between CD44(+)/CD24(-/low) and ALDH1 expression with clinical-pathologic tumor characteristics, tumor molecular subtype, and survival in a well characterized collection of familial breast cancer cases. 364 familial breast cancers from the Ontario Familial Breast Cancer Registry (58 BRCA1-associated, 64 BRCA2-associated, and 242 familial non-BRCA1/2 cancers) were studied. Each tumor had a centralized pathology review performed. TMA sections of all tumors were analyzed for the expression of ER, PR, HER2, CK5, CK14, EGFR, CD44, CD24, and ALDH1. The Chi square test or Fisher's exact test was used to analyze the marker associations with clinical-pathologic tumor variables, molecular subtype and genetic subtype. Analyses of the association of overall survival (OS) with marker status were conducted using Kaplan-Meier plots and log-rank tests. The CD44(+)/CD24(-/low) and ALDH1+ phenotypes were identified in 16% and 15% of the familial breast cancer cases, respectively, and associated with high-tumor grade, a high-mitotic count, and component features of the medullary type of breast cancer. CD44(+)/CD24(-/low) and ALDH1 expression in this series were further associated with the basal-like molecular subtype and the CD44(+)/CD24(-/low) phenotype was independently associated with BRCA1 mutational status. The currently accepted breast CSCs markers are present in a minority of familial breast cancers. Whereas the presence of these markers is correlated with several poor prognostic features and the basal-like subtype of breast cancer, they do not predict OS.

Lymphatic invasion (LVI) is associated with disease recurrence in axillary node-negative (ANN) breast cancer. Using gene expression profiling of 105 ANN tumors, we found that podocalyxin (PODXL) was more highly expressed in tumors with LVI (LVI+) than in those without LVI (LVI-). Differences in PODXL expression were validated using real-time polymerase chain reaction as well as by immunohistochemistry in an independent set of 652 tumors on tissue microarrays. Disease-free survival (DFS) analyses were conducted for association of high PODXL protein expression with risk of distant recurrence overall and within breast cancer subtypes using both Cox and cure-rate models. High PODXL expression was associated with poor prognosis features including large tumor size, high histological grade, estrogen and progesterone receptor negativity, and with clinical alterations characteristic of the basal-like breast cancer phenotype. Surprisingly, despite having other poor prognosis characteristics, women with high PODXL expressing tumors had better long-term DFS in multivariate analysis with traditional clinicopathologic factors including LVI and HER2 status (P = 0.001). PODXL has the potential to be a useful biomarker for identifying good prognosis patients in characteristically poor prognosis breast cancer groups and may impact treatment of women with this disease.

Energy restriction inhibits mammary tumor development in animal models. Epidemiologic studies in humans generally do not support an association between dietary energy intake and breast cancer risk, although some studies suggest a more complex interplay between measures of energy intake, physical activity, and body size. We examined the association between total energy intake jointly with physical activity and body mass index (BMI) and the risk of breast cancer among 1,775 women diagnosed with breast cancer between 1995 and 2006 and 2,529 of their unaffected sisters, enrolled in the Breast Cancer Family Registry. We collected dietary data using the Hawaii-Los Angeles Multiethnic Cohort food frequency questionnaire. Using conditional logistic regression to estimate the odds ratios (OR) and 95 % confidence intervals (CI) associated with total energy intake, we observed an overall 60-70 % increased risk of breast cancer among women in the highest quartile of total energy intake compared to those in the lowest quartile (Q4 vs. Q1: OR = 1.6, 95 % CI: 1.3-2.0; P (trend) < 0.0001); these associations were limited to pre-menopausal women or women with hormone receptor-positive cancers. Although the associations were slightly stronger among women with a higher BMI or lower level of average lifetime physical activity, we observed a positive association between total energy intake and breast cancer risk across different strata of physical activity and BMI. Our results suggest that within sisters, high energy intake may increase the risk of breast cancer independent of physical activity and body size. If replicated in prospective studies, then these findings suggest that reductions in total energy intake may help in modifying breast cancer risk.

Several common germline variants identified through genome-wide association studies of breast cancer risk in the general population have recently been shown to be associated with breast cancer risk for BRCA1 and/or BRCA2 mutation carriers. When combined, these variants can identify marked differences in the absolute risk of developing breast cancer for mutation carriers, suggesting that additional modifier loci may further enhance individual risk assessment for BRCA1 and BRCA2 mutation carriers. Recently, a common variant on 6p22 (rs9393597) was found to be associated with increased breast cancer risk for BRCA2 mutation carriers [hazard ratio (HR) = 1.55, 95 % confidence interval (CI) 1.25-1.92, p = 6.0 × 10(-5)]. This observation was based on data from GWAS studies in which, despite statistical correction for multiple comparisons, the possibility of false discovery remains a concern. Here, we report on an analysis of this variant in an additional 6,165 BRCA1 and 3,900 BRCA2 mutation carriers from the Consortium of Investigators of Modifiers of BRCA1/2 (CIMBA). In this replication analysis, rs9393597 was not associated with breast cancer risk for BRCA2 mutation carriers (HR = 1.09, 95 % CI 0.96-1.24, p = 0.18). No association with ovarian cancer risk for BRCA1 or BRCA2 mutation carriers or with breast cancer risk for BRCA1 mutation carriers was observed. This follow-up study suggests that, contrary to our initial report, this variant is not associated with breast cancer risk among individuals with germline BRCA2 mutations.

Epidemiologic studies of histologic types of breast cancer including mucinous, medullary, and tubular carcinomas have primarily relied on International Classification of Diseases-Oncology (ICD-O) codes assigned by local pathologists to define histology. Using data from the Breast Cancer Family Registry (BCFR), we compared histologic agreement between centralized BCFR pathology review and ICD-O codes available from local tumor registries among 3,260 breast cancer cases. Agreement was low to moderate for less common histologies; for example, only 55 and 26 % of cases classified as mucinous and medullary, respectively, by centralized review were similarly classified using ICD-O coding. We then evaluated risk factors for each histologic subtype by comparing each histologic case group defined by centralized review with a common set of 2,997 population-based controls using polytomous logistic regression. Parity [odds ratio (OR) = 0.4, 95 % confidence interval (95 % CI): 0.2-0.9, for parous vs. nulliparous], age at menarche (OR = 0.5, 95 % CI: 0.3-0.9, for age ≥13 vs. ≤11), and use of oral contraceptives (OCs) (OR = 0.5, 95 % CI: 0.2-0.8, OC use >5 years vs. never) were associated with mucinous carcinoma (N = 92 cases). Body mass index (BMI) (OR = 1.05, 95 % CI: 1.0-1.1, per unit of BMI) and high parity (OR = 2.6, 95 % CI: 1.1-6.0 for ≥3 live births vs. nulliparous) were associated with medullary carcinoma (N = 90 cases). We did not find any associations between breast cancer risk factors and tubular carcinoma (N = 86 cases). Relative risk estimates from analyses using ICD-O classifications of histology, rather than centralized review, resulted in attenuated, and/or more imprecise, associations. These findings suggest risk factor heterogeneity across breast cancer tumor histologies, and demonstrate the value of centralized pathology review for classifying rarer tumor types.

Overexpression of topoisomerase II protein (topo 2α) is postulated to be more closely associated with responsiveness to anthracycline-containing chemotherapy than human epidermal growth factor receptor type 2 (HER2) gene amplification or alterations in the topoisomerase II alpha gene (TOP2A). The authors used tissue microarrays from 477 of 710 premenopausal women with node-positive breast cancer randomized to CEF or CMF adjuvant chemotherapy in the NCIC Clinical Trials Group Mammary 5 (MA.5) trial. No significant interaction was found between treatment and continuous topo 2α level in either relapse-free (RFS) or overall survival (OS). In 136 patients (28.5%) whose tumors showed topo 2α overexpression by immunohistochemistry based on a cut-off of 13%, CEF was superior to CMF for RFS (adjusted HR 0.45; 95% CI 0.25-0.82; P = 0.009) and OS (adjusted HR 0.50; 95% CI 0.26-0.96; P = 0.04). When tumors lacked topo 2α overexpression, CEF was not superior for RFS (adjusted HR 0.88; 95% CI 0.64-1.22; P = 0.46) or OS (adjusted HR 0.95; 95% CI 0.66-1.38; P = 0.80). Interaction between topo 2α and treatment was borderline significant for RFS (P = 0.04) and OS (P = 0.05) and not substantially more significant than between TOP2A gene alteration (P (interaction) = 0.09 for RFS and 0.02 for OS) or HER2 overexpression (P (interaction) = 0.002 for RFS and 0.009 for OS). Topo 2α protein overexpression based on the cut-off identified in this study, TOP2A gene alterations and HER2 protein overexpression were each associated with responsiveness to anthracycline-containing chemotherapy. The topo 2α protein analysis was exploratory and will require further validation.

EMSY is a putative oncogene amplified in a minority of breast carcinomas, its protein product interacts with and transcriptionally silences BRCA2. We hypothesized that breast tumors from BRCA2 mutation carriers would be less likely than other familial breast cancers to exhibit EMSY amplification. As EMSY is located on 11q13 in proximity to CCND1, an established breast cancer oncogene, we also examined the amplification of CCND1 in the same tumor cohort. Amplification of EMSY and CCND1 were examined in 58 BRCA1-associated, 64 BRCA2-associated, and 242 familial non-BRCA1/BRCA2 breast cancers using fluorescent in situ hybridization (FISH). All tumors had a centralized pathology review and underwent molecular phenotyping by immunohistochemical profiling on tissue microarrays (TMAs). Tumors with amplification of EMSY and/or CCND1 were compared with non-amplified tumors for morphological appearance, molecular subtype, and overall survival. EMSY amplification was detected in 8% of BRCA1-associated, 0% of BRCA2-associated, and 9% of familial non-BRCA1/BRCA2 breast tumors (P = 0.036). CCND1 was amplified in 4% of BRCA1-associated, 13% of BRCA2-associated and 21% of non-BRCA1/BRCA2 breast tumors (P = 0.054). EMSY was amplified independently of CCND1 in 38% of cases. EMSY amplification was associated with increased tumor stage only; whereas CCND1 amplification was associated with high tumor grade, ER positivity, and inversely associated with the basal-like phenotype. There was a trend toward worse overall survival in ER-positive non-BRCA1/BRCA2 familial breast cancer patients whose tumors exhibited EMSY and CCND1 co-amplification. BRCA2-associated breast tumors are less likely than non-BRCA1/BRCA2 familial breast cancers to exhibit EMSY amplification. BRCA1-associated breast cancers are less likely than non-BRCA1/BRCA2 familial breast cancers to exhibit CCND1 amplification. EMSY amplification does occur independently of CCND1 amplification in a minority of familial breast cancers, supporting its role as a possible breast cancer oncogene.

Few studies have considered the joint association of body mass index (BMI) and physical activity, two modifiable factors, with all-cause mortality after breast cancer diagnosis. Women diagnosed with invasive breast cancer (n = 4,153) between 1991 and 2000 were enrolled in the Breast Cancer Family Registry through population-based sampling in Northern California, USA; Ontario, Canada; and Melbourne and Sydney, Australia. During a median follow-up of 7.8 years, 725 deaths occurred. Baseline questionnaires assessed moderate and vigorous recreational physical activity and BMI prior to diagnosis. Associations with all-cause mortality were assessed using Cox proportional hazards regression, adjusting for established prognostic factors. Compared with no physical activity, any recreational activity during the 3 years prior to diagnosis was associated with a 34% lower risk of death [hazard ratio (HR) = 0.66, 95% confidence interval (CI): 0.51-0.85] for women with estrogen receptor (ER)-positive tumors, but not those with ER-negative tumors; this association did not appear to differ by race/ethnicity or BMI. Lifetime physical activity was not associated with all-cause mortality. BMI was positively associated with all-cause mortality for women diagnosed at age > or =50 years with ER-positive tumors (compared with normal-weight women, HR for overweight = 1.39, 95% CI: 0.90-2.15; HR for obese = 1.77, 95% CI: 1.11-2.82). BMI associations did not appear to differ by race/ethnicity. Our findings suggest that physical activity and BMI exert independent effects on overall mortality after breast cancer.

Several studies suggest that higher circulating insulin-like growth factor I (IGF-I) levels are associated with premenopausal breast cancer risk. Breast cancer risk and circulating IGF-I concentration appear to be partly heritable, thus genetic variation at IGF1 could influence IGF-I levels and breast cancer risk. We investigated the association of IGF1 CA repeat variants with premenopausal breast cancer risk using a family-based design. The study sample included 840 families from the Ontario Familial Breast Cancer Registry (OFBCR) and the Australian Breast Cancer Family Registry (ABCFR). Three CA repeat variants, at 5', 3', and in intron 2 were genotyped (5'CA, 3'CA, In2CA). We found several nominally significant associations. The 5'CA-21 allele (P = 0.03) and In2CA-212 allele (P = 0.04) were associated with lower risk, and the In2CA-216 allele with higher risk (P = 0.04) for the combined ABCFR-OFBCR. These associations were not significant after taking into account multiple comparisons. In2CA-216 was more strongly associated with risk when we used a recessive instead of an additive model (P = 0.01). 5'CA alleles of repeat length 18-20 were associated with higher risk (P = 0.02), and 5'CA alleles of >20 repeats were associated with lower risk (P = 0.01). These associations were significant in the OFBCR (In2CA-216 recessive, P = 0.02; 5'CA 18-20 and >20 allele grouping, P = 0.01) but not strongly supported by the ABCFR (In2CA-216 recessive, P = 0.14; 5'CA 18-20, P = 0.25; 5'CA >20, P = 0.20). The associations we found could be due to chance as many comparisons were made. Our results do not strongly support an association between these IGF1 variants and breast cancer risk.

Background: BRCA1- and BRCA2-associated tumors appear to have distinct molecular signatures. BRCA1-associated tumors are predominantly basal-like cancers, whereas BRCA2-associated tumors have a predominant luminal-like phenotype. These two molecular signatures reflect in part the two cell types found in the terminal duct lobular unit of the breast. To elucidate novel genes involved in these two spectra of breast tumorigenesis we performed global gene expression analysis on breast tumors from germline BRCA1 and BRCA2 mutation carriers., Methodology: Breast tumor RNAs from 7 BRCA1 and 6 BRCA2 mutation carriers were profiled using UHN human 19K cDNA microarrays. Supervised univariate analyses were conducted to identify genes differentially expressed between BRCA1 and BRCA2-associated tumors. Selected discriminatory genes were validated using real time reverse transcription polymerase chain reaction in the tumor RNAs, and/or by immunohistochemistry (IHC) or by in situ hybridization (ISH) on tissue microarrays (TMAs) containing an independent set of 58 BRCA1 and 64 BRCA2-associated tumors., Results: Genes more highly expressed in BRCA1-associated tumors included stathmin, osteopontin, TGFbeta2 and Jagged 1 in addition to genes previously identified as characteristic of basal-like breast cancers. BRCA2-associated cancers were characterized by the higher relative expression of FGF1 and FGFR2. FGFR2 protein was also more highly expressed in BRCA2-associated cancers (P = 0.004)., Significance: BRCA1-associated tumours demonstrated increased expression of component genes of the Notch and TGFbeta pathways whereas the higher expression of FGFR2 and FGF1 in BRCA2-associated cancers suggests the existence of an autocrine stimulatory loop.

Although having a family history of breast cancer is a well established breast cancer risk factor, it is not known whether it influences mortality after breast cancer diagnosis. We studied 4,153 women with first primary incident invasive breast cancer diagnosed between 1991 and 2000, and enrolled in the Breast Cancer Family Registry through population-based sampling in Northern California, USA; Ontario, Canada; and Melbourne and Sydney, Australia. Cases were oversampled for younger age at diagnosis and/or family history of breast cancer. Carriers of germline mutations in BRCA1 or BRCA2 were excluded. Cases and their relatives completed structured questionnaires assessing breast cancer risk factors and family history of cancer. Cases were followed for a median of 6.5 years, during which 725 deaths occurred. Cox proportional hazards regression was used to evaluate associations between family history of breast cancer at the time of diagnosis and risk of all-cause mortality after breast cancer diagnosis, adjusting for established prognostic factors. The hazard ratios for all-cause mortality were 0.98 (95% confidence interval [CI] = 0.84-1.15) for having at least one first- or second-degree relative with breast cancer, and 0.85 (95% CI = 0.70-1.02) for having at least one first-degree relative with breast cancer, compared with having no such family history. Estimates did not vary appreciably when stratified by case or tumor characteristics. In conclusion, family history of breast cancer is not associated with all-cause mortality after breast cancer diagnosis for women without a known germline mutation in BRCA1 or BRCA2. Therefore, clinical management should not depend on family history of breast cancer.

The Breast Cancer Family Registry is a resource for interdisciplinary and translational studies of the genetic epidemiology of breast cancer. This resource is available to researchers worldwide for collaborative studies. Herein, we report the results of testing for germline mutations in BRCA1 and BRCA2. We have tested 4,531 probands for mutations in BRCA1 and 4,084 in BRCA2. Deleterious mutations in BRCA1 and BRCA2 were identified for 9.8% of probands tested [233/4,531 (5.1%) for BRCA1 and 193/4,084 (4.7%) for BRCA2]. Of 1,385 Ashkenazi Jewish women tested for only the three founder mutations, 17.4% carried a deleterious mutation. In total, from the proband and subsequent family testing, 1,360 female mutation carriers (788 in BRCA1, 566 in BRCA2, 6 in both BRCA1 and BRCA2) have been identified. The value of the resource has been greatly enhanced by determining the germline BRCA1 and BRCA2 mutation statuses of nearly 6,000 probands.

Background: The transforming growth factor beta-1 gene (TGFB1) is a plausible candidate for breast cancer susceptibility. The L10P variant of TGFB1 is associated with higher circulating levels and secretion of TGF-beta, and recent large-scale studies suggest strongly that this variant is associated with breast cancer risk in the general population., Methods: To evaluate whether TGFB1 L10P also modifies the risk of breast cancer in BRCA1 or BRCA2 mutation carriers, we undertook a multi-center study of 3,442 BRCA1 and 2,095 BRCA2 mutation carriers., Results: We found no evidence of association between TGFB1 L10P and breast cancer risk in either BRCA1 or BRCA2 mutation carriers. The per-allele HR for the L10P variant was 1.01 (95%CI: 0.92-1.11) in BRCA1 carriers and 0.92 (95%CI: 0.81-1.04) in BRCA2 mutation carriers., Conclusions: These results do not support the hypothesis that TGFB1 L10P genotypes modify the risk of breast cancer in BRCA1 or BRCA2 mutation carriers.

High-density oligonucleotide microarrays containing a large number of single nucleotide polymorphisms (SNPs) have enabled genome-wide association (GWA) analysis to become a reality. We used the early access Affymetrix Mendel Nsp 250K chips in a GWA case-control pilot study to identify genomic regions associated with breast cancer. We included 30 randomly sampled incident invasive breast cancer cases aged <45 years without deleterious mutations in the BRCA1 or BRCA2 genes, and 30 population controls individually matched on age, ethnicity and geographical area. The overall genotype call rate was 97.13+/-1.33% for controls and 97.48+/-1.42% for cases. Comparison was made between cases and controls for 203,477 genotyped SNPs using (a) unconditional logistic regression (ULR), (b) conditional logistic regression (CLR) models with adjustment for the matched pairs, (c) allelic tests for single marker tests and (d) haplotype trend regression (HTR). Genomic control and EIGENSTRAT methods were used for correction of population stratification in appropriate models. We demonstrate the similarity and dissimilarity of results from different statistical analyses. We found several possible significant regions harboring biologically meaningful known candidate genes, such as genes encoding fibroblast growth factor, transforming growth factor, epidermal growth factor, and estrogen synthesis enzymes to be associated with early-onset breast cancer. In single marker analysis, none of the SNPs were statistically significant after correction for multiple testing. However, haplotype association tests, using 90730 tag-SNPs, suggested two regions in GLG1 and UGT1 genes retaining significance even after Bonferroni correction. Nevertheless, without systematic replication, findings from this pilot study, especially the associations of breast cancer in relation to specific SNPs, should be interpreted with great caution.

Expression of the JAG1 Notch ligand has previously been shown to correlate with poor overall survival in women with advanced breast cancer. We undertook to test whether expression of JAG1 is associated with reduced disease free survival (DFS) in 887 samples from a prospectively accrued LNN cohort with a median follow-up greater than 8 years. Moderate to high JAG1 mRNA expression was associated with reduced DFS in univariate analysis (hazard ratio of 1.58; 95% confidence interval, 1.03-2.40; P = 0.034) and correlated with large tumor size, ER and PgR negativity, high tumor grade, and p53 antibody reactivity. Although elevated risk of reduced DFS in patients with high JAG1 mRNA did not persist with adjustment for other prognostic factors, it did in combination with HER2. JAG1 mRNA was positively associated with expression of basal breast cancer markers, however, in contrast to the finding that basal gene expression is most strongly associated with reduced DFS in the first 36 months of follow-up, JAG1 mRNA expression was associated with reduced DFS through the full follow-up period. Also, tumors expressing high levels of both mRNA and protein showed reduced DFS as compared to all other groups in univariate analysis (hazard ratio of 1.73; 95% confidence interval, 1.09-2.74; P = 0.020). Thus, JAG1 expression is associated with poor DFS in LNN breast cancer. As JAG1 is a target of several oncogenic signaling pathways, and is a ligand for Notch, these data provide novel insights into signaling that may contribute to progression of early stage breast cancer.

Moderate alcohol intake has been consistently associated with a modest (30-50%) increase in breast cancer risk, but it remains unclear if certain individuals have higher susceptibility to the harmful effects of alcohol intake. Individuals differ in their ability to metabolize alcohol through genetic differences in alcohol dehydrogenase (ADH), the enzyme that catalyzes the oxidation of approximately 80% of ethanol to acetaldehyde, a known carcinogen. Using data from the Breast Cancer Family Registry (n = 811 sister sets), we examined whether sisters with breast cancer differ with respect to alcohol consumption and alcohol metabolism (measured by polymorphisms in ADH1B and ADH1C) compared to their sisters without breast cancer. Neither alcohol drinking nor alcohol metabolizing ADH1B and ADH1C genotypes were associated with breast cancer risk. However, only 19% and 42% of sisters were discordant by ADH1B and ADH1C, respectively, and even fewer were discordant by both genotype and alcohol intake, making it difficult to detect differences if they existed.

Alterations of TP53 and ERBB2 have been shown to play important roles in the prognosis of breast cancer. The primary objective of this study is to characterize TP53 mutation types in node negative breast cancer and investigate their prognostic value, alone and in combination with ERBB2 amplification status. TP53 mutational status (exons 2-10) and ERBB2 amplification status were determined in tumor specimens from a prospective cohort of 543 women with node-negative breast cancer. During a median follow-up of 120 months, there were 111 disease recurrences, and 81 disease-related deaths (3 with cancer; 78 from cancer). Of 543 women, 133 (24.5%) carried mutations in exons 4-9 of the TP53 gene. Seventy-one (53.4%) of these mutations were missense; whereas 62 (46.6%) were protein-truncating mutations. Women whose tumors had missense TP53 mutations were found to be at significantly higher risk of recurrence and death compared to those with wild type TP53, and they also tended to have worse prognosis compared to those with truncating mutations. Those with short truncated proteins tended to have good prognosis compared to those with long truncated proteins, but the risk of recurrence and death did not differ between those whose tumors exhibited conserved versus non-conserved mutations. Missense mutations, in combination with ERBB2 amplification, were observed in 4.6% of the tumors and dramatically affected the disease-specific survival (DSS) and disease-free survival (DFS) of the breast cancer patients. Our study suggests that the type of TP53 mutation, especially missense mutation, is a strong prognostic indicator for DFS and DSS in node-negative breast cancer, particularly in combination with ERBB2 amplification.

Purpose: The objective of this study was to examine the association of: (i) diagnosis at age

p53 mutation is a common event in sporadic breast cancer being found in 15-50% of invasive carcinomas. The purpose of this study was to determine the earliest histologic stage at which p53 mutation could be detected with a widely used anti-p53 antibody (DO7, Novocastra) which recognizes both wild type and mutant forms. p53 expression was assessed immunohistochemically in 12 primary breast carcinomas with known p53 mutations and in all pre-malignant epithelial lesions surrounding these invasive cancers. Strong p53 nuclear staining was found in all of the tumors known to have missense mutations and none of the tumors with truncation mutations. In cases with intense staining in the invasive carcinoma, a similar quality of staining was also seen in all areas of DCIS (ductal carcinoma in situ) and was representative of missense p53 mutations. Lighter nuclear staining intensity was observed in up to 40% of cells in areas of hyperplasia and in up to 30% of normal breast lobules irrespective of the type of mutation found in the invasive carcinoma. This weak staining was not specific to mutated p53 and may indicate increased amounts of normal p53 protein. We conclude that p53 inactivation occurs prior to invasion in breast carcinogenesis, with mutations being uniformly identified in DCIS associated with p53-mutated invasive carcinomas. In contrast, there is no evidence that epithelial hyperplasia or epithelial cells of the terminal duct lobular unit harbor the same mutations as their associated invasive carcinoma.