Nashville Breast Health Study Research Articles

Multiplex immunohistochemistry and high-throughput image analysis for evaluation of spatial tumor immune cell markers in human breast cancer.

The clinicopathological significance of spatial tumor-infiltrating lymphocytes (TILs) subpopulations is not well studied due to lack of high-throughput scalable methodology for studies with large human sample sizes. Establishing a cyclic fluorescent multiplex immunohistochemistry (mIHC/IF) method coupled with computer-assisted high-throughput quantitative analysis to evaluate associations of six TIL markers (CD3, CD8, CD20, CD56, FOXP3, and PD-L1) with clinicopathological factors of breast cancer. Our 5-plex mIHC/IF staining was shown to be reliable and highly sensitive for labeling three biomarkers per tissue section. Through repetitive cycles of 5-plex mIHC/IF staining, more than 12 biomarkers could be detected per single tissue section. Using open-source software CellProfiler, the measurement pipelines were successfully developed for high-throughput multiplex evaluation of intratumoral and stromal TILs. In analyses of 188 breast cancer samples from the Nashville Breast Health Study, high-grade tumors showed significantly increased intratumoral CD3+CD8+ cytotoxic T lymphocyte density (P= 0.0008, false discovery rate (FDR) adjusted P= 0.0168) and intratumoral PD-L1 expression (P= 0.0061, FDR adjusted P= 0.0602) compared with low-grade tumors. The high- and low-grade breast cancers exhibit differential immune responses which may have clinical significance. The multiplexed imaging quantification strategies established in this study are reliable, cost-efficient and applicable in regular laboratory settings for high-throughput tissue biomarker studies, especially retrospective and population-based studies using archived paraffin tissues.

Abstract C075: Association of obesity with breast cancer subtypes among non-Hispanic Black women

Abstract Although non-Hispanic White and non-Hispanic Black women have similar breast cancer incidence rates, Black women are more likely to be diagnosed with aggressive or late-stage disease and have higher mortality rates compared to White women. Clinical studies have demonstrated that tumors lacking estrogen receptor (ER), progesterone receptor (PR) and/or human epidermal growth factor receptor 2 (HER2) (referred to as “triple negative breast cancers” or TNBC), are associated with an aggressive pathology and poor prognosis and distinct risk factor profiles, compared to those with receptor positivity. Through the current study, we examined the effect of obesity in the development of breast cancer subtypes; specifically, ER- tumors and TNBC. To examine the association between body mass index (BMI) and breast cancer subtype, we utilized a pooled dataset of Black women with breast cancer from four studies: Black Women Etiology and Survival of Triple-negative Breast Cancers Study, Southern Community Cohort Study, Nashville Breast Health Study and the Tri-State Breast Study. For all four studies, information on tumor ER, PR and HER2 status was abstracted from state cancer registry records and supplemented by available pathology reports and medical records. We calculated BMI as kg/m2, based on self-reported weight and height, and classified it as normal (18.5-<25), overweight (25-<30), obese I (30-<35), obese II (35-<40) and obese III (40+). Case-only logistic regression models were used to estimate odds ratios (ORs) and 95% confidence intervals (CIs) for the association between obesity and breast cancer subtype, using ER+ as the reference group for ER- and TNBC. Since the association between obesity and breast cancer has been shown to differ between pre- and postmenopausal women, all analyses were stratified by menopausal status at breast cancer diagnosis. Approximately 70% of the 2,801 women included in the study were postmenopausal at breast cancer diagnosis. Overall, 69% of the tumors were ER+, with similar proportions among pre- and post-menopausal women. In total, 25% were TNBC, of which a third were among pre-menopausal patients. In multivariable analyses, using premenopausal women with ER+ breast cancer as the reference group, those with ER- breast cancer (OR 2.16, 95% CI 1.20-3.88) or TNBC (OR 2.13, 95% CI 1.06-4.28) were more likely to be in the obese II category. Among postmenopausal women, there was no clear association between obesity and breast cancer subtype, although the obese class I category was non-significantly associated with lower risk of TNBC compared to ER+ subtype (OR 0.79, 95% CI 0.53-1.17). Our study represents one of the largest studies to assess the differential impact of obesity on breast cancer subtypes among Black women. Our findings suggest that higher BMI is associated with increased risk of TNBC for premenopausal women and demonstrates that premenopausal obese Black women are more likely to be diagnosed with ER- and TNBC compared with ER+ tumors. Citation Format: Jaleesa Moore, Maureen Sanderson, Tuya Pal, Mary Kay Fadden, Steffie-Ann Dujon, Sonya Reid, Anne Tezak, Loren Lipworth. Association of obesity with breast cancer subtypes among non-Hispanic Black women [abstract]. In: Proceedings of the Twelfth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2019 Sep 20-23; San Francisco, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2020;29(6 Suppl_2):Abstract nr C075.

Gene expression in triple-negative breast cancer in relation to survival

The identification of biomarkers related to the prognosis of triple-negative breast cancer (TNBC) is critically important for improved understanding of the biology that drives TNBC progression. We evaluated gene expression in total RNA isolated from formalin-fixed paraffin-embedded tumor samples using the NanoString nCounter assay for 469 TNBC cases from the Shanghai Breast Cancer Survival Study. We used Cox regression to quantify Hazard Ratios (HR) and corresponding confidence intervals (CI) for overall survival (OS) and disease-free survival (DFS) in models that included adjustment for breast cancer intrinsic subtype. Of 302 genes in our discovery analysis, 22 were further evaluated in relation to OS among 134 TNBC cases from the Nashville Breast Health Study and the Southern Community Cohort Study; 16 genes were further evaluated in relation to DFS in 335 TNBC cases from four gene expression omnibus datasets. Fixed-effect meta-analysis was used to combine results across data sources. Twofold higher expression of EOMES (HR 0.90, 95% CI 0.83-0.97), RASGRP1 (HR 0.89, 95% CI 0.82-0.97), and SOD2 (HR 0.80, 95% CI 0.66-0.96) was associated with better OS. Twofold higher expression of EOMES (HR 0.89, 95% CI 0.81-0.97) and RASGRP1 (HR 0.87, 95% CI 0.81-0.95) was also associated with better DFS. On the contrary, a doubling of FA2H (HR 1.14, 95% CI 1.06-1.22) and GSPT1 (HR 1.33, 95% CI 1.14-1.55) expression was associated with shorter DFS. We identified five genes (EOMES, FA2H, GSPT1, RASGRP1, and SOD2) that may serve as potential prognostic biomarkers and/or therapeutic targets for TNBC.

Abstract 2272: Association of G-protein estrogen receptor (GPER) in primary breast cancer in Caucasian and Black women by tumor immunophenotype and menopausal status

Abstract Background: Assignment of breast cancer therapies is largely defined by immunohistochemical analyses that evaluate steroid hormone and growth factor receptor status in primary tumor biopsies. Patients whose tumors are devoid of these receptors have fewer treatment options, and poor survival. This outcome is more commonly measured among Black versus Caucasian patients, and in patients with intact ovarian function. The G-protein coupled estrogen receptor, GPER, holds significance for breast cancer biology and treatment as 17β-estradiol and ER antagonists trigger the release of epidermal growth factor (EGF)-ligands and GPER associates directly with clinicopathological factors that predict advanced disease. Experimental design: GPER was measured by immunohistochemical analysis and a semi-quantitative scoring method in 521 primary breast tumors obtained from the NCI Cooperative Breast Cancer Tissue Resource and the Nashville Breast Health Study, including tumor specimens derived from 389 Caucasian and 108 Black patients. GPER was correlated with disease progression variables and evaluated by race, menopausal status and tumor immunophenotype. Results: GPER is expressed in more than two-thirds of invasive breast carcinomas in this study with no difference observed between Caucasian and Black women (p = 0.69). Coexpression of GPER and ER was measured in less than half of all tumors (48%). Additionally, GPER is commonly expressed in ER-negative tumors (107/171; 62%) and in triple negative breast cancer (TNBC) (42/56; 75%) and did not differ by race for this newly defined measure of estrogen responsiveness (p = 0.42). Moreover, while ER-negative tumors are more common in pre- (79/153; 52%) versus postmenopausal women (92/336; 27%), GPER expression is not influenced by menopausal status. GPER is positively associated with tumor size (particularly in premenopausal women) in both studies, and with frank metastases at first diagnosis in the NCI population. No significant association was seen with nodal invasion. Survival data in NBHS patients found that women who died of breast cancer are more likely to have GPER-positive tumors than survivors. Conclusion: GPER is expressed in the majority of invasive breast tumors (> 60%) at diagnosis and is commonly retained in patients with ER-negative or triple-negative disease regardless of race or menopause. Unlike ER, which varies inversely with variables that predict advanced disease, GPER is positively associated with these same prognostic parameters. These data support GPER as an independent marker of estrogen responsiveness and breast cancer progression. Most importantly, our results suggest that GPER represents a promising therapeutic target since its expression is constant across multiple types of invasive breast cancer, including patients with TNBC and in tumors from pre- and postmenopausal women. Citation Format: Sandra L. Deming-Halverson, Carl Graeber, Jason Machan, Edmond Sabo, Wei Zheng, Edward J. Filardo. Association of G-protein estrogen receptor (GPER) in primary breast cancer in Caucasian and Black women by tumor immunophenotype and menopausal status. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2272.

ALDH1A1 mRNA expression in association with prognosis of triple-negative breast cancer

ALDH1 is a crucial element in the retinoic acid signaling pathway regulating the self-renewal and differentiation of normal stem cells, and may play an important role in cancer progression. However, research on ALDH1 gene expression and breast cancer prognosis has yielded conflicting results. We evaluated the association between tumor tissue ALDH1A1/ALDH1A3 mRNA expression and triple-negative breast cancer (TNBC) prognosis in the Shanghai Breast Cancer Survival Study (SBCSS, N=463), Nashville Breast Health Study (NBHS, N=86), and Southern Community Cohort Study (SCCS, N=47). Gene expression was measured in RNA isolated from breast cancer tissues. In the SBCSS, higher ALDH1A1 mRNA level was associated with improved disease-free (HR=0.87, 95% CI: 0.80-0.95, per log unit change) and overall survival (HR=0.85, 95% CI: 0.78-0.93 per log unit change) independent of age at diagnosis, TNM stage and treatment. We replicated the findings for overall survival in the NBHS and SCCS (HR = 0.27, 95% CI: 0.10-0.73) and for disease-free survival by a meta-analysis of four publicly-available gene expression datasets (HR = 0.86, 95% CI: 0.76-0.97). No significant association was found for ALDH1A3.Our study suggests high expression of ALDH1A1 mRNA in tumor tissues may be an independent predictor of a favorable TNBC outcome.

Abstract 2773: Association of ALDH1A1 gene expression with survival of triple-negative breast cancer

Abstract Aldehyde dehydrogenases (ALDH) are a family of enzymes that catalyze aldehyde into carboxylic acids via the NAD(P)+-dependent oxidation. Several ALDH family members have been identified in humans, including ALDH1A1, ALDH1A3, ALDH2, ALDH3A1, and ALDH4A1. ALDH1A1 is a crucial element in the retinoic acid signaling pathway regulating the self-renewal and differentiation of normal stem cells. It ALDH1A1 has been suggested as one of breast cancer stem cell markers and may play an important role in breast cancer prognosis. However, previous research on ALDH1A1 and breast cancer prognosis has yielded conflicting results. We analyzed the association between mRNA expression of ALDH1A1 in tumor tissues and survival of triple-negative breast cancer (TNBC: ER-/PR-/HER2-) using data and samples from three cohorts of breast cancer patients, including 469 cases from the Shanghai Breast Cancer Survival Study (SBCSS), 86 cases from the Nashville Breast Health Study (NBHS), and 47 cases from the Southern Community Cohort Study (SCCS). Gene expression levels were measured in total RNA isolated from microdissected archival formalin-fixed paraffin-embedded breast cancer tissues using Nanostring nCounter assays. The associations between ALDH1A1 mRNA expression levels and recurrence/breast cancer mortality and overall mortality were evaluated by multivariate survival analysis using the Cox regression model. Data from the SCCS and NBHS were combined in the analysis because both studies were conducted among US women and had a small sample size. The levels of ALDH1A1 mRNA expression were consistently and positively associated with disease free and overall survival independent of age at diagnosis and TNM stage in both Chinese and US women with TNBC. Compared to those with ALDH1A1 mRNA expression below the median, TNBC patients with ALDH1A1 mRNA expression above the median had a reduced risk of recurrence/breast cancer mortality (HR = 0.60, 95% CI: 0.39-0.92) and overall mortality (HR = 0.70, 95% CI: 0.47-1.05) in the SBCSS and reduced overall mortality in the SCCS/NBHS (HR = 0.27,95% CI: 0.10-0.74). Additional analyses of the SBCSS cohort stratified by basal-like breast cancer subtype showed that ALDH1A1 mRNA expression was similarly associated with reduced risk of recurrence/breast cancer mortality and total mortality in both basal-like and non-basal like TNBC patients, although the point estimates were not significant due to the smaller sample size. In addition, we found that patients with a higher tumor grade had a lower level of ALDH1A1 mRNA expression but did not observe an association with TNM stage. Our study suggests that the mRNA expression of ALDH1A1 in tumor tissue may be associated with favorable clinical outcomes in patients with TNBC. Further investigations are needed to understand the biological mechanism(s) underlying the observed association. Citation Format: Yan Liu, Qiuyin Cai, Ying Zheng, Michelle L. Baglia, Yinghao Su, Sarah Nechuta, Ping-Ping Bao, William Blot, Wei Zheng, Xiao-Ou Shu. Association of ALDH1A1 gene expression with survival of triple-negative breast cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2773. doi:10.1158/1538-7445.AM2015-2773

Energy-Related Indicators and Breast Cancer Risk among White and Black Women.

Energy-related indicators, including physical activity, energy intake, body mass index (BMI) and adult weight change, have been linked to breast cancer risk. Very few studies of these associations have been conducted among black women, therefore we used the Nashville Breast Health Study (NBHS) to determine whether similar effects were seen in black and white women. The NBHS is a population-based case-control study of breast cancer among women age 25 to 75 years conducted between 2001 and 2010 in and around the Nashville Metropolitan area. Telephone interviews and self-administered food frequency questionnaires were completed with 2,614 incident breast cancer cases ascertained through hospitals and the statewide cancer registry, and 2,306 controls selected using random digit dialing. Among premenopausal white and black women, there was little effect of adult exercise or other energy-related indicators on breast cancer risk, regardless of tumor estrogen receptor (ER) status. The beneficial effect of adult exercise on postmenopausal breast cancer appeared to be comparable between white and black women (highest tertile relative to none - white odds ratio [OR] 0.8, 95% confidence interval [CI] 0.6-1.0, p for trend=0.05; black OR 0.7, 95% CI 0.4-1.1, p for trend=0.07); however, among black women the reduction was limited to those with ER-positive disease. White and black women should be encouraged to engage in more physical activity to reduce their risk of postmenopausal breast cancer.

Abstract B30: Indicators of energy balance and breast cancer risk among white and black women

Abstract Introduction: Physical activity has been linked to reduced breast cancer risk, especially among women diagnosed postmenopausally. Since the majority of studies of this association have been conducted in white women, we used the Nashville Breast Health Study (NBHS) to determine whether similar effects were seen in black women. Methods: The NBHS is a population-based case-control study of breast cancer among women age 25 to 75 conducted between 2001 and 2010 in and around the Nashville Metropolitan area. Telephone interviews were completed with 1,866 incident breast cancer cases ascertained through hospitals and the statewide cancer registry, and 2,306 randomly selected controls (with respective response rates of 58% and 79%). Results: A greater percentage of white (premenopausal 59%, postmenopausal 54%) than black (premenopausal 51%, postmenopausal 43%) women reported they had engaged in leisure-time physical activity or exercise in the past 10 years. Among premenopausal white and black women, there was little effect of exercise on breast cancer risk; however, occupational activity, defined as standing and walking at work, was associated with an increased risk of breast cancer. After adjusting for confounding and dividing the group of women who had engaged in exercise into tertiles (MET hours per day), there was a suggested reduction in postmenopausal breast cancer in women of both races (highest tertile relative to none - white odds ratio [OR] 0.8, 95% confidence interval [CI] 0.6-1.0, p for trend=0.058; black OR 0.7, 95% CI 0.4-1.1, p for trend=0.098). Similar point estimates were found for occupational activity. When considering the joint effects of caloric intake, obesity and weight gain since age 18 on the association of exercise and breast cancer, the greatest effect was seen for weight gain since 18 with increased risks in most categories relative to women in the highest tertile of exercise who had lost or maintained weight. This pattern was most pronounced among white women diagnosed postmenopausally, but among black women diagnosed premenopausally. Discussion: Our results indicate that although postmenopausal black women report engaging in very little leisure-time physical activity, its' effect on breast cancer risk is comparable to white women. However, the joint effect of adult weight gain on the exercise and breast cancer association may differ by race and menopausal status. White and black women should be encouraged to engage in leisure-time physical activity regardless of the amount of work- and housework-related physical activity done to help reduce their risk of breast cancer and other chronic diseases. Citation Format: Maureen Sanderson, Sandra L. Deming-Halverson, Alecia M. Fair, Martha J. Shrubsole, Alicia Beeghly-Fadiel, Sarah Nechuta, Yong Cui, David Shen-Miller, Heather O'Hara, Nia Foderingham, Xiao-Ou Shu, Wei Zheng. Indicators of energy balance and breast cancer risk among white and black women. [abstract]. In: Proceedings of the Sixth AACR Conference: The Science of Cancer Health Disparities; Dec 6–9, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2014;23(11 Suppl):Abstract nr B30. doi:10.1158/1538-7755.DISP13-B30

Abstract 3272: Association between estrogen-metabolizing genetic risk scores and breast cancer risk

Abstract Purpose: Estrogens are metabolized by phase I enzymes to intermediate metabolites, which are conjugated by phase II enzymes to be excreted. Differences in estrogen metabolism by phase I and phase II enzymes may contribute to breast cancer risk. The levels and activities of these enzymes are controlled, in part, by genetic polymorphisms. We evaluated three genetic risk scores constructed to reflect the contributions of phase I, phase II and combined phase I/II estrogen-metabolizing activity in association with breast cancer risk. Methods: Data arise from 1296 White breast cancer cases and 1295 White controls enrolled in the Nashville Breast Health Study, a population-based case-control study conducted during 2001-2008. Telephone interviews obtained data on breast cancer risk factors. Saliva samples were obtained to determine genotypes for 12 functional variants in genes involved in estrogen metabolism (AHR, CYP1A1, CYP1A2, CYP3A4, NQO1, COMT, SULT1A1, UGT2B17, GSTM1 and GSTT1). Risk scores were formed according to the hypothesis that women with high phase I and II estrogen metabolizing profiles were at lower breast cancer risk, irrespective of menopausal status. Risk alleles in single nucleotide polymorphisms (SNPs) and copy number variants (CNVs) were summed to form risk scores containing phase I, phase II or combined phase I/II genetic variants. The combined phase I/II estrogen metabolizing score used subjects in the quartile for highest phase II estrogen metabolizing as the reference group. All other participants were separated at the median according to phase I enzyme activity. Adjusted odds ratios (OR) and 95% confidence intervals (CI) were calculated using logistic regression to test associations between genetic scores and breast cancer risk. Likelihood ratio tests p-values were obtained for interactions between the phase I/II combined score and risk factors that may modify the association between endogenous estrogen metabolism and breast cancer, including age at menarche, menopausal status, age at menopause, body mass index and well-done red meat consumption. Results: Lower phase I and phase II estrogen metabolizing activity scores were associated with non-significant increased breast cancer risk. Women with low phase I/II estrogen metabolizing risk scores had an increased OR of 1.22 (CI: 1.01-1.47) compared to participants with high phase II estrogen metabolizing scores. Lower copy numbers of the UGT2B17 variant were associated with increased risk (OR: 1.16, CI: 1.03-1.30). No other individual CNVs or SNPs were associated with breast cancer risk. Effect modification was not observed by other risk factors (all P>0.05). Conclusions: Lower phase I and phase II metabolism activity as measured by functional genetic variants is associated with a modest increase in breast cancer risk; this relationship is not modified by established breast cancer risk factors. Citation Format: Shaneda N. Warren Andersen, Guoliang Li, Qiuyin Cai, Alicia Beeghly-Fadiel, Martha J. Shrubsole, Xiao-Ou Shu, Wei Zheng. Association between estrogen-metabolizing genetic risk scores and breast cancer risk. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3272. doi:10.1158/1538-7445.AM2014-3272

Associations of Hormone-Related Factors With Breast Cancer Risk According to Hormone Receptor Status Among White and African American Women

BackgroundCauses of racial disparities in breast cancer incidence and mortality between white and African American women remain unclear. This study evaluated associations of menstrual and reproductive factors with breast cancer risk by race and cancer subtypes. Patients and MethodsIncluded in the study were 1866 breast cancer cases and 2306 controls recruited in the Nashville Breast Health Study, a population-based case-control study. Multivariable logistic regression was used to estimate odds ratios (ORs) and 95% confidence intervals (CIs). ResultsAfrican American women were more likely to have estrogen receptor-negative (ER−), progesterone receptor-negative (PR−), and triple-negative (ER−PR−HER2−) breast cancer than white women. Age at menarche (≥ 14 years) and multiparity (≥ 3 live births) were inversely associated with ER+ tumors only, whereas late age at first live birth (> 30 years) and nulliparity were associated with elevated risk; such associations were predominantly seen in white women (OR = 0.70, 95% CI = 0.55-0.88; OR = 0.72, 95% CI = 0.56-0.92; OR = 1.42, 95% CI = 1.13-1.79; OR = 1.32, 95% CI = 1.06-1.63, respectively). Age at menopause between 47 and 51 years was associated with elevated risk of ER− tumors in both white and African American women. Among women who had natural menopause, positive association between ever-use of hormone replacement therapy and breast cancer risk was seen in white women only (OR = 1.39, 95% CI = 1.03-1.87). ConclusionThis study suggests that certain hormone-related factors are differentially associated with risk of breast cancer subtypes, and these associations also differ by race.

Interactions of Hormone Replacement Therapy, Body Weight, and Bilateral Oophorectomy in Breast Cancer Risk

To examine potential modifying effects of body weight and bilateral oophorectomy on the association of hormone replacement therapy (HRT) with risk of breast cancer, overall and by subtypes according to status of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (Her2) among postmenopausal women. This analysis included 2,510 postmenopausal white women recruited in the Nashville Breast Health Study, a population-based case-control study of breast cancer. Multivariable logistic regression was used to estimate ORs and 95% confidence intervals (CI) for associations between HRT use and risk of breast cancer overall and by subtypes, adjusted for age and education. Among women with natural menopause and body mass index (BMI) < 25 kg/m(2), ever-use of HRT was associated with increased breast cancer risk (OR, 1.95; 95% CI, 1.32-2.88). Risk was elevated with duration of HRT use (P for trend = 0.002). Similar association patterns were found for ER(+), ER(+)PR(+), and luminal A cancer subtypes but not ER(-), ER(-)PR(-), and triple-negative cancer. In contrast, ever-HRT use in overweight women (BMI ≥ 25 kg/m(2)) showed no association with risk of breast cancer overall or by subtypes; interaction tests for modifying effect of BMI were statistically significant. Ever-HRT use was associated with decreased breast cancer risk (OR, 0.70; 95% CI, 0.38-1.31) among women with prior bilateral oophorectomy but elevated risk (OR, 1.45; 95% CI, 0.92-2.29) among those with hysterectomy without bilateral oophorectomy (P for interaction = 0.057). Similar associations were seen for virtually all breast cancer subtypes, although interaction tests were statistically significant for ER(+) and luminal A only. Body weight and bilateral oophorectomy modify associations between HRT use and breast cancer risk, especially the risk of hormone receptor-positive tumors.

Evaluating 17 breast cancer susceptibility loci in the Nashville breast health study

Genome-wide association studies have discovered multiple genetic loci associated with breast cancer risk. Investigating these loci would be helpful to evaluate previous findings and identify causal variants for breast cancer. We evaluated index SNPs in 17 of these loci in a study of 1,511 cases and 1,454 controls of European descent. We investigated the overall association with breast cancer and among subtypes defined as ER+ (estrogen receptor positive), ER- (estrogen receptor negative) and triple-negative breast cancer (TNBC). Combined effects of SNPs on breast cancer risk were assessed via a genetic risk score. We evaluated the contribution of both genetic variants and traditional risk factors to a breast cancer risk assessment model. Five of the 17 SNPs were significantly associated (P≤0.05) with overall breast cancer in the same direction as previously reported: rs13387042 (2q35/TNP1), rs4973768 (3p24/SLC4A7), rs2046210 (6q25/ESR1), rs1219648 (10q26/FGFR2), and rs4784227 (16q12/TOX3). When stratified by breast cancer subtype, all five SNPs were associated (P<0.05) with ER+ cancer, three with ER- cancer (rs13387042, rs1219648, and rs4784227), and one with TNBC (rs1219648). A GRS, based on those five significant SNPs, showed strong association with overall breast cancer with ORs (95% CI) of 1.48 (1.22-1.79), 1.85 (1.52-2.25) and 2.26 (1.82-2.80), respectively, for each quartile, (P=2.0×10(-15)). Traditional risk factors, including previous benign breast disease, breast cancer family history and parity, were significantly associated with breast cancer risk in the present study. These factors, together with the GRS, were used to build a breast cancer risk assessment model with a c statistic of 0.6321 from receiver operating characteristic analysis. The contribution of the GRS to the model was greater than prior benign breast disease, family history and parity with the c statistic change of 0.0374, 0.0324, 0.0103, 0.0012, respectively. Our study demonstrates that five SNPs were associated with overall breast cancer, with stronger association for ER+ than ER- cancer as previously reported, and suggests that a risk assessment model incorporating the GRS from five loci is useful in identifying women at high risk of breast cancer.

APOBEC3 deletion polymorphism is associated with breast cancer risk among women of European ancestry

Copy number variations occur frequently in the genome and are a significant source of human genetic variation accounting for disease. Recently, we discovered a common deletion located in the APOBEC3A and APOBEC3B genes significantly associated with breast cancer in Chinese women. Investigating this locus in other populations would be an expedient way to evaluate the generalizability of the novel finding. We analyzed the APOBEC3 deletion in a large study of 3273 European-ancestry women (including 1671 breast cancer cases and 1602 controls) from the population-based Nashville Breast Health Study. All participants were genotyped using real-time qualitative PCR. Logistic regression was used to derive odds ratios (ORs) and 95% confidence intervals (CIs) for the associations between the deletion polymorphism and breast cancer risk. The APOBEC3 deletion was observed in 12.4% of cases and 10.4% of controls. The deletion was significantly associated with breast cancer risk, with ORs and 95% CIs of 1.21 (1.02-1.43) associated with one-copy deletion and 2.29 (1.04-5.06) associated with two-copy deletion compared with women with no deletion (P for trend = 0.005). The positive association of the APOBEC3 deletion with breast cancer risk was similar for estrogen receptor-positive and estrogen receptor-negative breast cancer and was not modified by known breast cancer risk factors. Results from this study confirmed the association of the APOBEC3 deletion with breast cancer risk among women of European ancestry.

Evaluating genome-wide association study-identified breast cancer risk variants in African-American women.

Genome-wide association studies (GWAS), conducted mostly in European or Asian descendants, have identified approximately 67 genetic susceptibility loci for breast cancer. Given the large differences in genetic architecture between the African-ancestry genome and genomes of Asians and Europeans, it is important to investigate these loci in African-ancestry populations. We evaluated index SNPs in all 67 breast cancer susceptibility loci identified to date in our study including up to 3,300 African-American women (1,231 cases and 2,069 controls), recruited in the Southern Community Cohort Study (SCCS) and the Nashville Breast Health Study (NBHS). Seven SNPs were statistically significant (P≤0.05) with the risk of overall breast cancer in the same direction as previously reported: rs10069690 (5p15/TERT), rs999737 (14q24/RAD51L1), rs13387042 (2q35/TNP1), rs1219648 (10q26/FGFR2), rs8170 (19p13/BABAM1), rs17817449 (16q12/FTO), and rs13329835 (16q23/DYL2). A marginally significant association (P<0.10) was found for three additional SNPs: rs1045485 (2q33/CASP8), rs4849887 (2q14/INHBB), and rs4808801 (19p13/ELL). Three additional SNPs, including rs1011970 (9p21/CDKN2A/2B), rs941764 (14q32/CCDC88C), and rs17529111 (6q14/FAM46A), showed a significant association in analyses conducted by breast cancer subtype. The risk of breast cancer was elevated with an increasing number of risk variants, as measured by quintile of the genetic risk score, from 1.00 (reference), to 1.75 (1.30–2.37), 1.56 (1.15–2.11), 2.02 (1.50–2.74) and 2.63 (1.96–3.52), respectively, (P = 7.8×10–10). Results from this study highlight the need for large genetic studies in AAs to identify risk variants impacting this population.

Abstract 2616: Association of UGT2B17 gene copy number variation with breast cancer risk: The Nashville Breast Health Study

Abstract UDP-glucuronosyltransferase 2B17 (UGT2B17) catalyzes the transfer of glucuronic acid from uridine diphosphoglucuronic acid to a variety of substrates, including estrogen, a hormone that plays an important role in the etiology of breast cancer. Glucuronidated estrogens are more polar and water soluble; thus, they are more easily excreted in the bile and urine. The UGT2B17 gene displays variations in copy number (0 copies, 1 copy, and 2 copies), and the null genotype (0 copies) is associated with lower glucuronidating activity in human liver microsomes. In this study, we evaluated the association of UGT2B17 gene copy number variation with breast cancer risk. Included in the study were 1,536 breast cancer cases and 1,205 matched controls recruited into the Nashville Breast Health Study, a population-based, case-control study based in Nashville, Tennessee. Copy number of the UGT2B17 gene was evaluated by real-time PCR by using ABI's TaqMan Copy Number Assays reagents. UGT2B17 gene copy number variation was associated with breast cancer risk with odds ratios of 1.17 (95% CI: 1.00-1.38) and 1.33 (95% CI: 1.02-1.74), respectively, for 1 and 0 copies of the UGT2B17 gene, compared with 2 copies of the gene. The association varied little by menopausal status. The association was consistent in both whites and African Americans. Our study suggests that UGT2B17 gene copy number variation may play a role in the etiology of breast cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2616. doi:1538-7445.AM2012-2616

Well-done meat intake and meat-derived mutagen exposures in relation to breast cancer risk: the Nashville Breast Health Study

Previous studies of the association of meat intake and meat-derived mutagen exposure with breast cancer risk have produced inconsistent results. We evaluated this association in a population-based case-control study of incident breast cancer conducted in Nashville, Tennessee, United States, including 2,386 breast cancer cases and 1,703 healthy women controls. Telephone interviews were conducted to obtain information related to meat intake including amount, cooking methods, and doneness levels, as well as other known or hypothesized risk factors for breast cancer. Unconditional logistic regression was used to derive odds ratios (ORs) after adjusting for potential confounders. High intake of red meat was associated with a significantly elevated risk of breast cancer (P-trend < 0.001). The association was particularly strong for high intake of well-done red meat (P-trend < 0.001), with an adjusted OR of 1.5 (95% CI = 1.3-1.9) for the highest versus the lowest quartile. Associations between red meat and breast cancer risk were slightly stronger for postmenopausal women than for premenopausal women. Meat-derived mutagens such as 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline and 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline, were significantly associated with increased breast cancer risk among postmenopausal women only (P-trend = 0.002 and 0.003, respectively). The results from this study provide strong support for the hypotheses that high red meat intake and meat-derived mutagen exposure may be associated with an increase in breast cancer risk.

Abstract 3718: Well-done meat intake and meat-derived mutagen exposures in relation to breast cancer risk: The Nashville Breast Health Study

Abstract Previous studies of the association of meat intake and meat-derived mutagen exposure with breast cancer risk have produced inconsistent results. We evaluated this association in a population-based case-control study of incident breast cancer conducted in Nashville,TN, United States, including 2,386 breast cancer cases and 1,703 healthy women controls. Telephone interviews were conducted to obtain information related to meat intake including amount, cooking methods, and doneness levels, as well as other known or hypothesized risk factors for breast cancer. Unconditional logistic regression was used to derive odds ratios (ORs) after adjusting for potential confounders. High intake of red meat was associated with a significantly elevated risk of breast cancer (P-trend &amp;lt;0.001). The association was particularly strong for high intake of well-done red meat (P-trend &amp;lt;0.001), with an adjusted OR of 1.5 (95% CI = 1.3-1.9) for the highest versus the lowest quartile. Associations between red meat and breast cancer risk were stronger for postmenopausal women than for premenopausal women. Meat-derived mutagens such as 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) and 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline (DiMeIQx), were significantly associated with increased breast cancer risk among postmenopausal women only (P-trend = 0.002 and 0.003, respectively). The results from this study provide strong support for the hypotheses that high red meat intake and meat-derived mutagen exposure may be associated with an increase in breast cancer risk. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3718. doi:10.1158/1538-7445.AM2011-3718