Michelle Roberts

Conducting Molecular Epidemiological Research in the Age of HIPAA: A Multi-Institutional Case-Control Study of Breast Cancer in African-American and European-American Women

Breast cancer in African-American (AA) women occurs at an earlier age than in European-American (EA) women and is more likely to have aggressive features associated with poorer prognosis, such as high-grade and negative estrogen receptor (ER) status. The mechanisms underlying these differences are unknown. To address this, we conducted a case-control study to evaluate risk factors for high-grade ER- disease in both AA and EA women. With the onset of the Health Insurance Portability and Accountability Act of 1996, creative measures were needed to adapt case ascertainment and contact procedures to this new environment of patient privacy. In this paper, we report on our approach to establishing a multicenter study of breast cancer in New York and New Jersey, provide preliminary distributions of demographic and pathologic characteristics among case and control participants by race, and contrast participation rates by approaches to case ascertainment, with discussion of strengths and weaknesses.

Variants in the vitamin D pathway, serum levels of vitamin D, and estrogen receptor negative breast cancer among African-American women: a case-control study.

IntroductionAmerican women of African ancestry (AA) are more likely than European Americans (EA) to have estrogen receptor (ER)-negative breast cancer. 25-hydroxyvitamin D (25OHD) is low in AAs, and was associated with ER-negative tumors in EAs. We hypothesized that racial differences in 25OHD levels, as well as in inherited genetic variations, may contribute, in part, to the differences in tumor characteristics.MethodsIn a case (n = 928)-control (n = 843) study of breast cancer in AA and EA women, we measured serum 25OHD levels in controls and tested associations between risk and tag single nucleotide polymorphisms (SNPs) in VDR, CYP24A1 and CYP27B1, particularly by ER status.ResultsMore AAs had severe vitamin D deficiency (< 10 ng/ml) than EAs (34.3% vs 5.9%), with lowest levels among those with the highest African ancestry. Associations for SNPs differed by race. Among AAs, VDR SNP rs2239186, associated with higher serum levels of 25OHD, decreased risk after correction for multiple testing (OR = 0.53, 95% CI = 0.31-0.79, p by permutation = 0.03), but had no effect in EAs. The majority of associations were for ER-negative breast cancer, with seven differential associations between AA and EA women for CYP24A1 (p for interaction < 0.10). SNP rs27622941 was associated with a > twofold increased risk of ER-negative breast cancer among AAs (OR = 2.62, 95% CI = 1.38-4.98), but had no effect in EAs. rs2209314 decreased risk among EAs (OR = 0.38, 95% CI = 0.20-0.73), with no associations in AAs. The increased risk of ER-negative breast cancer in AAs compared to EAs was reduced and became non-significant (OR = 1.20, 95% CI = 0.80-1.79) after adjusting for these two CYP24A1 SNPs.ConclusionsThese data suggest that genetic variants in the vitamin D pathway may be related to the higher prevalence of ER-negative breast cancer in AA women.

Single-nucleotide polymorphisms in DNA repair genes and association with breast cancer risk in the web study

Base excision repair (BER) and nucleotide excision repair (NER) pathways repair damaged DNA, and polymorphisms in these genes might affect breast cancer susceptibility. We evaluated associations between seven single-nucleotide polymorphisms in four DNA repair genes (ERCC4 rs1799801, XPC rs2227998, rs2228001, rs2228000, OGG1 rs1052133 and XRCC1 rs25487 and rs25486) and breast cancer risk, examining modification by smoking and alcohol consumption, using data from the Western New York Exposures and Breast Cancer Study. Women aged 35-79 years with incident breast cancer (n = 1170) and age- and race-matched controls (n = 2115) were enrolled. Genotyping was performed using matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Unconditional logistic regression was used to estimate odds ratios (OR) and 95% confidence intervals (CIs). No significant associations were observed in premenopausal women. Among postmenopausal women, rs25487 and rs25486 (OR = 1.24; 95% CI 1.01-1.51 and OR = 1.23; 95% CI 1.01-1.49, respectively, for combined heterozygous and homozygous variant compared with reference) were associated with increased risk of breast cancer. Postmenopausal women carrying the variant allele of the synonymous XPC polymorphism (rs2227998) were also at borderline significantly increased risk (OR = 1.24; 95% CI 1.01-1.52, heterozygous variant compared with reference; OR = 1.22; 95% CI 1.01-1.48, for combined heterozygous and homozygous variant compared with reference). There was no evidence of genotype-smoking and genotype-alcohol consumption interactions for pre- and postmenopausal women. These results indicate that some of the variants in BER and NER genes may influence risk of postmenopausal breast cancer.

Cytokine and cytokine receptor genes of the adaptive immune response are differentially associated with breast cancer risk in American women of African and European ancestry.

Disparities in breast cancer biology are evident between American women of African ancestry (AA) and European ancestry (EA) and may be due, in part, to differences in immune function. To assess the potential role of constitutional host immunity on breast carcinogenesis, we tested associations between breast cancer risk and 47 single nucleotide polymorphisms (SNPs) in 26 cytokine‐related genes of the adaptive immune system using 650 EA (n = 335 cases) and 864 AA (n = 458 cases) women from the Womens Circle of Health Study (WCHS). With additional participant accrual to the WCHS, promising SNPs from the initial analysis were evaluated in a larger sample size (1,307 EAs and 1,365 AAs). Multivariate logistic regression found SNPs in genes important for T helper type 1 (Th1) immunity (IFNGR2 rs1059293, IL15RA rs2296135, LTA rs1041981), Th2 immunity (IL4R rs1801275), and T regulatory cell‐mediated immunosuppression (TGFB1 rs1800469) associated with breast cancer risk, mainly among AAs. The combined effect of these five SNPs was highly significant among AAs (P‐trend = 0.0005). When stratified by estrogen receptor (ER) status, LTA rs1041981 was associated with ER‐positive breast cancers among EAs and marginally among AAs. Only among AA women, IL15 rs10833 and IL15RA rs2296135 were associated with ER‐positive tumors, and IL12RB1 rs375947, IL15 rs10833 and TGFB1 rs1800469 were associated with ER‐negative tumors. Our study systematically identified genetic variants in the adaptive immune response pathway associated with breast cancer risk, which appears to differ by ancestry groups, menopausal status and ER status.

Innate Immunity Pathways and Breast Cancer Risk in African American and European-American Women in the Women’s Circle of Health Study (WCHS)

African American (AA) women are more likely than European American (EA) women to be diagnosed with early, aggressive breast cancer. Possible differences in innate immune pathways (e.g., inflammatory responses) have received little attention as potential mechanisms underlying this disparity. We evaluated distributions of selected genetic variants in innate immune pathways in AA and EA women, and examined their associations with breast cancer risk within the Women’s Circle of Health Study (WCHS). In stage I of the study (864 AA and 650 EA women) we found that genotype frequencies for 35 of 42 tested SNPs (18 candidate genes) differed between AAs and EAs (corroborated by ancestry informative markers). Among premenopausal AA women, comparing variant allele carriers to non-carriers, reduced breast cancer risk was associated with CXCL5-rs425535 (OR=0.61, P=0.02), while among EA women, there were associations with TNFA-rs1799724 (OR =2.31, P =0.002) and CRP-rs1205 (OR=0.54, P=0.01). For postmenopausal women, IL1B-rs1143627 (OR=1.80, P=0.02) and IL1B-rs16944 (OR=1.85, P =0.02) were associated with risk among EA women, with significant associations for TNFA-rs1799724 limited to estrogen receptor (ER) positive cancers (OR=2.0, P =0.001). However, none of the SNPs retained significance after Bonferroni adjustment for multiple testing at the level of P0.0012 (0.05/42) except for TNFA-rs1799724 in ER positive cancers. In a stage II validation (1,365 AA and 1,307 EA women), we extended evaluations for four SNPs (CCL2-rs4586, CRP-rs1205, CXCL5-rs425535, and IL1RN-rs4251961), which yielded similar results. In summary, distributions of variants in genes involved in innate immune pathways were found to differ between AA and EA populations, and showed differential associations with breast cancer according to menopausal or ER status. These results suggest that immune adaptations suited to ancestral environments may differentially influence breast cancer risk among EA and AA women.

Genetic variants in one-carbon metabolism genes and breast cancer risk in European American and African American women.

Folate‐mediated one‐carbon metabolism plays critical roles in DNA synthesis, repair and DNA methylation. The impact of single nucleotide polymorphisms (SNPs) in folate‐metabolizing enzymes has been investigated in risk of breast cancer among European or Asian populations, but not among women of African ancestry. We conducted a comprehensive analysis of SNPs in eleven genes involved in one‐carbon metabolism and risk of breast cancer in 1,275 European‐American (EA) and 1,299 African‐American (AA) women who participated in the Womens Circle of Health Study. Allele frequencies varied significantly between EA and AA populations. A number of these SNPs, specifically in genes including MTR, MTRR, SHMT1, TYMS and SLC19A1, were associated with overall breast cancer risk, as well as risk by estrogen receptor (ER) status, in either EA or AA women. Associations appeared to be modified by dietary folate intake. Although single‐SNP associations were not statistically significant after correcting for multiple comparisons, polygenetic score analyses revealed significant associations with breast cancer risk. Per unit increase of the risk score was associated with a modest 19 to 50% increase in risk of breast cancer overall, ER positive or ER negative cancer (all p < 0.0005) in EAs or AAs. In summary, our data suggest that one‐carbon metabolizing gene polymorphisms could play a role in breast cancer and that may differ between EA and AA women.

Variants of estrogen-related genes and breast cancer risk in European and African American women

It has been observed previously that compared with women of European ancestry (EA), those of African ancestry (AA) are more likely to develop estrogen receptor (ER)-negative breast cancer, although the mechanisms have not been elucidated. We tested the associations between breast cancer risk and a targeted set of 20 genes known to be involved in estrogen synthesis, metabolism, and response and potential gene-environment interactions using data and samples from 1307 EA (658 cases) and 1365 AA (621 cases) participants from the Womens Circle of Health Study (WCHS). Multivariable logistic regression found evidence of associations with single-nucleotide polymorphisms (SNPs) in the ESR1 gene in EA women (rs1801132, odds ratio (OR)=1.47, 95% CI=1.20-1.80, P=0.0002; rs2046210, OR=1.24, 95% CI=1.04-1.47, P=0.02; and rs3020314, OR=1.43, 95% CI=1.19-1.70, P=0.00009), but not in AA women. The only other gene associated with breast cancer risk was CYP1A2 in AA women (rs2470893, OR=1.42, 95% CI=1.00-2.02, P=0.05), but not in EA women. When stratified by ER status, ESR1 rs1801132, rs2046210, and rs3020314 showed stronger associations in ER-positive than in ER-negative breast cancer in only EA women. Associations with the ESR1 SNPs in EA women also appeared to be stronger with longer endogenous estrogen exposure or hormonal replacement therapy use. Our results indicate that there may be differential genetic influences on breast cancer risk in EA compared with AA women and that these differences may be modified by tumor subtype and estrogen exposures. Future studies with a larger sample size may determine the full contribution of estrogen-related genes to racial/ethnic differences in breast cancer.

Genetic variations, reproductive aging, and breast cancer risk in African American and European American women: The Women's Circle of Health Study

Reproductive aging phenotypes, including age at menarche (AM) and age at natural menopause (ANM), are well-established risk factors for breast cancer. In recent years, many genetic variants have been identified in association with AM and ANM in genome-wide association studies among European populations. Using data from the Women’s Circle of Health Study (WCHS) of 1,307 European-American (EA) and 1,365 African-American (AA) breast cancer cases and controls, we aimed to replicate 53 earlier GWAS variants for AM and ANM in AA and EA groups and to perform analyses on total and net reproductive lifespan (TRLS; NRLS). Breast cancer risk was also examined in relation to a polygenic risk score (PRS) for each of the reproductive aging phenotypes. We replicated a number of variants in EA women, including rs7759938 in LIN28B for AM and rs16991615 in MCM8 for ANM; whereas in the AA group, only one SNP (rs2947411 in TMEM18) for AM was directionally consistent and nominally significant. In analysis of TRLS and NRLS, several SNPs were significant, including rs466639 in RXRG that was associated with both phenotypes in both AA and EA groups. None of the PRS was associated with breast cancer risk. Given the paucity of data available among AA populations, our study contributes to the literature of genetics of reproductive aging in AA women and highlights the importance of cross population replication of GWAS variants.

Single nucleotide variants in metastasis‐related genes are associated with breast cancer risk, by lymph node involvement and estrogen receptor status, in women with European and African ancestry

Single nucleotide polymorphisms (SNPs) in pathways influencing lymph node (LN) metastasis and estrogen receptor (ER) status in breast cancer may partially explain inter‐patient variability in prognosis. We examined 154 SNPs in 12 metastasis‐related genes for associations with breast cancer risk, stratified by LN and ER status, in European‐American (EA) and African‐American (AA) women. Two‐thousand six hundred and seventy‐one women enrolled in the Womens Circle of Health Study were genotyped. Pathway analyses were conducted using the adaptive rank truncated product (ARTP) method, with pARTP ≤ 0.10 as significant. Multi‐allelic risk scores were created for the ARTP‐significant gene(s). Single‐SNP and risk score associations were modeled using logistic regression, with false discovery rate (FDR) P‐value adjustment. Although single‐SNP associations were not significant at pFDR < 0.05, several genes were significant in the ARTP analyses. In AA women, significant ARTP gene‐level associations included CDH1 with LN+ (pARTP = 0.10; multi‐allelic OR = 1.13, 95%CI 1.07–1.19, pFDR = 0.0003) and SIPA1 with ER− breast cancer (pARTP = 0.10; multi‐allelic OR = 1.16, 95%CI 1.02–1.31, pFDR = 0.03). In EA women, MTA2 was associated with overall breast cancer risk (pARTP = 0.004), regardless of ER status, and with LN− disease (pARTP = 0.01). Also significant were SATB1 in ER− (pARTP = 0.03; multi‐allelic OR = 1.12, 95%CI 1.05–1.20, pFDR = 0.003) and KISS1 in LN− (pARTP = 0.10; multi‐allelic OR = 1.18, 95%CI 1.08–1.29, pFDR = 0.002) analyses. Among LN+ cases, significant ARTP associations were observed for SNAI1, CD82, NME1, and CTNNB1 (multi‐allelic OR = 1.09, 95%CI 1.04–1.14, pFDR = 0.001). Our findings suggest that variants in several metastasis genes may affect breast cancer risk by LN or ER status, although verification in larger studies is required.

Abstract 3593: Tumor size and lymph node metastases in African-American and European-American women with breast cancer

Few studies have evaluated lymph node metastasis in African-American (AA) women with breast cancer, who are more likely to be diagnosed at an advanced stage and with lymph node positive tumors. Likelihood of nodal involvement increases with tumor size, although recent data have indicated that this may not be true for AA breast cancer patients and patients with basal-like tumors. Nodal metastases are also more likely in premenopausal AA patients compared to either premenopausal European-American (EA) patients or postmenopausal AA and EA patients. We examined risk factors for lymph node metastasis at breast cancer diagnosis in AA and EA women, and investigated the contributions of race, tumor subtype, and menopausal status to the tumor size-lymph node metastasis relationship. This analysis included 805 women diagnosed with primary, incident breast cancer enrolled in the Women9s Circle of Health Study, a case-control study of AA and EA breast cancer patients and healthy women. Cases were identified using hospital-based ascertainment in New York City hospitals with high referral patterns for AA women and through population-based ascertainment in New Jersey using the State Cancer Registry. Eligible cases were self-identified AA and EA women age 20-75 with no previous history of cancer other than nonmelanoma skin cancer. In-person interviews were conducted and consent to review pathology reports was obtained. Tumor size was categorized as tumors 2 cm or less (small tumors) and tumors greater than 2 cm (large tumors). Unconditional logistic regression was used to estimate odds ratios (OR) and 95% confidence intervals (CI). AAs with small tumors were more likely to be node positive compared to EAs (OR=1.24, 95% CI 0.81-1.88) while among patients with large tumors, AAs were less likely to be node positive (OR=0.83, 95% CI 0.49-1.41). When grouped by race and tumor subtype, we found that the triple negative subtype was associated with a decreased risk of nodal metastases among EA women with small tumors (OR=0.17, 95% CI 0.04-0.79) and a nonsignificantly decreased risk among AA women with large tumors, using the luminal A subtype as the referent group. Associations were null in EA women with large tumors and AA women with small tumors. When grouped by race and ER status, ER negativity was associated with a decreased risk of nodal metastases among AA women with large tumors (OR=0.41, 95% CI 0.20-0.84), while AA women with small tumors were at increased risk (OR=1.90, 95% CI 0.92-3.91). Our data suggest an effect of race and tumor subtype on the relationship between tumor size and likelihood of lymph node metastases. Tumor size appears to affect lymph node metastasis differently by race, a mechanism that is modified by tumor biology. Our findings support the hypothesis that in AA breast cancer patients, large tumors may not be more likely to give rise to metastatic lymph nodes. 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 3593. doi:1538-7445.AM2012-3593