Identification of a novel percent mammographic density locus at 12q24.

Percent mammographic density adjusted for age and body mass index (BMI) is one of the strongest risk factors for breast cancer and has a heritable component that remains largely unidentified. We performed a three-stage genome-wide association study (GWAS) of percent mammographic density to identify novel genetic loci associated with this trait. In stage 1, we combined three GWASs of percent density comprised of 1241 women from studies at the Mayo Clinic and identified the top 48 loci (99 single nucleotide polymorphisms). We attempted replication of these loci in 7018 women from seven additional studies (stage 2). The meta-analysis of stage 1 and 2 data identified a novel locus, rs1265507 on 12q24, associated with percent density, adjusting for age and BMI (P = 4.43 × 10(-8)). We refined the 12q24 locus with 459 additional variants (stage 3) in a combined analysis of all three stages (n = 10 377) and confirmed that rs1265507 has the strongest association in the 12q24 region (P = 1.03 × 10(-8)). Rs1265507 is located between the genes TBX5 and TBX3, which are members of the phylogenetically conserved T-box gene family and encode transcription factors involved in developmental regulation. Understanding the mechanism underlying this association will provide insight into the genetics of breast tissue composition.

No evidence for association of inherited variation in genes involved in mitosis and percent mammographic density.

INTRODUCTION: Increased mammographic breast density is one of the strongest risk factors for breast cancer. While two-thirds of the variation in mammographic density appears to be genetically influenced, few variants have been identified. We examined the association of inherited variation in genes from pathways that mediate cell division with percent mammographic density (PMD) adjusted for age, body mass index (BMI) and postmenopausal hormones, in two studies of healthy postmenopausal women. METHODS: We investigated 2,058 single nucleotide polymorphisms (SNPs) in 378 genes involved in regulation of mitosis for associations with adjusted PMD among 484 unaffected postmenopausal controls (without breast cancer) from the Mayo Clinic Breast Cancer Study (MCBCS) and replicated the findings in postmenopausal controls (n = 726) from the Singapore and Sweden Breast Cancer Study (SASBAC) study. PMD was assessed in both studies by a computer-thresholding method (Cumulus) and linear regression approaches were used to assess the association of SNPs and PMD, adjusted for age, BMI and postmenopausal hormones. A P-value threshold of 4.2 × 10-5 based on a Bonferroni correction of effective number of independent tests was used for statistical significance. Further, a pathway-level analysis was conducted of all 378 genes using the self-contained gene-set analysis method GLOSSI. RESULTS: A variant in PRPF4, rs10733604, was significantly associated with adjusted PMD in the MCBCS (P = 2.7 × 10-7), otherwise, no single SNP was associated with PMD. Additionally, the pathway analysis provided no evidence of enrichment in the number of associations observed between SNPs in the mitotic genes and PMD (P = 0.60). We evaluated rs10733604 (PRPF4), and 73 other SNPs at P < 0.05 from 51 genes in the SASBAC study. There was no evidence of an association of rs10733604 (PRPF4) with adjusted PMD in SASBAC (P = 0.23). There were, however, consistent associations (P < 0.05) of variants at the putative locus, LOC375190, Aurora B kinase (AURKB), and Mini-chromosome maintenance complex component 3 (MCM3) with adjusted PMD, although these were not statistically significant. CONCLUSIONS: Our findings do not support a role of inherited variation in genes involved in regulation of cell division and adjusted percent mammographic density in postmenopausal women.

Evaluation of chromosome 6p22 as a breast cancer risk modifier locus in a follow-up study of BRCA2 mutation carriers.

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.

Allele-specific expression of APC in adenomatous polyposis families.

BACKGROUND & AIMS: Germline mutations in the APC gene cause of most cases of familial adenomatous polyposis (FAP) and a lesser proportion of attenuated FAP (AFAP). Systematic analysis of APC at the RNA level could provide insight into the pathogenicity of identified mutations and the molecular basis of FAP/AFAP in families without identifiable mutations. Here, we analyzed the prevalence of imbalances in the allelic expression of APC in polyposis families with germline mutations in the gene and without detectable mutations in APC and/or MUTYH. METHODS: Allele-specific expression (ASE) was determined by single nucleotide primer extension using an exon 11 polymorphism as an allele-specific marker. In total, 52 APC-mutation-positive (36 families) and 24 APC/MUTYH-mutation-negative (23 families) informative patients were analyzed. Seventy-six controls also were included. RESULTS: Of the APC-mutation-positive families, most of those in whom the mutation was located before the last exon of the gene (12 of 14) had ASE imbalance, which is consistent with a mechanism of nonsense-mediated decay. Of the APC/MUTYH-mutation-negative families, 2 (9%) had ASE imbalance, which might cause the disease. Normal allele expression was restored shortly after lymphocytes were cultured with puromycin, supporting a 'nonsense-mediated' hypothesis. CONCLUSIONS: ASE analysis might be used to determine the pathogenesis of some cases of FAP and AFAP in which APC mutations are not found. ASE also might be used to prioritize the order in which different areas of APC are tested. RNA-level studies are important for the molecular diagnosis of FAP.

Evaluation of associations between common variation in mitotic regulatory pathways and risk of overall and high grade breast cancer.

Mitotic regulatory pathways insure proper timing of mitotic entry, sister chromatid cohesion and separation, and cytokinesis. Disruption of this process results in inappropriate chromosome segregation and aneuploidy, and appears to contribute to cancer. Specifically, disregulation and somatic mutation of mitotic regulators has been observed in human cancers, and overexpression of mitotic regulators is common in aggressive and late stage tumors. However, the role of germline variation in mitotic pathways and risk of cancer is not well understood. We tested 1,084 haplotype-tagging and functional variants from 164 genes in mitotic regulatory pathways in 791 Caucasian women with breast cancer and 843 healthy controls for association with risk of overall and high grade breast cancer. Sixty-one single nucleotide polymorphisms (SNPs) from 40 genes were associated (P < 0.05) with risk of breast cancer in a log-additive model. In addition, 60 SNPs were associated (P < 0.05) with risk of high grade breast cancer. However, none of these associations were significant after Bonferroni correction for multiple testing. In gene-level analyses, CDC25C, SCC1/RAD21, TLK2, and SMC6L1 were associated (P < 0.05) with overall breast cancer risk, CDC6, CDC27, SUMO3, RASSF1, KIF2, and CDC14A were associated with high grade breast cancer risk, and EIF3S10 and CDC25A were associated with both. Further investigation in breast and other cancers are needed to understand the influence of inherited variation in mitotic genes on tumor grade and cancer risk.

Genetic susceptibility to triple-negative breast cancer.

Triple-negative breast cancers (TNBC), defined by the absence of estrogen receptor, progesterone receptor, and HER-2 expression, account for 12% to 24% of all breast cancers. TNBC is associated with early recurrence of disease and poor outcome. Germline mutations in the BRCA1 and BRCA2 breast cancer susceptibility genes have been associated with up to 15% of TNBC, and TNBC accounts for 70% of breast tumors arising in BRCA1 mutation carriers and 16% to 23% of breast tumors in BRCA2 carriers. Whether germline mutations in other breast cancer susceptibility genes also predispose to TNBC remains to be determined. Common variation in a subset of the 72 known breast cancer susceptibility loci identified through genome-wide association studies and other large-scale genotyping efforts have also been associated with risk of TNBC (TOX3, ESR1, RAD51L1, TERT, 19p13.1, 20q11, MDM4, 2p24.1, and FTO). Furthermore, variation in the 19p13.1 locus and the MDM4 locus has been associated with TNBC, but not other forms of breast cancer, suggesting that these are TNBC-specific loci. Thus, TNBC can be distinguished from other breast cancer subtypes by a unique pattern of common and rare germline predisposition alleles. Additional efforts to combine genetic and epidemiologic data are needed to better understand the etiology of this aggressive form of breast cancer, to identify prevention and therapeutic targets, and to impact clinical practice through the development of risk prediction models.

Contemporary outcomes of surgical ventricular septal defect closure.

OBJECTIVES: Surgical closure of ventricular septal defects remains the most common pediatric cardiac surgical procedure. No studies, however, have comprehensively analyzed risk factors and drivers of nonmortality outcomes in the current era. The purpose of this study was to assess both baseline characteristics and outcomes of children undergoing surgical repair of ventricular septal defects in a contemporary cohort. METHODS: This retrospective study examined a consecutive series of 369 ventricular septal defect closures at a single institution. Because mortality is low in nearly all centers for repair of these defects, we focused on morbidity and identified drivers of risk via multivariable linear regression modeling. RESULTS: For children younger than age 6 months undergoing ventricular septal defect closure, every extra kilogram in operative weight results in a 2.3-day shorter length of stay. In an analysis of composite risk, patients younger than age 6 months undergoing ventricular septal defect repair exhibited a 1.8-fold increase in composite risk for each kilogram decrease in weight, whereas patients older than age 6 months experienced no significant difference. CONCLUSIONS: Even in the current surgical era, weight remains a significant predictor of morbidity and driver or length of stay in young infants undergoing ventricular septal defect closure. Weight still should be considered when discussing operative risks for children younger than age 6 months undergoing this procedure, irrespective of the indication for operation.

Genetic modifiers of menopausal hormone replacement therapy and breast cancer risk: a genome-wide interaction study.

Women using menopausal hormone therapy (MHT) are at increased risk of developing breast cancer (BC). To detect genetic modifiers of the association between current use of MHT and BC risk, we conducted a meta-analysis of four genome-wide case-only studies followed by replication in 11 case-control studies. We used a case-only design to assess interactions between single-nucleotide polymorphisms (SNPs) and current MHT use on risk of overall and lobular BC. The discovery stage included 2920 cases (541 lobular) from four genome-wide association studies. The top 1391 SNPs showing P values for interaction (Pint) <3.0 × 10(-3) were selected for replication using pooled case-control data from 11 studies of the Breast Cancer Association Consortium, including 7689 cases (676 lobular) and 9266 controls. Fixed-effects meta-analysis was used to derive combined Pint. No SNP reached genome-wide significance in either the discovery or combined stage. We observed effect modification of current MHT use on overall BC risk by two SNPs on chr13 near POMP (combined Pint≤8.9 × 10(-6)), two SNPs in SLC25A21 (combined Pint≤4.8 × 10(-5)), and three SNPs in PLCG2 (combined Pint≤4.5 × 10(-5)). The association between lobular BC risk was potentially modified by one SNP in TMEFF2 (combined Pint≤2.7 × 10(-5)), one SNP in CD80 (combined Pint≤8.2 × 10(-6)), three SNPs on chr17 near TMEM132E (combined Pint≤2.2×10(-6)), and two SNPs on chr18 near SLC25A52 (combined Pint≤4.6 × 10(-5)). In conclusion, polymorphisms in genes related to solute transportation in mitochondria, transmembrane signaling, and immune cell activation are potentially modifying BC risk associated with current use of MHT. These findings warrant replication in independent studies.

Common variation in Nemo-like kinase is associated with risk of ovarian cancer.

BACKGROUND: Overexpression of mitotic kinases has been associated with prognosis, histologic grade, and clinical stage in ovarian cancer, but the relationship between inherited variation in these genes and ovarian cancer risk has not been well defined. METHODS: We measured associations between 397 single nucleotide polymorphisms (SNPs) from 67 mitotic kinases and invasive epithelial ovarian cancer risk in two case-control studies (n = 671 cases; n = 939 controls). Thirty-six candidate SNPs (P < 0.05) were assessed in a replication analysis consisting of three additional studies (n = 1,094 cases; n = 829 controls). RESULTS: In initial analysis, thirty-six SNPs were suggestive of association with risk of serous ovarian cancer, all subtypes of ovarian cancer, or both (P < 0.05). Replication analyses suggested an association between rs2125846 in the Nemo-like kinase (NLK) gene and ovarian cancer (serous OR = 1.36, 95% CI: 1.11-1.67, P = 1.77 × 10(-3); all subtypes OR = 1.30, 95% CI: 1.08-1.56, P = 2.97 × 10(-3)). Furthermore, rs2125846 was associated with risk in the combined discovery and replication sets (serous OR = 1.33, 95% CI: 1.15-1.54; all subtypes OR = 1.27, 95% CI: 1.12-1.45). CONCLUSIONS: Variation in NLK may be associated with risk of invasive epithelial ovarian cancer. Further studies are needed to confirm and understand the biologic relationship between this mitotic kinase and ovarian cancer risk. IMPACT: An association between SNPs in NLK and ovarian cancer may provide biologic insight into the development of this disease.

19p13.1 is a triple-negative-specific breast cancer susceptibility locus.

The 19p13.1 breast cancer susceptibility locus is a modifier of breast cancer risk in BRCA1 mutation carriers and is also associated with the risk of ovarian cancer. Here, we investigated 19p13.1 variation and risk of breast cancer subtypes, defined by estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor-2 (HER2) status, using 48,869 breast cancer cases and 49,787 controls from the Breast Cancer Association Consortium (BCAC). Variants from 19p13.1 were not associated with breast cancer overall or with ER-positive breast cancer but were significantly associated with ER-negative breast cancer risk [rs8170 OR, 1.10; 95% confidence interval (CI), 1.05-1.15; P = 3.49 × 10(-5)] and triple-negative (ER-, PR-, and HER2-negative) breast cancer (rs8170: OR, 1.22; 95% CI, 1.13-1.31; P = 2.22 × 10(-7)). However, rs8170 was no longer associated with ER-negative breast cancer risk when triple-negative cases were excluded (OR, 0.98; 95% CI, 0.89-1.07; P = 0.62). In addition, a combined analysis of triple-negative cases from BCAC and the Triple Negative Breast Cancer Consortium (TNBCC; N = 3,566) identified a genome-wide significant association between rs8170 and triple-negative breast cancer risk (OR, 1.25; 95% CI, 1.18-1.33; P = 3.31 × 10(-13)]. Thus, 19p13.1 is the first triple-negative-specific breast cancer risk locus and the first locus specific to a histologic subtype defined by ER, PR, and HER2 to be identified. These findings provide convincing evidence that genetic susceptibility to breast cancer varies by tumor subtype and that triple-negative tumors and other subtypes likely arise through distinct etiologic pathways.

Common breast cancer susceptibility loci are associated with triple-negative breast cancer.

Triple-negative breast cancers are an aggressive subtype of breast cancer with poor survival, but there remains little known about the etiologic factors that promote its initiation and development. Commonly inherited breast cancer risk factors identified through genome-wide association studies display heterogeneity of effect among breast cancer subtypes as defined by the status of estrogen and progesterone receptors. In the Triple Negative Breast Cancer Consortium (TNBCC), 22 common breast cancer susceptibility variants were investigated in 2,980 Caucasian women with triple-negative breast cancer and 4,978 healthy controls. We identified six single-nucleotide polymorphisms, including rs2046210 (ESR1), rs12662670 (ESR1), rs3803662 (TOX3), rs999737 (RAD51L1), rs8170 (19p13.1), and rs8100241 (19p13.1), significantly associated with the risk of triple-negative breast cancer. Together, our results provide convincing evidence of genetic susceptibility for triple-negative breast cancer.

Common variation in ISL1 confers genetic susceptibility for human congenital heart disease.

Congenital heart disease (CHD) is the most common birth abnormality and the etiology is unknown in the overwhelming majority of cases. ISLET1 (ISL1) is a transcription factor that marks cardiac progenitor cells and generates diverse multipotent cardiovascular cell lineages. The fundamental role of ISL1 in cardiac morphogenesis makes this an exceptional candidate gene to consider as a cause of complex congenital heart disease. We evaluated whether genetic variation in ISL1 fits the common variant-common disease hypothesis. A 2-stage case-control study examined 27 polymorphisms mapping to the ISL1 locus in 300 patients with complex congenital heart disease and 2,201 healthy pediatric controls. Eight genic and flanking ISL1 SNPs were significantly associated with complex congenital heart disease. A replication study analyzed these candidate SNPs in 1,044 new cases and 3,934 independent controls and confirmed that genetic variation in ISL1 is associated with risk of non-syndromic congenital heart disease. Our results demonstrate that two different ISL1 haplotypes contribute to risk of CHD in white and black/African American populations.

Network modeling links breast cancer susceptibility and centrosome dysfunction.

Many cancer-associated genes remain to be identified to clarify the underlying molecular mechanisms of cancer susceptibility and progression. Better understanding is also required of how mutations in cancer genes affect their products in the context of complex cellular networks. Here we have used a network modeling strategy to identify genes potentially associated with higher risk of breast cancer. Starting with four known genes encoding tumor suppressors of breast cancer, we combined gene expression profiling with functional genomic and proteomic (or 'omic') data from various species to generate a network containing 118 genes linked by 866 potential functional associations. This network shows higher connectivity than expected by chance, suggesting that its components function in biologically related pathways. One of the components of the network is HMMR, encoding a centrosome subunit, for which we demonstrate previously unknown functional associations with the breast cancer-associated gene BRCA1. Two case-control studies of incident breast cancer indicate that the HMMR locus is associated with higher risk of breast cancer in humans. Our network modeling strategy should be useful for the discovery of additional cancer-associated genes.