Sellers, Thomas A.

Abstract Introduction: Mammographic breast density is one of the strongest risk factors for breast cancer and appears to be modifiable. However, whether longitudinal changes in density are associated with breast cancer risk is unknown. We hypothesize that decreases in breast density are associated with reduced breast cancer risk. Methods: The Mayo Mammography Health Study cohort enrolled 19,924 women having a screening mammogram (mgm) at the Mayo Clinic, Rochester, between 2003 and 2006. Participants were 35 years and older, had no prior breast cancer and lived in the tri-state area (MN, IA and WI). Risk factor data were collected through self-administered questionnaire and clinic databases. Incident breast cancers were identified through tumor registries (Mayo Clinic, MN, IA, WI). The BI-RADS, 4-category clinical density measure (fatty, scattered density, heterogeneous density and extreme density) was available on all mgms performed after 1996. A case-cohort design consisting of incident cases (n=241) and a random selection of the cohort (n=2300, 28 developed cancer) was used to examine the association between change in BI-RADS density and breast cancer. BI-RADS densities measured from the earliest available, and enrollment mgms (average 6 years apart) were used to assess density change. Proportional hazards regression was performed to estimate the hazard ratio (HR) associated with a change in one or more BI-RADS density categories over time, relative to staying within the same category. Age was used as the time scale, and the subcohort was weighted by the inverse of the sampling proportion. Analyses were adjusted for age, years between mgms, BI-RADS density at earliest mgm, and changes in BMI, postmenopausal hormone use, and menopausal status. Results: Longitudinal analyses were conducted on the 219 cases and 1900 non-cases in the subcohort with at least one mgm available prior to enrollment mgm. Compared to the subcohort, cases were older (56.8 vs. 52.9 years), more likely postmenopausal (73.4% vs. 61.6%), had higher BMI (29.0 vs 28.1) and were more frequently screened (8.0 vs. 6.9 pre-enrollment mgms). As expected, cases were more likely to have extreme density (BI-RADS=4) at earliest mgm than non-cases (16.2% vs 14.3%). Cases were less likely than non-cases to experience a reduction of one density category or more (37.0% vs 38.6%) following earliest mgm. Adjusting for potential confounders, women who decreased one BI-RADS category or more over an average 6 years were at reduced risk of breast cancer (HR=0.72, 95%CI: 0.50-0.99) compared to those whose density was unchanged. However, women who increased one or more BI-RADS categories had suggestion of increased risk (HR=1.52, 95%CI: 0.97-2.4). Conclusion: Women with a decrease in BI-RADS density category over 6 years may have decreased breast cancer risk relative to women whose breast density category remains stable. Two measures of breast density may inform women's risk beyond a measure at one point in time. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4828.

Abstract Recent GWAS have identified and confirmed associations between several SNPs in the gene desert located on chromosome 8q24 with risk of cancers of the breast, prostate, glioma, bladder and colorectum. Evidence supporting an association with ovarian cancer is limited and inconsistent. Here we evaluate preliminary results in the 8q24 region from stage I of an ongoing invasive epithelial ovarian cancer GWAS among 1864 cases (1096 serous cases) and 1912 controls. All subjects were self-reported non-Hispanic non-Jewish Caucasian, and were genotyped with the Illumina 610quad Chip. Samples and SNPs with call rates less than 95% were excluded (9.6% and 9.9%, respectively) as well as 128 subjects with ambiguous gender, unresolved identical genotypes and less than 80% European ancestry were also excluded. Our most significant hit in the region was rs6651252 located at 8q24.21 with p = 0.000125 for all ovarian cancer, and 1.02*10−5 for serous cases only. This region is located approximately 849 kb downstream of the previous 8q24.21 findings on other cancers, and was not in linkage disequilibrium with any of the haplotype blocks reported previously. We further examined the block structure of this region with eight genotyped SNPs located within 129.61 to 129.64 MB (rs10088218, rs1516973, rs1516975, rs16903097, rs10098821, rs6651252, rs1561928, and rs1561927) and performed haplotype analysis to evaluate the association with ovarian cancer risk. One haplotype (AAGCAGGG, estimated frequency=0.11) was significantly associated with decreased ovarian cancer risk with an odds ratio of 0.67 (all cases, 95% CI: 0.78-0.91, p=0.0026, empirical p= 0.0020). This region contains no known genes, but it is located upstream and overlaps with a known copy number variation (CNV) reported by Redon et al. (2006). Previous published literature on array CGH and CNV results from TCGA (The Cancer Genome Atlas) also indicate that the 8q24.21 region is associated with frequent gains/amplification in ovarian tumors. Combined analysis of SNPs and CNV identified in this region is underway to elucidate the joint effect of these two types of genetic variation on ovarian cancer risk. Our result indicates a novel locus on 8q24.21 is associated with epithelial ovarian cancer susceptibility. Haplotype analysis reveals one specific haplotype that is associated with decreased ovarian cancer risk compared to the most common haplotype. Given previous 8q24 findings from other cancers and evidence from publicly available resources, fine-mapping and resequencing of this region is warranted. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4724.

Abstract Background: We previously identified a panel of genes associated with outcome of ovarian cancer. The purpose of the current study was to assess whether variants in these genes correlated with ovarian cancer risk. Methods and Findings: Women with and without invasive ovarian cancer (749 cases, 1,041 controls) were genotyped at 136 single nucleotide polymorphisms (SNPs) within 13 candidate genes. Risk was estimated for each SNP and for overall variation within each gene. At the gene-level, variation within MSL1 (male-specific lethal-1 homolog) was associated with risk of serous cancer (p=0.03); haplotypes within PRPF31 (PRP31 pre-mRNA processing factor 31 homolog) were associated with risk of invasive disease (p=0.03). MSL1 rs7211770 was associated with decreased risk of serous disease (OR 0.81, 95% CI 0.66-0.98; p=0.03). SNPs in MFSD7, BTN3A3, ZNF200, PTPRS, and CCND1A were inversely associated with risk (p<0.05), and there was increased risk at HEXIM1 rs1053578 (p=0.04, OR 1.40, 95% CI 1.02-1.91). Conclusions: Tumor studies can reveal novel genes worthy of follow-up for cancer susceptibility. Here, we found that inherited markers in the gene encoding MSL1, part of a complex that modifies the histone H4, may decrease risk of invasive serous ovarian cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4718.

Abstract Genome-wide association analyses have identified an ovarian cancer susceptibility locus near BNC2 at 9p22.2 (OR 0.82, p 5.1 × 10−19, Nat Genet 41:996-1000, 2009), based on 1,772 and 2,354 Stage I cases and controls, 4,803 and 5,237 Stage 2 cases and controls, and extensive Stage 3 analysis in independent samples. Here, we report on Stage 3 analysis of nine additional regions with Stage 2 results at p < 10−4. Stage 1 samples were genotyped using the Illumina 610Quad and imputed at up to 2 million SNPs; Stage 2 samples were genotyped on an Illumina iSelect of 22,790 SNPs. Stage 3 samples from 4,335 cases (including 2,197 serous subtype) and 5,951 controls (14 studies) were genotyped using Illumina or Sequenom at up to 32 SNPs on chromosomes 1, 2, 3, 7, 8, 11, and 17. No evidence of heterogeneity by study site was observed. Logistic regression revealed four regions with ordinal p ≤ 10−7 in all cases or serous subtype only. The most significant finding for all cases was for rs2072590 at 2q31.1 (all cases OR 1.15, p=8.1 × 10−13; serous only OR 1.19 p=9.0 × 10−13) which is located 3.8 kb 3-prime of HOXD3, a homeobox gene which also had an intronic SNP (rs2301301) with p=3.1 × 10−5 (all cases). Second, we observed a locus at 8q24.21 highlighted by rs10088218 (all cases OR 0.84, p=1.3 × 10−9; serous only OR 0.75 p=2.2 × 10−14). This locus resides on the opposite side of MYC and 1.3 Mb from the 8q24 regions harboring loci for other cancers, and is independent of the other cancer-associated variants (r2 < 0.02). Third, at 3q25.31, promising results were observed at rs2665390, a SNP intronic to TIPARP (TiPARP, TCDD-inducible poly(ADP-ribose) polymerase; all cases OR 1.19, p=8.7 × 10−7; serous only OR 1.26, p=3.2 × 10−8). Finally, at 17q21.32, rs9303542 was associated with risk particularly among serous cases (all cases OR 1.13, p=1.5 × 10−6; serous only OR 1.16 p=4.3 × 10−7); this SNP is intronic to SKAP1 (src kinase associated phosphoprotein 1). We evaluated the functional role of genes at these loci in ovarian cancer and normal ovarian epithelial cells and in an in vitro model of neoplastic initiation and progression from normal ovarian epithelial cells after CYMC and KRAS activation. Both BNC2 and TiPARP were down regulated in cancer versus normal cells (p=5 × 10−9 and p=3.4 × 10−5, respectively) suggesting that loss of function is required for cancer development. Over-expressing CMYC (at 8q24) induced neoplastic transformation of normal ovarian cells. Both BNC2 and TiPARP were down-regulated following CMYC over-expression and showed significant loss of function with further transformation after mutant KRAS activation (p=7.8 × 10−7 and p=6.3 × 10−4, respectively). These results emphasize the importance of genome-wide association studies and consortia for the identification of moderate-risk alleles in rare cancers. Importantly, our findings unravel the genetic components of this disease and suggest functionally-significant novel targets. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3860.

Abstract Introduction: Alterations in components of the stromal tissue that underlie the epithelium can initiate, promote or inhibit epithelial tumorigenesis. Decorin (DCN) and lumican (LUM) are stromal genes with reduced expression in serous ovarian cancer. We hypothesized that single nucleotide polymorphisms (SNPs) in these genes may influence susceptibility to serous ovarian cancer. Methods: To test this hypothesis, we compared genotype frequencies of 11 tagSNPs in DCN and LUM between 397 Caucasian cases with primary serous ovarian cancer and 920 Caucasian controls in two studies conducted at Mayo Clinic and Duke University (discovery set). Associations were evaluated further in a third Caucasian population of 436 cases and 1,098 controls in Australia (replication set). TagSNPs were selected from unrelated Caucasian samples within HapMap based on the following criteria: (i) minor allele frequency ≥ 0.05, (ii) pairwise linkage disequilibrium (LD) of r2 ≥ 0.8, and (iii) location within, and 5kb upstream and downstream of, each gene region. Genotyping was performed using the Illumina GoldenGate™ assay and associations between SNPs and serous cancer were evaluated using unconditional logistic regression to estimate per allele odds ratios (OR) and 95% confidence intervals (95%CI). Untyped genotypes for DCN rs13312816 in the discovery set, and for DCN rs3138165 and LUM rs17018765 in the replication set, were imputed using the MACH software. Results: In the discovery set, we observed inverse associations between DCN rs3138165 (OR=0.7; 95%CI=0.5-1.0), DCN rs13312816 (OR=0.7; 95%CI=0.5-1.0), DCN rs516115 (OR=0.8; 95%CI=0.7-1.0) and LUM rs17018765 (OR=0.7; 95%CI=0.5-1.0) and risk of serous ovarian cancers (all Ptrend=0.06). Associations were confirmed in the replication set for DCN rs3138165 (OR=0.6; 95%CI=0.4-0.9; Ptrend=0.009), DCN rs13312816 (OR=0.7; 95%CI=0.5-0.9; Ptrend=0.01) and LUM rs17018765 (OR=0.6; 95%CI=0.4-0.9; Ptrend=0.008), but not for DCN rs516115 (Ptrend=0.20). To increase statistical power, data from the discovery and replication sets were combined (833 cases and 2,013 controls) following confirmation of no statistical heterogeneity in the ORs between study sites. In the combined sample, associations were strengthened for DCN rs3138165 (OR=0.7; 95%CI=0.5-0.9; Ptrend=0.002), DCN rs13312816 (OR=0.7; 95%CI=0.5-0.9; Ptrend=0.002), DCN rs516115 (OR=0.9; 95%CI=0.8-1.0; Ptrend=0.03) and LUM rs17018765 (OR=0.6; 95%CI=0.5-0.8; Ptrend=0.001). All four SNPs were in the same LD block on chromosome 12, suggesting that serous ovarian cancer susceptibility may be driven by a single, correlated variant in the region. Conclusion: The findings of this study suggest that genetic variation in DCN and LUM may influence susceptibility to serous ovarian cancer and contribute to the mounting evidence that supports the role of the stroma in epithelial ovarian cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2854.

AbstractObesity is a controversial risk factor for colorectal cancer (CRC) in older women. We evaluated associations between multiple body size parameters and incident CRC in the prospective, population-based Iowa Women's Health Study (IWHS). IWHS participants, ages 55 to 69 years, provided data regarding height; weight; weight at ages 50, 40, 30, 18 years; hip circumference; and waist circumference at baseline (1986). Derived variables included body mass index (BMI), waist-to-hip ratio (WHR), and “overweight years” (OWY; conceptually similar to cigarette pack-years). Incident CRC cases (n = 1,464) were ascertained from the State Health Registry of Iowa, through 2005. Multivariable Cox regression models were fit to estimate body size–associated CRC risks. Among 36,941 women (619,961 person-years), baseline height, weight, BMI, hip circumference, waist circumference, and WHR were all positively associated with incident CRC (Ptrend ≤ 0.003 for each). Baseline BMI yielded the highest CRC risk estimates (obese III versus normal, RR = 1.56; 95% CI = 1.10–2.22; Ptrend < 0.001) and was more closely associated with distal than proximal tumors (Ptrend < 0.001 versus 0.06). Conversely, height was more closely associated with proximal than distal tumors (Ptrend < 0.001 versus 0.04). Other body size parameters were less predictive of incident CRC. These data strongly support a positive association between increased body size and CRC risk among older women. Further investigation of when increased body size has the greatest effect on CRC risk (i.e., early adulthood versus later adulthood) might also be informative, particularly with respect to defining subsite-specific pathways of colorectal carcinogenesis. Cancer Prev Res; 3(12); 1608–20. ©2010 AACR.

Purpose Mammographic density and lobular involution are both significant risk factors for breast cancer, but whether these reflect the same biology is unknown. We examined the involution and density association in a large benign breast disease (BBD) cohort. Patients and Methods Women in the Mayo Clinic BBD cohort who had a mammogram within 6 months of BBD diagnosis were eligible. The proportion of normal lobules that were involuted was categorized by an expert pathologist as no (0%), partial (1% to 74%), or complete involution (≥ 75%). Mammographic density was estimated as the four-category parenchymal pattern. Statistical analyses adjusted for potential confounders and evaluated modification by parity and age. We corroborated findings in a sample of women with BBD from the Mayo Mammography Health Study (MMHS) with quantitative percent density (PD) and absolute dense and nondense area estimates. Results Women in the Mayo BBD cohort (n = 2,667) with no (odds ratio, 1.7; 95% CI, 1.2 to 2.3) or partial (odds ratio, 1.3; 95% CI, 1.0 to 1.6) involution had greater odds of high density (DY pattern) than those with complete involution (P trend < .01). There was no evidence for effect modification by age or parity. Among 317 women with BBD in the MMHS study, there was an inverse association between involution and PD (mean PD, 22.4%, 21.6%, 17.2%, for no, partial, and complete, respectively; P trend = .04) and a strong positive association of involution with nondense area (P trend < .01). No association was seen between involution and dense area (P trend = .56). Conclusion We present evidence of an inverse association between involution and mammographic density.

Abstract Aberrant glycosylation is a well-described hallmark of cancer. In a previous ovarian cancer case control study that examined polymorphisms in 26 glycosylation-associated genes, we found strong statistical evidence (P = 0.00017) that women who inherited two copies of a single-nucleotide polymorphism in the UDP-N-acetylgalactosamine:polypeptide N-acetylgalactosaminyltransferase, GALNT1, had decreased ovarian cancer risk. The current study attempted to replicate this observation. The GALNT1 single-nucleotide polymorphism rs17647532 was genotyped in 6,965 cases and 8,377 controls from 14 studies forming the Ovarian Cancer Association Consortium. The fixed effects estimate per rs17647532 allele was null (odds ratio, 0.99; 95% confidence interval, 0.92-1.07). When a recessive model was fit, the results were unchanged. Test for hetero geneity of the odds ratios revealed consistency across the 14 replication sites but significant differences compared with the original study population (P = 0.03). This study underscores the need for replication of putative findings in genetic association studies. Cancer Epidemiol Biomarkers Prev; 19(2); 600–4

Abstract The p53 protein is critical for multiple cellular functions including cell growth and DNA repair. We assessed whether polymorphisms in the region encoding TP53 were associated with risk of invasive ovarian cancer. The study population includes a total of 5,206 invasive ovarian cancer cases (2,829 of which were serous) and 8,790 controls from 13 case-control or nested case-control studies participating in the Ovarian Cancer Association Consortium (OCAC). Three of the studies performed independent discovery investigations involving genotyping of up to 23 single nucleotide polymorphisms (SNP) in the TP53 region. Significant findings from this discovery phase were followed up for replication in the other OCAC studies. Mixed effects logistic regression was used to generate posterior median per allele odds ratios (OR), 95% probability intervals (PI), and Bayes factors (BF) for genotype associations. Five SNPs showed significant associations with risk in one or more of the discovery investigations and were followed up by OCAC. Mixed effects analysis confirmed associations with serous invasive cancers for two correlated (r2 = 0.62) SNPs: rs2287498 (median per allele OR, 1.30; 95% PI, 1.07–1.57) and rs12951053 (median per allele OR, 1.19; 95% PI, 1.01–1.38). Analyses of other histologic subtypes suggested similar associations with endometrioid but not with mucinous or clear cell cancers. This large study provides statistical evidence for a small increase in risk of ovarian cancer associated with common variants in the TP53 region. [Cancer Res 2009;69(6):2349–57]

Abstract Polymorphisms in genes critical to cell cycle control are outstanding candidates for association with ovarian cancer risk; numerous genes have been interrogated by multiple research groups using differing tagging single-nucleotide polymorphism (SNP) sets. To maximize information gleaned from existing genotype data, we conducted a combined analysis of five independent studies of invasive epithelial ovarian cancer. Up to 2,120 cases and 3,382 controls were genotyped in the course of two collaborations at a variety of SNPs in 11 cell cycle genes (CDKN2C, CDKN1A, CCND3, CCND1, CCND2, CDKN1B, CDK2, CDK4, RB1, CDKN2D, and CCNE1) and one gene region (CDKN2A-CDKN2B). Because of the semi-overlapping nature of the 123 assayed tagging SNPs, we performed multiple imputation based on fastPHASE using data from White non-Hispanic study participants and participants in the international HapMap Consortium and National Institute of Environmental Health Sciences SNPs Program. Logistic regression assuming a log-additive model was done on combined and imputed data. We observed strengthened signals in imputation-based analyses at several SNPs, particularly CDKN2A-CDKN2B rs3731239; CCND1 rs602652, rs3212879, rs649392, and rs3212891; CDK2 rs2069391, rs2069414, and rs17528736; and CCNE1 rs3218036. These results exemplify the utility of imputation in candidate gene studies and lend evidence to a role of cell cycle genes in ovarian cancer etiology, suggest a reduced set of SNPs to target in additional cases and controls. (Cancer Epidemiol Biomarkers Prev 2009;18(3):935–44)