Schwartz, Ann G.

Abstract Epithelial ovarian cancer (EOC) is a rare but deadly disease for which there is notably poorer survival in women of African Ancestry (AA) compared to women of European Ancestry (EA). Previous EA-based genome-wide association studies (GWAS) have identified 30 common, low penetrant EOC susceptibility alleles. Using the custom-designed 533,631 SNP Illumina OncoArray and imputation to ~12 million genetic variants in the 1000 Genomes Phase 3, we conducted a GWAS in 755 AA EOC cases, including 537 high-grade serous ovarian cancer (HGSOC) cases, and 1,235 AA controls from the Ovarian Cancer Association Consortium (OCAC). We identified novel susceptibility loci with suggestive evidence of association with EOC, based on a threshold of P<1x10-6, at three distinct loci including 10p15.1 (lead SNP rs4525119, intronic to AKR1C3, P=4.9 x 10-7, minor allele frequency [MAF]=0.33), 3p25.3 (lead SNP rs7643459, intronic to LOC101927394, P=8.4 x 10-7, MAF=0.36), and 4q13.3 (lead SNP rs4286604, 15 kb 3’ of UGT2A1, P=8.5x10-7, MAF=0.27). In analysis of HGSOC, we identified suggestive evidence of association at six distinct loci including 5q11.2 (lead SNP rs37792, 132 kb 5’ of FST, P=6.0x10-8, MAF=0.34), Xq27.2 (lead SNP rs57403204, 81 kb 3' of MAGEC1, P=1.7x10-7, MAF=0.06), 10p15.1 (lead SNP rs79079890, LOC105376360 intronic, P=3.0x10-7, MAF=0.03), 17p25.1 (lead SNP rs66459581, 5 kb 5’ of PRPSAP1, P=5.1x10-7, MAF=0.23), 15p12 (lead SNP rs116046250, GABRG3 intronic, P=8.7x10-7, MAF=0.05), and 4q21.21 (lead SNP rs192876988, 30 kb 3’ of GK2, P=9.2x10-7, MAF=0.05). These SNPs showed no evidence of association with risk in the OCAC EA GWAS including up to 23,543 EOC cases and 29,444 controls. Of the SNPs identified with suggestive evidence of association in our GWAS, two are located near genes known to regulate hormones and previously reported in relation to diseases of the ovary (AKR1C3 and FST), two have been linked to cancer (AKR1C3 and MAGEC1) and one represents testis-specific expression (GK2). Of the 30 SNPs identified previously in a EA EOC GWAS, we observed nominally significant associations (P<0.05) with consistent direction of effect for eight SNPs in AA women. Our investigation presents evidence of (1) variants for EOC shared among EA and AA women and (2) novel EOC risk loci of importance for AA women. Citation Format: Ani Manichaikul, Lauren C. Peres, Xin-Qun Wang, Kate Lawrenson, Sarah Abbott, Ann G. Schwartz, Anna H. Wu, Edward Peters, Patricia G. Moorman, Michele L. Cote, Melissa Bondy, Linda Kelemen, Ellen L. Goode, Jill Barnholtz-Sloan, Simon A. Gather, Andrew Berchuck, Jennifer A. Doherty, Paul Pharoah, Joellen Schildkraut. Identification of novel epithelial ovarian cancer loci in women of African ancestry from the Ovarian Cancer Association Consortium [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1307. doi:10.1158/1538-7445.AM2017-1307

Lung cancer is the deadliest cancer in the United States, killing roughly one of four cancer patients in 2016. While it is well-established that lung cancer is caused primarily by environmental effects (particularly tobacco smoking), there is evidence for genetic susceptibility. Lung cancer has been shown to aggregate in families, and segregation analyses have hypothesized a major susceptibility locus for the disease. Genetic association studies have provided strong evidence for common risk variants of small-to-moderate effect. Rare and highly penetrant alleles have been identified by linkage studies, including on 6q23–25. Though not common, some germline mutations have also been identified via sequencing studies. Ongoing genomics studies aim to identify additional high penetrance germline susceptibility alleles for this deadly disease.

Abstract Several papers have implicated mutations in the AT-rich interactive domain containing protein 1B (ARID1B) in carcinoma formation in multiple tissue types, including the lung. The ARID1B protein functions as part of SWI/SNF, a multi-subunit chromatin remodeling complex that repositions nucleosomes and may act as a tumor suppressor in cancer. ARID1B is located in a region in 6q that was previously shown to have strong evidence of linkage with lung cancer risk. As it is one of the more promising candidates within the target region, this study presents the viability of ARID1B as a candidate gene for familial lung cancer. Here, we chose individuals from families in our previous linkage study that showed the strongest linkage signal on 6q. 75 individuals from 9 families were sequenced for 37Mb of chromosome 6 from 130Mb to 167Mb using Illumina technology and a custom Agilent kit. Genotypes were imported into GoldenHelix SVS 7 for quality control and dropped if read depth < 10, quality score < 10 or a quality score to read depth ratio < 0.5. Variants with more than 1 Mendelian error were dropped. Variants with a single Mendelian inconsistency were dropped only for that family. Two-point parametric linkage analysis at theta = 0 was performed using R and an implementation of the Elston-Stewart algorithm in the paramlink package using a penetrance model of 0.01/0.1/0.1 (dd/Dd/DD) and a disease allele (D) frequency of 0.01. Family 102 showed strong evidence for linkage near ARID1B. The highest LOD score (1.34) out of all sequenced variants was located near this gene in a known conserved intergenic region between ARID1B and NOX3. Four other LOD scores > 0.7 were found in the intergenic region between these two genes. Two are transcription factor binding sites; two are conserved genomic elements. We also observed two linked intronic variants of ARID1B in this family; an enhancer (LOD = 0.89) and a transcribed region (LOD = 0.73). Two other families (47 and 59) displayed evidence of linkage in ARID1B. Here, the interest is not the value of the LOD scores (top scores were 0.64 and 0.54), but the centering of the linked haplotype around the gene. Family 47 has 60 variants with LODs between 0.64 and 0.61; 45 located within ARID1B or known regulatory regions. Family 59 has 61 variants with LODs between 0.54 and 0.51; 29 located within ARID1B or known regulatory regions The variants appear to be spread evenly throughout ARID1B, suggesting the presence of linked haplotypes. Analyses are currently underway to reconstruct possible haplotypes in these families. The results of this study show that variants in the ARID1B gene at least partially account for the linkage signal on 6q in three of the nine most strongly linked families in our cohort and thus we believe that ARID1B is a plausible candidate gene for further studies involving familial lung cancer. Replication studies using 27 new extended families are currently underway to confirm this initial result. Citation Format: Anthony M. Musolf, Claire L. Simpson, Mariza de Andrade, Diptasri Mandal, Colette Gaba, Ping Yang, Ming You, Elena Y. Kupert, Marshall W. Anderson, Ann G. Schwartz, Susan M. Pinney, Christopher I. Amos, Joan E. Bailey-Wilson. Chromatin remodeling gene ARID1B is linked to familial lung cancer. [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 2542.

Abstract Individuals from pancreatic cancer families are at increased risk, not only of pancreatic cancer, but also of melanoma, breast, ovarian, and colon cancers. While some of the increased risk may be due to mutations in high-penetrance genes (i.e., BRCA2, PALB2, ATM, p16/CDKN2A or DNA mismatch repair genes), common genetic variants may also be involved. In a high-risk population of cases with either a family history of pancreatic cancer or early-onset pancreatic cancer (diagnosis before the age of 50 years), we examined the role of genetic variants previously associated with risk of pancreatic, breast, ovarian, or prostate cancer. We genotyped 985 cases (79 early-onset cases, 906 cases with a family history of pancreatic cancer) and 877 controls for 215,389 SNPs using the iSelect Collaborative Oncological Gene-Environment Study (iCOGS) array with custom content. Logistic regression was performed using a log-linear additive model. We replicated several previously reported pancreatic cancer susceptibility loci, including recently identified variants on 2p13.3 and 7p13 (2p13.3, rs1486134: OR = 1.36; 95% CI, 1.13–1.63; P = 9.29 × 10−4; 7p13, rs17688601: OR = 0.76; 95% CI, 0.63–0.93; P = 6.59 × 10−3). For the replicated loci, the magnitude of association observed in these high-risk patients was similar to that observed in studies of unselected patients. In addition to the established pancreatic cancer loci, we also found suggestive evidence of association (P < 5 × 10−5) to pancreatic cancer for SNPs at HDAC9 (7p21.1) and COL6A2 (21q22.3). Even in high-risk populations, common variants influence pancreatic cancer susceptibility. Cancer Epidemiol Biomarkers Prev; 25(7); 1185–91. ©2016 AACR.

Abstract Pancreatic cancer is projected to become the second leading cause of cancer-related death in the United States by 2020. A familial aggregation of pancreatic cancer has been established, but the cause of this aggregation in most families is unknown. To determine the genetic basis of susceptibility in these families, we sequenced the germline genomes of 638 patients with familial pancreatic cancer and the tumor exomes of 39 familial pancreatic adenocarcinomas. Our analyses support the role of previously identified familial pancreatic cancer susceptibility genes such as BRCA2, CDKN2A, and ATM, and identify novel candidate genes harboring rare, deleterious germline variants for further characterization. We also show how somatic point mutations that occur during hematopoiesis can affect the interpretation of genome-wide studies of hereditary traits. Our observations have important implications for the etiology of pancreatic cancer and for the identification of susceptibility genes in other common cancer types. Significance: The genetic basis of disease susceptibility in the majority of patients with familial pancreatic cancer is unknown. We whole genome sequenced 638 patients with familial pancreatic cancer and demonstrate that the genetic underpinning of inherited pancreatic cancer is highly heterogeneous. This has significant implications for the management of patients with familial pancreatic cancer. Cancer Discov; 6(2); 166–75. ©2015 AACR. This article is highlighted in the In This Issue feature, p. 109

Abstract The aim of this study was to estimate the effect of exposure to secondhand tobacco smoke on the incidence of lung adenocarcinoma in situ/minimally invasive adenocarcinoma (AIS/MIA). Data from seven case–control studies participating in the International Lung Cancer Consortium (ILCCO) were pooled, resulting in 625 cases of AIS/MIA and 7,403 controls, of whom 170 cases and 3,035 controls were never smokers. Unconditional logistic regression was used to estimate adjusted ORs (ORadj) and 95% confidence intervals (CI), controlling for age, sex, race, smoking status (ever/never), and pack-years of smoking. Study center was included in the models as a random-effects intercept term. Ever versus never exposure to secondhand tobacco smoke was positively associated with AIS/MIA incidence in all subjects (ORadj = 1.48; 95% CI, 1.14–1.93) and in never smokers (ORadj = 1.45; 95% CI, 1.00–2.12). There was, however, appreciable heterogeneity of ORadj across studies (P = 0.01), and the pooled estimates were largely influenced by one large study (40% of all cases and 30% of all controls). These findings provide weak evidence for an effect of secondhand tobacco smoke exposure on AIS/MIA incidence. Further studies are needed to assess the impact of secondhand tobacco smoke exposure using the newly recommended classification of subtypes of lung adenocarcinoma. Cancer Epidemiol Biomarkers Prev; 24(12); 1902–6. ©2015 AACR.

Abstract Several studies have identified common genetic factors that influence susceptibility to lung cancer but no large studies have systematically studied familial lung cancers. The Genetic Epidemiology of Lung Cancer Consortium completed a genome-wide association study (GWAS) using samples from 757 cases and 796 controls, all of European American ancestry, and frequency matched on age and sex. Genotyping was performed by the Center for Inherited Disease Research (CIDR) using an Illumina Human OmniExpressExome-8v1 array. The most significantly associated SNPs were identified in the cases as a whole, and were also stratified by histology (adenocarcinoma and squamous cell carcinoma) and family history (0-1 and 2-5 affected family members). Analysis of all cases and controls identified a large number of SNPs around the CFTR gene that are strongly associated with lung cancer risk. Upon imputation with European Ancestry 1000 genomes data, the most significant SNP in this region is rs43034 (p = 3.89E-07, OR = 0.657) in the promoter region. In addition, an exonic missense SNP, rs213950 (p = 6.32E-07, OR = 0.663), and an intronic SNP with a possible splicing effect, rs213935 (p = 5.20E-07, OR = 0.660) were also identified as possible protective alleles. Analysis by histological subtype identified several rare and exonic SNPs that were specific by histology. We highlight two SNPs, rs114719990 (p = 1.194E-04, OR = 336.8, SETD2, c.3229A>G, p.Thr1077Ala) and rs199710487 (p = 1.124E-04, OR = 366, NTN5, c.656G>A, p.Arg219Gln), that were identified as strongly associated SNPs in cases with histologically confirmed adenocarcinoma, and two SNPs, rs144465058 (p = 1.029E-04, OR = 484.6, ANK3, c.6555G>T, p.Gln2185His) and rs140216112 (p = 1.087E-04, OR = 476, GRIK4, c.1013C>G, p.Ala338Gly), that were identified as top SNPs in cases with histologically confirmed squamous cell carcinoma. The SETD2 gene is thought to be a tumor suppressor gene and variants in this gene have previously been associated with leukemia and glioma (Zhu X et al. 2014, Fontebasso AM et al. 2013), and may now have implications in lung cancer, specifically adenocarcinoma development. In conclusion, we identified SNPs in the CFTR gene that may have a protective effect on lung cancer and several rare and exonic variants that were specific to histology subtype. SNPChrPositionGeneGenomic/ Nucleotide/ Amino acid changeOdds Ratiop-valueCFTR Protective Allelesrs430347117108449CFTRg.117108449T>C; 2.4kb upstream0.6573.89×10−7rs2139507117199533CFTRc.1408G>A; p.Val470Met0.6636.32×10−7rs2139357117178208CFTRc.869+1481A>G0.6605.20×10−7 Citation Format: Diana M. Toledo, Susan M. Pinney, Diptasri Mandal, Mariza de Andrade, Elena Kupert, Jennifer Franks, Colette Gaba, Claire L. Simpson, Ming You, Marshall W. Anderson, Joan E. Bailey-Wilson, Christopher I. Amos, Ann Schwartz. Genetic Epidemiology of Lung Cancer Consortium: genome-wide association study of familial lung cancer cases. [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 LB-189. doi:10.1158/1538-7445.AM2015-LB-189

Abstract Recently, Wilson et al. (doi:10.1038/onc.2013.396) found that EYA4 was frequently inactivated biallelically in sporadic lung cancer, displays tumor suppressor gene-like properties and affects DNA repair, and that 5 of 17 common SNPs in this 0.3Mb region showed nominal association to familial lung cancer (FLC, p<0.05) although these associations are not genome-wide significant after correction for multiple testing.. However, this is an interesting candidate gene for FLC risk because it is located on chromosome 6q in a region where we previously showed evidence of linkage to FLC (PMID: 15272417, 20215501). Here, we sequenced 75 individuals from our 9 most strongly 6q-linked families, using Illumina technology and a custom Agilent kit to capture 37Mb of chromosome 6 from 130Mb to 167Mb. We closely examined a region around EYA4, between 133Mb and 135Mb. This region contains a number of other genes in addition to EYA4, including the Vanin cluster; ribosomal protein S12; the tumor suppressor gene TCF21 and its antisense RNA TARID which directs methylation and activation of TCF21; solute carriers SLC18B1 and SLC2A12 and the serum/glucocorticoid regulated kinase SGK1. To detect sharing of variants among affecteds in each family while allowing for phenocopies, singlepoint affected-only linkage analysis was performed on lung cancer affection status for all sequence variants using the Elston-Stewart algorithm implemented in the R package paramlink assuming penetrance of 0.01, 0.1 and 0.1 for dd/Dd/DD and a disease allele (D) frequency of 0.01, at a recombination fraction (theta) of 0. The highest total LOD score (summed over all 9 families) in the entire 37Mb targeted region was not near EYA4. In only one family was there any variant in EYA4 that appeared on the linked haplotype with close to the highest family-specific LOD score: Family 30 had a LOD of 0.6514 at several EYA4 variants (max LOD across region 0.6519). However, many variants across the region had similar LODs. In the remaining families, all EYA4 variants had LODs much lower than the maximum family-specific LOD score in the region. Thus, EYA4 variants are unlikely to explain our 6q linkage peak in these 6q-linked families. The second highest total LOD score for the entire 37Mb targeted region was found near EYA4 at 133.9Mb (LOD = 3.82), in the middle of the TARID antisense RNA gene. This variant appears to be entirely novel, as it is not present in any of the online variant databases, such as NHLBI's ESP or 1000 Genomes. In two families, the family LOD score at this locus was at or close to the highest in the region: in Family 44 the LOD score was 1.206 (max LOD across entire region 1.207) and in Family 35 the LOD score was 0.3958 (max LOD across region 0.3959). Therefore this variant in the TARID antisense RNA is a candidate in some of our families most strongly linked to chromosome 6q. Citation Format: Joan E. Bailey-Wilson, Claire L. Simpson, Susan M. Pinney, Mariza de Andrade, Colette Gaba, Ping Yang, Ming You, Elena Y. Kupert, Ann G. Schwartz, Diptasri Mandal, Christopher I. Amos, Marshall W. Anderson. Evaluation of EYA4 as a candidate risk locus in familial lung cancer families linked to 6q. [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 2757. doi:10.1158/1538-7445.AM2015-2757

Abstract Children who are exposed to secondhand smoke (SHS) are faced with an increased risk of nicotine dependence later in life. We have previously shown that children exposed to SHS have an increased risk of lung cancer as adults. Furthermore, genetic variation plays a role in the susceptibility of individuals to lung cancer. The gene DRD1 mediates dopamine signaling following nicotine exposure, thus we reasoned that DRD1 may be related to lung cancer via modulation of the nicotine reward pathway. Our previous studies identified 4 SNPs (rs4809294, rs2292975, rs4861327, rs686) that were predicted to modulate a microRNA binding site and we assessed the association of these SNPs with lung cancer risk. Our current study investigated whether these SNPs were associated with nicotine dependence and whether this had a subsequent effect on lung cancer risk. We used the National Cancer Institute-Maryland lung cancer case-control study. Cases and controls were recruited from the greater Baltimore metropolitan area. We used ordinal logistic regression to examine the association between SNPs and nicotine dependence. Logistic regression adjusted for age, gender and smoking was used to assess the association between SNPs and lung cancer risk. No significant associations were found between genotypes in the 4 SNPs and nicotine dependence levels (n=1,450). Individuals with the heterozygous allele (AG) exposed to SHS during childhood had increased odds of lung cancer compared to those with the homozygous wild type allele (AA) for rs686 (OR=1.52, CI= 1.12, 2.07). Individuals with the homozygous variant allele (GG) exposed to childhood SHS had increased odds of lung cancer compared to individuals with the homozygous wild type allele (AA) (OR=2.11, CI= 1.45, 3.10). Individuals with the heterozygous allele (GA) and the highest levels of nicotine dependence who were also exposed to childhood SHS, had higher odds of lung cancer than those with the homozygous recessive allele (AA) (OR=3.50, CI=1.37, 8.96). Consistent with earlier findings, heterozygotes (AG) had increased odds of lung cancer but this was not statistically significant (OR=1.90, CI= 0.82, 4.40). To further explore this association, we examined the effect of race on this relationship. Caucasians with the heterozygous allele (AG), the highest levels of nicotine dependence and childhood SHS, had increased odds of lung cancer compared to those with the homozygous recessive allele (AA) for rs686 (OR=1.47, CI=1.06, 2.05). Caucasians with the homozygous allele (GG) who were exposed to childhood SHS, had higher odds of lung cancer compared to individuals with the homozygous recessive allele (AA) for rs686 (OR=1.84, CI=1.15, 2.98). No association was found among African Americans. Although none of the SNPs examined were found to be associated with nicotine dependence, it appears that variations in rs686 may modify an individual's susceptibility to lung cancer due to SHS exposure. Citation Format: Victoria A. Zigmont, Brid M. Ryan, Jin Jen, Ana I. Robles, Cain McClary, Kara Calhoun, Elise D. Bowman, Kirsi Vähäkangas, K. Leigh Greathouse, Wang Yi, Angela S. Wenzlaff, Bo Deng, Ping Yang, Ann G. Schwartz, Curtis C. Harris, Susan Olivo Marston. Childhood exposure to secondhand smoke, nicotine dependence, and DRD1 are associated with lung 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 2156. doi:10.1158/1538-7445.AM2014-2156

Abstract Lung cancer has a familial component which suggests a genetic contribution to its etiology. Given the strong evidence linking smoking with lung cancer, we studied miRNA-related loci in genes associated with smoking behavior. CHRNA, CHRNB gene families, CYP2A6, and DRD1 (dopamine receptor D1) were mined for SNPs that fell within the seed region of miRNA binding sites and then tested for associations with risk in a three-stage validation approach. A 3′UTR (untranslated region) SNP in DRD1 was associated with a lower risk of lung cancer among individuals exposed to secondhand smoke during childhood [OR, 0.69; 95% confidence interval (CI), 0.60–0.79; P < 0.0001]. This relationship was evident in both ever (OR, 0.74; 95% CI, 0.62–0.88; P = 0.001) and never smokers (OR, 0.61; 95% CI, 0.47–0.79; P < 0.0001), European American (OR, 0.65; 95% CI, 0.53–0.80; P < 0.0001), and African American (OR, 0.73; 95% CI, 0.62–0.88; P = 0.001) populations. Although much remains undefined about the long-term risks associated with exposure to secondhand smoke and heterogeneity between individuals in regard to their susceptibility to the effects of secondhand smoke, our data show an interaction between an SNP in the 3′UTR of DRD1 and exposure to secondhand smoke during childhood. Further work is needed to explore the mechanistic underpinnings of this SNP and the nature of the interaction between DRD1 and exposure to secondhand smoke during childhood. Cancer Prev Res; 7(12); 1210–8. ©2014 AACR.