Kathryn L. Penney

Assessing the impact of population stratification on genetic association studies

Population stratification refers to differences in allele frequencies between cases and controls due to systematic differences in ancestry rather than association of genes with disease. It has been proposed that false positive associations due to stratification can be controlled by genotyping a few dozen unlinked genetic markers. To assess stratification empirically, we analyzed data from 11 case-control and case-cohort association studies. We did not detect statistically significant evidence for stratification but did observe that assessments based on a few dozen markers lack power to rule out moderate levels of stratification that could cause false positive associations in studies designed to detect modest genetic risk factors. After increasing the number of markers and samples in a case-cohort study (the design most immune to stratification), we found that stratification was in fact present. Our results suggest that modest amounts of stratification can exist even in well designed studies.

Admixture mapping identifies 8q24 as a prostate cancer risk locus in African-American men

A whole-genome admixture scan in 1,597 African Americans identified a 3.8 Mb interval on chromosome 8q24 as significantly associated with susceptibility to prostate cancer [logarithm of odds (LOD) = 7.1]. The increased risk because of inheriting African ancestry is greater in men diagnosed before 72 years of age (P < 0.00032) and may contribute to the epidemiological observation that the higher risk for prostate cancer in African Americans is greatest in younger men (and attenuates with older age). The same region was recently identified through linkage analysis of prostate cancer, followed by fine-mapping. We strongly replicated this association (P < 4.2 × 10−9) but find that the previously described alleles do not explain more than a fraction of the admixture signal. Thus, admixture mapping indicates a major, still-unidentified risk gene for prostate cancer at 8q24, motivating intense work to find it.

Transferability of tag SNPs in genetic association studies in multiple populations

A general question for linkage disequilibrium–based association studies is how power to detect an association is compromised when tag SNPs are chosen from data in one population sample and then deployed in another sample. Specifically, it is important to know how well tags picked from the HapMap DNA samples capture the variation in other samples. To address this, we collected dense data uniformly across the four HapMap population samples and eleven other population samples. We picked tag SNPs using genotype data we collected in the HapMap samples and then evaluated the effective coverage of these tags in comparison to the entire set of common variants observed in the other samples. We simulated case-control association studies in the non-HapMap samples under a disease model of modest risk, and we observed little loss in power. These results demonstrate that the HapMap DNA samples can be used to select tags for genome-wide association studies in many samples around the world.

Familial Risk and Heritability of Cancer Among Twins in Nordic Countries

IMPORTANCE Estimates of familial cancer risk from population-based studies are essential components of cancer risk prediction. OBJECTIVE To estimate familial risk and heritability of cancer types in a large twin cohort. DESIGN, SETTING, AND PARTICIPANTS Prospective study of 80,309 monozygotic and 123,382 same-sex dizygotic twin individuals (N = 203,691) within the population-based registers of Denmark, Finland, Norway, and Sweden. Twins were followed up a median of 32 years between 1943 and 2010. There were 50,990 individuals who died of any cause, and 3804 who emigrated and were lost to follow-up. EXPOSURES Shared environmental and heritable risk factors among pairs of twins. MAIN OUTCOMES AND MEASURES The main outcome was incident cancer. Time-to-event analyses were used to estimate familial risk (risk of cancer in an individual given a twins development of cancer) and heritability (proportion of variance in cancer risk due to interindividual genetic differences) with follow-up via cancer registries. Statistical models adjusted for age and follow-up time, and accounted for censoring and competing risk of death. RESULTS A total of 27,156 incident cancers were diagnosed in 23,980 individuals, translating to a cumulative incidence of 32%. Cancer was diagnosed in both twins among 1383 monozygotic (2766 individuals) and 1933 dizygotic (2866 individuals) pairs. Of these, 38% of monozygotic and 26% of dizygotic pairs were diagnosed with the same cancer type. There was an excess cancer risk in twins whose co-twin was diagnosed with cancer, with estimated cumulative risks that were an absolute 5% (95% CI, 4%-6%) higher in dizygotic (37%; 95% CI, 36%-38%) and an absolute 14% (95% CI, 12%-16%) higher in monozygotic twins (46%; 95% CI, 44%-48%) whose twin also developed cancer compared with the cumulative risk in the overall cohort (32%). For most cancer types, there were significant familial risks and the cumulative risks were higher in monozygotic than dizygotic twins. Heritability of cancer overall was 33% (95% CI, 30%-37%). Significant heritability was observed for the cancer types of skin melanoma (58%; 95% CI, 43%-73%), prostate (57%; 95% CI, 51%-63%), nonmelanoma skin (43%; 95% CI, 26%-59%), ovary (39%; 95% CI, 23%-55%), kidney (38%; 95% CI, 21%-55%), breast (31%; 95% CI, 11%-51%), and corpus uteri (27%; 95% CI, 11%-43%). CONCLUSIONS AND RELEVANCE In this long-term follow-up study among Nordic twins, there was significant excess familial risk for cancer overall and for specific types of cancer, including prostate, melanoma, breast, ovary, and uterus. This information about hereditary risks of cancers may be helpful in patient education and cancer risk counseling.

Vitamin D–Related Genetic Variation, Plasma Vitamin D, and Risk of Lethal Prostate Cancer: A Prospective Nested Case–Control Study

BACKGROUND The association of vitamin D status with prostate cancer is controversial; no association has been observed for overall incidence, but there is a potential link with lethal disease. METHODS We assessed prediagnostic 25-hydroxyvitamin D [25(OH)D] levels in plasma, variation in vitamin D-related genes, and risk of lethal prostate cancer using a prospective case-control study nested within the Health Professionals Follow-up Study. We included 1260 men who were diagnosed with prostate cancer after providing a blood sample in 1993-1995 and 1331 control subjects. Men with prostate cancer were followed through March 2011 for lethal outcomes (n = 114). We selected 97 single-nucleotide polymorphisms (SNPs) in genomic regions with high linkage disequilibrium (tagSNPs) to represent common genetic variation among seven vitamin D-related genes (CYP27A1, CYP2R1, CYP27B1, GC, CYP24A1, RXRA, and VDR). We used a logistic kernel machine test to assess whether multimarker SNP sets in seven vitamin D pathway-related genes were collectively associated with prostate cancer. Tests for statistical significance were two-sided. RESULTS Higher 25(OH)D levels were associated with a 57% reduction in the risk of lethal prostate cancer (highest vs lowest quartile: odds ratio = 0.43, 95% confidence interval = 0.24 to 0.76). This finding did not vary by time from blood collection to diagnosis. We found no statistically significant association of plasma 25(OH)D levels with overall prostate cancer. Pathway analyses found that the set of SNPs that included all seven genes (P = .008) as well as sets of SNPs that included VDR (P = .01) and CYP27A1 (P = .02) were associated with risk of lethal prostate cancer. CONCLUSION In this prospective study, plasma 25(OH)D levels and common variation among several vitamin D-related genes were associated with lethal prostate cancer risk, suggesting that vitamin D is relevant for lethal prostate cancer.

Evaluation of the 8q24 Prostate Cancer Risk Locus and MYC Expression

Polymorphisms at 8q24 are robustly associated with prostate cancer risk. The risk variants are located in nonprotein coding regions and their mechanism has not been fully elucidated. To further dissect the function of this locus, we tested two hypotheses: (a) unannotated microRNAs (miRNA) are transcribed in the region, and (b) this region is a cis-acting enhancer. Using next generation sequencing, 8q24 risk regions were interrogated for known and novel miRNAs in histologically normal radical prostatectomy tissue. We also evaluated the association between the risk variants and transcript levels of multiple genes, focusing on the proto-oncogene, MYC. RNA expression was measured in histologically normal and tumor tissue from 280 prostatectomy specimens (from 234 European American and 46 African American patients), and paired germline DNA from each individual was genotyped for six 8q24 risk single nucleotide polymorphisms. No evidence was found for significant miRNA transcription within 8q24 prostate cancer risk loci. Likewise, no convincing association between RNA expression and risk allele status was detected in either histologically normal or tumor tissue. To our knowledge, this is one of the first and largest studies to directly assess miRNA in this region and to systematically measure MYC expression levels in prostate tissue in relation to inherited risk variants. These data will help to direct the future study of this risk locus.

Fatty Acid Synthase Polymorphisms, Tumor Expression, Body Mass Index, Prostate Cancer Risk, and Survival

PURPOSE Fatty acid synthase (FASN) regulates de novo lipogenesis, body weight, and tumor growth. We examined whether common germline single nucleotide polymorphisms (SNPs) in the FASN gene affect prostate cancer (PCa) risk or PCa-specific mortality and whether these effects vary by body mass index (BMI). METHODS In a prospective nested case-control study of 1,331 white patients with PCa and 1,267 age-matched controls, we examined associations of five common SNPs within FASN (and 5 kb upstream/downstream, R(2) > 0.8) with PCa incidence and, among patients, PCa-specific death and tested for an interaction with BMI. Survival analyses were repeated for tumor FASN expression (n = 909). RESULTS Four of the five SNPs were associated with lethal PCa. SNP rs1127678 was significantly related to higher BMI and interacted with BMI for both PCa risk (P(interaction) = .004) and PCa mortality (P(interaction) = .056). Among overweight men (BMI > or = 25 kg/m(2)), but not leaner men, the homozygous variant allele carried a relative risk of advanced PCa of 2.49 (95% CI, 1.00 to 6.23) compared with lean men with the wild type. Overweight patients carrying the variant allele had a 2.04 (95% CI, 1.31 to 3.17) times higher risk of PCa mortality. Similarly, overweight patients with elevated tumor FASN expression had a 2.73 (95% CI, 1.05 to 7.08) times higher risk of lethal PCa (P(interaction) = .02). CONCLUSION FASN germline polymorphisms were significantly associated with risk of lethal PCa. Significant interactions of BMI with FASN polymorphisms and FASN tumor expression suggest FASN as a potential link between obesity and poor PCa outcome and raise the possibility that FASN inhibition could reduce PCa-specific mortality, particularly in overweight men.

Systematic Evaluation of Genetic Variation at the Androgen Receptor Locus and Risk of Prostate Cancer in a Multiethnic Cohort Study

Repeat length of the CAG microsatellite polymorphism in exon 1 of the androgen receptor (AR) gene has been associated with risk of prostate cancer in humans. This association has been the focus of >20 primary epidemiological publications and multiple review articles, but a consistent and reproducible association has yet to be confirmed. We systematically addressed possible causes of false-negative and false-positive association in >4,000 individuals from a multiethnic, prospective cohort study of prostate cancer, comprehensively studying genetic variation by microsatellite genotyping, direct resequencing of exons in advanced cancer cases, and haplotype analysis across the 180-kb AR genomic locus. These data failed to confirm that common genetic variation in the AR gene locus influences risk of prostate cancer. A systematic approach that assesses both coding and noncoding genetic variation in large and diverse patient samples can help clarify hypotheses about association between genetic variants and disease.

The Heritability of Prostate Cancer in the Nordic Twin Study of Cancer

Background: Prostate cancer is thought to be the most heritable cancer, although little is known about how this genetic contribution varies across age. Methods: To address this question, we undertook the worlds largest prospective study in the Nordic Twin Study of Cancer cohort, including 18,680 monozygotic (MZ) and 30,054 dizygotic (DZ) same-sex male twin pairs. We incorporated time-to-event analyses to estimate the risk concordance and heritability while accounting for censoring and competing risks of death, essential sources of biases that have not been accounted for in previous twin studies modeling cancer risk and liability. Results: The cumulative risk of prostate cancer was similar to that of the background population. The cumulative risk for twins whose co-twin was diagnosed with prostate cancer was greater for MZ than for DZ twins across all ages. Among concordantly affected pairs, the time between diagnoses was significantly shorter for MZ than DZ pairs (median, 3.8 versus 6.5 years, respectively). Genetic differences contributed substantially to variation in both the risk and the liability [heritability = 58% (95% confidence interval, 52%–63%)] of developing prostate cancer. The relative contribution of genetic factors was constant across age through late life with substantial genetic heterogeneity even when diagnosis and screening procedures vary. Conclusions: Results from the population-based twin cohort indicate a greater genetic contribution to the risk of developing prostate cancer when addressing sources of bias. The role of genetic factors is consistently high across age. Impact: Findings affect the search for genetic and epigenetic markers and frame prevention efforts. Cancer Epidemiol Biomarkers Prev; 23(11); 2303–10. ©2014 AACR.

Analysis of the 10q11 Cancer Risk Locus Implicates MSMB and NCOA4 in Human Prostate Tumorigenesis

Genome-wide association studies (GWAS) have established a variant, rs10993994, on chromosome 10q11 as being associated with prostate cancer risk. Since the variant is located outside of a protein-coding region, the target genes driving tumorigenesis are not readily apparent. Two genes nearest to this variant, MSMB and NCOA4, are strong candidates for mediating the effects of rs109939934. In a cohort of 180 individuals, we demonstrate that the rs10993994 risk allele is associated with decreased expression of two MSMB isoforms in histologically normal and malignant prostate tissue. In addition, the risk allele is associated with increased expression of five NCOA4 isoforms in histologically normal prostate tissue only. No consistent association with either gene is observed in breast or colon tissue. In conjunction with these findings, suppression of MSMB expression or NCOA4 overexpression promotes anchorage-independent growth of prostate epithelial cells, but not growth of breast epithelial cells. These data suggest that germline variation at chromosome 10q11 contributes to prostate cancer risk by influencing expression of at least two genes. More broadly, the findings demonstrate that disease risk alleles may influence multiple genes, and associations between genotype and expression may only be observed in the context of specific tissue and disease states.