Sex hormones, particularly the androgens, are important for the growth of the prostate gland and have been implicated in prostate cancer carcinogenesis, yet the determinants of endogenous steroid hormone levels remain poorly understood. Twin studies suggest a heritable component for circulating concentrations of sex hormones, although epidemiologic evidence linking steroid hormone gene variants to prostate cancer is limited. Here we report on findings from a comprehensive study of genetic variation at the CYP19A1 locus in relation to prostate cancer risk and to circulating steroid hormone concentrations in men by the Breast and Prostate Cancer Cohort Consortium (BPC3), a large collaborative prospective study. The BPC3 systematically characterized variation in CYP19A1 by targeted resequencing and dense genotyping; selected haplotype-tagging single nucleotide polymorphisms (htSNP) that efficiently predict common variants in U.S. and European whites, Latinos, Japanese Americans, and Native Hawaiians; and genotyped these htSNPs in 8,166 prostate cancer cases and 9,079 study-, age-, and ethnicity-matched controls. CYP19A1 htSNPs, two common missense variants and common haplotypes were not significantly associated with risk of prostate cancer. However, several htSNPs in linkage disequilibrium blocks 3 and 4 were significantly associated with a 5% to 10% difference in estradiol concentrations in men [association per copy of the two-SNP haplotype rs749292-rs727479 (A-A) versus noncarriers; P = 1 × 10-5], and with inverse, although less marked changes, in free testosterone concentrations. These results suggest that although germline variation in CYP19A1 characterized by the htSNPs produces measurable differences in sex hormone concentrations in men, they do not substantially influence risk of prostate cancer. (Cancer Epidemiol Biomarkers Prev 2009;18(10):2734–44)

Prostate cancer is the most commonly diagnosed cancer in males in developed countries, yet the etiologic risk factors for the disease are not well understood. The only established risk factors for the disease are age, family history of prostate cancer, and ethnicity. There has been considerable interest in the potential role of sex hormones in prostate cancer carcinogenesis, with a particular focus on androgens, which are important for the development, growth, and maintenance of the prostate gland. Finasteride, which blocks the metabolism of testosterone within the prostate, has been found to reduce the risk of prostate cancer (although the increased incidence of high-grade tumors on biopsy has led to controversy; ref. 1) and prostate tumors can be induced when testosterone, either alone or together with estradiol, is administered to laboratory animals (2). Estrogens have also been implicated in prostate biology and in the development of prostate cancer, via direct estrogen receptor–mediated effects and indirect effects, although data suggest the role of estrogens may vary with disease progression (3, 4). However, a recent re-analysis of the worldwide prospective data found no large associations between circulating androgen and estrogen concentrations in humans and prostate cancer risk (5).

Twin studies suggest a heritable component for circulating concentrations of sex hormones in men (6). The CYP19A1 gene has been identified as a candidate locus that may influence circulating sex hormone concentrations and risk of hormone-related cancers (7). CYP19A1 encodes aromatase, an enzyme that catalyses the conversion of the C19 androgens, androstenedione and testosterone, to the C18 estrogens, estrone and estradiol, respectively. This cytochrome P450 enzyme is expressed primarily in the gonads, as well as in peripheral sites including the prostate (8). In postmenopausal women, common variants in the CYP19A1 gene have been found to be associated with a 10% to 20% difference in circulating estrogen levels, and a number of studies have assessed the relationship of variants in CYP19A1 with risk of cancers of the breast and endometrium (9-12). However, published data on CYP19A1 in men in relation to sex hormones and prostate cancer are relatively limited, partly due to incomplete characterization of genetic variation at the locus of interest and small sample sizes (13-23).

In the present study, we examined the contribution of common genetic variation at the CYP19A1 locus to prostate cancer risk and to concentrations of serum sex hormones and sex hormone binding globulin (SHBG) in a large, collaborative investigation, the Breast and Prostate Cancer Cohort Consortium (BPC3; ref. 24).

Study Population

The BPC3 has been described in detail elsewhere (24). Briefly, the consortium includes large well-established cohorts assembled in the United States and Europe that have DNA for genotyping and extensive questionnaire data from cohort members. For prostate cancer, analyses include men from seven cohort studies: the American Cancer Society Cancer Prevention Study II (ACS CPS-II; ref. 25); the Alpha-Tocopherol, Beta-Carotene Cancer Prevention (ATBC) Study (26); the European Prospective Investigation into Cancer and Nutrition (EPIC) Cohort (itself comprising cohorts from Denmark, Germany, Great Britain, Greece, Italy, the Netherlands, Spain, and Sweden; ref. 27); the Health Professionals Follow-up Study (HPFS; ref. 28); the Hawaii/Los Angeles Multi-ethnic Cohort Study (MEC; ref. 29); the Physicians' Health Study (PHS; ref. 30); and the Prostate, Lung, Colorectal, and Ovarian (PLCO) Cancer Screening Trial (31). With the exception of the MEC and PLCO, these cohorts are composed predominantly of whites of European descent. The MEC contributes African American, Latino, Japanese American, and Native Hawaiian cases and controls recruited from Los Angeles and Hawaii. The PLCO includes over 400 African American participants. Cases of prostate cancer were identified through population-based cancer registries or self-report confirmed by medical records. The BPC3 data for prostate cancer consist of a series of matched nested case-control studies from each cohort; controls were matched within each cohort to cases on a number of potential confounding factors, including age (within 5 y), ethnicity, and in some cohorts, additional criteria, such as region of recruitment in EPIC (for further details on the selection of controls in each cohort see Supplementary Methods). The study protocol was approved by all institutional review boards.

SNP Discovery and Selection of Haplotype-Tagging Single Nucleotide Polymorphisms

We used a multistage approach to characterize genetic variation by initially resequencing the coding exons and exon-intron boundaries of CYP19A1 in a multiethnic panel of 95 advanced prostate cancer cases to identify putative functional alleles directly. The linkage disequilibrium (LD) structure of the locus was determined empirically in a multiethnic panel of 349 individuals from the MEC by genotyping a dense network of single nucleotide polymorphisms (SNP), including SNPs selected from dbSNP22

and the Celera SNP database23 and common missense SNPs found during resequencing (although not novel and can also be found in dbSNP) using the Sequenom and Illumina genotyping platforms at the Broad Institute/MIT Center for Genome Research. Haplotype-tagging SNPs (htSNP) for each haplotype block, determined by the confidence interval method of Gabriel et al. (32, 33), were chosen based on Rh2, a measure of the correlation between observed and predicted haplotypes based on the htSNP genotypes (34), to select a minimum set of SNPs that would achieve a Rh2 ≥0.7 for all common haplotypes among whites with an estimated frequency of ≥5%.

The LD pattern across CYP19A1 has been previously shown (9). Briefly, the genomic structure of the region consists of four LD blocks spanning 181 kb which was determined by 107 SNPs with minor allele frequency ≥0.05 among whites, and two missense SNPs with a frequency of >1% among whites in the multiethnic panel. In total 19 htSNPs (Supplementary Table S1) were selected to provide high predictability (R2H ≥0.7; ref. 35) of 27 common haplotypes (≥0.05 frequency in at least one ethnic group among the five ethnic groups in the multiethnic panel) across the four LD block regions (33, 36) with interblock distances <6 kb. Haplotype frequencies were similar for whites across cohorts (data not shown), whereas some differences in haplotype frequencies were seen among whites, African Americans, Japanese Americans, and Native Hawaiians.

Genotyping

Genotyping of the htSNPs and two missense SNPs in the 17,245 case patient and control participant samples was conducted using the TaqMan assay (Applied Biosystems) in four BPC3 laboratories. Initial quality control checks of the SNP assays were done at the manufacturer (ABI); an additional 500 test reactions were run by the BPC3 on the multiethnic reference panel; >99.5% concordance was observed across genotyping platforms. Assay characteristics for the htSNPs for CYP19A1 are available on the public website.24

Sequence validation for each SNP assay was done, and 100% concordance was observed25 (37). To assess interlaboratory variation, each center ran assays on a designated set of 94 samples from the SNP 500 cancer panel, showing completion and concordance rates of >99% (37). The internal quality of genotype data at each center was assessed by 5% to 10% blinded samples in duplicate or triplicate (depending on study), and intralaboratory concordance rates >99.5% were observed. Empty water wells were also included on each plate and positioned according to the individual laboratory's protocols. Hardy-Weinberg Equilibrium checks were carried out among the controls in each study and stratified by ethnicity in the MEC and by country for EPIC. No deviation in Hardy-Weinberg Equilibrium was observed (P < 0.01) across more than one study for any given assay.

Of the 8,248 prostate cancer cases and 9,312 controls sent for genotyping, at least 1 SNP was successfully genotyped for 8,166 (94%) cases and 9,079 (94%) controls. This study therefore comprises 8,166 case patients and 9,079 control participants. Among these men, we evaluated the relationship of prostate cancer risk with the htSNPs, with the common CYP19A1 haplotypes predicted among whites by the 19 htSNPs, and with the two common missense SNPs at the CYP19A1 locus, including SNPs R264C (rs700519) and T201M (rs28757184). In a subset of control participants from ATBC, EPIC, HPFS, PHS, and PLCO, we also investigated whether variation at this locus contributes to interindividual differences in circulating levels of sex hormones and SHBG, using measurements of steroid hormones made previously by individual cohorts (38-41).

Statistical Analyses

We used conditional logistic regression to estimate odds ratios (OR) for prostate cancer associated with carrying either one or two versus zero copies of the minor allele for each SNP. We estimated haplotype-specific OR using an expectation-substitution approach to account for haplotype uncertainty given unphased genotype data (42, 43). Haplotype frequencies and subject-specific expected haplotype indicators were calculated separately for each cohort (and country within EPIC or ethnicity in the MEC and PLCO). To test the global null hypothesis of no association between variation in common CYP19A1 haplotypes and risk of prostate cancer, and to control for type-I error over all the SNPs and haplotypes considered, we used a likelihood ratio test comparing a model with additive effects for each common haplotype (treating the most common haplotype as the referent) with the intercept-only model. For all cancer risk analyses, we tested for significance associations at the 0.01 level to minimize the chance of both false-positive and false-negative results.

We tested for heterogeneity in OR estimates across cohorts among white participants and by ethnicity. In addition, we calculated risk stratum–specific OR and tested for departures from a multiplicative interaction model to assess whether other risk factors for prostate cancer modify the association with htSNPs, missense SNPs, or common haplotypes, including age (at diagnosis), body mass index (BMI), and family history of prostate cancer. To assess the influence of genetic variation in CYP19A1 on prostate cancer diagnosed with an aggressive phenotype, we calculated stratum-specific OR for high-grade prostate cancer (defined as poorly differentiated or Gleason score ≥8), advanced-stage prostate cancer (stage C or D), and aggressive disease (defined as high-grade, advanced-stage prostate cancer, or death from prostate cancer).

We used fixed-effect models to evaluate associations of circulating steroid hormone and SHBG concentrations with CYP19A1 htSNPs, missense SNPs, and common haplotypes in cases and controls, with study cohort included as a fixed effect with a suitable nesting of batches within study where appropriate, and adjustment for age (in 5-year age groups). For these analyses, hormone and SHBG concentrations were logarithmically transformed for statistical analyses to approximately normalize their distributions. We report the P value from the test of trend across genotype groups.

Characteristics of Prostate Cancer Case Patients and Controls

The demographic and other characteristics of cases and controls from the seven cohorts are shown in Table 1. Most study participants were U.S. or European whites (75%), followed by African Americans (11%), Latinos (7%), Japanese Americans (5%), and Native Hawaiians (1%). Among participants, 14% of cases and 9% of controls reported a father or a brother with prostate cancer. Cases and controls were similar with respect to age, BMI, and height. Data on tumor stage and grade were available from six of the seven participating cohorts (all but PLCO). Stage information was available on 70% of genotyped prostate cancer cases, and of these, 19% had advanced disease (defined as stage C or D disease at diagnosis). Gleason score was recorded for 65% of genotyped cases, and 18% of cases had a Gleason sum of ≥8.

Table 1.

Characteristics of the study population by study, BPC3

ACS CPS-IIATBCEPICHPFS
CasesControlsCasesControlsCasesControlsCasesControls
Number 1,176 1,177 1,048 1,055 760 1,179 700 698 
Ethnicity (%)* 
    White 99 99 100 100 100 100 94 94 
    African American 
    Native Hawaiian 
    Japanese Americans 
    Latino 
Mean age at diagnosis(y) 70 70 70 69 65 65 69 69 
Mean BMI 26 26 26 26 27 27 25 26 
Family history available (n1,176 1,177 914 927 700 698 
Family history (% yes) 21 11 N/A N/A 20 15 
For cases: 
    Years of diagnoses 1992-2002 1986-2003 1991-2003 1994-2000 
    Stage info available (n1,142 651 424 607 
    Stage (% ≥ C) 11 31 16 15 
    Gleason score available 1,009 632 101 618 
    Gleason score (% ≥8) 11 25 16 10 
 
 MEC PHS PLCO TOTAL 
Cases Controls Cases Controls Cases Controls Cases Controls 
Number 2,320 2,290 887 1,052 1,275 1,628 8,166 9,079 
Ethnicity (%)* 
    White 20 20 95 94 92 80 75 75 
    African American 29 28 20 10 11   
    Native Hawaiian     
    Japanese Americans 20 21     
    Latino 28 28     
Mean age at diagnosis (y) 68 66 70 70 67 67 68 68 
Mean BMI 27 27 25 25 27 28 26 27 
Family history available (n2,111 2,096 1,275 1,628 6,176 6,526 
Family history (% yes) 12 N/A N/A 11 14 
For cases: 
    Years of diagnoses (range) 1995-2002 1982-2000 1994-2001 1982-2003 
    Stage info avail 2,180 698 5,702 
    Stage (% ≥C) 18 31 N/A 19 
    Gleason score available 2,234 703 5,279 
    Gleason score (% ≥8) 24 13 N/A 18 
ACS CPS-IIATBCEPICHPFS
CasesControlsCasesControlsCasesControlsCasesControls
Number 1,176 1,177 1,048 1,055 760 1,179 700 698 
Ethnicity (%)* 
    White 99 99 100 100 100 100 94 94 
    African American 
    Native Hawaiian 
    Japanese Americans 
    Latino 
Mean age at diagnosis(y) 70 70 70 69 65 65 69 69 
Mean BMI 26 26 26 26 27 27 25 26 
Family history available (n1,176 1,177 914 927 700 698 
Family history (% yes) 21 11 N/A N/A 20 15 
For cases: 
    Years of diagnoses 1992-2002 1986-2003 1991-2003 1994-2000 
    Stage info available (n1,142 651 424 607 
    Stage (% ≥ C) 11 31 16 15 
    Gleason score available 1,009 632 101 618 
    Gleason score (% ≥8) 11 25 16 10 
 
 MEC PHS PLCO TOTAL 
Cases Controls Cases Controls Cases Controls Cases Controls 
Number 2,320 2,290 887 1,052 1,275 1,628 8,166 9,079 
Ethnicity (%)* 
    White 20 20 95 94 92 80 75 75 
    African American 29 28 20 10 11   
    Native Hawaiian     
    Japanese Americans 20 21     
    Latino 28 28     
Mean age at diagnosis (y) 68 66 70 70 67 67 68 68 
Mean BMI 27 27 25 25 27 28 26 27 
Family history available (n2,111 2,096 1,275 1,628 6,176 6,526 
Family history (% yes) 12 N/A N/A 11 14 
For cases: 
    Years of diagnoses (range) 1995-2002 1982-2000 1994-2001 1982-2003 
    Stage info avail 2,180 698 5,702 
    Stage (% ≥C) 18 31 N/A 19 
    Gleason score available 2,234 703 5,279 
    Gleason score (% ≥8) 24 13 N/A 18 

Abbreviation: N/A, not applicable.

*May not add to 100% due to missing data.

Associations of CYP19A1 htSNPs, Missense SNPs, and Common Haplotypes with Prostate Cancer Risk

Genotype specific ORs for the 21 SNPs tested, including the two missense SNPs, are shown in Table 2 for analyses pooling all participants. There was no evidence of an association of the two missense SNPs with risk for prostate cancer (P > 0.50 for both for analyses). None of the htSNPs showed a significant association with prostate cancer risk at the 0.01 level. The SNP-specific ORs did not show significant heterogeneity across ethnicities or across cohorts among whites at the 0.01 level. Global tests of association between CYP19A1 common haplotypes and prostate cancer were also not significant at the 0.01 level (Table 3), and no statistically significant associations were observed between individual common haplotypes in each block and risk of prostate cancer (P > 0.01 for all).

Table 2.

CYP19A1 htSNPs, missense SNPs, and prostate cancer risk among all participants, BPC3

SNP (minor allele)Homozygote major alleleHeterozygoteHomozygote minor alleleP value for association*P value for heterogeneity
All subjects
rs2446405 (T) Cases/Controls 5,364/4,889 2,981/2,652 632/539  0.234 
OR (99% CI) 1 (ref) 0.98 (0.90-1.07) 0.97 (0.81-1.15) 0.782  
rs2445765 (C) Cases/Controls 5,932/5,373 2,705/2,451 321/264  0.408 
OR (99% CI) 1 (ref) 1.01 (0.92-1.10) 0.92 (0.74-1.16) 0.616  
rs2470144 (C) Cases/Controls 2,082/1,941 4,236/3,794 2,615/2,315  0.092 
OR (99% CI) 1 (ref) 0.96 (0.87-1.06) 0.98 (0.87-1.10) 0.558  
rs2445762 (C) Cases/Controls 4,501/4,140 3,641/3,105 702/681  0.162 
OR (99% CI) 1 (ref) 0.93 (0.85-1.01) 1.08 (0.92-1.25) 0.014  
rs1004984 (A) Cases/Controls 3,437/3,202 4,208/3,709 1,285/1,121  0.458 
OR (99% CI) 1 (ref) 0.95 (0.87-1.04) 0.96 (0.85-1.09) 0.361  
rs1902584 (T) Cases/Controls 7,523/6,801 1,413/1,214 68/65  0.174 
OR (99% CI) 1 (ref) 0.93 (0.83-1.04) 1.10 (0.70-1.74) 0.222  
rs3751591 (G) Cases/Controls 6,280/5,659 2,348/2,096 262/212  0.667 
OR (99% CI) 1 (ref) 0.97 (0.88-1.06) 0.90 (0.70-1.15) 0.388  
rs28566535/CV1664178 (C) Cases/Controls 7,116/6,450 1,572/1,351 307/279  0.083 
OR (99% CI) 1 (ref) 0.96 (0.85-1.08) 1.02 (0.80-1.31) 0.564  
rs2445759 (T) Cases/Controls 7,742/6,970 1,176/996 52/53  0.934 
OR (99% CI) 1 (ref) 0.95 (0.85-1.08) 1.14 (0.68-1.89) 0.487  
rs936306 (T) Cases/Controls 5,685/5,178 2,694/2,369 583/520  0.060 
OR (99% CI) 1 (ref) 0.97 (0.89-1.07) 1.06 (0.88-1.28) 0.469  
rs1902586 (A) Cases/Controls 7,162/6509 1,549/1,317 292/253  0.041 
OR (99% CI) 1.00 (ref) 0.95 (0.84-1.07) 1 (0.77-1.29) 0.536  
rs749292 (A) Cases/Controls 2,899/2,623 4,434/3,889 1,598/1,535  0.356 
OR (99% CI) 1 (ref) 0.97 (0.88-1.06) 1.07 (0.95-1.21) 0.055  
rs6493494 (A) Cases/Controls 3,247/2,895 4,268/3,728 1,332/1,318  0.381 
OR (99% CI) 1 (ref) 0.97 (0.89-1.07) 1.11 (0.98-1.25) 0.017  
rs1008805 (G) Cases/Controls 3,210/2,953 4,250/3,745 1,491/1,346  0.246 
OR (99% CI) 1 (ref) 0.94 (0.86-1.03) 0.95 (0.84-1.07) 0.195  
rs727479 (C) Cases/Controls 3,896/3,504 3,943/3,592 1,078/930  0.139 
OR (99% CI) 1 (ref) 1 (0.92-1.10) 0.94 (0.83-1.08) 0.467  
rs2414096 (G) Cases/Controls 1,875/1,705 4,338/3,870 2,707/2,443  0.491 
OR (99% CI) 1 (ref) 0.98 (0.88-1.08) 1 (0.89-1.12) 0.731  
rs28757184(A) (Thr201Met) Cases/Controls 8,253/7,487 569/439 8/7  0.989 
OR (99% CI) 1 (ref) 0.86 (0.72-1.02) 0.99 (0.26-3.84) 0.075  
rs700519 (A) (Arg264Cys) Cases/Controls 7,900/7,112 934/840 85/86  0.331 
OR (99% CI) 1 (ref) 1 (0.87-1.14) 1.15 (0.76-1.75) 0.670  
rs17601241(A) Cases/Controls 7,548/6,741 1,353/1,254 65/78  0.703 
OR (99% CI) 1 (ref) 1.04 (0.93-1.16) 1.33 (0.86-2.07) 0.180  
rs10046 (G) Cases/Controls 2,070/1,845 4,364/3,915 2,466/2,233  0.424 
OR (99% CI) 1 (ref) 1 (0.90-1.11) 1.02 (0.91-1.15) 0.842  
rs4646 (A) Cases/Controls 4,570/4,156 3,626/3,193 762/705  0.065 
OR (99% CI) 1 (ref) 0.96 (0.89-1.05) 1 (0.86-1.16) 0.520  
SNP (minor allele)Homozygote major alleleHeterozygoteHomozygote minor alleleP value for association*P value for heterogeneity
All subjects
rs2446405 (T) Cases/Controls 5,364/4,889 2,981/2,652 632/539  0.234 
OR (99% CI) 1 (ref) 0.98 (0.90-1.07) 0.97 (0.81-1.15) 0.782  
rs2445765 (C) Cases/Controls 5,932/5,373 2,705/2,451 321/264  0.408 
OR (99% CI) 1 (ref) 1.01 (0.92-1.10) 0.92 (0.74-1.16) 0.616  
rs2470144 (C) Cases/Controls 2,082/1,941 4,236/3,794 2,615/2,315  0.092 
OR (99% CI) 1 (ref) 0.96 (0.87-1.06) 0.98 (0.87-1.10) 0.558  
rs2445762 (C) Cases/Controls 4,501/4,140 3,641/3,105 702/681  0.162 
OR (99% CI) 1 (ref) 0.93 (0.85-1.01) 1.08 (0.92-1.25) 0.014  
rs1004984 (A) Cases/Controls 3,437/3,202 4,208/3,709 1,285/1,121  0.458 
OR (99% CI) 1 (ref) 0.95 (0.87-1.04) 0.96 (0.85-1.09) 0.361  
rs1902584 (T) Cases/Controls 7,523/6,801 1,413/1,214 68/65  0.174 
OR (99% CI) 1 (ref) 0.93 (0.83-1.04) 1.10 (0.70-1.74) 0.222  
rs3751591 (G) Cases/Controls 6,280/5,659 2,348/2,096 262/212  0.667 
OR (99% CI) 1 (ref) 0.97 (0.88-1.06) 0.90 (0.70-1.15) 0.388  
rs28566535/CV1664178 (C) Cases/Controls 7,116/6,450 1,572/1,351 307/279  0.083 
OR (99% CI) 1 (ref) 0.96 (0.85-1.08) 1.02 (0.80-1.31) 0.564  
rs2445759 (T) Cases/Controls 7,742/6,970 1,176/996 52/53  0.934 
OR (99% CI) 1 (ref) 0.95 (0.85-1.08) 1.14 (0.68-1.89) 0.487  
rs936306 (T) Cases/Controls 5,685/5,178 2,694/2,369 583/520  0.060 
OR (99% CI) 1 (ref) 0.97 (0.89-1.07) 1.06 (0.88-1.28) 0.469  
rs1902586 (A) Cases/Controls 7,162/6509 1,549/1,317 292/253  0.041 
OR (99% CI) 1.00 (ref) 0.95 (0.84-1.07) 1 (0.77-1.29) 0.536  
rs749292 (A) Cases/Controls 2,899/2,623 4,434/3,889 1,598/1,535  0.356 
OR (99% CI) 1 (ref) 0.97 (0.88-1.06) 1.07 (0.95-1.21) 0.055  
rs6493494 (A) Cases/Controls 3,247/2,895 4,268/3,728 1,332/1,318  0.381 
OR (99% CI) 1 (ref) 0.97 (0.89-1.07) 1.11 (0.98-1.25) 0.017  
rs1008805 (G) Cases/Controls 3,210/2,953 4,250/3,745 1,491/1,346  0.246 
OR (99% CI) 1 (ref) 0.94 (0.86-1.03) 0.95 (0.84-1.07) 0.195  
rs727479 (C) Cases/Controls 3,896/3,504 3,943/3,592 1,078/930  0.139 
OR (99% CI) 1 (ref) 1 (0.92-1.10) 0.94 (0.83-1.08) 0.467  
rs2414096 (G) Cases/Controls 1,875/1,705 4,338/3,870 2,707/2,443  0.491 
OR (99% CI) 1 (ref) 0.98 (0.88-1.08) 1 (0.89-1.12) 0.731  
rs28757184(A) (Thr201Met) Cases/Controls 8,253/7,487 569/439 8/7  0.989 
OR (99% CI) 1 (ref) 0.86 (0.72-1.02) 0.99 (0.26-3.84) 0.075  
rs700519 (A) (Arg264Cys) Cases/Controls 7,900/7,112 934/840 85/86  0.331 
OR (99% CI) 1 (ref) 1 (0.87-1.14) 1.15 (0.76-1.75) 0.670  
rs17601241(A) Cases/Controls 7,548/6,741 1,353/1,254 65/78  0.703 
OR (99% CI) 1 (ref) 1.04 (0.93-1.16) 1.33 (0.86-2.07) 0.180  
rs10046 (G) Cases/Controls 2,070/1,845 4,364/3,915 2,466/2,233  0.424 
OR (99% CI) 1 (ref) 1 (0.90-1.11) 1.02 (0.91-1.15) 0.842  
rs4646 (A) Cases/Controls 4,570/4,156 3,626/3,193 762/705  0.065 
OR (99% CI) 1 (ref) 0.96 (0.89-1.05) 1 (0.86-1.16) 0.520  

*P value from 2 d.f. likelihood ratio test for association.

P value for likelihood ratio test of heterogeneity of odds ratios across ethnicity (all subjects).

Table 3.

CYP19A1 common haplotypes in LD blocks 1, 2, 3, and 4 and prostate cancer risk among all participants, BPC3

Haplotype*0 copies1 copy2 copiesP value for associationP value for heterogeneity
Block 1: global χ2 = 8.10 on 8 d.f.; P = 0.423 0.057 
AGTTGA Case/Controls 2,370/2,656 3,835/4,287 1,934/2,073   
OR (99% CI) 1 (ref) 0.97 (0.88-1.07) 1.01 (0.90-1.13) 0.501  
AGCTGA Case/Controls 7,271/8,098 839/884 29/34   
OR (99% CI) 1 (ref) 1.09 (0.95-1.25) 1 (0.50-1.99) 0.298  
AGCTAA Case/Controls 7,354/8,169 758/817 27/30   
OR (99% CI) 1 (ref) 1.03 (0.89-1.19) 0.98 (0.49-2) 0.865  
AGCCAA Case/Controls 5,451/5,902 2,375/2,811 312/303   
OR (99% CI) 1 (ref) 0.92 (0.84-1) 1.12 (0.90-1.40) 0.011  
TGCCGA Case/Controls 7,586/8,366 501/593 52/56   
OR (99% CI) 1 (ref) 0.91 (0.76-1.10) 1.05 (0.62-1.77) 0.431  
TCCTAA Case/Controls 7,432/8,186 686/807 21/22   
OR (99% CI) 1 (ref) 0.97 (0.84-1.13) 1.15 (0.51-2.58) 0.817  
TCCTAT Case/Controls 6,871/7,567 1,208/1,385 60/64   
OR (99% CI) 1 (ref) 0.94 (0.84-1.05) 1.07 (0.67-1.72) 0.333  
TCCCGA Case/Controls 7,269/8,106 848/877 22/33   
OR (99% CI) 1 (ref) 1.09 (0.95-1.24) 0.74 (0.36-1.53) 0.175  
TGCTAA Case/Controls 8,020/8,881 114/134 5/1   
OR (99% CI) 1 (ref) 1.14 (0.78-1.67) 5.42 (0.26-111.40) 0.175  
Block 2: global χ2 = 2.24 on 4 d.f.; P = 0.691 0.132 
AAGCG Case/Controls 791/894 3,120/3,475 4,228/4,647   
OR (99% CI) 1 (ref) 0.96 (0.82-1.13) 0.97 (0.83-1.14) 0.842  
AATTG Case/Controls 7,653/8,473 473/530 13/13   
OR (99% CI) 1 (ref) 1.01 (0.84-1.22) 1.04 (0.37-2.94) 0.979  
ACGTA Case/Controls 7,271/7,993 727/851 141/172   
OR (99% CI) 1 (ref) 0.99 (0.83-1.17) 1 (0.71-1.40) 0.982  
GAGCG Case/Controls 7,001/7,766 1,104/1,202 34/47   
OR (99% CI) 1 (ref) 1 (0.89-1.13) 0.81 (0.45-1.47) 0.664  
GAGTA Case/Controls 7,462/8,241 647/738 30/37   
OR (99% CI) 1 (ref) 0.92 (0.79-1.08) 0.85 (0.44-1.63) 0.351  
Block 3: global χ2 = 2.39 on 3 d.f.; P = 0.495 0.019 
GGG Case/Controls 3,019/3,270 3,787/4,261 1,333/1,485   
OR (99% CI) 1 (ref) 0.94 (0.86-1.03) 0.94 (0.84-1.07) 0.213  
GGA Case/Controls 5,734/6,314 2,136/2,402 269/300   
OR (99% CI) 1 (ref) 0.98 (0.90-1.08) 1.04 (0.83-1.32) 0.788  
AGA Case/Controls 7,602/8,403 503/560 34/53   
OR (99% CI) 1 (ref) 1.06 (0.88-1.27) 0.76 (0.41-1.40) 0.350  
AAA Case/Controls 3,024/3,362 3,789/4,320 1,326/1,335   
OR (99% CI) 1 (ref) 0.97 (0.89-1.06) 1.10 (0.98-1.25) 0.017  
Block 4: global χ2 = 9.60 on 8 d.f.; P = 0.295 0.204 
AAGGGAC Case/Controls 2,860/3,201 3,800/4,254 1,479/1,561   
OR (99% CI) 1 (ref) 0.99 (0.90-1.08) 1.05 (0.93-1.19) 0.326  
AAAGGAC Case/Controls 7,694/8,454 438/554 6/8   
OR (99% CI) 1 (ref) 0.87 (0.73-1.04) 0.85 (0.21-3.50) 0.116  
AGGGGAC Case/Controls 7,719/8,517 414/491 6/8   
OR (99% CI) 1 (ref) 0.93 (0.77-1.12) 0.81 (0.20-3.33) 0.560  
AGGGGGC Case/Controls 7,745/8,550 351/420 43/46   
OR (99% CI) 1 (ref) 1.06 (0.83-1.34) 1 (0.55-1.82) 0.843  
AGGGGGA Case/Controls 8,008/8,847 127/164 4/4   
OR (99% CI) 1 (ref) 0.93 (0.65-1.32) 1.70 (0.26-11.29) 0.668  
AGGGAGA Case/Controls 6,814/7,598 1,250/1,351 75/66   
OR (99% CI) 1 (ref) 1.03 (0.92-1.15) 1.24 (0.79-1.93) 0.376  
AGGAGGC Case/Controls 7,221/8,009 849/938 70/69   
OR (99% CI) 1 (ref) 1 (0.87-1.15) 1.11 (0.69-1.76) 0.855  
CGGGGGC Case/Controls 5,994/6,679 1,959/2,131 185/206   
OR (99% CI) 1 (ref) 1.03 (0.93-1.13) 0.99 (0.75-1.29) 0.765  
CGGGGGA Case/Controls 5,397/5,913 2,429/2,745 313/358   
OR (99% CI) 1 (ref) 0.95 (0.87-1.04) 0.94 (0.76-1.16) 0.308  
Haplotype*0 copies1 copy2 copiesP value for associationP value for heterogeneity
Block 1: global χ2 = 8.10 on 8 d.f.; P = 0.423 0.057 
AGTTGA Case/Controls 2,370/2,656 3,835/4,287 1,934/2,073   
OR (99% CI) 1 (ref) 0.97 (0.88-1.07) 1.01 (0.90-1.13) 0.501  
AGCTGA Case/Controls 7,271/8,098 839/884 29/34   
OR (99% CI) 1 (ref) 1.09 (0.95-1.25) 1 (0.50-1.99) 0.298  
AGCTAA Case/Controls 7,354/8,169 758/817 27/30   
OR (99% CI) 1 (ref) 1.03 (0.89-1.19) 0.98 (0.49-2) 0.865  
AGCCAA Case/Controls 5,451/5,902 2,375/2,811 312/303   
OR (99% CI) 1 (ref) 0.92 (0.84-1) 1.12 (0.90-1.40) 0.011  
TGCCGA Case/Controls 7,586/8,366 501/593 52/56   
OR (99% CI) 1 (ref) 0.91 (0.76-1.10) 1.05 (0.62-1.77) 0.431  
TCCTAA Case/Controls 7,432/8,186 686/807 21/22   
OR (99% CI) 1 (ref) 0.97 (0.84-1.13) 1.15 (0.51-2.58) 0.817  
TCCTAT Case/Controls 6,871/7,567 1,208/1,385 60/64   
OR (99% CI) 1 (ref) 0.94 (0.84-1.05) 1.07 (0.67-1.72) 0.333  
TCCCGA Case/Controls 7,269/8,106 848/877 22/33   
OR (99% CI) 1 (ref) 1.09 (0.95-1.24) 0.74 (0.36-1.53) 0.175  
TGCTAA Case/Controls 8,020/8,881 114/134 5/1   
OR (99% CI) 1 (ref) 1.14 (0.78-1.67) 5.42 (0.26-111.40) 0.175  
Block 2: global χ2 = 2.24 on 4 d.f.; P = 0.691 0.132 
AAGCG Case/Controls 791/894 3,120/3,475 4,228/4,647   
OR (99% CI) 1 (ref) 0.96 (0.82-1.13) 0.97 (0.83-1.14) 0.842  
AATTG Case/Controls 7,653/8,473 473/530 13/13   
OR (99% CI) 1 (ref) 1.01 (0.84-1.22) 1.04 (0.37-2.94) 0.979  
ACGTA Case/Controls 7,271/7,993 727/851 141/172   
OR (99% CI) 1 (ref) 0.99 (0.83-1.17) 1 (0.71-1.40) 0.982  
GAGCG Case/Controls 7,001/7,766 1,104/1,202 34/47   
OR (99% CI) 1 (ref) 1 (0.89-1.13) 0.81 (0.45-1.47) 0.664  
GAGTA Case/Controls 7,462/8,241 647/738 30/37   
OR (99% CI) 1 (ref) 0.92 (0.79-1.08) 0.85 (0.44-1.63) 0.351  
Block 3: global χ2 = 2.39 on 3 d.f.; P = 0.495 0.019 
GGG Case/Controls 3,019/3,270 3,787/4,261 1,333/1,485   
OR (99% CI) 1 (ref) 0.94 (0.86-1.03) 0.94 (0.84-1.07) 0.213  
GGA Case/Controls 5,734/6,314 2,136/2,402 269/300   
OR (99% CI) 1 (ref) 0.98 (0.90-1.08) 1.04 (0.83-1.32) 0.788  
AGA Case/Controls 7,602/8,403 503/560 34/53   
OR (99% CI) 1 (ref) 1.06 (0.88-1.27) 0.76 (0.41-1.40) 0.350  
AAA Case/Controls 3,024/3,362 3,789/4,320 1,326/1,335   
OR (99% CI) 1 (ref) 0.97 (0.89-1.06) 1.10 (0.98-1.25) 0.017  
Block 4: global χ2 = 9.60 on 8 d.f.; P = 0.295 0.204 
AAGGGAC Case/Controls 2,860/3,201 3,800/4,254 1,479/1,561   
OR (99% CI) 1 (ref) 0.99 (0.90-1.08) 1.05 (0.93-1.19) 0.326  
AAAGGAC Case/Controls 7,694/8,454 438/554 6/8   
OR (99% CI) 1 (ref) 0.87 (0.73-1.04) 0.85 (0.21-3.50) 0.116  
AGGGGAC Case/Controls 7,719/8,517 414/491 6/8   
OR (99% CI) 1 (ref) 0.93 (0.77-1.12) 0.81 (0.20-3.33) 0.560  
AGGGGGC Case/Controls 7,745/8,550 351/420 43/46   
OR (99% CI) 1 (ref) 1.06 (0.83-1.34) 1 (0.55-1.82) 0.843  
AGGGGGA Case/Controls 8,008/8,847 127/164 4/4   
OR (99% CI) 1 (ref) 0.93 (0.65-1.32) 1.70 (0.26-11.29) 0.668  
AGGGAGA Case/Controls 6,814/7,598 1,250/1,351 75/66   
OR (99% CI) 1 (ref) 1.03 (0.92-1.15) 1.24 (0.79-1.93) 0.376  
AGGAGGC Case/Controls 7,221/8,009 849/938 70/69   
OR (99% CI) 1 (ref) 1 (0.87-1.15) 1.11 (0.69-1.76) 0.855  
CGGGGGC Case/Controls 5,994/6,679 1,959/2,131 185/206   
OR (99% CI) 1 (ref) 1.03 (0.93-1.13) 0.99 (0.75-1.29) 0.765  
CGGGGGA Case/Controls 5,397/5,913 2,429/2,745 313/358   
OR (99% CI) 1 (ref) 0.95 (0.87-1.04) 0.94 (0.76-1.16) 0.308  

*Alleles listed for htSNPs in 5′ to 3′ order: block 1 rs2446405, rs2445765, rs2470144, rs2445762, rs1004984, rs1902584; block 2 rs3751591, hCV1664178, rs2445759, rs936306, rs1902586; block 3, rs749292, rs64993494, rs1008805; block 4, rs727479, rs2414096, rs28757184 (Thr20Met), rs700519 (Arg264Cys), rs17601241, rs10046, rs4646.

P value from 2 d.f. likelihood ratio test for association.

P value for global test of heterogeneity of OR across ethnicity (all subjects).

Tumor Stage and Grade

We calculated ORs for risk of advanced disease by CYP19A1 htSNPs, missense SNPs, and common haplotypes, classifying disease severity by prostate cancer stage, grade, or a combined score of prostate tumor stage and histologic grade. For aggressive disease in relation to CYP19A1 htSNPs, rs2445762 in LD block 1 was significantly associated with high-grade disease with a significant decrease in risk being observed in heterozygotes [P2 d.f.test = 0.0002; OR in heterozygotes, 0.76; 99% confidence interval (99% CI), 0.62-0.93; OR in homozygotes, 1.15; 99% CI, 0.83-1.59], and similarly with the composite variable for aggressive disease (P = 0.0003). We found no significant association between other CYP19A1 htSNPs or missense SNPs and risk of tumor stage or grade or the composite score of aggressive disease at the 0.01 level. There was no evidence of any association between CYP19A1 common haplotypes and risk of aggressive prostate cancer, defined as high grade, advanced stage, or a composite variable.

Family History, Age at Diagnosis, and BMI

Tests for departures from multiplicative interaction models were null when we examined statistical interaction between CYP19A1 htSNPs, missense SNPs, or common haplotypes and prostate cancer risk with the following risk factors: family history (at least one first-degree relative diagnosed with prostate cancer versus none), age at diagnosis (≤65, >65 years), and BMI (<25, ≥25 <30, ≥30 kg/m2).

Associations of CYP19A1 htSNPs and Missense SNPs with Sex Hormone Concentrations

In analyses of genetic variation in CYP19A1 in relation to circulating sex hormone and SHBG concentrations, there were significant associations at the 0.01 level for a number of htSNPs with concentrations of estradiol, free estradiol, free testosterone, and androstanediol glucuronide. No significant associations were observed with concentrations of testosterone or SHBG, and neither of the missense SNPs were associated with concentrations of sex hormones or SHBG.

For estradiol, the most significant associations were observed for htSNPs in LD blocks 3 and 4, with Ptrend < 0.005 for every htSNP in block 3 and P ≤ 0.006 for rs727479 and rs10046 in block 4 (Table 4). These associations did not differ significantly among the three cohorts that measured estradiol, nor did they differ between cases and controls or by ethnicity, although only approximately 5% of the hormone data were from non-white participants. Percentage change in estradiol between homozygotes for the wild-type and the variant allele for these SNPs ranged from approximately 5% to 10%. There is a high degree of LD between previously defined LD blocks 3 and 4, and therefore many of the htSNPs are highly correlated (r2 ≥ 0.83 for SNP pairs rs749292 and rs6493494, and rs2414096 and rs10046; ref. 9). Haplotypes in blocks 3 and 4 were also strongly associated with estrogen levels, and the magnitude of the associations was similar to the independent tagging SNPs in these blocks (data not shown).

Table 4.

Geometric mean serum estradiol (E2) and testosterone (T) concentrations by htSNPs in LD blocks 3 and 4 and common nonsynonomous SNPs in CYP19A1 among cases and controls

RS numberGenotypeEstradiol, pmol/LFree estradiol, pmol/LTestosterone, nmol/LFree testosterone, nmol/L
nMean (95% CI)P trendnMean (95% CI)P trendnMean (95% CI)P trendnMean (95% CI)P trend
rs749292 G G 683 105.3 (98-113.1) 2.1 × 10-4 681 4.88 (4.48-5.32) 1.8 × 10-4 1,542 17.5 (16-19.3) 0.072 1,524 7.9 (7.35-8.49) 0.015 
G A 1,047 107.2 (99.9-115.1)  1,046 4.92 (4.55-5.36)  2,341 17.4 (15.8-19.1)  2,304 7.80 (7.28-8.39)  
A A 421 113 (104.9-121.7)  420 5.32 (4.88-5.80)  917 17 (15.4-18.7)  905 7.63 (7.115-8.25)  
rs6493494/hCV8234971 G G 695 105.5 (98.2-113.4) 3.8 × 10-3 692 4.88 (4.48- 5.32) 2.1 × 10-3 1,611 17.6 (16-19.3) 0.056 1,590 7.9 (7.35-8.53) 6.7 × 10-3 
G A 1,043 107.5 (100.2-115.3)  1042 4.95 (4.55-5.36)  2,307 17.3 (15.8-19)  2,273 7.8 (7.25-8.39)  
A A 388 111.6 (103.6-120.3)  387 5.25 (4.81-5.73)  846 17 (15.4- 18.7)  834 7.63 (7.07- 8.25)  
rs1008805 A A 743 109.8 (102.2-118) 3.3 × 10-3 742 5.14 (4.70-5.58) 3.1 × 10-3 1,602 17.1 (15.6-18.8) 0.089 1,581 7.73 (7.18-8.32) 0.011 
G A 1,041 106.8 (99.42-114.7)  1,040 4.92 (4.51-5.32)  2,351 17.6 (16.1- 19.4)  2,316 7.94 (7.35- 8.53)  
G G 356 104.1 (96.5-112.3)  353 4.84 (4.4-5.28)  841 17.5 (15.9-19.3)  830 7.94 (7.38-8.56)  
rs727479 A A 916 110.5 (103-118.6) 1.2 × 10-5 913 5.14 (4.7-5.58) 1.9 × 10-4 2,025 17.1 (15.5-18.7) 0.037 1,996 7.63 (7.07-8.22) 1.8 × 10-4 
A C 950 107.3 (100-115.3)  949 4.95 (4.55-5.39)  2,175 17.4 (15.8-19.1)  2,145 7.90 (7.35-8.52)  
C C 267 100.8 (93.3-108.9)  266 4.66 (4.26-5.14)  587 17.7 (16-19.5)  579 7.94 (7.35-8.6)  
rs2414096 A A 537 110 (102.2-118.4) 0.066 534 5.10 (4.70-5.58) 0.094 1,218 17.2 (15.6-18.9) 0.32 1,198 7.63 (7.07-8.22) 7.7 × 10-3 
G A 1,088 107.1 (99.8-114.9)  1,087 4.95 (4.55-5.36)  2,404 17.3 (15.7-19)  2,373 7.84 (7.28-8.42)  
G G 523 106.4 (98.9-114.5)  522 4.92 (4.51-5.39)  1,179 17.5 (15.9-19.2)  1,164 7.9 (7.35-8.53)  
rs28757184/Thr201Met G G 1,985 107.8 (100.6-115.5) 0.10 1,981 4.99 (4.59-5.43) 0.055 4,426 17.3 (15.8-19) 0.17 4,362 7.80 (7.25-8.39) 0.16 
G A 137 102.0 (93.7-111) 137 4.59 (4.15-5.1)  329 17.9 (16.1-19.8)  326 7.94 (7.32-8.6)  
A A 135.8 (96.7-190.5) 7.67 (5.1-11.49)  18.4 (12.7-26.6)  10.37 (7.77-13.8)  
rs700519/Arg264Cys G G 2,000 107.5 (100.2-115.3) 0.60 1,995 4.99 (4.59-5.39) 0.55 4,438 17.5 (15.9-19.2) 0.13 4,374 7.87 (7.32-8.46) 0.079 
G A 151 108.1 (99.8-117.1)  151 4.99 (4.51-5.47)  283 16.8 (15.2-18.6)  282 7.59 (7-8.22)  
A A 67.5 (48-95)  3.19 (2.13-4.77)  22 17.1 (14-20.7)  21 7.73 (6.62-9.01)  
rs17601241 G G 1,799 107 (99.8-114.8) 0.042 1,795 4.95 (4.55-5.36) 0.044 4,006 17.4 (15.8-19.1) 0.22 3,950 7.8 (7.28-8.42) 0.22 
G A 356 110.2 (102.2-118.8)  356 5.1 (4.66-5.58)  786 17.2 (15.6-18.9)  779 7.73 (7.18-8.36)  
A A 16 117 (997-137.3)  16 5.65 (4.7-6.86)  37 16.1 (13.7-18.8)  35 7.25 (6.38-8.25)  
rs10046 A A 572 111.5 (103.7-120) 5.7 × 10-3 569 5.21 (4.77-5.65) 0.014 1,300 17.2 (15.6-18.9) 0.45 1,279 7.7 (7.14-8.29) 0.033 
G A 1,070 106.5 (99.3-114.3)  1,069 4.92 (4.51-5.36)  2,380 17.3 (15.8-19)  2,347 7.80 (7.28-8.42)  
G G 485 106.3 (98.8-114.4)  484 4.95 (4.51-5.39)  1,082 17.4 (15.8-19.1)  1,070 7.90 (7.35-8.53)  
rs4646 C C 1,113 107.4 (100-115.2) 0.930 1,110 4.95 (4.55-5.39) 0.46 2,567 17.3 (15.8-19) 0.88 2,531 7.77 (7.21-8.36) 0.51 
A C 850 107.7 (100.2-115.8)  849 5.03 (4.62-5.47)  1,870 17.2 (15.7-18.9)  1,842 7.8 (7.25-8.39)  
A A 171 107.2 (98.8-116.4)  171 4.99 (4.51-5.51)  363 17.4 (15.7-19.2)  360 7.87 (7.25-8.53)  
RS numberGenotypeEstradiol, pmol/LFree estradiol, pmol/LTestosterone, nmol/LFree testosterone, nmol/L
nMean (95% CI)P trendnMean (95% CI)P trendnMean (95% CI)P trendnMean (95% CI)P trend
rs749292 G G 683 105.3 (98-113.1) 2.1 × 10-4 681 4.88 (4.48-5.32) 1.8 × 10-4 1,542 17.5 (16-19.3) 0.072 1,524 7.9 (7.35-8.49) 0.015 
G A 1,047 107.2 (99.9-115.1)  1,046 4.92 (4.55-5.36)  2,341 17.4 (15.8-19.1)  2,304 7.80 (7.28-8.39)  
A A 421 113 (104.9-121.7)  420 5.32 (4.88-5.80)  917 17 (15.4-18.7)  905 7.63 (7.115-8.25)  
rs6493494/hCV8234971 G G 695 105.5 (98.2-113.4) 3.8 × 10-3 692 4.88 (4.48- 5.32) 2.1 × 10-3 1,611 17.6 (16-19.3) 0.056 1,590 7.9 (7.35-8.53) 6.7 × 10-3 
G A 1,043 107.5 (100.2-115.3)  1042 4.95 (4.55-5.36)  2,307 17.3 (15.8-19)  2,273 7.8 (7.25-8.39)  
A A 388 111.6 (103.6-120.3)  387 5.25 (4.81-5.73)  846 17 (15.4- 18.7)  834 7.63 (7.07- 8.25)  
rs1008805 A A 743 109.8 (102.2-118) 3.3 × 10-3 742 5.14 (4.70-5.58) 3.1 × 10-3 1,602 17.1 (15.6-18.8) 0.089 1,581 7.73 (7.18-8.32) 0.011 
G A 1,041 106.8 (99.42-114.7)  1,040 4.92 (4.51-5.32)  2,351 17.6 (16.1- 19.4)  2,316 7.94 (7.35- 8.53)  
G G 356 104.1 (96.5-112.3)  353 4.84 (4.4-5.28)  841 17.5 (15.9-19.3)  830 7.94 (7.38-8.56)  
rs727479 A A 916 110.5 (103-118.6) 1.2 × 10-5 913 5.14 (4.7-5.58) 1.9 × 10-4 2,025 17.1 (15.5-18.7) 0.037 1,996 7.63 (7.07-8.22) 1.8 × 10-4 
A C 950 107.3 (100-115.3)  949 4.95 (4.55-5.39)  2,175 17.4 (15.8-19.1)  2,145 7.90 (7.35-8.52)  
C C 267 100.8 (93.3-108.9)  266 4.66 (4.26-5.14)  587 17.7 (16-19.5)  579 7.94 (7.35-8.6)  
rs2414096 A A 537 110 (102.2-118.4) 0.066 534 5.10 (4.70-5.58) 0.094 1,218 17.2 (15.6-18.9) 0.32 1,198 7.63 (7.07-8.22) 7.7 × 10-3 
G A 1,088 107.1 (99.8-114.9)  1,087 4.95 (4.55-5.36)  2,404 17.3 (15.7-19)  2,373 7.84 (7.28-8.42)  
G G 523 106.4 (98.9-114.5)  522 4.92 (4.51-5.39)  1,179 17.5 (15.9-19.2)  1,164 7.9 (7.35-8.53)  
rs28757184/Thr201Met G G 1,985 107.8 (100.6-115.5) 0.10 1,981 4.99 (4.59-5.43) 0.055 4,426 17.3 (15.8-19) 0.17 4,362 7.80 (7.25-8.39) 0.16 
G A 137 102.0 (93.7-111) 137 4.59 (4.15-5.1)  329 17.9 (16.1-19.8)  326 7.94 (7.32-8.6)  
A A 135.8 (96.7-190.5) 7.67 (5.1-11.49)  18.4 (12.7-26.6)  10.37 (7.77-13.8)  
rs700519/Arg264Cys G G 2,000 107.5 (100.2-115.3) 0.60 1,995 4.99 (4.59-5.39) 0.55 4,438 17.5 (15.9-19.2) 0.13 4,374 7.87 (7.32-8.46) 0.079 
G A 151 108.1 (99.8-117.1)  151 4.99 (4.51-5.47)  283 16.8 (15.2-18.6)  282 7.59 (7-8.22)  
A A 67.5 (48-95)  3.19 (2.13-4.77)  22 17.1 (14-20.7)  21 7.73 (6.62-9.01)  
rs17601241 G G 1,799 107 (99.8-114.8) 0.042 1,795 4.95 (4.55-5.36) 0.044 4,006 17.4 (15.8-19.1) 0.22 3,950 7.8 (7.28-8.42) 0.22 
G A 356 110.2 (102.2-118.8)  356 5.1 (4.66-5.58)  786 17.2 (15.6-18.9)  779 7.73 (7.18-8.36)  
A A 16 117 (997-137.3)  16 5.65 (4.7-6.86)  37 16.1 (13.7-18.8)  35 7.25 (6.38-8.25)  
rs10046 A A 572 111.5 (103.7-120) 5.7 × 10-3 569 5.21 (4.77-5.65) 0.014 1,300 17.2 (15.6-18.9) 0.45 1,279 7.7 (7.14-8.29) 0.033 
G A 1,070 106.5 (99.3-114.3)  1,069 4.92 (4.51-5.36)  2,380 17.3 (15.8-19)  2,347 7.80 (7.28-8.42)  
G G 485 106.3 (98.8-114.4)  484 4.95 (4.51-5.39)  1,082 17.4 (15.8-19.1)  1,070 7.90 (7.35-8.53)  
rs4646 C C 1,113 107.4 (100-115.2) 0.930 1,110 4.95 (4.55-5.39) 0.46 2,567 17.3 (15.8-19) 0.88 2,531 7.77 (7.21-8.36) 0.51 
A C 850 107.7 (100.2-115.8)  849 5.03 (4.62-5.47)  1,870 17.2 (15.7-18.9)  1,842 7.8 (7.25-8.39)  
A A 171 107.2 (98.8-116.4)  171 4.99 (4.51-5.51)  363 17.4 (15.7-19.2)  360 7.87 (7.25-8.53)  

Abbreviation: 95% CI, 95% confidence interval.

The two SNPs most significantly associated with estradiol concentrations, rs749292 and rs727479, were only modestly correlated with each other (r2 = 0.46), and each remained significantly associated with circulating levels when modeled concurrently (Ptrend = 0.002 and Ptrend = 0.006, respectively). A two-SNP haplotype (A-A) comprising these SNPs was found to be a more significant predictor of estradiol concentrations (Ptrend = 1 × 10-5; Table 5); however, it accounted for only 0.75% of the variation in estradiol concentrations, and when we examined risk of prostate cancer in relation to this two-SNP haplotype we observed no significant association with risk of disease at the 0.01 significance level (P = 0.028, OR in men with one copy of the A-A haplotype, 0.96; 99% CI, 0.88-1.06; OR for two copies, 1.08; 99% CI, 0.96-1.22).

Table 5.

Estradiol and testosterone concentrations* by haplotype A-A (rs749292-rs727479)

Number of copies of A-A haplotypeP trend
012
Estradiol 
    n 698 1,088 427 1 × 10-5 
    Mean 109.5 113.6 119.3  
    (95% CI) (57.3-209.2) (60.1-214.7) (61.7-230.6)  
Free estradiol 
    n 696 1,086 425 6.6 × 10-5 
    Mean 1.45 1.55 1.66  
    (95% CI) (0.46-4.55) (0.51-4.71) (0.55-4.97)  
Testosterone 
    n 1,573 2,423 929 0.099 
    Mean 16.2 16.0 15.7  
    (95% CI) (7.4-35.5) (7.2-35.7) (6.9-35.4)  
Free testosterone 
    n 1,555 2,385 918 0.020 
    Mean 2.29 2.32 2.26  
    (95% CI) (0.97-5.39) (1.01-5.3) (0.96-5.29)  
Free testosterone/free estradiol ratio 
    n 690 1,080 423 8 × 10-6 
    Mean 0.67 0.69 0.74  
    (95% CI) (0.30-1.48) (0.31-1.53) (0.33-1.67)  
Number of copies of A-A haplotypeP trend
012
Estradiol 
    n 698 1,088 427 1 × 10-5 
    Mean 109.5 113.6 119.3  
    (95% CI) (57.3-209.2) (60.1-214.7) (61.7-230.6)  
Free estradiol 
    n 696 1,086 425 6.6 × 10-5 
    Mean 1.45 1.55 1.66  
    (95% CI) (0.46-4.55) (0.51-4.71) (0.55-4.97)  
Testosterone 
    n 1,573 2,423 929 0.099 
    Mean 16.2 16.0 15.7  
    (95% CI) (7.4-35.5) (7.2-35.7) (6.9-35.4)  
Free testosterone 
    n 1,555 2,385 918 0.020 
    Mean 2.29 2.32 2.26  
    (95% CI) (0.97-5.39) (1.01-5.3) (0.96-5.29)  
Free testosterone/free estradiol ratio 
    n 690 1,080 423 8 × 10-6 
    Mean 0.67 0.69 0.74  
    (95% CI) (0.30-1.48) (0.31-1.53) (0.33-1.67)  

*Hormone values are geometric means and 95% confidence intervals.

Full results for all sex hormones and SHBG in relation to CYP19A1 htSNPs are shown in Supplementary Table S2. Findings for free estradiol were broadly similar to those observed for estradiol, both with respect to level of statistical significance and the percentage change in hormone level by genotype. Significant associations were also observed between CYP19A1 htSNPs in LD blocks 3 and 4 and the ratio of free estradiol to free testosterone, as an index of aromatase activity, with trends mirroring those observed for estradiol concentrations.

For testosterone, there were no significant associations with htSNPs at the 0.01 significance level, although weak associations that did not reach statistical significance were observed with the three htSNPs in LD block 3 (Ptrend < 0.09), and with rs727479 in LD block 4 (P = 0.04); changes in testosterone concentrations by genotype were the inverse of those seen for estradiol concentrations, and percentage changes in hormone concentrations were approximately 2% to 3% for htSNPs in LD block 3 and 4% for rs727479. For free testosterone concentrations, however, significant or borderline significant associations were observed with a number of htSNPs in LD blocks 3 and 4 (P < 0.01 for rs6493494, rs727479, and rs10046), with trends in free testosterone concentrations being the inverse of those observed for estradiol and free estradiol (Table 4) and percentage changes in free testosterone being approximately 3% to 4%.

For androstranediol glucuronide, significant associations were observed between androstanediol glucuronide and htSNPs rs28566535/CV1664178 and rs1902586 in LD block 2 (Ptrend = 3 × 10-4 and 0.001, respectively) and with rs727479 in LD block 4 (Ptrend = 0.009; Supplementary Table S2). The percentage changes in androstranediol glucuronide concentration between the wild-type and the variant allele were 18.7%, 13.9%, and 6.3%, respectively. We observed no significant associations between CYP19A1 htSNPs and concentrations of SHBG (Supplementary Table S2).

We investigated common genetic variation at the CYP19A1 locus in relation to prostate cancer risk and hormone levels in a large, collaborative investigation (BPC3; ref. 24). We evaluated both haplotype patterns in four well-characterized LD blocks and two common missense variants, and found no evidence that this gene harbors a prostate cancer susceptibility allele. In addition, this is the first large study that has comprehensively assessed genetic variation in CYP19A1 in relation to sex hormones and SHBG concentrations in men. Our results provide evidence for significant associations of a number of htSNPs in CYP19A1 with circulating concentrations of estradiol, free estradiol, free testosterone, and androstranediol glucuronide.

Ten studies on the relationship between genetic variation in CYP19A1 and risk of prostate cancer have been published to date (13-18, 20-23), with inconsistent findings. These studies only considered a limited number of variants across the locus, and with a few exceptions (15, 20, 21) were small, being underpowered to detect the modest magnitude of effect anticipated for a common low-penetrance susceptibility allele (relative risk <1.5). These variants include a tetranucleotide repeat (14, 16, 21-23), two nonsynonymous mutations (13, 15, 17, 18, 20), a G/A SNP in the 3`untranslated region (rs10046; ref. 20), and six other genetic variants identified in screening of prostate cancer patients (15). The null association with the missense mutation R264C in the current study contrasts with those from three small previous studies (13, 17, 18), but is in agreement with null results from two large recent studies (15, 20), as is our null finding for rs10046 (20). Our results suggesting no strong association with T201M, however, contrast with those from the one previous study, which reported an increase in risk, especially in association with low-grade organ-confined disease (15). The genome-wide association studies of prostate cancer also provide data on the CYP19A1 locus but have not implicated the region. For example, in a genome-wide association study in the Cancer Genetic Markers of Susceptibility (CGEMS) project of 1,172 prostate cases and 1,157 controls, 46 SNPs that met quality control metrics were evaluated across CYP19A1 (from 30 kb upstream of the ATG and 30 kb downstream of the polyA tail; ref. 44). Using HapMap Phase 2 CEU samples, the 46 SNPs genotyped tagged (using r2 >0.8) a total 164 SNPs in 40 bins, which represents 74.9% of all CEU HapMap SNPs with minor allele frequency >5%. In CGEMS, no SNPs had a P < 0.01 in the adjusted trend test; rs2124873 displayed the smallest p value (P ≥ 0.03) in CGEMS.

The findings from the current study on prostate cancer risk in relation to genetic variation in CYP19A1 provide no evidence that risks differ by ethnicity, family history, age, or BMI. Our findings also do not support a strong association with severity of disease, with one exception. We observed a statistically significant association between rs2445762 and risk of high-grade disease and a composite score of aggressive disease, with a decreased risk being observed in heterozygotes, but not in homozygotes, for the variant allele. Given the multiple-testing in this study and the small number of cases with two copies of the rs2445762 variant allele (91 and 191 cases with high-grade and aggressive disease, respectively), these associations with aggressive prostate cancer are likely due to chance; replication is warranted in further large prostate cancer studies that have detailed information on prostate cancer phenotype.

Despite the lack of evidence for an association between genetic variants and prostate cancer risk, we found evidence for an association between CYP19A1 variants and circulating sex hormone concentrations. The strongest associations between CYP19A1 variation and hormone concentrations were observed for htSNPs in LD blocks 3 and 4. These findings are consistent with those from a recent report on 5,531 men from Sweden and the United States that found that SNP rs2470152 in intron 1 of CYP19A1 was associated with serum estradiol (8% to 13% difference between AA and GG homozygotes in the three cohorts studied; P = 2 × 10-14 for all cohorts combined; ref. 45). For the two most strongly associated SNPs in the current study, the LD relationship with rs2470152 was strong for rs749292 (D' = 0.96, r2 = 0.65) and weaker LD for rs727479 (D' = 0.60, r2 = 0.19), and given these allelic associations, the findings with estradiol for these SNPs (rs749292, 7% difference between homozygotes, P = 2 × 10-4; rs727479, 9% difference, P = 1 × 10-5) are comparable with the Swedish data. Other published studies on sex hormones and CYP19A1 variation in men have had small sample sizes, and results have been conflicting (19, 46-48). Data from the current study for estradiol and free estradiol in relation to CYP19A1 variants are also broadly consistent with those previously reported for women (9-11, 49, 50), including findings from 3,400 postmenopausal women participating in BPC3 (9). For five of seven variants in CYP19A1 LD blocks 3 and 4 found to be significantly associated with estradiol concentrations in women in BPC3, we observed significant associations in men. However, the strength and the magnitude of these associations among men in the current study are somewhat weaker than those observed among postmenopausal women; in men we found variants to be significantly associated with a 5% to 10% difference in estradiol concentrations, whereas in postmenopausal women in BPC3 these variants were significantly associated with a 10% to 20% difference in endogenous estrogen levels (9). It is possible that although CYP19A1 variants influence aromatase activity, the impact on circulating hormone concentrations in men might be relatively small because of homeostasis of free testosterone by the hypothalamic-pituitary-gonadal feedback loop.

We found no evidence for a significant association of total testosterone with CYP19A1 variation; however, we found several variants in LD blocks 3 and 4 to be significantly associated with a 2% to 4% difference in levels of free testosterone, with the trend in hormone concentration being the reverse of that seen for estradiol. An explanation for these findings may lie in the fact that free testosterone is the substrate for conversion to estradiol by aromatase. The association found between androstanediol glucuronide and genetic variation in LD blocks 2 and 4 of CYP19A1 is also compatible with an influence of CYP19A1 genetic variants on aromatase activity as androstanediol glucuronide is an end metabolite of testosterone (51).

Overall, our findings that variants in CYP19A1 were associated with small differences in circulating hormone concentrations but have no detectable effect on risk of prostate cancer are consistent with what is known on the association of hormones with risk. A recent collaborative re-analysis of the worldwide data reported no strong association between serum concentrations of sex hormones and risk of prostate cancer (5), with differences in serum sex hormone concentrations between the highest and lowest fourths of the distribution being in the order of 2-fold (38). Thus, the small differences (5-10%) in sex hormones in relation to CYP19A1 variants may be of insufficient magnitude to have a detectable influence on risk of prostate cancer. The relationship between circulating hormone concentrations and intraprostatic hormone levels, however, remains unclear, and the authors of the collaborative re-analysis concluded that any biological interpretation of their results for serum hormones must be viewed with caution (5). Aromatase is expressed in the prostate, and local estrogen production may lead to intraprostatic levels exceeding those in the circulation, with prostate cells being exposed to estradiol from the blood plus locally produced estradiol (3, 52, 53). However, the null association in the current study between CYP19A1 variants and prostate cancer risk does not lend support to the hypothesis that intraprostatic sex hormone metabolism by aromatase is strongly associated with risk because it would be anticipated that CYP19A1 variation might have a similar effect on intraprostatic sex hormone concentrations as on circulating hormones.

The strengths of this study lie in the scale of the analysis, both with respect to the comprehensive screening for prostate cancer susceptibility alleles across the entire gene region facilitated by the consortium's cost-effective haplotype-tagging approach, and with respect to the large sample size, which makes it possible to conduct adequately powered subgroup analyses by potential prostate cancer risk factors and by tumor characteristics. In this study, with over 8,000 case patients and 9,000 control participants, we have >95% power to detect a dominant effect or log-additive OR of 1.3 for an allele with 5% minor allele frequency at the 0.001 level. Furthermore, with respect to subgroup analyses we still have, for example, >95% power to detect a stratum-specific dominant OR of 1.7 for a 5% frequency variant at the 0.001 level when the stratum consists of only 20% of the sample. The current study, however, had less power to assess whether variation in CYP19A1 is associated with risk of prostate cancer in non-white ethnic groups. Further analyses may clarify associations in non-white populations as sample sizes are increased with longer follow-up or the addition of new cohorts.

We have studied the importance of variation of CYP19A1 to steroid hormone concentrations and to prostate cancer risk. Ultimately, however, the importance in variation in CYP19A1 on endogenous hormone concentrations should be assessed in the context of variation in the many other genes involved in steroid hormone biosynthesis and metabolism pathways, as well as in genes involved in steroid hormone-binding and -receptor pathways. Although variation in individual genes that encode enzymes in the hormone metabolism pathways may individually only result in modest differences in hormone concentrations, a combination of several genes with functional variants could result in a cumulative large effect that would endure over the course of a human lifespan (54). Indeed, twin studies suggest that additive genetic factors may account for up to 57% and 25% of the total variation in circulating testosterone and estradiol concentrations, respectively, in men (6). Thus, the development of a multifactorial score for the prediction of hormone concentrations within large-scale studies, such as is planned within BPC3, may provide important insights into the determinants of endogenous hormone concentrations and risk for the diseases and disorders that they influence. Such conditions are not limited to cancer and may also include, for example, vascular disease (55), diabetes (56), and those related to cognitive decline (57, 58), and to bone metabolism (19, 45, 46).

In summary, results from this study suggest that although germline mutations in CYP19A1 produce measurable differences in steroid hormone concentrations in men, they do not substantially influence the risk of prostate cancer. These findings are consistent with a reanalysis of worldwide data that found no large associations between serum concentrations of sex hormones and prostate cancer risk. Our results for steroid hormone concentrations in relation to variants in CYP19A1, however, may have wider relevance for other conditions of public health importance in men.

No potential conflicts of interest were disclosed.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

We gratefully acknowledge the participants in the component cohort studies.

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Supplementary data