Genome-wide association study of prostate cancer
identifies a second risk locus at 8q24
Meredith Yeager1,2, Nick Orr3, Richard B Hayes2, Kevin B Jacobs4, Peter Kraft5, Sholom Wacholder2,
Mark J Minichiello6, Paul Fearnhead7, Kai Yu2, Nilanjan Chatterjee2, Zhaoming Wang1,2, Robert Welch1,2,
Brian J Staats1,2, Eugenia E Calle8, Heather Spencer Feigelson8, Michael J Thun8, Carmen Rodriguez8,
Demetrius Albanes2, Jarmo Virtamo9, Stephanie Weinstein2, Fredrick R Schumacher5, Edward Giovannucci10,
Walter C Willett10, Geraldine Cancel-Tassin11, Olivier Cussenot11, Antoine Valeri11, Gerald L Andriole12,
Edward P Gelmann13, Margaret Tucker2, Daniela S Gerhard14, Joseph F Fraumeni Jr2, Robert Hoover2,
David J Hunter2,5, Stephen J Chanock2,3& Gilles Thomas2
Recently, common variants on human chromosome 8q24
were found to be associated with prostate cancer risk. While
conducting a genome-wide association study in the Cancer
Genetic Markers of Susceptibility project with 550,000 SNPs in
a nested case-control study (1,172 cases and 1,157 controls of
European origin), we identified a new association at 8q24 with
an independent effect on prostate cancer susceptibility. The
most significant signal is 70 kb centromeric to the previously
reported SNP, rs1447295, but shows little evidence of linkage
disequilibrium with it. A combined analysis with four
additional studies (total: 4,296 cases and 4,299 controls)
confirms association with prostate cancer for rs6983267 in
the centromeric locus (P ¼ 9.42 ? 10–13; heterozygote odds
ratio (OR): 1.26, 95% confidence interval (c.i.): 1.13–1.41;
homozygote OR: 1.58, 95% c.i.: 1.40–1.78). Each SNP
remained significant in a joint analysis after adjusting for
the other (rs1447295 P ¼ 1.41 ? 10–11; rs6983267 P ¼ 6.62 ?
10–10). These observations, combined with compelling evidence
for a recombination hotspot between the two markers, indicate
the presence of at least two independent loci within 8q24 that
contribute to prostate cancer in men of European ancestry. We
estimate that the population attributable risk of the new locus,
marked by rs6983267, is higher than the locus marked by
rs1447295 (21% versus 9%).
In developed countries, prostate cancer is the most common non-
cutaneous malignancy in men, yet a positive family history of prostate
cancer and ethnic background are the only established risk factors1–3.
In the USA, men of African descent are at greater risk than those of
European descent2. Two independent studies previously demonstrated
a single nucleotide polymorphism (SNP) in 8q24, rs1447295, is
associated with prostate cancer risk4,5. In one study, a stronger
association was observed in African Americans4, while the other
study reported a stronger association with aggressive prostate cancer5.
A third larger study, nested in seven USA and European cohorts and
including more than 7,000 prostate cancer cases and 8,000 matched
controls, reported an association between rs1447295 and increased
risk for prostate cancer in Caucasian men, regardless of age at
diagnosis (P ¼ 4.00 ? 10–19)6.
We conducted a genome-wide association study (GWAS) of 550,000
SNPs in 1,172 affected individuals (484 with nonaggressive prostate
cancer, Gleason o7 and stage A/B; 688 aggressive prostate cancer,
Gleason Z7 and/or stage C/D) and 1,157 controls using an incidence
density sampling strategy in the Prostate, Lung, Colon and Ovarian
(PLCO) Trial7,8(see Supplementary Methods online and the Cancer
Genetic Markers Susceptibility website). The GWAS confirmed the
association for rs1447295, located at physical position 128554220 in
NCBI genome build 36 (P ¼ 9.75 ? 10–5; heterozygote OR: 1.42, 95%
c.i.: 1.16–1.73; homozygote OR: 2.78, 95% c.i.: 1.32–5.86; Table 1).
Received 16 January 2006; accepted 7 March 2007; published online 1 April 2007; doi:10.1038/ng2022
1SAIC-Frederick, National Cancer Institute (NCI)-Frederick Cancer Research and Development Center, Frederick, Maryland 21702, USA.2Division of Cancer
Epidemiology and Genetics and3Pediatric Oncology Branch, Center for Cancer Research, NCI, US National Institutes of Health (NIH), Department of Health and
Human Services (DHHS), Bethesda, Maryland 20892, USA.4Bioinformed Consulting Services, Gaithersburg, Maryland 20877, USA.5Program in Molecular and
Genetic Epidemiology, Department of Epidemiology, Harvard School of Public Health, Boston, Massachusetts 02115, USA.6Wellcome Trust Sanger Institute,
Wellcome Trust Genome Campus, Cambridge CB10 1SA, UK.7Department of Mathematics and Statistics, Lancaster University, Lancaster LA1 4YF, UK.8Department
of Epidemiology and Surveillance Research, American Cancer Society, Atlanta, Georgia 30329, USA.9Department of Health Promotion and Chronic Disease
Prevention, National Public Health Institute, Helsinki, FIN-00300, Finland.10Department of Nutrition, Harvard School of Public Health, Boston, Massachusetts
02115, USA.11Centre de Recherche pour les Pathologies Prostatiques (CeRePP), Ho ˆpital Tenon, Assistance Publique-Ho ˆpitaux de Paris, 75970 Paris, France.
12Division of Urologic Surgery, Washington University School of Medicine, St. Louis, Missouri 63108, USA.13Division of Hematology and Oncology, Columbia
University, New York, New York 10032, USA.14Office of Cancer Genomics, NCI, NIH, DHHS, Bethesda, Maryland 20892, USA. Correspondence should be addressed
to S.J.C. (firstname.lastname@example.org).
NATURE GENETICS VOLUME 39 [ NUMBER 5 [ MAY 2007645
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It also identified four SNPs (rs6983267, rs7837328, rs7014346 and
rs12334695) significantly associated with prostate cancer in a second
region of low correlation (Fig. 1). These SNPs reside in a block9with
strong linkage disequilibrium bounded by markers rs10505476
(128477298) and rs6470517 (128529586) (Fig. 1). We investigated
one of the most significant associations (rs6983267) in a combined
analysis with four additional replication studies totaling 3,124 affected
individuals and 3,142 controls (the American Cancer Society Cancer
Prevention Study II10, 1,150 affected individuals and 1,151 controls;
the Health Professionals Follow-up Study11, 625 affected individuals
and 636 controls; the CeRePP French Prostate Case-Control Study12,
455 affected individuals and 459 controls; and the Alpha-Tocopherol,
Beta-Carotene Cancer Prevention Study13, 896 affected individuals
and 894 controls) (Table 1, Supplementary Table 1 and Supplemen-
tary Table 2 online). Our results show that rs6983267, which has an
overall population frequency of 50% in northern Europeans for the
‘at-risk’ G allele, replicated in all four studies (P ¼ 1.63 ? 10–8), thus
providing strong evidence for its contribution to prostate cancer risk.
An additional SNP, rs7837688, from the same bin (r2¼ 0.81 with
rs1447295 in the PLCO) seemed to be more significant in the GWAS,
but in the replication studies, its significance and magnitude of effect
were comparable to rs1447295 (Table 1) overall. Because rs1447295
has been established as the benchmark, based on replication of the two
initial studies6, we conducted our subsequent analyses with rs1447295.
Markers rs1447295 and rs6983257 are physically close, but the two
association signals correspond to two independent loci (Fig. 1).
Analysis of PLCO controls between SNPs rs10505476 (128,477,298)
and rs7837688 (128,608,542) using the SequenceLDhot program14
gives strong evidence for a hotspot of recombination between
rs7841264 and rs1447293 (between 128535996 and 128541502)
(P ¼ 5 ? 10–5). This corresponds to an inferred location of a
recombination hotspot in the HapMap data (data release 21)15,16.
We estimate that 90% of the meiotic recombination events occurring
in the 130-kb region bound by rs10505476 and rs7837688 take place in
the 5.5-kb region between rs7841264 and rs1447293. Specifically, the
population scaled recombination rate14within this hotspot is esti-
mated to be 260 (95% c.i.: 100–540), whereas the recombination rate
across the remaining region is approximately 30. As a rough guide to
these estimates, an effective population size of 10,000 yields a genetic
distance of 0.65 cM within the hotspot, and of 0.075 cM for the
remainder of the region. The population-scaled recombination rate
governs the amount of linkage disequilibrium that would be expected
across the hotspot within our population. Such a large value suggests
that there will be almost no linkage disequilibrium across the hotspot.
To further explore this region of 8q24, we performed analyses using
inferred ancestral recombination graphs (ARGs)17(Fig. 2 and Sup-
plementary Note online). We inferred 100 ARGs for 197 SNPs in the
region and tested the genealogies at each SNP position for evidence of
association using a four–degree of freedom test (one control pheno-
type, two case phenotypes and three genotypes), with significance
calculated using a maximum of 105permutations. The analysis
identified two close, but distinct, regions of strong association that
straddle the recombination hotspot marked by rs7841264. Within
each region, pairwise comparison of the genealogies at positions of
strong association uncovered high correlation. The genealogies were
not correlated when the loci were located in different regions. Thus,
the ARG procedure detected in each region a single and specific
genetic event. The ARGs were used to estimate the frequency of the
inferred predisposing alleles in the two regions. In the centromeric
region at rs6983267, the frequency of the inferred predisposing allele
Table 1 Results of the single-SNP analysis of rs6983267, rs1447295 and rs7837688 (per study and combined)
Risk allele frequency,
controlsP OR (GT)95% c.i. OR (GG) 95% c.i.
2.43 ? 10-05
3.16 ? 10-03
1.89 ? 10-03
1.17 ? 10-01
9.54 ? 10-03
9.42 ? 10-13
9.75 ? 10-05
2.26 ? 10-05
2.88 ? 10-02
4.35 ? 10-03
2.74 ? 10-03
1.53 ? 10-14
6.52 ? 10-06
6.82 ? 10-06
2.85 ? 10-01
3.45 ? 10-03
1.28 ? 10-03
1.85 ? 10-14
Analysis adjusted for age in five-year intervals and study. See Supplementary Table 1 for genotype distribution information and Supplementary Table 2 for complete results of each
study using unconstrained and multiplicative models. ACS: American Cancer Society Prevention Study II; ATBC: Alpha-Tocopherol, Beta-Carotene Prevention Study; FPCC: CeRePP
French Prostate Case-Control Study; HPFS ¼ Health Professionals Follow-up Study; PLCO ¼ Prostate, Lung, Colon, Ovarian Trial.
646VOLUME 39 [ NUMBER 5 [ MAY 2007 NATURE GENETICS
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in controls was 0.46 ± 0.13. In the telomeric region, the frequency was
lower (0.12 ± 0.17) at rs1447295 and was between 0.10 and 0.11
(±0.08) for the other five locations (rs4242382, rs4242384, rs7017300,
rs11988857 and rs7837688). These differences in estimated allele
frequency corroborate the existence of two distinct functional poly-
morphisms on either side of the recombination hotspot.
In order to identify the haplotypes harboring the deleterious allele
of the centromeric region, we phased genotypes from 20 SNPs to
determine the most likely pair of haplotypes present in each indivi-
dual18,19(Fig. 3). We then used these haplotypes to generate 100
ARGs17. At each SNP location, the position of the mutation on
inferred genealogies that best explains the disease status partitions
the haplotypes into two groups: those predicted to harbor the
protective allele versus those predicted to harbor the ‘at-risk’ allele.
At rs6983267, six haplotypes (with frequencies greater than 0.1%)
defined by 11 contiguous SNPs were predicted to harbor the protec-
tive allele more than 80% of the time and all included the T allele of
rs6983267. In the 19 remaining haplotypes, the deleterious allele was
predicted in more than 95 of 100 genealogies. All harbored the ‘at risk’
G allele of rs6983267 (Fig. 3). The diversity of the two haplotype
groups suggests that the protective allele of the centromeric region is
more recent and/or was positively selected. A similar analysis per-
formed with 27 SNPs of the telomeric region identified two haplo-
types, defined by 24 contiguous SNPs, carrying the susceptibility
alleles, whereas 25 haplotypes carried the protective allele (Fig. 3).
Taken together, these observations suggest that the telomeric muta-
tional event might have occurred more recently than the centromeric
mutation. Furthermore, our data suggest that the telomeric mutation
generated a deleterious allele, in contrast to the centromeric mutation.
To determine possible interaction between the two independent
SNPs, we investigated seven logistic models for the joint effect of both
rs6983267 and rs1447295 on prostate cancer risk, comparing all cases
with controls adjusted for study and/or study center and age in 5-year
intervals (Table 2 and Supplementary Table 3 online). The associa-
tion between each SNP and prostate cancer risk remained significant
after adjusting for the other SNP (in the unconstrained model,
rs6983267 P ¼ 6.62 ? 10–10(adjusted for rs1447295 in row 25 of
Supplementary Table 2); rs1447295 P ¼ 1.41 ? 10–11(adjusted for
rs6983267 in row 11 of Supplementary Table 2)). No compelling
evidence differentiated the unconstrained model (which estimated
that odds ratios for the eight nonreferent genotypes varied freely)
from the simple multiplicative allelic risk model (likelihood ratio
test comparing nested models, P 4 0.7). The estimated OR under
the multiplicative model is 3.17 (95% c.i.: 2.55–3.94) for individuals
who are homozygous for the risk alleles at both loci relative to
the referent category: namely, double homozygotes for the protective
alleles (P ¼ 9.18 ? 10–22).
In a polytomous logistic regression analysis of rs6983267 and
rs1447295, assuming a multiplicative allelic risk model, the estimated
genotype effects were comparable for aggressive and nonaggressive
prostate cancer at diagnosis (Supplementary Table 3) (P ¼ 0.17). We
also investigated the possible effect of age on risk in an analysis of all
126.5 Mb127 Mb127.5 Mb 128 Mb128.5 Mb
128.5 Mb128.55 Mb 128.6 Mb
Figure 1 Association analysis of SNPs across a
region of 8q24. The upper panel shows P values
for association testing drawn from a genome-wide
association study of prostate cancer in the PLCO
cohort across a region of 8q24 bounded by
rs4559257 and rs7387606 (chromosome 8:
126501167–128998553). The analysis is based
on the genome-wide association study using
incidence density sampling with a score test
(4 d.f.) adjusted for population stratification
(Supplementary Methods). Note that rs6983267
is close to a putative pseudogene, POU5F1P1.
Shaded region ‘A’ corresponds to the admixture
peak reported in ref. 4 and spans chromosome 8,
Shaded region ‘B’ was analyzed by ancestral
recombination graph17(see Figs. 2 and 3).
Shaded region ‘C’ includes a segment containing
the most significant P values bounded by
rs1562871 and rs4407842. Lower panel shows
an enlarged view of the region bounded by
rs1562871 and rs4407842 (chromosome 8:
128470954–128619305). We estimated the
squared correlation coefficient (r2) for each
pairwise comparison of SNPs in this region using
a modified version of TagZilla (see Methods).
Negative log10r2was plotted using Aabel (see
Methods) (for example, 0 indicates evidence for
strong LD, whereas –8 corresponds to minimal
LD). Purple dots represent the locations of the
three loci evaluated in this study, rs6983267,
rs1447295 and rs7837688 (r2for the latter two
is 0.81 in the PLCO study); red dots represent
the other SNPs that showed evidence of
association in the GWAS (rs7837328,
rs4242382, rs4242384, rs7017300, and
rs11988857). Black arrow indicates the site
of recombination discussed in the text.
NATURE GENETICS VOLUME 39 [ NUMBER 5 [ MAY 2007 647
© 2007 Nature Publishing Group http://www.nature.com/naturegenetics
prostate cancers in order to test for heterogeneity of genetic effects at
ages above and below 65 years (Supplementary Table 3). The
estimated genetic effects under a two-locus multiplicative allelic risk
model did not differ significantly between age groups (P ¼ 0.18).
Although the region of 8q24 analyzed in this report is frequently
amplified in prostate tumors20,21, it harbors few known or predicted
genes. Furthermore, we did not observe an association between SNPs
in the MYC gene (263 kb from rs1447295 in the telomeric direction)
and prostate cancer risk.
Our results demonstrate how multiple SNPs within a chromosomal
region in distinct blocks may be associated with disease risk. Although
the rs6983267 G allele is associated with a lower relative risk than the
rs1447295 A allele, it is substantially more frequent in populations of
European ancestry (50% versus 11%). Based on our five studies, an
estimate for the population attributable risk22,23(PAR) of prostate
cancer associated with rs6983267 G is 21%, whereas the PAR for
rs1447295 A is 9%; the estimated joint effects PAR for carriage of
either or both is 27% (Supplementary Table 4 online). The estimated
joint and individual PARs suggest that the two loci substantially
contribute to the population burden of prostate cancer. However,
comparisons between PARs based on markers can be misleading when
the correlation between the markers and the respective functional
It is worth noting that subtle variations may occur within the
European population24. In the Alpha-Tocopherol, Beta-Carotene
study of Finns, the additional SNP in the telomeric block,
rs7837688, that was tested across all studies showed a higher MAF
and did not replicate, whereas the other studies showed a consistent,
positive association (Table 1). This apparent discrepancy could be
used to better define the region(s) harboring the functional variant(s)
in the telomeric block similar to a previous approach to mapping
PRKCA and multiple sclerosis25,26.
Figure 3 Centromeric and telomeric haplotypes
of the 8q24 region associated with prostate
cancer susceptibility in PLCO. The best pair of
haplotypes for each individual was determined
using PHASE18,19in two independent
calculations for 20 and 27 SNPs located in
either region flanking the recombination hotspot.
For each region, the phased haplotypes were
used to generate 100 genealogies17, and the
positions of the putative functional mutations
were estimated for the eight locations indicated
in Figure 1 and Figure 2. The frequency with
which each haplotype was predicted to carry the
‘at-risk’ mutation was then computed. Results are
shown for rs6983276 (haplotypes with
population frequencies larger than 0.001 defined
by 11 contiguous SNPs: rs10956365,
rs10505476, rs10808555, rs17467139,
rs6983267, rs10505473, rs7837328,
rs7014346, rs12375310, rs6995633,
rs6999921) and rs7837688 (haplotypes with population frequencies larger than 0.002 defined by 24 contiguous SNPs: rs7830412, rs1447293,
rs921146, rs4871799, rs1447295, rs9297758, rs13363309, rs11775749, rs16902169, rs13253127, rs6985504, rs16902173, rs12155672,
rs1562432, rs1562431, rs4871808, rs4242382, rs4242384, rs7017300, rs11988857, rs9656816, rs7814251, rs7837688, rs6991990). Results
from genealogies at position rs7837328 and rs1447295 were less contrasted. Those from positions rs4242382, rs4242384, rs7017300 and rs11988857
were identical to those of rs7837688, with the exception of the haplotype marked by an asterisk, predicted to carry an ‘at-risk’ mutation with a frequency
of 0.36 for genealogies located at rs11988857. Positions in almost perfect linkage disequilibrium (r2B1) with the imputed functional mutation are
highlighted in yellow. ‘Hap. freq.’ ¼ frequency of haplotype in all samples from PLCO; ‘prediction’ ¼ frequency predicted to carry the ‘at-risk’ mutation.
Figure 2 Association signal in the 8q24 region detected by ancestral
recombination graph (ARG). Unphased genotypes of rs1447295 and 196
flanking SNPs were used to infer 100 ARGs17, each one describing a
possible mutation and recombination history for this region jointly for the
controls, the nonaggressive cases and the aggressive cases of the PLCO
cohort. An ARG gives a genealogy for every SNP position, and these
genealogies were tested for association by placing putative causative
mutations on the branches and performing a w2test on a nine-cell
contingency table including three phenotypes (aggressive and nonaggressive
cases and controls) and three genotypes (4-d.f. w2test)17. After combining
this analysis across all genealogies at a position, we determined the
significance by random permutation of the phenotypes (maximum number
of permutations, 105). The log base10 of this evaluation, called
log10(permutation P value), is plotted as a function of the SNP position
along the chromosome. The eight locations that provided permutation
P values o 10–3are indicated by rs numbers at this position. The vertical
orange line indicates the position of the region with an estimated high
recombination rate. The horizontal green and red lines indicate the position of the centromeric and telomeric haplotypes, respectively, that were
reconstructed using the program PHASE and further studied for association using an ARG strategy (see main text and Supplementary Note). The positions
of three notable genes are also indicated (horizontal red arrows).
128.4 Mb128.6 Mb128.8 Mb 129.0 Mb
log10 (permutation P value)
Haplotype Hap. freq. Prediction
648VOLUME 39 [ NUMBER 5 [ MAY 2007 NATURE GENETICS
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Although it is possible that additional variants in the region of 8q24 Download full-text
could further modulate the risk of prostate cancer, the results of this
GWAS did not identify other highly significant loci in men of
European ancestry (Fig. 1). However, a previous admixture scan
identified peaks in the region that could be important in men of
other ancestral backgrounds4. We note that for the centromeric SNP,
rs6983267, the ‘at-risk’ G allele has an estimated frequency of 0.98 in
the Yoruban and 0.37 in the East Asian samples of HapMap compared
with 0.50 in the controls of our combined studies (Table 1)16. These
observations could partially explain the known ethnic disparities in
prostate cancer incidence2. Further work is needed to identify com-
mon and uncommon variants across both regions (particularly in
populations with different underlying genetic structures) to determine
the optimal candidates for functional studies designed to confirm the
causal variants in the 8q24 region.
Detailed descriptions of the methods are provided in Supplementary Methods.
URLs. Cancer Genetic Markers of Susceptibility project: http://cgems.cancer.
gov; HapMap: http://hapmap.org/; TagZilla: http://tagzilla.nci.nih.gov/; Aabel:
Note: Supplementary information is available on the Nature Genetics website.
The HPFS study is supported by NIH grants CA CA55075 and 5U01CA098233-
04. The ACS study is supported by U01 CA098710. The ATBC study is supported
by NIH contracts N01-CN-45165, N01-RC-45035 and N01-RC-37004. F.R.S. is
supported by an NRSA training grant (T32 CA 09001). P.F. is supported by a UK
Engineering and Physical Sciences Research Council Grant (GR/S18786). M.M. is
supported by the Wellcome Trust. N.O., R.B.H., S.W., K.Y., N.C., M.T., J.F.F.,
R.H., S.J.C. and G.T. are supported by the Intramural Research Program of the
National Cancer Institute (US National Institutes of Health, Department of
Health and Human Services).
COMPETING INTERESTS STATEMENT
The authors declare no competing financial interests.
Published online at http://www.nature.com/naturegenetics
Reprints and permissions information is available online at http://npg.nature.com/
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Table 2 Odds ratios and 95% confidence intervals for the two-SNP
unconstrained and multiplicative interaction models for all studies
Observed overall P ¼ 9.18 ? 10–22. Multiplicative overall P ¼ 1.69 ? 10–25. Analysis
adjusted for age in 5-year intervals and study. O ¼ observed (unconstrained); M ¼
multiplicative model. See Supplementary Table 3 for complete results of each study
using unconstrained and multiplicative models.
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