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A two-stage genome-wide association study (GWAS) of the Cancer Genetic Markers of Susceptibility (CGEMS) initiative identified single nucleotide polymorphisms (SNP) in 150 regions across the genome that may be associated with prostate cancer (PCa) risk. We filtered these results to identify 43 independent SNPs where the frequency of the risk allele was consistently higher in cases than in controls in each of the five CGEMS study populations. Genotype information for 22 of these 43 SNPs was obtained either directly by genotyping or indirectly by imputation in our PCa GWAS of 500 cases and 500 controls selected from a population-based case-control study in Sweden [Cancer of the Prostate in Sweden (CAPS)]. Two of these 22 SNPs were significantly associated with PCa risk (P<0.05). We then genotyped these two SNPs in the remaining cases (n=2,393) and controls (n=1,222) from CAPS and found that rs887391 at 19q13 was highly associated with PCa risk (P=9.4 x 10(-4)). A similar trend of association was found for this SNP in a case-control study from Johns Hopkins Hospital (JHH), albeit the result was not statistically significant. Altogether, the frequency of the risk allele of rs887391 was consistently higher in cases than controls among each of seven study populations examined, with an overall P=3.2 x 10(-7) from a combined allelic test. A fine-mapping study in a 110-kb region at 19q13 among CAPS and JHH study populations revealed that rs887391 was the most strongly associated SNP in the region. Additional confirmation studies of this region are warranted.

To search for genetic variants that are associated with prostate cancer risk in the genome, we combined the data from our genome-wide association study (GWAS) in a population-based case-control study in Sweden with publicly available GWAS data from the Cancer Genetic Markers of Susceptibility (CGEMS) study. We limited the cases to those with aggressive disease in an attempt to identify risk variants that are associated with this most clinically relevant form of the disease. Among the most likely candidate single nucleotide polymorphisms (SNP) identified from the two GWAS, we sequentially confirmed one SNP at 22q13 in two independent study populations: the remaining subjects in Cancer of the Prostate in Sweden and a hospital-based case-control study at Johns Hopkins Hospital. Association of aggressive prostate cancer with the SNP at 22q13 was also observed in the publicly available data of four additional study populations from the second stage of the CGEMS study. In all seven study populations examined, the frequency of allele "C" of rs9623117 at 22q13 was consistently higher in aggressive cases than in controls. The combined allelic test was highly significant, with P = 5.0 x 10(-7). The odds ratio (OR) of allele C for aggressive prostate cancer was estimated to be 1.18 [95% confidence interval (95% CI), 1.11-1.26]. However, the SNP was also associated with nonaggressive prostate cancer, with an estimated OR of 1.11 (95% CI, 1.04-1.19; P = 0.004). The risk-associated variants are located within the genomic region of TNRC6B, a gene involved in miRNA-mediated mRNA degradation. Additional studies are warranted to further confirm the association.

NKX3.1, a gene mapped to 8p21, is a member of the NK class of homeodomain proteins and is expressed primarily in the prostate. NKX3.1 exerts a growth-suppressive and differentiating effect on prostate epithelial cells. Because of its known functions and its location within a chromosomal region where evidence for prostate cancer linkage and somatic loss of heterozygosity is found, we hypothesize that sequence variants in the NKX3.1 gene increase prostate cancer risk. To address this, we first resequenced the NKX3.1 gene in 159 probands of hereditary prostate cancer families recruited at Johns Hopkins Hospital; each family has at least three first-degree relatives affected with prostate cancer. Twenty-one germ-line variants were identified in this analysis, including one previously described common nonsynonymous change (R52C), two novel rare nonsynonymous changes (A17T and T164A), and a novel common 18-bp deletion in the promoter. Overall, the germ-line variants were significantly linked to prostate cancer, with a peak heterogeneity logarithm of odds of 2.04 (P = 0.002) at the NKX3.1 gene. The rare nonsynonymous change, T164A, located in the homeobox domain of the gene, segregated with prostate cancer in a family with three affected brothers and one unaffected brother. Importantly, nuclear magnetic resonance solution structure analysis and circular dichroism studies showed this specific mutation to affect the stability of the homeodomain of the NKX3.1 protein and decreased binding to its cognate DNA recognition sequence. These results suggest that germ-line sequence variants in NKX3.1 may play a role in susceptibility to hereditary prostate cancer and underscore a role for NKX3.1 as a prostate cancer gatekeeper.

Prostate cancer is a leading and increasingly prevalent cause of cancer death in men. Whereas family history of disease is one of the strongest prostate cancer risk factors and suggests a hereditary component, the predisposing genetic factors remain unknown. We first showed that KLF6 is a tumor suppressor somatically inactivated in prostate cancer and since then, its functional loss has been further established in prostate cancer cell lines and other human cancers. Wild-type KLF6, but not patient-derived mutants, suppresses cell growth through p53-independent transactivation of p21. Here we show that a germline KLF6 single nucleotide polymorphism, confirmed in a tri-institutional study of 3,411 men, is significantly associated with an increased relative risk of prostate cancer in men, regardless of family history of disease. This prostate cancer-associated allele generates a novel functional SRp40 DNA binding site and increases transcription of three alternatively spliced KLF6 isoforms. The KLF6 variant proteins KLF6-SV1 and KLF6-SV2 are mislocalized to the cytoplasm, antagonize wtKLF6 function, leading to decreased p21 expression and increased cell growth, and are up-regulated in tumor versus normal prostatic tissue. Thus, these results are the first to identify a novel mechanism of self-encoded tumor suppressor gene inactivation and link a relatively common single nucleotide polymorphism to both regulation of alternative splicing and an increased risk in a major human cancer.

The loss of cell cycle control is believed to be an important mechanism in the promotion of carcinogenesis. CDKN1B (p27) belongs to the Cip/Kip family and functions as an important cell cycle gatekeeper. Several lines of evidence from clinical studies and laboratory experiments demonstrate that CDKN1B is an important tumor suppressor gene in prostate cancer etiology. In addition, a case-control study has shown that the 326T/G (V109G) polymorphism in CDKN1B is associated with advanced prostate cancer. In light of the evidence for linkage between the chromosomal location of the CDKN1B gene (12p13) and prostate cancer susceptibility in several hereditary prostate cancer (HPC) populations, we hypothesized that sequence variants of CDKN1B play a role in HPC. To test this hypothesis, we first resequenced this gene in 96 HPC probands to identify germ-line mutations and sequence variants. We then genotyped the identified sequence variants among all family members of 188 HPC families and tested for their cosegregation with prostate cancer. In total, 10 sequence variants were identified, including three nonsynonymous changes. A family-based test, which is free from the effects of population stratification, revealed a significant association between single nucleotide polymorphism (SNP) -79C/T and prostate cancer (with a nominal P of 0.0005). The C allele of -79C/T was overtransmitted from parents to their affected offspring. Evidence for this association was primarily contributed by affected offspring whose age at diagnosis was <65 years. Together with the previous association study in a sporadic prostate cancer population, our new findings additionally suggest that germ-line variants of this gene play a role in prostate cancer susceptibility.

The enzyme alpha-methylacyl-CoA racemase (AMACR) plays an important role in peroxisomal beta-oxidation of branched-chain fatty acid and therefore is relevant to carcinogenesis. The involvement of AMACR in prostate cancer (CaP) is implicated by the recent observation that expression of AMACR is consistently and extensively up-regulated in CaP. This observation is of particular interest, given previous findings from epidemiological studies that red meat and dairy products, major sources of branched-chain fatty acid, are associated with CaP risk and from linkage studies that the AMACR gene region at 5p13 is linked to a CaP susceptibility gene. In this study, we hypothesize that sequence variants in AMACR may alter the risk for CaP. To test this hypothesis, we sequenced all five exons, exon-intron junctions, the promoter region, and 3'-untranslated region of AMACR in germ-line DNA samples of 96 probands from hereditary CaP (HPC) families. Seventeen sequence variants, including five novel (R118Q, V185A, P238S, Q239H, and L250R) and five known (M9V, S52P, D175G, S201L, and K277E) missense changes, were identified. Six of these variants are at conserved residues among the rat and mouse AMACR. Eleven of these single nucleotide polymorphisms were genotyped in a total of 159 HPC probands, 245 sporadic cases, and 211 unaffected controls to assess their association with CaP risk. Significantly different genotype frequencies between HPC probands and unaffected controls were found for several missense changes, including M9V (P = 0.03), G1175D (P = 0.02), S291L (P = 0.02), and K277E (P = 0.02). Haplotype analysis provided stronger evidence for association (P = 0.001). Furthermore, the AMACR sequence variants strongly cosegregate with CaP in HPC families (log of odds = 3.78; P = 0.00006), especially in the subset of families whose probands carry the "A-A" haplotype of M9V and D175G (log of odds = 4.34; P = 0.000008). These results suggest that sequence variants in AMACR may be associated with CaP risk.

8-Hydroxyguanine is a mutagenic base lesion produced by reactive oxygen species. The hOGG1 gene encodes a DNA glycosylase/AP lyase that can suppress the mutagenic effects of 8-hydroxyguanine by catalyzing its removal from oxidized DNA. A population-based (245 cases and 222 controls) and family-based (159 hereditary prostate cancer families) association study was performed to test the hypothesis that sequence variants of hOGG1 increase susceptibility to prostate cancer. We found that the genotype frequency of two sequence variants (11657A/G and Ser326Cys) was significantly different between cases and controls. The association with 11657A/G is confirmed and strengthened by our family-based association study. These results suggest that sequence variants in this gene are associated with prostate cancer risk, presumably through defective DNA repair function of hOGG1.

3beta-hydroxysteroid dehydrogenases (HSD3Bs), encoded by the HSD3B gene family at 1p13, have long been hypothesized to have a major role in prostate cancer susceptibility. The recent reports of a prostate cancer linkage at 1p13 provided additional evidence that HSD3B genes may be prostate cancer susceptibility genes. To evaluate the possible role of HSD3B genes in prostate cancer, we screened a panel of DNA samples collected from 96 men with or without prostate cancer for sequence variants in the putative promoter region, exons, exon-intron junctions, and 3'-untranslated region of HSD3B1 and HSD3B2 genes by direct sequencing. Eleven single nucleotide polymorphisms (SNPs) were identified, four of which, including a missense change (B1-N367T), were informative. These four SNPs were further genotyped in a total of 159 hereditary prostate cancer probands, 245 sporadic prostate cancer cases, and 222 unaffected controls. Although a weak association between prostate cancer risk and a missense SNP (B1-N367T) was found, stronger evidence for association was found when the joint effect of the two genes was considered. Men with the variant genotypes at either B1-N367T or B2-c7519g had a significantly higher risk to develop prostate cancer, especially the hereditary type of prostate cancer. Most importantly, the subset of hereditary prostate cancer probands, whose families provided evidence for linkage at 1p13, predominantly contributed to the observed association. Additional studies are warranted to confirm these findings.