Christiane Maier

Multiple Novel Prostate Cancer Predisposition Loci Confirmed by an International Study: The PRACTICAL Consortium

A recent genome-wide association study found that genetic variants on chromosomes 3, 6, 7, 10, 11, 19 and X were associated with prostate cancer risk. We evaluated the most significant single-nucleotide polymorphisms (SNP) in these loci using a worldwide consortium of 13 groups (PRACTICAL). Blood DNA from 7,370 prostate cancer cases and 5,742 male controls was analyzed by genotyping assays. Odds ratios (OR) associated with each genotype were estimated using unconditional logistic regression. Six of the seven SNPs showed clear evidence of association with prostate cancer (P = 0.0007-P = 10−17). For each of these six SNPs, the estimated per-allele OR was similar to those previously reported and ranged from 1.12 to 1.29. One SNP on 3p12 (rs2660753) showed a weaker association than previously reported [per-allele OR, 1.08 (95% confidence interval, 1.00-1.16; P = 0.06) versus 1.18 (95% confidence interval, 1.06-1.31)]. The combined risks associated with each pair of SNPs were consistent with a multiplicative risk model. Under this model, and in combination with previously reported SNPs on 8q and 17q, these loci explain 16% of the familial risk of the disease, and men in the top 10% of the risk distribution have a 2.1-fold increased risk relative to general population rates. This study provides strong confirmation of these susceptibility loci in multiple populations and shows that they make an important contribution to prostate cancer risk prediction. (Cancer Epidemiol Biomarkers Prev 2008;17(8):2052–61)

REVEL: An Ensemble Method for Predicting the Pathogenicity of Rare Missense Variants

The vast majority of coding variants are rare, and assessment of the contribution of rare variants to complex traits is hampered by low statistical power and limited functional data. Improved methods for predicting the pathogenicity of rare coding variants are needed to facilitate the discovery of disease variants from exome sequencing studies. We developed REVEL (rare exome variant ensemble learner), an ensemble method for predicting the pathogenicity of missense variants on the basis of individual tools: MutPred, FATHMM, VEST, PolyPhen, SIFT, PROVEAN, MutationAssessor, MutationTaster, LRT, GERP, SiPhy, phyloP, and phastCons. REVEL was trained with recently discovered pathogenic and rare neutral missense variants, excluding those previously used to train its constituent tools. When applied to two independent test sets, REVEL had the best overall performance (p < 10-12) as compared to any individual tool and seven ensemble methods: MetaSVM, MetaLR, KGGSeq, Condel, CADD, DANN, and Eigen. Importantly, REVEL also had the best performance for distinguishing pathogenic from rare neutral variants with allele frequencies <0.5%. The area under the receiver operating characteristic curve (AUC) for REVEL was 0.046-0.182 higher in an independent test set of 935 recent SwissVar disease variants and 123,935 putatively neutral exome sequencing variants and 0.027-0.143 higher in an independent test set of 1,953 pathogenic and 2,406 benign variants recently reported in ClinVar than the AUCs for other ensemble methods. We provide pre-computed REVEL scores for all possible human missense variants to facilitate the identification of pathogenic variants in the sea of rare variants discovered as sequencing studies expand in scale.

Mutation screening and association study of RNASEL as a prostate cancer susceptibility gene.

To date, germline mutations have been found in three candidate genes for hereditary prostate cancer: ELAC2 at 17p11, RNASEL at 1q25 and MSR1 at 8p22. RNASEL, encoding the 2′,5′-oligoadenylate-dependant RNase L, seems to have rare mutations in different ethnicities, such as M1I in Afro-Americans, E265X in men of European descent and 471delAAAG in Ashkenazi Jews. In order to evaluate the relevance of RNASEL in the German population, we sequenced its open reading frame to determine the spectrum and frequency of germline mutations. The screen included 303 affected men from 136 Caucasian families, of which 45 met the criteria for hereditary prostate cancer. Variants were analysed using a family-based association test, and genotyped in an additional 227 sporadic prostate cancer patients and 207 controls. We identified only two sib pairs (1.4% of our families) cosegregating conspicuous RNASEL variants with prostate cancer: the nonsense mutation E265X, and a new amino-acid substitution (R400P) of unknown functional relevance. Both alleles were also found at low frequencies (1.4 and 0.5%, respectively) in controls. No significant association of polymorphisms (I97L, R462Q and D541E) was observed, neither in case–control analyses nor by family-based association tests. In contrast to previous reports, our study does not suggest that common variants (i.e. R462Q) modify disease risk. Our results are not consistent with a high penetrance of deleterious RNASEL mutations. Due to the low frequency of germline mutations present in our sample, RNASEL does not have a significant impact on prostate cancer susceptibility in the German population.

Detecting predictive androgen receptor modifications in circulating prostate cancer cells

Molecular modifications of the androgen receptor (AR) can cause resistance to androgen deprivation therapy (ADT) in prostate cancer patients. Since lack of representative tumor samples hinders therapy adjustments according to emerging AR-modifications, we evaluated simultaneous detection of the two most common AR modifications (AR-V7 splice variant and AR point mutations) in circulating tumor cells (CTCs). We devised a single-tube assay to detect AR-V7 splice variants and AR point mutations in CTCs using immunomagnetic cell isolation, followed by quantitative real-time PCR and DNA pyrosequencing. We prospectively investigated 47 patients with PSA progression awaiting therapy switch. Comparison of response to newly administered therapy and CTC-AR-status allowed effect size estimation. Nineteen (51%) of 37 patients with detectable CTCs carried AR-modifications. Seventeen patients carried the AR-V7 splice variant, one harbored a p.T878A point mutation and one harbored both AR-V7 and a p.H875Y mutation. We estimated a positive predictive value for response and non-response to therapy by AR status in CTCs of ~94%. Based on a conservative calculation, we estimated the effect size for molecularly-informed therapy switches for prospective clinical trial planning to ~27%. In summary, the ability to determine key resistance-mediating AR modifications in CTCs has the potential to considerably improve prostate cancer treatment.

Fine-mapping identifies multiple prostate cancer risk loci at 5p15, one of which associates with TERT expression

Associations between single nucleotide polymorphisms (SNPs) at 5p15 and multiple cancer types have been reported. We have previously shown evidence for a strong association between prostate cancer (PrCa) risk and rs2242652 at 5p15, intronic in the telomerase reverse transcriptase (TERT) gene that encodes TERT. To comprehensively evaluate the association between genetic variation across this region and PrCa, we performed a fine-mapping analysis by genotyping 134 SNPs using a custom Illumina iSelect array or Sequenom MassArray iPlex, followed by imputation of 1094 SNPs in 22 301 PrCa cases and 22 320 controls in The PRACTICAL consortium. Multiple stepwise logistic regression analysis identified four signals in the promoter or intronic regions of TERT that independently associated with PrCa risk. Gene expression analysis of normal prostate tissue showed evidence that SNPs within one of these regions also associated with TERT expression, providing a potential mechanism for predisposition to disease.

POU5F1P1, a putative cancer susceptibility gene, is overexpressed in prostatic carcinoma.

Association between genetic variants located on human chromosome 8q24.21 with an increased risk for prostatic carcinoma has been well established. POU5F1P1, a processed pseudogene homologous to the pluripotency factor OCT4, is the only sequence with coding capacity in this region. The objective of this study was to investigate the POU5F1P1 expression in prostatic carcinoma and carcinoma surrounding prostatic tissue.

Role of the Nijmegen Breakage Syndrome 1 Gene in Familial and Sporadic Prostate Cancer

The Nijmegen breakage syndrome 1 (NBS1) gene, which participates in DNA double strand break repair, has been postulated to be a susceptibility factor for a number of cancers, including prostate cancer. Numerous mutations have been identified in NBS1, including the founder mutation 657del5. In this study, a number of analyses were done to determine whether mutations in NBS1 are associated with an increased risk for prostate cancer. The frequency of the 657del5 mutation in both familial prostate cancer cases (1,819 affected men among 909 families) and sporadic prostate cancer cases (1,218 affected men) collected from five centers participating in the International Consortium for Prostate Cancer Genetics were compared with that found in 697 normal controls. Seven individuals were identified to carry the mutation among the 3,037 cases screened: four in the familial group (three from one family and one from another) and three in the sporadic cases. The carrier frequency was 0.22% (2 of 909) for the probands and 0.25% (3 of 1,218) for the sporadic cases of prostate cancer. The 657del5 mutation was not detected in either the 293 unaffected members of the prostate cancer families or in the 697 control samples tested. The entire NBS1 gene was also sequenced in 20 of the youngest affected individuals from the Finnish group of familial cases to identify the presence of possible mutations in this high-risk group. One rare (D95N) and one common (E185Q) missense alteration was identified. More detailed analyses of the E185Q polymorphism, along with a third rare variant (R215W), failed to show an association with prostate cancer. Because the 657del5 mutation was absent from the control population, we are unable to determine if this alteration predisposes to prostate cancer. However, our data does suggest that mutations within NBS1, and in particular, 657del5, do not significantly contribute to the overall prostate cancer burden within our patient samples. (Cancer Epidemiol Biomarkers Prev 2006;15(5):935–8)

Multiple novel prostate cancer susceptibility signals identified by fine-mapping of known risk loci among Europeans

Genome-wide association studies (GWAS) have identified numerous common prostate cancer (PrCa) susceptibility loci. We have fine-mapped 64 GWAS regions known at the conclusion of the iCOGS study using large-scale genotyping and imputation in 25 723 PrCa cases and 26 274 controls of European ancestry. We detected evidence for multiple independent signals at 16 regions, 12 of which contained additional newly identified significant associations. A single signal comprising a spectrum of correlated variation was observed at 39 regions; 35 of which are now described by a novel more significantly associated lead SNP, while the originally reported variant remained as the lead SNP only in 4 regions. We also confirmed two association signals in Europeans that had been previously reported only in East-Asian GWAS. Based on statistical evidence and linkage disequilibrium (LD) structure, we have curated and narrowed down the list of the most likely candidate causal variants for each region. Functional annotation using data from ENCODE filtered for PrCa cell lines and eQTL analysis demonstrated significant enrichment for overlap with bio-features within this set. By incorporating the novel risk variants identified here alongside the refined data for existing association signals, we estimate that these loci now explain ∼38.9% of the familial relative risk of PrCa, an 8.9% improvement over the previously reported GWAS tag SNPs. This suggests that a significant fraction of the heritability of PrCa may have been hidden during the discovery phase of GWAS, in particular due to the presence of multiple independent signals within the same region.

Baseline Brain Perfusion and the Serotonin Transporter Promoter Polymorphism

BACKGROUND The serotonin transporter length repeat polymorphism (5-HTTLPR) in the promoter region of the serotonin transporter gene (SLC6A4) has been associated in healthy subjects with changes in basal perfusion levels in the limbic system and ventral prefrontal areas, regions involved in the pathophysiology of depression and anxiety, suggesting the existence of a neurobiological trait predisposing to these disorders. We reassess the findings of an increased baseline perfusion in the amygdala and ventral prefrontal areas in healthy carriers of the risk genotype in a much larger sample than in previous studies. METHODS A cohort of 183 healthy European individuals underwent perfusion imaging with continuous arterial spin-labeling (CASL) while resting quietly in the scanner for 8 minutes. Participants were genotyped to assess the occurrence of the short allele and the Lg and La variants of the long repeat. RESULTS No association between the 5-HTTLPR polymorphism and baseline brain perfusion was detected in the regions of interest or elsewhere in the brain. In the amygdala, variability in baseline perfusion was explained in large part by global cerebral flow levels (between 50% and 55%), in minor part by sex (between 4% and 5%), but not by genotype (less than .5%). Power analyses showed that the study was of sufficient size to be informative. CONCLUSIONS The findings did not confirm the existence of a biological marker of the effect of 5-HTTLPR polymorphism in the amygdala or in the orbitofrontal cortex.

A genomewide linkage analysis for prostate cancer susceptibility genes in families from Germany

Prostate cancer is a complex disease with a substantial genetic contribution involved in the disease risk. Several genomewide linkage studies conducted so far have demonstrated a strong heterogeneity of susceptibility. In order to assess candidate regions that are particularly relevant for the German population, we performed a genomewide linkage search on 139 prostate cancer families. A nonparametric method (Zlr scores), using GENEHUNTERPLUS, was applied at 500 markers (panel P1400, deCODE), with an average spacing of 7.25 cM. In the entire family collection, linkage was most evident at 8p22 (Zlr=2.47, P=0.0068), close to the previously identified susceptibility gene MSR1. Further local maxima with Zlr>2 (P<0.025) were observed at 1q, 5q and 15q. In a subgroup of 47 families, which matched the Johns Hopkins criteria of hereditary prostate cancer, suggestive linkage was found on 1p31 (Zlr=3.37, P=0.00038), a previously not described candidate region. The remaining 92 pedigrees, with no strong disease history, revealed a maximum Zlr=3.15 (P=0.00082) at 8q13, possibly indicating a gene with reduced penetrance or recessive inheritance. Our results suggest pronounced locus heterogeneity of prostate cancer susceptibility in Germany. In the present study population, the MSR1 gene could play a significant role. Other conspicuous loci, like 1p31 and 8q13, need further investigation in order to verify their relevance and to identify candidate genes.