Petra J. de Verdier

Sequence variants at the TERT-CLPTM1L locus associate with many cancer types

The common sequence variants that have recently been associated with cancer risk are particular to a single cancer type or at most two. Following up on our genome-wide scan of basal cell carcinoma, we found that rs401681[C] on chromosome 5p15.33 satisfied our threshold for genome-wide significance (OR = 1.25, P = 3.7 × 10−12). We tested rs401681 for association with 16 additional cancer types in over 30,000 cancer cases and 45,000 controls and found association with lung cancer (OR = 1.15, P = 7.2 × 10−8) and urinary bladder, prostate and cervix cancer (ORs = 1.07−1.31, all P < 4 × 10−4). However, rs401681[C] seems to confer protection against cutaneous melanoma (OR = 0.88, P = 8.0 × 10−4). Notably, most of these cancer types have a strong environmental component to their risk. Investigation of the region led us to rs2736098[A], which showed stronger association with some cancer types. However, neither variant could fully account for the association of the other. rs2736098 corresponds to A305A in the telomerase reverse transcriptase (TERT) protein and rs401681 is in an intron of the CLPTM1L gene.

Sequence variant on 8q24 confers susceptibility to urinary bladder cancer

We conducted a genome-wide SNP association study on 1,803 urinary bladder cancer (UBC) cases and 34,336 controls from Iceland and The Netherlands and follow up studies in seven additional case-control groups (2,165 cases and 3,800 controls). The strongest association was observed with allele T of rs9642880 on chromosome 8q24, 30 kb upstream of MYC (allele-specific odds ratio (OR) = 1.22; P = 9.34 × 10−12). Approximately 20% of individuals of European ancestry are homozygous for rs9642880[T], and their estimated risk of developing UBC is 1.49 times that of noncarriers. No association was observed between UBC and the four 8q24 variants previously associated with prostate, colorectal and breast cancers, nor did rs9642880 associate with any of these three cancers. A weaker signal, but nonetheless of genome-wide significance, was captured by rs710521[A] located near TP63 on chromosome 3q28 (allele-specific OR = 1.19; P = 1. 15 × 10−7).

Genetic variation in the prostate stem cell antigen gene PSCA confers susceptibility to urinary bladder cancer

We conducted a genome-wide association study on 969 bladder cancer cases and 957 controls from Texas. For fast-track validation, we evaluated 60 SNPs in three additional US populations and validated the top SNP in nine European populations. A missense variant (rs2294008) in the PSCA gene showed consistent association with bladder cancer in US and European populations. Combining all subjects (6,667 cases, 39,590 controls), the overall P-value was 2.14 × 10−10 and the allelic odds ratio was 1.15 (95% confidence interval 1.10–1.20). rs2294008 alters the start codon and is predicted to cause truncation of nine amino acids from the N-terminal signal sequence of the primary PSCA translation product. In vitro reporter gene assay showed that the variant allele significantly reduced promoter activity. Resequencing of the PSCA genomic region showed that rs2294008 is the only common missense SNP in PSCA. Our data identify rs2294008 as a new bladder cancer susceptibility locus.

TERT promoter mutations in bladder cancer affect patient survival and disease recurrence through modification by a common polymorphism

Significance This study shows that the telomerase reverse transcriptase (TERT) promoter mutations, which create de novo E-twenty six/ternary complex factors (Ets/TCF) transcription binding sites, besides being the most common somatic genetic lesions, influence both survival and disease recurrence in bladder cancer patients. The effect of the TERT promoter mutations on both survival and recurrence is modified by a common polymorphism within the preexisting Ets binding site in the TERT promoter. The data were supported by the results from reporter assays carried out in two urothelial carcinoma cell lines. The findings of the study suggest that the TERT promoter mutations in conjunction with the common polymorphism have potential of being used as clinical biomarkers in bladder cancer. The telomerase reverse transcriptase (TERT) promoter, an important element of telomerase expression, has emerged as a target of cancer-specific mutations. Originally described in melanoma, the mutations in TERT promoter have been shown to be common in certain other tumor types that include glioblastoma, hepatocellular carcinoma, and bladder cancer. To fully define the occurrence and effect of the TERT promoter mutations, we investigated tumors from a well-characterized series of 327 patients with urothelial cell carcinoma of bladder. The somatic mutations, mainly at positions −124 and −146 bp from ATG start site that create binding motifs for E-twenty six/ternary complex factors (Ets/TCF), affected 65.4% of the tumors, with even distribution across different stages and grades. Our data showed that a common polymorphism rs2853669, within a preexisting Ets2 binding site in the TERT promoter, acts as a modifier of the effect of the mutations on survival and tumor recurrence. The patients with the mutations showed poor survival in the absence [hazard ratio (HR) 2.19, 95% confidence interval (CI) 1.02–4.70] but not in the presence (HR 0.42, 95% CI 0.18–1.01) of the variant allele of the polymorphism. The mutations in the absence of the variant allele were highly associated with the disease recurrence in patients with Tis, Ta, and T1 tumors (HR 1.85, 95% CI 1.11–3.08). The TERT promoter mutations are the most common somatic lesions in bladder cancer with clinical implications. The association of the mutations with patient survival and disease recurrence, subject to modification by a common polymorphism, can be a unique putative marker with individualized prognostic potential.

A sequence variant at 4p16.3 confers susceptibility to urinary bladder cancer

Previously, we reported germline DNA variants associated with risk of urinary bladder cancer (UBC) in Dutch and Icelandic subjects. Here we expanded the Icelandic sample set and tested the top 20 markers from the combined analysis in several European case-control sample sets, with a total of 4,739 cases and 45,549 controls. The T allele of rs798766 on 4p16.3 was found to associate with UBC (odds ratio = 1.24, P = 9.9 × 10−12). rs798766 is located in an intron of TACC3, 70 kb from FGFR3, which often harbors activating somatic mutations in low-grade, noninvasive UBC. Notably, rs798766[T] shows stronger association with low-grade and low-stage UBC than with more aggressive forms of the disease and is associated with higher risk of recurrence in low-grade stage Ta tumors. The frequency of rs798766[T] is higher in Ta tumors that carry an activating mutation in FGFR3 than in Ta tumors with wild-type FGFR3. Our results show a link between germline variants, somatic mutations of FGFR3 and risk of UBC.

Polymorphisms in XPD, XPC and the risk of death in patients with urinary bladder neoplasms

We conducted a follow-up study on 311 patients with urinary bladder neoplasms to investigate the association of polymorphisms in DNA repair and cell growth regulatory genes with the clinical outcomes of this disease. We found that patients carrying the variant allele of XPD (K751Q) polymorphism were at lower risk of death (p = 0.04) than the non-carriers. Patients that were simultaneous carriers of variant alleles from XPD (K751Q) and XPC (K939Q) polymorphisms, showed lower risk of death than the other patients (p = 0.001). The variant allele carriers of MSH6 (G39E) polymorphism showed a higher risk for highly malignant disease (TaG3+T1) than the non-carriers (p = 0.03). The variant allele carriers of XRCC1 (R399Q) polymorphism showed lower risk for recurrence (TaG2; p = 0.05) and death (T2+; p = 0.03) after instillation and radiotherapy than the non-carriers. After radiotherapy, an inverse association of the variant allele of OGG1 (S326C) polymorphism was observed with the risk of death (T2+; p = 0.04). A significant low-risk for stage progression (p = 0.03) was observed in patients carrying the variant allele of H-ras (H27H) polymorphism. Our results are consistent with the notion that the XPD (K751Q) polymorphism either individually or in combination with the XPC (K939Q) polymorphism modulates the risk of death in patients with urinary bladder neoplasms.

Genome-wide association study yields variants at 20p12.2 that associate with urinary bladder cancer.

Genome-wide association studies (GWAS) of urinary bladder cancer (UBC) have yielded common variants at 12 loci that associate with risk of the disease. We report here the results of a GWAS of UBC including 1670 UBC cases and 90 180 controls, followed by replication analysis in additional 5266 UBC cases and 10 456 controls. We tested a dataset containing 34.2 million variants, generated by imputation based on whole-genome sequencing of 2230 Icelanders. Several correlated variants at 20p12, represented by rs62185668, show genome-wide significant association with UBC after combining discovery and replication results (OR = 1.19, P = 1.5 × 10(-11) for rs62185668-A, minor allele frequency = 23.6%). The variants are located in a non-coding region approximately 300 kb upstream from the JAG1 gene, an important component of the Notch signaling pathways that may be oncogenic or tumor suppressive in several forms of cancer. Our results add to the growing number of UBC risk variants discovered through GWAS.

Mutations in TERT promoter and FGFR3 and telomere length in bladder cancer.

Mutations in the promoter of the telomerase reverse transcriptase (TERT) and fibroblast growth factor receptor 3 (FGFR3) genes constitute the most recurrent somatic alterations in urothelial carcinoma of bladder. In this study, we screened DNA from 327 urothelial bladder carcinomas from well‐documented patients, with different stages and grades and known TERT promoter mutational status, for FGFR3 alterations and measured relative telomere length (RTL). Although, the frequency of the TERT promoter mutations was higher than those in FGFR3; however, the alterations at the two loci occurred together more frequently than per chance [Odds ratio (OR) = 4.93, 95% CI = 2.72–8.92, p < 0.0001]. While tumors with TERT promoter and FGFR3 mutations had shorter RTL than those without mutations (p < 0.0001), the TERT promoter mutations in conjunction with the common allele of the rs2853669 polymorphism defined sub‐group of patients with an observed decreased overall survival (OR = 2.15, 95% CI = 1.00–4.61) and increased recurrence in patients with TaG1+TaG2 disease categories (OR = 3.68, 95%CI = 1.12–12.05). The finding of shorter telomeres in tumors with TERT promoter and/or FGFR3 mutations than without mutations implies mechanistic relevance of telomere biology in cancer progression. The observed association with recurrence and survival shows that the TERT promoter mutations can potentially be used as markers to refine selection of patients for different treatments. The overwhelming frequency of the TERT promoter mutations also represents a case for development of an eventual therapeutic target.

Polymorphisms in NQO1 and the clinical course of urinary bladder neoplasms.

Objective. Urinary bladder neoplasms differ considerably in biological potential, and tumor morphology alone cannot predict their clinical behaviors. Polymorphisms in xenobiotic metabolic genes reportedly modulate susceptibility to bladder neoplasms and may affect the clinical course and outcomes of the disease. This study was conducted to determine the effect of polymorphisms in the xenobiotic metabolic genes on the disease course and clinical outcomes of urinary bladder neoplasms. Material and methods. Patients with urinary bladder neoplasms who had been followed up for a 5-year period were genotyped for NQO1 (R139W, P187S), NAT (rapid/slow), GSTP1 (I105V), GSTT1 and GSTM1 (non-null/null) and MTHFR (A222V, E429A) polymorphisms. Results. Variant allele carriers of the NQO1 (P187S) polymorphism showed a higher risk for high-stage disease than non-carriers at diagnosis [relative risk (RR)=1.4; 95% CI 1.0–1.8). A higher risk for highly malignant disease (T2+) was also observed in variant allele carriers than non-carriers of the GSTP1 (I105V) polymorphism (RR=1.6; 95% CI 1.1–2.5). NQO1 (R139W) variant allele carrier patients with intermediate malignant disease (TaG3+T1) had shorter disease-free survival than non-carriers (p=0.05). In contrast, carriers of the variant allele for the MTHFR (A222V) polymorphism had significantly longer disease-free survival than non-carriers (p=0.02). Conclusions. Our data are consistent with the notion that NQO1 polymorphisms influence the course and clinical outcomes of urinary bladder neoplasms. However, our results need to be confirmed in a large study as most of the associations detected were only of marginal statistical significance, and would be lost on correction for multiple comparisons.

Polymorphisms in nitric-oxide synthase 3 may influence the risk of urinary-bladder cancer

Nitric oxide (NO) is an important biological messenger known to influence several types of human cancers. NO formation is catalyzed by three different nitric oxide synthase (NOS) enzymes. In this study we analyzed if the NOS3 promoter polymorphism -786T>C (rs2070744) and the NOS3 Glu298Asp polymorphism in exon 7 (rs1799983) influence risk and pathogenesis of urinary-bladder cancer. Allelic discrimination and DNA sequencing were used to determine the -786T>C and the Glu298Asp NOS3 genotypes in 359 urinary-bladder cancer patients, from a population-based patient material, and 164 population controls. Patient genotypes were combined with information on tumor stage, grade, stage and grade progression and cancer-specific death, using a 5-year clinical follow-up. A threefold increased odds ratio for bladder cancer was found in homozygous carriers of the C allele of the -786T>C promoter polymorphism (p=0.017). No increased bladder cancer risk was found for the Glu298Asp polymorphism, but there was an association between the Glu298Asp and tumor grade (p=0.040). Our results suggest that the NOS3 promoter polymorphism -786T>C may influence bladder cancer risk.