Angela Carta

GST, NAT, SULT1A1, CYP1B1 genetic polymorphisms, interactions with environmental exposures and bladder cancer risk in a high‐risk population

Tobacco smoking and occupation are major risk factors of bladder cancer via exposure to polycyclic aromatic hydrocarbons (PAHs) and aromatic amines. Glutathione S‐transferase (GST) M1, T1 and P1 are involved in the detoxification of PAH reactive metabolites. Two N‐acetyltransferase isozymes, NAT2 and NAT1, have major roles in catalyzing the N‐acetylation and O‐acetylation of aromatic amines. Cytochrome P450 1B1 (CYP1B1) and sulfotransferase 1A1 (SULT1A1) are also involved in the metabolism of PAHs and aromatic amines. It is hypothesized that the genetic polymorphisms of these metabolic enzymes have an effect on the individual susceptibility to bladder cancer in particular by interacting with relevant environmental exposures. A hospital‐based case‐control study among men in Brescia, Northern Italy recruited 201 incidence cases and 214 controls from 1997–2000. Occupational exposures were blindly coded by occupational physicians. Genotyping of polymorphisms were carried out with PCR‐RFLP method. Unconditional multivariate logistic regression was applied to model the association between genetic polymorphisms and bladder cancer risk. Effect modifications by age of onset, smoking and occupational exposures to PAHs and aromatic amines were evaluated. We also conducted an analysis of interaction between genetic factors. GSTM1 and GSTT1 null genotype were associated with an increased risk of bladder cancer with an odds ratio (OR) of 1.69 (95% confidence interval [CI] = 1.11–2.56) and 1.74 (95% CI = 1.02–2.95), respectively. The effect of GSTM1 null was seen particularly in heavy smokers, and there was a combined effect with occupational exposure of aromatic amines (OR = 2.77, 95% CI = 1.08–7.10). We observed a trend (p‐value < 0.01) of increasing cancer risk comparing subjects with normal GSTM1 and T1 activity to subjects with one (OR = 1.82, 95% CI = 1.16–2.85) or both null genotypes (OR = 2.58, 95% CI = 1.27–5.23). NAT2 slow acetylator was associated with marginally increased risk of bladder cancer (OR = 1.50, 95% CI = 0.99–2.27), and the OR for the joint effect with occupational exposure of aromatic amines was 3.26 (95% CI = 1.06–9.95). SULT1A1 Arg213His polymorphism showed a marginal protective effect. These findings suggest that individual susceptibility to bladder cancer may be modulated by GSTM1, GSTT1 and NAT2 polymorphisms.

Polymorphisms in DNA repair genes, smoking, and bladder cancer risk: findings from the International Consortium of Bladder Cancer

Tobacco smoking is the most important and well-established bladder cancer risk factor and a rich source of chemical carcinogens and reactive oxygen species that can induce damage to DNA in urothelial cells. Therefore, common variation in DNA repair genes might modify bladder cancer risk. In this study, we present results from meta-analyses and pooled analyses conducted as part of the International Consortium of Bladder Cancer. We included data on 10 single nucleotide polymorphisms corresponding to seven DNA repair genes from 13 studies. Pooled analyses and meta-analyses included 5,282 cases and 5,954 controls of non-Latino white origin. We found evidence for weak but consistent associations with ERCC2 D312N [rs1799793; per-allele odds ratio (OR), 1.10; 95% confidence interval (95% CI), 1.01-1.19; P = 0.021], NBN E185Q (rs1805794; per-allele OR, 1.09; 95% CI, 1.01-1.18; P = 0.028), and XPC A499V (rs2228000; per-allele OR, 1.10; 95% CI, 1.00-1.21; P = 0.044). The association with NBN E185Q was limited to ever smokers (interaction P = 0.002) and was strongest for the highest levels of smoking dose and smoking duration. Overall, our study provides the strongest evidence to date for a role of common variants in DNA repair genes in bladder carcinogenesis.

Genome-wide association study identifies multiple loci associated with bladder cancer risk

Candidate gene and genome-wide association studies (GWAS) have identified 11 independent susceptibility loci associated with bladder cancer risk. To discover additional risk variants, we conducted a new GWAS of 2422 bladder cancer cases and 5751 controls, followed by a meta-analysis with two independently published bladder cancer GWAS, resulting in a combined analysis of 6911 cases and 11 814 controls of European descent. TaqMan genotyping of 13 promising single nucleotide polymorphisms with P < 1 × 10(-5) was pursued in a follow-up set of 801 cases and 1307 controls. Two new loci achieved genome-wide statistical significance: rs10936599 on 3q26.2 (P = 4.53 × 10(-9)) and rs907611 on 11p15.5 (P = 4.11 × 10(-8)). Two notable loci were also identified that approached genome-wide statistical significance: rs6104690 on 20p12.2 (P = 7.13 × 10(-7)) and rs4510656 on 6p22.3 (P = 6.98 × 10(-7)); these require further studies for confirmation. In conclusion, our study has identified new susceptibility alleles for bladder cancer risk that require fine-mapping and laboratory investigation, which could further understanding into the biological underpinnings of bladder carcinogenesis.

Bladder cancer, GSTs, NAT1, NAT2, SULT1A1, XRCC1, XRCC3, XPD genetic polymorphisms and coffee consumption: a case–control study

The aim of the study was to investigate NAT1, NAT2, GSTM1, GSTT1, GSTP1, SULT1A1, XRCC1, XRCC3 and XPD genetic polymorphisms, coffee consumption and risk of bladder cancer (BC) through a hospital-based case–control study. The study population included 197 incident BC cases and 211 controls. The association between genetic polymorphisms, coffee drinking and BC risk was assessed by logistic regression taking into account age, education, tobacco smoking and occupational exposures to polycyclic aromatic hydrocarbons and aromatic amines. No association was found between the genetic polymorphisms investigated and BC risk according to coffee consumption apart of a significant increased BC risk among GSTP1 105-114 Val carriers heavy coffee drinkers (>3 cups/day) (OR 3.18, 95%CI 1.06–9.55). In conclusion our findings suggest a very limited role, if any, of genetic polymorphisms investigated in modulating the BC risk in coffee drinkers.

Primary liver cancer and occupation in men: A case‐control study in a high‐incidence area in northern Italy

The objective of our study was to evaluate the association between occupation and risk of liver cancer. A hospital‐based case‐control study was carried out during 1997–1999 in the Province of Brescia, a highly industrialized area in Northern Italy with a high incidence of this neoplasm. The cases were 144 male patients with incident liver cancer (96% hepatocellular carcinoma). Controls were 283 male patients, matched to cases on age (±5 years), period and hospital of admission. Information on lifetime occupational history and alcohol consumption was obtained via interview. Specific occupational exposures to pesticides, solvents and other suspected hepatocarcinogens were evaluated. A blood sample was collected to detect hepatitis B and C infections. Odds ratios (OR) of occupational exposure and 95% confidence intervals (CI), adjusted for age, residence, education, heavy alcohol intake, hepatitis B surface antigen and hepatitis C virus antibodies positivity were computed. A statistically significant increased OR was observed for employment in repair of motor vehicles (OR 3.7; 95% CI 1.1–12.3; 9 exposed cases, 10 exposed controls). Increased ORs, although not statistically significant, were found for field‐crop farm workers, food and beverage processors, blacksmiths and machine‐tool operators, electrical fitters, clerical workers, manufacture of industrial machinery and personal and household services. A slightly increased OR was noted in workers exposed to toluene and xylene (OR 1.4; 95% CI 0.7–3.0, 23 cases, 36 controls); the OR was 2.8 (95% CI 1.0–7.6, 11 cases, 12 controls) for 20 or more years of exposure and 2.0 (95% CI 0.9–4.1, 21 cases, 28 controls) for 30 or more years of time since first exposure. The increase in OR seemed to be independent from that of alcohol or viral infections. Our study showed that the role of occupational exposures in liver carcinogenesis is limited. However, prolonged exposure to organic solvents such as toluene and xylene may represent a risk factor for liver cancer.

Genome-wide interaction study of smoking and bladder cancer risk

Bladder cancer is a complex disease with known environmental and genetic risk factors. We performed a genome-wide interaction study (GWAS) of smoking and bladder cancer risk based on primary scan data from 3002 cases and 4411 controls from the National Cancer Institute Bladder Cancer GWAS. Alternative methods were used to evaluate both additive and multiplicative interactions between individual single nucleotide polymorphisms (SNPs) and smoking exposure. SNPs with interaction P values < 5 × 10(-) (5) were evaluated further in an independent dataset of 2422 bladder cancer cases and 5751 controls. We identified 10 SNPs that showed association in a consistent manner with the initial dataset and in the combined dataset, providing evidence of interaction with tobacco use. Further, two of these novel SNPs showed strong evidence of association with bladder cancer in tobacco use subgroups that approached genome-wide significance. Specifically, rs1711973 (FOXF2) on 6p25.3 was a susceptibility SNP for never smokers [combined odds ratio (OR) = 1.34, 95% confidence interval (CI) = 1.20-1.50, P value = 5.18 × 10(-) (7)]; and rs12216499 (RSPH3-TAGAP-EZR) on 6q25.3 was a susceptibility SNP for ever smokers (combined OR = 0.75, 95% CI = 0.67-0.84, P value = 6.35 × 10(-) (7)). In our analysis of smoking and bladder cancer, the tests for multiplicative interaction seemed to more commonly identify susceptibility loci with associations in never smokers, whereas the additive interaction analysis identified more loci with associations among smokers-including the known smoking and NAT2 acetylation interaction. Our findings provide additional evidence of gene-environment interactions for tobacco and bladder cancer.

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.

The 19q12 bladder cancer GWAS signal: association with cyclin E function and aggressive disease.

A genome-wide association study (GWAS) of bladder cancer identified a genetic marker rs8102137 within the 19q12 region as a novel susceptibility variant. This marker is located upstream of the CCNE1 gene, which encodes cyclin E, a cell-cycle protein. We performed genetic fine-mapping analysis of the CCNE1 region using data from two bladder cancer GWAS (5,942 cases and 10,857 controls). We found that the original GWAS marker rs8102137 represents a group of 47 linked SNPs (with r(2) ≥ 0.7) associated with increased bladder cancer risk. From this group, we selected a functional promoter variant rs7257330, which showed strong allele-specific binding of nuclear proteins in several cell lines. In both GWASs, rs7257330 was associated only with aggressive bladder cancer, with a combined per-allele OR = 1.18 [95% confidence interval (CI), 1.09-1.27, P = 4.67 × 10(-5)] versus OR = 1.01 (95% CI, 0.93-1.10, P = 0.79) for nonaggressive disease, with P = 0.0015 for case-only analysis. Cyclin E protein expression analyzed in 265 bladder tumors was increased in aggressive tumors (P = 0.013) and, independently, with each rs7257330-A risk allele (P(trend) = 0.024). Overexpression of recombinant cyclin E in cell lines caused significant acceleration of cell cycle. In conclusion, we defined the 19q12 signal as the first GWAS signal specific for aggressive bladder cancer. Molecular mechanisms of this genetic association may be related to cyclin E overexpression and alteration of cell cycle in carriers of CCNE1 risk variants. In combination with established bladder cancer risk factors and other somatic and germline genetic markers, the CCNE1 variants could be useful for inclusion into bladder cancer risk prediction models.

Evidence of cross-reactive immunity to 2009 pandemic influenza A virus in workers seropositive to swine H1N1 influenza viruses circulating in Italy.

Background Pigs play a key epidemiologic role in the ecology of influenza A viruses (IAVs) emerging from animal hosts and transmitted to humans. Between 2008 and 2010, we investigated the health risk of occupational exposure to swine influenza viruses (SIVs) in Italy, during the emergence and spread of the 2009 H1N1 pandemic (H1N1pdm) virus. Methodology/Principal Findings Serum samples from 123 swine workers (SWs) and 379 control subjects (Cs), not exposed to pig herds, were tested by haemagglutination inhibition (HI) assay against selected SIVs belonging to H1N1 (swH1N1), H1N2 (swH1N2) and H3N2 (swH3N2) subtypes circulating in the study area. Potential cross-reactivity between swine and human IAVs was evaluated by testing sera against recent, pandemic and seasonal, human influenza viruses (H1N1 and H3N2 antigenic subtypes). Samples tested against swH1N1 and H1N1pdm viruses were categorized into sera collected before (n. 84 SWs; n. 234 Cs) and after (n. 39 SWs; n. 145 Cs) the pandemic peak. HI-antibody titers ≥10 were considered positive. In both pre-pandemic and post-pandemic peak subperiods, SWs showed significantly higher swH1N1 seroprevalences when compared with Cs (52.4% vs. 4.7% and 59% vs. 9.7%, respectively). Comparable HI results were obtained against H1N1pdm antigen (58.3% vs. 7.7% and 59% vs. 31.7%, respectively). No differences were found between HI seroreactivity detected in SWs and Cs against swH1N2 (33.3% vs. 40.4%) and swH3N2 (51.2 vs. 55.4%) viruses. These findings indicate the occurrence of swH1N1 transmission from pigs to Italian SWs. Conclusion/Significance A significant increase of H1N1pdm seroprevalences occurred in the post-pandemic peak subperiod in the Cs (p<0.001) whereas SWs showed no differences between the two subperiods, suggesting a possible occurrence of cross-protective immunity related to previous swH1N1 infections. These data underline the importance of risk assessment and occupational health surveillance activities aimed at early detection and control of SIVs with pandemic potential in humans.

Complex Relationships between Occupation, Environment, DNA Adducts, Genetic Polymorphisms and Bladder Cancer in a Case-Control Study Using a Structural Equation Modeling

DNA adducts are considered an integrate measure of carcinogen exposure and the initial step of carcinogenesis. Their levels in more accessible peripheral blood lymphocytes (PBLs) mirror that in the bladder tissue. In this study we explore whether the formation of PBL DNA adducts may be associated with bladder cancer (BC) risk, and how this relationship is modulated by genetic polymorphisms, environmental and occupational risk factors for BC. These complex interrelationships, including direct and indirect effects of each variable, were appraised using the structural equation modeling (SEM) analysis. Within the framework of a hospital-based case/control study, study population included 199 BC cases and 213 non-cancer controls, all Caucasian males. Data were collected on lifetime smoking, coffee drinking, dietary habits and lifetime occupation, with particular reference to exposure to aromatic amines (AAs) and polycyclic aromatic hydrocarbons (PAHs). No indirect paths were found, disproving hypothesis on association between PBL DNA adducts and BC risk. DNA adducts were instead positively associated with occupational cumulative exposure to AAs (p = 0.028), whereas XRCC1 Arg 399 (p<0.006) was related with a decreased adduct levels, but with no impact on BC risk. Previous findings on increased BC risk by packyears (p<0.001), coffee (p<0.001), cumulative AAs exposure (p = 0.041) and MnSOD (p = 0.009) and a decreased risk by MPO (p<0.008) were also confirmed by SEM analysis. Our results for the first time make evident an association between occupational cumulative exposure to AAs with DNA adducts and BC risk, strengthening the central role of AAs in bladder carcinogenesis. However the lack of an association between PBL DNA adducts and BC risk advises that these snapshot measurements are not representative of relevant exposures. This would envisage new scenarios for biomarker discovery and new challenges such as repeated measurements at different critical life stages.