Protein-altering germline mutations implicate novel genes related to lung cancer development.

Few germline mutations are known to affect lung cancer risk. We performed analyses of rare variants from 39,146 individuals of European ancestry and investigated gene expression levels in 7,773 samples. We find a large-effect association with an ATM L2307F (rs56009889) mutation in adenocarcinoma for discovery (adjusted Odds Ratio = 8.82, P = 1.18 × 10-15) and replication (adjusted OR = 2.93, P = 2.22 × 10-3) that is more pronounced in females (adjusted OR = 6.81 and 3.19 and for discovery and replication). We observe an excess loss of heterozygosity in lung tumors among ATM L2307F allele carriers. L2307F is more frequent (4%) among Ashkenazi Jewish populations. We also observe an association in discovery (adjusted OR = 2.61, P = 7.98 × 10-22) and replication datasets (adjusted OR = 1.55, P = 0.06) with a loss-of-function mutation, Q4X (rs150665432) of an uncharacterized gene, KIAA0930. Our findings implicate germline genetic variants in ATM with lung cancer susceptibility and suggest KIAA0930 as a novel candidate gene for lung cancer risk.

Identification of susceptibility pathways for the role of chromosome 15q25.1 in modifying lung cancer risk.

Genome-wide association studies (GWAS) identified the chromosome 15q25.1 locus as a leading susceptibility region for lung cancer. However, the pathogenic pathways, through which susceptibility SNPs within chromosome 15q25.1 affects lung cancer risk, have not been explored. We analyzed three cohorts with GWAS data consisting 42,901 individuals and lung expression quantitative trait loci (eQTL) data on 409 individuals to identify and validate the underlying pathways and to investigate the combined effect of genes from the identified susceptibility pathways. The KEGG neuroactive ligand receptor interaction pathway, two Reactome pathways, and 22 Gene Ontology terms were identified and replicated to be significantly associated with lung cancer risk, with P values less than 0.05 and FDR less than 0.1. Functional annotation of eQTL analysis results showed that the neuroactive ligand receptor interaction pathway and gated channel activity were involved in lung cancer risk. These pathways provide important insights for the etiology of lung cancer.

Traffic-derived particulate matter exposure and histone H3 modification: A repeated measures study.

BACKGROUND: Airborne particulate matter (PM) may induce epigenetic changes that potentially lead to chronic diseases. Histone modifications regulate gene expression by influencing chromatin structure that can change gene expression status. We evaluated whether traffic-derived PM exposure is associated with four types of environmentally inducible global histone H3 modifications. METHODS: The Beijing Truck Driver Air Pollution Study included 60 truck drivers and 60 office workers examined twice, 1-2 weeks apart, for ambient PM10 (both day-of and 14-day average exposures), personal PM2.5, black carbon (BC), and elemental components (potassium, sulfur, iron, silicon, aluminum, zinc, calcium, and titanium). For both PM10 measures, we obtained hourly ambient PM10 data for the study period from the Beijing Municipal Environmental Bureau's 27 representatively distributed monitoring stations. We then calculated a 24h average for each examination day and a moving average of ambient PM10 measured in the 14 days prior to each examination. Examinations measured global levels of H3 lysine 9 acetylation (H3K9ac), H3 lysine 9 tri-methylation (H3K9me3), H3 lysine 27 tri-methylation (H3K27me3), and H3 lysine 36 tri-methylation (H3K36me3) in blood leukocytes collected after work. We used adjusted linear mixed-effect models to examine percent changes in histone modifications per each µg/m3 increase in PM exposure. RESULTS: In all participants each µg/m3 increase in 14-day average ambient PM10 exposure was associated with lower H3K27me3 (ß=-1.1%, 95% CI: -1.6, -0.6) and H3K36me3 levels (ß=-0.8%, 95% CI: -1.4, -0.1). Occupation-stratified analyses showed associations between BC and both H3K9ac and H3K36me3 that were stronger in office workers (ß=4.6%, 95% CI: 0.9, 8.4; and ß=4.1%, 95% CI: 1.3; 7.0 respectively) than in truck drivers (ß=0.1%, 95% CI: -1.3, 1.5; and ß=0.9%, 95% CI: -0.9, 2.7, respectively; both pinteraction <0.05). Sex-stratified analyses showed associations between examination-day PM10 and H3K9ac, and between BC and H3K9me3, were stronger in women (ß=10.7%, 95% CI: 5.4, 16.2; and ß=7.5%, 95% CI: 1.2, 14.2, respectively) than in men (ß=1.4%, 95% CI: -0.9, 3.7; and ß=0.9%, 95% CI: -0.9, 2.7, respectively; both pinteraction <0.05). We observed no associations between personal PM2.5 or elemental components and histone modifications. CONCLUSIONS: Our results suggest a possible role of global histone H3 modifications in effects of traffic-derived PM exposures, particularly BC exposure. Future studies should assess the roles of these modifications in human diseases and as potential mediators of air pollution-induced disease, in particular BC exposure.

Inflammatory Markers and Genetic Polymorphisms in Workers Exposed to Flour Dust.

OBJECTIVE: The aim of this study is to evaluate inflammatory markers and pro-inflammatory CD14 and Toll-like Receptor 4 (TLR4) polymorphisms in workers exposed to flour dust. METHODS: Polymorphisms in TLR4 and CD14 were identified in our study population of 167 workers that included 63 healthy subjects (HS), 45 atopic subjects (A), and 59 subjects diagnosed clinically with occupational asthma/rhinitis (OAR). Endpoint measures in this study included fractional exhaled nitric oxide and serum concentrations of interleukin IL-6, IL-8, and tumor necrosis factor-alpha (TNF-α). RESULTS: We identified a polymorphism in CD14 (rs2569190) that may be differentially expressed (P = 0.06). IL-6 concentrations in the serum were significantly higher in the A and OAR groups (P < 0.01) than in subjects in the HS group, while IL-8 concentrations were significantly elevated only in the OAR group (P < 0.01). Interestingly, TNF-α concentrations in the OAR group were significantly reduced when compared with subjects in the HS group (P < 0.01). CONCLUSION: Cytokines are likely a defensive response in atopic and healthy workers. A protective genotype is hypothesized for occupational asthma.

Blood DNA methylation, nevi number, and the risk of melanoma.

Germline mutations determining increased cutaneous malignant melanoma (CMM) risk have been identified in familial and sporadic CMM cases, but they account only for a small proportion of CMM cases. Recent evidence suggests that germline epimutations (e.g. DNA methylation alterations), which can be inherited similarly to genomic mutations and can be detected in normal body cells (including blood), might increase susceptibility to cancer. The aim of the study was to identify germline epimutations of genes that were found to be mutated in familial CMM (p16, p14, CDK4, MC1R, hTERT), immune and inflammatory genes (ICAM-1, TNFα), DNA mismatch repair gene (MLH1), and repetitive elements (ALU, LINE-1, HERV-w). We measured DNA methylation using bisulfite pyrosequencing in peripheral blood mononuclear cells from 167 CMM cases and 164 sex-matched and age-matched controls. We used multivariable logistic regression models to evaluate the association between methylation levels and CMM status or presence of dysplastic nevi. We found an association between the risk of CMM and peripheral blood mononuclear cell methylation levels of TNFα [odds ratio (OR)=1.11, 95% confidence interval (CI)=1.03-1.18], CDK4 (OR=0.76, 95% CI=0.64-0.91), and MLH1 (OR=1.12, 95% CI=1.02-1.22). In control participants, the risk of developing dysplastic nevi was associated with methylation levels of TNFα (OR=0.81, 95% CI=0.69-0.95), hTERT (OR=0.90, 95% CI=0.82-0.99), and ALU (OR=1.56, 95% CI=1.02-2.39). Epimutations in CMM susceptibility genes and in genes involved in response to oxidative damage are associated with the risk of developing CMM or dysplastic nevi. Further studies measuring methylation levels of these genes in prospectively collected samples are warranted to further elucidate their role in the development and progression of CMM.

Airborne concentrations of chrysotile asbestos in serpentine quarries and stone processing facilities in Valmalenco, Italy.

Asbestos may be naturally present in rocks and soils. In some cases, there is the possibility of releasing asbestos fibres into the atmosphere from the rock or soil, subsequently exposing workers and the general population, which can lead to an increased risk of developing asbestos-related diseases. In the present study, air contaminated with asbestos fibres released from serpentinites was investigated in occupational settings (quarries and processing factories) and in the environment close to working facilities and at urban sites. The only naturally occurrence of asbestos found in Valmalenco area was chrysotile; amphibole fibres were never detected. An experimental cut-off diameter of 0.25 µm was established for distinguishing between Valmalenco chrysotile and antigorite single fibres using selected area electron diffraction analyses. Air contamination from chrysotile fibres in the examined occupational settings was site-dependent as the degree of asbestos contamination of Valmalenco serpentinites is highly variable from place to place. Block cutting of massive serpentinites with multiple blades or discs and drilling at the quarry sites that had the highest levels of asbestos contamination generated the highest exposures to (i.e. over the occupational exposure limits) asbestos. Conversely, working activities on foliated serpentinites produced airborne chrysotile concentrations comparable with ambient levels. Environmental chrysotile concentrations were always below the Italian limit for life environments (0.002 f ml(-1)), except for one sample collected at a quarry property boundary. The present exposure assessment study should encourage the development of an effective and concordant policy for proper use of asbestos-bearing rocks and soils as well as for the protection of public health.

Inhalable metal-rich air particles and histone H3K4 dimethylation and H3K9 acetylation in a cross-sectional study of steel workers.

BACKGROUND: Epidemiology investigations have linked exposure to ambient and occupational air particulate matter (PM) with increased risk of lung cancer. PM contains carcinogenic and toxic metals, including arsenic and nickel, which have been shown in in vitro studies to induce histone modifications that activate gene expression by inducing open-chromatin states. Whether inhalation of metal components of PM induces histone modifications in human subjects is undetermined. OBJECTIVES: We investigated whether the metal components of PM determined activating histone modifications in 63 steel workers with well-characterized exposure to metal-rich PM. METHODS: We determined histone 3 lysine 4 dimethylation (H3K4me2) and histone 3 lysine 9 acetylation (H3K9ac) on histones from blood leukocytes. Exposure to inhalable metal components (aluminum, manganese, nickel, zinc, arsenic, lead, iron) and to total PM was estimated for each study subject. RESULTS: Both H3K4me2 and H3K9ac increased in association with years of employment in the plant (p-trend = 0.04 and 0.006, respectively). H3K4me2 increased in association with air levels of nickel [ß = 0.16; 95% confidence interval (CI), 0.03-0.3], arsenic (ß = 0.16; 95% CI, 0.02-0.3), and iron (ß = 0.14; 95% CI, 0.01-0.26). H3K9ac showed nonsignificant positive associations with air levels of nickel (ß = 0.24; 95% CI, -0.02 to 0.51), arsenic (ß = 0.21; 95% CI, -0.06 to 0.48), and iron (ß = 0.22; 95% CI, -0.03 to 0.47). Cumulative exposures to nickel and arsenic, defined as the product of years of employment by metal air levels, were positively correlated with both H3K4me2 (nickel: ß = 0.16; 95% CI, 0.01-0.3; arsenic: ß = 0.16; 95% CI, 0.03-0.29) and H3K9ac (nickel: ß = 0.27; 95% CI, 0.01-0.54; arsenic: ß = 0.28; 95% CI, 0.04-0.51). CONCLUSIONS: Our results indicate histone modifications as a novel epigenetic mechanism induced in human subjects by long-term exposure to inhalable nickel and arsenic.

Exposure to particulate air pollution and risk of deep vein thrombosis.

BACKGROUND: Particulate air pollution has been linked to heart disease and stroke, possibly resulting from enhanced coagulation and arterial thrombosis. Whether particulate air pollution exposure is related to venous thrombosis is unknown. METHODS: We examined the association of exposure to particulate matter of less than 10 microm in aerodynamic diameter (PM10) with deep vein thrombosis (DVT) risk in 870 patients and 1210 controls from the Lombardy region in Italy, who were examined between 1995 and 2005. We estimated exposure to PM10 in the year before DVT diagnosis (cases) or examination (controls) through area-specific mean levels obtained from ambient monitors. RESULTS: Higher mean PM10 level in the year before the examination was associated with shortened prothrombin time (PT) in DVT cases (standardized regression coefficient [beta] = -0.12; 95% confidence interval [CI], -0.23 to 0.00) (P = .04) and controls (beta = -0.06; 95% CI, -0.11 to 0.00) (P = .04). Each increase of 10 microg/m3 in PM10 was associated with a 70% increase in DVT risk (odds ratio [OR], 1.70; 95% CI, 1.30 to 2.23) (P < .001) in models adjusting for clinical and environmental covariates. The exposure-response relationship was approximately linear over the observed PM10 range. The association between PM10 level and DVT risk was weaker in women (OR, 1.40; 95% CI, 1.02 to 1.92) (P = .02 for the interaction between PM10 and sex), particularly in those using oral contraceptives or hormone therapy (OR, 0.97; 95% CI, 0.58 to 1.61) (P = .048 for the interaction between PM10 level and hormone use). CONCLUSIONS: Long-term exposure to particulate air pollution is associated with altered coagulation function and DVT risk. Other risk factors for DVT may modulate the effect of particulate air pollution.

Changes in DNA methylation patterns in subjects exposed to low-dose benzene.

Aberrant DNA methylation patterns, including global hypomethylation, gene-specific hypermethylation/hypomethylation, and loss of imprinting (LOI), are common in acute myelogenous leukemia (AML) and other cancer tissues. We investigated for the first time whether such epigenetic changes are induced in healthy subjects by low-level exposure to benzene, a widespread pollutant associated with AML risk. Blood DNA samples and exposure data were obtained from subjects with different levels of benzene exposure, including 78 gas station attendants, 77 traffic police officers, and 58 unexposed referents in Milan, Italy (personal airborne benzene range, < 6-478 microg/m(3)). Bisulfite-PCR pyrosequencing was used to quantitate DNA methylation in long interspersed nuclear element-1 (LINE-1) and AluI repetitive elements as a surrogate of genome-wide methylation and examine gene-specific methylation of MAGE-1 and p15. Allele-specific pyrosequencing of the H19 gene was used to detect LOI in 96 subjects heterozygous for the H19 imprinting center G/A single-nucleotide polymorphism. Airborne benzene was associated with a significant reduction in LINE-1 (-2.33% for a 10-fold increase in airborne benzene levels; P = 0.009) and AluI (-1.00%; P = 0.027) methylation. Hypermethylation in p15 (+0.35%; P = 0.018) and hypomethylation in MAGE-1 (-0.49%; P = 0.049) were associated with increasing airborne benzene levels. LOI was found only in exposed subjects (4 of 73, 5.5%) and not in referents (0 of 23, 0.0%). However, LOI was not significantly associated with airborne benzene (P > 0.20). This is the first human study to link altered DNA methylation, reproducing the aberrant epigenetic patterns found in malignant cells, to low-level carcinogen exposure.

Monitoring low benzene exposure: comparative evaluation of urinary biomarkers, influence of cigarette smoking, and genetic polymorphisms.

Benzene is a human carcinogen and an ubiquitous environmental pollutant. Identification of specific and sensitive biological markers is critical for the definition of exposure to low benzene level and the evaluation of the health risk posed by this exposure. This investigation compared urinary trans,trans-muconic acid (t,t-MA), S-phenylmercapturic acid, and benzene (U-benzene) as biomarkers to assess benzene exposure and evaluated the influence of smoking and the genetic polymorphisms CYP2E1 (RsaI and DraI) and NADPH quinone oxidoreductase-1 on these indices. Gas station attendants, urban policemen, bus drivers, and two groups of controls were studied (415 subjects). Median benzene exposure was 61, 22, 21, 9 and 6 microg/m(3), respectively, with higher levels in workers than in controls. U-benzene, but not t,t-MA and S-phenylmercapturic acid, showed an exposure-related increase. All the biomarkers were strongly influenced by cigarette smoking, with values up to 8-fold higher in smokers compared with nonsmokers. Significant correlations of the biomarkers with each other and with urinary cotinine were found. A possible influence of genetic polymorphism of CYP2E1 (RsaI and/or DraI) on t,t-MA and U-benzene in subjects with a variant allele was found. Multiple linear regression analysis correlated the urinary markers with exposure, smoking status, and CYP2E1 (RsaI; R(2) up to 0.55 for U-benzene). In conclusion, in the range of investigated benzene levels (<478 micro/m(3) or <0.15 ppm), smoking may be regarded as the major source of benzene intake; among the study indices, U-benzene is the marker of choice for biomonitoring low-level occupational and environmental benzene exposure.

Urinary t,t-muconic acid, S-phenylmercapturic acid and benzene as biomarkers of low benzene exposure.

This research compared the capability of urinary trans,trans-muconic acid (t,t-MA), S-phenylmercapturic acid (S-PMA) and benzene excreted in urine (U-benzene) to monitor low benzene exposure and evaluated the influence of smoking habit on these indices. Gasoline attendants, urban policemen, bus drivers and two groups of referents working in two large Italian cities (415 people) were studied. Median benzene exposure was 61, 22, 21, 9 and 6 microg/m3, respectively, with higher levels in workers than in referents. U-benzene, but not t,t-MA and S-PMA, showed an exposure-related increase. All the biomarkers were strongly influenced by cigarette smoking, with values up to five-fold higher in smokers compared to non-smokers. In conclusion, in the range of investigated benzene exposure (<478 microg/m3 or <0.15 ppm), the smoking habit may be regarded as a major source of benzene intake; among the study indices, U-benzene is the marker of choice for the biological monitoring of occupational and environmental exposure.