OverviewHuman exposure to chemical carcinogens can result in cancer. What dictates this outcome is relatively predictable but highly variable. Factors governing the outcome include type of exposure, amount of exposure, time of exposure, and genetic makeup of the human host. The latter is comprised of variations in single nucleotides within genes (e.g., single nucleotide polymorphisms in DNA repair genes), as well as the metabolomic, proteomic, microbiomic, transcriptomic, and epigenomic background of the individual. It is becoming increasingly clear that these endpoints and signatures can be modified by chemical carcinogens and that the inflammatory load influences outcome. To this end, the past decade has seen an explosion of extremely sensitive and highly accurate technology for measuring the impact of carcinogens. Linking this technology to the rapid development of bioinformatics has enabled us to begin merging the totality of lifetime exposure (“exposome”) with the totality of metabolomic, proteomic, microbiomic, transcriptomic, epigenomic, and other “omic” profiles. We feel optimistic that the next decade will bring the development of tools to identify an individual's weighted risk signature as a biomarker for cancer risk and develop a personalized and precise approach to cancer chemoprevention and treatment.

Sarcoidosis and chronic beryllium disease (CBD) are phenocopies, however the latter one has a clear trigger factor that is beryllium exposure. This study analyses single nucleotide polymorphisms (SNPs) in a large cohort for beryllium-exposed persons. SNPs were chosen for their relevance in sarcoidosis. Even though one of largest cohorts of beryllium-exposed persons was analysed, no statistically relevant association between any SNP and CBD could be verified. Notably, some SNPs exhibit inverse OR for beryllium sensitization and CBD with nominally statistical significance, which allows hypothesizing about pathophysiological role of genes for the disease triggering and development.

Currently lung cancer accounts for 1.7 million deaths worldwide and more than 150,000 deaths in the United States yearly. The molecular events responsible for the development of lung carcinogenesis are becoming elucidated with continuing research. Carcinogens in tobacco, tobacco smoke, vehicle exhaust, and industrial effluents are inhaled mostly and act either directly or following biotransformation to damage DNA. DNA damage leads to dysregulation of cell growth and renewal, which in turn is followed by mutagenesis, hyperplasia, and malignancy. This chapter describes the multistage process of human chemical carcinogenesis through the example of lung cancer. Major players in the carcinogenic process leading to human lung cancer include chemicals of the classes: (1) polycyclic aromatic hydrocarbons (PAHs), (2) tobacco-specific nitrosamines (TSNs), and (3) diesel engine exhaust fumes (DEEF). Evidence of exposure to these classes of chemicals can be found through DNA adduct formation in normal lung tissue, mutational spectra in lung tumor DNA, and changes in gene expression specific to lung tumor pathology. Knowledge of the dysregulated biological pathways involved in carcinogenesis, including proliferative mechanisms, tyrosine kinase upregulation, motility, differentiation, angiogenesis, and metastasis, have led to development of individualized or precision therapies. In the future, early detection of lung cancer will likely involve low-dose computed tomography, which will allow successful implementation of personized therapeutic approaches.

Malignant mesothelioma is a neoplasm associated with occupational and environmental inhalation exposure to asbestos* fibers and other elongate mineral particles (EMPs) (1-3). Patients have a median survival of approximately 1 year from the time of diagnosis (1). The latency period from first causative exposure to malignant mesothelioma development typically ranges from 20 to 40 years but can be as long as 71 years (2,3). Hazardous occupational exposures to asbestos fibers and other EMPs have occurred in a variety of industrial operations, including mining and milling, manufacturing, shipbuilding and repair, and construction (3). Current exposures to commercial asbestos in the United States occur predominantly during maintenance operations and remediation of older buildings containing asbestos (3,4). To update information on malignant mesothelioma mortality (5), CDC analyzed annual multiple cause-of-death records† for 1999-2015, the most recent years for which complete data are available. During 1999-2015, a total of 45,221 deaths with malignant mesothelioma mentioned on the death certificate as the underlying or contributing cause of death were reported in the United States, increasing from 2,479 deaths in 1999 to 2,597 in 2015 (in the same time period the age-adjusted death rates§ decreased from 13.96 per million in 1999 to 10.93 in 2015). Malignant mesothelioma deaths increased for persons aged ≥85 years, both sexes, persons of white, black, and Asian or Pacific Islander race, and all ethnic groups. Despite regulatory actions and the decline in use of asbestos the annual number of malignant mesothelioma deaths remains substantial. The continuing occurrence of malignant mesothelioma deaths underscores the need for maintaining measures to prevent exposure to asbestos fibers and other causative EMPs and for ongoing surveillance to monitor temporal trends.

Objective:Beryllium workers may better understand their genetic susceptibility to chronic beryllium disease (CBD) expressed as population-based prevalence, rather than odds ratios from case-control studies.Methods:We calculated CBD prevalences from allele-specific DNA sequences of 853 workers for Human Leukocyte Antigen (HLA)-DPB1 genotypes and groups characterized by number of E69-containing alleles and by calculated surface electronegativity of HLA-DPB1.Results:Of 18 groups of at least 10 workers with specific genotypes, CBD prevalence was highest, 72.7%, for the HLA-DPB102:01:02/DPB117:01 genotype. Population-based grouped genotypes with two E69 alleles wherein one allele had -9 surface charge had a beryllium sensitization (BeS) of 52.6% and a CBD prevalence of 42.1%.Conclusions:The high CBD and BeS prevalences associated with -9-charged E69 alleles and two E69s suggest that workers may benefit from knowing their genetic susceptibility in deciding whether to avoid future beryllium exposure.

Silicosis is a potentially fatal but preventable occupational lung disease caused by inhaling respirable crystalline silica (silica). Chronic silicosis, the most common form, occurs after exposure to relatively low silica concentrations for >10 years. Accelerated silicosis occurs after 5-10 years of exposure to higher silica levels, and acute silicosis can occur after only weeks or months of exposure to extremely high silica concentrations. New national mortality data for silicosis have become available since a previous report on silicosis surveillance was published earlier this year. CDC reviewed multiple cause-of-death mortality files from the National Center for Health Statistics to analyze deaths from silicosis (International Classification of Diseases, 10th Revision diagnosis code J62: a pneumoconiosis due to dust containing silica) reported during 1999-2013. Each record lists one underlying cause of death (the disease or injury that initiated the chain of events that led directly and inevitably to death), and up to 20 contributing causes of death (other significant conditions contributing to death but not resulting in underlying cause). Available death certificates from 35 states were reviewed for the period 2004-2006 to identify occupations associated with silicosis among decedents aged 15-44 years. Results indicate that despite substantial progress in eliminating silicosis, silicosis deaths continue to occur. Of particular concern are silicosis deaths in young adults (aged 15-44 years). These young deaths likely reflect higher exposures than those causing chronic silicosis mortality in older persons, some of sufficient magnitude to cause severe disease and death after relatively short periods of exposure. A total of 12 such deaths occurred during 2011-2013, with nine that had silicosis listed as the underlying cause of death.

Silicosis is a preventable occupational lung disease caused by the inhalation of respirable crystalline silica dust and can progress to respiratory failure and death. No effective specific treatment for silicosis is available; patients are provided supportive care, and some patients may be considered for lung transplantation. Chronic silicosis can develop or progress even after occupational exposure has ceased. The number of deaths from silicosis declined from 1,065 in 1968 to 165 in 2004. Hazardous occupational exposures to silica dust have long been known to occur in a variety of industrial operations, including mining, quarrying, sandblasting, rock drilling, road construction, pottery making, stone masonry, and tunneling operations. Recently, hazardous silica exposures have been newly documented during hydraulic fracturing of gas and oil wells and during fabrication and installation of engineered stone countertops. To describe temporal trends in silicosis mortality in the United States, CDC analyzed annual multiple cause-of-death data for 2001-2010 for decedents aged ≥15 years. During 2001-2010, a total of 1,437 decedents had silicosis coded as an underlying or contributing cause of death. The annual number of silicosis deaths declined from 164 (death rate† = 0.74 per 1 million population) in 2001 to 101 (0.39 per 1 million) in 2010 (p = 0.002). Because of new operations and tasks placing workers at risk for silicosis, efforts to limit workplace exposure to crystalline silica need to be maintained.

The polycyclic aromatic hydrocarbon (PAH) benzo(a)pyrene (BP) is thought to bind covalently to DNA, through metabolism by cytochrome P450 1A1 (CYP1A1) and CYP1B1, and other enzymes, to form r7, t8, t9-trihydroxy-c-10-(N (2) -deoxyguanosyl)-7,8,9,10-tetrahydro-benzo[a]-pyrene (BPdG). Evaluation of RNA expression data, to understand the contribution of different metabolic enzymes to BPdG formation, is typically presented as fold-change observed upon BP exposure, leaving the actual number of RNA transcripts unknown. Here, we have quantified RNA copies/ng cDNA (RNA cpn) for CYP1A1 and CYP1B1, as well as NAD(P)H:quinone oxidoreductase 1 (NQO1), which may reduce formation of BPdG adducts, using primary normal human mammary epithelial cell (NHMEC) strains, and the MCF-7 breast cancer cell line. In unexposed NHMECs, basal RNA cpn values were 58-836 for CYP1A1, 336-5587 for CYP1B1 and 5943-40112 for NQO1. In cells exposed to 4.0 A mu M BP for 12h, RNA cpn values were 251-13234 for CYP1A1, 4133-57078 for CYP1B1 and 4456-55887 for NQO1. There were 3.5 (mean, range 0.2-15.8) BPdG adducts/10(8) nucleotides in the NHMECs (n = 16), and 790 in the MCF-7s. In the NHMECs, BP-induced CYP1A1 RNA cpn was highly associated with BPdG (P = 0.002), but CYP1B1 and NQO1 were not. Western blots of four NHMEC strains, chosen for different levels of BPdG adducts, showed a linear correlation between BPdG and CYP1A1, but not CYP1B1 or NQO1. Ethoxyresorufin-O-deethylase (EROD) activity, which measures CYP1A1 and CYP1B1 together, correlated with BPdG, but NQO1 activity did not. Despite more numerous levels of CYP1B1 and NQO1 RNA cpn in unexposed and BP-exposed NHMECs and MCF-7cells, BPdG formation was only correlated with induction of CYP1A1 RNA cpn. The higher level of BPdG in MCF-7 cells, compared to NHMECs, may have been due to a much increased induction of CYP1A1 and EROD. Overall, BPdG correlation was observed with CYP1A1 protein and CYP1A1/1B1 enzyme activity, but not with CYP1B1 or NQO1 protein, or NQO1 enzyme activity.

Carcinogen–DNA adducts can form following endogenous or exogenous exposure to reactive chemical species, usually electrophiles, that modify the structure of DNA. The DNA repair system comprises the products of more than 150 genes that form discrete units or an apparatus of one to eight proteins, which can detect DNA damage, effect cell cycle arrest if necessary, eliminate the damage, patch the DNA molecule, and reconstitute the DNA helix (usually by ligation). The carcinogen–DNA adduct formation is a DNA-damaging event that potentiates the development of mutations, which constitute the molecular basis of a disease, including cancer. The process of DNA repair can mitigate the molecular basis of disease, but it is complex and sometimes imperfect. This article is an attempt to outline some of the many ways that carcinogen–DNA adducts are formed, and to describe the basic mechanisms of DNA repair.

Multiple epidemiologic studies demonstrate associations between chronic beryllium disease (CBD), beryllium sensitization (BeS), and HLA-DPB1 alleles with a glutamic acid residue at position 69 (E69). Results suggest that the less-frequent E69 variants (non-*0201/*0202 alleles) might be associated with greater risk of CBD. In this study, we sought to define specific E69-carrying alleles and their amino acid sequences in the DP peptide binding groove, as well as their relationship to CBD and BeS risk, using the largest case control study to date. We enrolled 502 BeS/CBD subjects and 653 beryllium-exposed controls from three beryllium industries who gave informed consent for participation. Non-Hispanic white cases and controls were frequency-matched by industry. HLA-DPB1 genotypes were determined using sequence-specific primer PCR. The E69 alleles were tested for association with disease individually and grouped by amino acid structure using logistic regression. The results show that CBD cases were more likely than controls to carry a non-*02 E69 allele than an *02 E69, with odds ratios (95% confidence interval) ranging from 3.1 (2.1–4.5) to 3.9 (2.6–5.9) (p < 0.0001). Polymorphic amino acids at positions 84 and 11 were associated with CBD: DD versus GG, 2.8 (1.8–4.6), p < 0.0001; GD versus GG, 2.1 (1.5–2.8), p < 0.0001; LL versus GG, 3.2 (1.8–5.6), p < 0.0001; GL versus GG, 2.8 (2.1–3.8), p < 0.0001. Similar results were found within the BeS group and CBD/BeS combined group. We conclude that the less frequent E69 alleles confer more risk for CBD than does *0201. Recent studies examining how the composition and structure of the binding pockets influence peptide binding in MHC genes, as well of studies showing the topology of the TCR to likely bind DPB1 preferentially, give plausible biological rationale for these findings.

Benzo[a]pyrene (BP) is a potent pro-carcinogen and ubiquitous environmental pollutant. Here, we examined the induction and modulation of CYP1A1 and CYP1B1 and 10-(deoxyguanosin-N2-yl)-7,8,9-trihydroxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPdG) adduct formation in DNA from 20 primary normal human mammary epithelial cell (NHMEC) strains exposed to BP (4μM) in the absence or presence of chlorophyllin (5μM). Real-time polymerase chain reaction (RT-PCR) analysis revealed strong induction of both CYP1A1 and CYP1B1 by BP, with high levels of inter-individual variability. Variable BPdG formation was found in all strains by r7, t8-dihydroxy-t-9, 10 epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE)-DNA chemiluminescence assay (CIA). Chlorophyllin mitigated BP-induced CYP1A1 and CYP1B1 gene expression in all 20 strains when administered with BP. Chlorophyllin, administered prior to BP-exposure, mitigated CYP1A1 expression in 18/20 NHMEC strains (p<0.005) and CYP1B1 expression in 17/20 NHMEC strains (p<0.005). Maximum percent reductions of CYP1A1 and CYP1B1 gene expression and BPdG adduct formation were observed when cells were pre-dosed with chlorophyllin followed by administration of the carcinogen with chlorophyllin (p<0.005 for CYP1A1 and CYP1B1 expression and p<0.0005 for BPdG adducts). Therefore, chlorophyllin is likely to be a good chemoprotective agent for a large proportion of the human population.

OBJECTIVE:To determine if single nucleotide polymorphisms (SNPs) in interleukin (IL) IL-1A, IL-1B, IL-1RN, IL-2, IL-9, and IL-9R were associated with chronic beryllium disease (CBD) and beryllium sensitization (BeS). METHODS:Forty SNPs in six IL genes were evaluated in 85 individuals with CBD, 61 individuals with BeS, and 730 individuals without BeS or CBD (nonsensitized) using a 5' nuclease polymerase chain reaction assay. Logistic regression was used to evaluate the association between IL SNPs, CBD, and BeS, adjusting for plant-site and HLA-DPB1Glu69 in additive, dominant, and recessive inheritance models. RESULTS:IL-1A-1142, IL-1A-3769, and IL-1A-4697 were significantly associated with CBD in both the additive and dominant models compared to individuals with BeS or the nonsensitized. CONCLUSIONS:These results indicate that genetic variations in the IL-1A gene may play a role in the development of CBD but not BeS.

Background Animal models provide powerful tools for dissecting dose-response relationships and pathogenic mechanisms and for testing new treatment paradigms. Mechanistic research on beryllium exposure-disease relationships is severely limited by a general inability to develop a sufficient chronic beryllium disease animal model. Discovery of the Human Leukocyte Antigen (HLA) DPB1Glu69 genetic susceptibility component of chronic beryllium disease permitted the addition of this human beryllium antigen presentation molecule to an animal genome which may permit development of a better animal model for chronic beryllium disease. Using FVB/N inbred mice, Drs. Rubin and Zhu, successfully produced three strains of HLA-DPB1 Glu 69 transgenic mice. Each mouse strain contains a haplotype of the HLADPB1 Glu 69 gene that confers a different magnitude of odds ratio (OR) of risk for chronic beryllium disease: HLA-DPB1*0401 (OR = 0.2), HLA-DPB1*0201 (OR = 15), HLA-DPB1*1701 (OR = 240). In addition, Drs. Rubin and Zhu developed transgenic mice with the human CD4 gene to permit better transmission of signals between T cells and antigen presenting cells. This project has maintained the colonies of these transgenic mice and tested the functionality of the human transgenes.

We hypothesized that chlorophyllin (CHLN) would reduce benzo[a]pyrene-DNA (BP-DNA) adduct levels. Using normal human mammary epithelial cells (NHMECs) exposed to 4 microM BP for 24 hr in the presence or absence of 5 microM CHLN, we measured BP-DNA adducts by chemiluminescence immunoassay (CIA). The protocol included the following experimental groups: BP alone, BP given simultaneously with CHLN (BP+CHLN) for 24 hr, CHLN given for 24 hr followed by BP for 24 hr (preCHLN, postBP), and CHLN given for 48 hr with BP added for the last 24 hr (preCHLN, postBP+CHLN). Incubation with CHLN decreased BPdG levels in all groups, with 87% inhibition in the preCHLN, postBP+CHLN group. To examine metabolic mechanisms, we monitored expression by Affymetrix microarray (U133A), and found BP-induced up-regulation of CYP1A1 and CYP1B1 expression, as well as up-regulation of groups of interferon-inducible, inflammation and signal transduction genes. Incubation of cells with CHLN and BP in any combination decreased expression of many of these genes. Using reverse transcription real time PCR (RT-PCR) the maximal inhibition of BP-induced gene expression, >85% for CYP1A1 and >70% for CYP1B1, was observed in the preCHLN, postBP+CHLN group. To explore the relationship between transcription and enzyme activity, the ethoxyresorufin-O-deethylase (EROD) assay was used to measure the combined CYP1A1 and CYP1B1 activities. BP exposure caused the EROD levels to double, when compared with the unexposed controls. The CHLN-exposed groups all showed EROD levels similar to the unexposed controls. Therefore, the addition of CHLN to BP-exposed cells reduced BPdG formation and CYP1A1 and CYP1B1 expression, but EROD activity was not significantly reduced.

Use of tamoxifen is associated with a 50% reduction in breast cancer incidence and an increase in endometrial cancer incidence. Here, we documented tamoxifen-induced gene expression changes in cultured normal human mammary epithelial cells (strains 5, 16, and 40), established from tissue taken at reduction mammoplasty from three individuals. Cells exposed to 0, 10, or 50 mu mol/L of tamoxifen for 48 hours were evaluated for (E)-alpha-(deoxyguanosine-N-2-yl)-tamoxifen (dG-N-2-TAM) adduct formation using TAM-DNA (DNA modified with dG-N2-TAM) chemiluminescence immunoassay, gene expression changes using National Cancer Institute DNA-oligonucleotide microarray, and real-time PCR. At 48 hours, cells exposed to 10 and 50 mu mol/L of tamoxifen were 85.6% and 48.4% viable, respectively, and there were no measurable dG-N-2-TAM adducts. For microarrays, cells were exposed to 10 mu mol/L of tamoxifen and genes with expression changes of >3-fold were as follows: 13 genes up-regulated and 1 down-regulated for strain 16; 17 genes up-regulated for strain 5, and 11 genes up-regulated for strain 40. Interferon-inducible genes (IFITM1, IFIT1, MXI, and GIP3), and a potassium ion channel (KCNJ1) were up-regulated in all three strains. No significant expression changes were found for genes related to estrogen or xenobiotic metabolism. Real-time PCR revealed the upregulation of IFNA1 and confirmed the tamoxifen-induced upregulation of the five other genes identified by microarray, with the exception of GIP3 and MX1, which were not upregulated in strain 40. Induction of IFN-related genes in the three normal human mammary epithelial cell strains suggests that, in addition to hormonal effects, tamoxifen exposure may enhance immune response in normal breast tissue. [Cancer Res 2009;69(3):1150-5]

Carcinogenic polycylic aromatic hydrocarbons can alter immune responses. Changes in immune response gene expression profiles in multiple human mammary cell strains exposed to benzo(alpha)pyrene (BP) (4 microM) in vitro, in the presence or absence of chlorophyllin (5 microM), were observed using Affymetrix gene arrays. Expressions of five immune response genes were altered ~3.0-fold by BP exposure and 24 genes by BP in the presence chlorophyllin. In silico pathway analysis revealed altered immune response genes form interactive gene networks with many cellular processes, suggesting their role in a complex multigenic response to toxins. Additionally, it was suggestive of the possible immunomodulatory potential of chlorophyllin apart from various other well-documented mechanisms of action. Gene expression matrices revealed consistent alteration patterns involving IL1B, SECTM1 and CXCL14 on exposure to BP, and IL1RN, CD86, IF144 and GIP2 in the presence of chlorophyllin and BP, suggesting some of these genes might constitute putative immune response biomarkers of PAH exposure. This study has therefore identified a battery of potential immune response biomarkers of PAH exposure, amidst several genes, for future validation studies.