Carcinogen Exposure Research Articles

Lifespan Kras mutation levels in lung and liver of B6C3F1 mice.

Somatic mutations accumulate in the human genome and are correlated with increased cancer incidence as humans age. The standard model for studying the carcinogenic effects of exposures for human risk assessment is the rodent 2-year carcinogenicity assay. However, there is little information regarding the effect of age on cancer-driver gene mutations in these models. The mutant fraction (MF) of Kras codon 12 GGT to GAT and GGT to GTT mutations, oncogenic mutations orthologous between humans and rodents, was quantified over the lifespan of B6C3F1 mice. MFs were measured in lung and liver tissue, organs that frequently develop tumors following carcinogenic exposures. The MFs were evaluated at 4, 6, 8, 12, 21, and 85 weeks, with the 12-week and 21-week time points being coincident with the conclusion of 28-day and 90-day exposure durations used in short-term toxicity testing. The highly sensitive and quantitative Allele-specific Competitive Blocker PCR (ACB-PCR) assay was used to quantify the number of mutant Kras codon 12 alleles. The mouse lung showed a slight, but significant trend increase in the Kras codon 12 GAT mutation over the 85-week period. The trend with age can be equally well-fit by several non-linear functions, but not by a linear function. In contrast, the liver GAT mutation did not increase, and the GTT mutation did not increase for either organ. Even with the slight increase in the lung GAT MFs, our results indicate that the future use of Kras mutation as a biomarker of carcinogenic effect will not be confounded by animal age. Environ. Mol. Mutagen. 59:715-721, 2018. Published 2018. This article is a U.S. Government work and is in the public domain in the USA.

Genomic case report of a low grade bladder tumor metastasis to lung

BackgroundWe present a rare case where distant metastasis of a low grade bladder tumor was observed. We carried out detailed genomic analysis and cell based experiments on patient tumor samples to study tumor evolution, possible cause of disease and provide personalized treatment strategies.Case presentationA man with a smoking history was diagnosed with a low-grade urothelial carcinoma of the bladder and a concurrent high-grade upper urinary tract tumor. Seven years later he had a lung metastasis. We carried out exome sequencing on all the patient’s tumors and peripheral blood (germline) to identify somatic variants. We constructed a phylogenetic tree to capture how the tumors are related and to identify somatic changes important for metastasis. Although distant metastasis of low-grade bladder tumor is rare, the somatic variants in the tumors and the phylogenetic tree showed that the metastasized tumor had a mutational profile most similar to the low grade urothelial carcinoma. The primary and the metastatic tumors shared several important mutations, including in the KMT2D and the RXRA genes. The metastatic tumor also had an activating MTOR mutation, which may be important for tumor metastasis. We developed a mutational signature to understand the biologic processes responsible for tumor development. The mutational signature suggests that the tumor mutations are associated with tobacco carcinogen exposure, which is concordant with the patient’s smoking history. We cultured cells from the lung metastasis to examine proliferation and signaling mechanisms in response to treatment. The mTOR inhibitor Everolimus inhibited downstream mTOR signaling and induced cytotoxicity in the metastatic tumor cells.ConclusionWe used genomic analysis to examine a rare case of low grade bladder tumor metastasis to distant organ (lung). Our analysis also revealed exposure to carcinogens found is tobacco as a possible cause in tumor development. We further validated that the patient might benefit from mTOR inhibition as a potential salvage therapy in an adjuvant or recurrent disease setting.

Abstract B35: Biomarkers for precision application of prostacyclin lung cancer chemoprevention

Abstract Lung cancer remains the leading cause of cancer death in the United States, and chemoprevention offers an appealing area of investigation in the face of limited therapeutic success. Improvement in endobronchial histology was demonstrated in former smokers after oral iloprost chemoprevention in a phase II clinical trial. Of the 48 patients who received iloprost in the chemoprevention trial, 23 had regressive histology and 25 had stable or progressive histology. Identifying markers that predict which patients will respond to treatment will help refine target populations for future trials and clinical applications. Markers of drug activity will also facilitate clinical application by allowing monitoring during treatment. Preclinical studies indicate that iloprost, a prostacyclin analogue, acts through peroxisome proliferator activated receptor gamma (PPAR) to reduce transformed lung epithelial cell growth and markers of epithelial-to-mesenchymal transition (EMT). We hypothesize that PPAR expression status predicts response to prostacyclin chemoprevention and that expression changes in known downstream targets of PPAR and prostacyclin indicate response to prostacyclin treatment. Prostacyclin may also induce expression of PPAR, leading to increased antitumor signaling. We found that expression of PPAR and markers of prostacyclin signaling (15pgdh, CES1, and PTGS2) and EMT markers (Snail, Vimentin, Ecadherin, and CRB3) changed in cultured human bronchial epithelial cells (HBEC) in response to cigarette smoke condensate (CSC) and that iloprost reversed these changes. Expression in mice also changed in response to cigarette smoke carcinogens and prostacyclin. miR-34c is inversely correlated with bronchial histology grade and suppresses lung cancer progression. In our study, the level of miR-34c was lower in the NSCLC cell line H322 compared to HBEC. In H322, miR-34c increased with PPAR agonist treatment and decreased with PPAR antagonist treatment. In HBEC and mouse models of lung cancer chemoprevention, miR-34c increased with prostacyclin and decreased with cigarette smoke carcinogen exposure. In cultured primary progressive dysplastic bronchial epithelial cells, treatment with iloprost resulted in similar expression changes in Vimentin, Snail, Ecadherin, CRB3, CES1, and miR-34c. This study has the potential to improve iloprost lung cancer chemoprevention by allowing future trials to more effectively target high-risk patients, by providing a clinical biomarker for identification of chemoprevention candidates, and by identifying biomarkers for intermediate monitoring of response. Future studies will test expression of these markers in lung tissue biopsy and liquid biopsy samples from the oral and inhaled iloprost chemoprevention trials. Citation Format: Melissa New, Debbie McArthur, Lori Nield, Dan Merrick, Robert L. Keith, Meredith A. Tennis. Biomarkers for precision application of prostacyclin lung cancer chemoprevention [abstract]. In: Proceedings of the Fifth AACR-IASLC International Joint Conference: Lung Cancer Translational Science from the Bench to the Clinic; Jan 8-11, 2018; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(17_Suppl):Abstract nr B35.

MicroRNAs and their role in environmental chemical carcinogenesis

MicroRNAs (miRNAs) are a class of small, noncoding RNA species that play crucial roles across many biological processes and in the pathogenesis of major diseases, including cancer. Recent studies suggest that the expression of miRNA is altered by certain environmental chemicals, including metals, organic pollutants, cigarette smoke, pesticides and carcinogenic drugs. In addition, extensive studies have indicated the existence and importance of miRNA in different cancers, suggesting that cancer-related miRNAs could serve as potential markers for chemically induced cancers. The altered expression of miRNA was considered to be a vital pathogenic role in xenobiotic-induced cancer development. However, the significance of miRNA in the etiology of cancer and the exact mechanisms by which environmental factors alter miRNA expression remain relatively unexplored. Hence, understanding the interaction of miRNAs with environmental chemicals will provide important information on mechanisms underlying the pathogenesis of chemically induced cancers, and effectively diagnose and treat human cancers resulting from chronic or acute carcinogen exposure. This study presents the current evidence that the miRNA deregulation induced by various chemical carcinogens, different cancers caused by environmental carcinogens and the potentially related genes in the onset or progression of cancer. For each carcinogen, the specifically expressed miRNA may be considered as the early biomarkers of the cancer process. In this review, we also summarize various target genes of the altered miRNA, oncogenes or anti-oncogenes, and the existing evidence regarding the gene regulation mechanisms of cancer caused by environmentally induced miRNA alteration. The future perspective of miRNA may become attractive targets for the diagnosis and treatment of carcinogen-induced cancer.

Joint effect of smoking andNQO1C609T polymorphism on undifferentiated nasopharyngeal carcinoma risk in a North African population

BackgroundNasopharyngeal carcinoma (NPC) has a higher incidence in North Africa than in most parts of the world. In addition to environmental factors such as Epstein–Barr virus infection and chemical carcinogen exposure, genetic susceptibility has been reported to play a key role in the development of NPC. NAD(P)H: quinone oxidoreductase 1 is a cytosolic enzyme that protects cells from oxidative damage. A C to T transition at position 609 in the NQO1 gene (OMIM: 125860) has been shown to alter the enzymatic activity of the enzyme and has been associated with increased risk to several cancers. This study investigates for the first time the effect of this polymorphism on NPC susceptibility in a North African population.MethodsThe NQO1 C609T polymorphism was genotyped using PCR‐RFLP in 392 NPC cases and 365 controls from Morocco, Algeria, and Tunisia.ResultsThe allele frequencies and distributions of genotypes did not differ between cases and controls (p > 0.05). When stratifying according to smoking status, we observed two‐fold higher NPC risk in ever‐smokers carrying the CT or TT genotype. Multiple logistic regression analysis revealed that there was a significant interaction between T allele and smoking status (OR = 1.95, 95% CI = 1.20–3.19; interaction p = 0.007).ConclusionIn this North African population, the functional NQO1 polymorphism was associated with a significantly higher risk of NPC among smokers and did not affect the risk among nonsmokers.

Prevalence of exposure to occupational carcinogens among farmers.

Farmers experience a range of carcinogenic exposures, including some pesticides, fuels, engine exhausts, metals, some organic solvents, silica, wood dusts and solar radiation. However many studies investigating the risk of cancer in farmers focus on pesticide exposure alone. The aim of this study was to determine which carcinogens Australian farmers are exposed to, the prevalence and circumstances of those exposures, and the use of protective equipment. The study used data from the Australian Work Exposures Study (AWES) a cross-sectional study conducted in 2012 that investigated the prevalence of carcinogen exposure among Australian workers. This was supplemented with data from AWES-Western Australia (WA), conducted in 2013, which followed the same methodology but in Western Australian workers only. A total of 5498 Australian workers were interviewed about the tasks they carry out in their workplace. The 166 participants who worked in farming (126 men and 40 women, with an age range of 18-65 years) are the focus of this article. On average, farmers had been exposed to five different carcinogens. Highest numbers of exposures occurred among men and those working on mixed crop and livestock farms. Solar radiation, diesel engine exhaust and certain solvents were the most prevalent exposures, each with over 85% of farmers exposed. The main tasks leading to exposure were working outdoors, using and repairing farming equipment and burning waste. Sun protection and closed cabs on machinery were the most frequently used forms of protection. Farmers are a high risk group in relation to carcinogen exposure. The variation in tasks that they undertake results in exposure to a wide variety of different carcinogens that require similarly varied control measures.

Contribution of Tobacco Use and 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone to Three Methyl DNA Adducts in Urine

Human urinary DNA adducts may be useful surrogate biomarkers to estimate carcinogen exposure and activation, particularly if such adducts are of high selectivity from a specific carcinogen source. In this report, we provided evidence supporting tobacco use and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) being the dominant source for 3-methyladenine (3-mA), while nontobacco factors contribute significantly to 7-methylguanine and 1-methyladenine in the urine. Upon confirmation in human urine samples from larger populations in the future, urinary 3-mA may be used to estimate NNK bioactivation in smokers and to facilitate the development of a chemopreventive agent against NNK-induced carcinogenesis.

Effects of Menthol Flavor Cigarettes or Total Urinary Menthol on Biomarkers of Nicotine and Carcinogenic Exposure and Behavioral Measures.

The effects of either menthol flavor cigarettes or total urinary menthol on nicotine dependence, biomarkers of addictive and carcinogenic exposure, and behavioral measures may inform differences and similarities of these two approaches. Stratified recruitment by cigarette (menthol flavor or regular) and race (African American and white) yielded a balanced sample of 136 adult smokers in a 36-hour inpatient protocol. Exposure measures assessed during 24-hour data collection included urinary menthol, total NNAL [4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol], 10 polycyclic aromatic hydrocarbon metabolites, baseline plasma cotinine, plasma nicotine pre- and post-smoking, exhaled carbon monoxide pre- and post-smoking, and cigarette puff volumes. The latter three were measured at four specified timepoints throughout the day. There were no significant differences between menthol flavor and regular cigarette smokers in measures of nicotine dependence, biomarkers of addictive and carcinogenic exposures, or behavioral measures. Significant race × cigarette type interaction effects were found for two biomarkers: plasma nicotine and 3-hydroxyphenanthrene. Total urinary menthol was significantly associated with higher levels of nearly all dependent variables including puff volume, exhaled carbon monoxide, plasma nicotine and cotinine, NNAL, and polycyclic aromatic hydrocarbons. The significant effects of total urinary menthol were sustained after adjusting for menthol flavor and regular cigarette type and other covariates (eg, number of cigarettes per day, baseline cotinine, and baseline nicotine). Urinary menthol is an independent predictive biomarker for nicotine dependence, addictive and carcinogenic exposure, and behaviors. Comparison of the effects of menthol flavor and total urinary menthol on nicotine dependence, biomarkers of addictive and carcinogenic exposure, and behavioral measures emphasizes the important significant contribution of total urinary menthol concentrations in contrast to no significant associations by dichotomous cigarette type with these biomarkers.

The inhibitory effect of Filipendula ulmaria (L.) Maxim. on colorectal carcinogenesis induced in rats by methylnitrosourea

Ethnopharmacological relevanceMeadowsweet (Filipendula ulmaria (L.) Maxim.) is a medicinal plant with a variety of therapeutic properties, traditionally used in various diseases including treatment and prevention of tumors. The aim of this study was to present an ethnomedicinal justification that a meadowsweet decoction is able to inhibit colorectal carcinogenesis induced by the methylnitrosourea (MNU) in rats. Materials and methodsThe chemical composition of meadowsweet extracts was studied by traditional methods. In animal experiments adult outbred female rats received four intrarectal instillations of MNU, one per week, at dose 4 mg in 0.5 ml saline (the total dose of MNU during the 4-week exposure was 16 mg/rat). After carcinogenic exposure one group (MNU) of rats continued to receive standard feed and tap water throughout life. In another group (MNU+meadowsweet), rats were given daily a decoction of the meadowsweet instead of drinking water and standard feed. ResultsMeadowsweet extracts showed a sufficiently high content of flavonoids and tannins and also some individual phenolic compounds and salicylic acid. In rats after administration of MNU the overall incidence of tumors was 91% with tumor multiplicity of 3.5. The majority of rats (86%) developed multiple tumors of large intestine (most often adenocarcinomas:88 from 107; index of multiplicity – 2.0). In rats from the group MNU+meadowsweet there was a statistically significant decrease of the overall tumor incidence and multiplicity–by 1.4 and 2.9 times, respectively, and the incidence and multiplicity of colon tumors – by 2.0 and 2.8 times, respectively; the incidence and multiplicity of malignant tumors of other localizations was also reduced–by 2.2 and 3.0 times, respectively. ConclusionsMeadowsweet extract is an effective inhibitor of colorectal carcinogenesis in experiment, that provides support for the traditional use of this plant in the oncology.

Speciation Theory of Carcinogenesis Explains Karyotypic Individuality and Long Latencies of Cancers.

It has been known for over 100 years that cancers have individual karyotypes and arise only years to decades after initiating carcinogens. However, there is still no coherent theory to explain these definitive characteristics of cancer. The prevailing mutation theory holds that cancers are late because the primary cell must accumulate 3–8 causative mutations to become carcinogenic and that mutations, which induce chromosomal instability (CIN), generate the individual karyotypes of cancers. However, since there is still no proven set of mutations that transforms a normal to a cancer cell, we have recently advanced the theory that carcinogenesis is a form of speciation. This theory predicts carcinogens initiate cancer by inducing aneuploidy, which automatically unbalances thousands of genes and thus catalyzes chain-reactions of progressive aneuploidizations. Over time, these aneuploidizations have two endpoints, either non-viable karyotypes or very rarely karyotypes of new autonomous and immortal cancers. Cancer karyotypes are immortalized despite destabilizing congenital aneuploidy by clonal selections for autonomy—similar to those of conventional species. This theory predicts that the very low probability of converting the karyotype of a normal cell to that of a new autonomous cancer species by random aneuploidizations is the reason for the karyotypic individuality of new cancers and for the long latencies from carcinogens to cancers. In testing this theory, we observed: (1) Addition of mutagenic and non-mutagenic carcinogens to normal human and rat cells generated progressive aneuploidizations months before neoplastic transformation. (2) Sub-cloning of a neoplastic rat clone revealed heritable individual karyotypes, rather than the non-heritable karyotypes predicted by the CIN theory. (3) Analyses of neoplastic and preneoplastic karyotypes unexpectedly identified karyotypes with sets of 3–12 new marker chromosomes without detectable intermediates, consistent with single-step origins. We conclude that the speciation theory explains logically the long latencies from carcinogen exposure and the individuality of cancers. In addition, the theory supports the single-step origins of cancers, because karyotypic autonomy is all-or-nothing. Accordingly, we propose that preneoplastic aneuploidy and clonal neoplastic karyotypes provide more reliable therapeutic indications than current analyses of thousands of mutations.

In Vivo Stable-Isotope Labeling and Mass-Spectrometry-Based Metabolic Profiling of a Potent Tobacco-Specific Carcinogen in Rats.

The tobacco-specific nitrosamine, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), is a potent lung carcinogen that exerts its carcinogenic effects upon metabolic activation. The identification and quantitation of NNK metabolites could identify potential biomarkers of bioactivation and detoxification of this potent carcinogen and may be used to predict lung cancer susceptibility among smokers. Here, we used in vivo isotope-labeling and high-resolution-mass-spectrometry-based methods for the comprehensive profiling of all known and unknown NNK metabolites. The sample-enrichment, LC-MS, and data-analysis workflow, including a custom script for automated d0- d4- m/ z-pair-peak detection, enabled unbiased identification of numerous NNK metabolites. The structures of the metabolites were confirmed using targeted LC-MS2 with retention-time ( tR) and MS2-fragmentation comparisons to those of standards when possible. Eleven known metabolites and unchanged NNK were identified simultaneously. More importantly, our workflow revealed novel NNK metabolites, including 1,3-Diol (13), α-OH-methyl-NNAL-Gluc (14), nitro-NK- N-oxide (15), nitro-NAL- N-oxide (16), γ-OH NNAL (17), and three N-acetylcysteine (NAC) metabolites (18a-c). We measured the differences in the relative distributions of a panel of nitroso-containing NNK-specific metabolites in rats before and after phenobarbital (PB) treatment, and this served as a demonstration of a general strategy for the detection of metabolic differences in animal and cell systems. Lastly, we generated a d4-labeled NNK-metabolite mixture to be used as internal standards ( d4-rat urine) for the relative quantitation of NNK metabolites in humans, and this new strategy will be used to assess carcinogen exposure and ultimately to evaluate lung-cancer risk and susceptibility in smokers.

Abstract 5726: Rearrangement-related peptides with neoantigenic potential in malignant pleural mesothelioma

Abstract Malignant pleural mesothelioma is a disease primarily associated with exposure to the carcinogen asbestos. Consistent with this carcinogenic exposure, cytogenetic analyses have identified multiple recurrent structural chromosomal abnormalities in this malignancy, but more recent high-throughput sequencing evaluations of point mutations suggest that there is a low mutational burden in mesothelioma. Since tumor mutational burden has been correlated with responses to treatment with immune checkpoint inhibitors such as nivolumab, it was not consistent that patients with mesothelioma and low mutation burdens would have similar response rates in clinical trials with immune checkpoint inhibitors as patients with non-small cell lung cancer which is associated with a high mutation burden. In order to reconcile these differences, and given the potential for an improved understanding of the molecular pathogenesis of mesothelioma to improve therapeutic options, we used mate-pair sequencing (MPseq) and RNA sequencing (RNAseq) to understand how structural variants affect the transcriptome. MPseq differs from standard next generation sequencing approaches by tiling the whole genome with larger fragments (2-5kb) to reliably detect structural variants such as insertions, deletions and rearrangements. Amongst 22 mesothelioma specimens there were 1535 chromosomal rearrangements (median 41, range 3-298 per specimen), that resulted in junctions or novel fusions of non-coding DNA or genes. Six-hundred thirty-seven of these rearrangements (median 22, range 5-103 range per specimen) resulted in novel fusions of genes. Many of these inter- or intra-chromosomal rearrangements were consistent with a pattern of chromoanagesis such as chromoplexy or chromothripsis. Chromosomal rearrangements detected by MPseq were used to guide analysis of RNAseq data and revealed that these chromosomal junctions resulted in the expression of 179 novel amino acid sequences (median 5, 0-51 range per specimen). To determine whether transcription of chromosomal rearrangement-related junctions have neoantigenic potential, we used in silico tools to determine whether any of the expressed junctions contained peptides that could be presented by patient-specific HLA molecules. The top candidate rearrangement-related peptides with neoantigenic potential bound patient-specific HLA molecules nearly as well or as well as a positive control in competitive binding assays. Our findings represent the discovery of potential neoantigen expression driven by structural chromosomal rearrangements. These results may have implications for the development of novel therapeutic strategies, the selection of patients to receive immunotherapy, and blood-based treatment monitoring strategies. Citation Format: Aaron S. Mansfield, Tobias Peikert, James B. Smadbeck, Julia B. Udell, Farhad Kosari, Stephen J. Murphy, Hongzheng Ren, Vishnu V. Serla, Janet L. Schaefer Klein, Giannoula Karagouga, Faye R. Harris, Carlos Sosa, Sarah H. Johnson, Wendy Nevala, Svetomir N. Markovic, Aaron O. Bungum, Eric S. Edell, Haidong Dong, John C. Cheville, Marie Christine Aubry, Jin Jen, George Vasmatzis. Rearrangement-related peptides with neoantigenic potential in malignant pleural mesothelioma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5726.

PO-389 Epigenomic and mutation determinants of carcinogen-driven primary epithelial cell immortalization

IntroductionBased on cancer genome sequencing, almost 600 genes have been implicated in cancer development to date, and recently, chromatin structural changes have emerged as a key mechanism for translating exogenous cues, such as carcinogen exposures, to the deregulation of biological processes implicated in tumorigenesis. However, the functional relationship between genetic and epigenetic alterations in the context of early events of carcinogen-driven cell transformation is not well understood.Material and methodsWe employ an in vitro carcinogen exposure system using primary human mammary epithelial cells (HMEC), which utilises a biological barrier bypass to mimic early steps of cell transformation, to introduce genetic and epigenetic alterations and to investigate their functional interplay during cell immortalization. A temporal, integrated (epi)genomic analysis is applied to benzo[a]pyrene (B[a]P)-exposed, immortalised HMEC, involving whole genome sequencing (WGS), analyses of chromatin structure (ChIP-seq, ATAC-seq), and patterns of gene regulation (DNA methylome 850K array, RNA-seq).Results and discussionsWGS of sequentially derived post-stasis and immortalised clones, as well as tumorigenic clones (generated by TP53 insertional mutagenesis of post-stasis lines), identified a prominent B[a]P mutational signature across all clones. While all sequenced clones had comparable SBS counts, we observed a gradual increase of structural variants, most notably copy number variations, from post-stasis to immortal and tumorigenic clones. Reproducibly distinct methylome patterns also correlated closely with the individual phenotypic stages of cell transformation. Chromatin accessibility analysis further showed changes characteristic to individual as well as multiple exposed clones, such as in the breast cancer tumour suppressor gene BRCA2 or the promoter region of PCDH18, a gene frequently inactivated by promoter methylation in colorectal cancer. Cell type-matched histone modification and RNA expression patterns are being established to enable a thorough integrated analysis of genetic and epigenetic determinants of carcinogen-driven HMEC transformation.ConclusionWe propose that in-depth, integrated (epi)genomic analysis of this innovative model system can reveal novel molecular determinants of carcinogen-driven cellular immortalization, thus improving our understanding of cancer causes, development and progression.Funding INCa-INSERM Plan Cancer 2015 Grant.

The Airway Transcriptome as a Biomarker for Early Lung Cancer Detection.

Lung cancer remains the leading cause of cancer-related death due to its advanced stage at diagnosis. Early detection of lung cancer can be improved by better defining who should be screened radiographically and determining which imaging-detected pulmonary nodules are malignant. Gene expression biomarkers measured in normal-appearing airway epithelium provide an opportunity to use lung cancer-associated molecular changes in this tissue for early detection of lung cancer. Molecular changes in the airway may result from an etiologic field of injury and/or field cancerization. The etiologic field of injury reflects the aberrant physiologic response to carcinogen exposure that creates a susceptible microenvironment for cancer initiation. In contrast, field cancerization reflects effects of "first-hit" mutations in a clone of cells from which the tumor ultimately arises or the effects of the tumor on the surrounding tissue. These fields might have value both for assessing lung cancer risk and diagnosis. Cancer-associated gene expression changes in the bronchial airway have recently been used to develop and validate a 23-gene classifier that improves the diagnostic yield of bronchoscopy for lung cancer among intermediate-risk patients. Recent studies have demonstrated that these lung cancer-related gene expression changes extend to nasal epithelial cells that can be sampled noninvasively. While the bronchial gene expression biomarker is being adopted clinically, further work is necessary to explore the potential clinical utility of gene expression profiling in the nasal epithelium for lung cancer diagnosis, lung cancer risk assessment, and precision medicine for lung cancer treatment and chemoprevention. Clin Cancer Res; 24(13); 2984-92. ©2018 AACR.

Genotyping of single nucleotide polymorphisms using the SNP-RFLP method.

Genetic polymorphisms, including single nucleotide polymorphisms (SNPs), are responsible for inter-individual variability in susceptibility to cancer and other disorders. Both environmental factors (e.g., smoking or carcinogen exposure) and genetic variation underlie the development of cancer; however, studies of twins suggest that genetic variation is more important. Hence, the identification of SNPs makes an important contribution to cancer research. In this study, 13 SNPs in 12 genes were genotyped in HEK 293 and HeLa cells using the simple and inexpensive SNP-RFLP method. Sanger sequencing was performed for one SNP to validate the SNP-RFLP results. Of the 13 SNPs, 10 were homozygous and three were heterozygous (rs10937405, rs12296850, and rs3814113) in HEK 293 cells, while 12 were homozygous and one was heterozygous (rs995030) in HeLa cells. The cells carried eight disease-associated risk alleles (32% of typed alleles), including rs2853677, rs995030, rs2736100, and rs6010620 in HEK 293 cells, and rs10937405, rs3814113, rs4767364, and rs6010620 in HeLa cells. Four SNP loci were homozygous for different alleles in each cell line, with HEK 293 cells having a CC genotype at rs2853677, GG at rs2736100 and rs4767364, and TT at rs3819197, while HeLa cells had TT genotypes at rs2853677 and rs2736100, AA at rs4767364, and CC at rs3819197. In conclusion, these results are potentially applicable for testing of novel gene therapeutic approaches in future experiments where the non-risk alleles of the eight identified risk alleles are substituted into HEK 293 or HeLa cells.

Case Studies of Gastric, Lung, and Oral Cancer Connect Etiologic Agent Prevalence to Cancer Incidence.

Obtaining detailed individual-level data on both exposure and cancer outcomes is challenging, and it is difficult to understand and characterize how temporal aspects of exposures translate into cancer risk. We show that, in lieu of individual-level information, population-level data on cancer incidence and etiologic agent prevalence can be leveraged to investigate cancer mechanisms and to better characterize and predict cancer trends. We use mechanistic carcinogenesis models [multistage clonal expansion (MSCE) models] and data on smoking, Helicobacter pylori (H. pylori), and HPV infection prevalence to investigate trends of lung, gastric, and HPV-related oropharyngeal cancers. MSCE models are based on the initiation-promotion-malignant conversion paradigm and allow for interpretation of trends in terms of general biological mechanisms. We assumed the rates of initiation depend on the prevalence of the corresponding risk factors. We performed two types of analysis, using the agent prevalence and cancer incidence data to estimate the model parameters and using cancer incidence data to infer the etiologic agent prevalence as well as the model parameters. By including risk factor prevalence, MSCE models with as few as three parameters closely reproduced 40 years of age-specific cancer incidence data. We recovered trends of H. pylori prevalence in the United States and demonstrated that cohort effects can explain the observed bimodal, age-specific pattern of oral HPV prevalence in men. Our results demonstrate the potential for joint analyses of population-level cancer and risk factor data through mechanistic modeling. This approach can be a first step in systematically testing relationships between exposures and cancer risk when individual-level data is lacking.Significance: Analysis of trends in risk-factor prevalence and cancer incidence can shed light on cancer mechanisms and the way that carcinogen exposure through time shapes the risk of cancer at different ages.Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/78/12/3386/F1.large.jpg Cancer Res; 78(12); 3386-96. ©2018 AACR.

Underlying Causes and Therapeutic Targeting of the Inflammatory Tumor Microenvironment.

Historically, the link between chronic inflammation and cancer has long been speculated. Only more recently, pre-clinical and epidemiologic data as well as clinical evidence all point to the role of the tumor microenvironment as inextricably connected to the neoplastic process. The tumor microenvironment (TME), a complex mix of vasculature, inflammatory cells, and stromal cells is the essential “soil” helping to modulate tumor potential. Increasingly, evidence suggests that chronic inflammation modifies the tumor microenvironment, via a host of mechanisms, including the production of cytokines, pro-inflammatory mediators, angiogenesis, and tissue remodeling. Inflammation can be triggered by a variety of different pressures, such as carcinogen exposure, immune dysfunction, dietary habits, and obesity, as well as genetic alterations leading to oncogene activation or loss of tumor suppressors. In this review, we examine the concept of the tumor microenvironment as related to both extrinsic and intrinsic stimuli that promote chronic inflammation and in turn tumorigenesis. Understanding the common pathways inherent in an inflammatory response and the tumor microenvironment may shed light on new therapies for both primary and metastatic disease. The concept of personalized medicine has pushed the field of oncology to drill down on the genetic changes of a cancer, in the hopes of identifying individually targeted agents. Given the complexities of the tumor microenvironment, it is clear that effective oncologic therapies will necessitate targeting not only the cancer cells, but their dynamic relationship to the tumor microenvironment as well.

Industry and geographic patterns of use and emission of carcinogens in Ontario, Canada, 2011\u20132015

ObjectivesThe goal of this study was to leverage data from two environmental regulatory initiatives, Ontario’s Toxics Reduction Act (TRA) and Canada’s National Pollutant Release Inventory (NPRI), to assess their ability to monitor trends in the use and emission of carcinogens by industry sector in Ontario.MethodsData reported to the TRA and NPRI by industrial facilities in Ontario were retrieved from 2011 to 2015. Twenty-six known and suspected carcinogens were identified (IARC) and the trends in the use and emission were evaluated by industry sector. The locations of industrial facilities that used and released carcinogens were mapped by Public Health Unit (PHU).ResultsChemical manufacturing and primary metal manufacturing sectors accounted for 84% of all reported industrial use of carcinogens during the period 2011–2015. The largest source of carcinogen emissions came from facilities in the primary metal manufacturing and paper manufacturing sectors. The largest number of industrial facilities that reported the use and release of carcinogens were located in the City of Toronto and Peel Region PHUs. Overall, the use of carcinogens across all sectors appeared to decrease by 8%, while emissions increased by about 2%.ConclusionThe results of this study show the need to reduce the use and emission of select carcinogens in priority industry sectors. Environmental reporting programs, such as the TRA and NPRI, can serve as important tools in cancer prevention by tracking potential carcinogen exposures in the environment and in the workplace.

A randomized clinical trial of counseling and nicotine replacement therapy for treatment of African American non-daily smokers: Design, accrual, and baseline characteristics

BackgroundNon-daily smokers (NDS) who smoke on some but not all days are a growing subset of United States (US) tobacco users. Racial/ethnic minorities are more likely to be NDS. African American NDS have strikingly high levels of nicotine and carcinogen exposure, making treatment of this high risk group a priority. MethodsThe current study is one of three ongoing federally-funded clinical trials of NDS and, to our knowledge the only RCT focused on racial/ethnic minority NDS. The design has been guided by input from Patient and Stakeholder Advisory Panels who helped develop the research questions, design the intervention, and select the outcomes. The objective is to compare the effectiveness of smoking cessation counseling alone (C) or smoking cessation counseling plus participant's choice of nicotine replacement therapy (NRT; C + NRT) for African American NDS. Two-hundred seventy-eight African American NDS will be randomized in a 2:1 fashion to C + NRT or C. All participants receive five sessions of smoking cessation counseling; those randomized to C + NRT receive their choice of nicotine gum, patch, and/or lozenge. Treatment in both groups lasts for 12 weeks. We hypothesize that C + NRT will be more effective than C on the primary outcome of biochemically-confirmed abstinence from smoking at week 12. Secondary aims will compare C + NRT and C on patient- and provider-desired outcomes including abstinence from smoking at week 26, change in biochemically-verified nicotine and carcinogen exposure, days abstinent, and treatment process measures (e.g., NRT use and side effects). Predictors of abstinence will also be explored. DiscussionFindings will illuminate effective treatment options for African American NDS and contribute to development of evidence-based guidelines for treating the 8.9 million US adult NDS for whom no guidelines currently exist.TRIAL REGISTRATION NUMBER: ClinicalTrials.gov: NCT02244918

Occupational exposures and cancer: a review of agents and relative risk estimates

ObjectivesThe contribution of occupational exposures to the cancer burden can be estimated using population-attributable fractions, which is of great importance for policy making. This paper reviews occupational carcinogens, and presents...