High tumor cell platelet-derived growth factor receptor beta expression is associated with shorter survival in malignant pleural epithelioid mesothelioma.

Malignant pleural mesothelioma (MPM) has a rich stromal component containing mesenchymal fibroblasts. However, the properties and interplay of MPM tumor cells and their surrounding stromal fibroblasts are poorly characterized. Our objective was to spatially profile known mesenchymal markers in both tumor cells and associated fibroblasts and correlate their expression with patient survival. The primary study cohort consisted of 74 MPM patients, including 16 patients who survived at least 60 months. We analyzed location-specific tissue expression of seven fibroblast markers in clinical samples using multiplexed fluorescence immunohistochemistry (mfIHC) and digital image analysis. Effect on survival was assessed using Cox regression analyses. The outcome measurement was all-cause mortality. Univariate analysis revealed that high expression of secreted protein acidic and cysteine rich (SPARC) and fibroblast activation protein in stromal cells was associated with shorter survival. Importantly, high expression of platelet-derived growth factor receptor beta (PDGFRB) in tumor cells, but not in stromal cells, was associated with shorter survival (hazard ratio [HR] = 1.02, p < 0.001). A multivariable survival analysis adjusted for clinical parameters and stromal mfIHC markers revealed that tumor cell PDGFRB and stromal SPARC remained independently associated with survival (HR = 1.01, 95% confidence interval [CI] = 1.00-1.03 and HR = 1.05, 95% CI = 1.00-1.11, respectively). The prognostic effect of PDGFRB was validated with an artificial intelligence-based analysis method and further externally validated in another cohort of 117 MPM patients. In external validation, high tumor cell PDGFRB expression associated with shorter survival, especially in the epithelioid subtype. Our findings suggest PDGFRB and SPARC as potential markers for risk stratification and as targets for therapy.

Lung cancer biomarker testing: perspective from Europe.

A questionnaire on biomarker testing previously used in central European countries was extended and distributed in Western and Central European countries to the pathologists participating at the Pulmonary Pathology Society meeting 26-28 June 2019 in Dubrovnik, Croatia. Each country was represented by one responder. For recent biomarkers the availability and reimbursement of diagnoses of molecular alterations in non-small cell lung carcinoma varies widely between different, also western European, countries. Reimbursement of such assessments varies widely between unavailability and payments by the health care system or even pharmaceutical companies. The support for testing from alternative sources, such as the pharmaceutical industry, is no doubt partly compensating for the lack of public health system support, but it is not a viable or long-term solution. Ideally, a structured access to testing and reimbursement should be the aim in order to provide patients with appropriate therapeutic options. As biomarker enabled therapies deliver a 50% better probability of outcome success, improved and unbiased reimbursement remains a major challenge for the future.

Reactive oxygen species-regulating proteins peroxiredoxin 2 and thioredoxin, and glyceraldehyde-3-phosphate dehydrogenase are differentially abundant in induced sputum from smokers with lung cancer or asbestos exposure.

Lung cancer is a deadly disease, typically caused by known risk factors, such as tobacco smoke and asbestos exposure. By triggering cellular oxidative stress and altering the antioxidant pathways eliminating reactive oxygen species (ROS), tobacco smoke and asbestos predispose to cancer. Despite easily recognizable high-risk individuals, lung cancer screening and its early detection are hampered by poor diagnostic tools including the absence of proper biomarkers. This study aimed to recognize potential lung cancer biomarkers using induced sputum noninvasively collected from the lungs of individuals in risk of contracting lung cancer. Study groups composed of current and former smokers, who either were significantly asbestos exposed, had lung cancer, or were unexposed and asymptomatic. Screening of potential biomarkers was performed with 52, and five differentially abundant proteins, peroxiredoxin 2 (PRDX2), thioredoxin (TXN), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), extracellular matrix protein 1 (ECM1), and protein S100 A8 (S100A8), were chosen to undergo validation, for their previously known connection with oxidative stress or cancer. Results from the validation in 123 sputa showed that PRDX2, TXN, and GAPDH were differentially abundant in sputa from individuals with lung cancer. TXN had a negative correlation with asbestos exposure, yet a positive correlation with smoking and lung cancer. Thus, tobacco smoking, asbestos exposure, and lung carcinogenesis may disturb the cellular redox state in different ways. A strong correlation was found among PRDX2, TXN, GAPDH, and S100A8, suggesting that these proteins may present a diagnostic biomarker panel to aid recognizing individuals at high risk of contracting lung cancer.

CDKN2A copy number and p16 expression in malignant pleural mesothelioma in relation to asbestos exposure.

BACKGROUND: Deletion of the CDKN2A locus is centrally involved in the development of several malignancies. In malignant pleural mesothelioma (MPM), it is one of the most frequently reported genomic alteration. MPM is strongly associated with a patients' asbestos exposure. However, the status of CDKN2A and the expression of the corresponding protein, p16, in relation to MPM patient's asbestos exposure is poorly known. Copy number alterations in 2p16, 9q33.1 and 19p13 have earlier been shown to accumulate in lung cancer in relation to asbestos exposure but their status in MPM is unclear. METHODS: We studied DNA copy numbers for CDKN2A using fluorescence in situ hybridization (FISH) and p16 expression by immunohistochemistry (IHC) in 92 MPM patients, 75 of which with known asbestos exposure status. We also studied, in MPM, copy number alterations in 2p16, 9q33.1 and 19p13 by FISH. RESULTS: We were unable to detect an association between p16 expression and pulmonary asbestos fiber count in MPM tumor cells. However, significantly more MPM patients with high pulmonary asbestos fiber count (> 1 million fibers per gram [f/g]) had stromal p16 immunoreactivity than MPM of patients with low exposure (≤ 0.5 million f/g) (51.4% vs 16.7%; p = 0.035, Chi-Square). We found that an abnormal copy number of CDKN2A in MPM tumor cells associated with a high pulmonary asbestos fiber count (p = 0.044, Fisher's Exact test, two-tailed). In contrast to our earlier findings in asbestos associated lung cancer, DNA copy number changes in 2p16, 9q33 and 19p13 were not frequent in MPM although single cases with variable copy numbers on those regions were seen. CONCLUSIONS: We found two instances where the gene locus CDKN2A or its corresponding protein expression, is associated with high asbestos exposure levels. This suggests that there may be biological differences between the mesotheliomas with high pulmonary asbestos fiber count and those with low fiber count.

Stromal Caveolin-1 and Caveolin-2 Expression in Primary Tumors and Lymph Node Metastases.

The expression of caveolin-1 (CAV1) in both tumor cell and cancer-associated fibroblasts (CAFs) has been found to correlate with tumor aggressiveness in different epithelial tumor entities, whereas less is known for caveolin-2 (CAV2). The aim of this study was to investigate the clinicopathological significance and prognostic value of stromal CAV1 and CAV2 expression in lung cancer. The expression of these two genes was investigated at protein level on a tissue microarray (TMA) consisting of 161 primary tumor samples. 50.7% of squamous cell lung cancer (SCC) tumors showed strong expression of CAV1 in the tumor-associated stromal cells, whereas only 15.1% of adenocarcinomas (AC) showed a strong CAV1 expression (p < 0.01). A strong CAV2 stromal expression was found in 46.0% of the lung tumor specimens, with no significant difference between the subtypes. Neither CAV1 nor CAV2 stromal expression was associated with any other clinicopathological factor including survival. When the stromal expression in matched primary tumors and lymph node metastases was compared, both CAV1 and CAV2 expressions were frequently found lost in the corresponding stroma of the lymph node metastasis (40.6%, p = 0.003 and 38.4%, p = 0.001, resp.). Loss of stromal CAV2 in the lymph node metastases was also significantly associated with earlier death (p = 0.011). In conclusion, in contrast to the expression patterns in the tumor tissue of lung cancer, stromal expression of CAV1 in primary tumors was not associated with clinical outcome whereas the stromal expression of especially CAV2 in the metastatic lymph nodes could be associated with lung cancer pathogenesis.

Asbestos-associated genome-wide DNA methylation changes in lung cancer.

Previous studies have revealed a robust association between exposure to asbestos and human lung cancer. Accumulating evidence has highlighted the role of epigenome deregulation in the mechanism of carcinogen-induced malignancies. We examined the impact of asbestos on DNA methylation. Our genome-wide studies (using Illumina HumanMethylation450K BeadChip) of lung cancer tissue and paired normal lung from 28 asbestos-exposed or non-exposed patients, mostly smokers, revealed distinctive DNA methylation changes. We identified a number of differentially methylated regions (DMR) and differentially variable, differentially methylated CpGs (DVMC), with individual CpGs further validated by pyrosequencing in an independent series of 91 non-small cell lung cancer and paired normal lung. We discovered and validated BEND4, ZSCAN31 and GPR135 as significantly hypermethylated in lung cancer. DMRs in genes such as RARB (FDR 1.1 × 10-19 , mean change in beta [Δ] -0.09), GPR135 (FDR 1.87 × 10-8 , mean Δ -0.09) and TPO (FDR 8.58 × 10-5 , mean Δ -0.11), and DVMCs in NPTN, NRG2, GLT25D2 and TRPC3 (all with p <0.05, t-test) were significantly associated with asbestos exposure status in exposed versus non-exposed lung tumors. Hypomethylation was characteristic to DVMCs in lung cancer tissue from asbestos-exposed subjects. When DVMCs related to asbestos or smoking were analyzed, 96% of the elements were unique to either of the exposures, consistent with the concept that the methylation changes in tumors may be specific for risk factors. In conclusion, we identified novel DNA methylation changes associated with lung tumors and asbestos exposure, suggesting that changes may be present in causal pathway from asbestos exposure to lung cancer.

Accumulation of genomic alterations in 2p16, 9q33.1 and 19p13 in lung tumours of asbestos-exposed patients.

We have previously demonstrated an association between genomic alterations in 19p13, 2p16, and 9q33.1 and asbestos exposure in patients' lung tumours. This study detected allelic imbalance (AI) in these regions in asbestos-exposed lung cancer (LC) patients' histologically normal pulmonary epithelium. We extended the analyses of tumour tissue to cover a large LC patient cohort and studied DNA copy number alteration (CNA) and AI in 19p13, 2p16, and 9q33.1 for the first time in combination. We found both CNA and AI in ≥2/3 of the regions to be significantly and dose-dependently (P < 0.001) associated with pulmonary asbestos fibre count. Twenty percent of the exposed patients' LC showed CNA in ≥2/3 of the regions, whereas none of the non-exposed patients' LC showed CNA in more than one region. AI was evident in 89% of the exposed and in only 26% of the non-exposed patients' LC. The genomic alterations in 19p13, 2p16, and 9q33.1 in compilation identified asbestos-exposed patients' lung tumours better than each of the regions alone. These alterations form the basis for the development of a combinatorial molecular assay that could be used to identify asbestos-related LC.

Peroxiredoxins and tropomyosins as plasma biomarkers for lung cancer and asbestos exposure.

The prognosis of lung cancer is poor due to late diagnosis, the lack of established screening programs, and the paucity of early biomarkers for high-risk populations. Plasma proteome analysis was used to identify novel biomarkers for diagnosing lung cancer, and to unravel the mechanisms of underlying pathogenesis. Plasma proteins obtained from asbestos-exposed lung cancer cases detected by CT screening, asbestos-exposed subjects, clinical lung cancer patients, and healthy tobacco smokers, 5-6 cases in each group, were separated by two-dimensional gel electrophoresis, and identified with tandem mass spectrometry (LC-MS/MS). Nine proteins were selected for immunological confirmation in a test or validation set of plasma samples from an additional 49 clinical lung cancer cases, 66 asbestos-exposed patients, and 107 healthy tobacco smokers. Twenty-eight unique proteins were differentially expressed between the four study groups (p<0.05). Peroxiredoxin 1 (PRX1) was detected as a novel plasma marker for lung cancer (p=0.001). We also confirmed the previously found association of serum amyloid A with lung cancer (p<0.001). High plasma levels of tropomyosin 4 (TPM4: p<0.001) and peroxiredoxins 1 and 2 (PRX2: p<0.001) correlated with asbestos exposure or a diagnosis of asbestosis. PRX1 and PRX2 exhibited an inverse correlation with tobacco smoking (p<0.001). Plasma peroxiredoxins 1 and 2, and tropomyosin 4 were shown to associate with asbestos-exposure, and peroxiredoxin 1 with lung cancer. High plasma levels of peroxiredoxin 1 may result from genetic damage caused by reactive oxygen species. This study has identified several biomarkers worthy of further investigation in lung cancer and asbestos-related diseases.

Epithelioid lesions of the serosa.

CONTEXT: Diagnosing epithelioid serosal lesions remains a challenge because numerous different processes-primary or secondary, benign or malignant-occur in body cavities, some of which are very rare. OBJECTIVES: To review the newest literature and to describe the morphologic criteria and immunohistochemical markers that are useful for distinguishing epithelioid serosal lesions. DATA SOURCES: Previously published literature concentrating on the newest research findings. Earlier reviews are principally referred to for established diagnostic criteria. CONCLUSIONS: Immunohistochemistry with a panel of antibodies has made the diagnosis of epithelioid serosal lesions very reliable. When deciding on antibodies used in differential diagnosis, it is important to consider tumor location, clinical and radiologic information, and morphologic features. Immunohistochemistry is less useful in the differential diagnosis of benign versus malignant mesothelial lesions. The diagnosis of benign versus malignant mesothelial proliferations still relies on the histologic criteria of invasion.

Similar DNA methylation pattern in lung tumours from smokers and never-smokers with second-hand tobacco smoke exposure.

Tobacco smoke causes lung cancer in smokers and in never-smokers exposed to second-hand tobacco smoke (SHS). Nonetheless, molecular mechanisms of lung cancer in SHS-exposed never-smokers are still elusive. We studied lung cancers from current smokers (n = 109), former smokers (n = 56) and never-smokers (n = 47) for promoter hypermethylation of five tumour suppressor genes--p16, RARB, RASSF1, MGMT and DAPK1--using methylation-specific polymerase chain reaction. Lung tumours from ever-smokers suggested an increased risk of p16 hypermethylation as compared to never-smokers (P = 0.073), with former smokers having the highest frequency of p16 hypermethylation (P = 0.044 versus current smokers and P = 0.009 versus never-smokers). In the never-smoking group, p16 hypermethylation was seen in lung tumours from SHS-exposed individuals (4/33; 12%) but in none of the non-exposed individuals (0/9). The overall occurrence of hypermethylation (measured both as methylation index and as number of genes affected) was similar in those ever exposed to tobacco smoke (smokers, SHS-exposed never-smokers) and differed from non-exposed never-smokers. In multivariate analysis, p16 hypermethylation was more prevalent in lung tumours from male than female patients (P = 0.018) and in squamous cell carcinomas than in adenocarcinomas (P = 0.025). Occurrence of TP53 mutation in the tumour was associated with hypermethylation of at least one gene (P = 0.027). In all, our data suggest that promoter hypermethylation pattern in SHS-exposed never-smokers resembles that observed in smokers. Association between TP53 mutation, a hallmark of smokers' lung cancer, and methylation of one or more of the lung cancer-related genes studied, provides further evidence for common tobacco smoke-related origin for both types of molecular alterations.

Integrative analysis of microRNA, mRNA and aCGH data reveals asbestos- and histology-related changes in lung cancer.

Lung cancer has the highest mortality rate of all of the cancers in the world and asbestos-related lung cancer is one of the leading occupational cancers. The identification of asbestos-related molecular changes has long been a topic of increasing research interest. The aim of this study was to identify novel asbestos-related molecular correlates by integrating miRNA expression profiling with previously obtained profiling data (aCGH and mRNA expression) from the same patient material. miRNA profiling was performed on 26 tumor and corresponding normal lung tissue samples from highly asbestos-exposed and non-exposed patients, and on eight control lung tissue samples. Data analyses on miRNA expression, and integration of miRNA and previously obtained mRNA data were performed using Chipster. A separate analysis was used to integrate miRNA and previously obtained aCGH data. Both known and new lung cancer-associated miRNAs and target genes with inverse correlation were discovered. Furthermore, DNA copy number alterations (e.g., gain at 12p13.31) were correlated with the deregulated miRNAs. Specifically, thirteen novel asbestos-related miRNAs (over-expressed: miR-148b, miR-374a, miR-24-1*, Let-7d, Let-7e, miR-199b-5p, miR-331-3p, and miR-96 and under-expressed: miR-939, miR-671-5p, miR-605, miR-1224-5p and miR-202) and inversely correlated target genes (e.g., GADD45A, LTBP1, FOSB, NCALD, CACNA2D2, MTSS1, EPB41L3) were identified. In addition, over-expression of the well known squamous cell carcinoma-associated miR-205 was linked to down-regulation of the DOK4 gene. The miRNAs/genes presented here may represent interesting targets for further investigation and could eventually have potential diagnostic implications.

Cytochrome P450-mediated pulmonary metabolism of carcinogens: regulation and cross-talk in lung carcinogenesis.

Lung cancer is strongly associated with exogenous risk factors, in particular tobacco smoking and asbestos exposure. New research data are accumulating about the regulation of the metabolism of tobacco carcinogens and the metabolic response to oxidative stress. These data provide mechanistic details about why well known risk factors cause lung cancer. The purpose of this review is to evaluate the present knowledge of the role of cytochrome P450 (CYP) enzymes in the metabolism of tobacco carcinogens and associations with tobacco and asbestos carcinogenesis. Major emphasis is placed on human data and regulatory pathways involved in CYP regulation and lung carcinogenesis. The most exciting new research findings concern cross-talk of the CYP-regulating aryl hydrocarbon receptor with other transcription factors, such as nuclear factor-erythroid 2-related factor 2, involved in the regulation of xenobiotic metabolism and antioxidant enzymes. This cross-talk between transcription factors may provide mechanistic evidence for clinically relevant issues, such as differences in lung cancers between men and women and the synergism between tobacco and asbestos as lung carcinogens.

Molecular alterations at 9q33.1 and polyploidy in asbestos-related lung cancer.

PURPOSE: Asbestos causes DNA damage and the fibers, together with tobacco smoke, have a synergistic effect on lung cancer risk. We recently identified 18 chromosomal regions that showed differences in DNA copy number between the lung tumors of asbestos-exposed and nonexposed patients. One of the previously identified asbestos-associated chromosomal regions at 9q was further analyzed for allelic imbalance and DNA copy number alterations (CNA) in the lung tumors of asbestos-exposed and nonexposed patients. In addition, the ploidy level of the tumors was studied. EXPERIMENTAL DESIGN: Allelic imbalance was analyzed at 9q31.3-34.3 with 15 microsatellite markers in 52 lung tumor samples from asbestos-exposed and nonexposed patients. CNA at 9q32-34.3 were characterized by fluorescent in situ hybridization (FISH) with six bacterial artificial chromosome probes in 95 lung tumors. The ploidy level was analyzed in 100 lung tumors with FISH using three to five centromere probes. RESULTS: Allelic imbalance at 9q31.3-q34.3 was found in all asbestos-exposed patient tumors (100%, 17 of 17) compared with 64% (14 of 22) in the nonexposed cases (P = 0.005). The most significant difference was detected at 9q33.1 (P = 0.002). FISH results showed that also CNA were more frequent at 9q33.1 in the three major histologic types of non-small-cell lung tumors of exposed patients, and the association showed a dose-dependent trend (P = 0.03). Furthermore, we detected more frequent polyploidy among the exposed (48%, 28 of 58) than among the nonexposed (29%, 12 of 42) patient tumors (P < 0.05). CONCLUSIONS: These results provide a basis for the development of a method to identify asbestos-related lung cancer on a molecular level.

Pathways affected by asbestos exposure in normal and tumour tissue of lung cancer patients.

BACKGROUND: Studies on asbestos-induced tumourigenesis have indicated the role of, e.g., reactive oxygen/nitrogen species, mitochondria, as well as NF-kappaB and MAPK signalling pathways. The exact molecular mechanisms contributing to asbestos-mediated carcinogenesis are, however, still to be characterized. METHODS: In this study, gene expression data analyses together with gene annotation data from the Gene Ontology (GO) database were utilized to identify pathways that are differentially regulated in lung and tumour tissues between asbestos-exposed and non-exposed lung cancer patients. Differentially regulated pathways were identified from gene expression data from 14 asbestos-exposed and 14 non-exposed lung cancer patients using custom-made software and Iterative Group Analysis (iGA). Western blotting was used to further characterize the findings, specifically to determine the protein levels of UBA1 and UBA7. RESULTS: Differences between asbestos-related and non-related lung tumours were detected in pathways associated with, e.g., ion transport, NF-kappaB signalling, DNA repair, as well as spliceosome and nucleosome complexes. A notable fraction of the pathways down-regulated in both normal and tumour tissue of the asbestos-exposed patients were related to protein ubiquitination, a versatile process regulating, for instance, DNA repair, cell cycle, and apoptosis, and thus being also a significant contributor of carcinogenesis. Even though UBA1 or UBA7, the early enzymes involved in protein ubiquitination and ubiquitin-like regulation of target proteins, did not underlie the exposure-related deregulation of ubiquitination, a difference was detected in the UBA1 and UBA7 levels between squamous cell carcinomas and respective normal lung tissue (p = 0.02 and p = 0.01) without regard to exposure status. CONCLUSION: Our results indicate alterations in protein ubiquitination related both to cancer type and asbestos. We present for the first time pathway analysis results on asbestos-associated lung cancer, providing important insight into the most relevant targets for future research.

Aberrations of chromosome 19 in asbestos-associated lung cancer and in asbestos-induced micronuclei of bronchial epithelial cells in vitro.

Exposure to asbestos is known to induce lung cancer, and our previous studies have suggested that specific chromosomal regions, such as 19p13, are preferentially aberrant in lung tumours of asbestos-exposed patients. Here, we further examined the association between the 19p region and exposure to asbestos using array comparative genomic hybridization and fluorescence in situ hybridization (FISH) in lung tumours and FISH characterization of asbestos-induced micronuclei (MN) in human bronchial epithelial BEAS 2B cells in vitro. We detected an increased number of 19p losses in the tumours of asbestos-exposed patients in comparison with tumours from non-exposed subjects with similar distribution of tumour histology in both groups (13/33; 39% versus 3/25; 12%, P = 0.04). In BEAS 2B cells, a 48 h exposure to crocidolite asbestos (2.0 microg/cm(2)) was found to induce centromere-negative MN-harbouring chromosomal fragments. Furthermore, an increased frequency of rare MN containing a 19p fragment was observed after the crocidolite treatment in comparison with untreated controls (6/6000 versus 1/10 000, P = 0.01). The results suggest that 19p has significance in asbestos-associated carcinogenesis and that asbestos may be capable of inducing specific chromosome aberrations.

Molecular and genetic changes in asbestos-related lung cancer.

Asbestos-exposure is associated with an increased risk of lung cancer, one of the leading causes of cancer deaths worldwide. Asbestos is known to induce DNA and chromosomal damage as well as aberrations in signalling pathways, such as the MAPK and NF-kappaB cascades, crucial for cellular homeostasis. The alterations result from both indirect effects through e.g. reactive oxygen/nitrogen species and direct mechanical disturbances of cellular constituents. This review describes the current knowledge on genomic and pathway aberrations characterizing asbestos-related lung cancer. Specific asbestos-associated molecular signatures can assist the development of early biomarkers, molecular diagnosis, and molecular targeted treatments for asbestos-exposed lung cancer patients.

Gene expression profiles in asbestos-exposed epithelial and mesothelial lung cell lines.

BACKGROUND: Asbestos has been shown to cause chromosomal damage and DNA aberrations. Exposure to asbestos causes many lung diseases e.g. asbestosis, malignant mesothelioma, and lung cancer, but the disease-related processes are still largely unknown. We exposed the human cell lines A549, Beas-2B and Met5A to crocidolite asbestos and determined time-dependent gene expression profiles by using Affymetrix arrays. The hybridization data was analyzed by using an algorithm specifically designed for clustering of short time series expression data. A canonical correlation analysis was applied to identify correlations between the cell lines, and a Gene Ontology analysis method for the identification of enriched, differentially expressed biological processes. RESULTS: We recognized a large number of previously known as well as new potential asbestos-associated genes and biological processes, and identified chromosomal regions enriched with genes potentially contributing to common responses to asbestos in these cell lines. These include genes such as the thioredoxin domain containing gene (TXNDC) and the potential tumor suppressor, BCL2/adenovirus E1B 19kD-interacting protein gene (BNIP3L), GO-terms such as "positive regulation of I-kappaB kinase/NF-kappaB cascade" and "positive regulation of transcription, DNA-dependent", and chromosomal regions such as 2p22, 9p13, and 14q21. We present the complete data sets as Additional files. CONCLUSION: This study identifies several interesting targets for further investigation in relation to asbestos-associated diseases.

Evidence of gene gene interactions in lung carcinogenesis in a large pooled analysis.

To test the hypothesis of interaction among genetic variants in increasing the individual risk of cancer, we have studied the cumulative effect on lung cancer risk of variants in three metabolic genes, CYP1A1, GSTM1 and GSTT1, which are involved in the metabolism of the tobacco smoke constituents and environmental contaminants, polycyclic aromatic hydrocarbons and of other lung carcinogens. We have selected from the Genetic Susceptibility to Environmental Carcinogens pooled analysis all the studies on lung cancer conducted after 1991 in which all variants were available. The data set includes 611 cases and 870 controls. We found a cumulative effect of the combination of the a priori 'at-risk' alleles for these genes (P for trend 0.004). The risk of lung cancer was increased with the combination of CYP1A1*2B or CYP1A1*4 alleles and the double deletion of both GSTM1 and GSTT1 up to an odds ratio (OR) of 8.25 (95% confidence interval 2.29-29.77) for the combination including CYP1A1*4; among never smokers, the latter combination was associated with an OR of 16.19 (1.90-137). Estimates did not change after adjustment by the number of cigarettes smoked and duration of smoking were consistent across ethnicities and were approximately the same for adenocarcinomas and squamous cell carcinomas. These observations from a large pooled analysis strongly suggest the existence of gene-gene interactions in lung carcinogenesis. People with rare combinations of common gene variants have a high risk of cancer and can be assimilated to subjects with highly penetrant mutations.

COX-2 is widely expressed in metaplastic epithelium in pulmonary fibrous disorders.

Idiopathic usual interstitial pneumonia/idiopathic pulmonary fibrosis (UIP/IPF) and asbestosis represent progressive and often fatal pulmonary fibrous disorders, whereas cryptogenic organizing pneumonia (COP), desquamative interstitial pneumonia (DIP), and respiratory bronchiolitis-interstitial lung disease (RB-ILD) usually are reversible or nonprogressive conditions. Prostaglandin E2 (PGE2) inhibits fibroblast proliferation and myofibroblast transition, its production depending on cyclooxygenase-2 (COX-2). In patients with UIP/IPF, levels of PGE2 and COX-2 are reduced in fibroblasts, and levels of PGE2 in bronchioalveolar lavage fluid may be lowered. We analyzed the immunohistochemical expression of COX-2 in UIP/IPF, asbestosis, COP, DIP, and RB-ILD. Our results show that the metaplastic epithelium in UIP/IPF, asbestosis, and COP is widely COX-2+, whereas COX-2 positivity is scant in DIP and RB-ILD. The mesenchymal cells remained negative. Our results suggest that irrespective of the underlying disease, lung injury that causes extensive fibrosis induces wide expression of COX-2 in the regenerating metaplastic epithelium.

Copy number gains on 5p15, 6p11-q11, 7p12, and 8q24 are rare in sputum cells of individuals at high risk of lung cancer.

UNLABELLED: Lung cancer specimens display recurrent copy number aberrations in distinguished chromosomal regions as compared with normal lung cells. Such alterations have been utilized in design of fluorescence in situ hybridization (FISH) probe sets in attempts to improve the cytological diagnosis of lung cancer. One of such probe sets, LAVysion, detects copy number changes in the centromeric region of chromosome 6 (CEP6), and regions 5p15, 8q24, and 7p12, often gained in lung cancer. METHODS: We evaluated the feasibility of the LAVysion multi-color probe set in detection of individuals at high risk of lung cancer by applying the FISH probe set on smears prepared of induced sputa obtained from 20 lung cancer patients, 43 asbestos-exposed workers, 21 heavy tobacco smokers, and 15 healthy never-smokers. The hybridized sputum smears were examined using fluorescence microscopy and the number of signals in epithelial cells was examined throughout the hybridized area. Additionally, we review here the previous studies using LAVysion probe set. RESULTS: The FISH probe set was slightly more sensitive than cytology alone in detecting lung cancer. No significant differences in copy number gain were found between high-risk individuals and healthy never-smokers. The proportions of individuals with copy number gains in sputa among the lung cancer patients, asbestos-exposed workers, tobacco smokers, and never-smokers were 50, 20, 12, and 27%, respectively, when three or more cells with a copy number gain detected by at least two different probes was used as the cut-off point. In comparison, the sensitivity of cytology in detecting lung cancer was 44%. In the lung cancer patients the number of abnormal cells by FISH correlated well with the cytologic atypia class (Spearman rank correlation coefficient 0.77, p<0.01). Using multivariant variance analysis, gains in CEP6, 5p15, 8q24 and 7p12 were not associated with smoking or asbestos exposure. CONCLUSIONS: FISH did not significantly exceed the sensitivity of sputum cytology in finding lung cancers. Significant differences were not found between sputa of asbestos-exposed individuals, heavy-smokers and never-smokers. More sensitive methods are needed for the follow-up of populations at high risk of contracting lung cancer.