Malignant Lung Tissues Research Articles

Uncovering Growth-Suppressive MicroRNAs in Lung Cancer

MicroRNA (miRNA) expression profiles improve classification, diagnosis, and prognostic information of malignancies, including lung cancer. This study uncovered unique growth-suppressive miRNAs in lung cancer. miRNA arrays were done on normal lung tissues and adenocarcinomas from wild-type and proteasome degradation-resistant cyclin E transgenic mice to reveal repressed miRNAs in lung cancer. Real-time and semiquantitative reverse transcription-PCR as well as in situ hybridization assays validated these findings. Lung cancer cell lines were derived from each transgenic line (designated as ED-1 and ED-2 cells, respectively). Each highlighted miRNA was independently transfected into these cells. Growth-suppressive mechanisms were explored. Expression of a computationally predicted miRNA target was examined. These miRNAs were studied in a paired normal-malignant human lung tissue bank. miR-34c, miR-145, and miR-142-5p were repressed in transgenic lung cancers. Findings were confirmed by real-time and semiquantitative reverse transcription-PCR as well as in situ hybridization assays. Similar miRNA profiles occurred in human normal versus malignant lung tissues. Individual overexpression of miR-34c, miR-145, and miR-142-5p in ED-1 and ED-2 cells markedly repressed cell growth. Anti-miR cotransfections antagonized this inhibition. The miR-34c target, cyclin E, was repressed by miR-34c transfection and provided a mechanism for observed growth suppression. miR-34c, miR-145, and miR-142-5p were repressed in murine and human lung cancers. Transfection of each miRNA significantly repressed lung cancer cell growth. Thus, these miRNAs were growth suppressive and are proposed to exert antineoplastic effects in the lung.

Identification of the novel KI Polyomavirus in paranasal and lung tissues

KI is a novel polyomavirus identified in the respiratory secretions of children with acute respiratory symptoms. Whether this reflects a causal role of the virus in the human respiratory disease remains to be established. To investigate the presence of KIV in the respiratory tissue, we examined 20 fresh lung cancer specimens and surrounding normal tissue along with one paranasal and one lung biopsy from two transplanted children. KIV-VP1 gene was detected in 9/20 lung cancer patients and 2/2 transplanted patients. However, amplification of the sequence coding for the C-terminal part of the early region of KIV performed on the 11 positive cases was successful only in two malignant lung tissues, one surrounding normal tissue, and 1/2 biopsies tested. Phylogenetic analysis performed on the early region of KIV (including the four Italian isolates), BKV and JCV revealed the presence of three distinct clades. Within the KIV clade two sub-clades were observed. A sub-clade A containing the four Italian strains, and a sub-clade B comprising the Swedish and Australian isolates. Interestingly, the two Italian strains identified in normal tissue clustered together, whereas those detected in malignant tissue fell outside this cluster. In vitro studies are needed to investigate the transforming potential of KIV strains.

Abstract A135: Uncovering tumor suppressive and oncogenic microRNAs in lung cancer

Abstract A135 MicroRNAs (miRNAs) are noncoding small RNAs that regulate gene expression. Expression profiles of miRNAs are useful to improve classification, diagnosis, and prognostic information of specific human malignancies, including lung cancer. We sought to uncover miRNAs preferentially repressed or over-expressed in pre-malignant and malignant lung lesions in recently described transgenic cyclin E mice. These transgenic mice expressed in the lung under control of the human surfactant C promoter wild-type or proteasome degradation-resistant cyclin E species. These mice developed pulmonary dysplasia and adenocarcinoma, recapitulating pre-malignant and malignant lung lesions frequently found in lung cancer patients. Comprehensive miRNA microarray analyses were conducted using independently harvested normal and malignant lung tissues from these transgenic mice. A cluster of miRNAs was preferentially repressed in transgenic lung cancers versus normal lung tissues including: miR-34c, miR-145, miR142-5p, and other miRNAs previously associated with lung carcinogenesis. Expression profiles were independently validated by semi-quantitative and real-time polymerase chain reaction assays. In transgenic mice, single cell expression profiles were studied in pre-malignant and malignant lung lesions by in situ hybridization assays. Concordant results were obtained after analyses of paired normal-malignant human lung tissues representing each histopathologic subtype of non-small cell lung cancer. To address functional roles of repressed miRNAs, novel lung cancer cell lines were derived from murine transgenic wild-type (ED-1 cells) or proteasome-degradation resistant (ED-2 cells) cyclin E expressing lung cancers. ED-1 and ED-2 cells each caused lung adenocarcinomas to form after tail-vein injections into syngeneic FVB mice. Engineered over-expression of each basally repressed miRNA in ED-1 as well as in ED-2 cells markedly (P < 0.001) repressed cell growth. Anti-miR co-transfections antagonized these effects. A mechanism for this growth suppression was found by showing cyclin E, a predicted miR-34c bioinformatic target, was significantly repressed in miR-34c transfectants. Other miRNAs were selected for knock-down in ED-1 and ED-2 cells by virtue of their high basal expression in these cells as well as in murine transgenic lung cancers and human lung cancers versus normal lung tissues. Knock-down of candidate oncogenic miRNAs repressed lung cancer cell growth, which was antagonized by over-expression of the same miRNA. Experiments are underway in miRNA transfected ED-1 cells to assess in vivo tumorigenicity in FVB mice. Taken together, these studies uncovered candidate tumor suppressive and oncogenic miRNAs. We propose these miRNAs are molecular pharmacologic targets for lung cancer therapy and chemoprevention. Citation Information: Cancer Prev Res 2008;1(7 Suppl):A135.

Expression of cyclin D1 splice variants is differentially associated with outcome in non-small cell lung cancer patients

Real-time reverse transcription polymerase chain reaction and immunohistochemistry were used to evaluate the messenger RNA (mRNA) and protein expression levels of total cyclin D1 and its splice variants (cyclin D1a and cyclin D1b) in 102 paired malignant and nonmalignant tissues from patients with non-small cell lung cancer, respectively. The expression levels of total cyclin D1 and its splice variants were significantly up-regulated in malignant tissues than in nonmalignant tissues at both mRNA and protein levels. Although the expression levels of cyclin D1a were higher than those of cyclin D1b, the relative expression ratios of cyclin D1b mRNA between malignant and nonmalignant lung tissues were obviously higher than those of cyclin D1a mRNA. Analysis of variance showed that cyclin D1b mRNA expression was significantly associated with the histologic grade, lymph node metastasis, distant metastasis, and tumor stage of patients, whereas cyclin D1a mRNA expression was not related to clinicopathologic characteristics except sex. Patients with cyclin D1b mRNA expression above the median value had shorter survival than those below the median value (P = .033). Similarly, cyclin D1b immunopositivity was also associated with histologic grade, and patients with immunostaining positivity for cyclin D1b showed poor survival (P = .005). Multivariate analysis demonstrated that cyclin D1b immunopositivity was an independent risk factor in survival of patients with non-small cell lung cancer (P = .018). Our data show that cyclin D1b, rather than canonical cyclin D1a, might contribute to the development of non-small cell lung cancer. Cyclin D1b would be a better prognostic indicator for non-small cell lung cancer as compared to total cyclin D1 or cyclin D1a.

Increased Expression of Id Family Proteins in Small Cell Lung Cancer and its Prognostic Significance

To study the molecular pathology of human small cell lung cancer (SCLC), molecular biology approaches were used to identify genes involved in malignant progression of the cancer cells. Microquantity differential display was used initially to identify genes expressed differentially between normal and malignant cell lines. The differences were verified by Western blot. Immunohistochemical analysis was done on paired normal and malignant lung tissues and on tissues taken by biopsy to assess the expression status of candidate genes and their prognostic significance. Inhibitor of DNA/differentiation (Id)1 gene was up-regulated in SCLC cells. Levels of Id1 in 8 of 10 cell lines were increased by 1.7- to 21.4-fold when compared with the benign cells. A similar increase was also found in levels of Id2 and Id3. On 26 pairs of lung tissues, all four Id proteins were significantly (Wilcoxon Signed Rank Test, P < 0.001-0.005) overexpressed in cytoplasm of the malignant cells. In nuclei of SCLC cells, Id1 expression was significantly reduced, whereas the levels of Id2, Id3, and Id4 were significantly (Wilcoxon Signed Rank Test, P < 0.001) increased. Immunohistochemical staining on biopsy specimens showed that the increased expression of Id2 in cytoplasm of cancer cells, not the other three proteins, was significantly associated with the increased survival of SCLC patients. Changed expression profiles of Id proteins may play important roles in malignant progression of SCLC, and the increased Id2 in cytoplasm is a novel prognostic factor to predict the patient outcomes.

Expression of Leukemia/Lymphoma Related Factor (LRF) in Human Breast Carcinoma and Comparison with other Cancers

Leukemia/lymphoma‐related factor (LRF), a member of the POK family, has a vital function in cellular transformation. Apart from its role in lymphomas, very little is known about its expression in most solid tumors, including breast carcinoma. In this study, LRF mRNA transcripts and protein expression was quantified in benign and malignant breast, colon, liver, lung, renal, and mesothelioma tissues by performing RT‐PCR, Western blot, immunohistochemistry, and immunocytochemistry. All malignant breast tissues expressed significantly more LRF mRNA transcripts and protein than the benign breast tissues. LRF was expressed more strongly in fibroadenoma, invasive lobular carcinoma and ductal carcinoma in situ than in other breast diseases. The ducts in ductal hyperplasia tissues of the breast were weakly positive. In general, a significantly higher expression of LRF was seen in malignant tissue than in the corresponding benign or normal tissue. LRF expression in the carcinoma of the colon, liver, and lung was higher than in the corresponding normal or benign tissues. A strong expression of LRF was observed in normal prostate but not in prostate cancer. These data suggest that LRF could be a marker for malignancy or proliferation of the cancers.(Supported by the Claire Booth Luce Fellowship to A.A.)

Malignant and non-malignant lung tissue areas are differentially populated by natural killer cells and regulatory T cells in non-small cell lung cancer

Even though the lung represents a special immune compartment with the capacity of a high inflammatory response, ineffective anti-tumour immunity is common in lung-associated malignancies. We asked whether a differential composition of the immune cell infiltrate in malignant (MLTAs) and non-malignant lung tissue areas (N-MLTAs) exists and might potentially contribute to this effect. We performed a comparative analysis of immune cells residing in MLTAs and N-MLTAs of non-small cell lung cancer (NSCLC) patients. To this end, we used immunophenotyping and functional analyses on directly isolated immune cells and tissue arrays on archived paraffin-embedded specimens. A strong T cell infiltration was prominent in both tissue compartments whereas CD4(+)CD25(+)CD127(-) T regulatory cells were present in MLTAs only. Nonetheless, concurrent functional ex vivo T cell analyses revealed no significant difference between T cells of MLTA and N-MLTA, suggesting that tumour-infiltrating T cells were not functionally impaired. Interestingly, T cell infiltration was less pronounced in specimens with a high neutrophilic infiltrate. NK cell infiltration was strikingly heterogenous between MLTA and N-MLTA. While NK cells were almost absent in the malignant tissue regions, non-malignant counterparts were selectively populated by NK cells and those NK cells showed strong cytotoxic activity ex vivo. We report that malignant and non-malignant tissue areas in NSCLC are selectively infiltrated by certain immune cell types with NK cells being displaced from the tumour tissue. These phenomena have important implications for tumour immunology of NSCLC and should be considered for the development of future immunologic intervention therapies.

Indoleamine 2,3-dioxygenase (IDO) expression in lung cancer

BACKGROUND: The expression of indoleamine 2,3-dioxygenase (IDO) by tumor cells has been considered as a major tumor immune escape mechanism. The aim of this study was to investigate the expression of IDO in lung cancer cell lines as well as in surgically resected lung cancer specimens comparing the latter, to the expression in autologous sam-ples from the corresponding non malignant lung tissue. Correlations of IDO expression with clinicopathological parameters of the disease were performed.METHODS: Nine human lung cancer cell lines and 28 patients with various types of primary lung cancer were enrolled in the study. IDO expression was determined by quantitative real-time PCR using a sample of lung hamartoma as reference.RESULTS: IDO expression was detected in all but 3 patients’ tumor samples, in all but 4 autologous non malignant lung tissues and in 3 out of the 9 cell lines that were exam-ined. The relative expression of IDO in lung cancer cell lines (4.7±11.1) was significantly lower than that of all patients’ tumor samples (p=0.006) as well as than that of the autologous non affected lung tissues (p=0.027). No statistically significant differences were noted between ADC and SCC regarding either the tumor samples or the autologous non affected samples. No significant correlations between IDO expression and clinico-pathological parameters were found.CONCLUSION: Direct evidence is provided demonstrating that IDO mRNA can be constitutively expressed by lung cancer cells. The higher IDO expression observed in patients’ samples can be attributed to the production of the enzyme by other cells recruited in the tumor microenvironment and the peritumoral lung area and/or to its induction by soluble factors of tumor origin.

Identification and Validation of Differences in Protein Levels in Normal, Premalignant, and Malignant Lung Cells and Tissues Using High-Throughput Western Array and Immunohistochemistry

The identification of proteins, which exhibit different levels in normal, premalignant, and malignant lung cells, could improve early diagnosis and intervention. We compared the levels of proteins in normal human bronchial epithelial (NHBE) and tumorigenic HBE cells (1170-I) by high-throughput immunoblotting (PowerBlot Western Array) using 800 monoclonal antibodies. This analysis revealed that 87 proteins increased by >2-fold, and 45 proteins decreased by >2-fold, in 1170-I compared with NHBE cells. These proteins are involved in DNA synthesis and repair, cell cycle regulation, RNA transcription and degradation, translation, differentiation, angiogenesis, apoptosis, cell adhesion, cytoskeleton and cell motility, and the phosphatidylinositol 3-kinase signaling pathway. Conventional Western blotting using lysates of normal, immortalized, transformed, and tumorigenic HBEs and non-small cell lung cancer cell lines confirmed some of these changes. The expression of several of these proteins has been then analyzed by immunohistochemistry in tissue microarrays containing 323 samples, including normal bronchial epithelium, hyperplasia, squamous metaplasia, dysplasias, squamous cell carcinomas, atypical adenomatous hyperplasia, and adenocarcinomas from 144 patients. The results of the immunohistochemical studies correlated with the Western blotting findings and showed gradual increases (caspase-8, signal transducers and activators of transcription 5, and p70s6K) or decrease (E-cadherin) in levels with tumor progression. These results indicate that the changes in proteins detected in this study may occur early in lung carcinogenesis and persist in lung cancer. In addition, some of the proteins detected by this approach may be novel biomarkers for early detection of lung cancer and novel targets for chemoprevention or therapy.

059 Différentiel expression of the kallikrein 5 and kallikrein 7 gènes in non-small cell lung cancers subtypes

Human tissue kallikreins (hKs) are secreted serine protéases with diverse expression patterns and physiological rôles. Several members of the kallikrein gène (KLK) family, which now includes 15 gènes, are differentially expressed in various cancers and some émerge as useful diagnostic/ prognostic markers for hormone dépendent cancers. Although clinically effective non-invasive meth-ods for early détection and screening of lung cancer are lacking, analysis of kallikrein gène expression has never been performed in lung. The aim of this study was to investigate KLK5 and KLK7 gene expression in non small-cell lung cancers (NSCLC). We used both RT-PCR and Western blotting to analyse KLK5 and KLK7 gène expression and their respective products (hK5 and hK7) in lung. Gène expression was then quantified by real-time PCR in matched malignant and nonmalignant lung tissue samples obtained from 56 patients with NSCLC and results were compared to clinico-pathological parameters. We detected for the first time the expression of both KLK5 and KLK7 gènes and their protein products (hK5 and hK7) in tumoral and nontumoral lung tissues. A significant increase in KLK5 expression was observed in squamous cell carcinoma (P = 0.02) compared to matched nontumoral tissue, whereas KLK7 expression was decreased in adenocarcinoma (P = 0.003). In non-cancerous tissues, multivariate analysis revealed a négative corrélation between KLK5 expression and the metastatic status of the paired tumors (P = 0.05) and, a positive corrélation between KLK7 expression and tumor differentiation (P = 0.02). In cancerous tissues, KLK5 and KLK7 expressions were mainly associated with squamous cell carcinoma (P = 0.04 and P = 0.01, respectively). Fur-thermore, KLK7 expression correlated positively with differentiation (P = 0.03) and negatively with adenocarcinoma histotype (P = 0.003). Our results demonstrate that KLK5 and KLK7 are differentially expressed in NSCLC subtypes and indicate that the variability in KLK5 and KLK7 expressions hâve to be taken into account not only to better understand the lung tumorogenesis but also for clinical purposes.

Involvement of multiple developmental genes on chromosome 1p in lung tumorigenesis

Lung cancer is the leading cause of cancer death in North America. Despite advances in lung cancer treatment, the overall 5 year survival rate for those diagnosed with the disease is bleak presumably due to the late stage of diagnosis. Owing to the difficulty of early detection, preneoplastic specimens are rare. However, studying both preinvasive and invasive stages of disease is necessary to fully understand lung cancer progression. Aberration of chromosome arm 1p is common in lung and other cancers. In this study, we used a genomic array with complete tiling coverage of 1p to profile preinvasive and invasive squamous non-small cell lung carcinoma samples. With this technology, multiple novel submegabase alterations were identified. Three of the 1p alterations harbored genes belonging to gene families known to be involved in cancer development through either the Wnt or the Notch developmental pathways. Our finding of a 0.4 Mb amplified region at 1p36.12 containing WNT4 in preinvasive lung cancer, coupled with the identification of three additional alterations in invasive tumors that also contain genes related to the Notch and Wnt pathways, strongly suggests an intricate role of these pathways in early and late stages of lung cancer development. Furthermore, ectopic expression of DVL1, LRP8 and Notch2 in malignant lung tissue validates the biological impact of these genetic alterations. Importantly, this implication of pathways known only to be activated in fetal lung development lends support to the proposed model of lung cancer ontology whereby tumors arise from dysregulated pleuripotent stem cells.

Expression pattern of the novel gene EG-1 in cancer.

We recently discovered a novel gene responsive to tumor-conditioned media: endothelial-derived gene 1 (EG-1). Its transcript has been shown to be present in epithelial cells, as well as in endothelial cells. In this study, we examined the levels of EG-1 protein expression in breast, colon, prostate, and lung cancers, which constitute the four most common solid malignancies in the United States. Polyclonal antibodies were generated that recognize the EG-1 peptide. These antibodies were used in immunoblot analysis, as well as immunohistochemistry of multiple human clinical specimens of cancer. In immunoblots of whole cell lysates, EG-1 antibodies revealed the presence of a 22-kDa peptide. Immunohistochemistry of breast, colon, and prostate specimens showed higher levels of EG-1 peptides in cancer tissues, in comparison with their benign counterparts. However, EG-1 expression was minimal in both benign and malignant lung tissues. Here, we demonstrated that the expression of EG-1 is elevated in cancerous in comparison to benign epithelial cells, as seen in immunohistochemistry of human pathological specimens. These observations collectively support the hypothesis that the novel gene EG-1 is associated with the malignant phenotype of the common epithelial-derived cancers of the breast, colon, and prostate.

In vivo expression of p53 and Bcl-2 and their role in programmed cell death in premalignant and malignant lung lesions

Forty-four specimens of non-malignant and malignant human lung tissue, taken from patients with non-small cell lung cancer (NSCLC), were examined for the expression of wild-type p53, mutant p53, and bcl-2 and the occurrence of programmed cell death (apoptosis). Wild-type p53 expression peaked in peritumoral and metaplastic samples, whereas mutant p53, bcl-2 and apoptosis were first detected in metaplasia and increased with progression to carcinoma. Bcl-2 positive samples had lower levels of apoptosis than bcl-2 negative samples and was independent of wild-type or mutant p53 expression. These results suggest that the over-expression of wild-type p53 may be an early cellular response to an alteration in normal cellular homeostasis. The ensuing increase in apoptosis appears to be relatively independent of mutant or wild-type p53 expression, but does not occur in cells expressing bcl-2.

Simultaneous Quantitation of Topoisomerase II α and β Isoform mRNAs in Lung Tumor Cells and Normal and Malignant Lung Tissue

Certain drugs used in the treatment of lung cancer and other human malignancies are cytotoxic because of their ability to interact with the two isoforms of topoisomerase II (topo II), topo IIα and topo IIβ. As part of an effort to evaluate the contribution of topo II alterations to drug sensitivity and resistance in lung cancer, we have developed a semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) assay to measure levels of topo II α and β mRNAs simultaneously using a single pair of primers with sequences common to both isoforms. The PCR products derived from the topo II α and β mRNAs are both 446 bp but have different electrophoretic mobilities in a nondenaturing polyacrylamide gel, allowing sensitive, rapid quantitation when the products are radiolabeled with [35S]-dATP. Using this RT-PCR method, poly(A+) RNA from 13 non-small cell lung cancer (NSCLC) cell lines was analyzed. The results obtained indicated that the cell lines express a wide range of topo IIα mRNA levels (12-fold) and topo IIβ mRNA levels (5.5-fold). Tumor and normal lung tissues from 25 patients with NSCLC were also examined. In the tumor samples, the levels of the topo II α and β mRNAs were similar. However, mean topo IIα mRNA levels in the tumors were approximately 7-fold higher than those of the paired normal lung tissues. In contrast, topo IIβ mRNA levels were similar in both tumor and normal lung. Topo II α and β mRNA levels were both significantly lower in the squamous cell tumors than in the adenocarcinoma samples. Topo IIβ mRNA levels in the squamous cell tumors were also significantly lower than those in paired normal lung tissue. The RT-PCR method described is reliable and convenient, and for the first time, makes the rapid simultaneous direct comparison of topo IIα and topo IIβ mRNA levels feasible in large numbers of clinical samples.

Trace element load in cancer and normal lung tissue

Samples of malignant and benign human lung tissues were analysed by two complementary methods, i.e., particle induced X-ray emission (PIXE) and total reflection X-ray fluorescence (TRXRF). The concentration of trace elements of P, S, K, Ca, Ti, Cr, Mn, Fe, Cu, Zn, Se, Sr, Hg and Pb was determined in squamous cancer of lung tissue from 65 people and in the benign lung tumour tissue from 5 people. Several elements shows enhancement in cancerous lung tissue of women in comparison to men, i.e., titanium show maximum enhancement by 48% followed by Cr (20%) and Mn (36%). At the same time trace element concentration of Sr and Pb are declaimed by 30% and 20% in women population. Physical basis of used analytical methods, experimental set-up and the procedure of sample preparation are described.

Distribution of peptidyl-glycine alpha-amidating mono-oxygenase (PAM) enzymes in normal human lung and in lung epithelial tumors.

C-terminal alpha-amidation is a post-translational modification necessary for the biological activity of many regulatory peptides produced in the respiratory tract. This modification is a two-step process catalyzed by two separate enzyme activities, both derived from the peptidyl-glycine alpha-amidating mono-oxygenase (PAM) precursor. The distribution of these two enzymes, peptidyl-glycine alpha-hydroxylating monoxygenase (PHM) and peptidyl-alpha-hydroxyglycine a amidating lyase (PAL), was studied in the normal lung and in lung tumors using immunocytochemical methods and in situ hybridization. In normal lung the enzymes were located in some cells of the airway epithelium and glands, the endothelium of blood vessels, some chondrocytes of the bronchial cartilage, the alveolar macrophages, smooth muscle cells, neurons of the intrinsic ganglia, and in myelinated nerves. A total of 24 lung tumors of seven different histological types were studied. All cases contained PAM-immunoreactive cells with various patterns of distribution. All immunoreactive cells were positive for the PHM antiserum but only some of them for the PAL antiserum. The distribution of PAM co-localizes with some other previously described amidated peptides, suggesting that amidation is an important physiological process taking place in the normal and malignant human lung tissue.

LACK OF TISSUE-SPECIFICITY OF MUCIN MARKERS IN A LUNG-CANCER MODEL - BIOCHEMICAL APPROACH

Mucin-associated epitopes are recognized by monoclonal antibodies in the immunometric assays used for the diagnosis and monitoring of cancer. The recently developed new assays measure mucins as tumor markers, assuming that each mucin is associated with a particular tumor site, i.e. CA 15.3 and MCA with breast cancer, CA 125 with ovarian cancer, CA 19.9 and CA 195 with colon and pancreatic cancer. These associations are based on the frequency and the intensity of expression of the single markers for a certain organ. However, this theoretical organ specificity is not absolute, since the mucins are expressed also by tumors other than those mentioned above and they may also be present in inflammatory conditions and in normal tissues. These observations were confirmed by the present study, which used an experimental model consisting of a pool of 20 lung tissue samples (10 normal and 10 cancer). The tissue concentrations of the mucins MCA, CA 15.3, CA 125, CA 19.9, and of the glycoprotein CEA, were measured both in malignant tissue samples and in their normal counterparts. The marker levels were detected by immunometric assays in mucin fractions separated from the tissue extract by chromatographic methods. The comparison of the chromatographic profiles and the evaluation of the mucin levels in normal and malignant lung tissue specimens confirmed the absence of tissue specificity of these biochemical parameters. Recent developments in molecular biology and the discovery of genes coding for several apomucins may open new perspectives towards the understanding of the mechanisms regulating mucin pathways.

Genetics of lung cancer: implications for early detection and prevention.

Human lung cancer is a complex disease originating in different cell types. In general, tumorigenesis in the lung appears to involve a complex series of molecular, genetic, and histopathologic events, leading to the transformation of normal cells to malignant cells. It is generally believed that it is the accumulation of biologic changes, not necessarily the order in which they occur, that could best describe the transformation of cells. Our knowledge of genetic changes during tumorigenesis in the lung is limited and comes from studies involving animal models and frankly malignant lung tissues. Such studies can be questioned as to their applicability in primary prevention. A vast majority of genetic changes occurring during the preclinical stage are yet to be documented. Study of these preclinical changes would offer a better opportunity for early detection and prevention. Oncogenes and tumor suppressor genes have potential for use as biomarkers in identifying high-risk groups and in early detection. The interaction between nature and nurture is nowhere as evident as in cancers of the lung, which are influenced by environmental exposure, familial inheritance, and metabolic phenotypes. The gene-environment interaction provides several targets for prevention and early detection. Biomarkers, if validated for specificity, sensitivity, and predictive values, could act as markers of exposure, susceptibility, progression, and disease. Validation of these markers, however, would require well-characterized, prospectively collected samples, representing normal, premalignant, malignant, invasive, and metastatic conditions. The NCI has started an initiative for a tissue bank of prostate, bladder, colorectum, head and neck, and lung cancers. Clinical and epidemiological information are also collected for each sample. Study of genetic and molecular changes in collected samples should shed light on the involvement of these changes in the transformation of normal to malignant tissues.

Peripheral blood immune responses to surgically induced lung tissue injury.

Surgically induced lung tissue trauma reveals an immune response in peripheral blood, the degree of which could depend on extent and severity of the organ trauma and the presence of lung malignancies. The study included 34 patients who underwent elective thoracic surgery because of benign and malignant lung tissue diseases. Flow-cytometric phenotyping of lymphocyte subsets shows a clear shift to reduced immunocompetent cells after lung tissue injury. Patients with lung tumors reveal a postoperative activation of the macrophage system as indicated by increased plasma levels of neopterin. Increased levels of soluble interleukin-2 receptor in the plasma of these patients may be the result of cellular shedding of the receptor. Lung tissue trauma is also followed by reduced immunoglobulin levels which are most pronounced in the presence of lung malignancies. These results suggest that lung tissue injury leads to postoperative immunosuppression especially in tumor patients.

Reciprocal expression of pleiotrophin and midkine in normal versus malignant lung tissues.

Abundant evidence suggests that growth factors are important mediators of non-small cell lung cancer (NSCLC) growth. Although multiple growth factors have been found to be produced by NSCLC tissues, little is known about possible differences in growth factor expression between malignant and adjacent normal lung tissues. Variation in growth factor expression between normal and malignant lung tissues could be potentially useful diagnostically and therapeutically. In studies reported here, the expression of the angiogenic growth factor pleiotrophin (PTN) and homolog midkine (MK) was assessed in resected normal and malignant lung tissues. Primers specific for the two growth factors were used to amplify reverse transcriptase-produced DNA copies of RNA transcripts harvested from the tissues. This analysis revealed that all normal lung tissues examined (n = 17) expressed PTN but only two expressed MK. Conversely, all of the resected lung cancers (n = 20) expressed MK but only one expressed PTN. These results demonstrated a striking reciprocal expression pattern of MK and PTN in normal versus malignant lung tissue.