Molecular Pathogenesis Of Lung Cancer Research Articles

Tissue specific epigenetic silencing of the distinct tumor suppressor genes in lung cancer

The role of aberrant methylation of the cytosine-guanine dinucleotide islands in the promoter region of tumor suppressor genes in lung cancer development is increasingly recognized. DNAs extracted from cancer tissue biopsies of 40 patients with lung cancer were used. Methylated sites of tumor suppressor geneswere examined by bisulfate treatment and methylation-specific PCR method. Strip Assay was used to access the promoter profiles of cytosine-guanine dinucleotide islands. Some of the tumor specimens evaluated showed fully inactive methylated pattern profiles for the tumor suppressor genes that were studied and some showed partially inactive methylated patterns. In lung cancers, the prevalence of SFRP2, p16, DAPK1, HIC1, MGMT promoter methylation status were 11/40(27.5), 8/40(20), 30/40(75), 16/40(40%) and 10/40(25%), respectively. DAPK1 gene inactivation was seen in all (5/5, 100%) the adenocarcinoma type of tumors and was fully hypermethylated in 13/19 squamous cell type (68.4%), 6/9 SCLC (66.7%), and 4/7 malign epithelial (57.1%) tumors. Our results confirm the importance of methylation in the molecular pathogenesis of lung cancer with the majority of tumors having one or more tumor suppressor genes islands methylated for some tumor suppressor genes promoters. Key words: Lung carcinoma, tumor suppressor genes, epigenetic alterations.

Abstract SY33-03: Role of telomerase in normal and neoplastic stem cells

Abstract We have used hTERT (the catalytic component of telomerase) to immortalize a variety of human cell types (sometimes in combination with Cdk4 to bypass cell culture stress). Cell types immortalized include skin keratinocytes and fibroblasts, muscle satellite cells, breast epithelial and stromal cells, corneal epithelial cells and fibroblasts (keratocytes), and human colonic epithelial cells. In addition, we have immortalized human bronchial epithelial cells (HBEC) and have determined that these cells can terminally differentiate into both central and peripheral lung cell types. These immortalized HBECs have been used to study the molecular pathogenesis of lung cancer by stable “knock down” of TP53 and also by over-expression of C-myc and mutant K-rasV12. When these cells containing multiple genetic alterations are introduced into immunosuppressed mice, the experimentally transformed cells make tumors that represent several distinct histological types. This suggests that these immortalized and transformed HBECs have bronchiolar-alveolar stem-like characteristics that can differentiate into multiple lineages. Stem cells are defined by both their ability to make more stem cells (self renewal) and their ability to produce cells that can differentiate. Experimentally immortalized human bronchial epithelial cells fulfill this definition of normal stem cells by continuous self renewal and by retaining the capability of differentiating into several cell types. Since experimentally transformed cells make lung tumors representing several major lineages, this is also an indication that the HBECs are derived from a multi-potent lung stem cell. Similar to normal stem cells, cancer (initiating) stem cells also have the ability to self-renew as well as undergo differentiation to give rise to phenotypically diverse types of cancer cells. There is mounting evidence that these rare cancer stem cells may be multidrug resistant and responsible for tumor relapse and metastasis. Targeted cancer therapeutic approaches seek to identify pathways that are more tumor specific, resulting in fewer side-effects and that may produce long-term durable responses. Telomerase is a novel cancer therapeutic target since it is activated in the vast majority of human cancers and telomeres of almost all human tumor cells are maintained at short but stable lengths. In addition, telomerase is not expressed or is expressed at levels that do not fully maintain telomeres in normal tissues, and telomeres are generally longer in normal stem cells compared to cancer cells. This potentially provides a therapeutic advantage for targeting telomerase over approaches that affect both normal and cancer cells equally. We have previously reported that telomerase positive cancer cells that are experimentally induced to undergo quiescence, down regulate telomerase. As part of our anti-telomerase therapeutics program, we have addressed the following questions: Are putative cancer stem cell populations quiescent and do they have short or long telomeres? While the molecular characteristics of cancer stem cells are not completely defined and subject to some controversies, we have isolated and examined cells expressing these cancer stem cell putative markers reported for breast, brain, prostate, pancreas, and lung cancer. In each case we have observed that purified cancer stem cell populations are positive for telomerase activity, indicating they are not quiescent. In addition, a telomerase inhibitor currently being tested in clinical trials robustly inhibits the activity of telomerase in these sorted sub-populations of putative cancer stem cells as well as the mass population of cancer cells. Finally, we have observed that cancer stem cells have short telomeres in comparison to normal stem cells. These findings support the idea that there may be a therapeutic window of opportunity to target cancer stem cells by inhibiting telomerase, thus driving telomeres progressively shorter leading to cancer stem cell death, potentially without irreversible damage to normal stem cells. Cancer remains a major cause of death in spite of substantial progress towards understanding the molecular basis of many types of cancers. The discovery of new drugs is a high priority, and telomerase inhibitors have the potential to act by a novel mechanism that will provide new options for cancer therapy. A review of ongoing anti-telomerase clinical trials will be presented. In summary, telomerase inhibitors might not only directly limit or stop the growth of human tumors including cancer stem cells, but might also act in an additive or synergistic fashion with existing therapies to amplify their effectiveness. Citation Format: Jerry W. Shay. Role of telomerase in normal and neoplastic stem cells [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY33-03

Abstract 2362: Defining the role of ZEB1 in the molecular pathogenesis of lung cancer

Abstract Background: Epithelial-to-mesenchymal transition (EMT) is an important biologic process in both early development and cancer whereby an epithelial cell transforms to a mesenchymal phenotype resulting in enhanced migration, invasiveness, and resistance to apoptosis. The transcription factor ZEB1 promotes EMT by repressing E-cadherin, and has been shown to promote tumor metastasis and increase resistance to standard chemotherapies in several human cancers. Its role in the tumorigenic progression of lung cancer however is less understood. Methods and Results: We analyzed genome-wide mRNA expression profiles across a cohort of 77 non small cell lung cancer (NSCLC) lines representing all common NSCLC histologies and oncogenotypes using Affymetrix GeneChip U133 Plus 2.0 arrays. Stratification of NSCLC lines by high/low ZEB1 expression identified 640 genes with significantly (p<0.001) different expression between the two groups. E-cadherin (CDH1) was significantly higher in cell lines with low ZEB1 expression, as was family member CDH3 and ERBB3. Mesenchymal markers vimentin, N-cadherin, (CDH2) as well as neuroendocrine markers (RTN1, NCAM1 and NEUROD1) were significantly up-regulated in cell lines with high ZEB1 expression. E-cadherin (r=-0.75) and Claudin 7 (r=-0.74) demonstrated the most significant indirect correlation with ZEB1 expression. To investigate the ability of ZEB1 to transform ‘normal’ epithelial cells we analyzed ZEB1 expression in an isogenic series of human bronchial epithelial cells (HBECs) - immortalized without viral oncoproteins - genetically manipulated to have single and multiple oncogenic changes common in lung cancer such as those for KRAS, p53, EGFR, and c-MYC. ZEB1 mRNA and protein expression was significantly increased in HBECs with combined oncogenic KRASV12 and p53 knockdown that had also been clonally selected for growth in soft agar. These transformed HBEC(KRASP53), which are tumorigenic in NOD/SCID mice in vivo, exhibit a decrease in epithelial markers and an increase in mesenchymal markers when compared to parental cells, indicating the cells have undergone EMT. To determine whether the observed increase in ZEB1 expression was causal to tumorigenicity or a bystander effect, ZEB1 expression was knocked down in HBEC(KRASP53) with RNA interference resulting in a significant decrease in colony formation, invasiveness and cell motility (p<0.005). Conclusion: These findings indicate that in NSCLC, ZEB1 expression is highly variable, is associated with an mRNA gene expression signature and is inversely correlated with E-cadherin expression, a known negative target of ZEB1. analyzing an isogenic series of progressively transformed HBECs we found ZEB1 increased with tumorigenic progression and an invasive phenotype. This phenotype could in turn be reversed by knockdown of ZEB1 suggesting ZEB1 plays an important role in tumorigenic transformation of human bronchial epithelial cells. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2362.

Smoking-related Genomic Signatures in Non–Small Cell Lung Cancer

Tobacco smoking is responsible for 85% of all lung cancers. To further our understanding of the molecular pathogenesis of lung cancer, we determined whether smoking history leads to the emergence of specific genomic alterations found in non-small cell lung cancer (NSCLC). To identify gene copy number alterations in NSCLCs associated with smoking history or DNA repair capacity. Seventy-five NSCLCs were selected for this study from patients with current, none, or past smoking history, including pack year information. Tissue sections were microdissected, and DNA was extracted, purified, and labeled by random priming before hybridization onto bacterial artificial chromosome (BAC) arrays. Normalized ratios were correlated with smoking history and DNA repair capacity was measured by an in vitro lymphocyte assay in the same patients. We identified smoking-related genomic signatures in NSCLCs that could be predicted with an overall 74% accuracy. Lung tumors arising from current-smokers had the greatest number of copy number alterations. The genomic regions most significantly associated with smoking were located within 60 regions and were functionally associated with genes controlling the M phase of the cell cycle, the segregation of chromosomes, and the methylation of DNA. Verification of the data is provided from data in the public domain and by quantitative real-time polymerase chain reaction. The associations between genomic abnormalities and DNA repair capacity did not reach statistical significance. These findings indicate that smoking history leaves a specific genomic signature in the DNA of lung tumors and suggest that these alterations may reflect new molecular pathways to cancer development.

SiRNA Targeting against EGFR, a Promising Candidate for a Novel Therapeutic Application to Lung Adenocarcinoma

Objective: To understand the molecular pathogenesis of lung cancer and to establish a novel therapeutic application, we examined the genetic alterations in lung cancer, and studied the effects of gefitinib and siRNA-mediated knockdown of EGFR on lung cancer. Methods: We analyzed mutations in EGFR, KRAS, TP53, and ERBB2 in 198 surgically resected lung cancer specimens. We then analyzed the effects of gefitinib and siRNA treatment on lung adenocarcinoma cell lines. Results: Mutations in EGFR were found only in adenocarcinoma (35 of 106 adenocarcinoma), mainly in females (73%). Mutually exclusive mutations of EGFR and KRAS genes were observed.Mutations of EGFR were well associated with a positive response to gefitinib. Cells with EGFR mutations were very sensitive to gefitinib as well as siRNA-mediated knockdown of EGFR, those with KRAS mutations responded poorly, and those without mutations of KRAS and EGFR showed moderate responses to both treatments. Conclusions: Our present results imply that (1) mutation analyses of EGFR and KRAS provide valuable information about whether or not to apply treatments targeting against EGFR and the selection of dosage for such treatments, and (2) siRNA-mediated knockdown is effective in lung adenocarcinomas with EGFR mutation, probably in those with resistance to gefitinib by acquired mutation in EGFR.

M15-02: Molecular pathogenesis of lung cancer and its precursor lesions

Molecular pathogenesis of lung cancer and its precursor lesions From histological and biological perspectives, lung cancer is a complex neoplasm. It can be divided into centrally arising tumors arising from the major bronchi and their divisions (squamous cell and small cell carcinomas) and peripherally arising adenocarcinomas (mainly adenocarcinomas and large cell carcinomas). In the bronchi the stem cell is thought to be the basal cell (for squamous cell carcinomas) and neuroendocrine cells (for small cell carcinomas). A peripheral airway cell has been identified in the mouse, and apparently is the common origin for clara cells and type 2 pneumocytes which are believed to be the precursor cells for bronchioles and alveoli respectively. Although the sequential preneoplastic changes have been defined for centrally arising squamous carcinomas of the lung, they have been poorly documented for the other major forms of lung cancers, including small cell lung carcinoma and adenocarcinomas. There are three main morphologic forms of preneoplastic lesions recognized in the lung: squamous dysplasias, atypical adenomatous hyperplasia (AAH), and diffuse idiopathic pulmonary neuroendocrine cell hyperplasia. However, these lesions account for the development of only a subset of lung cancers. Of interest, many changes associated with peripheral adenocarcinomas including KRAS and EGFR mutations have been demonstrated in AAH lesions. Some oncogenes such as EGFR may be activated by both mutations and/or amplification. Preliminary studies indicate that mutation occurs first, followed by preferential amplification of the mutant locus. Several studies have provided information regarding the molecular characterization of lung preneoplastic changes, especially for squamous cell carcinoma. These molecular changes have been detected in the histologically normal and abnormal respiratory epithelium of smokers. They indicate that molecular changes appear very early in the lengthy preneoplastic process, even before histologically detectable changes. Many histologically detected preneoplastic lesions are very small in size (1-5 mm). The small size and difficulties of identifying such lesions complicates their study. Fluorescence endoscopy aids the identification of such lesions, although most are fixed in formalin and paraffin embedded, rendering molecular studies more difficult. Approaches to the study of small preneoplastic lesions include microdissection of the epithelium, morphometric studies, allelic losses, gene mutations and amplifications, protein expression by immunostaing or in situ hybridization, gene expression at RNA or protein levels etc. The advent of CT screening approaches for early detection of lung cancer has led to the identification of small ground glass opacity (GGO) type of lesions, some of which are AAH lesions. Thus these lesions can be biopsied or removed and information about their natural history obtained. The natural history of preneoplastic lesions is complex, and cancers may appear at sites previously biopsied and having minimal or no morphologic changes. Thus preneoplastic lesions may be indicators of smoke damaged epithelium with a greater propensity to progress to high grade lesions or invasive cancer, although the latter may not arise directly from morphologically identified lesions. Global approaches to studying preneoplasia include microarray expression data and comparative genomic hybridization. These techniques clearly demonstrate progressive changes from normal epithelium to dysplasia to CIS. CIS is also somewhat different from invasive squamous carcinomas. In addition, the bronchial epithelium of smokers and never smokers demonstrate many changes. A novel method to study preneoplasia is the introduction of tumor associated genes into bronchial immortalized preneoplastic cell cultures. Bronchial epithelial cultures are immortalized by infection with CDK2 and hTERT. Such cells are immortal but retain minimal cytogenetic and molecular changes. After introduction of oncogenes such as mutated KRAS or p53 knowcksown, or both, they develop morphologic and biologic features suggestive of preneoplastic respiratory epithelium. These altered cell lines demonstrate squamous dysplastic changes and the ability to invade the underlying matrix. However they usually lack tumorigenicity in mice. They appear to be useful models for studying preneoplasia. Advances in biology are providing insights into the complex and lengthy preneoplastic process. These advances may impact on prevention measures and identification of individuals at greatly increased risk.

M10-01: Molecular pathogenesis of lung cancer with translation to the clinic

M10-01: Molecular pathogenesis of lung cancer with translation to the clinic

A Translational View of the Molecular Pathogenesis of Lung Cancer

Molecular genetic studies of lung cancer have revealed that clinically evident lung cancers have multiple genetic and epigenetic abnormalities, including DNA sequence alterations, copy number changes, and aberrant promoter hypermethylation. Together, these abnormalities result in the activation of oncogenes and inactivation of tumor-suppressor genes. In many cases these abnormalities can be found in premalignant lesions and in histologically normal lung bronchial epithelial cells. Findings suggest that lung cancer develops through a stepwise process from normal lung epithelial cells towards frank malignancy, which usually occurs as a result of cigarette smoking. Lung cancer has a high morbidity because it is difficult to detect early and is frequently resistant to available chemotherapy and radiotherapy. New, rationally designed early detection, chemoprevention, and therapeutic strategies based on the growing understanding of the molecular changes important to lung cancer are under investigation. For example, methylated tumor DNA sequences in sputum or blood are being investigated for early detection screening, and new treatments that specifically target molecules such as vascular endothelial growth factor and the epidermal growth factor receptor are becoming available. Meanwhile, global gene expression signatures from individual tumors are showing potential as prognostic and therapeutic indicators, such that molecular typing of individual tumors for therapy selection is not far away. Finally, the recent development of a model system of immortalized human bronchial epithelial cells, along with a paradigm shift in the conception of cancer stem cells, promises to improve the situation for patients with lung cancer. These advances highlight the translation of molecular discoveries on lung cancer pathogenesis from the laboratory to the clinic.

Genetics of Preneoplasia: Lessons from Lung Cancer

From biological, histopathologic, and clinical perspectives, lung cancer is a highly complex neoplasm probably having multiple preneoplastic pathways. The sequence of histopathologic changes in the bronchial mucosa that precedes the development of squamous carcinomas of the lung has been identified. For the other major forms of lung cancer, however, such sequences have been poorly documented. This review summarizes the current knowledge regarding the molecular and histopathologic pathogenesis of lung cancer and discusses the complexity of identifying novel molecular mechanisms involved in the development of the lung premalignant disease, and their relevance to the development of new strategies for early detection and chemoprevention. Although our current knowledge of the molecular pathogenesis of lung cancer is still meager, work over the last decade has taught several important lessons about the molecular pathogenesis of this tumor, including the following: a) Better characterization of the high-risk population is needed. b) There are several histopathologic and molecular pathways associated with the development of the major types of non-small cell lung cancer. c) Although there is a field effect phenomenon for lung preneoplastic lesions, recent data suggest that there are at least two distinct lung airway compartments (central and peripheral) for lung cancer pathogenesis. d) Inflammation may play an important role in lung cancer development and could be an important component of the field effect phenomenon. e) For lung adenocarcinoma, at least two pathways (smoking-related and nonsmoking-related) have been identified. f) Finally, the identification of deregulated molecular signaling pathways in lung cancer preneoplasias may provide a rationale for designing novel strategies for early detection and targeted chemoprevention of lung cancer.

Multiple bronchioloalveolar carcinomas in acromegaly: A potential role of insulin-like growth factor I in carcinogenesis

The molecular pathogenesis of lung cancer, especially multiple and synchronous bronchioloalveolar carcinomas (BACs), is still unknown. Here, we report two cases of multiple BACs associated with acromegaly, and discuss about the possible relationship between these two pathological condition. The first patient was a 52-year-old female with a history of Hardy's surgery for pituitary growth hormone cell adenoma 2 years earlier. The second patient was a 57-year-old female with acromegaly and obstructive sleep apnea syndrome. Both patients were non-smokers and showed a high serum level of insulin-like growth factor I (IGF-I) at the time of admission, even though the level of growth hormone had decreased. High-resolution computed tomography (HRCT) revealed multiple small nodules with pure ground-glass opacity (GGO) in both lungs of the first patient and a small nodule with pure GGO in the right lung of the second one. Partial resection for these tumors were performed under video-assisted thoracoscopic surgery. Resected lung specimens of the first case revealed one papillary adenocarcinoma, seven BACs, and 11 atypical adenomatous hyperplasias (AAHs). The second case showed two foci of BACs. Immunohistochemically, all BACs were strongly positive for IGF-IR which is a specific receptor for IGF-I, and all AAHs were also weakly positive for IGF-IR. Since IGF-I is known as a potent growth factor for normal as well as cancerous cells, it might play an important role for tumorigenesis and/or tumor progression of BACs through its interaction with and/or upregulation of IGF-IR. In addition, much attention should be paid to detect lung lesions in acromegaly with high serum level of IGF-I.

Immortalization of Human Bronchial Epithelial Cells in the Absence of Viral Oncoproteins

By expressing two genes (hTERT and Cdk4), we have developed a method to reproducibly generate continuously replicating human bronchial epithelial cell (HBEC) lines that provide a novel resource to study the molecular pathogenesis of lung cancer and the differentiation of bronchial epithelial cells. Twelve human bronchial epithelial biopsy specimens obtained from persons with and without lung cancer were placed into short-term culture and serially transfected with retroviral constructs containing cyclin-dependent kinase (Cdk) 4 and human telomerase reverse transcriptase (hTERT), resulting in continuously growing cultures. The order of introduction of Cdk4 and hTERT did not appear to be important; however, transfection of either gene alone did not result in immortalization. Although they could be cloned, the immortalized bronchial cells did not form colonies in soft agar or tumors in nude mice. The immortalized HBECs have epithelial morphology; express epithelial markers cytokeratins 7, 14, 17, and 19, the stem cell marker p63, and high levels of p16(INK4a); and have an intact p53 checkpoint pathway. Cytogenetic analysis and array comparative genomic hybridization profiling show immortalized HBECs to have duplication of parts of chromosomes 5 and 20. Microarray gene expression profiling demonstrates that the Cdk4/hTERT-immortalized bronchial cell lines clustered together and with nonimmortalized bronchial cells, distinct from lung cancer cell lines. We also immortalized several parental cultures with viral oncoproteins human papilloma virus type 16 E6/E7 with and without hTERT, and these cells exhibited loss of the p53 checkpoint and significantly different gene expression profiles compared with Cdk4/hTERT-immortalized HBECs. These HBEC lines are a valuable new tool for studying of the pathogenesis of lung cancer.

Multimodality Therapy in the Treatment of Lung Cancer

Lung cancer is the leading cause of cancer death in the world, causing more than one million deaths each year. The incidence and mortality rates are highest in the United States and Europe and continue to increase in developing nations. Non-small cell lung cancer (NSCLC) accounts for 80 to 85% of all new cases of lung cancer. The majority of patients with NSCLC present with advanced disease at the time of diagnosis. Although the prognosis of advanced NSCLC is very poor, current chemotherapy combinations have been shown to improve 1-year survival and quality of life for these patients. Approximately one third of patients with NSCLC are diagnosed with locally advanced disease. Although cure rates are modest and variable in locally advanced NSCLC, multimodality therapy (chemotherapy in combination with surgery or radiotherapy) has resulted in statistically significant improvement in 5-year survival when compared with surgery or radiotherapy alone. Patients with early-stage NSCLC have the best long-term survival rates following surgical resection; however, systemic recurrences remain a problem in the majority of these patients. The rationale for treating patients with early-stage NSCLC with combined-modality therapy (chemotherapy and surgery) is compelling, and several randomized trials are currently in progress. Although progress has been slow, when we consider the recent advances in smoking prevention, smoking cessation, staging classification, imaging and diagnostic techniques, screening and therapeutic modalities, and multidisciplinary care, as well as in the understanding of the molecular pathogenesis of lung cancer, the future, in my opinion, is very promising.

Genome-scale analysis of lung cancer progression.

Among cancers, lung cancer is the single biggest killer in the US. It is estimated that lung cancer was responsible for 171900 newly diagnosed cases of cancer in the US in 2003, and for 157200 deaths. Over many years, however, there has been little improvement in the clinical outcome of lung cancer, and any improvement in the incidence or mortality from lung cancer can largely be attributed to smoking cessation and not to the success of therapy. The histopathology of lung cancer reveals that it is a disease with many faces. Lung cancer is often nonresponsive to traditional therapy, leaving few, if any, alternatives in the management of the advanced stages of the disease. The molecular pathogenesis of lung cancer, only recently illuminated, involves numerous molecular and cell biological changes revealing a very complex disease progression. Large-scale mRNA expression analysis has been recently used to classify lung cancers molecularly. These techniques have been used successfully to differentiate lung cancer histotypes based on patterns of genes expressed. The use of protein analysis to this end has also been attempted, with limited correlation with RNA experiments. This likely reflects the limited sensitivity of the technologies and complex, poorly understood post-synthesis protein modifications. In any event, there have been great strides made in understanding the nature of lung cancer from a molecular perspective; these effects represent a great advancement in the diagnosis and prognosis of lung cancer. Moreover, these advances may lead to the improvement of patient survival by guiding the choice of more efficacious therapy.

Proteomic analysis of a neoplastic mouse lung epithelial cell line whose tumorigenicity has been abrogated by transfection with the gap junction structural gene for connexin 43, Gja1.

In order to examine how tumorigenicity is abrogated by gap junctional intercellular communication (GJIC), protein expression was analyzed in four related mouse lung epithelial cell lines that vary in their GJIC status and neoplastic potential. Since alterations in protein expression underlie neoplastic behavior, this proteomic analysis provides insights into the molecular pathogenesis of lung cancer. E10, an immortalized but non-tumorigenic cell line derived from alveolar type II pneumocytes, has functional GJIC. E9, a spontaneous transformant of E10, is GJIC-deficient and is tumorigenic upon injection into a syngeneic mouse. Stable transfection of E9 with Gja1, the gene for the gap junctional protein, connexin 43, re-established GJIC and rendered this line (designated E9-2) non-tumorigenic; the vector transfection control line, E9-41, remains tumorigenic. Proteins extracted from these cell lines were separated by two-dimensional electrophoresis (2DE) and visualized by Coomassie blue staining. We consistently observed differential expression of 27 proteins between E10 and E9 and identified 11 of these by peptide mass mapping. The functions of these proteins include stress response, cytoskeletal structure, signal transduction, apoptosis, immune response, pre-mRNA processing, and carbohydrate metabolism. Gja1 transfection affected the concentrations of four of these proteins, viz. PDI, alpha-enolase, aldolase A, and gelsolin-like protein. PDI concentration was most profoundly affected; E10 cells contain twice as much PDI as E9, and PDI was restored to E10-like levels in the E9-2 transfectant line while remaining at E9-like levels in the vector control E9-41 cells. An association between connexin 43 and PDI expression was also observed in a second set of independently derived type II cell lines. The PDI superfamily has multiple cellular roles including chaperoning assembled glycoproteins, regulating the activities of transcription factors, and regulating disulfide bond formation.

Thyroid transcription factor-1 (TTF-1) expression in human lung carcinomas: its prognostic implication and relationship with wxpressions of p53 and Ki-67 proteins.

This study was aimed to evaluate the prevalence and prognostic implication of thyroid transcription factor-1 (TTF-1) immunoreactivity in 81 human lung carcinomas, including 65 cases of non-small cell lung carcinoma (NSCLC) and 16 cases of small cell lung carcinoma (SCLC); and also to investigate its relationship with the cell proliferation and regulation by immunostaining of Ki-67 and p53 proteins, respectively. The immunohistochemical staining for TTF-1 (clone 8G7G3/1) was performed and several clinicopathologic variables and the follow-up data were obtained. The immunostaining results for TTF-1 were semiquantitatively interpreted as negative and positive. Of NSCLCs, TTF-1 is highly expressed in adenocarcinomas (76%), whereas squamous cell carcinomas revealed no immunoreactivity (0%). SCLCs showed strong TTF-1 expression (88%). In NSCLC, TTF-1 expression was inversely correlated with Ki-67 proliferative activity and independent of p53 overexpression. TTF-1 (+) group tended to show better survival than TTF-1 (-) group in NSCLC. Conclusively, these observations suggest that TTF-1 is a sensitive and specific diagnostic marker for pulmonary adenocarcinomas and SCLCs; that TTF-1 might have a good prognostic implication based on its inverse correlation with Ki-67 proliferative activity and tendency for better survival in NSCLC; that this cell lineage marker may play a role in the molecular pathogenesis of lung cancers at the level of transcription.

Gene therapy for lung cancer.

Lung cancer continues to be the largest killer of Americans due to cancer. Although progress has been made, with advances in chemotherapy, the majority of patients diagnosed with lung cancer ultimately succumb to the disease. A better understanding of the molecular pathogenesis of lung cancer is demonstrating how alterations in oncogenes and tumor suppressor genes control lung cancer initiation, growth, and survival. In this article, attempts to target molecular alterations in lung cancer using gene therapy techniques are reviewed. These include introducing suicide genes into tumor cells, replacement of defective tumor suppressor genes, inactivating oncogenes, and immunotherapy-based approaches using gene therapy technology. The major barrier for these techniques continues to be the inability to specifically target tumor cells while sparing normal cells. Nonetheless, these approaches are likely to yield important biologic and clinical data which will further the progress of lung cancer treatment.

Molecular pathogenesis of lung cancer.

Lung cancer is the most common cause of cancer death in the United States, killing more than 156,000 people every year. In the past two decades, significant progress has been made in understanding the molecular and cellular pathogenesis of lung cancer. Abnormalities of proto-oncogenes, genetic and epigenetic changes of tumor suppressor genes, the role of angiogenesis in the multistage development of lung cancer, as well as detection of molecular abnormalities in preinvasive respiratory lesions, have recently come into focus. Efforts are ongoing to translate these findings into new clinical strategies for risk assessment, chemoprevention, early diagnosis, treatment selection, and prognosis and to provide new targets and methods of treatment for lung cancer patients. All these strategies should aid in reducing the number of newly diagnosed lung cancer cases and in increasing the survival and quality of life of patients with lung cancer.

Molecular genetic abnormalities in the pathogenesis of human lung cancer.

In the past few years our knowledge of the molecular pathogenesis of lung cancer has significantly increased. There are several molecular mechanisms involved in the multistage carcinogenesis through which respiratory epithelial cells become preneoplastic and then invasive cancer. In this review we summarize some of these changes including, genomic alterations such as loss of heterozygosity and microsatellite alterations, autocrine-paracrine loops, alterations in oncogenes and tumor suppressor genes, tumor angiogenesis, aberrant promoter methylation and inherited predisposition to lung cancer. Translation of these findings to the clinic is also discussed.

Molecular analysis of the mitotic checkpoint genes BUB1, BUBR1 and BUB3 in human lung cancers

Our previous studies showed that mitotic checkpoint impairment is present in about 40% of human lung cancer cell lines but that mutations in the MAD mitotic checkpoint genes are infrequent. In the present study, we examined 44 lung cancer cases for the potential involvement of the other gene family involved in the mitotic checkpoint, i.e. BUB. We found that the BUB gene family members including BUB1, BUBR1 and BUB3 are not frequent targets for mitotic checkpoint defects in lung cancers, if present at all. Further studies are thus warranted to elucidate the molecular basis for the acquisition of mitotic checkpoint defects in order to better understand the molecular pathogenesis of lung cancers.

Symposium on molecular pathogenesis of respiratory diseases and its clinical implication. 4. Molecular pathogenesis of lung cancer and its molecular targeted therapy.

Symposium on molecular pathogenesis of respiratory diseases and its clinical implication. 4. Molecular pathogenesis of lung cancer and its molecular targeted therapy.