Genetics of preneoplasia and lung cancer development

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Abstracts

developing mammalian lung. The abundance of fetal expression of hnRNP A2/B1 is tightly associated with periods of peak lung development and then is restricted to low levels with maturation. The expression of this antigen is also frequently elevated in bronchial biopsies obtained from the airways of smokers with bronchial metaplasia. The frequent expression of hnRNP A2/B1 in this setting contrasts with the much more selective expression of this antigen in exfoliated sputum. The difference in the two settings suggests that the exfoliated sputum cell may be a particularly informative specimen source. The biological process that leads to exfoliation in general is an injury response. In the setting of field carcinogenesis, the cells that have undergone progressive genetic injury are known to evolve through a process of metastatic competence that is associated with progressive loss of contact with neighboring cell and the basement membrane. As a consequence of becoming invasive, these same cells have been shown also to be more resistant to mechanical stress and proteolysis. The implication is that the success of the sputum immunohistochemical assays is dependent both on the expression status of hnRNP A2/B1 and the presence of exfoliated bronchial epithelial cells. In the reports of the cohorts studied with this assay to date, the sensitivity of the test is around 70%. Measures to improve this performance include refining the assay procedure, adding the use of other markers, evaluating the benefit of multiple assay determinations and systematically evaluating the basis for assay failure. For lung cancer, the path to improved mortality outcomes is to improve the detection of the disease prior to metastatic dissemination. Along with spiral CT, sputum immunocytochemistry is the type of technology with the a priori characteristics that make population-based application feasible. Rigorous clinical trials are required to evaluate these new approaches in the routine detection of very early lung cancer. Further work will be required to refine the management of the very early phase of cancer that will be routinely detected with these new approaches. We are evaluating the use of aerosolized drug delivery strategies to determine if that approach is useful in allowing for high therapeutic index strategies to control the natural history of very early lung cancer. References: 1. Hirsch FR, Brambilla E, Gray N et a1. Prevention and Early Detection of Lung CancerClinical Aspects. Lung Cancer, 17:163-174, 1997. 2. Mulshine JL, Zhou J, Treston AM et al. New Approaches to the Integrated Management of Early Lung Cancer. Hematol Oncol Clin NA, 11 :235-252, 1997. 3. Mulshine JL, Tockman MS. Transition to Early Lung Cancer Management. IASLC Textbook of Lung Cancer. In press, 1999. 4. Tockman, MS, Mulshine JL, Piantadosi S et al. Prospective detection of preclinical lung cancer results from two studies of hnRNP overexpression. Clin. Cancer Res.,3:2237-2246, 1997. 5. Tockman MS, Gupta PK, Myers JD et al. Sensitive and Specific Monoclonal Antibody Recognition of Human Lung Cancer Antigen on Preserved Sputum Cells: A New Approach to Early Lung Cancer Detection. J Clin Oncol 6:1685-1693, 1988. Genetics of preneoplasia and lung cancer development Adi F. Gazdar, Hamon Cancer Center, UT Southwestern Medical Center, Dallas, TX Three important concepts are important in understanding the pathogenesis of lung cancer: 1) lung cancer is preceded by multiple preneoplastic changes that take many years to evolve into invasive cancer; 2) multiple molecular changes involving recessive and dominant oncogenes are present in lung cancers; and 3) these changes commence early in the pathogenesis. These concepts can be

explained by the field cancerization theory that states that much of the upper aerodigestive tract has been damaged by exposure to carcinogens present in tobacco smoke. The study of lung carcinogenesis is complicated by the inability to identify and isolate most preneoplastic lesions except by histopathological examination. The use of fluorescence bronchoscopy has greatly increased the ability to identify dysplastic lesions in those at increased risk (i.e. current and former smokers), permitting reversal of dysplasia to be used as an indicator for following the efficacy of chemoprevention regimens. These studies also indicate that women smokers (who are at increased risk for developing cancer) have fewer dysplastic lesions than male smokers do. Apparently tobacco causes more damage in the peripheral parts of the lung in women, resulting in adenocarcinomas, while the damage in male smokers is more central, resulting in squamous cell and small cell carcinomas. We have studied normal and abnormal bronchial epithelium from smokers and patients with lung cancer. Most of the lesions from smokers were obtained by fluorescent bronchoscopy, and consist of archival paraffin embedded biopsies. However, we have also developed a simple procedure known as EASI, in which scraped epithelium is fixed in methanol. EASI preps permit a wide variety of DNA and RNA assays on histologically identified lesions. Our studies have helped to identify multiple regions of allelic losses in tumors, presumably the sites of known and unknown tumor suppressor genes. The changes are more extensive in squamous cell and small cell carcinomas than in adenocarcinomas. However, ras mutations are associated with a subset of adenocarcinomas. Of interest, in cancers arising in Japanese patients, the changes in male and female lifetime non-smokers are very similar to those present in smokers. In both current and former smokers, the changes commence very early during pathogenesis, in histologically normal epithelium. They progressively increase with increasing pathological stage. In small cell carcinomas the changes in normal epithelium are far more extensive than in other cancer types, suggesting that small cell cancers may arise directly from histologically normal or slightly abnormal epithelium. We are slowly ascertaining the sequence of molecular events preceding cancers. Discreet losses at one of more chromosome 3p and 9p regions are the earliest detected changes, followed by loss at chromosome 8p. Losses at the p53 gene and mutations of the p53 and ras genes are later events. Most other changes are even later, arising in severe dysplasia or carcinoma in situ, which are characterized by loss of all or almost the entire 3p arm. The patterns of loss may be important in predicting which smokers are at highest risk for the development of cancer. The molecular changes are very similar in both current and former smokers and persist for decades after smoking cessation. Hypermethylation of the promoter regions of certain genes is a frequent event during carcinogenesis. Each tumor type has its own specific set of genes that are inactivated by this mechanism. We, in collaboration with James Herman and Stephen Baylin (Johns Hopkins Oncology Center) have determined the panel of genes frequently inactivated in non-small cell lung cancer, and are currently investigating the panel of genes involved with neuroendocrine lung tumors. Inactivation by hypermethylation commences early during lung cancer pathogenesis. Presence of aberrantly methylated genes in the plasma may be a clinically specific and sensitive test for early detection of invasive cancers. Chemoprevention trials require sequential biopsies of the same area to detect changes in morphology and molecular markers. Thus, it is important to determine the precise size of the clonal patches present in the bronchial epithelium. To investigate this, we utilized utilizing

Abstracts cross sections of bronchi from lobectomy specimens of patients with lung cancer. We carefully microdissected ~200 cell pieces of normal and slightly abnormal bronchial mucosa from near the proximal resection margin, and determined whether molecular changes were present in them. These two dimensional studies indicate that most clonal patches are surprisingly small, 200 Ñ 400 cells in size. No patch larger than 600 cells in size was identified. These studies indicate that very careful sampling of the bronchial mucosa must be performed if reproducible data from sequential biopsies is to be generated. Several markers are being investigated to determine whether they aid in the diagnosis of early cancers. These markers may be present in sputum, bronchoalveolar lavages and blood. While several markers appear promising, they are not yet ready for mass screening approaches. In summary, we have learnt a great deal about the molecular pathogenesis of lung cancers, and the information is being translated into clinical applications including risk assessment, chemoprevention trials and early diagnosis. References 1. Wistuba, I. I., Lam, S., Behrens, C., Virmani, A. K., Fong, K. M., LeRiche, J., Samet, J. M., Srivastava, S., D., M. J. and Gazdar, A. F. Molecular damage in the bronchial epithelium of current and former smokers. J. Natl. Cancer Inst., 89: 1366-1372, 1997. 2. Yashima, K., Litzky, L. A., Kaiser, L., Rogers, T., Lam, S., Wistuba, I.I, Milchgrub, S., Srivastava, S., Piatyszek, M. A., Shay, J. W. and Gazdar, A. F. Telomerase expression in respiratory epithelium during the multistage pathogenesis of lung carcinomas. Cancer Res, 57: 2373-7, 1997. 3. Wistuba, I. I. and Gazdar, A. F. Molecular abnormalities in the sequential development of lung carcinoma. In: S. Srivastava, D. E. Henson and A. F. Gazdar (eds.), Molecular Pathology of Early Cancer, pp. 265-276. Amsterdam: IOS Press, 1999. 4. Wistuba, I. I., Behrens, C., Virmani, A., Milchgrub, S., Syed, S., Lam, S., Mackay, B., Minna, J. D. and Gazdar, A. F. Allelic losses at chromosome 8p21-23 are early and frequent events in the pathogenesis of lung cancers. Cancer Res., 59: 1973-1979, 1999. 5. Onuki, N., Wistuba, I. I., Travis, W. D., Virmani, A. K., Yashima, Y., Brambilla, E., Hasleton, P. and Gazdar, A. F. Genetic changes in the spectrum of neuroendocrine lung tumors. Cancer, 85: 600-607, 1999. 6. Shivapurkar, N., Virmani, A. K., Wistuba, I. I., Milchgrub, S., Mackay, B., Minna, J. D. and Gazdar, A. F. Deletions of chromosome 4 at multiple sites are frequent in malignant mesothelioma and small cell lung carcinoma. Clinical Cancer Res, 5: 17-23, 1999. 7. Wistuba, I. I., Behrens, C., Milchgrub, S., Bryant, D., Hung, J., Minna, J. D. and Gazdar, A. F. Sequential molecular abnormalities are involved in the multistage development of squamous cell lung carcinoma. Oncogene, 18: 643-650, 1999. 8. Esteller, M., SanchezCespedes, M., Rosell, R., Sidransky, D., Baylin, S. B. and Herman, J. G. Detection of aberrant promoter hypermethylation of tumor suppressor genes in serum DNA from non-small cell lung cancer patients. Cancer Res, 59: 67-70, 1999. Intensive care for lung cancer patients Jean-Paul Sculier, Institut Jules Bordet, Brussels, Belgium. Introduction: Intensive care has taken an active part in the management of cancer patients at any disease stage during the last decade. Major cancer hospitals have developed intensive therapy units that take care not only of surgical patients but also of medical patients. In an international inquiry (1) about intensive care in anticancer centres, we found that there was at least one oncological intensive care unit (ICU) in 70% of the hospitals. The presumption, that often predominates in general hospitals, considering that patients with active malignant disease should not be admitted in

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ICU, has today to be looked as a conservative attitude that has been surpassed by the development of oncology. There are however few data in the medical literature about intensive care in oncology and they are practically absent for specific solid tumours like lung cancer. There are three main indications to admit a cancer patient in ICU: postoperative recovery, critical complications of cancer and its treatment, intensive anticancer treatment administration and/or monitoring. Postoperative recovery: Cancer patients can be admitted in ICU for postoperative care after extensive surgical procedures. Potential benefits of intensive care for such patients are availability of continuous haemodynamic monitoring, early identification of cardiovascular and respiratory disturbances, facilities for respiratory support and constant skilled nursing care. In fact, these advantages are the same as those for any high risk postoperative patient. Postsurgery care will often be not specific to the cancer disease but will depend much more on the type of surgical traumatic procedure that is performed. Recovery after surgery for lung cancer is a main indication for intensive care. Surgical mortality for the Òaverage riskÓ patients, as determined by review of multiple surgical experiences, is about 3% following lobectomy and 7% for pneumonectomy. These figures might be minimised by appropriate intensive care environment. In the immediate postoperative period, the priority concern should be the continuing function of the patientÕs remaining pulmonary tissue, because any further deterioration of the lung function might not be tolerated. Maintenance of the functional residual capacity includes less right-to-left shunt, better oxygenation and less work of breathing. Intravenous fluid therapy should be appropriate for the patientÕs cardiovascular and renal functions. A Swan-Ganz catheter should be inserted in patients with compromised heart function or in whom the clinical assessment of the intravascular volume status required optimum monitoring of left heart filling pressures. Critical complications of cancer and its treatment: Intensive care of cancer patients with lifethreatening problems (2) has specific characteristics related to the spectrum of complications (coronary problems are not frequent for example), to special emergencies mainly seen in oncology (severe hypercalcaemia is a common cause of admission for instance) and the presence of a severe chronic underlying disease with systemic effects (the cancer disease). Moreover, in lung cancer patients, severe comorbidity related to smoking can be present like chronic obstructive pulmonary disease or arterial diseases. In our experience, main types of problems requiring ICU admissions are respiratory (23%), cardiac (25%), digestive (8%), renal and metabolic (13%), neurologic (10%) and infectious, haematological and shock (21%). Respiratory problems include respiratory failure due to tumoral obstruction, infectious pneumonia, diffuse pneumopathy, ARDS (adults respiratory distress syndrome), superior vena cava syndrome, pleural effusion, pneumothorax and fistulas. Endoscopic laser therapy is often required in case of respiratory distress due to obstructive bronchial tumors (3). Cardiac complications are thromboembolic disease, arrhythmia, syncope, heart failure, cardiac arrest, coronary complications, acute thoracic pain and pericardial disease. In lung cancer, metastases to the pericardium or heart are quite frequent, detectable at autopsy in one third of the patients and explaining why cardiac arrhythmia might be frequent in this disease and why pericardial complications are not rare. Lung cancer is a main cause of superior vena cava syndrome, a well known oncological emergency. Metabolic problems are often due to hypercalcaemia that can be related to bone metastases or a paraneoplastic syndrome. In lung cancer, renal complications are often related to