The epigenomics of cancer.

This article reviews the mechanisms of gene silencing in cancer and clinical applications of this phenomenon. The silencing of genes, especially tumor-suppressor genes, is a key event in the development of cancer. The silencing can be effected by a disabling mutation or by a shutting down of the promoter region, the site at which transcription of the gene begins.

Gene Silencing in Cancer in Association with Promoter Hypermethylation

Since its discovery in 1983, the epigenetics of human cancer has been in the shadows of human cancer genetics. But this area has become increasingly visible with a growing understanding of specific epigenetic mechanisms and their role in cancer, including hypomethylation, hypermethylation, loss of imprinting and chromatin modification. This timeline traces the field from its conception to the present day. It also addresses the genetic basis of epigenetic changes — an emerging area that promises to unite cancer genetics and epigenetics, and might serve as a model for understanding the epigenetic basis of human disease more generally.

The history of cancer epigenetics.

We are in an era where the potential exists for deriving comprehensive profiles of DNA alterations characterizing each form of human cancer. Such profiles would provide invaluable insight into mechanisms underlying the evolution of each tumor type and will provide molecular markers, which could radically improve cancer detection. To date, no one type of DNA change has been defined which accomplishes this purpose. Herein, by using a candidate gene approach, we show that one category of DNA alteration, aberrant methylation of gene promoter regions, can enormously contribute to the above goals. We have now analyzed a series of promoter hypermethylation changes in 12 genes (p16(INK4a), p15(INK4b), p14(ARF), p73, APC,(5) BRCA1, hMLH1, GSTP1, MGMT, CDH1, TIMP3, and DAPK), each rigorously characterized for association with abnormal gene silencing in cancer, in DNA from over 600 primary tumor samples representing 15 major tumor types. The genes play known important roles in processes encompassing tumor suppression, cell cycle regulation, apoptosis, DNA repair, and metastastic potential. A unique profile of promoter hypermethylation exists for each human cancer in which some gene changes are shared and others are cancer-type specific. The hypermethylation of the genes occurs independently to the extent that a panel of three to four markers defines an abnormality in 70-90% of each cancer type. Our results provide an unusual view of the pervasiveness of DNA alterations, in this case an epigenetic change, in human cancer and a powerful set of markers to outline the disruption of critical pathways in tumorigenesis and for derivation of sensitive molecular detection strategies for virtually every human tumor type.

https://cancerres.aacrjournals.org/content/canres/61/8/3225.full.pdf

A Gene Hypermethylation Profile of Human Cancer

The complexity of tobacco smoke leads to some confusion about the mechanisms by which it causes lung cancer. Among the multiple components of tobacco smoke, 20 carcinogens convincingly cause lung tumors in laboratory animals or humans and are, therefore, likely to be involved in lung cancer induction. Of these, polycyclic aromatic hydrocarbons and the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone are likely to play major roles. This review focuses on carcinogens in tobacco smoke as a means of simplifying and clarifying the relevant information that provides a mechanistic framework linking nicotine addiction with lung cancer through exposure to such compounds. Included is a discussion of the mechanisms by which tobacco smoke carcinogens interact with DNA and cause genetic changes--mechanisms that are reasonably well understood--and the less well defined relationship between exposure to specific tobacco smoke carcinogens and mutations in oncogenes and tumor suppressor genes. Molecular epidemiologic studies of gene-carcinogen interactions and lung cancer--an approach that has not yet reached its full potential--are also discussed, as are inhalation studies of tobacco smoke in laboratory animals and the potential role of free radicals and oxidative damage in tobacco-associated carcinogenesis. By focusing in this review on several important carcinogens in tobacco smoke, the complexities in understanding tobacco-induced cancer can be reduced, and new approaches for lung cancer prevention can be envisioned.

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Tobacco Smoke Carcinogens and Lung Cancer

The p16INK4a (p16) tumor suppressor gene can be inactivated by promoter region hypermethylation in many tumor types including lung cancer, the leading cause of cancer-related deaths in the U.S. We have determined the timing of this event in an animal model of lung carcinogenesis and in human squamous cell carcinomas (SCCs). In the rat, 94% of adenocarcinomas induced by the tobacco specific carcinogen 4-methylnitrosamino-1-(3-pyridyl)-1-butanone were hypermethylated at the p16 gene promoter; most important, this methylation change was frequently detected in precursor lesions to the tumors: adenomas, and hyperplastic lesions. The timing for p16 methylation was recapitulated in human SCCs where the p16 gene was coordinately methylated in 75% of carcinoma in situ lesions adjacent to SCCs harboring this change. Moreover, the frequency of this event increased during disease progression from basal cell hyperplasia (17%) to squamous metaplasia (24%) to carcinoma in situ (50%) lesions. Methylation of p16 was associated with loss of expression in both tumors and precursor lesions indicating that both alleles were functionally inactivated. The potential of using assays for aberrant p16 methylation to identify disease and/or risk was validated by detection of this change in sputum from three of seven patients with cancer and 5 of 26 cancer-free individuals at high risk. These studies show for the first time that an epigenetic alteration, aberrant methylation of the p16 gene, can be an early event in lung cancer and may constitute a new biomarker for early detection and monitoring of prevention trials.

https://www.pnas.org/content/pnas/95/20/11891.full.pdf

Aberrant methylation of p16(INK4a) is an early event in lung cancer and a potential biomarker for early diagnosis.

The p16 INK4a (p16) tumor suppressor gene can be inactivated by promoter region hypermethylation in many tumor types including lung cancer, the leading cause of cancer-related deaths in the U.S. We have determined the timing of this event in an animal model of lung carcinogenesis and in human squamous cell carcinomas (SCCs). In the rat, 94% of adenocarcinomas induced by the tobacco specific carcinogen 4-methylnitrosamino-1-(3-pyridyl)-1-butanone were hypermethylated at the p16 gene promoter; most impor- tant, this methylation change was frequently detected in precursor lesions to the tumors: adenomas, and hyperplastic lesions. The timing for p16 methylation was recapitulated in human SCCs where the p16 gene was coordinately methylated in 75% of carcinoma in situ lesions adjacent to SCCs harboring this change. Moreover, the frequency of this event increased during disease progression from basal cell hyperplasia (17%) to squamous metaplasia (24%) to carcinoma in situ (50%) lesions. Methylation of p16 was associated with loss of expres- sion in both tumors and precursor lesions indicating that both alleles were functionally inactivated. The potential of using assays for aberrant p16 methylation to identify disease andyor risk was validated by detection of this change in sputum from three of seven patients with cancer and 5 of 26 cancer-free individuals at high risk. These studies show for the first time that an epigenetic alteration, aberrant methylation of the p16 gene, can be an early event in lung cancer and may constitute a new biomarker for early detection and monitoring of pre- vention trials.

Aberrant methylation of p16 INK4a is an early event in lung cancer and a potential biomarker for early diagnosis (p16 methylationysputumyNNKysquamous cell carcinoma)

OBJECTIVE: A procedure was developed to allow concurrent detection of chromosome aberrations and identification of bronchial epithelial cells. STUDY DESIGN: Fluorescence in situ hybridization for chromosome 7 and immunocytochemistry for cytokeratin were performed on exfoliated bronchial epithelial cells in a sputum sample from a cancer patient. RESULTS: The Spectrum Orange-labeled alpha satellite probe for chromosome 7 produced red fluorescence, nuclei were counter-stained with 4,6-diamidino2-phenylindole (blue), and cytokeratin was visualized using a fluorescein isothiocyanate (FITC)-conjugated secondary antibody (green). CONCLUSION: This procedure allowed the rapid identification of airway epithelial cells with numerical chromosome aberrations in this sample. Ultimately, this procedure could increase the sensitivity and specificity of sputum cytology as a laboratory diagnostic tool for the early detection of lung cancer.

Concurrent fluorescence in situ hybridization and immunocytochemistry for the detection of chromosome aberrations in exfoliated bronchial epithelial cells.

Ruth Michels

Papers

Aberrant methylation of p16(INK4a) is an early event in lung cancer and a potential biomarker for early diagnosis.

Aberrant methylation of p16 INK4a is an early event in lung cancer and a potential biomarker for early diagnosis (p16 methylationysputumyNNKysquamous cell carcinoma)

Concurrent fluorescence in situ hybridization and immunocytochemistry for the detection of chromosome aberrations in exfoliated bronchial epithelial cells.