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Портал Begell Электронная Бибилиотека e-Книги Журналы Справочники и Сборники статей Коллекции
Journal of Environmental Pathology, Toxicology and Oncology
Импакт фактор: 1.625 5-летний Импакт фактор: 1.63 SJR: 0.402 SNIP: 0.613 CiteScore™: 2.3

ISSN Печать: 0731-8898
ISSN Онлайн: 2162-6537

Выпуски:
Том 39, 2020 Том 38, 2019 Том 37, 2018 Том 36, 2017 Том 35, 2016 Том 34, 2015 Том 33, 2014 Том 32, 2013 Том 31, 2012 Том 30, 2011 Том 29, 2010 Том 28, 2009 Том 27, 2008 Том 26, 2007 Том 25, 2006 Том 24, 2005 Том 23, 2004 Том 22, 2003 Том 21, 2002 Том 20, 2001

Journal of Environmental Pathology, Toxicology and Oncology

DOI: 10.1615/JEnvironPatholToxicolOncol.v28.i2.10
pages 89-98

MET, HGF, EGFR, and PXN Gene Copy Number in Lung Cancer Using DNA Extracts from FFPE Archival Samples and Prognostic Significance

Rajani Kanteti
Department of Medicine, Section of Hematology and Oncology University of Chicago, Chicago, IL
Soheil Yala
Department of Medicine, Section of Hematology and Oncology University of Chicago, Chicago, IL
Mark K. Ferguson
Department of Surgery; University of Chicago, Chicago, IL
Ravi Salgia
Department of Medical Oncology and Therapeutics Research, Comprehensive Cancer Center, City of Hope National Medical Center

Краткое описание

Gene copy number analysis for some of the important molecules in lung tumorogenesis, such as MET, hepatocyte growth factor [(HGF), ligand for MET), epidermal growth factor receptor (EGFR), and paxillin (PXN), is likely to determine both the type of treatment and prognosis. Formalin-fixed paraffin-embedded (FFPE) archival tumor tissue samples are an excellent source for determining key molecular changes in the OncoGenome; however, existing extraction procedures yield relatively poor quality genomic DNA fragments. Although FISH is the method of choice for determining amplification of a gene, a more rapid quantitative poly-merase chain reaction (qPCR) technique to determine gene copy number can be used when reasonably good quality genomic DNA is available. We report here a relatively rapid method based on microwave/chelex-100 treatment that gives rise to genomic DNA fragments ranging from 1 to 12 Kb and beyond, thereby attesting to its superior quality. Genomic PCR for β-globin gene gave reliable and reproducible results. The number of steps for extracting the DNA was kept to a minimum, and instead of precipitating the DNA, we preserved the genomic DNA extracts so as to prevent a loss in DNA yield. We found the extracts to be stable and amenable to qPCR and mutational analysis. Using lung adenocarcinoma FFPE samples and cell lines derived from lung adenocarcinomas, we demonstrated that the gene copy number for MET in lung adenocarcinoma tissue samples was preferentially increased over EGFR, HGF, and PXN and that it positively correlated with a better prognosis. In contrast, the genomic DNA extracted from 25 NSCLC cell lines gave a relatively higher gene copy number for all four genes evaluated. Our results indicate that the microwave/chelex-100-based method yields good-quality genomic DNA extracts that can be used for complex DNA analysis, such as determination of gene copy number. In addition, our data demonstrated that the adenocarcinoma cell lines potentially evolved under ex vivo conditions, and therefore, in genetic studies it is imperative to use primary tumors for generalized conclusions about lung tumors.

Ключевые слова: genomic DNA, FFPE, MET, EGFR, paxillin, gene copy number

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