Screening of Epidermal Growth Factor Receptor Gene Mutation in Non-small Cell Lung Cancer Using a New PCR-based Enzymatic Digestion Method
Young T. Kim2, Sun J. Park2, Joo-yeon Park2, Hyun C. Wi2, Chang H. Kang1, Sook W. Sung2, Joo H. Kim1; 1Thoracic and Cardiovascular Surgery, Seoul National University Hospital, Seoul, South Korea; 2Cancer Research Institite, Seoul National University, Seoul, South Korea
Comment on this Abstract
Objective: Currently available methods for detection of Epidermal Growth Factor Receptor (EGFR) mutation rely on direct DNA sequencing. We developed a simplified method using PCR-based enzymatic digestion for the detection of exons 19 and 21 mutations and validated its usefulness as a screening tool.
Methods: We selected 74 samples of adenocarcinoma of the lung whose EGFR exons 19 and 21 had been previously sequenced. Based on the sequencing result, we designed PCR primers and chose DNA restriction enzyme. The PCR products were tested on the agarous gel directly for exon 19 and after enzymatic digestion for exon 21. To validate its accuracy, we set up the second sets of 74 lung cancer samples. For those samples, PCR-based method was performed first and the result was validated by DNA sequencing.
Results: In the first sample group, we found 15 (20.3%) and 9 (12.2%) mutations for exons 19 and 21 using sequencing method, respectively. By using PCR-based method, we were able to identify all the mutated samples detected by sequencing method. At the same time, we could detect mutations in additional 3 and 1 samples for exons 19 and 21, respectively. Those additionally detected 4 mutations were confirmed by performing a repeated sequencing. In the second set of samples, PCR-based method detected 10 (13.5%) and 7 (9.5%) mutations for exons 19 and 21, respectively. Additional mutations of exon 19 were identified in 2 samples by sequencing method. However, sequencing method failed to identify mutation of exon 21 in one sample.
Conclusion: The sensitivity of PCR-based enzymatic digestion method seems to be comparable to that of the traditional sequencing method for detecting EGFR mutations. As our new method is simple, cheap, rapid and sensitive, it can be widely used as a screening test for patient selection who may benefit from EGFR targeted therapy.
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