Detection of EGFR Exon 19 and 21 Mutations in Pleural Effusion from Non-Small-Cell Lung Cancer Patients by Mutant Enriched PCR Assay_Chinese Journal of Respiratory and Critical Care Medicine

Authors：

HE Chen , LIU Ming ,  XU Jun.

Guangzhou Institute of Respiratory Disease. First Affiliated Hospital of Guangzhou Medical College, State Key Lab of Respiratory Disease. Guangzhou, Guangdong, 510120, China Corresponding Author: XU Jun, E-mail: xufeili@ vip. 163. com;

Corresponding author：

XU Jun., Email: xufeili@vip.163.com

Keywords：

Mutant-enriched PCR assay; Pleural effusion; Epidermal growth factor receptor; Mutation; Non-small-cell lung cancer; Epidermal growth factor receptor tyrosine kinase inhibitor; Gefitinib

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Objective To investigate the feasibility of detection of epidermal growth factor receptor ( EGFR) exon 19 deletions and exon 21 L858R mutations in pleural effusion fromnon-small-cell lung cancer ( NSCLC) patients by mutant enriched PCR assay. Methods The mutations of exon 19 and 21 of EGFR gene in pleural samples fromthirty NSCLC patients were analyzed using both the mutant-enriched PCR assay and the non-enriched PCR assay. Results Ten ( 33. 3% , 10/ 30) exon 19 deletions and five ( 16. 7% , 5/30) exon 21 L858R mutation were detected by the mutant-enriched PCR assay, while only 6 cases ( 20. 0% ) and 1 case ( 3. 3% ) were detected by the non-enriched PCR assay respectively. The difference of mutation detection rate of EGFR gene between the two methods was statistically significant ( P = 0. 032) . Mutations were detected in all of partial responders ( 2 /4) among the four patients who received gefitinib therapy. Conclusions Mutant-enriched PCR assay can detect EGFR exon 19 deletions and exon 21 L858R mutation in pleural effusion from NSCLC patients effectively, economically and accurately. It may be a valuable biomarker for gefitinib therapy in advanced NSCLC.

Citation： HE Chen,LIU Ming,XU Jun.. Detection of EGFR Exon 19 and 21 Mutations in Pleural Effusion from Non-Small-Cell Lung Cancer Patients by Mutant Enriched PCR Assay. Chinese Journal of Respiratory and Critical Care Medicine, 2009, 09(5): 451-455. doi: Copy

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2.	Benlloch S, Galbis-Caravajal JM,Martin C, et al. Potential diagnostic value of methylation profile in pleural fluid and serum from cancer patients with pleural effusion. Cancer, 2006, 107: 1859-1865.
3.	Nakamoto M, Teramoto H, Matsumoto S, et al. K-ras and rho A mutations in malignant pleural effusion. Int J Oncol, 2001, 19 : 971 - 976.
4.	Lee JH, Hong YS, Ryu JS, et al. P53 and FHIT mutations and microsatellite alterations in malignancy-associated pleural effusion. Lung Cancer, 2004, 44: 33-42.
5.	Zhang X, Zhao Y, Wang M, et al. Detection and comparison of epidermal growth factor receptor mutations in cells and fluid of malignant pleural effusion in non-small cell lung cancer. Lung Cancer, 2008, 60: 175-182.
6.	Kahn SM, Jiang W, Culbertson TA, et al. Rapid and sensitive nonradioactive detection of mutant K-ras genes via ‘enriched’PCR amplification. Oncogene, 1991 , 6: 1079 -1083 .
7.	Chen J, Viola MV. A method to detect ras point mutations in small subpopulations of cells. Anal Biochem, 1991, 195: 51-56.
8.	Toyooka S, Tsukuda K, OuchidaM, et al. Detection of codon 61 point mutations of the K-ras gene in lung and colorectal cancers by enriched PCR. Oncol Rep, 2003, 10: 1455-1459.
9.	Asano H, Toyooka S, Tokumo M, et al. Detection of EGFR gene mutation in lung cancer by mutant-enriched polymerase chain reaction assay. Clin Cancer Res, 2006, 12: 43-48.
10.	Soh J, Toyooka S, Aoe K, et al. Usefulness of EGFR mutation screening in pleural fluid to predict the clinical outcome of gefitinib treated patients with lung cancer. Int J Cancer, 2006 , 119: 2353 - 2358.
11.	Kimura H, Fujiwara Y, Sone T, et al. EGFR mutation status in tumour-derived DNA from pleural effusion fluid is a practical basis for predicting the response to gefitinib. Br J Cancer, 2006, 95: 1390- 1395.
12.	Kimura H, Fujiwara Y, Sone T, et al. High sensitivity detection of epidermal growth factor receptor mutations in the pleural effusion of non-small cell lung cancer patients. Cancer Sci, 2006, 97: 642-648.
13.	Wu SG, Gow CH, Yu CJ, et al. Frequent epidermal growth factor receptor gene mutations in malignant pleural effusion of lung adenocarcinoma. Eur Respir J, 2008, 32: 924-930.
14.	Kimura H, Kasahara K, Kawaishi M, et al. Detection of epidermal growth factor receptor mutations in serum as a predictor of the response to gefitinib in patients with non-small-cell lung cancer. Clin Cancer Res, 2006, 12: 3915-3921.
15.	Kimura H, Suminoe M, Kasahara K, et al. Evaluation of epidermal growth factor receptor mutation status in serum DNA as a predictor of response to gefitinib( IRESSA) . Br J Cancer, 2007, 97: 778-784.
16.	董强刚, 黄进肃, 韩宝惠, 等. 晚期肺癌患者血清游离DNA 中 EGFR 外显子19 的基因突变研究. 肿瘤, 2006, 26: 59-63.

1. Sharma SV, Bell DW, Settleman J, et al. Epidermal growth factor receptor mutations in lung cancer. Nat Rev Cancer, 2007, 7: 169-181.
2. Benlloch S, Galbis-Caravajal JM,Martin C, et al. Potential diagnostic value of methylation profile in pleural fluid and serum from cancer patients with pleural effusion. Cancer, 2006, 107: 1859-1865.
3. Nakamoto M, Teramoto H, Matsumoto S, et al. K-ras and rho A mutations in malignant pleural effusion. Int J Oncol, 2001, 19 : 971 - 976.
4. Lee JH, Hong YS, Ryu JS, et al. P53 and FHIT mutations and microsatellite alterations in malignancy-associated pleural effusion. Lung Cancer, 2004, 44: 33-42.
5. Zhang X, Zhao Y, Wang M, et al. Detection and comparison of epidermal growth factor receptor mutations in cells and fluid of malignant pleural effusion in non-small cell lung cancer. Lung Cancer, 2008, 60: 175-182.
6. Kahn SM, Jiang W, Culbertson TA, et al. Rapid and sensitive nonradioactive detection of mutant K-ras genes via ‘enriched’PCR amplification. Oncogene, 1991 , 6: 1079 -1083 .
7. Chen J, Viola MV. A method to detect ras point mutations in small subpopulations of cells. Anal Biochem, 1991, 195: 51-56.
8. Toyooka S, Tsukuda K, OuchidaM, et al. Detection of codon 61 point mutations of the K-ras gene in lung and colorectal cancers by enriched PCR. Oncol Rep, 2003, 10: 1455-1459.
9. Asano H, Toyooka S, Tokumo M, et al. Detection of EGFR gene mutation in lung cancer by mutant-enriched polymerase chain reaction assay. Clin Cancer Res, 2006, 12: 43-48.
10. Soh J, Toyooka S, Aoe K, et al. Usefulness of EGFR mutation screening in pleural fluid to predict the clinical outcome of gefitinib treated patients with lung cancer. Int J Cancer, 2006 , 119: 2353 - 2358.
11. Kimura H, Fujiwara Y, Sone T, et al. EGFR mutation status in tumour-derived DNA from pleural effusion fluid is a practical basis for predicting the response to gefitinib. Br J Cancer, 2006, 95: 1390- 1395.
12. Kimura H, Fujiwara Y, Sone T, et al. High sensitivity detection of epidermal growth factor receptor mutations in the pleural effusion of non-small cell lung cancer patients. Cancer Sci, 2006, 97: 642-648.
13. Wu SG, Gow CH, Yu CJ, et al. Frequent epidermal growth factor receptor gene mutations in malignant pleural effusion of lung adenocarcinoma. Eur Respir J, 2008, 32: 924-930.
14. Kimura H, Kasahara K, Kawaishi M, et al. Detection of epidermal growth factor receptor mutations in serum as a predictor of the response to gefitinib in patients with non-small-cell lung cancer. Clin Cancer Res, 2006, 12: 3915-3921.
15. Kimura H, Suminoe M, Kasahara K, et al. Evaluation of epidermal growth factor receptor mutation status in serum DNA as a predictor of response to gefitinib( IRESSA) . Br J Cancer, 2007, 97: 778-784.
16. 董强刚, 黄进肃, 韩宝惠, 等. 晚期肺癌患者血清游离DNA 中 EGFR 外显子19 的基因突变研究. 肿瘤, 2006, 26: 59-63.

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