Highly sensitive and quantitative evaluation of the EGFR T790M mutation by nanofluidic digital PCR
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Eiji Iwama1,2, Koichi Takayama2, Taishi Harada2, Isamu Okamoto3, Fumihiko Ookubo4, Junji Kishimoto5, Eishi Baba1, Yoshinao Oda6 and Yoichi Nakanishi2,3
1 Faculty of Medical Sciences, Department of Comprehensive Clinical Oncology, Kyushu University, Fukuoka, Japan
2 Research Institute for Diseases of the Chest, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan
3 Center for Clinical and Translational Research, Kyushu University Hospital, Fukuoka, Japan
4 Division of Diagnostic Pathology, Kyushu University Hospital, Fukuoka, Japan
5 Department of Research and Development of Next Generation Medicine, Kyushu University, Fukuoka, Japan
6 Department of Anatomic Pathology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan
Eiji Iwama, email:
Keywords: EGFR, T790M, digital PCR, highly sensitive detection, quantification
Received: January 09, 2015 Accepted: April 20, 2015 Published: May 10, 2015
The mutation of T790M in EGFR is a major mechanism of resistance to treatment with EGFR-TKIs. Only qualitative detection (presence or absence) of T790M has been described to date, however. Digital PCR (dPCR) analysis has recently been applied to the quantitative detection of target molecules in cancer with high sensitivity. In the present study, 25 tumor samples (13 obtained before and 12 after EGFR-TKI treatment) from 18 NSCLC patients with activating EGFR mutations were evaluated for T790M with dPCR. The ratio of the number of T790M alleles to that of activating mutation alleles (T/A) was determined. dPCR detected T790M in all 25 samples. Although T790M was present in all pre-TKI samples from 13 patients, 10 of these patients had a low T/A ratio and manifested substantial tumor shrinkage during treatment with EGFR-TKIs. In six of seven patients for whom both pre- and post-TKI samples were available, the T/A ratio increased markedly during EGFR-TKI treatment. Highly sensitive dPCR thus detected T790M in all NSCLC patients harboring activating EGFR mutations whether or not they had received EGFR-TKI treatment. Not only highly sensitive but also quantitative detection of T790M is important for evaluation of the contribution of T790M to EGFR-TKI resistance.
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