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Clin Cancer Res. 2015 Aug 1;21(15):3552-60. doi: 10.1158/1078-0432.CCR-14-2151. Epub 2015 Apr 16.

Ultra-Sensitive Detection of the Pretreatment EGFR T790M Mutation in Non-Small Cell Lung Cancer Patients with an EGFR-Activating Mutation Using Droplet Digital PCR.

Author information

1
Department of Respiratory Medicine and Medical Oncology, National Hospital Organization Nagoya Medical Center, Aichi, Japan. Third Department of Internal Medicine, Wakayama Medical University, Kimiidera, Wakayama, Japan. Drug Discovery and Development Division, Shizuoka Cancer Center Research Institute, Shizuoka, Japan.
2
National Hospital Organization Kinki-Chuo Chest Medical Center, Osaka, Japan. Department of Respiratory Medicine, Graduate School of Medicine, Osaka City University, Osaka, Japan.
3
Clinical Research Center, Department of Lung Cancer Research, National Hospital Organization Kinki-chuo Chest Medical Center, Osaka, Japan.
4
Center for Advanced Medicine and Clinical Research, Nagoya University Hospital, Aichi, Japan.
5
Department of Internal Medicine, National Hospital Organization Kinki-chuo Chest Medical Center, Osaka, Japan.
6
Department of Respiratory Medicine and Medical Oncology, National Hospital Organization Nagoya Medical Center, Aichi, Japan. Division of Respiratory Medicine and Allergology, Aichi Medical University School of Medicine, Aichi, Japan.
7
Department of Respiratory Medicine and Medical Oncology, National Hospital Organization Nagoya Medical Center, Aichi, Japan.
8
Department of Thoracic Surgery, Clinical Research Institute, National Hospital Organization Kyushu Medical Center, Fukuoka, Japan.
9
Department of Thoracic Surgery, National Hospital Organization Hokkaido Cancer Center, Hokkaido, Japan.
10
Department of Thoracic Surgery, National Hospital Organization Nagasaki Medical Center, Nagasaki, Japan.
11
Department of Thoracic Surgery, National Hospital Organization Nishigunma National Hospital, Gunma, Japan.
12
Department of Thoracic Surgery, National Hospital Organization Shikoku Cancer Center, Ehime, Japan.
13
Department of Thoracic Surgery, National Hospital Organization Iwakuni Clinical Center, Yamaguchi, Japan.
14
Clinical Research Institute, National Kyushu Cancer Center, Fukuoka, Japan.
15
Department of General Thoracic Surgery, National Hospital Organization Toneyama National Hospital, Osaka, Japan.
16
Department of Respiratory Surgery, National Hospital Organization Yokohama Medical Center, Kanagawa, Japan.
17
Department of Surgery, National Hospital Organization Kinki-chuo Chest Medical Center, Osaka, Japan.
18
Department of Respiratory Medicine and Medical Oncology, National Hospital Organization Nagoya Medical Center, Aichi, Japan. Third Department of Internal Medicine, Wakayama Medical University, Kimiidera, Wakayama, Japan. Drug Discovery and Development Division, Shizuoka Cancer Center Research Institute, Shizuoka, Japan. ykoh@wakayama-med.ac.jp.

Abstract

PURPOSE:

The resistance to the EGFR tyrosine kinase inhibitors (TKI) is a major concern in non-small cell lung cancer (NSCLC) treatment. T790M mutation in EGFR accounts for nearly 50% of the acquired resistance to EGFR-TKIs. Earlier studies suggested that T790M mutation was also detected in TKI-naïve NSCLCs in a small cohort. Here, we use an ultra-sensitive droplet digital PCR (ddPCR) technique to address the incidence and clinical significance of pretreatment T790M in a larger cohort.

EXPERIMENTAL DESIGN:

ddPCR was established as follows: wild-type or T790M mutation-containing DNA fragments were cloned into plasmids. Candidate threshold was identified using wild-type plasmid, normal human genomic DNA, and human A549 cell line DNA, which expresses wild type. Surgically resected tumor tissues from 373 NSCLC patients with EGFR-activating mutations were then examined for the presence of T790M using ddPCR.

RESULTS:

Our data revealed a linear performance for this ddPCR method (R(2) = 0.998) with an analytical sensitivity of approximately 0.001%. The overall incidence of the pretreatment T790M mutation was 79.9% (298/373), and the frequency ranged from 0.009% to 26.9%. The T790M mutation was detected more frequently in patients with a larger tumor size (P = 0.019) and those with common EGFR-activating mutations (P = 0.022), as compared with the others.

CONCLUSIONS:

The ultra-sensitive ddPCR assay revealed that pretreatment T790M was found in the majority of NSCLC patients with EGFR-activating mutations. ddPCR should be utilized for detailed assessment of the impact of the low frequency pretreatment T790M mutation on treatment with EGFR-TKIs.

PMID:
25882755
DOI:
10.1158/1078-0432.CCR-14-2151
[Indexed for MEDLINE]
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