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Detection of Rare Mutations in CtDNA Using Next Generation Sequencing
Published on: August 24, 2017
A Prospective Evaluation of Circulating Tumor Cells and Cell-Free DNA in EGFR-Mutant Non-Small Cell Lung Cancer
Masahiko Yanagita1,2, Amanda J Redig1,2,3, Cloud P Paweletz1,4
1Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts.
Purpose:
Genotype-directed therapy is the standard of care for advanced non-small cell lung cancer (NSCLC), but obtaining tumor tissue for genotyping remains a challenge. Circulating tumor cell (CTC) or cell-free DNA (cfDNA) analysis may allow for noninvasive evaluation. This prospective trial evaluated CTCs and cfDNA in EGFR-mutant NSCLC patients treated with erlotinib until progression.
Experimental Design:
EGFR-mutant NSCLC patients were enrolled in a phase II trial of erlotinib. Blood was collected at baseline, every 2 months on study, and at disease progression. Plasma genotyping was performed by droplet digital PCR for EGFR19del, L858R, and T790M. CTCs were isolated by CellSave, enumerated, and analyzed by immunofluorescence for CD45 and pan-cytokeratin and EGFR and MET FISH were also performed. Rebiopsy was performed at disease progression.
Results:
Sixty patients were enrolled; 44 patients discontinued therapy for disease progression. Rebiopsy occurred in 35 of 44 patients (80%), with paired CTC/cfDNA analysis in 41 of 44 samples at baseline and 36 of 44 samples at progression. T790M was identified in 23 of 35 (66%) tissue biopsies and 9 of 39 (23%) cfDNA samples. CTC analysis at progression identified MET amplification in 3 samples in which tissue analysis could not be performed. cfDNA analysis identified T790M in 2 samples in which rebiopsy was not possible. At diagnosis, high levels of cfDNA but not high levels of CTCs correlated with progression-free survival.
Conclusions:
cfDNA and CTCs are complementary, noninvasive assays for evaluation of acquired resistance to first-line EGFR TKIs and may expand the number of patients in whom actionable genetic information can be obtained at acquired resistance. Serial cfDNA monitoring may offer greater clinical utility than serial monitoring of CTCs. Clin Cancer Res; 22(24); 6010-20. ©2016 AACR.
Insights
Cell-free DNA (cfDNA) and circulating tumor cells (CTCs) offer noninvasive evaluation of resistance to EGFR TKIs in non-small cell lung cancer. Serial cfDNA monitoring shows greater clinical utility than CTC monitoring for treatment resistance.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Genotype-directed therapy is standard for advanced non-small cell lung cancer (NSCLC).
- Obtaining tumor tissue for genotyping is challenging.
- Circulating tumor cells (CTCs) and cell-free DNA (cfDNA) offer noninvasive alternatives.
Purpose of the Study:
- To evaluate CTCs and cfDNA in EGFR-mutant NSCLC patients treated with erlotinib.
- To assess the utility of these markers in identifying acquired resistance.
- To compare the clinical utility of serial cfDNA and CTC monitoring.
Main Methods:
- Prospective phase II trial of erlotinib in EGFR-mutant NSCLC patients.
- Blood collection at baseline, every 2 months, and at progression.
- Plasma genotyping (EGFR19del, L858R, T790M) by droplet digital PCR.
- CTC isolation, enumeration, and immunofluorescence analysis.
- EGFR and MET FISH performed.
- Rebiopsy at disease progression.
Main Results:
- T790M mutation detected in 66% of tissue biopsies vs. 23% of cfDNA samples.
- CTC analysis identified MET amplification in cases where tissue analysis was not possible.
- cfDNA identified T790M in cases where rebiopsy was not possible.
- High cfDNA levels at diagnosis correlated with progression-free survival, unlike CTC levels.
Conclusions:
- cfDNA and CTCs are complementary noninvasive assays for evaluating acquired resistance to EGFR TKIs.
- These assays can increase the number of patients with actionable genetic information at acquired resistance.
- Serial cfDNA monitoring may provide greater clinical utility than serial CTC monitoring.
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