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. 2011 Jul;73(1):96-102.
doi: 10.1016/j.lungcan.2010.10.014. Epub 2010 Dec 3.

Detection of EGFR mutations in plasma DNA from lung cancer patients by mass spectrometry genotyping is predictive of tumor EGFR status and response to EGFR inhibitors

Marie Brevet  1 , Melissa L Johnson, Christopher G Azzoli, Marc Ladanyi
Affiliations

Detection of EGFR mutations in plasma DNA from lung cancer patients by mass spectrometry genotyping is predictive of tumor EGFR status and response to EGFR inhibitors

Marie Brevet et al. Lung Cancer. 2011 Jul.

Abstract

Aims: EGFR mutations now guide the clinical use of EGFR-targeted therapy in lung cancer. However, standard EGFR mutation analysis requires a minimum amount of tumor tissue, which may not be available in certain situations. In this study, we combined a mass spectrometry genotyping assay (Sequenom) with a mutant-enriched PCR (ME-PCR) to detect EGFR mutations in free plasma DNA from patients with lung cancer.

Method: DNAs were extracted from 31 plasma samples from 31 patients and analyzed by both methods for EGFR Exon 19 deletion and EGFR L858R mutation. Results in plasma DNA samples were compared with EGFR mutation status obtained in tumor DNA (18/31 EGFR mutant). The relationship of EGFR mutation status in tumor and/or plasma samples to overall survival was assessed.

Results: The EGFR mutation status in plasma DNA was identical to the primary tumor in 61% of patients (19/31). By mass spectrometry genotyping, the plasma samples contained mutant DNA corresponding to 5/14 EGFR Exon 19 deletions and 3/4 EGFR L858R mutations previously diagnosed in the matched tumors. Two samples were positive in plasma DNA but negative in primary tumor tissue. Results were similar for samples studied by ME-PCR. For patients treated with erlotinib, overall survival was correlated with the presence of EGFR mutation in plasma and/or tumor tissue (p=0.002), with the two patients positive only in plasma DNA showing responses and favorable outcomes.

Conclusion: The detection of EGFR mutations in plasma DNA samples by mass spectrometry genotyping and ME-PCR is feasible. A positive EGFR result in plasma DNA has a high predictive value for tumor EGFR status and for favorable clinical course on EGFR-targeted therapy and could therefore be useful in guiding clinical decisions in patients with insufficient or unavailable tumor specimens.

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Conflict of interest statement

Conflict of Interest statement: None declared

Figures

Figure 1
Figure 1
Sequenom assay results for EGFR exon 19 deletion in two plasma samples (patients Ex19-6 and Ex19-4) with the EGFR-M11R (A1 and A2), EGFR-M02F (B1 and B2) and EGFR-M03R (C1 and C2) assays. The control DNA provides a negative control for each assay and only the germline call is visible, indicated by an asterisk (A1, B1, C1). The mutant call is indicated by an arrow. The mass of the product is specific for each assay and is listed in table S1 (Supplementary file). Sample Ex19-6 shows a mutant call for the EGFR-M11R assay (A2) indicating a 9-bp deletion. Sample Ex19-4 shows a mutant call for the EGFR-M02F (B2) and the EGFR-M03R assays (B3) indicating a 15-bp deletion size.
Figure 2
Figure 2
Sequenom assay results for EGFR L858R mutations in two plasma samples (patients Ex21-1 and Ex21-2) with the EGFR-2573 assay. The control DNA provides a negative control and only the germline call is visible, indicated by an asterisk (a T call at 5402.6 Da). The mutant calls are visible at 5378.5 Da (G call) and are indicated by an arrow.
Figure 3
Figure 3
Overall survival curves for the 25 patients treated with erlotinib. Survival curves were evaluated by the Kaplan-Meier method and log-rank test. 3A “Patients with EGFR mutation in tumor DNA (n=18)” versus “Patients without EGFR mutation in tumor DNA (n=7)”, p=0.01. 3B “Patients with EGFR mutation in plasma or tumor DNA (n=20)” versus “Patients without EGFR mutation in either tumor or plasma DNA (n=5)”, p=0.002.
See this image and copyright information in PMC

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