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. 2020 Oct 16;12(10):3002.
doi: 10.3390/cancers12103002.

Mass Spectrometry as a Highly Sensitive Method for Specific Circulating Tumor DNA Analysis in NSCLC: A Comparison Study

Pierre-Jean Lamy  1 , Paul van der Leest  2 , Nicolas Lozano  1 , Catherine Becht  3 , Frédérique Duboeuf  3 , Harry J M Groen  4 , Werner Hilgers  5 , Nicolas Pourel  5 , Naomi Rifaela  2 , Ed Schuuring  2 , Catherine Alix-Panabières  6
Affiliations

Mass Spectrometry as a Highly Sensitive Method for Specific Circulating Tumor DNA Analysis in NSCLC: A Comparison Study

Pierre-Jean Lamy et al. Cancers (Basel). .

Abstract

Plasma-based tumor mutational profiling is arising as a reliable approach to detect primary and therapy-induced resistance mutations required for accurate treatment decision making. Here, we compared the FDA-approved Cobas® EGFR Mutation Test v2 with the UltraSEEK™ Lung Panel on the MassARRAY® System on detection of EGFR mutations, accompanied with preanalytical sample assessment using the novel Liquid IQ® Panel. 137 cancer patient-derived cell-free plasma samples were analyzed with the Cobas® and UltraSEEK™ tests. Liquid IQ® analysis was initially validated (n = 84) and used to determine ccfDNA input for all samples. Subsequently, Liquid IQ® results were applied to harmonize ccfDNA input for the Cobas® and UltraSEEK™ tests for 63 NSCLC patients. The overall concordance between the Cobas® and UltraSEEK™ tests was 86%. The Cobas® test detected more EGFR exon19 deletions and L858R mutations, while the UltraSEEK™ test detected more T790M mutations. A 100% concordance in both the clinical (n = 137) and harmonized (n = 63) cohorts was observed when >10 ng of ccfDNA was used as determined by the Liquid IQ® Panel. The Cobas® and UltraSEEK™ tests showed similar sensitivity in EGFR mutation detection, particularly when ccfDNA input was sufficient. It is recommended to preanalytically determine the ccfDNA concentration accurately to ensure sufficient input for reliable interpretation and treatment decision making.

Keywords: EGFR mutation; circulating DNA; liquid biopsy; non-small cell lung cancer; tyrosine kinase inhibitors.

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Conflict of interest statement

P.-J.L. is consultant in advisory boards for Agena Bioscience, AstraZeneca, BMS, Foundation medicine, Lilly, Novartis and Roche, and received research grants from Agena Bioscience, Beckman Coulter and Cepheid. E.S. has performed lectures for Bio-Rad, Novartis, Roche, Biocartis, Illumina, Pfizer, AstraZeneca, and Agena Bioscience, is consultant in advisory boards for AstraZeneca, Roche, Pfizer, Novartis, Bayer, Lilly, BMS, Amgen, BioCartis, Illumina, Agena Bioscience and MSD/Merck, and received research grants from Pfizer, Biocartis, Agena Bioscience, BMS, Bio-Rad, Roche, Boehringer Ingelheim. N.R. is employed by the Dutch Cancer Society (KWF, Alpe D’HuZes research grant RUG 2013-6355). The other authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Comparison of mutation detection between UltraSEEK™ and Cobas®. Pie charts representing the samples which were concordant between UltraSEEK™ and Cobas® (blue), samples in which UltraSEEK™ detected more mutations (green), and samples in which Cobas® detected more mutations (red) for (A) the mutations detectable on both panels and (B) all clinically relevant mutations across BRAF, EGFR, KRAS, ERBB2 and PIK3CA. (C) Bar graph illustrating the total amount of mutations detected in all 137 samples. The blue bar represents the number of mutations that could be detected on both panels; the green bar represents detected mutations unique for the UltraSEEK™ Panel. One EGFR exon19 deletion genotype was detected with Cobas® that is not represented on the UltraSEEK™ Panel (red bar). (D) Bar graph illustrating frequency of detection per mutation with UltraSEEK™ (green) and Cobas (red).
Figure 2
Figure 2
Correlation between Liquid IQ® and Qubit™ results of Streck and EDTA plasma samples. The correlation between Liquid IQ® and Qubit™ for the ccfDNA extracted from EDTA (black; R2 = 0.91) and Streck (red; R2 = 0.82) separately showed similar results.
Figure 3
Figure 3
Correlation of Liquid IQ® and a secondary DNA quantification assay. Correlation of the ccfDNA concentration as determined by the amplifiable copies measured with Liquid IQ® and the quantification based on (A) LabChip® performed on the samples of the MTP cohort and (B) Qubit™ performed on the samples of the GRO cohort. Red-colored results are not included in the correlation due to failure of either quantification assay or was considered an outlier.
Figure 4
Figure 4
Comparison of mutation detection between UltraSEEK™ and Cobas® with harmonized sample input. Pie charts representing the samples which were concordant between UltraSEEK™ and Cobas® (blue), samples in which UltraSEEK™ detected more mutations (green), samples in which Cobas® detected more mutations (red), and sample of which the Cobas® analysis was invalid supposedly due to insufficient input (purple) for (A) the mutations detectable on both panels and (B) all clinically relevant mutations. (C) Bar graph illustrating the total amount of mutations detected in all samples. The blue bar represents the number of mutations that could be detected on both panels; the green bar represents detected mutations unique for the UltraSEEK™ Panel. No mutations only covered by the Cobas® Panel were detected (red bar). (D) Bar graph illustrating frequency of detection per mutation with UltraSEEK™ (green) and Cobas (red).
Figure 5
Figure 5
Flowchart of data collection. * Reference material from Horizon Discovery, Waterbeach, UK. † Reference material from SeraCare Life Sciences Inc, Milford, MA, USA.
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