Plasma-based early screening and monitoring of EGFR mutations in NSCLC patients by a 3-color digital PCR assay

Noninvasive plasma-based detection of EGFR mutations using digital PCR promises a fast, sensitive and reliable approach to predicting the efficiency of EGFR-TKI. However, the low throughput and high cost of digital PCR restricts its clinical application. We designed a digital PCR assay, which can simultaneously detect 39 mutations of exons 18–21 of the EGFR gene. To assess overall performance, retrospective FFPE tissues from 30 NSCLC patients and plasma from 33 NSCLC patients were collected and analysed. The LoD of the EGFR mutations was as low as 0.308 copies/μL, and the linear correlation between the detected and expected values at different concentrations (0.01–10%) was low as well. Compared to ARMS-PCR in FFPE, the accuracy values of the dEGFR39 assay in plasma from 33 patients was 87.88% (29/33, 95% CI 72.67–95.18%). While monitoring the 33 patients, the EGFR mutation load as assessed by dEGFR39 was associated with the objective response to treatment. Thirteen samples from eight patients were identified by dEGFR39 to harbour the T790M mutation over time; of these patients, only nine (69%) were detected using SuperARMS. Our results indicate that dEGFR39 assay is reliable, sensitive and cost-efficient. This method is beneficial for profiling EGFR mutations for precision therapy and prognosis after TKI treatment, especially in patients with insufficient tissue biopsy samples.