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Updated: Apr 8, 2026
Single Droplet Digital Polymerase Chain Reaction for Comprehensive and Simultaneous Detection of Mutations in Hotspot Regions
Published on: September 25, 2018
Detection of epidermal growth factor receptor mutation in lung cancer by droplet digital polymerase chain reaction
Qing Xu1, Yazhen Zhu2, Yali Bai1
1Translational Bioscience and Diagnostics, WuXi AppTec, Shanghai, People's Republic of China.
Background:
Two types of epidermal growth factor receptor (EGFR) mutations in exon 19 and exon 21 (ex19del and L858R) are prevalent in lung cancer patients and sensitive to targeted EGFR inhibition. A resistance mutation in exon 20 (T790M) has been found to accompany drug treatment when patients relapse. These three mutations are valuable companion diagnostic biomarkers for guiding personalized treatment. Quantitative polymerase chain reaction (qPCR)-based methods have been widely used in the clinic by physicians to guide treatment decisions. The aim of this study was to evaluate the technical and clinical sensitivity and specificity of the droplet digital polymerase chain reaction (ddPCR) method in detecting the three EGFR mutations in patients with lung cancer.
Methods:
Genomic DNA from H1975 and PC-9 cells, as well as 92 normal human blood specimens, was used to determine the technical sensitivity and specificity of the ddPCR assays. Genomic DNA of formalin-fixed, paraffin-embedded specimens from 78 Chinese patients with lung adenocarcinoma were assayed using both qPCR and ddPCR.
Results:
The three ddPCR assays had a limit of detection of 0.02% and a wide dynamic range from 1 to 20,000 copies measurement. The L858R and ex19del assays had a 0% background level in the technical and clinical settings. The T790M assay appeared to have a 0.03% technical background. The ddPCR assays were robust for correct determination of EGFR mutation status in patients, and the dynamic range appeared to be better than qPCR methods. The ddPCR assay for T790M could detect patient samples that the qPCR method failed to detect. About 49% of this patient cohort had EGFR mutations (L858R, 15.4%; ex19del, 29.5%; T790M, 6.4%). Two patients with the ex19del mutation also had a naïve T790M mutation.
Conclusion:
These data suggest that the ddPCR method could be useful in the personalized treatment of patients with lung cancer.
Insights
Droplet digital PCR (ddPCR) accurately detects EGFR mutations in lung cancer, outperforming qPCR for resistance mutations. This method aids personalized lung cancer treatment by identifying key biomarkers.
Area of Science:
- Molecular diagnostics
- Oncology
- Biomarker discovery
Background:
- Epidermal growth factor receptor (EGFR) mutations (ex19del, L858R) are common in lung cancer and targeted by therapy.
- The T790M resistance mutation emerges during treatment, complicating therapy.
- Accurate detection of these EGFR mutations is crucial for personalized lung cancer treatment.
Purpose of the Study:
- To evaluate the sensitivity and specificity of droplet digital PCR (ddPCR) for detecting EGFR mutations.
- To compare ddPCR with quantitative PCR (qPCR) in a clinical setting for lung cancer patients.
Main Methods:
- Genomic DNA from cell lines and normal blood specimens were used for technical validation.
- Formalin-fixed, paraffin-embedded tumor tissues from 78 lung adenocarcinoma patients were analyzed using both ddPCR and qPCR.
Main Results:
- ddPCR demonstrated high sensitivity (0.02% limit of detection) and a wide dynamic range.
- ddPCR accurately identified EGFR mutations, including T790M, which qPCR missed in some cases.
- EGFR mutations were found in 49% of the patient cohort, with specific frequencies for L858R, ex19del, and T790M.
Conclusions:
- ddPCR is a robust and sensitive method for detecting clinically relevant EGFR mutations in lung cancer.
- The ddPCR assay shows potential for improving personalized treatment strategies in lung cancer patients.
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