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Updated: Feb 28, 2026
Next Generation Sequencing for the Detection of Actionable Mutations in Solid and Liquid Tumors
Published on: September 20, 2016
Most T790M mutations are present on the same EGFR allele as activating mutations in patients with non-small cell lung
Noriko Hidaka1, Eiji Iwama2, Naoki Kubo3
1Research Institute for Diseases of the Chest, Graduate School of Medical Sciences, Kyushu University, Fukuoka 8128582, Japan.
Objectives:
The T790M and C797S mutations of the epidermal growth factor receptor gene (EGFR) confer resistance to first- and third-generation EGFR tyrosine kinase inhibitors (TKIs), respectively, in patients with non-small cell lung cancer (NSCLC) harboring activating mutations of EGFR. C797S has been identified in cis or in trans with T790M in tumor specimens from patients who experienced treatment failure with first- and third-generation EGFR-TKIs. The allelic relation between T790M and activating mutations of EGFR has not been well characterized, however. We have now developed a digital polymerase chain reaction (dPCR)-based method for determination of the allelic relation between two types of EGFR mutation (T790M and either C797S or an activating mutation).
Materials And Methods:
Seven clinical NSCLC specimens and two NSCLC cell lines harboring both an activating mutation and T790M were analyzed with this new method to identify the allelic relation between these EGFR mutations.
Results:
The median ratio of the number of alleles positive for both an activating mutation and T790M to the number of T790M-positive alleles was 97.1% (range, 90.0-100%). Confirmatory analysis by next-generation sequencing yielded a corresponding value of 96.7% (range, 89.1-99.5%). Our dPCR method thus reliably identifies the allelic relation between two EGFR mutations in a quantitative manner.
Conclusions:
Almost all T790M mutations were detected in cis with activating mutations of EGFR regardless of the de novo or acquired status of T790M, with cancer cells harboring T790M and activating mutations on the same allele appearing to be selected and enriched during EGFR-TKI treatment.
Insights
A new digital PCR method reliably determines the allelic relationship between epidermal growth factor receptor (EGFR) mutations, T790M and activating mutations. This finding is crucial for understanding non-small cell lung cancer (NSCLC) treatment resistance.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Epidermal growth factor receptor (EGFR) mutations drive non-small cell lung cancer (NSCLC).
- EGFR tyrosine kinase inhibitors (TKIs) are standard treatment, but resistance develops via mutations like T790M and C797S.
- The allelic relationship between T790M and activating EGFR mutations is not well understood, impacting treatment strategies.
Purpose of the Study:
- To develop and validate a digital polymerase chain reaction (dPCR)-based method for determining the allelic relationship between EGFR mutations.
- To investigate the allelic status of T790M in relation to activating EGFR mutations in NSCLC.
Main Methods:
- Development of a novel digital PCR (dPCR) assay.
- Analysis of seven clinical NSCLC specimens and two NSCLC cell lines.
- Quantification of EGFR mutation allelic ratios and validation with next-generation sequencing.
Main Results:
- The dPCR method accurately quantified the allelic relationship between EGFR T790M and activating mutations.
- A median ratio of 97.1% (range, 90.0-100%) was observed for T790M mutations occurring in cis with activating mutations.
- Next-generation sequencing confirmed these findings with a median ratio of 96.7% (range, 89.1-99.5%).
Conclusions:
- The T790M mutation is predominantly found in cis with activating EGFR mutations in NSCLC.
- This allelic configuration appears to be selected and enriched during EGFR-TKI therapy.
- The developed dPCR method provides a reliable tool for assessing EGFR mutation allelic status in NSCLC.
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