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Published on: September 25, 2018
Multiplex Droplet Digital PCR Assay for Detection of MET and HER2 Genes Amplification in Non-Small Cell Lung Cancer
Igor P Oscorbin1, Maria A Smertina1, Ksenia A Pronyaeva1,2
1Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences (ICBFM SB RAS), 630090 Novosibirsk, Russia.
Abstract:
Non-small-cell lung cancer (NSCLC), a subtype of lung cancer, remains one of the most common tumors with a high mortality and morbidity rate. Numerous targeted drugs were implemented or are now developed for the treatment of NSCLC. Two genes, HER2 and MET, are among targets for these specific therapeutic agents. Alterations in HER2 and MET could lead to primary or acquired resistance to commonly used anti-EGFR drugs. Using current methods for detecting HER2 and MET amplifications is time and labor-consuming; alternative methods are required for HER2 and MET testing. We developed the first multiplex droplet digital PCR assay for the simultaneous detection of MET and HER2 amplification in NSCLC samples. The suitability of qPCR was assessed for the optimization of multiplex ddPCR. The optimal elongation temperature, reference genes for DNA quantification, and amplicon length were selected. The developed ddPCR was validated on control samples with various DNA concentrations and ratios of MET and HER2 genes. Using ddPCR, 436 EGFR-negative NSCLC samples were analyzed. Among the tested samples, five specimens (1.15%) showed a higher ratio of MET, and six samples (1.38%) showed a higher ratio of HER2. The reported multiplex ddPCR assay could be used for the routine screening of MET and HER2 amplification in NSCLC samples.
Insights
A new multiplex droplet digital PCR assay allows simultaneous detection of MET and HER2 gene amplification in non-small-cell lung cancer (NSCLC) patients. This method offers a faster alternative for identifying potential resistance to targeted therapies.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Non-small-cell lung cancer (NSCLC) is a leading cause of cancer-related deaths worldwide.
- Targeted therapies, including those against HER2 and MET, are crucial for NSCLC treatment.
- HER2 and MET alterations can confer resistance to anti-EGFR drugs, necessitating accurate detection methods.
Purpose of the Study:
- To develop and validate a novel multiplex droplet digital PCR (ddPCR) assay for simultaneous detection of MET and HER2 amplification in NSCLC.
- To establish an efficient and accurate method for identifying gene amplifications that may predict or cause resistance to targeted therapies.
Main Methods:
- Development of a multiplex ddPCR assay for simultaneous detection of MET and HER2 amplification.
- Optimization of the ddPCR assay using quantitative PCR (qPCR) for parameters like elongation temperature and reference genes.
- Validation of the ddPCR assay on control samples with varying DNA concentrations and gene ratios.
- Analysis of 436 EGFR-negative NSCLC samples using the developed ddPCR assay.
Main Results:
- The study successfully developed and validated a multiplex ddPCR assay for simultaneous MET and HER2 amplification detection.
- MET amplification was detected in 1.15% (5/436) of EGFR-negative NSCLC samples.
- HER2 amplification was detected in 1.38% (6/436) of EGFR-negative NSCLC samples.
Conclusions:
- The developed multiplex ddPCR assay is a suitable and efficient method for routine screening of MET and HER2 amplification in NSCLC.
- This assay can aid in identifying NSCLC patients who may benefit from or develop resistance to specific targeted therapies.
- The findings support the clinical utility of ddPCR for precise genetic profiling in NSCLC management.
