| Author: | Shek, Chiu Man |
| Title: | A novel digital PCR assay for accurate detection and differentiation of focal and non-focal subtypes of mesenchymal-epithelial transition (MET) gene amplification in lung cancer |
| Advisors: | Siu, Gilman Kit-hang (HTI) |
| Degree: | DHSc |
| Year: | 2025 |
| Department: | Faculty of Health and Social Sciences |
| Pages: | xv, 86 pages : color illustrations |
| Language: | English |
| Abstract: | The Mesenchymal–epithelial transition (MET) gene amplification is a critical molecular biomarker in non-small cell lung cancer (NSCLC), playing a pivotal role in treatment decisions and prognostic evaluations. MET amplification drives tumor progression, metastasis, and resistance to targeted therapies, particularly in patients with EGFR-mutant NSCLC who develop resistance to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs). Accurate detection of MET amplification, especially the differentiation between focal amplification (a therapeutically actionable biomarker) and polysomy (a non-actionable finding), is essential for guiding MET-targeted therapies such as capmatinib and tepotinib. However, current diagnostic methods, including fluorescence in situ hybridization (FISH) and next-generation sequencing (NGS), face significant limitations in speed, cost, and specificity, hindering their widespread clinical utility. This study introduces a novel digital PCR (dPCR) assay designed to address these limitations by providing a rapid, cost-effective, and objective method for MET amplification detection and subtype differentiation. The assay was developed and validated using 55 NSCLC samples with known MET amplification statuses (26 positive and 29 negative), as confirmed by FISH and NGS. The dPCR assay demonstrated exceptional diagnostic performance, achieving a sensitivity of 96.0% and specificity of 96.7%. Notably, it showed 100% concordance with FISH in differentiating focal MET amplification from polysomy, a critical distinction for therapy selection. Furthermore, the assay exhibited excellent precision, accuracy, and linearity (R² = 0.9951) in MET copy number quantification, surpassing NGS in diagnostic reliability. One of the key advantages of the dPCR assay is its significantly reduced turnaround time (TAT). While FISH requires at least 2 days for processing and analysis, the dPCR assay delivers results in approximately 3 hours, enabling timely clinical decision-making. Additionally, the assay is highly cost-effective, with reagent costs of approximately USD$60 per test, compared to USD$250 for FISH and USD$650 for NGS. This cost efficiency stems from its simplified workflow, reduced hands-on time, and minimal reliance on specialized expertise or infrastructure. The dPCR assay's ability to provide quantitative and objective results makes it particularly suitable for clinical laboratories with limited molecular expertise. Its robustness and reproducibility were further validated through rigorous cross-reactivity checks and performance comparisons with FISH and NGS. While rare discordant cases near MET copy number (CN) cutoffs highlight the value of complementary testing with FISH or NGS in ambiguous scenarios, the assay's overall performance underscores its potential as a standalone diagnostic tool. In conclusion, this study highlights the transformative potential of the dPCR assay in advancing precision oncology. By enabling timely, accurate, and cost-effective detection of MET amplification and subtype differentiation, the assay addresses critical gaps in current diagnostic workflows. Future studies should focus on validating the assay in larger cohorts, refining MET CN thresholds, and correlating results with therapeutic outcomes to fully establish its clinical utility. This work represents a significant step forward in optimizing treatment strategies for NSCLC patients, ultimately improving patient outcomes in the era of personalized medicine. |
| Rights: | All rights reserved |
| Access: | restricted access |
Files in This Item:
| File | Description | Size | Format | |
|---|---|---|---|---|
| 8654.pdf | For All Users (off-campus access for PolyU Staff & Students only) | 1.84 MB | Adobe PDF | View/Open |
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