Hybridization-based sensor with large dynamic range for detection of circulating tumor DNA in clinical samples
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Updated: May 14, 2025
Detection and Monitoring of Tumor Associated Circulating DNA in Patient Biofluids
Published on: June 8, 2019
Yannick Stulens1, Rebekka Van Hoof2, Karen Hollanders3
1UHasselt, Data Science Institute, Theory Lab, Agoralaan, Diepenbeek, 3590, Belgium.
Abstract:
In a liquid biopsy approach, targeted mutation analysis of circulating tumor DNA (ctDNA) is a valuable tool for diagnosis, monitoring and personalization of therapy. The ctDNA usually makes up only a small fraction of the total circulating free DNA (cfDNA), and ctDNA often only differs from cfDNA at a single nucleotide. This sets strong requirements on the analytical performance of hybridization-based biosensors, which is the focus of this paper. We use clinical samples and apply the concept of wild-type target depletion. Along with this, we develop an accurate thermodynamic theory for the competitive hybridization and use it for selecting optimal experimental conditions and for data analysis. The result is a biosensor with improved quantification of ctDNA mutations, both the sensitivity and dynamic range are improved by an order of magnitude. As reference techniques, we used a clinically-validated real-time PCR assay and digital PCR for absolute quantification. Our approach can be applied to a broad range of hybridization-based biosensors, providing a robust and effective method to improve the performance of existing biosensors.
Insights
This study enhances liquid biopsy by improving circulating tumor DNA (ctDNA) detection. Wild-type target depletion and thermodynamic theory boost biosensor sensitivity and dynamic range for accurate cancer mutation analysis.
Area of Science:
- Biotechnology
- Molecular Diagnostics
- Genomics
Background:
- Liquid biopsies offer non-invasive cancer detection and monitoring via circulating tumor DNA (ctDNA).
- ctDNA constitutes a small fraction of cell-free DNA (cfDNA), often differing by a single nucleotide, posing analytical challenges.
- Hybridization-based biosensors require high analytical performance for accurate ctDNA mutation detection.
Purpose of the Study:
- To improve the analytical performance of hybridization-based biosensors for ctDNA mutation quantification.
- To enhance the sensitivity and dynamic range of ctDNA detection in liquid biopsies.
- To develop a robust method applicable to various hybridization-based biosensing platforms.
Main Methods:
- Utilized clinical samples for analysis.
- Implemented wild-type target depletion strategy.
- Developed and applied a thermodynamic theory for competitive hybridization.
- Optimized experimental conditions and data analysis based on the theory.
Main Results:
- Achieved an order of magnitude improvement in both sensitivity and dynamic range for ctDNA mutation quantification.
- Demonstrated enhanced performance compared to clinically-validated real-time PCR and digital PCR assays.
- Validated the approach using clinical samples.
Conclusions:
- The developed method significantly improves ctDNA quantification in liquid biopsies.
- Wild-type target depletion combined with thermodynamic theory offers a robust solution for enhancing biosensor performance.
- This approach is broadly applicable to various hybridization-based biosensors for improved clinical diagnostics.
