Hybridization-based sensor with large dynamic range for detection of circulating tumor DNA in clinical samples

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.