Tumor Nucleic Acid Detection Using Circularized Concatemers
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Solution Overview
Problem
Current methods for detecting tumor nucleic acids in cell-free biological samples face challenges in achieving low limit of detection (LOD) and high sensitivity, particularly in tumor-informed approaches, due to logistical complexities and high error rates in sequencing, which are costly and time-consuming.
Innovation Solution
The method involves circularizing nucleic acids from the sample to create concatemers, followed by amplification and sequencing at a controlled depth, combined with selection techniques to remove non-target sequences, utilizing polymerases with strand-displacement activity, and employing concatemer sequencing for error correction.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional sequencing methods are used for tumor nucleic acid detection, then sequencing depth can be increased to improve detection sensitivity, but cost and turnaround time increase significantly
Solution Approach 1:
The patent applies preliminary action by performing circularization of nucleic acids before sequencing. This pre-processing step creates concatemers that can be sequenced at lower depth while maintaining detection sensitivity, thereby reducing turnaround time and cost without sacrificing measurement precision.
Solution Approach 2:
The patent changes the parameter of nucleic acid structure from linear to circular form through circularization. This structural parameter change enables the use of lower sequencing depth (reading 1-5x coverage instead of conventional higher depth) while maintaining or improving detection sensitivity through concatemer-based error correction.
2Measurement precision
If conventional sequencing methods are used for tumor nucleic acid detection, then sequencing depth can be increased to improve detection sensitivity, but cost increases significantly
Solution Approach 1:
The patent applies preliminary action by performing circularization of nucleic acids before sequencing. This pre-processing step creates concatemers that can be sequenced at lower depth while maintaining detection sensitivity, thereby reducing turnaround time and cost without sacrificing measurement precision.
Solution Approach 2:
The patent changes the parameter of nucleic acid structure from linear to circular form through circularization. This structural parameter change enables the use of lower sequencing depth (reading 1-5x coverage instead of conventional higher depth) while maintaining or improving detection sensitivity through concatemer-based error correction.
3Measurement precision
If high sequencing depth is used to detect tumor-specific variants, then detection accuracy improves, but error rates in sequencing increase
Solution Approach 1:
The patent applies preliminary action by performing circularization of nucleic acids before sequencing. This pre-processing step creates concatemers that can be sequenced at lower depth while maintaining detection sensitivity, thereby reducing turnaround time and cost without sacrificing measurement precision.
Solution Approach 2:
The patent uses copying through concatemer formation, where multiple copies of the circularized nucleic acid are generated and sequenced. This copying approach allows error correction by comparing multiple reads of the same sequence, improving reliability without requiring extremely high sequencing depth.
4Productivity
If shallow sequencing is used to reduce cost and time, then detection sensitivity decreases, but the method becomes cost-effective and fast
Solution Approach 1:
The patent changes the parameter of nucleic acid structure from linear to circular form through circularization. This structural parameter change enables the use of lower sequencing depth (reading 1-5x coverage instead of conventional higher depth) while maintaining or improving detection sensitivity through concatemer-based error correction.
Solution Approach 2:
The patent uses copying through concatemer formation, where multiple copies of the circularized nucleic acid are generated and sequenced. This copying approach allows error correction by comparing multiple reads of the same sequence, improving reliability without requiring extremely high sequencing depth.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This approach enables efficient and cost-effective genome-wide error suppression, allowing for sensitive and fast detection of tumor-specific sequence variants, even at low tumor burden, with improved LOD and reduced turnaround time.
Implementation Method 1
amplifying the circularized nucleic acid is effected by a polymerase having strand-displacement activity
Implementation Method 2
amplifying the circularized nucleic acid is effected by a polymerase having 5′ to 3′ exonuclease activity
Implementation Method 3
circularizing comprises ligating ends of the nucleic acid or a derivative thereof to one another
Data Source
AI summary
Provided herein are methods of detecting tumor nucleic acids in a biological sample of a subject.