Liquid Biopsy bTMB Detection with ctFE Threshold Filtering
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Solution Overview
Problem
Conventional liquid biopsy assays struggle to accurately determine tumor mutational burden (TMB) due to the diluted nature of circulating tumor DNA and the use of small targeted-panels that enrich genomic regions, leading to inaccuracies in determining blood TMB (bTMB) for precision oncology.
Innovation Solution
A method and system for determining bTMB using a panel-enriched sequencing reaction with a threshold circulating tumor fraction (ctFE) and filtering criteria to identify relevant mutations, incorporating a panel that enriches for up to 150 genes and performs sequencing at a read depth of at least 1,000X, followed by normalization and reporting of bTMB.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If small targeted-panel sequencing is used to enrich genomic regions, then sequencing depth and cost-effectiveness are improved, but measurement precision of tumor mutational burden deteriorates due to diluted circulating tumor DNA
Solution Approach 1:
The patent applies parameter changes by implementing a threshold circulating tumor fraction (ctFE) criterion and adjusting sequencing depth parameters (minimum 100x average depth) to optimize the balance between productivity and measurement precision. The method transforms the approach by changing the parameters for calculating bTMB based on ctFE thresholds, thereby improving accuracy while maintaining the efficiency of targeted-panel sequencing.
Solution Approach 2:
The patent replaces the conventional mechanical/enumerative counting method with a computational substitution approach. Instead of directly counting mutations without filters, the system uses ctFE threshold calculations and computational algorithms to identify and count only relevant mutations, substituting physical enumeration with intelligent computational filtering to resolve the accuracy-efficiency contradiction.
2Reliability
If comprehensive genomic evaluation is performed to improve precision oncology outcomes, then treatment matching accuracy is improved, but device complexity and assay cost increase
Solution Approach 1:
The patent extracts only the essential and relevant mutations needed for treatment matching by applying ctFE thresholds and filtering criteria. Instead of analyzing all genomic data comprehensively, the method selectively extracts mutations that meet specific criteria (ctFE ≥ threshold), thereby maintaining high treatment matching accuracy while reducing assay complexity and cost.
Solution Approach 2:
The patent applies partial action by performing sequencing and analysis at a controlled depth (minimum 100x average depth) rather than exhaustive whole-genome sequencing. This partial approach focuses resources on the most informative regions and mutations, achieving sufficient reliability for treatment matching without the excessive complexity and cost of complete genomic evaluation.
3Ease of operation
If liquid biopsy assay is used to minimize invasiveness, then ease of operation and patient comfort are improved, but measurement precision of TMB deteriorates due to diluted circulating tumor DNA
Solution Approach 1:
The patent introduces ctFE (circulating tumor fraction) as an intermediary parameter to bridge the gap between liquid biopsy simplicity and TMB accuracy. The ctFE serves as a mediator that quantifies the diluted tumor DNA signal, allowing the system to adjust calculations and apply appropriate thresholds to compensate for the dilution effect, thereby maintaining measurement precision while preserving the ease of liquid biopsy operation.
Solution Approach 2:
The patent implements a feedback mechanism where ctFE measurements inform subsequent bTMB calculations. The ctFE value feeds back into the analysis pipeline to determine appropriate thresholds and adjustment factors, creating a closed-loop system that continuously optimizes TMB determination accuracy based on the actual circulating tumor DNA levels detected in each patient sample.
Data Source
Figure 1A
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Figure 1C~1D
AI summary
Systems and methods for determining a blood tumor mutational burden (bTMB) for a test subject are provided in which there is obtained, from a panel-enriched sequencing reaction, a plurality of nucleic acid sequences. The plurality of nucleic acid sequences comprises a corresponding sequence for each cell-free DNA fragment in a plurality of cell-free DNA fragments obtained from a liquid biopsy sample from the test subject. Each respective cell-free DNA fragment in the plurality of cell-free DNA fragments corresponds to a respective probe sequence in a plurality of probe sequences used to enrich cell-free DNA fragments in the liquid biopsy sample in the panel-enriched sequencing reaction. There is deteremined, using the panel-enriched sequencing reaction, that a circulating tumor fraction (ctFE) is above a threshold ctFE value. Responsive to this determination, the bTMB is calculated for the test subject from the panel-enriched sequencing reaction and reported.