Digital PCR Biomarker Panel for MSI Detection
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Solution Overview
Problem
Current MSI testing methods for cancers, such as colorectal, gastric, and endometrial tumors, are complex, time-consuming, and costly, with limited sensitivity and reproducibility, requiring specialized equipment and skilled personnel, which hinders their adoption as routine diagnostic tools.
Innovation Solution
A novel diagnostic marker panel using short homopolymeric MSI markers and a robust method for detecting homonucleotide insertions or deletions (indels) that can be performed using standard laboratory equipment, allowing for automation and easy interpretation, thereby simplifying the detection of MSI in cancer samples.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional MSI testing methods are used, then measurement precision is improved, but device complexity increases and requires specialized equipment
Solution Approach 1:
The patent uses digital PCR to create digital copies of DNA molecules, allowing individual molecules to be amplified and counted separately. This digital copying approach enables precise MSI detection through simple binary outcomes (amplification yes/no) rather than complex continuous measurements, resolving the contradiction between precision and complexity
Solution Approach 2:
The patent replaces complex mechanical sequencing equipment with a simpler digital counting system. Instead of using sophisticated mechanical systems to sequence and analyze DNA, the invention uses digital PCR with fluorescent detection to count amplified DNA molecules, substituting mechanical complexity with optical detection and digital analysis
2Measurement precision
If conventional MSI testing methods are used, then measurement precision is improved, but loss of time increases due to extensive manipulation
Solution Approach 1:
The patent performs preliminary whole-genome amplification of the tumor DNA sample before the actual MSI analysis. This preliminary action creates sufficient DNA material in advance, eliminating the need for time-consuming DNA extraction and purification steps during the critical measurement phase, thus reducing total manipulation time while maintaining precision
Solution Approach 2:
The patent combines multiple steps into a single digital PCR reaction: DNA amplification, fragment separation by size, and fluorescent detection all occur in one integrated assay. This merging of steps eliminates the need for separate manipulation steps required by conventional methods, significantly reducing laboratory handling time
3Measurement precision
If conventional MSI testing methods are used, then measurement precision is improved, but productivity decreases due to skilled personnel requirements
Solution Approach 1:
The patent employs automated digital PCR systems that perform the entire MSI analysis process without human intervention after sample loading. The system automatically amplifies DNA, separates fragments, detects fluorescent signals, and generates results, making the assay self-sufficient and eliminating the need for skilled personnel to perform manual operations, thus increasing productivity while maintaining precision
Solution Approach 2:
The patent uses automated image analysis software that provides real-time feedback on amplification results, automatically determining MSI status based on predefined criteria. This feedback mechanism eliminates the need for skilled personnel to interpret complex data, allowing routine operation by less specialized staff while maintaining high measurement precision
4Reliability
If comprehensive MMR gene mutation testing is performed, then reliability is improved, but device complexity and cost increase
Solution Approach 1:
The patent extracts and analyzes only the specific microsatellite regions that are indicative of MMR deficiency, rather than testing for all possible MMR gene mutations. By taking out and focusing on these key genomic markers, the invention achieves reliable detection of MMR status with a simpler, more cost-effective digital PCR system rather than requiring comprehensive gene sequencing
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
The method provides highly sensitive and consistent results, capable of identifying MSI-H tumors with high accuracy, reducing turnaround time to less than 3 hours and facilitating implementation in clinical settings, improving the management and prognosis of cancer patients.
Implementation Method 1
determining the number of nucleotides in the following homopolymeric repeats
Implementation Method 2
capillary electrophoresis, fluorescent PCR is used to amplify specific genomic regions
Data Source
Figure 1A~1D
Figure 2A~2D
AI summary
The present invention generally relates to the field of cancer, in particular to cancers having microsatellite instability (MSI) and/or mismatch repair (MMR-) deficiency. Examples of such cancers include many colorectal, gastric, and endometrial tumors. Accordingly, the present invention provides a novel diagnostic marker panel for analyzing MSI loci, together with methods and kits of using said panel in the detection of cancers having microsatellite instability (MSI) and/or mismatch repair (MMR-) deficiency.