FLT3 Primer Composition for Simultaneous ITD and TKD Detection
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Solution Overview
Problem
Current methods for detecting FLT3 gene mutations in acute myeloid leukemia (AML) have low sensitivity and cannot accurately measure mutation length or sequence, particularly in the ITD and TKD regions, limiting effective diagnosis, prognosis prediction, and drug resistance determination.
Innovation Solution
A composition using specific primer sets to amplify and analyze the ITD and TKD regions of the FLT3 gene, enabling high-sensitivity and accurate detection of mutations through next-generation sequencing (NGS).
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If PCR and electrophoresis are used to detect FLT3-ITD mutations, then the detection process is simple and fast, but the sensitivity is very low (10%) and cannot accurately measure mutation length and sequence
Solution Approach 1:
The patent combines multiple detection capabilities (ITD detection, TKD mutation detection, sequence analysis, and length measurement) into a single NGS-based assay system. By merging these functions into one comprehensive platform, the invention achieves high sensitivity (at least 10^-4) while providing accurate mutation characterization that neither PCR nor fragment analysis can deliver alone.
Solution Approach 2:
The invention creates a universal detection system that can simultaneously perform multiple functions: detecting ITD mutations, identifying TKD point mutations, determining mutation sequences, and measuring ITD length. This multi-functional platform addresses the limitations of specialized methods by providing a single assay that handles all FLT3 mutation detection needs with high sensitivity and accuracy.
2Reliability
If fragment analysis is used to detect FLT3-ITD mutations, then the sensitivity improves to about 3% and ITD length can be measured, but sequence analysis is still incapable and sensitivity is insufficient for MRD detection (requires at least 10^-4)
Solution Approach 1:
The patent replaces the mechanical separation and sizing approach of fragment analysis with NGS technology. Instead of relying on physical fragment separation to infer mutation characteristics, the invention uses sequence-based detection to directly identify and characterize mutations, thereby recovering complete sequence information while achieving the required sensitivity for MRD detection.
Solution Approach 2:
The invention changes the detection parameter from fragment size (used in fragment analysis) to sequence composition. By using NGS to read the actual nucleotide sequences rather than merely measuring fragment lengths, the system recovers complete sequence information and achieves superior sensitivity through precise molecular identification rather than statistical size-based detection.
3Reliability
If NGS is used to detect FLT3 mutations with high sensitivity, then sensitivity reaches 5×10^-5 and sequence analysis is available, but it cannot simultaneously detect mutations in the TKD region
Solution Approach 1:
The patent divides the FLT3 gene into distinct functional regions (ITD region and TKD region) and designs specific primer sets for each region. This segmentation allows the NGS assay to simultaneously target and detect mutations in both regions with equal sensitivity, overcoming the limitation of previous methods that could only focus on one region at a time.
Solution Approach 2:
The invention creates a universal NGS-based FLT3 mutation detection platform that can simultaneously identify both ITD and TKD mutations with high sensitivity. By designing a multi-primer system that targets all relevant FLT3 regions, the assay achieves comprehensive coverage while maintaining the sensitivity required for MRD detection, making it more versatile than region-specific methods.
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
Enables rapid and precise diagnosis of AML, prediction of prognosis, and determination of drug resistance with improved sensitivity and accuracy, facilitating targeted treatment strategies.
Implementation Method 1
A composition containing primer sets for simultaneously amplifying an ITD detection region and a TKD mutation region of FLT3 genes
Data Source
AI summary
The present invention relates to a composition for amplifying a FLT3 gene, and uses thereof, and, more particularly, to a composition comprising a primer set capable of simultaneously amplifying an ITD detection region and a TKD mutation region of the FLT3 gene, and uses thereof. The composition for gene amplification, according to the present invention, enables the simultaneous performance of: diagnosis of acute myeloid leukemia (AML) in patients having FLT3-ITD mutations; determination of targeted anticancer treatment prescription for AML patients having FLT3-ITD mutations; detection of minimal residual disease (MRD) in AML patients; prognosis prediction in AML patients; and identification of drug resistance to AML tyrosine kinase inhibitors, and thus, shortens the time to derive analysis results from samples and enables efficient testing. The present invention enables the selection of correct and rapid diagnosis and treatment methods in the treatment of patients with acute myeloid leukemia, and thus is useful for early treatment and recurrence prevention.