Thyroid Nodule Genomic Screening for Degraded FFPE and FNA Samples
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Solution Overview
Problem
Current diagnostic methods for thyroid nodules with indeterminate cytology, such as FNA cytology and molecular techniques, have low accuracy and are hindered by DNA degradation in FFPE tissues and inadequate quality in FNA samples, making it difficult to distinguish between benign and cancerous nodules and determine the need for diagnostic surgery.
Innovation Solution
A highly sensitive PCR-based NGS assay that requires minimal nucleic acid input from FFPE or FNA samples to detect mutations and gene fusions in thyroid cancer-related genes, using hot start DNA polymerases with reduced 5'-3' exonuclease activity and adapter sequences for amplicon generation.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If FNA cytology and molecular techniques are used for diagnostic screening, then diagnostic accuracy is improved, but the methods are hindered by DNA degradation in FFPE tissues and inadequate quality in FNA samples
Solution Approach 1:
The patent modifies the PCR reaction parameters by using hot start DNA polymerases with reduced 5'-3' exonuclease activity and optimizing the cycling conditions to work effectively with degraded DNA templates from FFPE tissues and FNA samples, thereby maintaining diagnostic accuracy despite poor DNA quality
Solution Approach 2:
The patent introduces adapter sequences as intermediaries that facilitate the amplification of degraded DNA fragments. These adapters enable the PCR reaction to successfully amplify target sequences even when the DNA is fragmented, thus bridging the gap between degraded sample quality and diagnostic accuracy
2Measurement precision
If standard PCR-based NGS assay is used, then mutation detection sensitivity is improved, but it requires significant nucleic acid input that is unavailable in degraded FFPE or FNA samples
Solution Approach 1:
The patent optimizes the PCR amplification parameters including cycle number, polymerase concentration, and primer design to maximize amplification efficiency from minimal nucleic acid input, enabling sensitive mutation detection in FFPE and FNA samples with limited DNA quantity
Solution Approach 2:
The patent performs preliminary optimization of the PCR conditions and adapter design before applying the assay to clinical samples, ensuring that the method is pre-calibrated to work effectively with minimal nucleic acid input from degraded samples
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 allows for rapid and accurate detection of mutations and gene fusions in thyroid nodules, enabling informed decisions on diagnostic surgery and predicting malignancy risk with high sensitivity, even in degraded DNA samples.
Implementation Method 1
A highly sensitive PCR-based NGS assay that requires minimal nucleic acid input from FFPE or FNA samples to detect mutations and gene fusions in thyroid cancer-related genes
Implementation Method 2
using hot start DNA polymerases with reduced 5'-3' exonuclease activity and adapter sequences for amplicon generation
Data Source
AI summary
The present technology relates to methods for determining whether a patient having thyroid nodules with indeterminate cytology will benefit from diagnostic surgery, e.g., lobectomy. These methods are based on screening a patient's thyroid nodules and detecting alterations in target nucleic acid sequences corresponding to a specific set of thyroid cancer-related genes. Kits for use in practicing the methods are also provided.