2024 World Conference on Lung Cancer (WCLC) - ePosters-EP.12A.13 CRISPR with Double Mismatch Guide RNA Enhances Detection EGFR Mutation in Circulating Cell-Free DNA of Lung Cancer Patients

EP.12A.13 CRISPR with Double Mismatch Guide RNA Enhances Detection EGFR Mutation in Circulating Cell-Free DNA of Lung Cancer Patients

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This document discusses the development and application of a CRISPR/Cas12a-based detection method for identifying EGFR mutations in circulating cell-free DNA (cfDNA) from patients with non-small cell lung cancer (NSCLC). The targeted mutations include the L858R mutant allele and the exon 19 deletion, both significant biomarkers for NSCLC. Traditional methods like next-generation sequencing (NGS) are not always sensitive enough to detect low-abundance mutant alleles from cfDNA. Therefore, this CRISPR/Cas12a system offers a more sensitive tool for selective removal of normal alleles and specific enrichment of mutant alleles. <br /><br />The methodology leverages CRISPR/Cas12a for two main processes: enrichment of mutant alleles and deletion of normal alleles. This system enables enhanced detection via NGS and fluorescence-based collateral effects. It increases the sensitivity limit from 0.1% in traditional NGS to a much lower 0.001% abundance. This heightened sensitivity offers a significant advantage in detecting low-frequency mutations, which are often missed by less sensitive techniques.<br /><br />In clinical trials, this method demonstrated increased detection rates of targeted mutations in the cfDNA of 48 NSCLC patients, allowing for more reliable diagnosis and monitoring. Notably, the collateral effect-based fluorescence method allows these detections to be made within one hour, offering a quick and efficient alternative to more time-consuming sequencing methods. Additionally, using crRNA with a double mismatch to the normal DNA further enhances the assay's specificity and sensitivity.<br /><br />In conclusion, the CRISPR/Cas12a system represents a promising advancement for the sensitive detection of pathogenic mutations in cfDNA, potentially improving diagnostic capabilities and treatment selection for NSCLC and other diseases characterized by specific genetic mutations.

Asset Subtitle

Boksoon Chang

Meta Tag

Speaker Boksoon Chang

Topic Metastatic Non-small Cell Lung Cancer – Targeted Therapy

Keywords

CRISPR/Cas12a

EGFR mutations

circulating cell-free DNA

non-small cell lung cancer

L858R mutant allele

exon 19 deletion

next-generation sequencing

mutant allele enrichment

fluorescence detection

clinical trials