CRISPR/Cas13a-based supersensitive circulating tumor DNA assay for detecting EGFR mutations in plasma

作者：Li Wang, Xiaosha Wen, Yang Yang, Zheng Hu, Jing Jiang, Lili Duan, Xiaofen Liao, Yan He, Yaru Liu, Jing Wang, Zhikun Liang, Xiaoya Zhu, Quan Liu, Tiancai Liu, Dixian Luo · 发表于：Communications Biology · 年份：2024 · DOI：10.1038/s42003-024-06368-2 · 被引用次数：19 · 研究领域：CRISPR and Genetic Engineering、Cancer Genomics and Diagnostics、Advanced biosensing and bioanalysis techniques

Despite recent technological advancements in cell tumor DNA (ctDNA) mutation detection, challenges persist in identifying low-frequency mutations due to inadequate sensitivity and coverage of current procedures. Herein, we introduce a super-sensitivity and specificity technique for detecting ctDNA mutations, named HiCASE. The method utilizes PCR-based CRISPR, coupled with the restriction enzyme. In this work, HiCASE focuses on testing a series of EGFR mutations to provide enhanced detection technology for non-small cell lung cancer (NSCLC), enabling a detection sensitivity of 0.01% with 40 ng cell free DNA standard. When applied to a panel of 140 plasma samples from 120 NSCLC patients, HiCASE exhibits 88.1% clinical sensitivity and 100% specificity with 40 μL of plasma, higher than ddPCR and Super-ARMS assay. In addition, HiCASE can also clearly distinguish T790M/C797S mutations in different positions at a 1% variant allele frequency, offering valuable guidance for drug utilization. Indeed, the established HiCASE assay shows potential for clinical applications.