Mismatch-Guided Deoxyribonucleic Acid Assembly Enables Ultrasensitive and Multiplex Detection of Low-Allele-Fraction Variants in Clinical Samples.

Autor:	Huang D; Key Laboratory of Green Chemistry and Technology of Ministry of Education, College of Chemistry, Sichuan University, Chengdu, Sichuan 610061, P. R. China., Deng H; Department of Respiratory and Critical Care Medicine, Targeted Tracer Research and Development Laboratory, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P. R. China., Zhou J; Department of Laboratory Medicine, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, China., Wang GA; Key Laboratory of Green Chemistry and Technology of Ministry of Education, College of Chemistry, Sichuan University, Chengdu, Sichuan 610061, P. R. China., Lei Q; Department of Respiratory and Critical Care Medicine, Targeted Tracer Research and Development Laboratory, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P. R. China., Guo C; Key Laboratory of Green Chemistry and Technology of Ministry of Education, College of Chemistry, Sichuan University, Chengdu, Sichuan 610061, P. R. China., Peng W; Key Laboratory of Green Chemistry and Technology of Ministry of Education, College of Chemistry, Sichuan University, Chengdu, Sichuan 610061, P. R. China., Liang P; Department of Laboratory Medicine, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, China.; Med+X Center for Manufacturing, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P. R. China., Shen C; Department of Laboratory Medicine, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, China.; Med+X Center for Manufacturing, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P. R. China., Ying B; Department of Laboratory Medicine, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, China.; Med+X Center for Manufacturing, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P. R. China., Li W; Department of Respiratory and Critical Care Medicine, Targeted Tracer Research and Development Laboratory, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P. R. China., Li F; Key Laboratory of Green Chemistry and Technology of Ministry of Education, College of Chemistry, Sichuan University, Chengdu, Sichuan 610061, P. R. China.; Med+X Center for Manufacturing, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P. R. China.
Jazyk:	angličtina
Zdroj:	Journal of the American Chemical Society [J Am Chem Soc] 2023 Sep 20; Vol. 145 (37), pp. 20412-20421. Date of Electronic Publication: 2023 Aug 31.
DOI:	10.1021/jacs.3c05879
Abstrakt:	Somatic mutations are important signatures in clinical cancer treatment. However, accurate detection of rare somatic mutations with low variant-allele frequencies (VAFs) in clinical samples is challenging because of the interference caused by high concentrations of wild-type (WT) sequences. Here, we report a post amplification SNV-specific DNA assembly (PANDA) technology that eliminates the high concentration pressure caused by WT through a mismatch-guided DNA assembly and enables the ultrasensitive detection of cancer mutations with VAFs as low as 0.1%. Because it generates an assembly product that only exposes a single-stranded domain with the minimal length for signal readout and thus eliminates possible interferences from secondary structures and cross-interactions among sequences, PANDA is highly versatile and expandable for multiplex testing. With ultrahigh sensitivity, PANDA enabled the quantitative analysis of EGFR mutations in cell-free DNA of 68 clinical plasma samples and four pleuroperitoneal fluid samples, with test results highly consistent with NGS deep sequencing. Compared to digital PCR, PANDA returned fewer false negatives and ambiguous cases of clinical tests. Meanwhile, it also offers much lower upfront instrumental and operational costs. The multiplexity was demonstrated by developing a 3-plex PANDA for the simultaneous analysis of three EGFR mutations in 54 pairs of tumor and the adjacent noncancerous tissue samples collected from lung cancer patients. Because of the ultrahigh sensitivity, multiplexity, and simplicity, we anticipate that PANDA will find wide applications for analyzing clinically important rare mutations in diverse devastating diseases.
Databáze:	MEDLINE
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