Anchored multiplex PCR for targeted next-generation sequencing

By Zongli Zheng, M. Liebers, Boryana H. Zhelyazkova, Yi Cao, Divya Panditi, Kerry D Lynch, Juxiang Chen, Hayley E Robinson, H. Shim, J. Chmielecki and 4 more. Nature Medicine, 2014

832 citations recorded by the source corpus.

We describe a rapid target enrichment method for next-generation sequencing, termed anchored multiplex PCR (AMP), that is compatible with low nucleic acid input from formalin-fixed paraffin-embedded (FFPE) specimens. AMP is effective in detecting gene rearrangements (without prior knowledge of the fusion partners), single nucleotide variants, insertions, deletions and copy number changes. Validation of a gene rearrangement panel using 319 FFPE samples showed 100% sensitivity (95% confidence limit: 96.5–100%) and 100% specificity (95% confidence limit: 99.3–100%) compared with reference assays. On the basis of our experience with performing AMP on 986 clinical FFPE samples, we show its potential as both a robust clinical assay and a powerful discovery tool, which we used to identify new therapeutically important gene fusions: ARHGEF2-NTRK1 and CHTOP-NTRK1 in glioblastoma, MSN-ROS1, TRIM4-BRAF, VAMP2-NRG1, TPM3-NTRK1 and RUFY2-RET in lung cancer, FGFR2-CREB5 in cholangiocarcinoma and PPL-NTRK1 in thyroid carcinoma. AMP is a scalable and efficient next-generation sequencing target enrichment method for research and clinical applications.

Subfields: Molecular Biology, Surgery

View Paper: https://www.semanticscholar.org/paper/f17065d471033cad038e886348403d208db650b0

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Citations in the corpus

Enhancing the accuracy of next-generation sequencing for detecting rare and subclonal mutations (2018, background): “In addition, emerging hybrid biochemical methods that combine the benefits of PCR-based targeting on library preparation speed and conversion efficiency with the advantages of adapter-based molecular tagging are likely to further narrow current performance trade-offs 105,106 .”
Tumor cells can follow distinct evolutionary paths to become resistant to epidermal growth factor receptor inhibition (2016, neutral): “The gene targets covered by this assay are as follows (exons): AKT1 (3), ALK (22, 23, 25), APC (16), BRAF (11, 15), CDH1 (1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11,12, 13, 14, 15, 16), CDKN2A (1, 2, 3), CTNNB1 (3), DDR2 (12, 13, 14, 15, 16, 17, 18), EGFR (7, 15, 18, 19, 20, 21), ERBB2 (10, 20), ESR1 (8), FB”
STK11/LKB1 Mutations and PD-1 Inhibitor Resistance in KRAS-Mutant Lung Adenocarcinoma (2018, neutral): “55(0) 42(11) 20(32) 11(41) 6(44) 2(47) 0(49) 3(4) 0(6) 0(6) 0(6) 0(6) 0(6)”
Updated Molecular Testing Guideline for the Selection of Lung Cancer Patients for Treatment With Targeted Tyrosine Kinase Inhibitors: Guideline From the College of American Pathologists, the International Association for the Study of Lung Cancer, and the Association for Molecular Pathology (2018, methodology): “A capture-based sequencing approach, involving DNA or RNA, may be more sensitive and more readily integrated into a large multigene panel.(76) 5.”
GUIDE-Seq enables genome-wide profiling of off-target cleavage by CRISPR-Cas nucleases (2014, neutral): “For AMP validation of GUIDE-Seq detected DSBs, primers were designed to regions flanking inferred double-stranded breakpoints as described previously 37 , with the addition of an 8-nt molecular molecular index.”
Single-cell transcriptomic analysis of primary and metastatic tumor ecosystems in head and neck cancer (2017, methodology): “In addition, SnAPShot next generation sequencing v2 assay was performed on FFPE samples at the MGH Center for Integrated Diagnostics per standard protocols as previously described (Zheng et al., 2014).”