Next Generation-Targeted Amplicon Sequencing (NG-TAS): an optimised protocol and computational pipeline for cost-effective profiling of circulating tumour DNA

Genome Med. 2019 Jan 4;11(1):1. doi: 10.1186/s13073-018-0611-9.

Abstract

Circulating tumour DNA (ctDNA) detection and monitoring have enormous potential clinical utility in oncology. We describe here a fast, flexible and cost-effective method to profile multiple genes simultaneously in low input cell-free DNA (cfDNA): Next Generation-Targeted Amplicon Sequencing (NG-TAS). We designed a panel of 377 amplicons spanning 20 cancer genes and tested the NG-TAS pipeline using cell-free DNA from two HapMap lymphoblastoid cell lines. NG-TAS consistently detected mutations in cfDNA when mutation allele fraction was > 1%. We applied NG-TAS to a clinical cohort of metastatic breast cancer patients, demonstrating its potential in monitoring the disease. The computational pipeline is available at https://github.com/cclab-brca/NGTAS_pipeline .

Keywords: Cancer; Computational pipeline; Deep sequencing; Heterogeneous; Liquid biopsy; Multiplexing; Mutation; NG-TAS; ctDNA.

Publication types

  • Clinical Trial
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Biomarkers, Tumor / blood
  • Biomarkers, Tumor / genetics*
  • Cell Line, Tumor
  • Circulating Tumor DNA / blood
  • Circulating Tumor DNA / genetics*
  • Costs and Cost Analysis
  • Female
  • High-Throughput Nucleotide Sequencing / economics
  • High-Throughput Nucleotide Sequencing / methods*
  • Humans
  • Mutation
  • Sequence Analysis, DNA / economics
  • Sequence Analysis, DNA / methods*
  • Software*

Substances

  • Biomarkers, Tumor
  • Circulating Tumor DNA