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Updated: Oct 27, 2025
Detection and Monitoring of Tumor Associated Circulating DNA in Patient Biofluids
Published on: June 8, 2019
Validation of a Circulating Tumor DNA-Based Next-Generation Sequencing Assay in a Cohort of Patients with Solid
Wael Al Zoughbi1,2, Jesse Fox3, Shaham Beg1,2
1Department of Pathology and Laboratory Medicine, Weill Cornell Medicine, New York, New York, USA.
Background:
Characterization of circulating tumor DNA (ctDNA) has been integrated into clinical practice. Although labs have standardized validation procedures to develop single locus tests, the efficacy of on-site plasma-based next-generation sequencing (NGS) assays still needs to be proved.
Materials And Methods:
In this retrospective study, we profiled DNA from matched tissue and plasma samples from 75 patients with cancer. We applied an NGS test that detects clinically relevant alterations in 33 genes and microsatellite instability (MSI) to analyze plasma cell-free DNA (cfDNA).
Results:
The concordance between alterations detected in both tissue and plasma samples was higher in patients with metastatic disease. The NGS test detected 77% of sequence alterations, amplifications, and fusions that were found in metastatic samples compared with 45% of those alterations found in the primary tumor samples (p = .00005). There was 87% agreement on MSI status between the NGS test and tumor tissue results. In three patients, MSI-high ctDNA correlated with response to immunotherapy. In addition, the NGS test revealed an FGFR2 amplification that was not detected in tumor tissue from a patient with metastatic gastric cancer, emphasizing the importance of profiling plasma samples in patients with advanced cancer.
Conclusion:
Our validation experience of a plasma-based NGS assay advances current knowledge about translating cfDNA testing into clinical practice and supports the application of plasma assays in the management of oncology patients with metastatic disease. With an in-house method that minimizes the need for invasive procedures, on-site cfDNA testing supplements tissue biopsy to guide precision therapy and is entitled to become a routine practice.
Implications For Practice:
This study proposes a solution for decentralized liquid biopsy testing based on validation of a next-generation sequencing (NGS) test that detects four classes of genomic alterations in blood: sequence mutations (single nucleotide substitutions or insertions and deletions), fusions, amplifications, and microsatellite instability (MSI). Although there are reference labs that perform single-site comprehensive liquid biopsy testing, the targeted assay this study validated can be established locally in any lab with capacity to offer clinical molecular pathology assays. To the authors' knowledge, this is the first report that validates evaluating an on-site plasma-based NGS test that detects the MSI status along with common sequence alterations encountered in solid tumors.
