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Lung Cancer. 2019 Aug;134:96-99. doi: 10.1016/j.lungcan.2019.06.004. Epub 2019 Jun 5.

Sensitivity of next-generation sequencing assays detecting oncogenic fusions in plasma cell-free DNA.

Author information

1
Belfer Center for Applied Cancer Science, Dana-Farber Cancer Institute, 360 Longwood Avenue, Boston, MA, 02215, USA. Electronic address: julianna_supplee@dfci.harvard.edu.
2
Lowe Center for Thoracic Oncology, Dana-Farber Cancer Institute, 450 Brookline Avenue, Boston, MA, 02215, USA. Electronic address: MarinaS_Milan@dfci.harvard.edu.
3
Resolution Bioscience, Inc., 550 Kirkland Way, Kirkland, WA, 98033, USA. Electronic address: lplim@resolutionbio.com.
4
Resolution Bioscience, Inc., 550 Kirkland Way, Kirkland, WA, 98033, USA. Electronic address: kristy@resolutionbio.com.
5
Department of Pathology, Brigham and Women's Hospital, 75 Francis Street, Boston, MA, 02115, USA. Electronic address: lmsholl@bwh.harvard.edu.
6
Lowe Center for Thoracic Oncology, Dana-Farber Cancer Institute, 450 Brookline Avenue, Boston, MA, 02215, USA. Electronic address: geoffrey_oxnard@dfci.harvard.edu.
7
Belfer Center for Applied Cancer Science, Dana-Farber Cancer Institute, 360 Longwood Avenue, Boston, MA, 02215, USA. Electronic address: cloudp_paweletz@dfci.harvard.edu.

Abstract

OBJECTIVES:

Plasma genotyping represents an opportunity for convenient detection of clinically actionable mutations in advanced cancer patients, such has been well-documented in non-small cell lung cancer (NSCLC). Oncogenic gene fusions are complex variants that may be more challenging to detect by next-generation sequencing (NGS) of plasma cell-free DNA (cfDNA). Rigorous evaluation of plasma NGS assays in the detection of fusions is needed to maximize clinical utility.

MATERIALS AND METHODS:

Additional plasma was collected from patients with advanced NSCLC and ALK, ROS1, or RET gene fusions in tissue who had undergone clinical plasma NGS using Guardant360(G360, Guardant Health). We then sequenced extracted cfDNA with a plasma NGS kit focused on known driver mutations in NSCLC (ctDx-Lung, Resolution Bioscience) with cloud-based bioinformatic analysis and blinded variant calling.

RESULTS:

Of 16 patients assayed known to harbor anALK, ROS1, or RET in tumor, G360 detected fusions in 7 cases, ctDx-Lung detected fusions in 13 cases, and 3 cases were detected by neither. Of the 7 fusions detected by both assays, G360 reported lower mutant allelic fractions (AF). In cases missed by G360, tumor derived TP53 mutations were often detected confirming presence of tumor DNA. Raw sequencing data showed that inverted or out-of-frame variants were overrepresented in cases detected using ctDx-Lung but not by G360.

CONCLUSION:

Focusing on complex, clinically actionable mutations using tumor as a reference standard allows for evaluation of technical differences in plasma NGS assays that may impact clinical performance. Noting the heterogeneity of fusion sequences observed in NSCLC, we hypothesize that differences in hybrid capture techniques and bioinformatic calling may be sources of variations in sensitivity among these assays.

KEYWORDS:

Cell-free DNA; Gene fusions; NSCLC; Next-generation sequencing

PMID:
31320002
DOI:
10.1016/j.lungcan.2019.06.004
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