TY - JOUR
T1 - PCR-Free Shallow Whole Genome Sequencing for Chromosomal Copy Number Detection from Plasma of Cancer Patients Is an Efficient Alternative to the Conventional PCR-Based Approach
AU - Beagan, Jamie J.
AU - Drees, Esther E. E.
AU - Stathi, Phylicia
AU - Eijk, Paul P.
AU - Meulenbroeks, Laura
AU - Kessler, Floortje
AU - Middeldorp, Jaap M.
AU - Pegtel, D. Michiel
AU - Zijlstra, Josée M.
AU - Sie, Daoud
AU - Heideman, Daniëlle A. M.
AU - Thunnissen, Erik
AU - Smit, Linda
AU - de Jong, Daphne
AU - Mouliere, Florent
AU - Ylstra, Bauke
AU - Roemer, Margaretha G. M.
AU - van Dijk, Erik
N1 - Funding Information: Supported by Amsterdam University Medical Center intramural funds, including the Vrije Universiteit Medical Center Cancer Center Amsterdam Foundation . The funding body had no role in the study design, execution of the research activities, or the decision to submit the manuscript for publication. Publisher Copyright: © 2021 Association for Molecular Pathology and American Society for Investigative Pathology
PY - 2021/11/1
Y1 - 2021/11/1
N2 - Somatic copy number alterations can be detected in cell-free DNA (cfDNA) by shallow whole genome sequencing (sWGS). PCR is typically included in library preparations, but a PCR-free method could serve as a high-throughput alternative. To evaluate a PCR-free method for research and diagnostics, archival peripheral blood or bone marrow plasma samples, collected in EDTA- or lithium-heparin–containing tubes, were collected from patients with non–small-cell lung cancer (n = 10 longitudinal samples; 4 patients), B-cell lymphoma (n = 31), and acute myeloid leukemia (n = 15), or from healthy donors (n = 14). sWGS was performed on PCR-free and PCR library preparations, and the mapping quality, percentage of unique reads, genome coverage, fragment lengths, and copy number profiles were compared. The percentage of unique reads was significantly higher for PCR-free method compared with PCR method, independent of the type of collection tube: EDTA PCR-free method, 96.4% (n = 35); EDTA PCR method, 85.1% (n = 32); heparin PCR-free method, 94.5% (n = 25); and heparin PCR method, 89.4% (n = 10). All other evaluated metrics were highly comparable for PCR-free and PCR library preparations. These results demonstrate the feasibility of somatic copy number alteration detection by PCR-free sWGS using cfDNA from plasma collected in EDTA- or lithium-heparin–containing tubes and pave the way for an automated cfDNA analysis workflow for samples from cancer patients.
AB - Somatic copy number alterations can be detected in cell-free DNA (cfDNA) by shallow whole genome sequencing (sWGS). PCR is typically included in library preparations, but a PCR-free method could serve as a high-throughput alternative. To evaluate a PCR-free method for research and diagnostics, archival peripheral blood or bone marrow plasma samples, collected in EDTA- or lithium-heparin–containing tubes, were collected from patients with non–small-cell lung cancer (n = 10 longitudinal samples; 4 patients), B-cell lymphoma (n = 31), and acute myeloid leukemia (n = 15), or from healthy donors (n = 14). sWGS was performed on PCR-free and PCR library preparations, and the mapping quality, percentage of unique reads, genome coverage, fragment lengths, and copy number profiles were compared. The percentage of unique reads was significantly higher for PCR-free method compared with PCR method, independent of the type of collection tube: EDTA PCR-free method, 96.4% (n = 35); EDTA PCR method, 85.1% (n = 32); heparin PCR-free method, 94.5% (n = 25); and heparin PCR method, 89.4% (n = 10). All other evaluated metrics were highly comparable for PCR-free and PCR library preparations. These results demonstrate the feasibility of somatic copy number alteration detection by PCR-free sWGS using cfDNA from plasma collected in EDTA- or lithium-heparin–containing tubes and pave the way for an automated cfDNA analysis workflow for samples from cancer patients.
UR - http://www.scopus.com/inward/record.url?scp=85111084433&partnerID=8YFLogxK
UR - https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=85111084433&origin=inward
UR - https://www.ncbi.nlm.nih.gov/pubmed/34454114
U2 - https://doi.org/10.1016/j.jmoldx.2021.08.008
DO - https://doi.org/10.1016/j.jmoldx.2021.08.008
M3 - Article
C2 - 34454114
SN - 1525-1578
VL - 23
SP - 1553
EP - 1563
JO - Journal of molecular diagnostics
JF - Journal of molecular diagnostics
IS - 11
ER -