Accurate detection of circulating tumor DNA using nanopore consensus sequencing

Alessio Marcozzi; Myrthe Jager; Martin Elferink; Roy Straver; Joost H van Ginkel; Boris Peltenburg; Li-Ting Chen; Ivo Renkens; Joyce van Kuik; Chris Terhaard; Remco de Bree; Lot A Devriese; Stefan M Willems; Wigard P Kloosterman; Jeroen de Ridder

doi:10.1038/s41525-021-00272-y

Accurate detection of circulating tumor DNA using nanopore consensus sequencing

Alessio Marcozzi
, Myrthe Jager
, Martin Elferink
, Roy Straver
, Joost H van Ginkel
, Boris Peltenburg
, Li-Ting Chen
, Ivo Renkens
, Joyce van Kuik
, Chris Terhaard
, Remco de Bree
, Lot A Devriese
, Stefan M Willems
, Wigard P Kloosterman
, Jeroen de Ridder

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Abstract

Levels of circulating tumor DNA (ctDNA) in liquid biopsies may serve as a sensitive biomarker for real-time, minimally-invasive tumor diagnostics and monitoring. However, detecting ctDNA is challenging, as much fewer than 5% of the cell-free DNA in the blood typically originates from the tumor. To detect lowly abundant ctDNA molecules based on somatic variants, extremely sensitive sequencing methods are required. Here, we describe a new technique, CyclomicsSeq, which is based on Oxford Nanopore sequencing of concatenated copies of a single DNA molecule. Consensus calling of the DNA copies increased the base-calling accuracy ~60×, enabling accurate detection of TP53 mutations at frequencies down to 0.02%. We demonstrate that a TP53-specific CyclomicsSeq assay can be successfully used to monitor tumor burden during treatment for head-and-neck cancer patients. CyclomicsSeq can be applied to any genomic locus and offers an accurate diagnostic liquid biopsy approach that can be implemented in clinical workflows.

Original language	English
Article number	106
Pages (from-to)	1-11
Journal	npj Genomic Medicine
Volume	6
Issue number	1
DOIs	https://doi.org/10.1038/s41525-021-00272-y
Publication status	Published - 9 Dec 2021

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s41525-021-00272-yFinal published version, 1.87 MBLicence: CC BY

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Marcozzi, A., Jager, M., Elferink, M., Straver, R., van Ginkel, J. H., Peltenburg, B., Chen, L.-T., Renkens, I., van Kuik, J., Terhaard, C., de Bree, R., Devriese, L. A., Willems, S. M., Kloosterman, W. P., & de Ridder, J. (2021). Accurate detection of circulating tumor DNA using nanopore consensus sequencing. npj Genomic Medicine, 6(1), 1-11. Article 106. https://doi.org/10.1038/s41525-021-00272-y

@article{b00082e040b2474e8fd223ad8406e97e,

title = "Accurate detection of circulating tumor DNA using nanopore consensus sequencing",

abstract = "Levels of circulating tumor DNA (ctDNA) in liquid biopsies may serve as a sensitive biomarker for real-time, minimally-invasive tumor diagnostics and monitoring. However, detecting ctDNA is challenging, as much fewer than 5\% of the cell-free DNA in the blood typically originates from the tumor. To detect lowly abundant ctDNA molecules based on somatic variants, extremely sensitive sequencing methods are required. Here, we describe a new technique, CyclomicsSeq, which is based on Oxford Nanopore sequencing of concatenated copies of a single DNA molecule. Consensus calling of the DNA copies increased the base-calling accuracy \textasciitilde{}60×, enabling accurate detection of TP53 mutations at frequencies down to 0.02\%. We demonstrate that a TP53-specific CyclomicsSeq assay can be successfully used to monitor tumor burden during treatment for head-and-neck cancer patients. CyclomicsSeq can be applied to any genomic locus and offers an accurate diagnostic liquid biopsy approach that can be implemented in clinical workflows.",

author = "Alessio Marcozzi and Myrthe Jager and Martin Elferink and Roy Straver and \{van Ginkel\}, \{Joost H\} and Boris Peltenburg and Li-Ting Chen and Ivo Renkens and \{van Kuik\}, Joyce and Chris Terhaard and \{de Bree\}, Remco and Devriese, \{Lot A\} and Willems, \{Stefan M\} and Kloosterman, \{Wigard P\} and \{de Ridder\}, Jeroen",

note = "Funding Information: We would like to thank Manon Huibers for input on the study, Floris Reinders for input on the manuscript and the Utrecht Sequencing Facility for sequencing. The Utrecht Sequencing Facility is subsidized by the University Medical Center Utrecht, Hubrecht Institute, and Utrecht University. This study was financially supported by the Oncode Institute (project number P2018-004) and is part of the Oncode Institute, which is partly financed by the Dutch Cancer Society. Publisher Copyright: {\textcopyright} 2021, The Author(s).",

year = "2021",

month = dec,

day = "9",

doi = "10.1038/s41525-021-00272-y",

language = "English",

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journal = "npj Genomic Medicine",

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Marcozzi, A, Jager, M, Elferink, M, Straver, R, van Ginkel, JH, Peltenburg, B, Chen, L-T, Renkens, I, van Kuik, J, Terhaard, C , de Bree, R, Devriese, LA, Willems, SM, Kloosterman, WP & de Ridder, J 2021, 'Accurate detection of circulating tumor DNA using nanopore consensus sequencing', npj Genomic Medicine, vol. 6, no. 1, 106, pp. 1-11. https://doi.org/10.1038/s41525-021-00272-y

TY - JOUR

T1 - Accurate detection of circulating tumor DNA using nanopore consensus sequencing

AU - Marcozzi, Alessio

AU - Jager, Myrthe

AU - Elferink, Martin

AU - Straver, Roy

AU - van Ginkel, Joost H

AU - Peltenburg, Boris

AU - Chen, Li-Ting

AU - Renkens, Ivo

AU - van Kuik, Joyce

AU - Terhaard, Chris

AU - de Bree, Remco

AU - Devriese, Lot A

AU - Willems, Stefan M

AU - Kloosterman, Wigard P

AU - de Ridder, Jeroen

N1 - Funding Information: We would like to thank Manon Huibers for input on the study, Floris Reinders for input on the manuscript and the Utrecht Sequencing Facility for sequencing. The Utrecht Sequencing Facility is subsidized by the University Medical Center Utrecht, Hubrecht Institute, and Utrecht University. This study was financially supported by the Oncode Institute (project number P2018-004) and is part of the Oncode Institute, which is partly financed by the Dutch Cancer Society. Publisher Copyright: © 2021, The Author(s).

PY - 2021/12/9

Y1 - 2021/12/9

N2 - Levels of circulating tumor DNA (ctDNA) in liquid biopsies may serve as a sensitive biomarker for real-time, minimally-invasive tumor diagnostics and monitoring. However, detecting ctDNA is challenging, as much fewer than 5% of the cell-free DNA in the blood typically originates from the tumor. To detect lowly abundant ctDNA molecules based on somatic variants, extremely sensitive sequencing methods are required. Here, we describe a new technique, CyclomicsSeq, which is based on Oxford Nanopore sequencing of concatenated copies of a single DNA molecule. Consensus calling of the DNA copies increased the base-calling accuracy ~60×, enabling accurate detection of TP53 mutations at frequencies down to 0.02%. We demonstrate that a TP53-specific CyclomicsSeq assay can be successfully used to monitor tumor burden during treatment for head-and-neck cancer patients. CyclomicsSeq can be applied to any genomic locus and offers an accurate diagnostic liquid biopsy approach that can be implemented in clinical workflows.

AB - Levels of circulating tumor DNA (ctDNA) in liquid biopsies may serve as a sensitive biomarker for real-time, minimally-invasive tumor diagnostics and monitoring. However, detecting ctDNA is challenging, as much fewer than 5% of the cell-free DNA in the blood typically originates from the tumor. To detect lowly abundant ctDNA molecules based on somatic variants, extremely sensitive sequencing methods are required. Here, we describe a new technique, CyclomicsSeq, which is based on Oxford Nanopore sequencing of concatenated copies of a single DNA molecule. Consensus calling of the DNA copies increased the base-calling accuracy ~60×, enabling accurate detection of TP53 mutations at frequencies down to 0.02%. We demonstrate that a TP53-specific CyclomicsSeq assay can be successfully used to monitor tumor burden during treatment for head-and-neck cancer patients. CyclomicsSeq can be applied to any genomic locus and offers an accurate diagnostic liquid biopsy approach that can be implemented in clinical workflows.

UR - https://www.scopus.com/pages/publications/85121034107

U2 - 10.1038/s41525-021-00272-y

DO - 10.1038/s41525-021-00272-y

M3 - Article

C2 - 34887408

VL - 6

SP - 1

EP - 11

JO - npj Genomic Medicine

JF - npj Genomic Medicine

IS - 1

M1 - 106

ER -

Accurate detection of circulating tumor DNA using nanopore consensus sequencing

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