A Novel High-throughput Droplet Digital PCR-based Indel Quantification Method for the Detection of Circulating Donor-derived Cell-free DNA after Kidney Transplantation

Jeroen G.H.P. Verhoeven; Karin Boer; Annemiek M.A. Peeters; Marian C. Clahsen-Van Groningen; Joke I. Roodnat; Jacqueline Van De Wetering; Daan Nieboer; Douglas A. Bost; Carla C. Baan; Dennis A. Hesselink

doi:10.1097/TP.0000000000004078

A Novel High-throughput Droplet Digital PCR-based Indel Quantification Method for the Detection of Circulating Donor-derived Cell-free DNA after Kidney Transplantation

Jeroen G.H.P. Verhoeven^*, Karin Boer, Annemiek M.A. Peeters, Marian C. Clahsen-Van Groningen, Joke I. Roodnat, Jacqueline Van De Wetering, Daan Nieboer, Douglas A. Bost, Carla C. Baan, Dennis A. Hesselink

^*Corresponding author for this work

Research output: Contribution to journal › Article › Academic › peer-review

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Abstract

Background. Donor-derived cell-free DNA (ddcfDNA) is a promising minimally invasive biomarker for acute rejection (AR) in kidney transplant recipients. To assess the diagnostic value of ddcfDNA as a marker for AR, ddcfDNA was quantified at multiple time points after kidney transplantation with a novel high-throughput droplet digital PCR indel method that allowed for the absolute quantification of ddcfDNA. Methods. In this study, ddcfDNA in plasma samples from 223 consecutive kidney transplant recipients was analyzed pretransplantation; at 3, 7, and 180 d after transplantation; and at time of for-cause biopsies obtained within the first 180 d after transplantation. Results. Median (interquartile range) ddcfDNA concentration was significantly higher on day 3 (58.3 [17.7-258.3] copies/mL) and day 7 (25.0 [10.4-70.8] copies/mL) than on day 180 after transplantation (4.2 [0.0-8.3] copies/mL; P < 0.001 and P < 0.001, respectively). At time of biopsy-proven AR (BPAR), between day 11 and day 180 after transplantation, ddcfDNA concentration was significantly higher (50.0 [25.0-108.3] copies/mL) than those when biopsies showed non-AR (0.0 [0.0-15.6] copies/mL; P < 0.05). ddcfDNA concentration within the first 10 d after transplantation showed no significant difference between recipients with BPAR and those with non-AR in their biopsy or between recipients with BPAR and ddcfDNA measured at day 3 and day 7. Conclusions. Unfortunately, ddcfDNA concentration is not a good biomarker to detect AR within the first 10 d after transplantation; however, BPAR occurring after 10 d after transplantation can be detected in kidney transplant recipients by ddcfDNA using a novel and unique high-throughput droplet digital PCR indel method.

Original language	English
Pages (from-to)	1777-1786
Number of pages	10
Journal	Transplantation
Volume	106
Issue number	9
DOIs	https://doi.org/10.1097/TP.0000000000004078 https://doi.org/10.1097/TP.0000000000004078
Publication status	Published - 1 Sept 2022

Bibliographical note

Funding Information:
D.A.H. has received lecture and consulting fees from Astellas Pharma, Chiesi Farmaceutici SpA, and Novartis Pharma, as well as grant support from Astellas Pharma and Chiesi Farmaceutici SpA (paid to his institution). D.A.B. is chief executive officer of JETA Molecular, Utrecht, the Netherlands, that provided reagents for the ddcfDNA analysis.

Funding Information:
This research was funded by the Dutch Kidney Foundation (grant 17OI03). The Dutch Kidney Foundation is a non profit organization that subsidizes research and innovation in nephrology and renal transplantation care.

Publisher Copyright:
© 2022 Lippincott Williams and Wilkins. All rights reserved.

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Verhoeven, J. G. H. P., Boer, K., Peeters, A. M. A., Clahsen-Van Groningen, M. C., Roodnat, J. I., Van De Wetering, J., Nieboer, D., Bost, D. A., Baan, C. C., & Hesselink, D. A. (2022). A Novel High-throughput Droplet Digital PCR-based Indel Quantification Method for the Detection of Circulating Donor-derived Cell-free DNA after Kidney Transplantation. Transplantation, 106(9), 1777-1786. https://doi.org/10.1097/TP.0000000000004078, https://doi.org/10.1097/TP.0000000000004078

@article{74c55fc85a514121a166f80d1464c460,

title = "A Novel High-throughput Droplet Digital PCR-based Indel Quantification Method for the Detection of Circulating Donor-derived Cell-free DNA after Kidney Transplantation",

abstract = "Background. Donor-derived cell-free DNA (ddcfDNA) is a promising minimally invasive biomarker for acute rejection (AR) in kidney transplant recipients. To assess the diagnostic value of ddcfDNA as a marker for AR, ddcfDNA was quantified at multiple time points after kidney transplantation with a novel high-throughput droplet digital PCR indel method that allowed for the absolute quantification of ddcfDNA. Methods. In this study, ddcfDNA in plasma samples from 223 consecutive kidney transplant recipients was analyzed pretransplantation; at 3, 7, and 180 d after transplantation; and at time of for-cause biopsies obtained within the first 180 d after transplantation. Results. Median (interquartile range) ddcfDNA concentration was significantly higher on day 3 (58.3 [17.7-258.3] copies/mL) and day 7 (25.0 [10.4-70.8] copies/mL) than on day 180 after transplantation (4.2 [0.0-8.3] copies/mL; P < 0.001 and P < 0.001, respectively). At time of biopsy-proven AR (BPAR), between day 11 and day 180 after transplantation, ddcfDNA concentration was significantly higher (50.0 [25.0-108.3] copies/mL) than those when biopsies showed non-AR (0.0 [0.0-15.6] copies/mL; P < 0.05). ddcfDNA concentration within the first 10 d after transplantation showed no significant difference between recipients with BPAR and those with non-AR in their biopsy or between recipients with BPAR and ddcfDNA measured at day 3 and day 7. Conclusions. Unfortunately, ddcfDNA concentration is not a good biomarker to detect AR within the first 10 d after transplantation; however, BPAR occurring after 10 d after transplantation can be detected in kidney transplant recipients by ddcfDNA using a novel and unique high-throughput droplet digital PCR indel method.",

author = "Verhoeven, {Jeroen G.H.P.} and Karin Boer and Peeters, {Annemiek M.A.} and {Clahsen-Van Groningen}, {Marian C.} and Roodnat, {Joke I.} and {Van De Wetering}, Jacqueline and Daan Nieboer and Bost, {Douglas A.} and Baan, {Carla C.} and Hesselink, {Dennis A.}",

note = "Funding Information: D.A.H. has received lecture and consulting fees from Astellas Pharma, Chiesi Farmaceutici SpA, and Novartis Pharma, as well as grant support from Astellas Pharma and Chiesi Farmaceutici SpA (paid to his institution). D.A.B. is chief executive officer of JETA Molecular, Utrecht, the Netherlands, that provided reagents for the ddcfDNA analysis. Funding Information: This research was funded by the Dutch Kidney Foundation (grant 17OI03). The Dutch Kidney Foundation is a non profit organization that subsidizes research and innovation in nephrology and renal transplantation care. Publisher Copyright: {\textcopyright} 2022 Lippincott Williams and Wilkins. All rights reserved.",

year = "2022",

month = sep,

day = "1",

doi = "10.1097/TP.0000000000004078",

language = "English",

volume = "106",

pages = "1777--1786",

journal = "Transplantation",

issn = "0041-1337",

publisher = "Lippincott Williams & Wilkins",

number = "9",

}

Verhoeven, JGHP, Boer, K , Peeters, AMA , Clahsen-Van Groningen, MC , Roodnat, JI , Van De Wetering, J , Nieboer, D, Bost, DA, Baan, CC & Hesselink, DA 2022, 'A Novel High-throughput Droplet Digital PCR-based Indel Quantification Method for the Detection of Circulating Donor-derived Cell-free DNA after Kidney Transplantation', Transplantation, vol. 106, no. 9, pp. 1777-1786. https://doi.org/10.1097/TP.0000000000004078, https://doi.org/10.1097/TP.0000000000004078

A Novel High-throughput Droplet Digital PCR-based Indel Quantification Method for the Detection of Circulating Donor-derived Cell-free DNA after Kidney Transplantation. / Verhoeven, Jeroen G.H.P.; Boer, Karin ; Peeters, Annemiek M.A. et al.
In: Transplantation, Vol. 106, No. 9, 01.09.2022, p. 1777-1786.

Research output: Contribution to journal › Article › Academic › peer-review

TY - JOUR

T1 - A Novel High-throughput Droplet Digital PCR-based Indel Quantification Method for the Detection of Circulating Donor-derived Cell-free DNA after Kidney Transplantation

AU - Verhoeven, Jeroen G.H.P.

AU - Boer, Karin

AU - Peeters, Annemiek M.A.

AU - Clahsen-Van Groningen, Marian C.

AU - Roodnat, Joke I.

AU - Van De Wetering, Jacqueline

AU - Nieboer, Daan

AU - Bost, Douglas A.

AU - Baan, Carla C.

AU - Hesselink, Dennis A.

N1 - Funding Information: D.A.H. has received lecture and consulting fees from Astellas Pharma, Chiesi Farmaceutici SpA, and Novartis Pharma, as well as grant support from Astellas Pharma and Chiesi Farmaceutici SpA (paid to his institution). D.A.B. is chief executive officer of JETA Molecular, Utrecht, the Netherlands, that provided reagents for the ddcfDNA analysis. Funding Information: This research was funded by the Dutch Kidney Foundation (grant 17OI03). The Dutch Kidney Foundation is a non profit organization that subsidizes research and innovation in nephrology and renal transplantation care. Publisher Copyright: © 2022 Lippincott Williams and Wilkins. All rights reserved.

PY - 2022/9/1

Y1 - 2022/9/1

N2 - Background. Donor-derived cell-free DNA (ddcfDNA) is a promising minimally invasive biomarker for acute rejection (AR) in kidney transplant recipients. To assess the diagnostic value of ddcfDNA as a marker for AR, ddcfDNA was quantified at multiple time points after kidney transplantation with a novel high-throughput droplet digital PCR indel method that allowed for the absolute quantification of ddcfDNA. Methods. In this study, ddcfDNA in plasma samples from 223 consecutive kidney transplant recipients was analyzed pretransplantation; at 3, 7, and 180 d after transplantation; and at time of for-cause biopsies obtained within the first 180 d after transplantation. Results. Median (interquartile range) ddcfDNA concentration was significantly higher on day 3 (58.3 [17.7-258.3] copies/mL) and day 7 (25.0 [10.4-70.8] copies/mL) than on day 180 after transplantation (4.2 [0.0-8.3] copies/mL; P < 0.001 and P < 0.001, respectively). At time of biopsy-proven AR (BPAR), between day 11 and day 180 after transplantation, ddcfDNA concentration was significantly higher (50.0 [25.0-108.3] copies/mL) than those when biopsies showed non-AR (0.0 [0.0-15.6] copies/mL; P < 0.05). ddcfDNA concentration within the first 10 d after transplantation showed no significant difference between recipients with BPAR and those with non-AR in their biopsy or between recipients with BPAR and ddcfDNA measured at day 3 and day 7. Conclusions. Unfortunately, ddcfDNA concentration is not a good biomarker to detect AR within the first 10 d after transplantation; however, BPAR occurring after 10 d after transplantation can be detected in kidney transplant recipients by ddcfDNA using a novel and unique high-throughput droplet digital PCR indel method.

AB - Background. Donor-derived cell-free DNA (ddcfDNA) is a promising minimally invasive biomarker for acute rejection (AR) in kidney transplant recipients. To assess the diagnostic value of ddcfDNA as a marker for AR, ddcfDNA was quantified at multiple time points after kidney transplantation with a novel high-throughput droplet digital PCR indel method that allowed for the absolute quantification of ddcfDNA. Methods. In this study, ddcfDNA in plasma samples from 223 consecutive kidney transplant recipients was analyzed pretransplantation; at 3, 7, and 180 d after transplantation; and at time of for-cause biopsies obtained within the first 180 d after transplantation. Results. Median (interquartile range) ddcfDNA concentration was significantly higher on day 3 (58.3 [17.7-258.3] copies/mL) and day 7 (25.0 [10.4-70.8] copies/mL) than on day 180 after transplantation (4.2 [0.0-8.3] copies/mL; P < 0.001 and P < 0.001, respectively). At time of biopsy-proven AR (BPAR), between day 11 and day 180 after transplantation, ddcfDNA concentration was significantly higher (50.0 [25.0-108.3] copies/mL) than those when biopsies showed non-AR (0.0 [0.0-15.6] copies/mL; P < 0.05). ddcfDNA concentration within the first 10 d after transplantation showed no significant difference between recipients with BPAR and those with non-AR in their biopsy or between recipients with BPAR and ddcfDNA measured at day 3 and day 7. Conclusions. Unfortunately, ddcfDNA concentration is not a good biomarker to detect AR within the first 10 d after transplantation; however, BPAR occurring after 10 d after transplantation can be detected in kidney transplant recipients by ddcfDNA using a novel and unique high-throughput droplet digital PCR indel method.

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U2 - 10.1097/TP.0000000000004078

DO - 10.1097/TP.0000000000004078

M3 - Article

C2 - 35283452

SN - 0041-1337

VL - 106

SP - 1777

EP - 1786

JO - Transplantation

JF - Transplantation

IS - 9

ER -

Verhoeven JGHP, Boer K , Peeters AMA , Clahsen-Van Groningen MC , Roodnat JI , Van De Wetering J et al. A Novel High-throughput Droplet Digital PCR-based Indel Quantification Method for the Detection of Circulating Donor-derived Cell-free DNA after Kidney Transplantation. Transplantation. 2022 Sept 1;106(9):1777-1786. doi: 10.1097/TP.0000000000004078, 10.1097/TP.0000000000004078

A Novel High-throughput Droplet Digital PCR-based Indel Quantification Method for the Detection of Circulating Donor-derived Cell-free DNA after Kidney Transplantation

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