Background: Donor-derived cell-free DNA (dd-cfDNA) is considered a reliable marker of organ damage with potential applications in the follow-up of transplant recipients. Methods: In this work we present an assay based on the donor-recipient HLA-mismatch (human leukocyte antigen) at the HLA-DRB1 locus to monitor rejection by quantifying the percentage of dd-cfDNA using a droplet digital PCR (polymerase chain reaction) technique. A panel of probes targeting the HLA-DRB1 locus and covering >85% genetic variability was validated and used to assess dd-cfDNA levels in a prospective cohort of 19 adult heart transplant recipients (mean age 50.9±14.8 years). The assay was carried out on a total of 232 liquid biopsies collected at the same time as endomyocardial biopsy (EMB) during routine post-transplant follow-up. Results: Results show a significant increase of dd-cfDNA related to ischemia-reperfusion injury (2.22±2.09%) and to acute cellular rejection (1.71±3.10%) compared to stable conditions (0.43±1.04%, p < 0.0001). On the contrary, no increase was observed during infections or vascular complications, underlining the potential role of this biomarker for rejection monitoring. With a cut-off of 0.11%, the test showed 70.8% specificity (95% CI, 58.17% - 81.40%) and 64.2% sensitivity (95% CI, 49.80% - 76.86%) in discriminating acute rejection from no rejection. Conclusions: These data demonstrate that this HLA mismatch-based droplet digital PCR method is effective for monitoring rejection in heart transplant recipients. Compared to next generation sequencing approaches, it is far more flexible, less expensive and provides faster results.

HLA-DRB1 mismatch-based identification of donor-derived cell free DNA (dd-cfDNA) as a marker of rejection in heart transplant recipients: A single-institution pilot study.

Monica Sorbini
First
;
Gabriele Maria Togliatto;Erika Simonato;Massimo Boffini
;
Margherita Cappuccio;Alessandro Gambella;Francesca Arruga;Matteo Marro;Cristiana Caorsi;Morteza Mansouri;Mauro Giulio Papotti;Antonio Amoroso;Mauro Rinaldi;Tiziana Vaisitti
Co-last
;
Silvia Deaglio
Co-last
2021

Abstract

Background: Donor-derived cell-free DNA (dd-cfDNA) is considered a reliable marker of organ damage with potential applications in the follow-up of transplant recipients. Methods: In this work we present an assay based on the donor-recipient HLA-mismatch (human leukocyte antigen) at the HLA-DRB1 locus to monitor rejection by quantifying the percentage of dd-cfDNA using a droplet digital PCR (polymerase chain reaction) technique. A panel of probes targeting the HLA-DRB1 locus and covering >85% genetic variability was validated and used to assess dd-cfDNA levels in a prospective cohort of 19 adult heart transplant recipients (mean age 50.9±14.8 years). The assay was carried out on a total of 232 liquid biopsies collected at the same time as endomyocardial biopsy (EMB) during routine post-transplant follow-up. Results: Results show a significant increase of dd-cfDNA related to ischemia-reperfusion injury (2.22±2.09%) and to acute cellular rejection (1.71±3.10%) compared to stable conditions (0.43±1.04%, p < 0.0001). On the contrary, no increase was observed during infections or vascular complications, underlining the potential role of this biomarker for rejection monitoring. With a cut-off of 0.11%, the test showed 70.8% specificity (95% CI, 58.17% - 81.40%) and 64.2% sensitivity (95% CI, 49.80% - 76.86%) in discriminating acute rejection from no rejection. Conclusions: These data demonstrate that this HLA mismatch-based droplet digital PCR method is effective for monitoring rejection in heart transplant recipients. Compared to next generation sequencing approaches, it is far more flexible, less expensive and provides faster results.
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https://reader.elsevier.com/reader/sd/pii/S1053249821023159?token=2C4153CE52922B6789E5ECDD3C95A2CD092F53DE98BE3222513755EF305B382B47727A6F017CC5C6C507652645B972F0&amp;originRegion=eu-west-1&amp;originCreation=20210623162616
heart transplant;rejection;marker;HLA;cell-free circulatingDNA;droplet digital PCR
Monica Sorbini, Gabriele Maria Togliatto, Erika Simonato, Massimo Boffini, Margherita Cappuccio, Alessandro Gambella, Francesca Arruga, Nicola Mora, Matteo Marro, Cristiana Caorsi, Morteza Mansouri, Paola Magistroni, Luisa Delsedime, Mauro Giulio Papotti, Antonio Amoroso, Mauro Rinaldi, Tiziana Vaisitti, Silvia Deaglio
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/2318/1792061
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