Donor-Derived Cell-Free DNA Is a Novel Universal Biomarker for Allograft Rejection in Solid Organ Transplantation

Beck, Julia; Oellerich, Michael; Schulz, Uwe; Schauerte, Verena; Reinhard, Linda; Fuchs, U; Knabbe, Cornelius; Zittermann, Armin; Olbricht, Christoph J.; Gummert, Jan; Shipkova, Maria; Birschmann, Ingvíld; Wieland, Eberhard; Schütz, Ekkehard

doi:10.1016/j.transproceed.2015.08.035

Cited by 113 publications

(100 citation statements)

References 8 publications

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“…It is well known from cfDNA-based noninvasive prenatal testing that the body mass index (BMI) can also play a confounding role [28], e.g., higher values of cfDNA for patients with extremely low BMI and lower values for patients with high BMI. For such cases, the percentage values can be expressed as copies/milliliter plasma by adding a total cfDNA quantification [12,16]. Overall, for LTx this does not seem to be necessary, most likely because of the high percentage of graft DNA.…”

Section: Discussionmentioning

confidence: 99%

Graft-derived cell-free DNA, a noninvasive early rejection and graft damage marker in liver transplantation: A prospective, observational, multicenter cohort study

Schütz¹,

Fischer

Beck³

et al. 2017

PLoS Med

173

152

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BackgroundGraft-derived cell-free DNA (GcfDNA), which is released into the blood stream by necrotic and apoptotic cells, is a promising noninvasive organ integrity biomarker. In liver transplantation (LTx), neither conventional liver function tests (LTFs) nor immunosuppressive drug monitoring are very effective for rejection monitoring. We therefore hypothesized that the quantitative measurement of donor-derived cell-free DNA (cfDNA) would have independent value for the assessment of graft integrity, including damage from acute rejection.Methods and findingsTraditional LFTs were performed and plasma GcfDNA was monitored in 115 adults post-LTx at three German transplant centers as part of a prospective, observational, multicenter cohort trial. GcfDNA percentage (graft cfDNA/total cfDNA) was measured using droplet digital PCR (ddPCR), based on a limited number of predefined single nucleotide polymorphisms, enabling same-day turn-around. The same method was used to quantify blood microchimerism. GcfDNA was increased >50% on day 1 post-LTx, presumably from ischemia/reperfusion damage, but rapidly declined in patients without graft injury within 7 to 10 d to a median <10%, where it remained for the 1-y observation period. Of 115 patients, 107 provided samples that met preestablished criteria. In 31 samples taken from 17 patients during biopsy-proven acute rejection episodes, the percentage of GcfDNA was elevated substantially (median 29.6%, 95% CI 23.6%–41.0%) compared with that in 282 samples from 88 patients during stable periods (median 3.3%, 95% CI 2.9%–3.7%; p < 0.001). Only slightly higher values (median 5.9%, 95% CI 4.4%–10.3%) were found in 68 samples from 17 hepatitis C virus (HCV)–positive, rejection-free patients. LFTs had low overall correlations (r = 0.28–0.62) with GcfDNA and showed greater overlap between patient subgroups, especially between acute rejection and HCV+ patients. Multivariable logistic regression modeling demonstrated that GcfDNA provided additional LFT-independent information on graft integrity. Diagnostic sensitivity and specificity were 90.3% (95% CI 74.2%–98.0%) and 92.9% (95% CI 89.3%–95.6%), respectively, for GcfDNA at a threshold value of 10%. The area under the receiver operator characteristic curve was higher for GcfDNA (97.1%, 95% CI 93.4%–100%) than for same-day conventional LFTs (AST: 95.7%; ALT: 95.2%; γ-GT: 94.5%; bilirubin: 82.6%). An evaluation of microchimerism revealed that the maximum donor DNA in circulating white blood cells was only 0.068%. GcfDNA percentage can be influenced by major changes in host cfDNA (e.g., due to leukopenia or leukocytosis). One limitation of our study is that exact time-matched GcfDNA and LFT samples were not available for all patient visits.ConclusionsIn this study, determination of GcfDNA in plasma by ddPCR allowed for earlier and more sensitive discrimination of acute rejection in LTx patients as compared with conventional LFTs. Potential blood microchimerism was quantitatively low and had no significant influence on GcfDNA value. Further r...

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Section: Discussionmentioning

confidence: 99%

Graft-derived cell-free DNA, a noninvasive early rejection and graft damage marker in liver transplantation: A prospective, observational, multicenter cohort study

Schütz¹,

Fischer

Beck³

et al. 2017

PLoS Med

173

152

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“…Characterization of effects of miscellaneous other acute medical conditions is outside the scope of the current study, which identified patients who were stable and not having acute systemic inflammatory disorders such as sepsis. The very low levels of dd-cfDNA found were quantified in relation to the large amounts of cfDNA from the recipient; this method (ratio) has been used in the majority of published studies of dd-cfDNA, and reproducibly has been associated with rejection across organ types (2)(3)(4)(5)(6)(7)(8). Although determining levels of dd-cfDNA may not eliminate the need for biopsy to aid in the confirmation of a specific histopathology, its results could increase the prebiopsy probability of detecting injury, thereby making biopsy a more effective diagnostic tool.…”

Section: Limitations Of This Studymentioning

confidence: 99%

“…Observations of increased levels of ddcfDNA during acute rejection and correlation to severity of rejection have indicated the potential utility of dd-cfDNA as an early noninvasive indicator of allograft injury (2)(3)(4)(5)(6)(7)(8). Among the various strategies to measure dd-cfDNA, we have demonstrated the analytical validity of a targeted next-generation sequencing assay that uses 266 single-nucleotide polymorphisms (SNPs) to accurately quantify dd-cfDNA in the plasma of transplant recipients without the need for genotyping either the donor or the recipient (6).…”

mentioning

confidence: 99%

Biological Variation of Donor-Derived Cell-Free DNA in Renal Transplant Recipients: Clinical Implications

Bromberg

Brennan

Poggio

et al. 2017

The Journal of Applied Laboratory Medicine

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Background Previous studies have demonstrated that donor-derived cell-free DNA (dd-cfDNA) found in circulating blood of transplant recipients may serve as a noninvasive biomarker of allograft rejection. To better interpret the clinical meaning of dd-cfDNA, it is essential to understand the biological variation of this biomarker in stable healthy recipients. This report establishes the biological variation and clinical reference intervals of dd-cfDNA in renal transplant recipients by using an analytically validated assay that has a CV of 6.8%. Methods We sampled venous blood at patient surveillance visits (typically at posttransplant months 1–4, 6, 9, and 12) in a 14-center observational study. Patients with stable renal allograft function spanning ≥3 serial visits were selected. We used AlloSure®, a targeted next-generation sequencing-based approach, to measure dd-cfDNA in the plasma and computed the intraindividual CV (CVI) and interindividual CV (CVG), the index of individuality (II), and reference change value (RCV). Results Of 93 patients, 61% were men, 56% were Caucasian, mean age was 49 years, and 63% were deceased donor kidney recipients. Of 380 blood samples, the dd-cfDNA median value was 0.21% (interquartile range 0.12%–0.39%) and the 97.5th percentile was 1.20%. In 18 patients with an average of 4.1 tests, the CVI was 21%, CVG was 37%, II was 0.57, and RCV was 61%. Conclusions In a renal transplant recipient, a dd-cfDNA level above 1.2% is out of range and potentially abnormal. A serial increase of up to 61% in level of dd-cfDNA in a patient may be attributable to biological variation. Clinicaltrials.gov Identifier: NCT02424227

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“…5,7 In stable lung and liver transplant recipients, the level of ddcfDNA is higher than in stable heart transplant recipients, and it further increases in moderate-to-severe rejection. 8,9 Up to now, dd-cfDNA has been least studied in renal transplants; levels in stable kidney recipients are similar to those in heart transplant recipients, 4,10 and analyses of individual patients and a small single-center study identified higher levels during biopsy-proven acute rejection. 11 In kidney transplantation, there are no existing biomarkers that adequately measure the status of active injury to the allograft.…”

mentioning

confidence: 99%

Cell-Free DNA and Active Rejection in Kidney Allografts

Bloom

Bromberg²,

Poggio

et al. 2017

JASN

421

493

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Histologic analysis of the allograft biopsy specimen is the standard method used to differentiate rejection from other injury in kidney transplants. Donor-derived cell-free DNA (dd-cfDNA) is a noninvasive test of allograft injury that may enable more frequent, quantitative, and safer assessment of allograft rejection and injury status. To investigate this possibility, we prospectively collected blood specimens at scheduled intervals and at the time of clinically indicated biopsies. In 102 kidney recipients, we measured plasma levels of dd-cfDNA and correlated the levels with allograft rejection status ascertained by histology in 107 biopsy specimens. The dd-cfDNA level discriminated between biopsy specimens showing any rejection (T cell-mediated rejection or antibody-mediated rejection [ABMR]) and controls (no rejection histologically), P,0.001 (receiver operating characteristic area under the curve [AUC], 0.74; 95% confidence interval [95% CI], 0.61 to 0.86). Positive and negative predictive values for active rejection at a cutoff of 1.0% dd-cfDNA were 61% and 84%, respectively. The AUC for discriminating ABMR from samples without ABMR was 0.87 (95% CI, 0.75 to 0.97). Positive and negative predictive values for ABMR at a cutoff of 1.0% dd-cfDNA were 44% and 96%, respectively. Median dd-cfDNA was 2.9% (ABMR), 1.2% (T cell-mediated types $IB), 0.2% (T cellmediated type IA), and 0.3% in controls (P=0.05 for T cell-mediated rejection types $IB versus controls). Thus, ddcfDNA may be used to assess allograft rejection and injury; dd-cfDNA levels ,1% reflect the absence of active rejection (T cell-mediated type $IB or ABMR) and levels .1% indicate a probability of active rejection.

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Donor-Derived Cell-Free DNA Is a Novel Universal Biomarker for Allograft Rejection in Solid Organ Transplantation

Cited by 113 publications

References 8 publications

Graft-derived cell-free DNA, a noninvasive early rejection and graft damage marker in liver transplantation: A prospective, observational, multicenter cohort study

Graft-derived cell-free DNA, a noninvasive early rejection and graft damage marker in liver transplantation: A prospective, observational, multicenter cohort study

Biological Variation of Donor-Derived Cell-Free DNA in Renal Transplant Recipients: Clinical Implications

Cell-Free DNA and Active Rejection in Kidney Allografts

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