As the interest in telehealth and remote monitoring increases,1 many transplant practitioners have turned to donor-derived cell-free DNA (dd-cfDNA) to noninvasively monitor the immunologic status of allografts. As use of this testing grows, it is important to highlight limitations in its predictive value. There are several available dd-cfDNA tests. In kidney transplantation, one of the earliest to market was AlloSure (CareDx, Inc.), the results of which were published in JASN in 2017.2 Two features of the original Circulating Donor-Derived Cell-Free DNA in Blood for Diagnosing Acute Rejection in Kidney Transplant Recipients (DART) trial2 analysis may lead to confusion when interpreting dd-cfDNA testing. First, borderline rejections with allograft dysfunction were considered “no active rejection,” even though many centers treat these findings as acute rejection.3 Table 1 is adapted from figure 1 of the DART publication.2 When the study data are reanalyzed to reclassify patients with borderline rejection associated with graft dysfunction, there were 46 patients (43%) with rejection and 61 (57%) without rejection. The assay’s sensitivity for all rejection decreases to only 41%, with specificity of 82% and negative likelihood ratio of 0.72. As a result, the test will only have a reasonable negative predictive value when the pretest probability of rejection is very low.
Table 1.
Reassignment of DART trial cohort status, classifying patients with borderline changes and graft dysfunction as having T cell–mediated rejection
| Diagnosis | Total | dd-cfDNA<1% | dd-cfDNA>1% |
|---|---|---|---|
| No rejection | 61 | 50 (82%) | 11 (18%) |
| All rejection | 46 | 27 (59%) | 19 (41%) |
| Borderline | 19 | 17 (89%) | 2 (11%) |
| 1A | 5 | 5 (100%) | 0 (0%) |
| 1B | 5 | 2 (40%) | 3 (60%) |
| 2A | 1 | 1 (100%) | 0 (0%) |
| AMR | 8 | 0 (0%) | 8 (100%) |
| Mixed ACR/AMR | 8 | 2 (25%)a | 6 (75%)b |
ACR, acute cellular rejection.
One patient had mixed 1A and AMR, and one patient had mixed 2A and AMR.
Three patients had mixed borderline and AMR, two patients had mixed 1B and AMR, and one patient had mixed 2A and AMR.
Second, acute cellular rejections were grouped together with antibody-mediated rejection (AMR), implying that the excellent performance of the dd-cfDNA assay in AMR also applies to acute cellular rejection. When excluding mixed rejections, only five of 30 purely cellular rejections test positive, including only half of patients with severe rejections (1B or worse). Because most kidney allograft rejections are cellular, using this test to rule out rejection will miss a significant proportion of patients.4 In our center, we diagnosed 187 kidney rejections from January 2016 to December 2019 (borderline: 82 [44%]; 1A: 33 [18%]; 1B: 28 [15%]; grade 2/3: 28 [15%]; AMR: 23 [12%]). If dd-cfDNA testing had been used to screen for immunologically quiescent patients, an overall sensitivity of 41% would have led to 110 missed rejections.
All tests have limitations. Kidney allograft biopsy, the gold standard, is invasive, uncomfortable, and subject to its own difficulties with interpretation. The dd-cfDNA assay has a role in transplantation, particularly when following AMR and microvascular inflammation. However, clinicians must recognize that many patients with rejection will have reassuring dd-cfDNA results, so negative tests must be received with caution. On the basis of currently available data, negative dd-cfDNA alone should not be considered to “rule out” rejection in cases where the pretest probability of rejection is not already low.
Disclosures
S. Ali Husain reports grants from the National Center for Advancing Translational Sciences, outside the submitted work. All remaining authors have nothing to disclose.
Funding
None.
Footnotes
Published online ahead of print. Publication date available at www.jasn.org.
References
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