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

Background: Previous studies have demonstrated that donor-derived cell-free DNA (dd-cfDNA) found in circulating blood of transplant recipientsmay serve as a noninvasive biomarker of allograft rejection. To better interpret the clinicalmeaning 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 posttransplantmonths 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:Of93patients, 61%weremen, 56%wereCaucasian,meanagewas49 years, and63%weredeceaseddonor kidney recipients. Of 380 blood samples, the dd-cfDNA median value was 0.21% (interquartile range 0.12%–0.39%) and the 97.5th percentilewas 1.20%. In 18 patientswith an average of 4.1 tests, the CVI was 21%, CVGwas 37%, II was 0.57, and RCVwas 61%. Conclusions: In a renal transplant recipient, add-cfDNA levels above1.2% isoutof rangeandpotentially 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