Combining Donor-derived Cell-free DNA Fraction and Quantity to Detect Kidney Transplant Rejection Using Molecular Diagnoses and Histology as Confirmation

Abstract

Background. Donor-derived cell-free DNA (dd-cfDNA) fraction and quantity have both been shown to be associated with allograft rejection. The present study compared the relative predictive power of each of these variables to the combination of the two, and developed an algorithm incorporating both variables to detect active rejection in renal allograft biopsies. Methods. The first 426 sequential indication biopsy samples collected from the Trifecta study ( ClinicalTrials.gov # NCT04239703) with microarray-derived gene expression and dd-cfDNA results were included. After exclusions to simulate intended clinical use, 367 samples were analyzed. Biopsies were assessed using the molecular microscope diagnostic system and histology (Banff 2019). Logistic regression analysis examined whether combining dd-cfDNA fraction and quantity adds predictive value to either alone. The first 149 sequential samples were used to develop a two-threshold algorithm and the next 218 to validate the algorithm. Results. In regression, the combination of dd-cfDNA fraction and quantity was found to be significantly more predictive than either variable alone ( P = 0.009 and P < 0.0001). In the test set, the area under the receiver operating characteristic curve of the two-variable system was 0.88, and performance of the two-threshold algorithm showed a sensitivity of 83.1% and specificity of 81.0% for molecular diagnoses and a sensitivity of 73.5% and specificity of 80.8% for histology diagnoses. Conclusions. This prospective, biopsy-matched, multisite dd-cfDNA study in kidney transplant patients found that the combination of dd-cfDNA fraction and quantity was more powerful than either dd-cfDNA fraction or quantity alone and validated a novel two-threshold algorithm incorporating both variables.

Anti-interleukin-6 Antibody Clazakizumab in Antibody-mediated Kidney Transplant Rejection: Effect on Donor-derived Cell-free DNA and C-X-C Motif Chemokine Ligand 10

“…Finally, a possible limitation could be that data on absolute levels of dd-cfDNA, suggested by some authors to add diagnostic accuracy, were not available for our study. 43,44 In conclusion, the results of our study suggest that IL-6 blockade in late AMR may not trigger a significant decline in dd-cfDNA[%] and CXCL10 levels, at least over a 9-12 mo treatment period. These 2 noninvasive biomarkers may not be sensitive enough to timely detect subtle treatment effects of IL-6 neutralization in patients with late AMR.…”

The Current State of Donor-Derived Cell-Free DNA Use in Allograft Monitoring in Kidney Transplantation

“…In short, the absence of ddcfDNA is a better predictor of the state of an allograft, in terms of rejection, than the presence of ddcfDNA, which may represent driving forces other than rejection. Outside of the landmark original validation studies, this finding has been reproduced by several groups, with NPV being consistently higher than PPV [ 18 , 19 , 20 , 21 , 22 ]. Additionally, the clinical performance of commercially available ddcfDNA tests appears to be similar (commonly used cutoff value of ~1%) despite different validation strategies [ 23 , 24 ].…”

Biomarkers of alloimmune events in pediatric kidney transplantation

“…In Trifecta (n = 367 adults), patients with dd-cfDNA > 1% or absolute dd-cfDNA > 78 copies/ml had a sensitivity of 74%, specificity of 81%, PPV of 71%, NPV of 83%, and AUROC of 0.82 for diagnosing clinical acute rejection by traditional histology. The authors speculated that the improved diagnostic performance of the two-threshold algorithm was related to dd-cfDNA quantity being more sensitive to acute rejection where systemic inflammation causes high total cfDNA levels and dd-cfDNA fraction being more sensitive in cases where acute rejection was the primary source of inflammation (28,31).…”