BACKGROUND

Monitoring donor-derived cell-free DNA (dd-cfDNA) is a promising noninvasive method for assessing allograft health in renal transplant recipients. This UK pilot study evaluated a novel insertion and deletion (INDEL)-based next-generation sequencing (NGS) assay for detecting dd-cfDNA and explored its association with potentially injurious concomitant pathologies, including donor-specific antibodies. Current methods are limited to first and only transplant recipients, as other assays cannot distinguish graft injury in the context of transplantation from multiple donors.

METHODS

Fourteen high-risk renal transplant recipients (level IV human leukocyte antigen mismatch, calculated reaction frequency >20%, retransplant) were recruited between October 2023 and July 2024 at Liverpool University Hospitals. Plasma samples were collected 6 months posttransplant, and cfDNA was extracted using QIAsymphony DSP Circulating DNA Kit (Qiagen). dd-cfDNA was quantified using the Devyser Accept cfDNA assay (Devyser), and NGS was performed using MiSeq (Illumina).

RESULTS

We present preliminary observations from the first 14 patients included in this proof-of-concept arm of the study. A dd-cfDNA level ≤0.5% correlated with stable graft function (n=11). Patients with dd-cfDNA ≥1.0% had supratherapeutic tacrolimus levels (n=2). Intermediate dd-cfDNA levels (0.5%-1.0%) were found in the setting of donor-specific antibody emergence (n=1). We were able to identify informative markers and derive interpretable results in a multitransplant recipient setting.

CONCLUSIONS

The INDEL-based NGS assay is a promising novel tool for detecting and monitoring dd-cfDNA in renal transplant recipients with an easy-to-implement workflow. These preliminary results support its clinical utility in a high-immunological-risk setting. These findings are consistent with emergent literature; however, longitudinal data and further validation in a larger cohort of patients are required.