Diagnostic Performance of Donor‐derived Cell‐free DNA Combined With Immune Activation Markers for Detecting Acute Rejection After Liver Transplantation

Abstract

BACKGROUND: Accurate differentiation of acute rejection (AR) from other causes of graft dysfunction after liver transplantation remains challenging in real-world clinical practice. Although donor-derived cell-free DNA (dd-cfDNA) has emerged as a promising non-invasive biomarker of graft injury, its specificity may be limited in clinically suspected rejection due to concurrent inflammatory and non-rejection-related graft injuries. Immune activation markers may provide complementary information by reflecting ongoing alloimmune responses. METHODS: In this prospective diagnostic study, we enrolled liver transplant recipients presenting with graft dysfunction and classified them as AR or non-rejection based on biopsy. Plasma dd-cfDNA levels and peripheral immune activation markers, including interleukin-2 receptor (IL-2R) and activated T-cell subsets, were measured. Diagnostic performance was evaluated using receiver operating characteristic analysis and multivariable models. RESULTS: A total of 33 patients were included, comprising 18 with biopsy-proven AR and 15 with non-rejection graft dysfunction. dd-cfDNA levels were significantly higher in AR patients. Selected immune activation markers demonstrated complementary diagnostic characteristics. A simplified combined model integrating dd-cfDNA percentage with activated T-cell subsets achieved improved discrimination compared with individual markers. Decision curve analysis indicated a higher net clinical benefit across a range of threshold probabilities. CONCLUSION: Integration of dd-cfDNA with immune activation markers provides a complementary non-invasive strategy for evaluating acute rejection in liver transplant recipients with clinically suspected graft dysfunction. This approach may improve real-world diagnostic assessment and assist clinical decision-making while reducing unnecessary liver biopsies.