Quantitative PCR PCR of INDEL INDEL s to measure donor‐derived cell‐free DNA DNA —a potential method to detect acute rejection in kidney transplantation: a pilot study

摘要

Summary The quantification of donor‐derived cell‐free DNA (ddcf DNA ) in recipient's plasma is a novel, but technically challenging noninvasive method to assist the diagnosis of acute rejection ( AR ). A quantitative real‐time PCR ( qPCR ) approach targeting insertion/deletion polymorphisms ( INDEL ) was adapted to measure ddcf NA in plasma samples from 29 kidney transplant recipients obtained at time of clinically indicated biopsies (eight patients with a histologically verified AR , nine with borderline rejection and 12 without evidence of rejection). Measured ddcf DNA levels of smaller INDEL amplicon targets differed significantly ( P ?=?0.016, Kruskal–Wallis H test) between recipients with biopsy‐proven AR (median 5.24%; range 1.00–9.03), patients without (1.50%; 0.41–6.50) and patients with borderline AR (1.91%; 0.58–5.38). Similarly, pairwise testing by Mann–Whitney U ‐tests revealed significant differences between recipients with AR and without AR ( P ?=?0.012) as well as patients with AR and borderline histology ( P ?=?0.015). Receiver operating characteristic ( ROC ) analysis revealed an area under the ROC curve for discriminating AR and non‐ AR biopsies of 0.84 (95% CI : 0.66–1.00). The determined cutoff value of 2.7% ddcf DNA showed a sensitivity of 0.88 (95% CI : 0.63–1.00) and specificity of 0.81 (95% CI : 0.64–0.98). INDEL qPCR represents a novel method to quantify ddcf DNA on standard qPCR instruments within 6–8?h with high sensitivity and specificity to detect AR.