Development of a multivariable gene-expression signature targeting T-cell-mediated rejection in peripheral blood of kidney transplant recipinets validated in cross sectional and longitudinal samples

Background: Acute T-cell mediated rejection (TCMR) is usually indicated by alteration in serum-creatinine measurements when considerable transplant damage has already occurred. There is, therefore, a need for non-invasive early detection of immune signals that would precede the onset of rejection, prior to transplant damage. Methods: We examined the RT-qPCR expression of 22 literature-based genes in peripheral blood samples from 248 patients in the Kidney Allograft Immune Biomarkers of Rejection Episodes (KALIBRE) study. To account for post-transplantation changes unrelated to rejection, we generated time-adjusted gene-expression residuals from linear mixed-effects models in stable patients. To select genes, we used penalised logistic regression based on 27 stable patients and 27 rejectors with biopsy-proven T-cell-mediated rejection, fulfilling strict inclusion/exclusion criteria. We validated this signature in i) an independent group of stable patients and patients with concomitant T-cell and antibody-mediated-rejection, ii) patients from an independent study, iii) cross-sectional pre-biopsy samples from non-rejectors and iv) longitudinal follow-up samples covering the first posttransplant year from rejectors, non-rejectors and stable patients. Findings: A parsimonious TCMR-signature (IFNG, IP-10, ITGA4, MARCH8, RORc, SEMA7A, WDR40A) showed cross-validated area-under-ROC curve 0·84 (0·77-0·88) (median, 2·5th-97·5th centile of fifty cross-validation cycles), sensitivity 0·67 (0·59-0·74) and specificity 0·85 (0·75-0·89). The estimated probability of TCMR increased seven weeks prior to the diagnostic biopsy and decreased after treatment. Gene expression in all patients showed pronounced variability, with up to 24% of the longitudinal samples in stable patients being TCMR-signature positive. In patients with borderline changes, up to 40% of pre-biopsy samples were TCMR-signature positive. Interpretation Molecular marker alterations in blood emerge well ahead of the time of clinically overt TCMR. Monitoring a TCMR-signature in peripheral blood could unravel Tcell-related pro-inflammatory activity and hidden immunological processes. This additional information could support clinical management decisions in cases of patients with stable but poor kidney function or with inconclusive biopsy results