Longitudinal monitoring of mRNA levels of regulatory T cell biomarkers by using noninvasive strategies to predict outcome in renal transplantation

Background. Acute T-cell mediated rejection (aTCMR) is still an issue in kidney transplantation, for it is associated with chronic rejection, graft loss, and overall worse outcomes. For these reasons, a standard non-invasive molecular tool to detect is desirable to offer a simpler monitoring of kidney transplant recipients (KTRs). The purpose of our study was to examine, in peripheral blood before and after transplantation, the expression patterns of regulatory T cell (Treg)-related genes: the forkhead box P3 (FOXP3) and the two CTLA-4 isoforms (full-length and soluble) to predict acute rejection onset, de novo donor-specific antibodies (DSA) development and renal dysfunction one year after transplantation. Methods. We profiled by using a relative quantification analysis (qRT-PCR) circulating mRNA levels of these biomarkers in peripheral blood of 89 KTRs within the first post-transplant year (at baseline and 15, 60 and 365 days, and when possible at the acute rejection) and compared also the results with 24 healthy controls. Results. The three mRNA levels drastically reduced 15 days after transplantation and gradually recovered at one year in comparison with baseline, with very low levels at the time of aTCMR for FOXP3 (RQ=0.445, IQR=0.086-1.264, p=0.040), maybe for the pro-apoptotic role of FOXP3 during inflammation. A multivariate Cox regression analysis evidenced a significant relation between aTCMR onset and thymoglobuline induction (HR=6.749 p=0.041), everolimus use (HR=7.017, p=0.007) and an increased risk from the solCTLA-4 expression at 15 days, mainly considering recipients treated with Mycophelolic acid (HR=13.94 p=0.038, 95%CI:1.157-167.87). Besides, solCTLA-4 also predisposed to graft dysfunction (eGFR<60mL/min/1.73m2) at one year (AOR=3.683, 95%CI=1.145-11.845, p=0.029). On the other hand, pre-transplant solCTLA-4 levels showed a protective association with de novo DSAs development (HR=0.189, 95%CI=0.078-0.459, p<0.001). Conclusions. mRNA levels of Treg-associated genes, mainly for solCTLA-4, in peripheral blood could put forward as candidate non-invasive biomarkers of cellular and humoral alloreactivity in clinical transplantation and might help shape immunosuppression, tailor monitoring and achieve better long-term outcomes of kidney transplantation in the wake of "precision medicine".