MTOR Inhibitors Maintain Low Levels of Immune Activation, Immune Senescence and EBV Load in Kidney Transplant Patients

Abstract

Introduction Post-transplant lymphoproliferative disorders (PTLD) represent a severe complication in transplant patients. Epstein-Barr Virus (EBV) is the main driver of PTLD, particularly of those occurring early after transplantation. Immune activation/inflammation, immune senescence, reactivation of EBV and expansion of EBV-positive B cells, which play a pivotal role in the onset of PTLD, may be modulated by immunosuppressive strategies. The aim of this study was to assess the impact of mTOR inhibitor (mTORi) on viral and host cofactors involved in the PTLD’s pathogenesis. Materials and Methods Sixty-two kidney transplant patients were enrolled in the study: 33 were treated with mycophenolic acid (MPA) and 29 with mTORi, both in combination with tacrolimus and steroids. Thymoglobulin was the induction in all cases. All patients were assessed at the time of transplant and after 1 year of follow-up. Markers of T-cell activation(CD3+CD8+CD38+), exhaustion(CD3+CD8+PD-1+), senescence (CD3+CD8+CD28-CD57+), and B-cell activation (CD19+CD10-CD21lowCD27+) were evaluated by flow cytometry. Plasma levels of PAMPs(microbial translocation marker 16s ribosomal(r)DNA) and DAMPs(mitochondrial(mt)DNA) were quantified by real-time PCR. Inflammatory cytokines (IL-6 and TNF-alpha) were quantified by Luminex platform. EBV was typed and quantified by multiplex real-time PCR, and telomere lengths in peripheral blood cells (PBMC) were estimated by telomere/single copy gene ratio (T/S), using multiplex real-time PCR. Results After 1 year of follow-up, plasma levels of 16s rDNA increased in MPA-treated patients (p=0.029), while remained stable in the mTORi group. Levels of mtDNA and pro- inflammatory cytokines tended to decrease in mTORi-treated patients (mtDNA, p=0.034; IL-6, p=0.08; TNF-alpha, p<0.001), while no significant changes were observed in the MPA group. Percentages of activated, exhausted and senescent T cells significantly increased in the MPA-treated patients (4.6[2.2-9.1] vs 17.8[12.1-23.3],p<0.001; 4.8[3.0-6.9] vs 11.6[8.7-16.8],p<0.001; and 7.9[4.4-14.5] vs 14.5[8.5-27.7],p=0.005), but remained stable in the mTORi group (2.4[1.8-6.1] vs 4.1[2.4-6.3], p=0.161; 6.4[4.9-12.1] vs 7.1[4.3-12.7],p=0.888; and 9.7[5.2-13.8]vs 10.8[5.6-16.2],p=0.460). The increase in immunosenescent cells in the MPA group was supported by telomere shortening, higher in MPA than in mTORi-treated patients (ΔT/S=-0.449 vs ΔT/S=-0.219). Percentages of activated B cells and levels of EBV-DNA significantly increased in the MPA group (10.3[6.4-16.9] vs 18.3[11.6-28.4],p=0.002, and 1[1-17] vs 27[1-118] EBV-DNA copies/105 PBMC, p=0.004), while remained stable in the mTORi group (12.8[6.8-18.8]vs 13.0[9.2-18.0], p=0.836, and 1[1-7]vs 6[1-26] EBV-DNA copies/105 PBMC, p=0.128). Conclusions mTORi maintained low levels of pro-inflammatory cytokines, immune activation and senescence, thus constraining EBV reactivation and expansion of EBV-positive B cells. Overall, these findings suggest that mTORi may reduce the risk of EBV-related malignancies in kidney transplant patients. AIRC grant IG2016 Id 19112.