Systemic immunoregulatory and proteogenomic effects of tacrolimus to sirolimus conversion in liver transplant recipients

Abstract

Immunosuppression withdrawal from calcineurin inhibitors is only possible in ~20% of liver transplant recipients. However, mTOR inhibitors (sirolimus) appear to be more immunoregulatory and might promote a tolerant state for withdrawal. Our purpose was to determine if systemic (blood, marrow, allograft) signatures of immunoregulation are promoted by conversion from tacrolimus to sirolimus. We therefore performed the following serial assays before and after sirolimus conversion in liver transplant recipients to test for enhanced markers of immunoregulation: 1) Flow cytometry immunophenotyping of PBMC and bone marrow aspirates for regulatory T cells (Tregs: CD4+CD25+++FOXP3+) and regulatory dendritic cells (DCregs: ILT3+/4+); 2) liver biopsy immunohistochemical staining (FOXP3:CD3, CD4:CD8 ratios) and immunophenotyping of biopsy-derived Tregs after growth in culture; 3) effects of pre-vs. post-conversion sera on Treg generation in mixed lymphocyte reactions; 4) peripheral blood non-specific CD4 responses (Cylex® ImmuKnow); 5) peripheral blood gene transcripts and proteomic profiles. We successfully converted 20 non-immune, non-viremic recipients (age 57.2±8; 3.5±2.1 years post-LT) from tacrolimus to sirolimus for renal dysfunction. Our results demonstrated significant increases in Tregs in PBMC and marrow and DCregs in PBMC (p<0.01) following conversion. In biopsy immunohistochemistry, FOXP3:CD3 and CD4:CD8 ratios were significantly higher after conversion and a number of biopsy cultures developed new or higher FOXP3+ cell growth. Non-specific CD4 responses (Cylex® ImmuKnow) did not change. Both pre- and post-conversion sera inhibited mixed lymphocyte reactions, although only tacrolimus sera suppressed Treg generation. Finally, 289 novel genes and 22 proteins, several important in immunoregulatory pathways, were expressed after conversion. Conclusions Tacrolimus to sirolimus conversion increases systemic Tregs, DCregs and immunoregulatory proteogenomic signatures in liver transplant recipients and may therefore facilitate immunosuppression minimization or withdrawal.

Differential Effects of Calcineurin and Mammalian Target of Rapamycin Inhibitors on Alloreactive Th1, Th17, and Regulatory T Cells

“…On the other hand, SRL markedly increased the proportion of FOXP3 + Tregs compared to a dose-dependent decline mediated by TAC ( Figure 4A), akin to the observations made in vivo in transplant patients converted from TAC to SRL treatment. [39][40][41][42] Additionally, SRL was able to abrogate the inhibition of Treg expansion mediated by TAC (Figure 4). However, both agents significantly inhibited the expansion of the absolute number of cells in the cultures ( Figure 4B).…”

Integrating canonical and metabolic signalling programmes in the regulation of T cell responses

“…For example, whereas calcineurin inhibitors (such as cyclosporine A and FK506) block the generation of T Reg cells, mTOR inhibition does not 118 . To this end, treating liver transplant recipients with siro-limus (an mTOR inhibitor), rather than FK506, increases the number of liver FOXP3 + T Reg cells 119 . Likewise, a calci neurin inhibitor-free regimen (using sirolimus) has been developed to promote stable mixed donor chimer-ism after non-myeloablative allogeneic haematopoietic stem cell transplantation for adult sickle cell disease 120 .…”

Impact of Immune-Modulatory Drugs on Regulatory T Cell

“…As a result, the OPTN/SRTR Annual Data Report in 2013 recommend against CNI withdrawal 82 . The finding that Treg are resistant to low-dose CNI and combing CNIs with sirolimus restore Treg 93,104 suggest that reducing CNI dose may not only spare patients from the nephrotoxicity, but also spare Treg.…”