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

Life-long immunosuppression (IS) is generally required after liver transplantation (LT). With the advent of calcineurin inhibitors (CNI), rejection rates have declined, yet toxicity due to CNI therapy has led to long term adverse outcomes (1). Complete IS withdrawal (operational tolerance) would be ideal, although this has thus far been possible in only ~20% of LT recipients (2). The inability to immunologically predict successful IS withdrawal has obligated long term CNI maintenance at therapeutic doses, despite toxicities. The identification of specific cell populations and pathways responsible for immunoregulation may give clues towards achieving tolerance in LT. Tolerance develops initially by the interaction of antigen-specific T cells with unique thymic antigen-presenting cells (APC) or regulatory dendritic cells (DCregs), respectively resulting in either clonal deletion, anergy or an active immunoregulatory process (3). Such DCregs are characterized by high surface expression of CD123 and/or immunoglobulin-like transcripts ILT3 or ILT4 that inhibit antigen presentation, i.e. reflecting immunoregulation (4, 5). As mentioned, this interaction can lead to the generation of Tregs (CD4+CD25high) that migrate peripherally to control immune responses. These Tregs typically express an intracellular protein FOXP3 which blocks transcription of T cell activation molecules such as IL-2 and the expression of CD127 (6, 7). Moreover, gene transcripts and protein expression patterns (antibodies, circulating and cell proteins) as markers for immunoregulation may also provide a window into the tolerant state. There is thus strong interest in cellular (Treg, DCreg), genomic and proteomic assays to assess immunoregulation and predict more reliably who might achieve IS withdrawal (8, 9). As the liver appears to be the most tolerogenic transplanted organ, high numbers of such cells residing in the liver may be regulatory and protect against rejection. Similarly, the bone marrow, as a formative organ of the immune system may also contain localized Treg niches that enhance immunoregulation (10–12). Thus, assessing tolerogenic signatures not only in the blood but also in the allograft and bone marrow may further differentiate regulatory from alloreactive states. In addition to the need for predictive assays, there have been few efforts to modulate the immune system to induce tolerance other than simultaneous hematopoietic cell and solid organ transplantation (13), a risky approach in patients with severe end stage liver disease undergoing LT. One potential reason for the low efficacy of IS withdrawal is CNI therapy itself, as most patients in such studies were being treated with CNIs at withdrawal (2). CNIs may hinder immunoregulation by inhibiting IL-2 production needed for Treg development and also by not inhibiting DC maturation (14, 15). In contrast, mTOR inhibitors such as sirolimus (SRL) might facilitate immunoregulation by increasing Treg percentages (does not block IL-2 production) and inhibiting DC maturation and function (16–18). We previously demonstrated that LT recipients on SRL monotherapy had significantly higher Treg percentages than those on tacrolimus (TAC) monotherapy (19). Thus, conversion to SRL may both minimize CNI toxicity (20) and facilitate IS withdrawal by promoting immunoregulation. We therefore hypothesized that prospectively studied LT recipients converted from CNI to SRL monotherapy would display enhanced systemic cellular, functional and proteogenomic immunoregulatory markers. This would support larger scale studies utilizing SRL conversion as an intermediate step toward more clinically successful IS withdrawal in LT.