Abstract
Pharmacokinetic immunosuppressive drug monitoring poorly correlates with clinical outcomes after solid organ transplantation. A promising method for pharmacodynamic monitoring of tacrolimus (TAC) in T cell subsets of transplant recipients might be the measurement of (phosphorylated) p38MAPK, ERK1/2 and Akt (activated downstream of the T cell receptor) by phospho-specific flow cytometry. Here, blood samples from n = 40 kidney transplant recipients (treated with either TAC-based or belatacept (BELA)-based immunosuppressive drug therapy) were monitored before and throughout the first year after transplantation. After transplantation and in unstimulated samples, p-p38MAPK and p-Akt were inhibited in CD8+ T cells and p-ERK in CD4+ T cells but only in patients who received TAC-based therapy. After activation with PMA/ionomycin, p-p38MAPK and p-AKT were significantly inhibited in CD4+ and CD8+ T cells when TAC was given, compared to pre-transplantation. Eleven BELA-treated patients had a biopsy-proven acute rejection, which was associated with higher p-ERK levels in both CD4+ and CD8+ T cells compared to patients without rejection. In conclusion, phospho-specific flow cytometry is a promising tool to pharmacodynamically monitor TAC-based therapy. In contrast to TAC-based therapy, BELA-based immunosuppression does not inhibit key T cell activation pathways which may contribute to the high rejection incidence among BELA-treated transplant recipients.
Highlights
T cells are the main target of most immunosuppressive drugs used in transplantation
Phospho-specific flow cytometry of the intracellular signaling molecules p38MAPK, ERK and Akt is a promising technique to monitor the pharmacodynamic effects of immunosuppressive drugs in whole-blood of kidney transplant patients[27]
Belatacept (BELA), a fusion protein consisting of the Fc-fragment of human immunoglobulin G1 linked to the extracellular domain of human cytotoxic T-lymphocyte antigen (CTLA)-4, was approved in 2011 for the prevention of acute rejection after kidney transplantation[30,31,32]
Summary
T cells are the main target of most immunosuppressive drugs used in transplantation. T cells become activated upon three separate stimulation signals: 1) antigen recognition by the T cell receptor (TCR) with the help of antigen presenting cells (APC); 2) co-stimulation, of which the interaction between CD28 molecules on T cells and CD80/86 molecules on the APC is the best known pathway, and 3) binding of cytokines[8]. Phospho-specific flow cytometry of the intracellular signaling molecules p38MAPK, ERK and Akt is a promising technique to monitor the pharmacodynamic effects of immunosuppressive drugs in whole-blood of kidney transplant patients[27]. Mycophenolic acid (MPA) was been found to decrease the phosphorylation of p38MAPK and ERK1/2 in vitro[29] These previous studies lacked an appropriate control group that did not receive TAC and could not exclude an effect of other, concomitantly used immunosuppressive drugs on T cell activation in these kidney transplant patients. Belatacept (BELA), a fusion protein consisting of the Fc-fragment of human immunoglobulin G1 linked to the extracellular domain of human cytotoxic T-lymphocyte antigen (CTLA)-4, was approved in 2011 for the prevention of acute rejection after kidney transplantation[30,31,32] It blocks the co-stimulation signal between CD80/86 and CD28 molecules (on APCs and T cells, respectively) and prevents T cell activation. One of the explanations for this phenomenon is the fact that memory CD8+ T cells lack CD28 expression and are not dependent on the co-stimulatory signal from CD80/8635,38–40
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