Kho, M.M.L.

Pharmacodynamic analysis of tofacitinib and basiliximab in kidney allograft recipients (Article)

2012-09-15T00:00:00Z

BACKGROUND: The common γ-chain (γc) cytokines signal through the Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway and play pivotal roles in lymphocyte activation. We investigated the effect of immunosuppressive drugs targeting this pathway, the JAK inhibitor tofacitinib (CP-690,550) and the anti-interleukin (IL)-2R antibody basiliximab, as part of a phase 2 study. METHODS: After whole-blood activation with the γccytokines IL-2, IL-7, and IL-15, STAT5 phosphorylation was determined in T cells of de novo kidney transplantation patients treated with tofacitinib/basiliximab (n=5), calcineurin inhibitor (CNI) (cyclosporine A)/basiliximab (n=4) or CNI (tacrolimus)-based immunosuppression (n=6). The IC50 for phosphorylated STAT (P-STAT) 5 inhibition by tofacitinib was determined in cytokine-activated CD4+and CD8+T cells from healthy individuals (n=4). RESULTS: IC50 was 26, 72, and 37 ng/mL for IL-2, IL-7, and IL-15 activation, in CD4+T cells, respectively; and 35, 61, and 76 ng/mL for IL-2, IL-7, and IL-15 activation, in CD8+T cells, respectively. In kidney transplantation patients, 7 days after starting tofacitinib/basiliximab treatment, cytokine-induced P-STAT5 was inhibited in CD4+T cells (92% for IL-2 activation, 60% for IL-7, and 75% for IL-15), which persisted for the 2-month study period. In contrast, CNI/basiliximab treatment did not affect IL-7-activated or IL-15-activated P-STAT5; only IL-2-activated P-STAT5 was reduced by 77% on day 7 and recovered to pretreatment levels within 2 months. CD8+T cells showed a comparable profile to CD4+T cells. P-STAT5 was not inhibited in CNI-treated control patients. CONCLUSIONS: Tofacitinib therapy strongly inhibits γccytokine-induced JAK/STAT5 activation, whereas basiliximab suppresses IL-2-stimulated activation only. Pharmacodynamic monitoring offers a unique tool to evaluate the biologic effects of immunosuppressive drugs. Copyright

A case of primary aldosteronism revealed after renal transplantation (Article)

2010-11-23T00:00:00Z

Background. A 57-year-old woman was referred to a nephrology clinic because of chronic hypokalemia. She had a history of polycystic kidney disease, resistant hypertension, atrial fibrillation, type 2 diabetes, stroke, and end-stage renal disease, and had received a kidney transplant from a deceased donor at the age of 48 years. At presentation, the patient described symptoms of chronic fatigue and muscle aches, but she did not report pareses. Her medications included four antihypertensive agents, glucose-lowering drugs, immunosuppressants, digoxin, a coumarin derivative, and potassium chloride.Investigations. Full history, physical examination, laboratory testing of blood and urine, including aldosterone-torenin ratio, and a saline infusion test.Diagnosis. Primary aldosteronism.Management. Treatment with spironolactone resulted in prompt control of hypertension and hypokalemia, allowing discontinuation of potassium chloride and reduction in antihypertensive medication.

The calcineurin inhibitor tacrolimus allows the induction of functional CD4+CD25+ regulatory T cells by rabbit anti-thymocyte globulins (Article)

2010-08-01T00:00:00Z

Rabbit anti-thymocyte globulins (rATG) induce CD4+CD25 +forkhead box P3 (FoxP3+) regulatory T cells that control alloreactivity. In the present study, we investigated whether rATG convert T cells into functional CD4+CD25+FoxP3+CD127 -/low regulatory T cells in the presence of drugs that may hamper their induction and function, i.e. calcineurin inhibitors. CD25neg T cells were stimulated with rATG or control rabbit immunoglobulin G (rIgG) in the absence and presence of tacrolimus for 24 h. Flow cytometry was performed for CD4, CD25, FoxP3 and CD127 and the function of CD25+ T cells was examined in suppression assays. MRNA expression profiles were composed to study the underlying mechanisms. After stimulation, the percentage CD4 +CD25+FoxP3+CD127-/low increased (from 2% to 30%, mean, P < 0·01) and was higher in the rATG samples than in control rIgG samples (2%, P < 0·01). Interestingly, FoxP3 +T cells were also induced when tacrolimus was present in the rATG cultures. Blockade of the interleukin (IL)-2 pathway did not affect the frequency of rATG-induced FoxP3+ T cells. The rATG tacrolimus-induced CD25+ T cells inhibited proliferative responses of alloantigen-stimulated effector T cells as vigorously as rATG-induced and natural CD4+CD25+FoxP3+CD127-/low T cells (67% ± 18% versus 69% ± 16% versus 45% ± 20%, mean ± standard error of the mean, respectively). At the mRNA-expression level, rATG-induced CD25+ T cells abundantly expressed IL-10, IL-27, interferon (IFN)-, perforin and granzyme B in contrast to natural CD25 + T cells (all P = 0·03), while FoxP3 was expressed at a lower level (P = 0·03). These mRNA data were confirmed in regulatory T cells from kidney transplant patients. Our findings demonstrate that tacrolimus does not negatively affect the induction, phenotype and function of CD4 +CD25+ T cells, suggesting that rATG may induce regulatory T cells in patients who receive tacrolimus maintenance therapy.

Characterization of rabbit antithymocyte globulins-induced CD25+ regulatory T cells from cells of patients with end-stage renal disease (Article)

2010-03-01T00:00:00Z

Background: Rabbit antithymocyte globulins (rATGs) are known to convert CD4CD25FoxP3 T cells from healthy individuals to CD4CD25FoxP3 T cells. In this study, we investigated the effect of rATG on the induction of regulatory T cells (Tregs) from blood cells of patients with end-stage renal disease who are candidates for transplantation and rATG-induction therapy. The induced Tregs were analyzed and compared with naturally occurring CD4CD25FoxP3T cells. Methods: The CD25 T cells of pretransplant patients (n=7) and healthy controls (n=4) were stimulated with rATG or control rabbit immunoglobulins for 24 hr. The phenotype of induced Tregs was examined by flow cytometry, and their function was studied in the conventional suppression assay. Further characterization was performed by mRNA analyses. Results: After 24 hr, the percentage of CD4CD25FoxP3CD127 T cells and CD8CD25FoxP3CD127 T cells became higher in the rATG-treated samples compared with the rabbit immunoglobulin-treated samples (P<0.01). The rATG-induced CD25T cells, whether CD4 or CD8 inhibited the allogeneic responses of CD25 effector T cells as vigorously as natural CD25T cells. However, the proportion of FoxP3 within the top 2% rATG-induced CD4CD25T-cells was lower than within the natural CD4CD25T-cells (11%±2% vs. 95%±5%, P<0.01). The mRNA-expression levels of interleukin-27, interleukin-10, interferon-γ, perforin, and granzyme B were markedly higher compared with natural CD25T-cells (all P=0.03), whereas CTLA4 (P=0.03), transforming growth factor-β (P=0.02), and RORγt (P=0.04) were lower. Conclusion: rATG allows the induction of Tregs from patient peripheral blood mononuclear cell in vitro. In comparison with natural Tregs, the rATG-induced Tregs are phenotypically distinct but have similar regulatory activities. rATG may beneficially contribute to the mechanisms that control alloreactivity.

The effect of rabbit anti-thymocyte globulin induction therapy on regulatory T cells in kidney transplant patients (Article)

2009-05-01T00:00:00Z

Background. Prevention of alloreactivity by rabbit anti-thymocyte globulins (rATG) may not only result from immunodepletion but also from the induction of T cells that control allogeneic immune responses. In the present prospective and controlled study, we investigated the effect of rATG on the frequency, function and phenotype of peripheral immunoregulatory CD4+ T cells in kidney transplant (KTx) patients.Methods. After transplantation, 16 patients received ATG-induction therapy and triple therapy consisting of tacrolimus, MMF and steroids. The control group (n = 18) received triple therapy only. By flow cytometry, T cells were analysed for CD25, FoxP3, CD127, CD45RO and CCR7. To study their suppressive capacities, CD25bright T cells were co-cultured with CD25-dim effector T cells (Teff) in mixed lymphocyte reactions (MLR), stimulated with donor and third party (3P) antigens.Results. Pre-transplant levels of FoxP3+CD127-low T cells were 6 of CD4+ T cells. One week post-ATG treatment, no measurable numbers of regulatory T cells were present (P < 0.01). After 4 weeks, the cell numbers of CD4+FoxP3+CD127-low T cells slowly reappeared and thereafter remained low (P < 0.01). At 14 weeks, a significant shift towards the CD45RO+CCR7+ (central memory) phenotype within CD4+FoxP3+ T cells was observed (P < 0.01). At 26 weeks, the proliferative alloresponses of the PBMC and CD25-dim Teff profoundly decreased compared to pre-transplant (P = 0.01 and P = 0.02 respectively), while the regulatory capacity of the CD25bright T cells, of which 90 consisted of FoxP3+CD127-low T cells, remained unaffected. The CD25bright T cells suppressed the anti-donor (94) and 3P responses (93).Conclusion. Our findings show that rATG therapy does not spare peripheral immunoregulatory T cells in vivo, but after regeneration preserves their suppressive activity.