Myriam Labalette

Predictive Factors of Acute Rejection after Early Cyclosporine Withdrawal in Renal Transplant Recipients Who Receive Mycophenolate Mofetil: Results from a Prospective, Randomized Trial

The aim of this randomized, open-labeled trial was to compare the incidence of acute rejection after an early (3 mo posttransplantation) withdrawal of cyclosporine (CsA) or mycophenolate mofetil (MMF) in renal transplantation. Among 218 eligible recipients, 108 nonsensitized, rejection-free patients who were under a triple drug regimen (CsA-MMF-prednisone) and had received a first kidney from a deceased donor were enrolled. At 3 mo after graft, they were gradually withdrawn from CsA (MMF group, n = 54) or MMF (CsA group, n = 54). A graft biopsy and a pharmacokinetic study of CsA and mycophenolic acid were systematically performed before the randomization. At 1 yr, graft and patient survival rates were 100% in each group. Renal function was improved in the MMF group compared with the CsA group (Cockcroft calculated clearance 64.7 +/- 18.7 versus 56.5 +/- 18.0 ml/min; P = 0.023). However, the probability of acute rejection was higher in the MMF group (18.5 versus 5.6%; P = 0.045). The 10 patients who developed acute rejection after CsA withdrawal had a significantly higher incidence of borderline changes on the randomization biopsy than the 44 rejection-free patients (five of 10 versus eight of 44; P = 0.034), and they displayed a lower area under the curve of mycophenolic acid (43 +/- 9 versus 58 +/- 22 mg/h per L; P = 0.045). Multivariate analysis confirmed that borderline changes and area under the curve of mycophenolic acid were significant risk factors of acute rejection after CsA discontinuation. It is concluded that a systematic graft biopsy and a pharmacokinetic study of mycophenolic acid are needed to reduce the risk for acute rejection after CsA withdrawal.

CYP3A5 and ABCB1 polymorphisms in donor and recipient: impact on Tacrolimus dose requirements and clinical outcome after renal transplantation

BACKGROUNDnThe effect of potentially relevant genetic polymorphisms, CYP3A5 6986A>G and ABCB1 3435C>T, on Tacrolimus pharmacokinetics and graft clinical outcome was investigated in donor and recipient DNA samples from 209 kidney transplant patients.nnnMETHODOLOGY/PRINCIPAL FINDINGSnThe mean follow-up was 21.8 ± 9 months. The Tacrolimus dose, trough blood concentrations (C0) and C0/dose ratio were only statistically correlated with the recipient CYP3A5 genotype. CYP3A5 and ABCB1 genotypes appeared to have no influence on the incidence of Biopsy Proven Acute Rejection and Delayed Graft Function. Renal function was not affected by CYP3A5 and ABCB1 genotypes. Histological evaluation of biopsies revealed also no significant association between Tacrolimus toxicity features and donor or recipient CYP3A5 and ABCB1 polymorphisms. Tacrolimus sparing appeared to be independent of CYP3A5 and ABCB1 genotypes.nnnCONCLUSIONS/SIGNIFICANCEnRecipient CYP3A5 6986A>G polymorphism explains part of the interindividual variability of the pharmacokinetics of Tacrolimus. The clinical outcome at 2-year follow-up does not appear to be related to the donor or recipient CYP3A5 6986A>G and/or ABCB1 3435C>T polymorphisms.

Recall response to cytomegalovirus in allograft recipients : Mobilization of CD57+, CD28+ cells before expansion of CD57+, CD28- cells within the CD8+ T lymphocyte compartment

BACKGROUNDnStrong correlations have been described between persistently elevated proportions of CD57+ (CD28-) CD8+high T lymphocytes and cytomegalovirus (CMV) infection, in healthy individuals as well as in transplant patients. We investigated whether secondary exposure to CMV triggers recall responses within the CD8 T cell compartment.nnnMETHODSnIn a longitudinal study in 123 kidney recipients, we compared 17 primary CMV infections with 27 secondary CMV infections. Subset composition of the CD8 compartment was analyzed by flow cytometry.nnnRESULTSnCD8 lymphocytosis occurred significantly earlier (by 17 days on average) in CMV reactivations than in primary infections. Both in primary and secondary infections, CD28+ CD8+high T lymphocytes were mainly recruited at the start. In formerly CMV-seropositive patients, preexisting CD57+ CD8+high T lymphocytes switched at the start from no expression of CD28 to high expression of CD28 and, concomitantly, from CD45RA to high expression of CD45RO. These cells reverted rapidly to a CD28- and CD45RA+ phenotype. Nevertheless, the accumulation of CD57+ (CD28-) CD8+high T cells was delayed similarly in primary and secondary CMV infection, progressing over a period between 2 and 8 weeks after the onset of CD8 lymphocytosis to plateau at 366 CD57+ CD8+high cells/ mm3 on average.nnnCONCLUSIONSnThe faster kinetics of CD8 lymphocytosis in secondary CMV infection suggests that a recall response triggers cycling memory cells within the CD28+ CD8+high subset, while preexistent CD57+ CD8+high T cells with a long-lived cell phenotype can also be mobilized, possibly through the transient acquisition of CD28 expression. The protracted accumulation of CD57+ (and CD28-) lymphocytes might then reflect an end-stage differentiation.

Using human CD20-transfected murine lymphomatous B cells to evaluate the efficacy of intravitreal and intracerebral rituximab injections in mice.

PURPOSEnThe treatment of primary central nervous system lymphoma (PCNSL) and its subset, primary intraocular lymphoma (PIOL), remains of limited efficiency, and salvage therapies are often used without prior testing in adequate animal models. Most PNCSL/PIOL are aggressive B-cell malignancies. Two animal models that closely mimic the human situation were established to evaluate the efficiency of intravitreal and intracerebral anti-CD20 monoclonal antibody (rituximab) injections.nnnMETHODSnHuman CD20-transfected murine B-lymphoma cells (38C13 CD20(+)) were inoculated in the vitreous through the pars plana or in the caudate nucleus with the use of a stereotaxic frame in immunocompetent syngeneic mice. Animals were monitored clinically and by funduscopic and histologic examination. Rituximab was injected intravitreally or intracerebrally. Occurrences of exophthalmia, neurologic disturbance, and weight loss were monitored over 2 months.nnnRESULTSnInoculation of 38C13 CD20(+) cells in the eye or the brain resulted in tumor occurrence after a median of 15 days or 22 days, respectively, with histologic characteristics closely resembling those of PIOL and PCNSL. Local rituximab injections eradicated tumor colonization in more than half the graft recipients and inhibited tumor progression significantly in the others compared with progression in mice that underwent grafting with the control 38C13 cell line (no human CD20 expression) and in mice that underwent grafting with 38C13 CD20(+) cells that received local injections of an irrelevant antibody (trastuzumab).nnnCONCLUSIONSnInoculation of native or human CD20-transfected murine 38C13 cells in the vitreous or the brain of immunocompetent mice provides useful novel models for evaluating the biology and treatment of PIOL and PCNSL. Intravitreal and intracerebral rituximab injections reduced tumor occurrence and growth in each model.

Plasma levels of IL-7 and IL-15 after reduced intensity conditioned allo-SCT and relationship to acute GVHD

To assess the impact of homeostatic expansion on the occurrence of acute GVHD after reduced intensity conditioning (RIC) transplantation, systemic levels of IL-7 and IL-15 and expression of their specific receptor chains were prospectively investigated in 45 fully HLA-matched allograft recipients. IL-7 and IL-15 levels peaked at four- to fivefold over pre-conditioning values. IL-7 levels were inversely correlated to absolute T-cell counts. Peak IL-15 levels positively correlated to concurrent CRP levels, but normalized earlier than IL-7. These results indicate that the kinetic course of IL-7 depends mainly on initiation of T-cell recovery, while IL-15 depends more on peri-transplant inflammation after RIC. Longer duration of the rise in IL-7 levels was associated with preservation of a normal CD4/CD8 ratio. In all, 16 (35%) patients developed grade 2–4 acute GVHD at a median of 42 days post graft, preceded by higher IL-7 levels and more downregulation of IL-7 receptor α chain on CD4+ T cells than in patients without acute GVHD, suggesting enhanced homeostatic expansion. In multivariate analysis, IL-7 level measured on day +30 was the foremost predictive factor for grade 2–4 acute GVHD (P=0.002). Measurement of IL-7 level after RIC transplantation might help predict risk of subsequent acute GvHD.

Influence of cytochrome P450 3A5 (CYP3A5) genetic polymorphism on the pharmacokinetics of the prolonged-release, once-daily formulation of tacrolimus in stable renal transplant recipients.

AbstractBackground and Objective: Tacrolimus is metabolized by cytochrome P450 (CYP) 3A5. The objective of this study was to investigate the influence of the genetic polymorphism of CYP3A5 on the pharmacokinetics of a new modified-release, once-daily formulation of tacrolimus (Advagraf®) after a switch from the immediate-release formulation (Prograf®)Patients and Methods: This was a prospective, single-centre, open-label study in stable kidney transplant recipients. Seventeen ‘expressor’ patients (CYP3A5*1/*3 or *1/*1) were matched to 15 ‘non-expressor’ patients (CYP3A5*3/*3). Exposure variables (concentrations and area under the blood concentration-time curve from 0 to 24 hours [AUC24]) were obtained before and 15 days after the switch. Delay since grafting was similar for both groups of patients (expressors: 49±24 months; non-expressors: 45±22 months).n Results: During administration of tacrolimus as Prograf® or Advagraf®, the mean tacrolimus daily dose was significantly higher and the dose-adjusted AUC24 was significantly lower in the expressor group. Following the switch to Advagraf®, there was a significant decrease in the dose-adjusted AUC24 for both non-expressor (5910 ± 3019 vs 5334 ± 2668 ng · h/mL per mg/kg/day; p = 0.041) and expressor patients (3701 ± 1409 vs 3273 ± 1372 ng · h/mL per mg/kg/day; p = 0.03). In the non-expressor group, mean blood trough concentration (C0) was comparable for both formulations while it decreased significantly in the expressor group after the switch (8.2 ± 2.2 vs 6.3 ± 2.5ng/mL; p = 0.02). However, a good correlation between AUC24 and C0 was observed for both Advagraf® and Prograf® regardless of CYP3A5 genotype.n Conclusion: Tacrolimus exposure significantly decreases after a switch from Prograf® to Advagraf®, on a milligram-for-milligram basis, in CYP3A5 expressor recipients. Consequently, these patients should be carefully monitored.

Characterization, quantification, and localization of passenger T lymphocytes and NK cells in human liver before transplantation

Quantification and localization of the main lymphocyte populations were studied in the livers of normal (n=8) and brain dead (n=8) subjects. Cytometric analysis performed on mononuclear cell suspensions obtained from liver biopsies was compared to an automatic image analysis of immunostained sections. The overall number of liver associated lymphocytes was in the usual range of peripheral blood content (2 to 9 · 109 cells). Phenotypic analysis showed predominant NK and CD8+ cells that highly expressed class II antigen and CD25 and CD69 activation markers. Quantitative mapping of these activated lymphocytes revealed their preferential localization in the portal tract and the perisinusoidal area as compared to the pericentrolobular zone, especially in donor livers. This strategic localization could suggest a possible early cooperation between donor lymphocytes and initial infiltrating cells from the recipient and could explain the special immunological status of allografted livers.

Assessment of the risk of chronic allograft dysfunction after renal transplantation in a randomized cyclosporine withdrawal trial.

Background. We report the two-year follow-up of a trial comparing the three-month postgraft discontinuation of either cyclosporine (CsA) or mycophenolate mofetil (MMF) from a triple-drug regimen after de novo renal transplantation. Methods. One hundred and eight patients were enrolled in this study and randomized to be withdrawn from CsA (MMF group, n=54) or MMF (CsA group, n=54). Results. Despite an increased risk of acute rejection and a lower, but nonsignificant, two-year graft survival, CsA withdrawal induced a sustained improvement of the renal function. At one year, the chronic allograft damage index was similar in both the MMF and CsA groups. However, CsA elimination resulted in a higher incidence of C4d deposits, irrespective of the occurrence of a prior acute rejection. While this finding could suggest a risk of chronic rejection in the MMF group, the outcome did not appear to be related to the C4d status. Moreover, logistic regression analysis showed that only two factors, acute rejection and the one-year glomerular filtration rate level, were predictive of a significant decline of the renal function at two years. Conclusions. These results point out the need to secure the minimization of the calcineurin inhibitors after renal transplantation, in order to reduce the risk of acute rejection in these patients, because this strategy allows the improvement of the one-year renal function which is predictive of a chronic allograft dysfunction.

The lymphoid variant of hypereosinophilic syndrome: study of 21 patients with CD3-CD4+ aberrant T-cell phenotype.

AbstractThe CD3-CD4+ aberrant T-cell phenotype is the most described in the lymphoid variant of hypereosinophilic syndrome (L-HES), a rare form of HES. Only a few cases have been reported, and data for these patients are scarce. To describe characteristics and outcome of CD3-CD4+ L-HES patients, we conducted a national multicentric retrospective study in the French Eosinophil Network. All patients who met the recent criteria of hypereosinophilia (HE) or HES and who had a persistent CD3-CD4+ T-cell subset on blood T-cell phenotyping were included. Clinical and laboratory data were retrospectively collected by chart review. CD3-CD4+ L-HES was diagnosed in 21 patients (13 females, median age 42 years [range, 5–75 yr]). Half (48%) had a history of atopic manifestations. Clinical manifestations were dermatologic (81%), superficial adenopathy (62%), rheumatologic (29%), gastrointestinal (24%), pulmonary (19%), neurologic (10%), and cardiovascular (5%). The median absolute CD3-CD4+ T-cell count was 0.35u2009G/L (range, 0.01–28.3), with a clonal TCR&ggr;&dgr; rearrangement in 76% of patients. The mean follow-up duration after HES diagnosis was 6.9 ± 5.1 years. All patients treated with oral corticosteroids (CS) (nu2009=u200918) obtained remission, but 16 required CS-sparing treatments. One patient had a T-cell lymphoma 8 years after diagnosis, and 3 deaths occurred during follow-up.In conclusion, clinical manifestations related to CD3-CD4+ T cell-associated L-HES are not limited to skin, and can involve all tissue or organs affected in other types of HE. Contrary to FIP1L1-PDGFRA chronic eosinophilic leukemia patients, CS are always effective in these patients, but CS-sparing treatments are frequently needed. The occurrence of T-cell lymphoma, although rare in our cohort, remains a major concern during follow-up.

Implication of cyclosporine in up-regulation of Bcl-2 expression and maintenance of CD8 lymphocytosis in cytomegalovirus-infected allograft recipients

T cell homeostasis and CD4/CD8 ratios are normally reestablished by apoptotic clearance of activated T cells after immune stimulation. In allograft recipients with cytomegalovirus infection, CD8 lymphocytosis persists after negativation of viral cultures, contrary to immunocompetent hosts. We investigated the expression of Bcl-2 protein, an intracellular suppressor of apoptosis, and of CD95 (APO-1/Fas), a membrane inducer of apoptosis, in peripheral blood lymphocytes from 45 solid organ recipients. During the viremic phase of CMV infection, we found absence or diminished expression of Bcl-2 protein and increased expression of CD95 antigen in activated CD8+ T cells. Opposite evolution of these molecular regulators of apoptosis was reflected by the presence of 10-25% of apoptotic lymphocytes with fragmented DNA, as shown by both in situ nick translation and electrophoresis. Normalization of Bcl-2 expression was progressive over several months but still lower than in uninfected allograft recipients. These results suggest that the initial evolution of CMV infection in allograft recipients resembles acute viral infection in immunocompetent hosts. Conversely, we showed that overexpression of Bcl-2 protein in lymphocytes from uninfected allograft recipients, and culture of unstimulated normal lymphocytes with 0.5 micrograms/ml cyclosporine led to an increase in the expression of intracellular Bcl-2. This up-regulation of Bcl-2 protein by cyclosporine suggests the acquisition of resistance to apoptosis. Thus, the reversion of balance between T cell death and survival after acute CMV infection might be impeded by cyclosporine. Combination of CMV latent infection and cyclosporine therapy appears therefore critical to shift the homeostatic maintenance of the peripheral lymphocyte compartment toward persistingly high numbers of CD8+ T cells.