Elsevier

Journal of Autoimmunity

Volume 88, March 2018, Pages 91-102
Journal of Autoimmunity

NEPHRUTIX: A randomized, double-blind, placebo vs Rituximab-controlled trial assessing T-cell subset changes in Minimal Change Nephrotic Syndrome

https://doi.org/10.1016/j.jaut.2017.10.006Get rights and content

Highlights

  • MCNS is associated with low IL-2 production and a downregulation of Tregulatory cells.

  • MCNS is an autoimmune disease, which can be stabilized by Rituximab treatment.

  • Rituximab downregulates the double negative invariant T-cell subset (TCR DN-TCRα24).

Abstract

Minimal-change nephrotic syndrome (MCNS) is an immune-mediated glomerular disease. We have analyzed the modifications on T-cell subsets in twenty-three patients who were highly steroid/calcineurin inhibitor and/or mycophenolate mofetil-dependent for frequently relapsing nephrotic syndrome (FRNS) and who were enrolled in a multicenter, double-blind, randomized, placebo vs Rituximab-controlled trial. Patients with FRNS entered the trial at remission and were randomly assigned to receive either Rituximab or placebo. In both groups, patient blood samples were analyzed at inclusion and then monthly until six months post-perfusion. Disclosure of patient's allocation code occurred in relapse or at the end of the trial. All patients under placebo displaying relapse were subsequently treated with Rituximab.

Despite the significant decrease of immunosuppressive drugs, remission was maintained in all patients included in the Rituximab group, except one (n = 9/10). On the other hand, relapses occurred within a few weeks (means ≈ 7.3 weeks) in all patients receiving placebo (n = 13).

At inclusion, before rituximab therapy, the frequency of different T-cell subsets were highly similar in both groups, except for CD8+ and invariant TCRVα24 T-cell subsets, which were significantly increased in patients of the Placebo group ((p = 0,0414 and p = 0.0428, respectively). Despite the significant decrease of immunosuppressive drugs, remission was maintained in all patients included in the Rituximab group (n = 10), except one. Relapses were associated with a significant decrease in CD4+CD25highFoxP3high Tregulatory cells (p = 0.0005) and IL2 expression (p = 0.0032), while CMIP abundance was significantly increased (p = 0.03). Remissions after Rituximab therapy were associated in both groups with significant decrease in the frequency of CD4+CD45RO+CXCR5+, invariant natural killer T-cells (INKT) and CD4CD8 (double-negative, DN) T-cells expressing the invariant Vα24 chain (DN-TCR Vα24) T-cells, suggesting that MCNS involves a disorder of innate and adaptive immune response, which can be stabilized by Rituximab treatment.

Introduction

Minimal change nephrotic syndrome (MCNS) is considered as a disease of immune origin, in which the hallmark is the sudden onset of massive proteinuria. MCNS is most frequent in children and young adults. The nature of immune disorders involved in MCNS pathogenesis remains incompletely elucidated [1]. Several decades ago, it was suggested that MCNS is a systemic disorder of T-cell function [2]. This hypothesis was supported by several clinical observations such as the rapid occurrence of relapses upon antigen challenge (infections or immunization). Allergic manifestations such as contact dermatitis, rhinitis, and asthma might be observed, particularly in MCNS children. Several T-cell populations have been found perturbed during the active phase of the disease [3], [4]. Studies on genetic polymorphisms in the variable region of the T-cell receptor (TCR) β-chain have revealed a selective recruitment of some variable β gene families in peripheral CD8+ T cells from nephrotic patients with frequent relapses [5]. Perturbations of immune system have been described during active MCNS including hyporesponsiveness of lymphocytes to mitogens, a decrease of delayed hypersensitivity [6], an increased suppressive cell activity, defective opsonization and abnormal immunoglobulin production [7]. Reactive oxygen species (ROS) production by polymorphonuclear cells (PMN) was found increased in MCNS relapse and restored to normal levels in remission [8].

Recently, the finding that Rituximab, a B-cell-depleting agent, maintains long-lasting MCNS remission points out a role for B lymphocyte in the mechanisms of the disease and adds complexity about the origin of immune disturbances [9], [10]. Remission following B-cell depletion suggests that glomerular disease could be induced by some B lymphocyte subsets such as autoreactive B-cells but some arguments advocate a more intricate mechanism: (i) immunofluorescence studies on renal biopsies consistently show the absence of Ig deposits in MCNS relapse and (ii) remission can be maintained despite complete recovery of peripheral B-cell compartment, while relapses can occur in presence of sustained depletion of B cells [11], [12]. Beside their role in antibody-mediated mechanisms, B cells are also involved in antigen presentation, T-cell activation/regulation and production of cytokines and growth factors. Thus, B cells may facilitate disease activity by sustaining pathogenic T-cell responses through antibody-independent mechanisms. Rituximab is effective in autoimmune diseases primarily mediated by T-cells, such as type1 diabetes [13], immune thrombocytopenia [14] and autoimmune hepatitis [15]. Therefore, although Rituximab may be considered as an innovative therapeutic agent in frequent relapsing, steroid-dependent MCNS, the mechanisms by which it interferes with T-cell disorders remain unclear. Different immune cell types recirculate between the peripheral blood and secondary lymphatic organs such as lymph nodes. They include CD4+/CD8+ T cells, natural killer T (NKT) cells, B cells, T follicular helper cells and T regulatory cells (Tregs), which interact together in a complex network of innate-adaptive immunity able to provide an adapted immune response. Because these interactions are thought to be disrupted by B-cell depletion, we have addressed the question of which T-cell subsets are particularly affected by Rituximab therapy in patients with frequent relapses and highly dependent on steroids, mycophenolate mofetil (MMF) and calcineurin inhibitors (CAI).

Section snippets

Patients

Patients were enrolled in a multicenter, double-blind, randomized, placebo-controlled trial (NEPHRUTIX trial, NCT01268033). Two patients were excluded because of wrongly inclusion (they were steroid-resistant) and one patient lost to follow up and excluded by decision of the investigator. Clinical characteristics of patients and diverse treatment that were administrated before and after inclusion were summarized in Table 1A, Table 1B. The strategy of randomization is outlined in Fig. 1.

Characteristics of patients and clinical course following rituximab therapy vs placebo

Twenty-three patients who were highly steroid/calcineurin inhibitor and/or mycophenolate mofetil-dependent and displaying frequently relapsing nephrotic syndrome (FRNS), were enrolled in this multicenter, double-blind, randomized, placebo-controlled trial (NEPHRITUX trial, NCT01268033). Renal biopsy was performed in all patients except one patient in each group. MCNS diagnosis was confirmed in all, except one patient who exhibited focal and segmental glomerulosclerosis (FSGS) lesion. One

Discussion

Rituximab has become an essential alternative for MCNS patients who exhibit frequent relapses requiring sustained steroid and/or immunosuppressive drugs to achieve remission [9], [10]. In many instances, Rituximab allows tapering or withdrawal of therapy, while it maintains long-lasting remissions. The immune mechanisms underlying the effect of B-cell depletion in MCNS are unclear, especially as the disease is not associated with antibody production and some arguments suggest as T-cell mediated

Competing interests

The authors declare that they do not have any competing financial, personal, or professional interests.

Conflict of interest

The authors have declared that no conflict of interest exists.

Acknowledgments

This work was supported by a grant from the French Ministry of Health (PHRC, project number 16-03/2009). P Vachin was supported by grants from the Ministère de la Recherche.

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    BA and PV contributed equally to this work.

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