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One-year Results of the Effects of Rituximab on Acute Antibody-Mediated Rejection in Renal Transplantation: RITUX ERAH, a Multicenter Double-blind Randomized Placebo-controlled Trial

Sautenet, Bénédicte MD; Blancho, Gilles MD, PhD; Büchler, Mathias MD, PhD; Morelon, Emmanuel MD, PhD; Toupance, Olivier MD; Barrou, Benoit MD, PhD; Ducloux, Didier MD, PhD; Chatelet, Valérie MD; Moulin, Bruno MD, PhD; Freguin, Caroline MD; Hazzan, Marc MD, PhD; Lang, Philippe MD, PhD; Legendre, Christophe MD, PhD; Merville, Pierre MD, PhD; Mourad, Georges MD, PhD; Mousson, Christine MD, PhD; Pouteil-Noble, Claire MD, PhD; Purgus, Raj MD; Rerolle, Jean-Philippe MD; Sayegh, Johnny MD; Westeel, Pierre-François MD; Zaoui, Philippe MD, PhD; Boivin, Hedia PharmD; Le Gouge, Amélie MSc; Lebranchu, Yvon MD, PhD

doi: 10.1097/TP.0000000000000958
Original Clinical Science—General
Free
SDC

Background Treatment of acute antibody-mediated rejection (AMR) is based on a combination of plasma exchange (PE), IVIg, corticosteroids (CS), and rituximab, but the place of rituximab is not clearly specified in the absence of randomized trials.

Methods In this phase III, multicenter, double-blind, placebo-controlled trial, we randomly assigned patients with biopsy-proven AMR to receive rituximab (375 mg/m2) or placebo at day 5. All patients received PE, IVIg, and CS. The primary endpoint was a composite of graft loss or no improvement in renal function at day 12.

Results Among the 38 patients included, at 1 year, no deaths occurred, but 1 graft loss occurred in each group. The primary endpoint frequency was 52.6% (10/19) and 57.9% (11/19) in the rituximab and placebo groups, respectively (P = 0.744). Renal function improved in both groups, as soon as day 12 with no difference in serum creatinine level and proteinuria at 1, 3, 6, and 12 months. Supplementary administration of rituximab and total number of IVIg and PE treatments did not differ between the 2 groups. Both groups showed improved histological features of AMR and Banff scores at 1 and 6 months, with no significant difference between groups but with a trend in favor of the rituximab group. Both groups showed decreased mean fluorescence intensity of donor-specific antibodies as soon as day 12, with no significant difference between them but with a trend in favor of the rituximab group at 12 months.

Conclusions After 1 year of follow-up, we observed no additional effect of rituximab in patients receiving PE, IVIg, and CS for AMR. Nevertheless, our study was underpowered and important differences between groups may have been missed. Complementary trials with long-term follow-up are needed.

Supplemental digital content is available in the text.

1 Service de Néphrologie-Immunologie Clinique, CHU Bretonneau, Tours, France.

2 Université François-Rabelais, Tours, France.

3 Institut de Transplantation, Urologie-Néphrologie (ITUN), CHU, Nantes, France.

4 Université François-Rabelais, Tours, France.

5 Service de Néphrologie, CHU Edouard Herriot, Lyon, France.

6 Service de Néphrologie, CHU Maison Blanche, Reims, France.

7 Service d'Urologie, CHU Pitié Salpêtrière, Paris, France.

8 Service de Néphrologie, CHU Jean Minjoz, Besançon, France.

9 Service de Néphrologie, CHU, Caen, France.

10 Service de Néphrologie, CHU Civil, Strasbourg, France.

11 Service de Néphrologie, CHU, Rouen, France.

12 Service de Néphrologie, CHU, Lille, France.

13 Service de Néphrologie, CHU Henri Mondor, Paris, France.

14 Service de Néphrologie, CHU Necker, Paris, France.

15 Service de Néphrologie-Transplantation-Dialyse, CHU Pellegrin, Bordeaux, France.

16 Service de Néphrologie, CHU, Montpellier, France.

17 Service de Néphrologie, CHU, Dijon, France.

18 Service de Néphrologie, CHU, Lyon, France.

19 Service de Néphrologie, CHU, Marseille, France.

20 Service de Néphrologie, CHU, Limoges, France.

21 Service de Néphrologie-Dialyse-Transplantation, CHU, Angers, France.

22 Service de Néphrologie, CHU, Amiens, France.

23 Service de Néphrologie, CHU, Grenoble, France.

24 Service de Pharmacologie Clinique, CHU Bretonneau, Tours, France.

25 INSERM, CIC 1415, CHU Bretonneau, Tours, France.

Received 27 April 2015. Revision received 8 July 2015.

Accepted 1 August 2015.

Supported by grants from the French Ministry of Health (PHRN07-YL RITUX-ERAH) and grants from the Roche laboratory.

The authors declare no conflicts of interest.

B.S. participated in performance of the research, data analysis and writing of the article. G.B. participated in performance of the research. M.B. participated in the research design, performance of the research, data analysis, and writing of the article. E.M. participated in performance of the research. O.T. participated in the performance of the research. B.B. participated in performance of the research. D.D. participated in performance of the research. V.C. participated in performance of the research. B.M. participated in performance of the research. C.F. participated in performance of the research. M.H. participated in performance of the research. P.L. participated in performance of the research. C.L. participated in performance of the research. P.M. participated in performance of the research. G.M. participated in performance of the research. C.M. participated in performance of the research. C.P.-N. participated in performance of the research. R.P. participated in performance of the research. J.-P.R. participated in performance of the research. J.S. participated in performance of the research. P.-F.W. participated in performance of the research. P.Z. participated in performance of the research. H.B. participated in data analysis. A.L.G. participated in data analysis. Y.L. participated in the research design, performance of the research, data analysis and writing of the article.

Correspondence: Bénédicte Sautenet, MD, MSc, Service de Néphrologie-Immunologie clinique, Hôpital Bretonneau, CHU Tours, Tours, 37044 cedex. ( benedicte.sautenet@univ-tours.fr).

Supplemental digital content (SDC) is available for this article. Direct URL citations appear in the printed text, and links to the digital files are provided in the HTML text of this article on the journal’s Web site (www.transplantjournal.com).

ClinicalTrials.gov Identifier: NCT1066689.

Acute antibody-mediated rejection (AMR) is characterized by acute allograft dysfunction, morphologic evidence of acute tissue injury, C4d deposition in peritubular capillaries, and the presence of donor-specific antibodies (DSAs).1-4 Because acute AMR is an important predictor of chronic allograft nephropathy and graft loss, prevention and treatment are key targets to improve long-term outcome after renal transplantation.5-8

International guidelines suggest the use of plasma exchange (PE), IVIg, rituximab, or lymphocyte-depleting antibodies for acute AMR.9 However, the quality of evidence for these treatments is low in the absence of randomized controlled trials (RCTs) with adequate statistical power to compare their safety and efficacy.10,11 In 2012, a systematic review of treatment of acute AMR found only 5 RCTs of the efficacy of PE or immunoadsorption (4 trials performed before 1990) and 7 other controlled studies.12 A meta-analysis was not possible because all RCTs had small sample sizes and differed in inclusion criteria and treatment regimens. The evidence supporting the treatment was “low” for PE and immunoadsorption and “very low” for all other interventions. Therefore, the classical approach to treat acute AMR is antibody removal and IVIg, generally associated with a treatment targeting the B-cell cascade with rituximab or bortezomib.13

Rituximab is a chimeric antibody recognizing the cell-surface marker CD20, which is expressed at most stages of B-cell development except the very early stages but not on plasma cells.14 Only 1 RCT studied the effect of rituximab, not exactly in AMR but in acute transplant rejection with B-cell infiltrates in 20 pediatric patients.15 All other reports have been single case reports or case series.16-29 The findings suggest that rituximab may have some beneficial effects in the treatment of acute AMR along with standard treatment of the condition.

With this multicentric RCT, we aimed to compare the efficacy and safety of rituximab and placebo combined with conventional therapy for treating acute AMR.

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MATERIALS AND METHODS

Study Design And Setting

The RITUX ERAH study was a prospective, multicenter (21 transplant centers in France), randomized, double-blind, placebo-controlled, phase III, parallel group trial of renal transplant recipients. The study protocol and any amendments were reviewed and approved by the Institutional Review Board/Independent Ethics Committee for each site before the initiation of the study in accordance with the current Declaration of Helsinki (EudraCT 2007-003213-13). All participants gave their informed consent to be included. A data and safety monitoring board regularly assessed cumulative safety and efficacy data throughout the trial.

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Participants

Eligible patients were adults 18 years or older who received renal transplants from a living or deceased donor and had acute AMR during the first year after transplantation. Patients were included between October 7, 2008, and October 7, 2011. Acute AMR was defined as deteriorated renal function assessed by an increase in serum creatinine level greater than 20% compared to the lowest value in the first 28 days after transplantation, or no significant decrease in serum creatinine level, and at least 2 of the following: (1) tissue damage according to Banff scores1-3 (acute tubular necrosis, presence of monocytes, or granulocytes in the peritubular capillary or glomeruli and/or capillary thrombosis, intimal arteritis, fibrinoid necrosis); (2) C4d labeling of peritubular capillary and/or presence of Ig or complement in lesions of fibrinoid necrosis; and (3) presence of donor-specific anti-HLA antibodies (DSAs) expressed as mean fluorescence intensity (MFI), with an MFI of 1500 or greater. We excluded patients who were pregnant or had multiple organ transplants, active infection (HIV, hepatitis C and B virus, tuberculosis), uncontrolled cardiac disease or rituximab injection within 3 months before inclusion.

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Randomization, Allocation Concealment, And Follow-Up

Eligible consenting patients were randomly allocated in a 1:1 ratio to receive rituximab or placebo. Randomization and allocation concealment were achieved by use of a centralized, computer-generated, interactive, Web-response system managed by the Roche laboratory, which had no role in recruitment. Randomization was stratified by center, with permutation blocks of variable sizes. Day 1 was the day of randomization. Included patients were observed until 12 months.

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Interventions

All patients received usual care with PE (plasma removal of 60 mL/kg and 5% albumin infusion), with at least 3 PE treatments between days 1 and 5, 48 hours without PE after rituximab or placebo infusion at day 5, then 3 PE until day 12; IVIg (100 mg/kg per day after each PE until day 4, then 1 g/kg per day on days 5 and 6); corticosteroids (CS) (infusion of 500 mg/day methylprednisolone during the first 3 days, then oral CS, 1 mg/kg per day); and daily immunosuppression with tacrolimus (trough level, 8-12 ng/mL) and mycophenolate mofetil (2 g/day). At day 5, patients received intravenous infusions of rituximab (375 mg/m2) or placebo. In case of insufficient efficacy of treatment for acute AMR, according to their own opinion, investigators could propose additional infusions of rituximab from day 12, with a limit of 2 infusions of rituximab and additional PE and IVIg infusion. To prevent patients in the rituximab group from receiving a third infusion of rituximab, the study was unblinded before administration of the third infusion.

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Study Outcomes

The primary outcome was a composite of treatment failure represented by graft loss or absence of improvement of renal function at day 12, defined by a decrease in serum creatinine level of less than 30% as compared with the peak serum creatinine level recorded at the time of AMR. Secondary outcomes were 1-year posttreatment patient and graft survival, renal function estimated by the Modification of Diet in Renal Disease (MDRD) equation,30 proteinuria, additional administration of rituximab, total number of intravenous IVIg and PE treatments, safety (total severe adverse events), detection of DSAs by Luminex SA (Luminex LABScreen Single Antigen, One-Lambda Inc., CA) for single-antigen class I and II and recorded as MFI, and renal histology at months 1 and 6 (Banff scores: glomerulitis, tubulitis, transplant glomerulopathy, chronic vascular lesions, interstitial inflammation, interstitial fibrosis, tubular atrophy, microcirculation inflammation, peritubular capillaritis, vasculitis).3,4

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Sample Size

Our trial was powered for superiority on the primary outcome at day 12. The expected rate of failure was 15% in the intervention group and 50% in the control group. A total of 64 patients had to be enrolled for 80% power with 2-sided α = 0.05.

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Statistical Analysis

We used both modified intent-to-treat (ITT) and per-protocol analyses. The modified ITT population comprised all randomized patients except those who did not receive the treatment. The per-protocol population comprised patients who received at least 1 rituximab infusion versus patients who received only placebo infusion. The rate of failure at day 12 was evaluated by χ2 test. For secondary outcomes, the number of supplementary administrations of rituximab, IVIg, and PE per patient was evaluated by negative binomial regression. A linear mixed model was used to assess changes in serum creatinine level, proteinuria and anti-HLA DSAs. This model was selected because it can impute missing values and handle the highly correlated nature of repeated measurements within and between individuals.31 Renal histology expressed in Banff scores was evaluated by Mann-Whitney U test. Patient and graft survivals were described. Statistical analyses involved SAS v9.2 (SAS Inst., Cary, NC) and R v2.12.1 (http://www.r-project.org).

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RESULTS

Of the 40 patients randomized, 38 were included in the modified ITT analysis and the per-protocol analysis (Figure 1): 1 patient received rituximab instead of placebo in the placebo group, and 1 patient received placebo instead of rituximab in the rituximab group. In the placebo group, 7 patients received 1 rescue injection of rituximab, and 1 patient received 2 rescue injections. In the rituximab group, 6 patients received 1 rescue injection. In the per-protocol analysis, the placebo group consisted of 11 patients who did not receive any rituximab during the study, and the rituximab group consisted of 27 patients, 7 receiving 2 injections, and 20 a single injection.

FIGURE 1

FIGURE 1

Baseline characteristics were similar between the 2 groups (Tables 1A and 1B). The AMR occurred within the first 3 months after transplantation for 57.9% (11/19) and 52.6% (10/19) in the rituximab and placebo groups, respectively. The DSAs with MFI ≥ 1 500 were identified in 17 of 19 and 15 of 19 patients in the rituximab and placebo groups, respectively, at the time of rejection, with a mean MFI of the immunodominant DSAs (iDSAs) of 7199 ± 4616 and 5538 ± 4365, respectively (P = 0.442). The iDSAs were anticlass I antibodies in 7 of 17 and 3 of 15 and anticlass II antibodies in 10 of 17 and 12 of 15 of the rituximab and placebo groups, respectively. In the rituximab group, the iDSAs were anti-A for 2, anti-B for 4, anti-C for 1, anti-DQ for 5, and anti-DR for 5. In the placebo group, the iDSAs were anti-A for none, anti-B for 2, anti-C for 1, anti-DQ for 6, and anti-DR for 6. The DSAs were retrospectively identified at the time of transplantation in 63.2% (12/19) and 47.4% (9/19) of the rituximab and placebo groups, respectively, all with a negative T-cell cytotoxic crossmatch. All patients underwent a graft biopsy that disclosed patterns of acute AMR; deposits of C4d were identified in 89.4% (17/19) and 78.9% (15/19) of the rituximab and placebo groups, respectively. Borderline injury or cell-mediated rejection patterns were present in 4 and 3 and 1 and 6 patients in the rituximab and placebo groups, respectively.

TABLE 1A

TABLE 1A

TABLE 1B

TABLE 1B

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Primary Outcome

The primary outcome was a composite outcome of graft loss or absence of improvement of renal function at day 12. Because the 2 groups showed no graft loss at day 12, the primary outcome was defined by the absence of significant improvement in renal function at day 12. The primary outcome was observed in 52.6% (10/19) and 57.9% (11/19) of the rituximab and placebo groups, respectively (P = 0.744) (Figure 2A). The percentage decrease in serum creatinine level at day 12 was similar in the 2 groups (median [Q1; Q3] 20.5 [0.0; 47.1] vs 19.7 [8.6; 35.3], P = 1.00) (Figure 2B).

FIGURE 2

FIGURE 2

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Secondary Outcomes

Patient And Graft Survival

No death occurred during the study period. One graft loss was observed in the placebo group at 6 months and 1 in the rituximab group at 12 months postacute AMR.

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Supplementary Treatment

The rituximab and placebo groups did not differ in the number of patients who received supplementary rituximab (31.6% [6/19] vs 42.1% [8/19], P = 0.501) or total number of IVIg infusions (median [Q1; Q3] 9.0 [7.0; 13.0] vs 7.0 [6.0; 16.0], P = 0.761) or PE treatments (median [Q1; Q3], 7.0 [6.0; 9.0] vs 7.0 [6.0; 10.0], P = 0.924).

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Renal Function

Both groups showed improved renal function, with significant decrease in median serum creatinine level during the study (−5.27; 95% confidence interval [95% CI] −10.30 to −0.24] μmol/L; P = 0.040), with no significant between-group differences (−25.24 μmol/L; 95% CI, −95.53 to 45.05; P = 0.480) (Figure 3). The decrease in serum creatinine was rapid and significant between day 0 and day 12 (82.79 μmol/L; 95% CI, 39.86-125.73; P = 0.0004) with no significant between-group differences. The median (Q1; Q3) serum creatinine level at 12 months for the rituximab and placebo groups was 157 (107; 224) and 158 (129; 217) μmol/L, respectively. The median estimated glomerular filtration rate (eGFR), calculated by the MDRD equation, was increased but not significantly during the study (0.06; 95% CI, −0.002 to 0.118] mL/min per 1.73 m2; P = 0.058), with no significant between-group differences (0.30; 95% CI, −0.56 to 1.16 mL/min per 1.73 m2; P = 0.495). At 12 months, the eGFR was greater than 30 mL/min per 1.73 m2 for 57.9% (11/19) versus 66.7% (12/18) for the rituximab and placebo groups, respectively (P = 0.833); the eGFR was greater than 60 mL/min per 1.73 m2 for only 10.5% (2/19) and 5.6% (1/18) of patients, respectively (P = 1.00). When comparing renal function at 12 months, the eGFR was greater than 30 mL/min per 1.73 m2 in 14 of 19 (73.7%) patients in whom acute AMR occurred within the first 3 months versus 6 of 16 (37.5%) in whom acute AMR occurred later (P = 0.031); the difference was more striking when we used a 45-mL/min per 1.73 m2 eGFR cutoff value: 7 of 19 (36.8%) versus 0 of 16 (0.0%) (P = 0.022). The rituximab and placebo groups did not differ in these 2 subgroups. The results were similar in per-protocol analyses (SDC, http://links.lww.com/TP/B205).

FIGURE 3

FIGURE 3

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Proteinuria

We found no significant decrease in median proteinuria during the study (−0.05; 95% CI, -0.12 to 0.02; P = 0.172), with no significant between-group differences (−0.73; 95% CI, −1.57 to 0.12; P = 0.090) (Figure 4). At 12 months, the rate of proteinuria less than 0.3 g/d for the rituximab and placebo groups was 66.7% (12/18) and 63.2% (12/19), respectively; the rate 0.3 to 1.0 g/d was 5.5% (1/18) and 26.3% (5/19); and the rate > 1.0 g/d was 27.8% (5/18) and 10.5% (2/19), with no significant difference between groups (P = 0.174). The results were similar in per-protocol analyses (SDC, http://links.lww.com/TP/B205).

FIGURE 4

FIGURE 4

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Histological Changes

Histological features of AMR improved at 1 month after treatment in both groups. Nevertheless, the rate of histological lesions of AMR persisted at both 1 month (47.1% [8/17] vs 50.0% [7/14], P = 0.871) and 6 months (31.3% [5/16] vs 41.7% [5/12], P = 0.569) in the rituximab and placebo groups, respectively.

The changes in Banff scores for acute and chronic histological features at 0, 1, and 6 months were similar between the rituximab and placebo groups, although with a significant decrease in microvascular inflammatory score (glomerulitis and peritubular capillaritis) at 6 months in the rituximab group and a significant increase in chronic injury (interstitial fibrosis and tubular atrophy) at 6 months in the placebo group (Table 2, Figures 5 and 6). C4d deposits in peritubular capillaries decreased in both groups after treatment, with no differences between groups. At 6 months, only 3 of 16 and 1 of 12 patients had histological features of transplant glomerulopathy in the rituximab and placebo groups, respectively. We did not find any difference in rate of chronic lesions between patients with acute AMR associated with cell-mediated rejection and those with acute AMR alone. The results were similar in per-protocol analyses (SDC, http://links.lww.com/TP/B205).

TABLE 2

TABLE 2

FIGURE 5

FIGURE 5

FIGURE 6

FIGURE 6

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Dsas

We found a rapid and significant decrease in median MFI of iDSAs during the study (−2.14; 95% CI, −3.23 to −1.05; P < 0.001), with no significant between-group differences (1.23; 95% CI, −18.21 to 20.67; P = 0.901) (Figure 7). The decrease in MFI of iDSAs was rapid and significant between days 0 and 12 (3580; 95% CI, 1960 to 5200], P < 0.0001) with no between-group differences. Nevertheless, MFI less than 1500 for the iDSAs was more frequent but not significantly in the rituximab than placebo group (9/13 [69.2%] vs 5/13 [38.5%], P = 0.11) at the last serum analysis of DSAs. The results were similar in per-protocol analyses (SDC, http://links.lww.com/TP/B205).

FIGURE 7

FIGURE 7

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Change In Blood Cd19 B-Cell Count

For patients who received at least 1 rituximab injection, the blood CD19+ B cell count was low within 1 month (<5/mm3: 19/22), with a median of 1.0 (range, 0-62). At the end of follow-up (≥6 months), 60.0% (12/20) of patients still had CD19+ B-cell count less than 10/mm3.

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Safety

In the per-protocol analysis, during the study, 37 serious adverse events were reported, 14 for patients receiving only placebo (corresponding to 7 patients), and 23 for patients receiving rituximab (corresponding to 16 patients) (Table 3). Infections were the most frequent serious adverse events, with more urinary tract infections in the placebo group and more opportunistic infections (cytomegalovirus infection, BK virus infection, nocardia) in the rituximab group. One suspected but unexpected serious adverse reaction, superficial spreading melanoma, was reported during the study in 1 patient who received rituximab.

TABLE 3

TABLE 3

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DISCUSSION

In the RITUX-ERAH trial, a phase III, multicenter, double-blind, placebo-controlled RCT, the addition of rituximab to standard treatment for acute AMR in renal transplantation did not improve short-term outcomes for graft function as compared with placebo. At 1 year, we found no significant differences in renal function or iDSA MFI between the 2 groups, but the rituximab group showed a decrease, although not significant, in iDSA MFI.

The RITUX-ERAH trial is the largest RCT of AMR treatment in renal transplantation and the first to study its efficacy and safety. Only case series were previously reported, and a systematic review of acute AMR treatment published in October 2012 showed insufficient data to guide treatment.12,16-29,32 We showed that 1-year graft survival for AMR treated with IVIg, PE, and CS with or without rituximab was excellent, and we could not demonstrate a supplementary efficacy of rituximab by graft survival or our primary endpoint. An additional beneficial effect of PE and rituximab on graft survival was reported in a group of 12 patients receiving IVIg, PE, and rituximab (92%) as compared with a historical group of 12 patients receiving only IVIg (50%)29; however, the respective impact of PE and rituximab could not be assessed, especially because graft survival was observed in about 80% of patients receiving IVIg and PE treatment in the literature.33 A better, 2-year graft survival was reported for patients receiving rituximab and PE (80%) than that in a PE cohort (60%).28 However, this retrospective analysis compared 2 heterogeneous historical groups of patients with chronic and acute humoral rejection, with and without DSAs, and the rituximab group was more frequently supplemented with IVIg than the group receiving PE alone.

In contrast with the good graft survival at 1 year, both groups showed persistent histological lesions of AMR and microvascular inflammation, with increased chronic lesion score. The persistence of signs of AMR and microvascular inflammation associated with the development of chronic lesions in many patients could predict a poor long-term graft outcome, especially because we observed poor 1-year renal function in most patients, with an MDRD greater than 60 mL/min per 1.73 m2 in only 3 patients. However, the rituximab group showed a trend to less microvascular inflammation and fewer chronic lesions as compared with the placebo group.

Rituximab was developed to treat hematologic malignancies and was later used to treat antibody-dependant diseases, such as AMR. Rituximab induces a depletion of blood CD20+ B cells followed by reconstitution over subsequent months and reduces the number of these cells in spleen and lymph nodes.34 It has been used for treatment and/or prevention of AMR to abrogate or reduce the humoral response, in particular the production of de novo DSAs.35,36 Its efficacy, combined with PE and IVIg, has been demonstrated to prevent AMR35,37-40 perhaps because rituximab prevents an anamnestic response in patients with cryptic sensitization to HLA antigens.41 Nevertheless, rituximab does not have a direct effect on antibody-producing plasma cells, which could explain the lower results for established AMR treatment than that for AMR prevention. Interestingly, we observed a rapid and significant decrease in iDSAs as soon as day 12 with EP and IVIg but with no additional effect of rituximab. The AMR is a highly inflammatory process, and our results highlight the need to use more potent anti-inflammatory agents, such as eculizumab, in the acute phase42 to rapidly treat the microvascular injuries. In some patients, we observed patterns of T-cell–mediated rejection. In these mixed cellular rejections, the use of a T-cell–depleting agent could be considered, as previously proposed, and needs to be assessed in future trials.43 Nevertheless, as in autoimmunity, rituximab often takes a number of months to work; we noted that at 12 months, the number of patients with iDSAs less than 1 500 was more frequent in the rituximab group than the placebo group. Decreased microvascular inflammation was observed in the rituximab group between rejection and 6 months, and increased chronic lesions was significant only in the placebo group. The demonstrated role of DSAs in the progressive process of microvascular injury and late kidney transplant failure44-46 highlights the need for a more prolonged follow-up for testing the efficacy of AMR treatments.

The use of rituximab is associated with several complications.47 In our study, the patients who received rituximab more frequently had cytomegalovirus infection, BK virus infection, and gastrointestinal disorders than did other patients. These complications have been previously described and must be taken into account.

Our study has several limitations. The number of patients included was lower than the planned sample size because during the period, rituximab was largely used to prevent AMR at the time of transplantation and prior use of rituximab was an exclusion criteria. This reduced number of patients could have underpowered our study, particularly in terms of histopathology score and iDSAs, MFI showing trends toward a better outcome in the rituximab group. As our study was underpowered, important differences between groups may have been missed. For most patients, we used a single injection of rituximab instead of the 4 injections used for treatment of hematological malignancies. Nevertheless, all patients showed profound B-cell depletion over several weeks. Because the design allowed for a rescue injection of rituximab, the number of patients who received only placebo during the study was decreased, but in the per-protocol analysis, all the results were similar with the ITT analysis, regardless of renal function, Banff scores, or DSAs. We found no significant differences in DSAs between the 2 treatment groups, but new assays detecting pathogenic antibody-fixing complement have been developed since, and we cannot exclude a potential effect of rituximab on these antibodies. Finally, the primary endpoint only captured the short-term effect of the treatment, and a 1-year survey is perhaps too short. A longer follow-up of this cohort may be more informative, in particular because we observed fewer chronic lesions at 6 months in the rituximab than placebo group.

Despite these limitations, this trial remains the largest RCT of treatment of AMR. At 1 year follow-up, we observed no additional effect of rituximab. Nevertheless, our study was underpowered and complementary trials with long-term follow-up are needed.

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ACKNOWLEDGMENTS

The authors are very grateful to all the anatomic pathology laboratories (Machet Marie- Christine, François Arnaud, Rouvier Philippe, Pinel Nicole, Moreau Anne, Felix Sophie, Copin Marie-Christine, Bluod David, Comoz François, Desvaux Dominique, Deminiere Colette, Daniel Laurent, MacGregor Brigitte, Marcellin Luc, Raynaud Pierre, Funes de la Vega Mathilde, Dijoud Frederique, Paraf François, Saint-André Jean-Paul, Coure Anne, Noel Laure-Hélène, Nochy Dominique, Vuiblet Vincent, Birembaut Philippe, Cordonnier Carole) and HLA laboratories (Lever Laurent, Magdelaine Charlotte, Hau Françoise, Have Virgine, Zahr Noël, Masson Dominique, Devys Anne, Dupont Isabelle, Lablette Myriam, Toutirais Olivier, Suberbielle Caroline, Guidicelli Gwendaline, Basire Agnes, Dubois Valérie, Parissiadis Anne, Ramounau-Pigot Annie, Dautin Guillaume, Durand, Dominique, Faye Gaelle, Coeffic Brigitte, Lechaton Sophie, Tabary Thierry, Guillaume Nicolas) included in this study. We thank the members of the Center of Clinical Investigation of Tours (Nathalie Juteau, Bruno Giraudeau, Adeline Fourmy, Carinne Coffre and Estelle Boivin). The authors thank the Roche laboratory and the French Ministry of Health (PHRN07-YL RITUX-ERAH). The authors thank Laura Smales for copyediting.

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