Bortezomib in Late Antibody-mediated Kidney Transplant Rejection
Condition: Late Rejection of Renal Transplant; Antibody-mediated Rejection
Study type: Interventional
Phase: Phase 2

Summary: 
      Late antibody-mediated rejection (AMR) after kidney transplantation is defined as a separate
      rejection entity. So far, no appropriate treatment has been established for this rejection
      type. One promising strategy could be the targeting of alloantibody-producing plasma cells.
      There is now accumulating evidence that the proteasome inhibitor Bortezomib may substantially
      affect the function and integrity of non-malignant alloantibody-secreting plasma cells. The
      impact of this compound on the course of late AMR , however, has not yet been systematically
      investigated. In the planned phase IIa study we will examine the effect of Bortezomib on late
      AMR after kidney transplantation. We plan an initial cross-sectional HLA antibody screening
      of 1000 kidney transplant recipients to identify patients with detectable donor-specific
      antibodies (DSA). DSA-positive recipients will be subjected to kidney allograft biopsy to
      detect morphological features consistent with AMR. Forty-four patients with late AMR will be
      included in a randomized double-blind placebo-controlled parallel-group intervention trial.
      Patients in the active group will receive two cycles of Bortezomib (4 x 1.3 mg/m2). The
      primary end point will be the course of estimated GFR over 24 months after randomization.
      Secondary endpoints are the course of DSA levels and protein excretion, measured GFR after 24
      months, transplant and patient survival, and the development of acute and chronic
      morphological lesions in 24-month protocol biopsies. Our study will clarify the impact of an
      innovative anti-humoral strategy on the deleterious effects of late AMR processes.
    

Detailed description: 
      Background

      Recent studies have underscored a dominant role of alloimmune injury as a leading cause of
      long-term graft loss in kidney transplantation. In this respect, the formation of antibodies
      against specific polymorphic donor antigens, commonly human leukocyte antigens (HLA), has
      turned out to be an important trigger of graft rejection (Colvin, RB et al. 2007).

      Humoral rejection (antibody-mediated rejection; AMR) of organ transplants has been
      established to constitute a separate rejection entity and in recent years accurate
      biopsy-based (C4d) and serological criteria for this rejection type have been defined to
      provide a valuable basis for targeted treatment (Sis, B et al. 2010). It has become evident
      that a considerable proportion of recipients develop features of AMR late after
      transplantation, a process culminating in chronic irreversible tissue damage, graft
      dysfunction and loss (Mauiyyedi, S et al. 2001). Indeed, there are studies suggesting that
      newly formed donor-specific antibodies (DSA) represent the primary cause of late graft loss
      (Einecke, G et al. 2009).

      In contrast to early acute AMR, where various treatment protocols including immunoadsorption,
      intravenous immunoglobulin (IVIG) or CD20 antibody (Rituximab) exist, appropriate targeted
      treatment for late AMR still remains to be established (Vo, A et al. 2008).

      In an earlier uncontrolled study of 11 kidney transplant recipients with advanced
      C4d-positive chronic AMR we found that conversion of Cyclosporine A and Azathioprine-based
      basal immunosuppression to Tacrolimus and Mycophenolate-Mofetil (MMF) did not have any impact
      on DSA levels and graft performance (Schwarz, C et al. 2006). Our observations, which are in
      some contrast to a previous small study (Theruvath, TP et al. 2001), suggested that simple
      conversion strategies may not suffice to ameliorate ongoing late AMR.

      Rationale

      More recently, we reported on the treatment with Bortezomib, a proteasome inhibitor, in a
      patient diagnosed with advanced chronic active C4d-positive AMR (Schwaiger, E et al. 2011).
      In this patient, a single cycle of Bortezomib led to an impressive decrease in DSA levels and
      a complete disappearance of capillary C4d deposits in a follow-up biopsy. Also a remarkable
      observation was the stable decrease in proteinuria. As expected, advanced chronic lesions
      (severe transplant glomerulopathy) in this patient remained unchanged. Nevertheless, 16
      months after treatment, kidney function stabilized and the protein/creatinine ratio was still
      half of the initial value of 4,000 mg/g.

      Objective

      The primary objective of this investigator-driven, randomized prospective double-blind,
      placebo- controlled trial is to assess the efficiency of the innovative concept of proteasome
      inhibition with Bortezomib in the treatment of late AMR. Our primary hypothesis is that, by
      inhibiting the DSA production of plasma cells, Bortezomib can halt the progression of ongoing
      graft injury and dysfunction.

      Study Design

      The study (anticipated duration time is 36 months) will be performed in two major steps, Part
      A and B.

      Part A:

      This part consists of a cross-sectional screening analysis of a large cohort of approximately
      1000 kidney transplant recipients from our outpatient clinic for the presence of late AMR.

      Part A: Objective

      We will screen kidney transplant recipients for the presence of circulating DSA and biopsy
      features of late AMR. Key inclusion criteria are a functioning graft at > 6 months
      post-transplantation and an eGFR above 20 ml/min/1.73m2. The GFR threshold was chosen to
      avoid inclusion of transplants showing a high degree of irreversible chronic damage. For
      patients with very advanced graft injury a sustainable treatment benefit can no longer be
      expected.

      Part A: Sample size considerations

      Approximately 90% of 1000 recipients available for a pre-screening will be eligible for HLA
      antibody testing. In at least 10% (n≥90) of tested patients DSA detection can be expected
      (Lachmann, N et al. 2009). We estimate biopsy-based late AMR features in at least 60% of the
      DSA-positive patients (Hidalgo, LG et al. 2009). Hence, at least 50 patients will be eligible
      for inclusion into Part B, the randomized controlled trial.

      Part A: Methodology

      Estimation of GFR eGFR will be calculated using the Mayo equation. For kidney transplants,
      this equation was reported to be superior with respect to estimations of GFR slopes (Rule, AD
      et al. 2004).

      DSA screening:

      Serum obtained in the context of a routine outpatient laboratory control (10 ml blood; no
      additional venipuncture) will be pre-screened for anti-HLA immunoglobulin G (IgG)
      alloantibodies using LabScreenMixed assays (One Lambda, Canoga Park, CA, USA). For
      identification of HLA antigen/allele specificities, pre-screen-positive samples will be
      subjected to single-antigen flow-bead (SAFB) testing (LABscreen Single Antigen assays, One
      Lambda). SAFB results will be documented as mean fluorescence intensities (MFI), and MFI
      levels >500 will be considered to be positive. Donor-specificity will be defined according to
      donor and recipient HLA typing results. Virtual panel-reactive antibody (PRA) levels will be
      calculated using specific software tools (http://www.eurotransplant.eu).

      Renal biopsies:

      All biopsies will be performed under local anesthesia (lidocaine) using ultrasound-guided
      percutaneous techniques (two cores per biopsy). After biopsy, patients will be monitored
      closely for at least 6 hours for any complications.

      Biopsies will be evaluated on standard paraffin-embedded sections and by electron microscopy.

      (i) Histomorphological, (ii) electron-microscopical and (iii) immunohistochemical criteria
      will be applied to assess a score for late AMR.

      Part B:

      This is a phase IIa, proof of concept, randomized prospective, double-blind,
      placebo-controlled trial.

      It will examine whether treatment with Bortezomib is capable to halt the progression of late
      AMR. We plan to include 44 patients into this trial.

      Part B: Objective

      To evaluate the efficiency and safety of Bortezomib on allograft outcome in recipients with
      late AMR.

      Part B: Randomization

      44 patients will be centrally randomized by computer assignment between two study arms
      (Bortezomib versus Placebo) using a 1:1 randomization scheme. To avoid the bias of unbalanced
      baseline variables that potentially affect treatment responses, stratification will be
      performed for eGFR (> versus <50 ml/min/1.73 m2) and the presence or absence of Banff I
      T-cell-mediated rejection (index biopsy).

      For each patient a study ID will be assigned. The participating investigators and the
      patients will be blinded to group allocation until the completion of the study.

      Part B: Intervention

      Treatment with Bortezomib or Placebo. Patients allocated to the intervention group will
      receive two cycles of Bortezomib at an interval of 3 months. Each cycle will consist of
      Bortezomib 1.3 mg/m2 administered twice weekly on days 1, 4 and days 8, 11. Bortezomib will
      be given by injection intravenously (within 3-5 seconds). Bortezomib-treated patients will
      receive oral antiviral prophylaxis to prevent the development of herpes zoster infection:
      Valacyclovir 500 mg per day (eGFR <30 ml/min/1.73 m2: 250 mg per day) for 3 weeks after
      initiation of each cycle. Patients allocated to the control group will receive oral Placebo
      instead of Valacyclovir prophylaxis.

      According to our center standard, upon diagnosis of late AMR, all recipients (both study
      groups) on therapy with a calcineurin inhibitor (Tacrolimus or Cyclosporine A) or a mammalian
      Target of Rapamycin (mTOR) inhibitor (Everolimus or Sirolimus), without Azathioprine or MMF,
      will receive MMF (initially 2 x 500 mg per day; in absence of gastrointestinal side effects
      and significant leukopenia or thrombocytopenia stepwise increase of dose to 2 x 1000 mg per
      day) to avoid underimmunosuppression. Recipients weaned off steroids will receive low dose
      Prednisolone (initiation with 10 mg/day, tapered to 5 mg/day within 4 weeks).

      Part B: Sample size calculations, power calculation, statistical methodology and interim
      analyses

      Pilot studies using data from the OEDTR (OEsterreichische Dialysis and Transplant Registry)
      were conducted to estimate the eGFR decline and variance of sequential eGFR determinations.
      When using only one eGFR determination at two years as primary endpoint, analyses showed an
      impracticable high sample size number under the assumption of a median treatment effect of
      Bortezomib (0.5 SD). Therefore, the difference in slope of half yearly determined eGFR
      between the two treatment groups will be used as quantitative outcomes measure. Mixed linear
      models for longitudinal data were used for analyses. Power calculation using an
      autoregressive covariance matrix of the first order using a correlation of 0.9, an alpha of
      0.05 for the interaction term of treatment and time and an attrition of 8% per year showed
      that 2 x 22 subjects would be required to uncover a difference in GFR slopes of 5ml/min/year
      with a power of 80%.

      To provide a solid background for our power analysis, we re-analyzed a large retrospective
      transplant cohort (transplantation and follow-up at the Medical University Vienna. Evaluating
      the impact of late (> 6 months) (C4d-positive) AMR on the clinical performance of kidney
      allografts a GFR slope of -8.2 ml/year (over a follow-up of 6 years), as compared to a slope
      of -1.8 ml/min/year in non-biopsied and -2.8 ml/min patients in biopsied C4d-negative
      subjects was observed.

      All analyses will be conducted according to the intention-to-treat principle. Continuous data
      will be analyzed by t-test, categorical data by a chi-square test or Fisher's exact test when
      appropriate. The analysis of the creatinine trajectories stratified by treatment (and
      eventually by baseline GFR and T-cell-mediated rejection) will be performed by a mixed linear
      model with time and therapy as the independent parameters. The most applicable covariance
      matrices will be determined by graphical analysis and evaluated by the log likely hood ratio.

      This study will be monitored by an independent data and safety monitoring board (DSMB) of the
      Medical University of Vienna (MUW). Interim analyses will be performed by the DSMB after
      completion of 10 and 20 cases in the treatment group. The Lan & DeMets extension of the
      O'Brian-Fleming stopping rules will be applied (DeMets, LD et al. 1994). The trial will be
      stopped if the observed p-value is <0.00001 (first interim analysis after 10 patients) or
      <0.00305 (second interim analysis after 20 patients).

      Part B: Adverse events (AEs)

      The most probable AEs caused by Bortezomib (usually transient) are mild to moderate
      thrombopenia and leukopenia, decreased appetite, gastrointestinal side effects (vomiting,
      nausea, diarrhea), fatigue, peripheral neuropathy and other neurological symptoms such as
      headache, dizziness, tremor and hypotension. Intensified immunosuppression may be associated
      with an increased risk of infection (common: herpes zoster, herpes simplex, pneumonia,
      bronchitis, sinusitis, nasopharyngitis). Hence, a careful patient follow-up will include a
      close monitoring for infectious complications (bacterial, viral and fungal infections).
      Valacyclovir adapted to kidney function will be added to prevent the occurrence of herpes
      virus infections (herpes zoster).

      The most probable AEs caused by Valacyclovir are headache, nausea, gastrointestinal side
      effects (vomiting, diarrhea, abdominal pain), dizziness, hallucinations, confusion, changes
      in blood cell counts (Leukocytopenia, thrombocytopenia, anemia), increased liver and kidney
      parameters and fever.
    

Intervention name(s): Bortezomib; Placebo
Intervention type(s): Drug; Drug
Intervention description(s): Patients will receive two cycles of Bortezomib (Velcade®) at an interval of three months.
Each cycle will consist of intravenously administered (within 3-5 seconds) Bortezomib 1.3 mg/m2 twice weekly on days 1, 4, 8 and 11.; Patients will receive two cycles of Placebo (NaCl solution) at an interval of three months.
Each cycle will consist of intravenously administered (within 3-5 seconds) Placebo twice weekly on days 1, 4, 8 and 11.
Other intervention name(s): Velcade®; NaCl solution
Intervention MeSH terms: Bortezomib
Allocation: Randomized
Intervention model: Parallel Assignment
Masking: Quadruple (Participant, Care Provider, Investigator, Outcomes Assessor)
Primary purpose: Treatment
Keywords: Kidney transplantation; Antibody formation; Graft rejection; Proteasome inhibitors; Randomized controlled trial