Low-Dose Rituximab Therapy for Antibody-Mediated Rejection in a Highly Sensitized Heart-Transplant Recipient

Abstract

Antibody-mediated rejection is the B-cell–mediated production of immunoglobulin G antibody against the transplanted heart. The currently available therapies for antibody-mediated rejection have had marginal success, and chronic manifestations of rejection can result in an increased risk of graft vasculopathy and perhaps require repeat transplantation. Rituximab, a monoclonal antibody directed against the CD20 receptor of B-lymphocytes and approved as therapy for lymphoma, can be used in heart-transplant patients for the management of antibody-mediated rejection.

We present the case of a 52-year-old woman with high allosensitization (pre-transplantation panel reactive antibody level, 72%) who underwent successful orthotopic heart transplantation. Postoperatively, her acute antibody-mediated rejection with concomitant cellular rejection was successfully treated with low-dose rituximab. The patient died 5 months later because of multiple other medical problems. The present case suggests a role for low-dose rituximab as therapy for antibody-mediated rejection in heart-transplant patients.

Antibody-mediated rejection (AMR) in heart-transplant recipients is mediated by donor-specific antibodies and is histologically defined by linear deposits of immunoglobulin (Ig) and complement in the myocardial capillaries.¹ Antibody-mediated rejection is often accompanied by hemodynamic compromise and is associated with diminished graft survival. Standard immunosuppressive therapy, designed to target T-cell immune function, is largely ineffective against this B-cell–driven process. Various therapies for AMR, although available, can be of marginal use secondary to patients' comorbidities.^2,3 We present the case of a woman with a history of ventricular assist device (VAD) implantation, dialysis dependence, and severe thrombocytopenia who responded well to the addition of anti-CD20 monoclonal antibody therapy with rituximab after heart transplantation.

Case Report

A 52-year-old black woman with a history of nonischemic dilated cardiomyopathy and a left ventricular ejection fraction (LVEF) of 0.15 was transferred to our tertiary care facility for management of advanced heart failure. Her medical history included diabetes mellitus, 5 pregnancies, no previous transfusions, and acute worsening of chronic kidney injury that required ongoing hemodialysis. Upon arrival, the patient was in cardiogenic shock and on vasopressor support, and she required intra-aortic balloon pump implantation with mechanical ventilation. Two weeks later, she underwent implantation of a Thoratec® PVAD biventricular paracorporeal assist device (Thoratec Corporation; Pleasanton, Calif). The patient had an extremely complex and protracted hospital course that included coagulopathy, severe thrombocytopenia, Clostridium difficile infection with pseudomembranous colitis, and VAD drive-line infection. After 12 weeks, she was discharged from the hospital.

One month before heart transplantation (7 months after VAD placement), the patient's panel reactive antibody (PRA) levels were high (77%) as measured by flow cytometry, which was performed with use of human leukocyte antigen (HLA) class II Luminex-coated beads. After pretreatment with plasmapheresis and intravenous Ig for desensitization, the cytomegalovirus (CMV)-positive patient underwent a CMV-negative orthotopic heart transplantation. Her PRA level was tested again at this time, and it had decreased from 77% to 53%. Intraoperatively, the patient became anuric and was started on continuous venovenous hemofiltration. Initial postoperative immunosuppressive therapy included intravenous methylprednisolone and mycophenolate mofetil. The patient was given 2 more treatments with intravenous Ig and plasmapheresis over the course of 5 days. The retrospective, flow-cytometric donor crossmatch was weakly positive for B cells. During the first postoperative week, she developed atrial tachyarrhythmia that required electrical and chemical cardioversion. An echocardiogram showed an LVEF of 0.60. The first 2 endomyocardial biopsies at postoperative weeks 1 and 2 (Fig. 1) were negative for acute cellular rejection (International Society for Heart & Lung Transplantation [ISHLT] grade 0). The patient was maintained on methylprednisolone and mycophenolate therapy (500–750 mg/d). Right-side heart catheterization revealed elevated right-side filling pressures with moderate pulmonary hypertension and a pulmonary artery pressure of 50 to 60 mmHg. The patient's diuretic agents were increased. The 4 subsequent weekly biopsies (weeks 3–6) revealed ISHLT grade 1R acute cellular rejection with no AMR. At week 3, the patient was given 75 mg of daclizumab, an interleukin-2 antagonist, which failed to yield any histologic improvement. This was followed by low-dose thymoglobulin (antithymocyte globulin: total, 75 mg) at week 5. Successive biopsies were negative for acute cellular rejection. However, the biopsy at postoperative week 10 showed ISHLT grade 2R acute cellular rejection with no AMR. The patient's immunosuppressive therapy was augmented with pulsed methylprednisolone and daclizumab. Biopsy a week later showed ISHLT grade 2R with immunofluorescence that suggested AMR (weakly positive co-localization of C4d in the interstitial capillaries) (Fig. 2). An echocardiogram showed a normal, well-preserved LVEF. Because the patient was dialysis-dependent and fluid-overloaded, she did not undergo plasmapheresis, and cyclophosphamide could not be given because of her ongoing severe thrombocytopenia (platelet count, 30–40 ×10⁹/L). The patient was given a repeat dose of thymoglobulin and pulsed steroid. Because of the multiple medical problems that precluded conventional therapy, it was decided to add specific anti-B-cell therapy with rituximab.

Fig. 1 Graph depicts the level of acute cellular rejection (based on endomyocardial biopsy results) and medical therapy after transplantation. Arrows show times when biopsy results were positive for antibody-mediated rejection. Baseline maintenance immunosuppressive therapy included steroids and mycophenolate mofetil (500–750 mg).

D = daclizumab; R = rituximab; T = thymoglobulin

Fig. 2 Photomicrographs, obtained at week 11 before rituximab. A) Focal moderate acute cellular rejection is indicated by diffuse interstitial infiltration of mononuclear cells (H & E, orig. ×400). B) Immunostaining with C4d monoclonal antibody shows weakly positive peritubular capillary deposition (immunoperoxidase stain, orig. ×400).

Rituximab therapy was started at a single low dose of 200 mg (100 mg/m²). The patient's CD20 level fell from 17% to zero 48 hours after the first dose. She was given another dose a week later, because a repeat biopsy continued to show C4d mild diffuse staining of the interstitial capillaries and the HLA class I and II antibodies were strong (Table I). Subsequent biopsies showed no AMR or cellular rejection, with complete suppression of the CD20 counts. The patient's LVEF continued to be well preserved. However, while awaiting discharge from the hospital 2 weeks after the last negative surveillance biopsy, she went into asystolic cardiac arrest and could not be resuscitated.

Table I. Flow Cytometry Determinations of PRA Levels and Strength of Antibody Subtypes in Our Patient

The postmortem analysis revealed no diagnostic evidence of AMR or cellular rejection. The heart was grossly and histologically unremarkable; however, the substantial amount of pulmonary, vascular, hepatic, and splenic congestion suggested fluid overload.

Discussion

Although AMR is much less frequent than cellular rejection, the clinical consequences are more overwhelming. Antibody-mediated rejection is associated with substantial allograft dysfunction and coronary artery vasculopathy.³ Despite strong evidence that AMR is clinically important, it is infrequently reported and poorly understood. Its diagnosis requires a high degree of suspicion.⁴ Antibody-mediated rejection can occur in conjunction with cellular rejection, but it can also occur without significant histologic abnormalities. Half of patients show no evidence of lymphocytic infiltrate (ISHLT grade 0); the others have grade IA-to-IB cellular rejection.⁵ To detect Ig and complement deposition, additional endomyocardial biopsy fragments must be taken for frozen sections at the time of endomyocardial biopsies.² Specific diagnostic criteria for AMR include capillary endothelial-cell swelling, interstitial hemorrhage, and interstitial edema with immunofluorescent evidence of Ig deposition (in particular, IgM, IgG, C3, C4d, CIQ, and HLA-DR) and immunoperoxidase staining for CD68 (macrophages) and CD34 (capillary endothelium).^1,3,6

Among therapies for AMR, intravenous Ig has a proven, important role. Numerous proposed mechanisms of action include modification of autoantibody and alloantibody levels through induction of anti-idiotypic circuits,^7,8 inhibition of cytokine gene activation and anti-cytokine activity,^7,9 anti-T-cell receptor activity,¹⁰ Fc receptor-mediated interactions with antigen-presenting cells to block T-cell activation,^7,11 anti-CD4 activity,¹² stimulation of cytokine receptor antagonists,⁷ and inhibition of complement activity.^13,14 Plasmapheresis with intravenous Ig also offers substantial benefits in the treatment of AMR—especially for patients with high-titer anti-HLA antibodies and for those who receive ABO-incompatible transplants. The mechanisms of action are not clearly understood, but they appear to relate to the depletion of high-titered antibody followed by the suppression of antibody synthesis from intravenous Ig replacement.¹⁵

Rituximab, a rather new therapy for AMR, is a chimeric humanized monoclonal antibody IgG kappa immunoglobulin that binds to the CD20 that is expressed on all B cells from the early pre-B-cell to the mature-B-cell stage of differentiation. CD20 is not expressed on pluripotent hematopoietic stem cells, pro-B cells, or terminally differentiated plasma cells. Rituximab produces lysis of CD20-positive B cells, depleting them from the peripheral circulation, the lymph nodes, and the bone marrow.¹⁶ It has been approved and is commonly used as therapy for low-grade B-cell non-Hodgkin lymphoma.

Rituximab's half-life is proportional to the dose. The drug is detectable in serum 3 to 6 months after therapy. Although secondary antibody deficiencies were not seen among lymphoma patients who were given rituximab, it has been suggested that antibody levels should be routinely measured after the administration of rituximab in solid-organ transplantation.¹ The reported toxicity profile of rituximab is mild; the most commonly reported side effects are fever, chills, and hives, mainly during the first infusion. The most substantial side effects—neutropenia, hypotension, and bronchospasm—have been reported in less than 2% of patients.¹⁷ Patients who undergo successful rituximab therapy for AMR require careful monitoring, because the B-cell population can rebound as long as 9 months after therapy ends. Apart from the treatment of AMR, rituximab might have a role in patients who are heavily sensitized before transplantation. A regimen of plasmapheresis followed by rituximab to reduce memory B cells has been suggested, to permit transplantation in patients who have unacceptably high PRA levels.⁴

There have been a few reports of the successful use of rituximab to treat AMR in patients who have received transplanted hearts. The dosage in those cases, 375 mg/m², was much higher than that given to our patient and comparable to the usual therapeutic dosage for lymphoma.^2,4 To our knowledge, ours is the first report of the successful treatment of AMR with such a low dose of rituximab (100 mg/m²) in a heart-transplant recipient. Vieira and colleagues¹⁸ revealed in their dose-escalation study of renal transplantation that a single dose of rituximab, even at 50 mg/m², depleted B cells from the peripheral blood as effectively as did a single dose of 375 mg/m². In that study, the B-cell count remained suppressed for at least 1 year, although recovery began in 6 months. Consistent with these results, dosages as low as 15 mg/m² have been reported to cause complete circulatory B-cell depletion in renal-transplant recipients.¹⁹ B-cell recovery began 3 to 6 months after rituximab administration, which was faster than B-cell recovery after the standard dose of 375 mg/m².²⁰

Our report adds to the evidence that supports the role of rituximab as therapy for AMR in solid-organ transplantation, including cardiac transplantation. Whether rituximab will be adopted on a widespread basis as therapy in heart transplantation will depend upon results of large-scale trials that are warranted in similar patient populations.