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Ann Thorac Surg 1995;60:372-376
© 1995 The Society of Thoracic Surgeons

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Original Articles: Cardiovascular

Antithymocyte Globulin and Methotrexate Therapy of Severe or Persistent Cardiac Allograft Rejection

Department of Surgery, Montreal Heart Institute, Montreal, Quebec, Canada

Accepted for publication March 28, 1995.

	Abstract

Top Footnotes Abstract Introduction Material and Methods Results Comment References

 
Background. The treatment of severe or persistent acute rejection remains difficult despite newer immunosuppressive agents available.

Methods. To evaluate the effectiveness of rabbit antithymocyte globulin and methotrexate as therapy for severe or persistent acute cardiac allograft rejection, we conducted a retrospective analysis of clinical and laboratory data from 150 consecutive heart transplant recipients between 1983 and 1994.

Results. Thirteen episodes of severe or refractory acute rejection were treated with rabbit antithymocyte globulin in 10 patients. Rabbit antithymocyte globulin (125 mg/day for 3 consecutive days) was effective in 90% of patients. Therapy was well tolerated, and contributed to one infectious complication, no malignancy, and long-term survival in 8 of 10 patients. Recurrent rejection developed in 60% of patients. Methotrexate (7.5 to 15 mg/wk for 16 weeks) was administered to 8 patients with persistent rejection documented on three consecutive endomyocardial biopsies. Therapy was effective in 6 of the 8 patients, with one infectious complication and no malignancy on follow-up. White blood cell count decreased significantly during therapy (p = 0.008). Seven of the 8 patients in the methotrexate group are long-term survivors.

Conclusions. Rabbit antithymocyte globulin is a valuable alternative in patients with severe or refractory acute rejection. Methotrexate is an important adjunct in patients with persistent rejection unresponsive to conventional immunosuppressive regimens.

	Introduction

Top Footnotes Abstract Introduction Material and Methods Results Comment References

 
See also page 376.

Cardiac transplantation is now an accepted treatment of end-stage heart disease. The use of cyclosporine as long-term maintenance immunosuppression has resulted in a significant improvement in patient survival [1–4]. Although cyclosporine has been effective in decreasing the incidence of acute rejection, it remains an important cause of morbidity after transplantation. Intravenous administration of methylprednisolone and high doses of oral prednisone usually are effective in controlling acute rejection. Nonetheless, a more aggressive approach occasionally is necessary in patients with severe, recurring or life-threatening episodes of acute rejection.

Rabbit antithymocyte globulin (RATG) is a potent immunosuppressive drug used in both the prevention and treatment of acute rejection. Methotrexate, a folic acid analogue with immunosuppressive properties, may be of benefit in the management of recurrent or persistent rejection processes. The objective of this study is to evaluate the effectiveness of RATG as therapy for severe acute cardiac allograft rejection and methotrexate as treatment for persistent episodes of rejection.

	Material and Methods

Top Footnotes Abstract Introduction Material and Methods Results Comment References

 
A retrospective analysis of clinical and laboratory data from 150 consecutive patients who underwent cardiac transplantation at the Montreal Heart Institute between 1983 and 1994 was carried out. In all patients, cardiac biopsy for surveillance of acute rejection was performed every week during the first month after transplantation, every other week during the next 3 months, and monthly thereafter until the end of the first year after transplantation. Thirteen episodes of severe or refractory acute rejections were treated with RATG in 10 patients during this period. Routine maintenance immunosuppression consisted of cyclosporine and prednisone in 2 patients, whereas the remaining eight received a triple regimen of cyclosporine, prednisone, and azathioprine with RATG induction in the immediate postoperative period [5]. All but one episode of rejection occurred during the first year after transplantation, and all were refractory or severe episodes (grade 3 or 4) according to Billingham and associates' histologic classification [6]. Two patients were in cardiogenic shock at the time of diagnosis and treatment.

Rabbit antithymocyte globulin (Institut Merieux, Lyon, France) at a dosage of 125 mg was infused intravenously over an 8-hour period for 3 consecutive days, once the diagnosis of severe, uncontrolled acute rejection was established at endomyocardial biopsy. In 1 patient, three additional doses of RATG (125 mg) were given every other day for a total of six doses over a 9-day period. Pretreatment with acetaminophen, diphenhydramine, and methylprednisolone (125 mg) was administered before each RATG infusion.

In 4 patients of the RATG group, the initial episode of moderate or severe acute rejection was treated with intravenous methylprednisolone (1 g daily for 3 days) or oral prednisone at a dose of 100 mg/day tapered to 20 mg/day in 14 days, and RATG was used as subsequent treatment of refractory or uncontrolled severe rejection. In the other 6 patients, RATG was used to treat the first severe acute rejection because the episode occurred less than 3 weeks after transplantation in 3 patients, for cardiogenic shock in 2 patients, and for severe corticosteroid side effects in the remaining patient.

Methotrexate has been administered since 1993 in 8 patients with acute persistent rejection. Maintenance immunosuppression in these patients consisted of the standard cyclosporine, prednisone, and azathioprine regimen with postoperative RATG induction. Episodes treated with methotrexate occurred within the first year of transplantation in all patients. Persistent rejection was defined as evidence of acute rejection on three consecutive endomyocardial biopsies. Five patients presented a persistent grade 2 rejection process, 2 patients had grade 3A rejection, and in 1 patient, biopsies showed grade 3B rejection. None of the patients in this group had hemodynamic instability.

Methotrexate was administered in oral doses of 2.5 or 5 mg three times daily, 1 day a week for 16 weeks. The total cumulative dosage per patient ranged from 90 to 240 mg. Patients were monitored for systemic complications, leukopenia (defined as white blood count <4,000/mL) and secondary infections. White blood cell counts were obtained weekly throughout the treatment period, and the dosage of methotrexate and azathioprine was adjusted accordingly. Variation in dosage of immunosuppressive agents and blood levels of cyclosporine during therapy are shown in Table 1. Previous therapy with intravenous steroids or oral prednisone (bolus and tapered) had been attempted in each of the 8 patients.

	Results

Top Footnotes Abstract Introduction Material and Methods Results Comment References

 
Patient Characteristics
The 10 patients who required treatment with RATG for severe acute rejection represented 7% (10/150) of the total number of transplant patients. There were 4 women and 6 men with a mean age of 44 ± 5 years. Before the RATG-treated episode, 4 patients had experienced a mean of 2 ± 1 previous episodes of rejection requiring pulse methylprednisolone or oral prednisone taper (Table 2). The time from transplantation to the treatment with RATG averaged 59 ± 20 days. In the methotrexate group, there were 4 women and 4 men with a mean age of 44 ± 3 years. The average number of previous rejection episodes requiring therapy was 2 ± 1. Episodes treated with methotrexate occurred a mean of 9 ± 3 months after transplantation.

Subsequent rejection episodes recurred after an average of 63 ± 15 days in 6 patients (60%) treated with RATG, whereas 4 patients remained free of recurrences. Retreatment with RATG was successful in 2 patients with recurrences, whereas prednisone taper was used for moderate rejection in the 4 other cases. Overall, 8 of the 10 patients in the RATG-treated group are long-term survivors and 2 patients died of acute rejection during the first year after transplantation.

Of the 8 patients treated with methotrexate, 6 showed marked regression of the rejection process at biopsy for a success rate of 66%. The grade of rejection improved from 2 or 3 to grade 0 or 1A in 6 patients. In 1 patient with grade 2 rejection, the biopsy showed no signs of improvement after 12 weeks of methotrexate. A biopsy performed 3 months after the end of therapy revealed a persistent grade 2. The patient, however, remained asymptomatic with no signs of graft dysfunction on echocardiography or scintigraphy. The last patient in this group did not respond to methotrexate and progressed from grade 2 to grade 3A shortly after the end of therapy. The patient eventually responded to an oral prednisone (bolus and tapered) regimen. Of the patients with a complete response, 1 presented signs of recurrent rejection on biopsy. In the months after the administration of methotrexate in the 8 patients, a recurrent rate of rejection of 0.1 ± 0.1 episode per patient was noted. In the methotrexate-treated group, 1 patient died of acute rejection 7 months after the end of his therapy.

Side Effects of Therapy
There were no immediate side effects after the administration of RATG in the patients initially treated or in the 2 patients who required retreatment with RATG. Pneumocystis carinii pneumonia developed in 1 patient 3 weeks after treatment of an acute episode of rejection with RATG. No clinical signs of a lymphoproliferative disorder developed at a mean follow-up of 45 ± 13 months.

Methotrexate therapy was well tolerated in all patients. Mild leukopenia was found in two patients (3,800 and 4,000 cells/µL) during the third month of treatment with methotrexate (Fig 1). The white blood cell count at the end of therapy averaged 6,500 ± 682 cells/µL, a significant decrease from the mean value before initiation of methotrexate (p = 0.008). One infectious complication was noted during the third month of therapy. A Pneumocystis carinii pneumonia was diagnosed and treated successfully in 1 patient. No patient presented signs of a malignancy with a mean follow-up of 11 ± 1 months.

View larger version (18K): [in this window] [in a new window]   Fig 1. . White blood cell count in patients receiving methotrexate. (p = 0.008; *p < 0.05 versus pretreatment, month 1, and month 2.)

	Comment

Top Footnotes Abstract Introduction Material and Methods Results Comment References

Rabbit antithymocyte globulin has been used for the prevention of acute rejection immediately after transplantation with satisfactory results, as well as for the treatment of acute allograft rejection [8]. In all but 1 of our patients, RATG was effective in controlling the severe and refractory episodes of acute rejection. However, recurrence of rejection was common during the following months.

Methotrexate, a folic acid analogue, interferes with DNA and purine synthesis by inhibiting intracellular enzymes, primarily dihydrofolate reductase. In addition, methotrexate possesses cytotoxic inhibitory effects on both cellular and humoral immunities, suppressing cell-mediated immune responses and the production of immunoglobulins [9]. Methotrexate also has been shown to reduce the incidence of graft versus host disease after bone marrow transplantation [10].

The treatment of cardiac allograft rejection with methotrexate was first described by Costanzo-Nordin and associates in 1988 [11]. They reported a 90% success rate in reversing the rejection process in transplant patients. In our series, treatment was effective in 6 of 8 patients with persistent rejection, whereas resolution rates greater than 90% have been reported by several authors in recent studies [12–14]. Methotrexate also has been effective in pediatric heart transplant recipients [14, 15]. Although a number of protocols show wide variations in cumulative doses (10 to 240 mg) and length of therapy (3 weeks to 4 months), their effectiveness seems comparable. Methotrexate therapy of persistent rejection, however, is associated with a significant rate of recurrent rejection process. Only 20% to 50% of patients actually remain free of recurrences for any considerable length of time [11, 12, 16].

Other polyclonal antibodies and the monoclonal antibody OKT3 also have been used in the treatment of acute rejection after transplantation. In a prospective, randomized study comparing Minnesota antilymphoblastic globulin and OKT3, Deeb and colleagues [17] found that both drugs were effective in resolving recurrent episodes of acute rejection, but that OKT3-treated patients had a higher death rate due to subsequent infections than those treated with Minnesota antilymphoblastic globulin. Many uncontrolled clinical studies report excellent results with the monoclonal antibody OKT3 in the treatment of severe and recurrent acute rejection after heart transplantation [18–20]. A recent metaanalysis of the clinical experience with OKT3 also demonstrated the value of OKT3 in these life-threatening clinical situations [21].

Adverse reactions associated with methotrexate are few and generally well tolerated. These include mucositis, gastrointestinal intolerance, hypersensitivity pneumonitis, and hepatotoxicity. These side effects, however, usually are seen in patients receiving large cumulative dosages of 1,500 to 3,000 mg [22]. Bone marrow suppression and secondary infections represent the most serious complications of methotrexate therapy. The incidence of severe leukopenia may be as high as 30% to 50% [11, 13]. In a multivariate analysis, Bourge and associates [13] identified a number of risk factors for leukopenia: lower white blood cell count before methotrexate therapy, higher number of rejection episodes before treatment, and a higher total dose of methotrexate. In our series, however, although a substantial cumulative dosage of methotrexate (90 to 240 mg) was administered, mild leukopenia developed in only 2 patients. Robinson and co-workers [23] have reported similar marrow-suppressive effects even with ``very low dose'' schedules (10 mg/month) of methotrexate. Robinson and co-workers suggest that impaired renal and hepatic excretion of methotrexate (possibly associated with cyclosporine toxicity) may result in a prolonged exposure and greater myelotoxicity. Infectious complications occur infrequently [11, 12, 15, 16] but remain an important cause of morbidity and mortality in some patients receiving methotrexate. Although methotrexate may contribute to this phenomenon, the use of a triple-drug immunosuppressive regimen before and during methotrexate therapy also may have a significant role.

In our experience, treatment of severe, refractory, and life-threatening acute rejection with RATG after cardiac transplantation has proved to be effective in most patients without immediate side-effects or complications. One patient had development of a serious pulmonary infection due to Pneumocystis carinii, 2 died of irreversible graft dysfunction, and no lymphoproliferative disorder was seen at follow-up. The satisfactory clinical outcome obtained with RATG in the treatment of severe acute rejection indicates that this polyclonal antibody is a valuable alternative in these desperate situations, at a considerably lower cost than that of treatment with the monoclonal antibody OKT3 [24]. The long-term effects of persistent or recurrent rejection on graft function or coronary artery disease are unknown. The possible benefits of therapy therefore must be weighed against the significant side effects of cumulative immunosuppression such as myelotoxicity, infection, and malignancy. Methotrexate is nonetheless an important adjunct in patients unresponsive to conventional steroid regimens with persistent or recurrent acute allograft rejection. In the presence of severe (grade 4) and refractory rejection after cardiac transplantation, we recommend RATG therapy in association with intravenous steroids.

	Footnotes

Top Footnotes Abstract Introduction Material and Methods Results Comment References

 
Address reprint requests to Dr Carrier, Montreal Heart Institute, 5000 Belanger St, Montreal, Que, H1T 1C8, Canada.

	References

Top Footnotes Abstract Introduction Material and Methods Results Comment References

 

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This Article Abstract Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Add to Personal Folders Download to citation manager Author home page(s): Pasquale Ferraro Michel Carrier Permission Requests Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Ferraro, P. Articles by Pelletier, L. C. Search for Related Content PubMed PubMed Citation Articles by Ferraro, P. Articles by Pelletier, L. C. Related Collections Related Article