In 2002, we opened a new avenue in the management of ABO-incompatible kidney transplantation (ABO-IKT) by introducing an anti-CD20 monoclonal antibody, rituximab (1, 2). Our initial intention was to perform ABO-IKT in nonresponders who were refractory to the conventional preoperative regimen, consisting of plasma-exchange (PEX) or double-filtration plasmapheresis (DFPP) plus splenectomy. Later, application of the preoperative regimen comprising rituximab and PEX or DFPP was extended, and the need for splenectomy was eliminated (3, 4).
In the clinical setting at the time, not many studies of kidney transplantation had attempted to use a monoclonal antibody to eliminate B cells, which produce anti-ABO blood type natural antibody (5). The chimeric antibody, rituximab, was originally developed for CD20-positive B-cell, non-Hodgkin lymphoma in 1997 (6), and has since been used for treatment of not only hematological malignancies but also various other diseases, such as rheumatoid arthritis (7) and systemic lupus erythematosus (8). Because rituximab has fewer side-effects and has been shown to have high ability to eliminate CD20-positive B cells, we started our program to apply this antibody for ABO-IKT, as donor sources are very limited in Japan, and at the time kidney transplantation had to be abandoned in patients who were refractory to the conventional preoperative regimen.
Application of rituximab has dramatically expanded the eligibility criteria for ABO-IKT recipients and reduced the invasiveness of the operative procedure (3). Furthermore, the clinical course, graft function, and outcome of rituximab-treated ABO-IKT recipients have been better than those of ABO-compatible kidney transplantation (ABO-CKT) recipients and ABO-IKT recipients not treated with rituximab.
After our successful trials, many reports of rituximab application to kidney transplantation have been published (3, 4, 9–12). Here, we report the 5-year outcome of ABO-IKT recipients treated with rituximab at our institution, which, although not based on a prospective, randomized study, nevertheless indicated the value and utility of this approach.
The patients' background factors are shown in Table 1. Mean patient ages in groups A, B, and C were 43.5±13.7 years, 46.0±12.9 years, and 52.6±12.1 years, respectively (P<0.0001), patients in groups B and C being significantly older than those in group A. Donor relationships showed different patterns among the groups. In group A, graft donations from parents were the most frequent (41.8%), whereas donations from spouses were more frequent in groups B (44.4%) and C (60.0%; P=0.0006). The most common ABO blood type of the recipients was A in group A (45.0%) and O in groups B (44.4%) and C (48.0%; P<0.0001). The most common ABO blood type of the donors was O in group A (42.9%), A and B in group B (36.7%), and A in group C (50.0%; P<0.0001).
Although the numbers of mismatches at class I loci did not differ among the groups (P=0.0749), the numbers of mismatches at class II loci were higher in groups B and C than in group A (P=0.0105). There were no significant differences in the duration of dialysis or history of diabetes among the three groups. Preoperative antidonor type antibody titers did not differ significantly between groups B and C.
Table 2 shows the data for donor conditions. Donor ages in groups A, B, and C were 54.8±11.3 years, 57.9±9.8 years, and 52.6±12.1 years, respectively (P=0.063). There were no significant differences in graft weight, warm ischemia time, total ischemia time, and time until first urination among the three groups.
The numbers of episodes of acute antibody-mediated rejection after KTx in groups A, B, and C were 7 (2.5%), 10 (15.9%), and 2 (4.0%), respectively (P=0.651; Kruskal-Wallis test). Although the number of such episodes in group A was significantly lower than in group B (P<0.0001; Fisher's exact test), the number merely tended to be lower in group C than in group B (P=0.0651; Fisher's exact test). There was no significant difference in the number of episodes of acute antibody-mediated rejection between groups A and C (P=0.221; Fisher's exact test).
The numbers of episodes of acute cellular rejection after KTx in groups A, B, and C were 40 (14.3%), 6 (9.5%), and 2 (4.0%), respectively (P=0.0495; Kruskal-Wallis test). The number of such episodes tended to be lower in group C than in group A (P=0.0618; Fisher's exact test), and did not differ significantly between groups A and B (P=0.4141; Fisher's exact test), and between groups B and C (P=0.2983; Fisher's exact test).
Median values of serum creatinine (sCr) in groups A, B, and C on postoperative day 5 were 1.23 (0.23–10.38) mg/dL, 1.30 (0.44–8.55) mg/dL, and 1.30 (0.68–15.8) mg/dL, respectively (P=0.7042; Kruskal-Wallis test). Median values of sCr in groups A, B, and C on postoperative day 30 were 1.46 (0.24–9.00) mg/dL, 1.55 (0.49–9.50) mg/dL, and 1.28 (0.76–3.72) mg/dL, respectively (P=0.3848; Kruskal-Wallis test).
Median values of sCr in groups A, B, and C at various times after transplantation were 1.34 (0.48–5.97) mg/dL, 1.33 (0.55–4.80) mg/dL, and 1.23 (0.63–3.16) mg/dL at 1 year (P=0.856), 1.38 (0.52–7.60) mg/dL, 1.39 (0.64–7.10) mg/dL, and 1.22 (0.71–3.29) mg/dL at 3 years (P=0.2683), and 1.43 (0.62–10.56) mg/dL, 1.38 (0.65–6.11) mg/dL, and 1.33 (0.94–2.68) mg/dL at 5 years (P=0.8141), respectively.
The graft survival rates in groups A, B, and C were 99.2%, 96.8%, and 100% at 1 year, 93.8%, 94.9%, and 100% at 3 years, and 88.4%, 90.3%, and 100% at 5 years after transplantation (P=0.726), respectively (Fig. 1). None of the patients in any of the three groups died during the observation period.
Figure 2 shows the sCr level relative to the dose of rituximab at 1 year after transplantation. Mean sCr values of the patients who were administered rituximab at a doses of 100, 200, and 500 mg were 1.20±3.25 mg/dL, 1.39±0.51 mg/dL, and 1.46±0.31 mg/dL, respectively (P=0.363).
Cytomegalovirus (CMV) infection was observed in 78 cases (27.9%) in group A, 28 (44.4%) in group B, and 13 (26.0%) in group C. There were no significant differences in the CMV infection rates among the three groups (P=0.1801; Kruskal-Wallis test). However, two-group comparisons showed significant differences in the CMV infection rate between groups A and B (P=0.0101; Fisher's exact test) and groups B and C (P=0.0428; Fisher's exact test), but no significant difference between groups A and C (P=0.7866; Fisher's exact test).
In this series, the 5-year graft survival rate tended to be higher in group C than in groups A and B, but without statistical significance (Fig. 1). The number of episodes of acute antibody-mediated rejection tended to be lower in groups A and C than in group B (Table 2). Two-group comparisons showed that the incidence of acute antibody-mediated rejection was significantly lower in group A than in group B, and tended to be lower in group C than in group B. There were fewer episodes of acute cellular rejection in groups B and C than in group A. Tydén et al. (13) reported the outcome of a randomized controlled study of kidney transplantation in rituximab-treated recipients. Although their series included cases of ABO-compatible kidney transplantation, they concluded that rituximab-treated recipients tended to have fewer episodes of, and milder, acute rejection than control patients. Their conclusions are in accord with ours.
The recipients in groups B and C of the present study were significantly older than those in group A. The main reason for this difference was the relationship of the donors. In group A, parents were the most frequent donors, whereas spouses were the most frequent donors in groups B and C. In general, in Japanese society, parents concentrate their efforts on raising their children until the children become adults. After their children graduate from university or high school, parents then feel free to act as donors for kidney transplantation.
Parents and children share basically one haplotype of the chromosome, and so the number of matched loci is higher for parent-child combinations than for spouse combinations. The reason for the differences in the number of mismatched loci between classes I and II found in the present study (Table 1) remains unknown. There were no differences in donor age, warm ischemia time, total ischemia time, or time until first urination among the three groups (Table 2). Thus, most of the factors of the recipients, donors, and operations did not differ among the groups. Furthermore, immunological factors such as mismatches in major histocompatibility complex were even tougher in groups B and C than in group A.
CMV infection is a serious complication after organ transplantation. Kamar et al. (14) reported that the occurrence of CMV infection did not differ between patients with or without rituximab therapy. In our series, CMV infection was observed in 27.9% of group A, 44.4% of group B, and 26.0% of group C. Although the study by Kamar et al. included various indications for rituximab therapy such as antibody-mediated acute rejection and the presence of anti-human leukocyte antigen antibodies, our result was compatible with theirs. The incidence of acute antibody-mediated rejection was highest in group B, and that of acute cellular rejection was highest in group A, and higher in group B than in group C. Steroid-pulse therapy was performed to treat antibody-mediated rejection and acute cellular rejection, so this might have increased the risk of CMV infection.
In group C, two patients developed pneumonia due to Pneumocystis carinii in the same season, 73 and 77 months after transplantation. Immunosuppression was tapered in these two patients and eventually their kidney grafts were rejected. At that time, a small epidemic of P. carinii pneumonia occurred, and this infection was observed in three patients in group A and one in group B.
As shown in Figure 2, there were no differences in sCr 1 year after transplantation, irrespective of the dose of rituximab. In the current protocol, patients are given rituximab at a dose of 100 mg/body only at 7 days before transplantation.
Our initial purpose was to explore the preconditioning regimen that would facilitate kidney transplantation in nonresponders, and we found that the rituximab-containing preconditioning regimen achieved broader effects that far exceeded our initial expectation. Patients who received the rituximab-containing preconditioning regimen had far lower incidences of acute antibody-mediated rejection and acute cellular rejection, and the regimen did not increase the incidence of CMV infection. The 5-year graft survival tended to be better in rituximab-treated patients than in ABO-CKT or ABO-IKT patients who were not treated with rituximab.
The results of the present study suggest that, in the long term, a preoperative regimen including rituximab, together with improvements in other forms of immunosuppression and patient management, yields a better outcome than ABO-CKT or rituximab-untreated ABO-IKT, without any increase in the risk of infection.
MATERIALS AND METHODS
Between January 2002 and December 2008, 408 patients with end-stage renal disease underwent living-related kidney transplantation at the Department of Surgery, Kidney Center, Tokyo Women's Medical University. The median follow-up period was 36.4±19.57 months. The patients were divided into three groups: group A (n=280) ABO-CKT, group B (n=63) ABO-IKT without rituximab induction, and group C (n=50) ABO-IKT with rituximab induction. All the patients in group B underwent splenectomy at the time of transplantation. In group C, only one patient underwent splenectomy at the time of transplantation. Rituximab was administered at 100 mg (n=6), 200 mg (n=26), and 500 to 1000 mg (n=18) one, two or three times, depending on the timing of kidney transplantation.
Basic immunosuppression in all three groups consisted of basiliximab, steroid, cyclosporine or tacrolimus, and mycophenolate mofetil (MMF). Basiliximab was given at a dose of 20 mg on the day of the operation and on postoperative day 4. Steroid at a dose of 250 mg was administered intravenously on the day of the operation, and then tapered. Cyclosporine or tacrolimus was started 2 days before the operation and continued throughout the postoperative course. Cyclosporine was given so as to maintain a blood trough level of 250 to 300 ng/mL during the first 7 days after surgery, 200 to 250 ng/mL during the first month, 150 to 200 ng/mL until 3 months, and 100 to 150 ng/mL thereafter. Tacrolimus was given so as to maintain a blood trough level of 15 to 20 ng/mL during the first 7 days after the operation, 10 to 15 ng/mL during the first month, 8 to 12 ng/mL until 3 months, and 5 to 10 ng/mL thereafter. MMF was given at a dose of 2 g/day, 2 days before the operation and continued thereafter.
Patients who were diagnosed as showing acute cellular rejection or acute antibody-mediated rejection were treated with a bolus injection of 500 mg of steroid for 2 consecutive days, which was then tapered.
Preconditioning Regimens in Groups B and C
In groups B and C, preoperative PEX or DFPP was performed to reduce the antidonor blood type antibody titer to ×16 or under as determined by the saline method and Coomb's method. Splenectomy was performed 7 days before transplantation, or simultaneously with transplantation.
Diagnosis of Rejection
Acute rejection was suspected when the sCr level was increased relative to the value on the previous day, together with an evident decrease of urine volume. Renal biopsy was performed in all the suspected cases, and histological diagnoses were made. Acute cellular rejection and acute antibody-mediated vascular rejection were diagnosed on the basis of the Banff criteria.
After discharge from the hospital, patients visited the outpatient clinic monthly. Graft survival was defined as the period from the day of transplantation to the time when patients were re-introduced to dialysis.
CMV infection was defined as positivity for CMV antigenemia (SRL, Tokyo, Japan), and treated with ganciclovir.
Data were expressed as means ± standard deviation, or median (minimum-maximum). Comparisons among the three groups were analyzed with Kruskal-Wallis test with post hoc test (Dunn's multiple comparison test). Comparison of contingency tables was performed with Fisher's exact test. The graft survival rates were estimated by the Kaplan-Meier method, tested by log-rank test, and analyzed by Cox regression. Differences at P less than 0.05 were considered significant.
1. Sawada T, Fuchinoue S, Teraoka S. Successful A1 to O, ABO- incompatible kidney transplantation
after a preconditioning regimen consisting of anti-CD20 monoclonal antibody infusions, splenectomy, and double filtration plasmapheresis. Transplantation
2002; 74: 1207.
2. Sawada T, Fuchinoue S, Kawase T, et al. Preconditioning regimen consisting of anti-CD20 monoclonal antibody infusions, splenectomy and DFPP-enabled non-responders to undergo ABO-incompatible kidney transplantation
. Clin Transplant
2004; 18: 254.
3. Saito K, Nakagawa Y, Suwa M, et al. Pinpoint targeted immunosuppression: Anti-CD20/MMF desensitization with anti-CD25 in successful ABO-incompatible kidney transplantation
without splenectomy. Xenotransplantation
2006; 13: 111.
4. Tanabe K, Ishida H, Shimizu T, et al. Evaluation of two different preconditioning regimens for ABO-incompatible
living kidney donor transplantation. A comparison of splenectomy vs. rituximab
-treated non-splenectomy preconditioning regimens. Contrib Nephrol
2009; 162: 61.
5. Aranda JM Jr, Scornik JC, Normann SJ, et al. Anti-CD20 monoclonal antibody (rituximab
) therapy for acute cardiac humoral rejection: A case report. Transplantation
2002; 73: 907.
6. Maloney DG, Grillo-López AJ, White CA, et al. IDEC-C2B8 (Rituximab
) anti-CD20 monoclonal antibody therapy in patients with relapsed low-grade non-Hodgkin's lymphoma. Blood
1997; 90: 2188.
7. De Vita S, Zaja F, Sacco S, De Candia A, et al. Efficacy of selective B cell blockade in the treatment of rheumatoid arthritis: Evidence for a pathogenic role of B cells. Arthritis Rheum
2002; 46: 2029.
8. Kneitz C, Wihelm M, Tony H. Effective B cell depletion with rituximab
in the treatment of autoimmune diseases. Immunobiology
2002; 206: 519.
9. Tydén G, Kumlien G, Genberg H, et al. ABO incompatible kidney transplantations without splenectomy, using antigen-specific immunoadsorption and rituximab
. Am J Transplant
2005; 5: 145.
10. Sonnenday CJ, Warren DS, Cooper M, et al. Plasmapheresis, CMV hyperimmune globulin, and anti-CD20 allow ABO-incompatible
renal transplantation without splenectomy. Am J Transplant
2004; 4: 1315.
11. Becker YT, Becker BN, Pirsch JD, et al. Rituximab
as treatment for refractory kidney transplant rejection. Am J Transplant
2004; 4: 996.
12. Alausa M, Almagro U, Siddiqi N, et al. Refractory acute kidney transplant rejection with CD20 graft infiltrates and successful therapy with rituximab
. Clin Transplant
2005; 19: 137.
13. Tydén G, Genberg H, Tollemar J, et al. A randomized, double-blind, placebo-controlled, study of single-dose rituximab
as induction in renal transplantation. Transplantation
2009; 87: 1325.
14. Kamar N, Milioto O, Puissant-Lubrano B, et al. Incidence and predictive factors for infectious disease after rituximab
therapy in kidney-transplant patients. Am J Transplant
2010; 10: 89.