Pretransplant Soluble CD30 Serum Concentration Does Not Affect Kidney Graft Outcomes 3 Years After Transplantation

Pretransplant Soluble CD30 Serum Concentration Does Not Affect Kidney Graft Outcomes 3 Years After Transplantation J. Kovac˘, M. Arnol, B. Vidan Jeras, A.F. Bren, and A. Kandus ABSTRACT Introduction. An elevated serum concentration of soluble the form of CD30 (sCD30), an activation marker of mainly TH2-type cytokines producing T lymphocytes, has been reported as a predictive factor for acute cellular rejection episodes and poor graft outcomes in kidney transplantation. This historic cohort study investigated the association of a pretransplant sCD30 serum concentrations with kidney graft function and graft survival 3 years posttransplantation in adult recipients of deceased donor kidney grafts, treated with monoclonal anti-CD25 antibodies as an induction treatment combined with a cyclosporine (CsA)-based maintenance triple therapy. Materials and Methods. The pretransplant sera of 296 recipients were tested for sCD30 content using a microsphere flow-cytometry assay. The estimated glomerular filtration rate (eGFR) was determined by the 4-variable Modification of Diet in Renal Disease equation. The incidences of graft loss were calculated with the use of Kaplan-Meier survival analysis and compared using the log-rank test. Results. According to the distribution of the pretransplant sCD30 levels concentration ⱖ2700 pg/mL was defined as high (n ⫽ 146) and concentration ⬍2700 pg/mL as low (n ⫽ 150). Three years posttransplantation, the eGFR was not significantly different in the recipients in high and low sCD30 groups (65 ⫾ 24 vs 67 ⫾ 21 mL/min/1.73 m2; P ⫽ .43); there was no association between the eGFR 3 years after transplantation and the pretransplant sCD30 levels (r2 ⫽ 0.002; P ⫽ .49). Graft survival 3 years after transplantation was also not different in the recipients in high and low sCD30 groups (P ⫽ .52). Conclusion. In our adult deceased-donor kidney graft recipients, the pretransplant sCD30 serum concentration was not a predictive factor of immunologic risk associated with the kidney graft function 3 years posttransplantation; neither did it affect graft survival 3 years after transplantation. The immunosuppression with anti-CD25 antibodies as an induction treatment combined with the CsA-based maintenance triple therapy could possibly be decisive for our findings. D30 SURFACE ANTIGEN is an activation marker of the subset of mainly TH2-type cytokines producing T lymphocytes. It is a 120-kD membrane glycoprotein, and a member of the tumor necrosis factor receptor superfamily. An 85-kD soluble form of CD30 molecule (sCD30) is present in serum and is released by CD30 positive cells. The serum concentration of sCD30 correlates with the expression of the CD30 surface antigen on activated T lymphocytes.1– 4 In kidney transplantation, an elevated serum concentration of sCD30 has been reported as a predictive

C

factor for acute rejection episodes (ARE) and poor graft outcomes.5–13 From the Department of Nephrology (J.K., A.F.B., A.K.), University Medical Center, and the Tissue Typing Center (M.A., B.V.J.), Blood Transfusion Centre of Slovenia, Ljubljana, Slovenia. Address reprint requests to Janko Kovac˘, MD, PhD, Department of Nephrology, University Medical Center Ljubljana, Zalos˘ka cesta 7, 1525 Ljubljana, Slovenia. E-mail: [email protected]

© 2010 by Elsevier Inc. All rights reserved. 360 Park Avenue South, New York, NY 10010-1710

0041-1345/–see front matter doi:10.1016/j.transproceed.2010.09.050

Transplantation Proceedings, 42, 4043– 4046 (2010)

4043

KOVAC˘, ARNOL, VIDAN-JERAS ET AL

4044

This historical cohort study investigated the association of a pretransplant sCD30 serum concentration with kidney graft function and graft survival in adult recipients of deceased donor kidney transplants 3 years thereafter. It was hypothesized that the pretransplant sCD30 serum concentration in kidney transplant recipients treated with current immunosuppression with anti-CD25 antibodies as an induction treatment combined with the cyclosporine (CsA)-based maintenance triple therapy was not a predictive factor of immunologic risk associated with the kidney graft function and survival 3 years after transplantation. PATIENTS AND METHODS From January 2000 through December 2006, 316 adult patients received kidney grafts from deceased donors at our medical center. Among these recipients, 20 (6.3%) were excluded from the analysis because there was no pretransplant sera available (n ⫽ 19) or transplantectomy was performed due to postmortem detection of a metastatic tumor in the donor (n ⫽ 1). A total of 296 patients (93.7%) were included in the study. Clinical data related to kidney transplant recipients and donors were retrieved from our clinical records and our Kidney Transplantation Database. The immunosuppressive regimen in all recipients consisted of a triple maintenance therapy with CsA microemulsion (CsA-Neoral), mycophenolate mofetil, and methylprednisolone, combined with monoclonal anti-CD25 antibodies (basiliximab or daclizumab) as an induction therapy.14,15 A continuous intravenous infusion of CsA (0.08 mg/kg/h) was started just before transplantation and was continued by CsA-Neoral, 3 mg/kg twice daily beginning 2 days after transplantation. Methylprednisolone at a dose of 0.4 mg/kg was infused intravenously before transplantation and oral mycophenolate mofetil was started on posttransplant day 1. The dose of CsA-Neoral was adjusted to maintain CsA blood trough level of 100 –170 ng/mL during the first 3 months and 70 –130 ng/mL thereafter. Oral methylprednisolone was tapered by 4 mg/wk to achieve a maintenance dose of 0.08 mg/kg per day. Pretransplantation, 20 mg of basiliximab were infused intravenously and on day 4 posttransplantation, or daclizumab was infused intravenously in a dose of 1 mg/kg immediately before and at the end of weeks 2, 4, 6, and 8 posttransplantation. ARE were always confirmed with kidney biopsy. Delayed graft function (DGF) was defined as the need for dialysis during the first week after transplantation. Kidney graft function was estimated by the 4-variable Modification of Diet in Renal Disease study equation and expressed as the estimated glomerular filtration rate (eGFR) in milliliters per minute per 1.73 m2 at 1 and 3 years after transplantation.16,17 Routine panel reactive antibody reactivity assays and a lymphocyte cross-match were performed immediately before transplantation using a standard complement-dependent lymphocytotoxic assay.18 A negative lymphocyte cross-match assay was mandatory for all transplants. Concentrations of sCD30 were measured in the pretransplant sera, collected for the lymphocyte cross-match assays, and stored at ⫺20°C. Serum sCD30 concentration measurements were performed with a commercially available microsphere flow-cytometry assay (CD30 [soluble], Singleplex Bead Kit, Invitrogen, Carlsbad, Calif). For validation of the assay, sCD30 serum concentrations were measured in sera of 12 local control subjects (age 30 –54 years) with normal kidney function (range, 864 – 6847 pg/mL;

median, 864 pg/mL; interquartile range, 864 –3336 pg/mL). With this method, the lowest detectable concentration was 864 pg/mL. Unless otherwise stated, data are presented as mean values with standard deviations (SD) or medians with interquartile ranges. Differences in the distributions of continuous variables were evaluated with the unpaired Student’s t test or the Kruskal–Wallis test for normally and nonnormally distributed data, respectively. Chisquare statistics were used to evaluate cross-tabulations of categorical variables. The association between eGFR 3 years after transplantation and pretransplant serum concentration of sCD30 was assessed with a least-squares linear regression model. The incidences of graft loss were calculated with the use of Kaplan-Meier survival analysis and compared with the log-rank test. P ⬍ .05 was considered significant. All statistics were performed using SPSS, version 16.0 (SPSS Inc., Chicago, Ill). The study was approved by the ethical committee at our institution, and informed consent was obtained from the patients.

RESULTS

In 296 recipients (174 men [age 47 ⫾ 11 years] and 122 women [age 46 ⫾ 10 years]) included in our study the median pretransplant sCD30 serum concentration was 2672 pg/mL (range, 864 –94,536 pg/mL; interquartile range, 864 – 5257 pg/mL). According to the distribution of pretransplant sCD30 serum concentrations, a concentration ⱖ2700 pg/mL was defined as high (n ⫽ 146), and ⬍2700 pg/mL as low (n ⫽ 150). Patients were divided into low and the high pretransplant sCD30 serum concentration groups. The groups did not differ with respect to majority of transplantation-related clinical characteristics, such as recipient gender, donor age, number of retransplants, degree of human leukocyte antigen mismatches, and panel reactive antibody reactivity (Table 1). However, patients in the high sCD30 group were significantly younger (46 ⫾ 11 vs 48 ⫾ 9 years; P ⫽ .020) and the cold ischemia time was slightly longer in this group of patients (22.0 ⫾ 6.8 vs 20.3 ⫾ 6.9 hours; P ⫽ .031; Table 1). There were no significant differences in the incidences of DGF and ARE between the high and low sCD30 groups (Table 2). The eGFR 3 years after transplantation was not Table 1. Transplantation-Related Clinical Characteristics According to Pretransplant Soluble CD30 (sCD30) Serum Concentrations* sCD30 (pg/mL) (n ⫽ 296) Clinical Characteristics

Recipient age (y) Recipient gender (female:male) Donor age (y) Graft number (1st:2nd) Cold ischemia time (hr) HLA-A, -B, and -DR mismatches PRA, last pretransplant (%)

⬍2700 (n ⫽ 150)

ⱖ2700 (n ⫽ 146)

P

48 ⫾ 9 67:83

46 ⫾ 11 55:91

.020 .239

39 ⫾ 14 136:14 20.3 ⫾ 6.9 2.8 ⫾ 1.0

37 ⫾ 14 135:11 22.0 ⫾ 6.8 2.6 ⫾ 1.1

.677 .677 .031 .106

6 ⫾ 14

6 ⫾ 11

.938

Abbreviations: HLA, human leukocyte antigen; PRA, panel reactive antibody. *Patients were divided into 2 groups according to distribution of the pretransplant sCD30. Data are total numbers of mean values ⫾ SD.

SOLUBLE CD30 SERUM CONCENTRATION

4045

Table 2. Delayed Graft Function, Incidence of Acute Rejection Episodes, Kidney Graft Function (eGFR), and Graft and Recipient Status at 3 Years According to Pretransplant Soluble CD30 (sCD30) Serum Concentrations* sCD30 (pg/mL) (n ⫽ 296) Clinical Characteristics

DGF (yes:no) Acute rejection (yes:no) eGFR at 3 y (mL/min per 1.73m2) Graft status at 3 y (nonfunctioning:functioning) Recipient status at 3 y (dead:alive)

⬍2700 (n ⫽ 150)

ⱖ2700 (n ⫽ 146)

P

38:112 12:138 67 ⫾ 21

36:110 15:131 65 ⫾ 24

.890 .549 .268

22:128

18:128

.612

14:136

12:134

.838

Abbreviations: eGFR, estimated glomerular filtration rate (MDRD equation; see text). *Patients were divided into 2 groups according to distribution of the pretransplant sCD30. Data are total numbers of mean values ⫾ SD.

different in patients in the high or low pretransplant sCD30 concentration groups. There were also no differences in the incidences of graft loss and patient death 3 years after transplantation (Table 2). Univariate linear regression analysis showed no correlation between pretransplant sCD30 serum concentrations and eGFR 3 years (r2 ⫽ 0.002; P ⫽ .47) after transplantation. Grafts survival rates 3 years after transplantation were not different in the recipients in high and low sCD30 groups (95.2% vs 90.0%; P ⫽ .52). DISCUSSION

The results of this study demonstrate that kidney graft function and graft survival 3 years after transplantation were not associated with pretransplant sCD30 serum concentration. This suggests that sCD30 serum concentration was not predictor of immunologic risk in our patient cohort. Some previous studies have suggested the pretransplant sCD30 serum concentration is a potential immunologic marker for posttransplant risk estimation in kidney graft recipients.5–13 In those studies, the elevated pretransplant sCD30 serum concentration was considered as a prognostic factor for the subsequent frequency and grade of ARE, and was associated with poorer graft survival. In this historic cohort study, we analyzed the association of pretransplant sCD30 serum concentrations with kidney graft function and graft survival 3 years posttransplantation in the recipients of deceased donor kidneys, all treated with a modern and effective immunosuppressive regimen. Kidney graft function was estimated using the Modification of Diet in Renal Disease equation, which is a better surrogate for estimating kidney graft function than serum creatinine concentration alone.19,20 As described by other authors,5,21 the serum sCD30 concentrations in our study patient population were significantly higher than in the healthy controls. However, absolute concentrations of the pretransplant sCD30 serum

concentration in our study cannot be compared with other studies. The reason for this is a novel flow cytometry-based methodology used to determine sCD30 serum concentration, whereas in all previous studies enzyme-linked immunosorbent assay was used. The high and low sCD30 patient groups were similar with respect to majority recipient clinical characteristics and their immunologic risk. Patients in the high sCD30 group were, however, slightly younger. This observation was also described by other investigators.5,6,11,21 Furthermore, we did not notice any significant differences between patient groups in the incidences of posttransplant DGF and ARE, which have all been previously recognized to significantly affect graft function and survival.22,23 We could speculate that similar incidences of DGF and ARE were related to similar clinical characteristics of the patients in both groups. In this study, we could not demonstrate any difference among groups in graft function 3 years after transplantation. Even more important, graft survival 3 years after transplantation was also similar in both patient groups. We therefore believe that the absence of an association between the sCD30 and graft function implies that the pretransplant sCD30 is not an important immunologic parameter affecting posttransplant deceased donor kidney graft function in our patient cohort. The majority of investigators linked detrimental effects of higher pretransplant sCD30 serum concentrations on kidney graft outcome with the increased incidence of an early ARE.6,9,10,13 With the immunosuppression regimen mentioned, we observed a low incidence of ARE and there was no association between the pretransplant sCD30 serum concentration and the incidence of ARE. Patients with ARE were also evenly distributed between the high and low sCD30 concentration groups, which further supports the lack of a potential detrimental effect of higher sCD30 serum concentration in our patients. In conclusion, this study shows that the pretransplant sCD30 serum concentration in our adult recipients of deceased donor kidney transplants is not a predictive factor of immunologic risk associated with the kidney graft function 3 years after transplantation; neither does it affect kidney graft survival. The immunosuppression with antiCD25 antibodies as an induction treatment combined with the CsA based maintenance triple therapy could possibly be decisive for our findings. REFERENCES 1. Josimovic-Alasevic O, Durkop H, Schwarting R, et al: Ki-1 (CD30) antigen is released by Ki-1 positive tumor-cells in vitro and in vivo. I. Partial characterization of soluble Ki-1 antigen and detection of the antigen in cell culture supernatants and in serum by an enzyme- linked immunosorbent assay. Eur J Immunol 19:157, 1989 2. Del Prete G, De Carli M, D’Elios MM, et al: CD30-mediated signaling promotes the development of human T helper type 2-like T cells. J Exp Med 182:1655, 1995 3. Del Prete G, De Carli M, Almerigogna F, et al: Preferential expression of CD30 by human CD4⫹ T cells producing Th2-type cytokines. FASEB J 9:81, 1995

4046 4. Romagnani S, Parronchi P, D’Elios MM, et al: An update on human Th1 and Th2 cells. Int Arch Allergy Immunol 113:153, 1997 5. Pelzl S, Opelz G, Wieser M, et al: Soluble CD30 as a predictor of kidney graft outcome. Transplantation 73:3, 2002 6. Süsal C, Pelzl S, Dohler B, et al: Identification of highly responsive kidney transplant recipients using pretransplant soluble CD30. J Am Soc Nephrol 13:1650, 2002 7. Süsal C, Pelzl S, Opelz G: Strong human leukocyte antigen matching effect in nonsensitized kidney recipients with high pretransplant soluble CD30. Transplantation 76:1231, 2003 8. Süsal C, Pelzl S, Simon T, et al: Advances in pre- and posttransplant immunologic testing in kidney transplantation. Transplant Proc 36:29, 2004 9. Cinti P, Pretagostini R, Arpino A, et al: Evaluation of pretransplant immunologic status in kidney-transplant recipients by panel reactive antibody and soluble CD30 determinations. Transplantation 79:599, 2005 10. Ravindra R, Naina J, Mira V, et al: High pre-transplant soluble CD30 levels are predictive of the grade of rejection. Am J Transplant 5:1922, 2005 11. Kim MS, Kim HJ, Kim SI, et al: Pretransplant soluble CD30 level has limited effect on acute rejection, but affects graft function in living donor kidney transplantation. Transplantation 82:1602, 2006 12. Vaidya S, Parlow D, Barnes T, et al: Pretransplant soluble CD30 is a better predictor of posttransplant development of donor-specific antibodies and acute vascular rejection than panel reactive antibodies. Transplantation 82:1606, 2006 13. Sengul S, Keven K, Gormez U, et al: Identification of patients at risk of acute rejection by pretransplantation and posttransplantation monitoring of soluble cd30 levels in kidney transplantation. Transplantation 81:1216, 2006

KOVAC˘, ARNOL, VIDAN-JERAS ET AL 14. Kandus A, Grego K, Arnol M, et al: Effective immunoprophylaxis with basiliximab plus triple therapy in renal transplantation: five-year single-center experience. Transplant Proc 38:2853, 2006 15. Kandus A, Grego K, Bren AF: Prevention of early acute rejection with daclizumab and triple immunosuppression in cadaveric renal allograft recipients. Ther Apher Dial 9:262, 2005 16. Pöge U, Gerhardt T, Palmedo H, et al: MDRD equations for estimation of GFR in renal transplant patients. Am J Transplant 5:1306, 2005 17. Poggio ED, Wang X, Weinstein DM, et al: Assessing glomerular filtration rate by estimation equations in kidney transplant recipients. Am J Transplant 6:100, 2006 18. Terasaki PI, McCleland J: Microdroplet assay of human serum cytotoxins. Nature 204:998, 1964 19. Kasiske BL, Vazquez MA, Harmon WE, et al: Recommendations for the outpatient surveillance of renal transplant recipients. J Am Soc Nephrol 11(suppl 15):S1, 2000 20. National Kidney Foundation: K/DOQI clinical practice guidelines for chronic kidney disease: evaluation, classification, and stratification. Kidney Disease Outcome Quality Initiative. Am J Kidney Dis 39(2 suppl 1):S1, 2002 21. Heinemann FM, Rebmann V, Witzke O, et al: Association of elevated pretransplant sCD30 levels with graft loss in 206 patients treated with modern immunosuppressive therapies after renal transplantation. Transplantation 83:706, 2007 22. Womer KL, Vella JP, Sayegh MH: Chronic allograft dysfunction: mechanisms and new approaches to therapy. Semin Nephrol 20:126, 2000 23. Ojo AO, Wolfe RA, Held PJ, et al: Delayed graft function: risk factors and implications for renal allograft survival. Transplantation 63:968, 1997