Changes in cyclosporine A levels in pediatric renal allograft recipients receiving recombinant human growth hormone therapy

Changes in Cyclosporine A Levels in Pediatric Renal Allograft Recipients Receiving Recombinant Human Growth Hormone Therapy C.P. Sanchez, M. Salem, and R.B. Ettenger

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ROWTH retardation remains a prominent feature in children after renal transplantation despite a wellfunctioning graft.1–3 Recombinant human growth hormone (rhGH) has been utilized to augment linear growth in pediatric renal allograft recipients.4 –7 Several studies have demonstrated a considerable increase in growth velocity and an improvement in standard deviation scores for height after twelve months of rhGH therapy; the response, however, declines after the second year of treatment. Several concerns have been raised regarding the safety of rhGH therapy in the pediatric renal transplant population. Acute rejection and graft loss have been reported after initiation of rhGH therapy.8 –10 In mixed lymphocyte cultures, growth hormone potentiates the proliferative and cytotoxic responses and interferon gamma mRNA expression.11 Growth hormone has also been demonstrated to induce glomerular hyperfiltration and may lead to glomerulosclerosis.12 Cyclosporine A (CsA), first identified in 1972, is currently used as one of the primary immunosuppressants in all organ transplantation. Just like rhGH, cyclosporine A is extensively metabolized by the cytochrome P450 enzyme in the liver.13–16 In transplant recipients, the concomitant use of drugs which interfere with the cytochrome P-450 system must be used with caution since stimulation of its activity increases the clearance of CsA leading to low pharmacologic activity. Growth hormone increases the activity and regulates the gene expression of the hepatic cytochrome P450 at the pretranslational level.17 The altered regulation of the hepatic cytochrome P450 system by growth hormone may modify CsA metabolism leading to suboptimal blood levels and possibly contribute to the acute rejection reported in pediatric renal allograft recipients receiving growth hormone therapy.18,19 We, therefore, evaluated the changes in CsA levels in stable growth retarded pediatric renal allograft recipients who were receiving rhGH therapy. METHODS Twenty-one prepubertal growth retarded pediatric kidney allograft recipients who were enrolled in a separate randomized controlled study to assess the skeletal response to growth hormone therapy at the University of California at Los Angeles (UCLA) pediatric renal transplant clinic were evaluated. Out of twenty-one patients, only

sixteen patients received growth hormone therapy for a duration of 12 months. Ten of the sixteen patients were initially randomized to receive rhGH treatment, and the remaining six patients received rhGH therapy after being in the control group for 12 months. Of the sixteen patients who received rhGH therapy for 12 months, eight patients were Caucasian, five patients were of Hispanic origin, two patients were of Asian descent and one patient was African-American. The mean age of the patients was 11 ⫾ 3.6 years (mean ⫾ SD); there were thirteen boys and three girls. At the time of entry into the study, the mean transplant time was 3.9 ⫾ 2.7 years; seven patients underwent living-related transplantation, and nine patients received cadaveric renal allografts. All patients underwent an initial and final bone biopsy as part of the study protocol. Standard deviation scores for height and weight were calculated for each subject every three months. The study was approved by the UCLA Human Subject Protection Committee, and informed consent was obtained from each patient and his/her guardian. As part of the induction therapy, all patients received either a monoclonal or polyclonal anti-T cell therapy and were given triple immunosuppressive medications consisting of prednisone, CsA, and azathioprine or mycophenolate mofetil. All patients were maintained on daily doses of prednisone at an average dose of 0.1 ⫾ 0.04 mg/kg per day. Nine patients were maintained on azathioprine at a mean dose of 1.3 ⫾ 0.2 mg/kg per day, and six patients received mycophenolate mofetil at a mean dose of 33 ⫾ 9.2 mg/kg per day. All patients were receiving CsA (Neoral™) at an average dose of 6.1 ⫾ 3.2 mg/kg per day every 12 hours. Trough CsA levels were obtained monthly to coincide with the patient’s clinic visit, and blood was drawn approximately 12 hours after the last dose. Whole blood CsA levels were determined using the TDx system utilizing a fluorescence polarization immunoassay technique with a monoclonal antibody (Abbot Laboratories, Abbot Park, Ill). Samples of blood were obtained every 3 months until the end of the study period for measurements of total calcium, phosphorus, creatinine, parathyroid hormone, osteocalcin, alkaline phosphatase, insulin-like growth factor, complete blood count,

From the Department of Pediatrics, University of Wisconsin Medical School, Madison, Wisconsin; and Department of Pediatrics, University of California, Los Angeles, CA, USA. Supported in part by MO1-RR00865, DK 56688, Genentech Foundation and by funds from the Casey Lee Ball Foundation. Address reprint requests to Dr C.P. Sanchez, Department of Pediatrics, 3590 MSC/Pediatrics, University of Wisconsin Medical School, 1300 University Avenue, Madison, WI 53706.

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Transplantation Proceedings, 32, 2807–2810 (2000)

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Fig 1. Mean CsA dose (䊐) calculated in mg/kg per day and mean CsA level (■) (ng/mL) in all patients obtained 9 months before the study period and during the 12 months of rhGH therapy, *P ⬍ .05 compared to baseline (0 month). SGOT, and SGPT. Glomerular filtration rate was estimated from serum creatinine values using the Schwartz formula20 every 3 months. None of the patients experienced acute rejection episode for at least 12 months prior to initiation of rhGH treatment. A percutaneous kidney biopsy was performed in patients whose serum creatinine increased by 20% from baseline. Acute rejection episodes were treated using a standardized protocol of intravenous methylprednisolone, 10 mg/kg per dose, for three consecutive days. None of the patients were receiving any other medications which are known to interfere with CsA metabolism. There was no history of skeletal fracture or liver disease in any of the patients. All results are expressed as means ⫾ standard deviation. Comparisons between or among groups were done using the unpaired t-tests, paired t-tests or one-way analysis of variance with contrasts.21

RESULTS

The mean annual growth velocity increased after 12 months of rhGH therapy, 5.7 ⫾ 2.1 cm/year to 8.6 ⫾ 2.9 cm/year, P ⬍ .006. Standard deviation scores for height improved after 3 months of rhGH treatment, 2.3 ⫾ 1.1 and ⫺2.0 ⫾ 1.2, respectively, P ⬍ .003, and continued to improve throughout the study period. When compared to baseline, the standard deviations scores for weight only increased after 9 months of rhGH treatment, ⫺0.4 ⫾ 1.2 vs ⫺0.19 ⫾ 1.3, P ⬍ .01, respectively. The mean CsA levels declined three months after the start of rhGH therapy without significant changes in the mean CsA dose calculated at mg/kg body weight (Fig 1). Although there was an increase in standard deviation scores for weight after 9 months of rhGH therapy, the mean CsA dose remained unchanged during treatment with rhGH (Fig 1). However, mean CsA levels were lower throughout the duration of rhGH therapy when compared to serum levels obtained prior to initiation of rhGH (Fig 1). Serum IGF-I levels increased 3 months after starting rhGH treatment and remained elevated throughout the study period, 377 ⫾ 156 ng/mL at baseline and 515 ⫾ 240 ng/mL at the

end of the study, P ⬍ .05. There were no changes in mean serum calcium, phosphorus, parathyroid hormone, alkaline phosphatase, osteocalcin, 1.25-dihydroxyvitamin D, SGOT, SGPT, total bilirubin, and albumin levels following the initiation of rhGH therapy. In addition, there were no changes in the mean dose of prednisone, azathioprine, and mycophenolate mofetil during the rhGH treatment period. The calculated glomerular filtration rate by Schwartz formula remained unchanged during 12 months of rhGH therapy, 60 ⫾ 19 mL/min/1.73 m2 and 57 ⫾ 17 mL/min/1.73 m2, P ⫽ NS. Two patients experienced acute rejection after 3 and 12 months of rhGH therapy confirmed by kidney biopsy; one rejection was clearly associated with non-compliance to the immunosuppressive medications. The first patient developed acute rejection after 3 months of rhGH therapy. His mean cyclosporine A level for nine months before the study period was 169 ⫾ 8 ng/mL and the mean serum level was 137 ⫾ 10 ng/mL 3 months after starting rhGH therapy, P ⫽ NS. His mean CsA dose before and during rhGH treatment did not differ, 3.4 ⫾ 0.2 and 4.2 ⫾ 0.7 mg/kg per day, respectively, P ⫽ NS. The patient, however, admitted missing doses of prednisone during the study period. In contrast, the second patient developed acute rejection after 12 months of rhGH treatment. His mean cyclosporin A level for 9 months prior to rhGH treatment was 333 ⫾ 19 ng/mL, which declined progressively throughout the study period to a mean level of 125 ⫾ 35 ng/mL at the end of the study, P ⬍ .02. The patient had a mean CsA dose of 5.0 ⫾ 1.4 and 4.0 ⫾ 0.5 mg/kg per day, before and during rhGH therapy, P ⫽ NS. Treatment with methylprednisolone pulse for three days reversed the rejection episodes. DISCUSSION

The current study demonstrates that rhGH therapy increased height velocity in growth impaired pediatric renal

PEDIATRIC RENAL ALLOGRAFT

allograft recipients as previously shown in earlier studies.6,22,23 Although rhGH is an effective treatment for growth retardation in children after kidney transplantation, the safety of rhGH therapy continues to raise concerns. Two of the sixteen patients (12%) in the study had acute rejection episodes which were reversed with corticosteroid therapy. One of the patients, however, admitted to noncompliance to the prednisone therapy. Hokken-Koelega and colleagues reported acute rejection in 11% of their patients who received rhGH; this percentage did not differ from the number of acute rejection experienced in the control group.6 Johansson and colleagues reported that approximately 10% of prepubertal and pubertal patients experienced an acute rise in serum creatinine during rhGH therapy which was attributed either to chronic rejection or biopsy proven acute rejection.24 Growth hormone has been demonstrated to affect the immune system.25 In mixed lymphocyte cultures obtained from normal human volunteers with normal renal function, growth hormone increased proliferation, cytotoxicity, and interferon-␥ mRNA expression.11 When administered to pediatric renal allograft recipients, rhGH increased in vitro proliferative and cytotoxic responses in two of twelve patients and patients with augmented responses had biopsy evidence of chronic rejection and not acute rejection.8 Certainly, growth hormone may play a role in the acute rejection episodes reported in previous studies through immunological mechanisms. On the other hand, growth hormone can potentially interfere with the effects of immunosuppressive agents used to maintain the renal allograft. In the current study, CsA levels declined considerably after starting growth hormone therapy despite maintaining stable doses in milligram per kilogram body weight. These findings suggest that growth hormone therapy may alter the metabolism of CsA by interfering with the cytochrome P450 enzyme system in the liver. As such, close monitoring of CsA levels needs to be done in the follow-up of pediatric patients on rhGH therapy. In the current study, there was a decline in the CsA levels 3 months after starting rhGH therapy despite the same cyclosporine dose. This cannot be attributed to increase in weight gain since standard deviation scores for weight did not increase until after the 9th month of rhGH therapy. Indeed, patients receiving rhGH therapy may require higher doses of CsA to maintain optimal blood levels to prevent acute rejection. Corticosteroids are frequently used in the maintenance of the renal allograft. Growth hormone has been shown in vivo and vitro studies to counteract the catabolic and growth– depressing effects of corticosteroids.26,27 However, corticosteroids, just like CsA, are also extensively metabolized in the liver by the cytochrome P450 enzyme system. Thus, growth hormone may also play a role in the reduction of the immunosuppressive effects of corticosteroids in maintaining the renal allograft. In growth retarded pediatric renal allograft recipients, increasing the dose of cortico-

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steroids may not be beneficial particularly in those children receiving rhGH therapy.28 The calculated glomerular filtration rate (Schwartz formula) did not change after 12 months of rhGH therapy in the current study. Previous studies have shown that rhGH therapy may lead to glomerular hyperfiltration and glomerulosclerosis resulting in the progressive decline of glomerular function.25 Other investigators have reported no differences in the progression of graft deterioration between rhGH treated and control groups.6 Furthermore, measurements of inulin clearance by To ¨nshoff and co-workers in 11 pediatric allograft recipients also did not show any deterioration in glomerular filtration rate.29 Similarly, Johansson and co-workers did not report any considerable changes in the glomerular filtration rate calculated by creatinine clearance.24 The current findings demonstrate that growth hormone therapy does improve growth in stable pediatric renal allograft recipients. Growth hormone therapy, however, should be closely followed since it has the potential to affect T cell function and alter the cytochrome P450 enzyme activity in the liver. Accurate cyclosporine levels should be obtained frequently during the growth hormone therapy. Not only does rhGH increase weight which may decrease the dose of drugs, but it may also directly affect the metabolism of these immunosuppressive drugs. Further controlled studies are needed to assess the efficacy and safety of growth hormone use in growth retarded children after kidney transplantation. REFERENCES 1. Kamil ES, Yadin O, Ettenger RB, et al: Clin Transplant 5:208, 1991 2. Fennell RS, Moles M, Iravani A, et al: Pediatr Nephrol 4:335, 1990 3. So SKS, Chang P-N, Najarian JS, et al: J Pediatr 110:343, 1987 4. Fine RN, Yadin O, Moulton L, et al: J Am Soc Nephrol 2 (Suppl 3):S274, 1992 5. Guest G, Berard E, Crosnier H, et al: Pediatr Nephrol 12:437, 1998 6. Hokken-Koelega ACS, Stijnen T, Ridder MAJd, et al: Lancet 343:1313, 1994 7. Mentser M, Breen TJ, Sullivan K, et al: J Pediatr 131:S20, 1997 8. Benfield MR, Vail A, Waldo FB, et al: Pediatr Nephrol 10:280, 1996 9. Bozzola M, Valtorta A, Moretta A, et al: J Pediatr 117:596, 1990 10. Chavers BM, Doherty L, Nevins TE, et al: Pediatr Nephrol 9:176, 1995 11. Benfield MR, Vail A, Weigent DA: Transplant Proc 26:84, 1994 12. Benfield MR, Parker KL, Waldo FB, et al: Transplantation 55:305, 1993 13. Liddle C, Goodwin BJ, George J, et al: Hepatology 20:185, 1994 14. Redmond GP, Bell JJ, Nichola PS, et al: Pediatr Pharmacol 1:63, 1980 15. Brunner LJ, Bennet WM, Koop DR: Kidney Int 54:216, 1998

2810 16. Ptachcinski RJ, Venkataramanan R, Rosenthal JT, et al: Clin Pharmacology 38:296, 1985 17. Liddle C, Goodwin BJ, George J, et al: J Clin Endocrinol Metab 83:2411, 1998 18. Levitsky LL, Schoeller DA, Lambert GH, et al: Dev Pharmacol Ther 12:90, 1989 19. Cheung NW, Liddle C, Coverdale S, et al: J Clin Endocrinol Metab 81:1999, 1996 20. Schwartz GJ, Brion LP, Spitzer A: Pediatr Clin North America 34:571, 1987 21. Dawson-Saunders B, Trapp RG. Basic and clinical biostatistics. Norwalk, Connecticut: Appleton & Lange, 1990. 22. Fine RN, Yadin O, Nelson PA, et al: Pediatr Nephrol 5:147, 1991

SANCHEZ, SALEM, AND ETTENGER 23. Mehls O, Wuhl E, Haffner D, et al: Nephrol Dial Transplant 11:1747, 1996 24. Johansson G, Sietnieks A, Janssens F, et al: Acta Pediatr Scan [Suppl] 370:36, 1990 25. Kelley KW: Ann NY Acad Sci 594:95, 1990 26. Allen DB, Goldberg BD: Pediatrics 89:416, 1992 27. Kovacs G, Fine RN, Worgall S, et al: Kidney Int 40:1032, 1991 28. Ingulli E, Singh A, Moazami S, Tejani A: Kidney Int 44 (Suppl 43):S65, 1993 29. Tonshoff B, Tonshoff C, Mehls O, et al: Eur J Pediatr 151:601, 1992