Background Tacrolimus is used off-label in the treatment of Henoch-Schönlein purpura nephritis (HSPN) in children, with limited evidence-based data. Based on clinical empirical experience and mechanism of action, tacrolimus might be promoted as treatment for childhood HSPN. The objectives of this pilot study were to assess its effectiveness and safety, and to explore the potential impact of CYP3A5 genotype.
Methods Children with HSPN receiving tacrolimus as empirical treatment were included in this prospective, observational study. Effectiveness was classified as complete remission, partial remission or non-response. General safety data analyses during and after study drug exposure included adverse events, reasons for discontinuation, deaths, laboratory data and vital signs. Trough concentration was determined using high-performance liquid chromatography with tandem mass spectrometry. Pharmacogenetic analysis was performed on the CYP3A5 gene.
Results A total of 20 patients with a mean age of 7.5 (SD 2.1) years participated in the whole process of the study. Twelve patients reached complete remission and eight patients reached partial remission at the end of 6-month treatment. No patients discontinued tacrolimus treatment due to adverse events, and no drug-related adverse events were shown to have a causal association with tacrolimus therapy. Dose-adjusted trough concentration was significantly higher in children with CYP3A5*1 allele as compared with patients with CYP3A5*3/*3 genotype (170.7±100.9 vs 79.8±47.4 (ng/mL)/(mg/kg)).
Conclusion This pilot study showed that tacrolimus might be an effective and well-tolerated drug for the treatment of HSPN in children. CYP3A5 polymorphism had a significant impact on tacrolimus concentration.
- henoch-schönlein purpura nephritis
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What is already known on this topic?
Tacrolimus is used off-label in the treatment of Henoch-Schönlein purpura nephritis in children.
CYP3A5 polymorphism significantly affected tacrolimus pharmacokinetics in solid organ transplantation patients .
What this study adds?
Tacrolimus might be an effective and well-tolerated drug for the treatment of HSPN in children.
CYP3A5 polymorphism has a significant impact on tacrolimus concentration in children with HSPN.
The choice of tacrolimus dose should be considered based on CYP3A5 genotype.
Henoch-Schönlein purpura (HSP), an IgA-mediated systemic small-vessel vasculitis, is the most common vasculitis in children, with an incidence of approximately 10:100 000 children and a slight male predominance (male to female ratio of 1.5:1).1 Henoch-Schönlein purpura nephritis (HSPN) is the principal cause of morbidity for HSP, and 1%–7% of patients with HSPN may progress to renal failure or end-stage renal disease.2 It represents 10%–15% of paediatric glomerulonephritis.3
Immunosuppressive therapy has become the standard treatment in children with HSPN; however, the use of these drugs is still mainly in an off-label manner in clinical practice.4–10 According to the Chinese guideline for the diagnosis and treatment of HSPN (2016), glucocorticoid combination with the calcineurin inhibitor is recommended to treat HSPN with nephrotic proteinuria, nephrotic syndrome, acute nephritic syndrome, or IIIb and IV pathological grade. In vitro tests already showed that the immunosuppressive capacity of tacrolimus is 100 times stronger than ciclosporin A, and tacrolimus had less renal toxicity than ciclosporin A. Although tacrolimus has been empirically used in the treatment of HSPN in children,11 12 the evidence-based clinical data are still limited. The pivotal tacrolimus mechanism of action involves inhibition of the redistribution of calcineurin at the slit diaphragm,13 which could potentially ameliorate proteinuria. In addition, CYP3A5 is the predominant enzyme for metabolism of tacrolimus.14 15 CYP3A5 gene polymorphism significantly affected tacrolimus pharmacokinetic and clinical outcomes in solid organ transplantation patients.16–21 No data were available in children with HSPN.
Given the potential benefits and unmet need in clinical practice, the purposes of this pilot study were to assess the effectiveness and safety of tacrolimus in children with HSPN and to evaluate the potential impact of CYP3A5.
Materials and methods
This trial was a prospective, open-label study of tacrolimus, conducted at the Department of Paediatric Nephrology in the Children’s Hospital of Hebei Province, Shijiazhuang, China. The inclusion criteria included children with HSPN with nephrotic proteinuria, nephrotic syndrome, acute nephritic syndrome, or IIIb and IV pathological grade aged less than 18 years; and receiving tacrolimus as initial immunosuppressive therapy. The exclusion criteria were children who received other immunosuppressive drug before the trial or other systemic trial drug therapy; and with a concomitant medical condition, whose participation, in the opinion of the investigator and/or medical advisor, may create an unacceptable additional risk. This trial was registered at www.clinicaltrials.gov with ID number NCT03222687.
Children with HSPN who were treated with tacrolimus from September 2015 to May 2017 were included. Informed consent forms, signed by the patients’ parents or guardians, were obtained prior to commencing tacrolimus therapy.
Renal biopsies were performed on all patients and were graded according to the International Study of Kidney Diseases in Children (ISKDC) classification: grade I, minimal alterations; grade II, mesangial proliferation; grade IIIa, focal or IIIb, diffuse proliferation or sclerosis with <50% crescents; grade IVa, focal or IVb, diffuse mesangial proliferation or sclerosis with 50%–75% crescents; grade Va, focal or Vb, diffuse mesangial proliferation or sclerosis with >75% crescents; and grade VI, membranoproliferative-like lesion.
Immunosuppressive therapy included tacrolimus and prednisone. Tacrolimus treatment was initiated at a dosage of 0.05–0.1 mg/kg/day twice daily and used for at least 6 months. Prednisone was started at 2 mg/kg/day and tapered off gradually after initiation of treatment. Tacrolimus blood trough samples were collected at steady-state condition. DNA is extracted from these blood samples. Assessment of proteinuria was performed using the standard 24-hour urine protein test. During the first 4 months, the patients were followed up every 2 weeks and then followed up once a month. All data from HSPN onset until the end of the follow-up were analysed.
Effective and safety analyses
Complete remission was defined if clinical symptoms and signs disappeared and proteinuria was less than 4 mg/hour/m2 within 6 months.22 Partial remission was defined if proteinuria was reduced to 4.1–40 mg/hour/m2 within 6 months. A non-responsive patient was defined if there was no improvement in clinical symptoms or signs 6 months after the therapy of tacrolimus with or without prednisone, or urinary protein remained more than 40 mg/hour/m2.
Clinical and biological parameters (renal and liver function parameters and haematology parameters) were monitored using routine clinical practice. General safety data analyses during and after study drug exposure included adverse events, reasons for discontinuation, deaths, laboratory data and vital signs. The WHO-Uppsala Moniroting Centre system was used for adverse events causality assessment. In particular, nephrotoxicity, as indicated by the twofold increase of creatinine or an increase by at least 0.6 mg/dL from the start and any time during tacrolimus therapy, was evaluated.
Analytical methods and genotyping
Tacrolimus trough blood concentrations were determined using high-performance liquid chromatography with tandem mass spectrometry. The calibration curve ranged from 1.0 to 100 ng/mL. The interday and intraday coefficients of variation of controls were <7.2% and 4.4%, respectively. The lower limit of quantification was 1.0 ng/mL.
Total genomic DNA was extracted from blood samples using a TIANamp Blood Clot DNA Kit (TIANGEN Biotech, Beijing) according to the manufacturer’s protocol. CYP3A5 A6986G polymorphism was determined using the TaqMan allelic discrimination technique with 3’-minor groove binding quencher probes on a Bio-Rad Fluorescence Quantitative PCR according to a standardised protocol.
Continuous data were summarised by mean±SD or median (range), and categorical data were summarised as the count of each category. The differences between subgroup were tested with the t-test. A P value <0.05 was considered statistically significant. All analyses were performed using the SPSS V.16.0 statistics software.
Twenty-five patients (18 boys, 7 girls) were initially included. All of the patients fulfilled the inclusion and exclusion criteria. Five patients were transferred to other hospitals and were unable to attend for evaluation at 6 months at our centre. They were therefore excluded from the final analysis. Thirteen patients had trough concentration blood and DNA samples for concentration and CYP3A5 genotype analysis. The trial flow chart is presented in figure 1.
Finally, 20 children with a mean age of 7.5 (SD 2.1) years were included in the effectiveness and safety analyses. According to the ISKDC classification, 3 patients were categorised as grade IIb, 2 as grade IIIa, 16 as grade IIIb and 1 as grade IV. The characteristics of the 20 children are presented in table 1. The mean follow-up was 11.2 (SD 3.7) months.
Effectiveness and safety
The mean proteinuria of 20 children was 92.1 (SD 59.0) mg/hour/m2 before starting the treatment. At the end of the 6-month treatment, the mean proteinuria was reduced to 4.2 (SD 2.8) mg/hour/m2 (P<0.001). The complete and partial remission rates were 60% and 40%, respectively.
A mild transient elevation of alanine transaminase was observed in two patients and was back to normal value during tacrolimus therapy. All of these patients have no history of hepatitis and their hepatitis antibodies tests were negative. Gastrointestinal symptoms were observed in three patients and resolved after symptomatic treatment. Tacrolimus-related nephrotoxicity was not observed in any patient. No patients discontinued the tacrolimus treatment due to adverse events, and no drug-related adverse events were shown to have a causal association with tacrolimus therapy.
Concentrations and CYP3A5 genotype
The mean tacrolimus trough concentration in 13 patients was 4.0 (SD 2.7) ng/mL. Eight patients had CYP3A5*1/*3 genotype and five patients with CYP3A5*3/*3 genotype. Patients’ characteristics and clinical outcomes according to CYP3A5 polymorphism are shown in table 2.
Dose-adjusted trough concentration (C0/dose) was significantly higher in children with CYP3A5*1 allele as compared with patients with CYP3A5*3/*3 genotype (170.7±100.9 vs 79.8±47.4 (ng/mL)/(mg/kg)) (P=0.048). There was no significant difference in clinical outcomes between the two subgroups within 6 months (table 2).
Our study for the first time evaluated the effectiveness and safety of tacrolimus in children with HSPN. To date, there is no consensus on optimal immunosuppressive therapy in patients with HSPN. Our previous experience and previously published studies have shown that steroid alone did not yield good results.23–25 Remission was achieved slowly and only in 53% of patients with methylprednisolone.26 For other immunosuppressive drugs, mycophenolate mofetil (MMF) showed the mean duration for return to negative proteinuria was 11±1.7 months.27 The remission rate in patients with HSPN treated with MMF and prednisone group was 72.7% at 6 months.28 In a retrospective data study of 141 children with HSPN with corticosteroid or immunosuppressive treatments (cyclophosphamide or azathioprine), 35% showed complete remission after 6 months.29 In our present study, after receiving 6 months of treatment of tacrolimus, the complete and partial remission rates were 60% and 40%, respectively. Although it is not possible to compare directly our results with previously published studies, our study showed significant improvement in effectiveness.
Tacrolimus is indicated for prophylaxis or treatment of graft rejection after transplantation. In fact, off-label use of tacrolimus is a common issue in paediatric clinical practice. Some pilots studies have already reported the effectiveness of tacrolimus in children with focal segmental glomerulosclerosis, lupus nephritis, nephrotic syndrome30–33 as well as IgA nephropathy.34–36 The pathological manifestations of HSPN are the same as those of IgA nephropathy. Tacrolimus can inhibit mediator release from basophils and mast cells.37 The major adverse events of tacrolimus are nephrotoxicity, neurotoxicity, post-transplant diabetes mellitus and malignant complications. In our study, tacrolimus was well-tolerated and there was no case of major adverse events. The long-term safety data still need to be evaluated in further studies.
Choosing the right dose ‘adapted to each individual patient’ is a central question for immunosuppressive therapy. In adult kidney transplant recipients, it has been shown that CYP3A5 genotype-based dosing regimen resulted in a more rapid achievement of target tacrolimus trough concentrations with less dose adjustments than the universal 0.1 mg/kg twice-daily dosing regimen.38 In our previous study on paediatric kidney transplant patients, a visual patient-tailored dosing chart, taking into consideration the child’s weight, recent haematocrit level and CYP3A5 genotype, was also developed.39 40 In the current study, CYP3A5 genotype also shows significant impact on C0/dose. C0/dose in the CYP3A5*1/*3 subgroup was significantly lower than that in the CYP3A5*3/*3 subgroup (P=0.048). However, CYP3A5 polymorphism has no significant effect in the short-term response to tacrolimus, which can be explained mainly by the limited number of patients. Further studies with larger, randomised patient series and longer follow-up are warranted. After standard treatment, large interindividual variability in C0 has been demonstrated. Dosage adaptation based on therapeutic drug monitoring (TDM) of tacrolimus is recommended in both adult and paediatric organ transplant recipients in order to keep the blood concentration of individual patient within the target range,40 and the clinical benefit of routine TDM in children with HSPN still needs to be confirmed.
Our study had some limitations. No control group was designed in our trial. This is mainly because no standard treatment was available in children with HSPN. In addition, proteinuria was decreased rapidly after initialising tacrolimus treatment, and we did not perform repeat renal biopsies to confirm the efficacy of tacrolimus.
This pilot study showed that tacrolimus might be an effective and well-tolerated drug for the treatment of HSPN in children. CYP3A5 polymorphism had a significant impact on tacrolimus concentration. The CYP3A5 genotyping was recommended to personalise tacrolimus starting dose. In future perspective, a controlled study is warranted to define the optimal dose and duration of tacrolimus therapy in childhood HSPN, and the clinical benefits of CYP3A5 genotyping still need to be confirmed.
D-FZ and G-XH contributed equally.
Contributors D-FZ and G-XH retrieved the data, carried out the initial analyses and drafted the initial manuscript. C-ZL, Y-JY, F-JL, LL, X-YY, R-HL and LD collected samples, followed up patients and recorded patient information. QD sorted out the samples. EJ-A gave advice on the project and manuscript. WZ conceptualised, designed and initiated the study.
Funding This work is supported by the National Natural Science Foundation of China (81503163), National Science and Technology Major Projects for ‘Major New Drugs Innovation and Development' (2017ZX09304029-002), and Young Taishan Scholars Program and Young Scholars Program of Shandong University.
Competing interests None declared.
Patient consent Parental/guardian consent obtained.
Ethics approval This study was approved by the institutional ethics board.
Provenance and peer review Not commissioned; externally peer reviewed.
Data sharing statement Data is available for sharing.
Presented at Preliminary results of this study were presented at the 16th meeting of the European Society for Developmental Perinatal and Paediatric Pharmacology Congress, 2017, Leuven, Belgium.
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