dependent idiopathic focal glomerulosclerosis

Nephrol Dial Transplant (2002) 17: 655–662

Preliminary Communication

Combined therapy of tacrolimus and corticosteroids in cyclosporin-resistant or -dependent idiopathic focal glomerulosclerosis: a preliminary uncontrolled study with prospective follow-upy Alfons Segarra1, Josefa Vila1, Leonor Pou2, Joaquim Majo´3, Antonia Arbo´s4, Teresa Quiles4 and Luis L. Piera1 1

Servicio de Nefrologia, 2Bioquı´mica, 3Anatomia Patolo´gica and 4CIBBIM, Hospital General Valle Hebroń, Barcelona, Spain

Abstract Background. Cyclosporin has improved the outcome for steroid-resistant patients with focal glomerulosclerosis, but there is a proportion of patients that are either cyclosporin-resistant or suffer relapses, needing long-term therapy to sustain the remission. In these cases, preliminary reports suggest that tacrolimus could be an alternative therapy, but to date the evidence is limited to small series of patients with no long-term follow-up. Methods. In this study we analysed the efficacy and safety of a combined therapy of tacrolimus and steroids in 25 patients (mean serum creatinine s 1.24"0.49 mgudl; mean proteinuria s 10.2"9.5 guday; mean serum albumin s2.4"0.58 gudl) with idiopathic primary focal glomerulosclerosis and proven resistance to or dependence on cyclosporin A. Results. After a 6 months trial of tacrolimus and steroids, proteinuria decreased in 17 patients (68%) (complete remission in 10 patients (40%), partial remission in two patients (8%) and a moderate reduction in proteinuria to levels -3 guday was seen in five additional patients (20%)). The only predictor of response to tacrolimus was a previous response to cyclosporin and prednisone, either as a complete or partial remission (remission rate 75% vs 15.3; P s0.036). Mean time to remission was 112"24 days. After tacrolimus discontinuation, 13u17 patients (76%) relapsed and were treated with a second trial of tacrolimus for 1 year, achieving complete remission in five patients (38.4%), partial remission in four patients (30.7%) and reduction of proteinuria -3 guday in four patients (30.7%). After 2 years of follow-up, Correspondence and offprint requests to: A. Segarra, Servicio de Nefrologia, Anexo planta 7a, Hospital Valle Hebroń, Paseo Valle Hebroń 119–129, E-08035 Barcelona, Spain. Email: [email protected] y This work was presented at the EDTA Congress of Nice September 2000 and has been published in abstract form in Nephrology Dialysis Transplantation. #

12 patients (48%) were on sustained remission. The main side effect was acute reversible nephrotoxicity (40%). Predictors of renal toxicity were age (P s 0.037), baseline creatinine (P s 0.046) and tacrolimus trough level (P s0.001). Conclusions. We conclude that combined therapy of tacrolimus and steroids induce sustained remission of proteinuria in a significant number of patients with idiopathic focal glomerulosclerosis whose disease was not controlled by the standard therapy of steroids and cyclosporin A. Keywords: corticosteroids; idiopathic focal glomerulosclerosis; nephrotic syndrome; tacrolimus

Introduction Patients with idiopathic focal glomerulosclerosis and nephrotic proteinuria develop end-stage renal failure almost invariably over a period of 6 –8 years w1–3x. These patients are considered candidates to therapy with steroids or immunosuppressive drugs based on the evidence that nephrotic patients who achieve remission of proteinuria have a significanlty improved long-term prognosis w4x. Although focal glomerulosclerosis was considered for long to be resistant to steroids, evidence from uncontrolled studies indicates that oral prednisone can induce a stable remission in a significant number of patients w5x. To date, the experience in adults shows that prednisone must be given for 6 months before concluding that the patient is steroid resistant. After prednisone is discontinued, only 20–30% of patients maintain a stable remission. The remaining patients are either steroid resistant or suffer subsequent relapses, which require re-treatment with steroids or other immunosuppressive drugs, and are therefore exposed to the harmful effects of prolonged steroid therapy w6x.

2002 European Renal Association–European Dialysis and Transplant Association

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The therapy for both steroid-resistant and relapsing nephrotic patients with primary focal glomerulosclerosis remains a major problem for nephrologists. Cyclosporin A has been tested in several uncontrolled and prospective randomized studies, with remission rates ranging from 25 to 30% w7–13x. One study reported remission rates of 50%, but its definition of steroid resistance was based on no response after 6 weeks of prednisone, which according to current evidence seems to be an inappropiately short time period w7x. The evidence seems to indicate that cyclosporin offers little benefit for steroid-resistant patients. Moreover, most steroid-dependent patients also become cyclosporin dependent and need prolonged therapy to maintain remission, which exposes them to the nephrotoxic effects of this drug. Tacrolimus (FK 506) is a macrolide antibiotic isolated from Streptomyces tsukubaensis. As a calcineurin inhibitor, the basic mechanism of action of tacrolimus is very similar to that of cyclosporin, inducing a selective inhibitory action on CD4 T-helper lymphocyte activation and proliferation. However, both in vitro and experimental studies have demonstrated that its immunosuppressive activity is nearly 100 times higher than that of cyclosporin w14x. In recent years, successful treatment of steroid- and cyclosporin-resistant patients with tacrolimus has been reported in single cases or small series of patients w15–17x. Although encouraging, these results should be analysed with caution because neither the rate of relapses after tacrolimus discontinuation nor the longterm nephrotoxic effects have yet been described. Our hypothesis was that tacrolimus, sharing the basic mode of action of cyclosporin and having superior immunosuppressive effects, would either induce remission in cyclosporin-resistant patients or maintain a more stable remission in relapsers. We decided to associate tacrolimus and prednisone based on previous data indicating that in steroid-resistant patients the combined use of a low dose of prednisone with cyclosporin enhanced the likelihood of remission w2x. In this study, we analysed the efficacy and safety of a combined therapy of tacrolimus and prednisone in a group of 25 patients with either cyclosporin-resistant or -dependent idiopathic focal glomerulosclerosis.

Subjects and methods The study design was a non-comparative, open, uncontrolled study.

2.

3. 4.

5. 6. 7.

Histopathological analysis of kidney biopsies All biopsies were stained with haematoxylin–eosin, PAS and Masson’s thrichromic for light microscopy evaluation, and with fluoresecent antibodies against IgG, IgM, IgA, C3 and fibrinogen for immunofluorescence analysis. In each biopsy, we determined the percentage of glomeruli with total sclerosis and the percentage of glomeruli with focal sclerosis. The amount of interstitial fibrosis was quantified using an image autoanalyser Olympus WCUE-2 (Olympus, Hamburg, Germany) in 5-mm trichromic-stained sections.

Treatment protocol All patients received a free diet with no added salt. To control the antiproteinuric effect of antihypertensive therapy, all patients started treatment with the ACE inhibitor enalapril (20 – 40 mguday) at least 6 months before being included in the study. Three patients suffered from cough after starting enalapril treatment and were therefore switched to the angiotensin receptor antagonist losartan (50 mguday). Other antihypertensive drugs were added if needed to achieve accurate control of blood pressure. Hyperlipidemia was treated with either simvastatin (20 – 40 mguday) or atorvastatin (10 –20 mguday) when serum LDL cholesterol was )200 mgudl. The treatment regime was as follows. 1.

Patients During a period of 4 years, we recruited a group of 25 patients from four different centres, who fulfilled all of the following criteria. 1.

Diagnosis of focal glomerulosclerosis by renal biopsy. All biopsies carried out in the different centres were reanalysed by a single pathologist who confirmed the diagnosis.

Exclusion of secondary forms, including reduction in renal mass, morbid obesity, AIDS, cocaine- and heroinassociated nephropathy, analgaesic-induced nephropathy, reflux nephropathy and hereditary diseases. Resistance to steroids defined as the persistence of nephrotic range proteinuria after 6 months of therapy with prednisone (1 mgukguday). Adequately defined cyclosporin resistance (primary or secondary) or cyclosporin dependence. Primary resistance to cyclosporin was defined as the persistence of nephrotic proteinuria after a 6-month trial with steroids (1 mgukguday) and cyclosporin (5 mgukguday). Secondary resistance to cyclosporin was defined as the relapse of nephrotic proteinuria and resistance to therapy with cyclosporin, in patients who initially achieved complete or partial remission. Cyclosporin dependence was defined as at least two consecutive cyclosporin-responsive relapses of nephrotic proteinuria while being tappered off or shortly after discontinuation of cyclosporin. Serum creatinine level -3 mgudl. No therapy with other immunosuppressives or nonsteroidal anti-inflammatory drugs 6 months prior to the inclusion in the study. Absence of contraindications to tacrolimus and steroid therapy including pregnancy, insulin-dependent diabetes mellitus, active infectious diseases, peptic ulcer or malignancy.

2.

Prednisone was started at a dose of 1 mgukguday (maximum daily dose 60 mg) administered po in a single morning dose for 4 weeks, and then administered 1 mgukg every other day until week 8. After week 8, prednisone was progressively tapered off at a rate of 0.05 mgukguweek until complete supression by week 24. Tacrolimus was started at a total daily dose of 0.15 mgukg, which was given in two divided doses at 12-h intervals. During follow-up, the dose was adjusted to achieve 12-h whole blood levels between 5 and 10 nguml. Blood tacrolimus levels were determined by

Tacrolimus in cyclosporin resistant focal glomerulosclerosis

the microparticle enzyme immunoassay (MEIA) in the lmx analyser (Abbott Laboratories, IL, USA) according to the manufacturer’s instructions. The betweeen-run precision and the limit of detection of the assay were -9.5% and 1.22 nguml, respectively. Every time a rise of )25% baseline serum creatinine was observed, the total daily dose of tacrolimus was reduced by 25% and serum creatinine was determined again within 1 week. If serum creatinine did not improve to baseline levels after this dose reduction, tacrolimus was then discontinued. Those patients who achieved complete remission of proteinuria within the first 6 months of treatment were progresively tapered off tacrolimus at a rate of 1 mguweek until supression. In all the other cases, the administration of tacrolimus continued for a maximum of 6 months. Relapses in proteinuria were treated following the same protocol. If partial or total remission of proteinuria was achieved with a second trial, tacrolimus was maintained for 1 year. After 1 year of therapy, tacrolimus was discontinued in all patients to avoid potential long-term nephrotoxicity. The study protocol was approved by the ethical committee of our centre and all patients gave their written consent prior to inclusion in the study.

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Follow-up After being included in this study, patients were controlled on an out-patient basis every week during the first 4 weeks. After the 4th week, controls were carried out every month during the first 6 months, every 2 months until the end of the first year and every 4 months until the end of the second year. In each control, we recorded the patient’s clinical status, the systolic and diastolic blood presssure, and we carried out biochemical controls of serum creatinine, 24-h creatinine clearance (CrC), electrolytes, lipids, liver function, glycaemia, 24-h urinary protein excretion and tacrolimus 12-h trough levels.

Outcome variables The primary outcome was the number of patients with complete or partial remission. Secondary analyses included the number of patients with a reduction in proteinuria to non-nephrotic levels, time to achieve a complete or partial remission and serum creatinine after 2 years follow-up. In addition, we analysed the evolution of the slopes of creatinine clearance over time in patients with and without complete or partial remission.

Definitions

Statistical analysis

Nephrotic proteinuria was defined as an urinary protein excretion )3.5 guday. Nephrotic syndrome was defined as a proteinuria )3 guday, hypoalbuminaemia -3.5 gudl and oedema. Complete remission was defined as a proteinuria -0.3 guday in three consecutive controls carried out within a period of 3 months. Partial remission was defined as an urinary protein excretion -2 guday and )0.3 guday. High blood pressure was defined as a systolic blood pressure )140 mmHg and diastolic blood pressure )85 mmHg. Resistance to tacrolimus was defined as the persistence of nephrotic proteinuria after a 6-month trial. Relapse of proteinuria was defined as an increase in urinary protein excretion )3.5 guday confirmed in two consecutive analyses after tacrolimus discontinuation. Chronic renal failure was defined as a glomerular filtration rate (GFR) -60 mlumin. End-stage renal failure was defined as a creatinine clearance -10 mlumin. Acute tacrolimus nephrotoxicity was defined as an increase in baseline serum creatinine )25% which improved after a 25% reduction in total daily dose of tacrolimus.

The results are given as the mean"SD. Comparisons between groups were carried out by the x2 test for proportions and non-parametric Mann–Whitney rank sum test for quantitative variables. The diferences in renal function outcome over time between patients with and without remission of proteinuria were compared by the students t-test of the slopes of creatinine clearance over the observation period of 2 years. A P value of -0.05 was considered statistically significant. Statistical analyses were carried out with the SPSS 8.0 software.

Results Baseline characteristics of the study group Clinical and biochemical characteristics. The main clinical and biochemical characteristics of the 25 patients included in the study are summarized in Table 1. Time between renal biopsy and tacrolimus

Table 1. Baseline clinical, histopathological and biochemical characteristics of study group

Age (years) Systolic blood pressure (mmHg) Diastolic blood pressure (mmHg) Interstitial fibrosis extent (%) % of sclerosed glomeruli % of glomeruli with focal sclerosis Serum creatinine (mgudl) GFR (mlumin) Proteinuria (guday) Glycaemia (mgudl) LDL-cholesterol (mgudl)

Mean

SD

Percentile 25

Percentile 75

44 129.74 74.60 12 14.60 31.36 1.2 75 10.3 106 260

14 15.06 9.99 8.5 12.25 15.38 0.5 15 9.5 23 62

32 120 68 5 0 20 0.8 65 4.0 86 208

54 140 80 20 25 47.5 1.6 90 12.0 110 332

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therapy ranged from 3 to 7 years with a median of 3.5 years. According to our selection criteria, all patients showed resistance to a 6-month trial of steroids and had been previously treated with a 6-month trial of steroids and cyclosporin. In addition, six patients had been unsuccessfully treated with a short term (8 weeks) chlorambucil trial and four patients with cyclophosphamide. Before tacrolimus treatment, 13 patients (52%) showed CrC -80 mlumin, and 15 patients (60%) were hypertensive and required antihypertensive therapy. Fourteen patients (56%) suffered from clinical nephrotic syndrome, whereas the remaining 11 patients suffered from nephrotic proteinuria but not nephrotic syndrome. Twelve patients (48%) had been previously responsive to cyclosporin and prednisone but developed either two consecutive relapses soon after cyclosporin was discontinued (cyclosporin dependence; ns 5), or relapses of proteinuria that did not respond to treatment with cyclosporin (secondary resistance; ns 7). The remaining 13 patients (52%) never responded to cyclosporin and steroids (primary resistance). Time between previous cyclosporin treatment and inclusion in the study ranged from 1 to 3 years with a median of 2.5 years. Histopathological analysis. According to morphological classification of FSGS, four patients showed hiliar lesions, five patients peripheral lesions, 12 patients hiliar and peripheral lesions, and four patients showed mesangial hypercelullarity. Immunofluorescence staining showed focal IgM deposits in 13 patients, focal IgM and C3 in seven patients, isolated focal deposits of C3 in three patients and fibrinogen in two patients. The main histopathological findings at diagnosis are summarized in Table 1. We found significant correlations between GFR and both the amount of glomeruli with total sclerosis (r s0.62, P-0.01) and the amount of interstitial fibrosis (rs 0.72, P-0.01).

Response after the first 6-month trial of tacrolimus and prednisone After starting treatment with tacrolimus and prednisone, complete remission was achieved in 10 patients (40%) and partial remission in two patients (8%). In five additional patients (20%), urinary protein excretion decreased to non-nephrotic levels but persisted with levels )2 guday. Overall, proteinuria decreased to levels -3.5 guday in 17 patients (68%). Table 2 summarizes the response to therapy when the patients were classified according to the inclusion criteria. Mean time to achieve complete or partial remission was 112"24 days (median s120 days; 95% confidence interval (CI) s 98–126 days). Table 3 summarizes the evolution of the main clinical and biochemical variables over the first year of follow-up. Predictors of response to tacrolimus and steroids Table 4 summarizes the results of the univariate analysis carried out to evaluate the clinical, biochemical and histological variables associated with response to tacrolimus. Remission of proteinuria was neither associated with age, sex, baseline proteinuria, nephrotic syndrome, amount of interstitial fibrosis, tacrolimus Table 2. Response after 6 months of a combined therapy of tacrolimus and prednisone in patients classified according to inclusion criteria CR

PR

RP -3 guday Total

CyA dep (n s 5) 5 (100%) Sec res (n s 7) 3 (42.8%) 2 (28.5%) 2 (28.5%) Prim res (n s 13) 2 (15.3%) 3 (23%)

5 (100%) 7 (100%) 5 (38.4%)

CyA dep, cyclosporin dependence; Sec res, secondary resistance to cyclosporin; Prim res, primary resistance to cyclosporin; CR, complete remission; PR, partial remission; RP -3 guday, reduction of proteinuria to non-nephrotic levels.

Table 3. Evolution of renal function, proteinuria, serum albumin and tacrolimus trough levels during the first year of follow-up

Basal serum creatinine (mgudl) Serum creatinine at 6 months (mgudl) Serum creatinine at 12 months (mgudl) Basal GFR (mlumin) GFR at 6 months (mlumin) GFR at 12 months (mlumin) Basal proteinuria (guday) Proteinuria at 6 months (guday) Proteinuria at 12 months (guday) Basal serum albumin (gudl) Serum albumin at 6 months (gudl) Serum albumin at 12 months (gudl) FK trough level (first week) (nguml) FK trough level (second week) (nguml) FK trough level (fourth week) (nguml) FK trough level (6 months) (nguml) FK trough level (12 months) (nguml)

Mean

SD

Percentile 25

Percentile 75

1.2 1.3 1.4 73 75 68 10.3 3.1 2.6 2.6 3.8 3.8 11.8 10.3 10.5 8.5 6.7

0.5 0.6 0.5 16 21 29 9.5 3.4 3.2 0.6 0.9 .9 3.8 1.8 1.4 2.2 3.0

0.8 0.9 0.9 65 56 49 4.0 0.4 0.1 2.2 2.9 2.5 8.8 9.2 9.8 6.1 4.0

1.6 1.8 1.9 90 95 95 12.0 6.5 4.2 3.0 4.5 4.6 14.0 12.1 11.1 10.2 10.0


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Table 4. Clinical, biochemical and histopathological characteristics of patients with complete or partial remission of proteinuria (R) and without response (NR) to tacrolimus and prednisone R

Age (years) Systolic blood pressure (mmHg) Diastolic blood pressure (mmHg) Time from renal biopsy (years) Basal creatinine mgudl Basal GFR mlumin Basal proteinuria guday Basal cholesterol mgudl Basal LDL-cholesterol mgudl Basal serum albumin gudl % of sclerosed glomeruli % of glomeruli with focal sclerosis Interstitial fibrosis surface FK trough level (first week) (nguml) FK trough level (second week) (nguml) FK trough level (fourth week) (nguml) FK trough level (6 months) (nguml)

NR

P value

Mean

SD

Mean

SD

44.6 135.9 74.1 3.7 1.2 78.6 8.5 348.4 234 2.4 14 24.9 10.4 10.6 9.2 10.3 8.9

15.1 14.8 10.4 1.2 0.54 19.5 4.3 64.1 38 0.3 10.7 15.6 6.8 4.5 1.9 1.4 2.3

43.7 130.3 80 3.9 1.2 71.4 11.5 402 280.8 2.5 16.5 28.5 14.5 13.5 10.6 10.9 9.3

13.1 15.6 7.0 1.07 0.46 11.3 12.2 96.1 69 0.7 11.2 15.1 10.5 3.6 1.5 1.1 1.6

trough levels nor with baseline renal function in the range of values showed by our patients. The only variable associated with response to tacrolimus and steroids was a previous response to cyclosporin either as a complete or as a partial remission (remission rate 75% for patients with previous response to cyclosporin vs 15.3% for patients unresponsive to cyclosporin, P s0.036). Relapses after tacrolimus and steroid discontinuation After tacrolimus was discontinued at 6 months, relapses were observed in 13u17 (76%) patients who were in remission (four patients with cyclosporin dependence, five with secondary resistance to cyclosporin, and four patients with primary resistance to cyclosporin). Time to relapse ranged from 1 to 4 months with a median of 3 months. After a second trial of tacrolimus, five of these 13 patients achieved complete remission (38.4%), four partial remission (30.7%) and four (30.7%) reduction of proteinuria to -3 guday in a mean time of 60 days (range 30 –120 days). Tacrolimus therapy was maintained at doses ranging from 0.02 to 0.05 mgukguday for 1 year and then discontinued in all these patients. After definitive tacrolimus discontinuation, five patients maintained complete or partial remission, and eight relapsed within the first 6 months and were not re-treated to avoid potential long-term nephrotoxicity associated with tacrolimus. Table 5 summarizes the renal outcome of our patients at the end of the second year of follow-up. Evolution of renal function over time in responders and non-responders During a mean follow-up of 3.5 years before tacrolimus therapy, slopes of CrC were 6.6"2.4 mluminu year in patients who afterwards achieved remission

0.8 0.11 0.37 0.67 0.96 0.25 0.45 0.12 0.14 0.76 0.49 0.50 0.72 0.09 0.11 0.22 0.58

Table 5. Renal outcome at the end of the second year of follow-up of patients clasified according to inclusion criteria

CyA dep (n s 5) Sec res (n s 7) Prim res (n s 5)

CR

PR

RP-3 guday

NProt

ESRD

3 3 2

2 2 1

0 0 4

0 0 4

0 2 2

CyA dep, cyclosporin dependence; Sec res, secondary resistance to cyclosporin; Prim res, primary resistance to cyclosporin; CR, complete remission; PR, partial remission; RP -3 guday, reduction of proteinuria to non-nephrotic levels; NProt, nephrotic proteinuria; ESRD, end-stage renal failure.

after tacrolimus and prednisone therapy, and 6"5.06 mluminuyear in non-responding patients (P s NS). Two years after tacrolimus therapy was started, the slope of creatinine clearance was 2.6"1.6 mluminuyear in patients with sustained partial or complete remission (n s12) and 7.2"3.41 mluminuyear in non-responding patients (n s13) (P-0.001). At the end of the second year, four of 14 (28.5%; 95% CI s 5–50%) patients with persistent proteinuria, and no patients with complete or partial remission, suffered end-stage renal failure (Table 5). Adverse effects All patients completed the first 6 months of treatment. During therapy with tacrolimus, increases in baseline serum creatinine fulfilling the criteria for reversible acute nephrotoxicity were observed in 10u25 patients (40%). All these episodes were observed within the first week of treatment, during the phase of adjustment of tacrolimus dose. Serum creatinine increases

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were mild, ranging from 25% to 30% (median 28%) and reversible in all cases. Acute nephrotoxicity was statistically associated with baseline serum creatinine (mean 1.41"0.58 mgudl vs 0.94"0.2 mgudl; P s0.046), age (mean 46.9"15.5 years vs 35.3"7 years; P s 0.037) and tacrolimus trough levels (mean 14.55"3 nguml vs 9.1"2.15 nguml; P s 0.001). After analysing the statistical correlations of the acute nephrotoxic episodes, we decided to reduce the starting dose of tacrolimus for the 13 patients who relapsed after the first trial and were re-treated to 0.08 mgukguday for patients with serum creatinine -1.4 mgudl and to 0.06 mgukguday for those with baseline serum creatinine )1.4 mgudl. These doses provided 12-h trough levels ranging from 5 to 9 nguml and 4 to 7 nguml, respectively. Using this lower dosing schedule, no new episodes of acute nephrotoxicity were observed. The other significant adverse effects observed during tacrolimus treatment were gastrointestinal symptoms in one patient, and new onset of high blood pressure requiring antihypertensive therapy in two patients.

Discussion In this preliminary uncontrolled study we analysed the efficacy and safety of combined therapy of tacrolimus and steroids in a group of 25 nephrotic patients with well defined steroid-resistant primary focal glomerulosclerosis who suffered resistance or relapsed when treated with cyclosporin and steroids. This report does not intend to address the many issues dealing with the critical evaluation of tacrolimus in the treatment of cyclosporin-resistant focal glomerulosclerosis. We believe that our results are important in so far as they represent the first clinical trial including an homogeneous group of patients treated by the same protocol with a prospective follow-up. Moreover, given the potential harmful effects of long-term treatment with both cyclosporin and tacrolimus and the known antiproteinuric effects of ACE inhibitors w18x, patients were included only if nephrotic proteinuria persisted after 6 months of therapy with the ACE inhibitor enalapril. Our data indicate that a combined therapy of tacrolimus and steroids may induce complete or partial remissions of proteinuria in certain patients whose disease could not be controlled by standard therapies of cyclosporin and steroids. The only variable predicting the response was the response to a previous trial of cyclosporin and steroids. These results are in concordance with previous studies and suggest that only a group of patients with FSGS are highly responsive to therapy with steroids or calcineurin inhibitors. Unfortunately, on the basis of clinical and biochemical variables it is not possible to predict which patients will benefit from therapy and which will not w18x. When tacrolimus was discontinued early,

A. Segarra et al.

most patients relapsed and needed to be re-treated. In these cases, we decided to maintain tacrolimus for 1 year based on the experience reported with cyclosporin in previous studies w10x. After this period, a significant number of patients sustained partial or complete remission of proteinuria. After 2 years of follow-up, the potential benefit of tacrolimus therapy was evident only in patients who developed cyclosporin resistance after an initial response (secondary resistance) and in a minority of patients who never responded to previous therapy with steroids and cyclosporin (primary resistance), as their disease could eventually be controlled. On the contrary, tacrolimus did not add any benefit for cyclosporin-dependent patients because, according to current evidence, in these cases remission could also have been sustained with prolonged cyclosporin therapy w10,11x. Furthermore, over an observation period of 2 years, the rate of decline of the slopes of CrC were significantly reduced only in patients with sustained partial or complete remission after therapy with tacrolimus and prednisone. It could be argued that patients who were able to sustain a remission suffered a more benign disease; however, neither the clinical data nor the rate of decline of renal function documented during follow-up prior to the inclusion in the study are in concordance with this hypothesis. These data are in agreement with two facts highlighted in previous studies in patients treated with cyclosporin: first, that responsive patients sustain remission only with prolonged therapy, and secondly that the improvement in renal outcome is dependent on a sustained remission of proteinuria w10–12x. The main adverse effect observed was reversible mild acute nephrotoxicity. Taking into account the statistical associations of the nephrotoxic episodes, the high percentage of acute nephrotoxicity observed in our patients can probably be attributed to an inadequate starting dose of tacrolimus in a group of patients with high frequency of preexisting renal failure. These data indicate that the expected acute nephrotoxicity of tacrolimus in nephrotic patients is similar to that described with cyclosporin and emphasize that tacrolimus must be used with the same caution as cyclosporin both in old patients and in patients with increased baseline serum creatinine. Although our study was not designed to carry out a dose-response analysis, our preliminary experience seems to indicate that starting doses of 0.08 mgukguday with trough levels ranging from 5 to 12 nguml are enough to induce remission in the majority of patients with normal renal function. In our opinion, for patients with pre-existing renal failure, tacrolimus should not be started at doses of )0.06 mgukguday, maintaning trough levels in the range of 5–10 nguml. Higher trough levels will increase the risk of nephrotoxicity with no further increases in the probability of response. The potential chronic nephrotoxicity of long-term tacrolimus therapy remains a major concern and prompted us to discontinue tacrolimus after 1 year of therapy. This approach could be criticized since certain patients were on


remission and relapsed after tacrolimus discontinuation. However, recent data from a randomized clinical study w12x seem to indicate that a sustained remission of proteinuria is not mandatory to improve the renal outcome of nephrotic patients with focal glomerulosclerosis treated with cyclosporin, whereas it has been clearly demonstrated that prolonged cyclosporin therapy may increase both glomerular and interstitial lesions of sclerosis even in FSGS in remission w10,11x. On the basis of the evolution of renal function over time we could argue that in our study group the renal outcome actually improved in the patients who sustained a remission while being treated with tacrolimus; however, from a histological point of view we cannot give an adequate answer to this point because we did not repeat the renal biopsies after tacrolimus therapy. An interesting point is why tacrolimus induced remission in certain patients previously unresponsive to cyclosporin. To date, we do not know the answer to this question. As clinical experience with tacrolimus has accumulated, it has become clear that the clinical profile of this drug differs in several points from that of cyclosporin. Although it has been demonstrated that tacrolimus has higher immunosuppressor effects than cyclosporin, presumably as a consequence of greater affinity of its complex with FK binding protein to calcineurin, it is unlikely that all the distinctive clinical features of tacrolimus derive from it. In addition to its inhibition of calcineurin, tacrolimus has a variety of effects on cellular functions, and intercellular and intracellular signalling events not shared with cyclosporin, which could be important for its clinical profile but remain to be more accurately defined w20–23x. In conclusion, our results showed that 28% and 48% of patients with previous resistance or relapses after cyclosporin treatment achieved stable remission of proteinuria 6 and 12 months after a combined therapy of steroids and tacrolimus, respectively. The results expected depend entirely on the indication used to try a therapy of tacrolimus and steroids. When prescribed for cyclosporin-dependent patients, tacrolimus did not add any significant benefit, as most patients suffered relapses when tacrolimus was discontinued and again needed prolonged therapy to sustain remission. The most interesting point of our study was the evidence that a significant number of patients who developed resistance to cyclosporin after having been initially responsive to it, and a minority of patients who never responded to steroids and cyclosporin, were able to sustain a partial or complete remission after therapy with tacrolimus and steroids. In these patients, tacrolimus would have the potential to improve the renal outcome, but this is still only an hypothesis that remains to be proven in randomized clinical trials with long-term follow-up. Finally, a certain number of patients whose disease was unresponsive to corticosteroids and cyclosporin but maintained remission after 1 year of therapy with tacrolimus, relapsed when this drug was discontinued to avoid potential chronic nephrotoxicity. In these patients, before arguing in

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favour of more prolonged tacrolimus therapy, the potential long-term nephrotoxic effects of tacrolimus should be further defined.

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