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Clinical Transplantation

Addition of inhaled corticosteroids to systemic immunosuppression after lung transplantation: a double-blind, placebo-controlled trial1

Whitford, Helen; Walters, E. Haydn; Levvey, Bronwyn; Kotsimbos, Tom; Orsida, Bernadette; Ward, Chris; Pais, Michael; Reid, Sue; Williams, Trevor; Snell, Greg2

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Abstract

Lung transplantation has become established as a recognized therapeutic option for patients with end-stage pulmonary parenchymal and vascular disease (1). In the years since the first successful lung transplant there have been significant advances in donor management, organ preservation (2), surgical techniques, and postoperative care. However despite this, 5-year survival remains approximately 50% (3). Long-term survival remains limited by opportunistic infections and the development of progressive airflow limitation, histologically characterized by obliterative bronchiolitis, and is thought to be a manifestation of chronic rejection (4). The clinical picture of progressive airflow obstruction in the absence of acute rejection, infection, or local stenosis is called bronchiolitis obliterans syndrome (BOS) (5).

It is widely believed that BOS is preceded by airway inflammation, yet we have previously described airway inflammation even in healthy non-BOS lung transplant recipients (6). BOS occurs despite significant immunosuppression (7). Treatment of BOS has often involved augmentation or alteration of immunosuppression with some positive responses. However, no treatment has been shown to be effective in a well-controlled trial (8,9).

Standard baseline regimes commonly use triple therapy with cyclosporine, azathioprine, and corticosteroids. All of these agents have significant systemic side effects. Targeted local immunosuppression has been considered in many solid organ transplants (10–12), with difficulty of delivery the main limiting factor in the clinical application of these agents. In contrast, with lung transplantation the inhaled route is a potential practical option, for which delivery devices are widely available.

Inhaled corticosteroids are now considered first line therapy in some conditions of chronic airway inflammation such as asthma (13). Although there continues to be a large amount of debate regarding the magnitude, if any efficacy in chronic obstructive pulmonary disease (COPD) (14,15); these agents are widely used in this condition. There are potentially major similarities between the pathophysiology of COPD and BOS after lung transplantation with infiltration of CD8-positive T lymphocytes, macrophages, and prominent neutrophils (16–18). In lung transplantation inflammation of the airways has been shown to be an important feature in established BOS, but is also present in stable lung transplant recipients (6,17).

There have been two publications looking at the role of inhaled corticosteroids in lung transplantation (19,20). Both of these small nonblinded studies showed improved lung function, and one suggested a reduction in the rate of BOS development in the treatment arm (19).

In this study we hypothesized that high dose inhaled corticosteroids would decrease the airway inflammation in stable lung transplant recipients and as a result reduce the subsequent incidence of BOS.

METHODS

Patients

Thirty lung transplant recipients were recruited in the 3–9 months after lung transplantation. A power calculation based on observed variability in a pilot study of lung transplant recipients suggested that detectable differences were not likely to improve beyond a sample size of 15 (given a conventional power of 80% and a significance level of 5%). Patients were enrolled when clinically stable and BOS stage zero (5). Enrollment period was from April 1997 to June 1998. All eligible patients transplanted at this institution were considered. All patients gave written informed consent before being considered for the study, which was approved by the Alfred Hospital Research Ethics Committee.

Study Design

This was a randomized, double-blind, placebo-controlled study. Patients randomized to receive either 750 μg of fluticasone propionate (FP) via metered dose inhaler and spacer device, twice daily or an identical-appearing placebo. A computer-generated allocation schedule was used initially from a block size of 20 and then blocks of 4. Patients were randomized sequentially by the pharmacy department staff from a list held by the pharmacy department comprising treatment numbers only.

The initial study period was 3 months, and this was later extended to 2 years, after lung transplantation. Twenty of the 30 recruited patients continued in the longer study. Reasons for noncontinuation are shown in Figure 1.

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Figure 1:
Study flow chart.

Bronchoscopy with bronchoalveolar lavage (BAL) and transbronchial biopsies were performed before commencement of study medication, at 3 months after commencement, and then as per routine follow-up protocol at this institution (i.e., at 3, 6, 9, 12, 18, and 24 months posttransplant and yearly thereafter), or when clinically indicated.

Endpoints in the short-term component of the study:

  • 1. Incidence of acute rejection.
  • 2. Development of early BOS.
  • 3. Change in lung function (FEV1).
  • 4. Episodes of fungal infection (both symptomatic, clinical diagnosis of oral candidiasis, and BAL growth of fungus).

Endpoints of the long-term component of the study:

  • 1. Freedom from BOS.
  • 2. Incidence of acute rejection.
  • 3. Incidence of fungal infection.

Study Exclusion Criteria

Patients were excluded if there was evidence of clinically significant infection before study commencement or lung function indicated BOS.

Immunosuppression

All patients initially received a standard immunosuppression regime. Consisting of cyclosporine (to achieve a blood level of 250–300 μg/L by EMIT assay [Syva, CA]), azathioprine (1–2 mg/kg maintaining white blood cells >5×109/L), and prednisolone (in a reducing dose posttransplant to a long-term maintenance dose of 7.5 mg/day). Alterations to immunosuppression were made as clinically indicated by the caring physician.

Pulmonary Function Testing

Spirometry was performed within the week before each bronchoscopy. A computerized rolling-seal spirometer (Sensor Medics922; Yorba Linda, CA) was used to measure forced expiratory volume in 1 sec (FEV1). Values were compared to an average of the highest two consecutive measurements 3–6 weeks apart, as per the International Society for Heart and Lung Transplantation classification of chronic dysfunction in lung allografts (5).

Fibreoptic Bronchoscopy with BAL and Transbronchial Biopsies

Bronchoscopy was performed using intravenous sedation with Midazolam (Roche, Basel, Switzerland), with or without the addition of Propofol (Zeneca, Melbourne, Australia). Pulse oximetry was performed throughout the procedure. Upper and lower airways were anesthetized with topical 1–2% lignocaine.

BAL was performed with the bronchoscope wedged in a subsegment of the right middle lobe or left lingula. 3×60 ml aliquots of phosphate buffered saline were instilled using a hand operated syringe, then aspirated into a siliconized container at controlled pressure of negative 80 mmHg. A representative 10-ml sample was sent for microbiological analysis. Total cell counts and differentials were performed on all samples.

Transbronchial biopsies were then performed using alligator forceps (Olympus, FB 15C; Tokyo, Japan) under fluoroscopic control and sent for standard histopathological assessment.

Statistical Analysis

Analysis was undertaken on an intention-to-treat basis using SPSS Version 10 (SPSS Inc., Chicago, IL). Continuous variables are described using mean and standard deviation and compared using Student’s t test. Nonparametric variables are described using median and range and compared using Mann-Whitney U test. Categorical variables are compared using chi-square test with Yates Correction as appropriate. Freedom from BOS and survival curves were calculated using Kaplan-Meier methods. Statistical significance was considered at the P <0.05 level.

RESULTS

In the initial short-term study, 30 stable lung transplant recipients were recruited: 13 were randomized to the placebo arm and 17 to the FP arm. The demographics of these groups are shown in Table 1. Lung function was very similar in the two groups, but there were other inadvertent baseline differences between the groups; the FP group was of a younger age (P =0.04) and had a greater proportion of females (P =0.02). Clinically, all patients were evaluated 3 months later at a mean of 29.9±6.4 weeks posttransplant for the placebo group and 37.8±13.6 weeks for the FP group (P =0.03). All patients continued on study medication for the duration of the study. One FP patient developed rapidly progressive BOS and did not undergo the second protocol directed bronchoscopy and is therefore not included in the paired analyses.

T1-16
Table 1:
Short-term placebo versus FP: patient demographics

Subsequently 19 patients were eligible and consented for the long-term study out to 2 years posttransplant. Eleven patients did not continue with the study and the primary reasons for noninclusion were: long distance from the transplant center (n=4), technical difficulties with the supply of blinded-study medication (n=3), nonconsent (n=2), development of BOS 1 (n=1), and the clinical decision to place the patient on inhaled therapy for falling lung function (n=1). The demographics of these groups are shown in Table 2. There were no statistical differences between these two groups. All patients were then followed for at least until 2 years posttransplant or until death. All FP patients continued on study medication for the duration of the study. Two placebo-treated patients were withdrawn from the study: one in the setting of other medical problems (Parkinson’s disease) and the other by personal choice (P =NS [not significant]).

T2-16
Table 2:
Long-term placebo versus FP: patient demographics

Clinical Outcomes

Over the period of the short-term study there were no statistical differences in terms of episodes of treated acute rejection, the development of BOS stage 1, lung function, the incidence of fungal infection, or overall survival.

The clinical outcomes of the long-term study are shown in Table 3. Figure 2 represents a plot of the freedom from BOS stage 1 for the FP and placebo groups. It is notable that these curves run out beyond the finish of the study at 2 years posttransplant. Although the medication phase of the study stopped at 2 years posttransplant, for completeness, up-to-date lung function is shown in all patients.

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Table 3:
Long-term placebo versus FP: clinical outcomes
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Figure 2:
A Kaplan-Meier plot of the freedom from developing BOS 1 for the patients on long-term therapy. The tick marks on the curves represent individual censored patients. Patients are censored from further inclusion in the curves when they reach the point of current survival from the time of study start.

Figure 3 shows survival of all patients from the study start. Again it is notable that the curve extends well beyond the immediate study period. There are no important differences between the groups.

F3-16
Figure 3:
A Kaplan-Meier survival plot of all patients from the start of the study. The tick marks on the curves represent individual censored patients. Patients are censored from further inclusion in the curves when they reach the point of current survival from the time of study start.

No patients in any of these study groups required therapy for fungal infection, and there were no particular side effects ascribed to the inhaled medication.

Bronchoscopic Outcomes

The short-term study noted no significant rejection or microbiological changes over the 3 months of the study as shown in Table 4.

T4-16
Table 4:
Short-term placebo versus FP: bronchoscopic outcomes

The longer-term study did demonstrate bronchoscopic differences (Table 5). Pooled bronchoscopic follow-up results note a lower BAL lymphocyte count in the placebo group compared with the FP group (P =0.004), although these seem to mirror a nonstatistical trend to a difference in baseline values. There is also a higher incidence of positive bacterial cultures in the placebo arm compared with the FP group (P =0.04) that also mirrors nonstatistical baseline differences.

T5-16
Table 5:
Long-term placebo versus FP: bronchoscopic outcomes

DISCUSSION

Lung transplantation represents a particularly complex patient group to study, which has resulted in only a handful of randomized, blinded, placebo-controlled immunosuppression trials having ever been undertaken. In this context it is not then surprising that although based only on limited evidence (19,20), inhaled corticosteroids have been widely used to treat, and as prophylaxis against, airway inflammation after lung transplant.

Immunosuppression targeted to the transplanted organ has considerable appeal as a mode of long-term control of allorejection. Although this has been attempted experimentally in the setting of liver (11,12) and renal transplantation (10) with limited success, utilization of the inhaled route of delivery for immunosuppression in lung transplantation has several very attractive features. First, it may be possible to achieve higher drug levels in the allograft with less systemic side effects. Second, it may be possible to get potent local airway effects not achievable with systemic administration. For example, as well as its obvious immunological activities, inhaled FP has potentially beneficial effects on local airway vascularity (21), a problem we have previously described in the lung transplant setting (22). An inhaled therapy is particularly relevant given the airway-based nature of lung transplant chronic rejection and the lack of efficacy of current treatments. Additionally, there are practical advantages in the availability of accurate delivery devices for use via the inhaled route.

The Pittsburgh Lung Transplant Group has produced encouraging preliminary data in lung transplant recipients regarding the use of inhaled cyclosporine as adjuvant therapy for acute rejection and BOS (23,24). However, the lipophilic nature of the drug leads to technical problems in achieving appropriate particle size and contributes to cough as a particular side effect.

The use of inhaled steroids has previously been described in lung transplantation in a small randomized but open-label trial in the setting of refractory acute rejection (19) and as a single patient case report as therapy for BOS (20). The study by Takao et al. (19) specifically targeted those with troublesome acute rejection using 1 g of nebulized budesonide twice daily for 12 months. Withdrawal of patients from oral steroids was attempted and was more successful in the treatment. This study demonstrated no difference in the in terms of acute rejection or FEV1 over the 1 year of the trial, but at 24–30 months after the study the FEV1 was lower in the control group. The open-labeled design, and the limitation of presenting data from only five patients in the active arm, significantly limits the power of this study. However, this background was sufficiently promising to encourage us to plan the current study. FP was chosen as the most potent inhaled steroid available and the added advantage of minimal systemic absorption (25,26). It was felt that a dose of 1.5 mg/day of fluticasone was greater than the 1 mg b.i.d. of budesonide used by Takao et al. (19) with evidence of benefit and thus was considered an effective dose. This dose comparison, however, does not take into account the differing efficiencies of the delivery devices. The one published study by O’Reilly et al. (27), which directly compares these two drugs and delivery methods, concluded that fluticasone via metered-dose inhaler and spacer is at least as efficacious and probably more so than nebulized budesonide.

Despite optimism engendered from previous reports, the current study shows no advantage of inhaled FP over placebo. Although the study is relatively small, and potentially greater numbers on therapy for a longer time may show differences, the absence of any trends or rebound effects does not encourage such a trial. It is also possible that inhaled steroids need to be started earlier (perhaps at the time of transplant) to switch off a BOS signal that was already in place by the time the current study started at approximately 6 months posttransplant. There is some evidence that early acute rejection is linked to chronic rejection in this fashion (6,28). During the study our patients had a relatively low incidence of acute rejection in either arm and perhaps we were too late to influence this relationship. Although well characterized in the normal and asthmatic airway, the site and extent of deposition of inhaled drug has received less attention in the COPD setting and no attention in the lung transplant setting. Future studies of inhaled therapy in lung transplant recipients may need to include deposition studies.

The absence of a clinical response with the inhaled FP in the lung transplant setting is consistent with other publications in related “models” of chronic airway disease. Wong et al. (29) showed no benefit of inhaled FP in infants recovering from acute bronchiolitis. Burge et al. (30) in the large ISOLDE inhaled FP trial showed no change in the rate of decline of FEV1 over 3 years in patients with moderate-to-severe COPD. Although not unanimous, the expanding COPD literature on the subject supports the view that such pathologies are not steroid responsive (14,31).

Thus, the concept for treating airway disease via inhaled therapy in COPD and BOS is based on the success of this mode of treatment in asthma studies. In fact, there is increasing evidence that there are fundamental differences in the airway inflammation (and potential subsequent therapeutic response) when comparing asthma to COPD and has extrapolation to BOS. Inhaled and oral corticosteroids are very effective at suppressing the eosinophilic inflammation that characterizes asthma (16). Conversely, inhaled and oral corticosteroids do not consistently reduce inflammatory cell numbers (neutrophils in particular) or inflammatory cytokines in COPD (32,33). Barnes (16) suggests that COPD may actually involve an active resistance to inhaled steroids, because in vivo steroid therapy fails to suppress cytokines such as tumor necrosis factor α and interleukin-8, which are suppressible in vitro. There is evidence that BOS can similarly be regarded as a steroid-resistant disease at both a clinical (8,9) and molecular level (34).

The study was blinded so as not to influence the prescribing of concomitant immunosuppressive and antifungal therapy. The study started 3–6 months posttransplant at a time when we felt that significant fungal complications were unlikely.

Fungal infection, in particular with Aspergillus, is associated with serious morbidity and mortality after lung transplantation (35). One potential concern with the use of inhaled steroid was the prospect of increased upper and potentially lower airway fungal infection. Fortunately, increased fungal problems were not noted with FP in this study, reflecting both the reasonable safety margin of inhaled FP and the generous use of nystatin mouthwash and oral antifungals.

In conclusion, despite airway inflammation as the basis for the fall in lung function, which characterizes chronic allorejection and defines the BOS after lung transplant, inhaled FP is an ineffective therapy for the prevention of this condition. From the current study it is not apparent whether this result represents inadequate local delivery, the timing of therapy relative to the transplant, or a situation of local steroid resistance. Although a negative study is disappointing, the safety and relative ease of local immunosuppressive therapy in lung transplantation should still encourage further work with existing and emerging agents, potentially delivered from the time of transplant.

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