INTRODUCTION

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Nedocromil sodium is effective at inhibiting bronchial responses to airway drying such as exercise (1) and cold air hyperventilation (2), and osmotic stimuli such as hypertonic saline (3) and distilled water (4). Both airway drying and osmotic stimuli are thought to cause airway narrowing indirectly through changing the osmolarity of the airway surface, leading to the release of mediators (5, 6).

Inhaled mannitol is a newly developed bronchial provocation test that identifies asthmatics responsive to exercise, hyperventilation, and hypertonic saline (7, 8). We have proposed that mannitol induces airway narrowing via a similar mechanism to other osmotic stimuli, by increasing the osmolarity of the airway surface, causing release of mediators and leading to bronchial smooth muscle contraction (7).

To improve our understanding of the mannitol challenge, we assessed the effect of nedocromil sodium on airway responsiveness to mannitol in subjects with clinically diagnosed asthma. Nedocromil sodium (8 mg) or its placebo were administered randomized, double blind 10 min before challenge testing with inhaled mannitol.

	METHODS

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An advertisement circulated in the local community was used to recruit asthmatics. Subjects were interviewed by telephone and were asked to attend the laboratory if they were nonsmokers and had no chest infection 4 wk before the study. Subjects were asked to refrain from taking short-acting bronchodilators for 6 h, long-acting bronchodilators for 48 h, and nedocromil sodium or sodium cromoglycate for 36 h before the study. No inhaled corticosteroids were taken on the day of the study and no antihistamines 3 d before the study day. No vigorous exercise was permitted on the study day before testing, and the dose of inhaled corticosteroids was to have remained unchanged for at least 4 wk before the study. On the first day of the study, the subject had a history taken and was examined by a staff respiratory physician and a clinical diagnosis of asthma was confirmed.

Thirty-nine subjects were recruited and 24 (11 females and 13 males) age between 15 and 46 yr met the inclusion criteria and were entered into the study (Table 1). All subjects had been prescribed medication for their asthma; 13 were taking inhaled corticosteroids on a daily basis and 23 were taking ₂-adrenoceptor agonists (Table 1). All subjects had at least one positive skin test (3-mm wheal or more) in response to a common aeroallergen.

The study was approved by the Central Sydney Area Health Service Ethics Committee (X98-0080) and all subjects signed a consent form prior to commencement of the study. The study was performed under the Clinical Trials Notification scheme No. 1998/217 of the Therapeutic Goods Administration of Australia.

Study Design

Subjects were asked to attend the laboratory on three occasions. The first day served as a control day for familiarization, and a mannitol challenge was performed to assess responsiveness. To enter the study, the subjects were required to have a 15% reduction in FEV₁ (PD₁₅) less than 350 mg. The second and third days consisted of pretreatment with the administration of the active drug, 8 mg of nedocromil sodium (4 × 2 mg actuations) or its placebo (Rhône Polenc Rorer, Baulkham Hills, NSW, Australia) from a metered-dose inhaler. There was a minimum of 48 h between each challenge and all challenges were completed in each subject within a 3-wk period (5 to 21 d).

Lung Function Measurements

Spirometry was performed using a Microlab 3300 Spirometer (Micro Medical Ltd, Kent, UK). Forced expiratory volume in one second (FEV₁) was used as an index of change in airway caliber. On the control day the baseline FEV₁ had to be at least 75% of predicted (9, 10).

Mannitol Capsule Challenge

The preparation of the dry powder mannitol has been described in detail previously (8). The mannitol powder used had 64% of the particles by mass under 7 µm in diameter measured by dispersion into a multistage liquid impinger (Astra Pharmaceuticals, Lund, Sweden). A Halermatic (Rhône Polenc Rorer, Collegeville, PA) was used for the delivery of the mannitol.

On arrival at the laboratory on the control day, each subject had their FEV₁ measured in triplicate and this was repeated 10 min later to confirm stability. On the treatment days spirometry was performed and the metered-dose inhaler was actuated 5 times before administration to the subject. Four separate inhalations were made while subjects were instructed to take a long, slow inhalation. At least 60 s separated each inhalation and 10 min after the first inhalation, spirometry was repeated.

A nose clip was applied and subjects then performed the challenge with doses consisting of 0 (empty capsule acting as a placebo), 5, 10, 20, 40, 80, 160, 160, and 160 mg of mannitol via the Halermatic. The 80 mg and 160 mg were given in multiple doses of 40-mg capsules. After inhalation of each capsule, patients were instructed to hold their breath for 5 s. At least two repeatable FEV₁ maneuvers were performed 60 s after each dose and the highest FEV₁ was used in the calculation. The FEV₁ value measured after the 0 mg capsule was taken as the prechallenge FEV₁ and used to calculate the percentage decrease in FEV₁ in response to the mannitol challenge. If the subject had a greater than 10% decrease in FEV₁ in response to a single dose, the same dose was repeated for reasons of safety. The challenge ceased when a 15% reduction in FEV₁ was documented or a cumulative dose of 635 mg had been administered. In subjects in whom a 15 to 20% decrease was recorded, the dose administered to cause that response was repeated to establish if an additional dose would lead to either further airway narrowing or a plateau in the response. The provoking dose of mannitol to cause a 15% decrease in FEV₁ (PD₁₅) was calculated by linear interpolation of the relationship between the percent fall in FEV₁ and the cumulative dose of mannitol required to provoke this.

Statistical Analysis

The geometric mean (Gmean) ± 95% confidence interval (CI) of the PD₁₅ and the response dose ratio (RDR) (final %fall in FEV₁/total dose administered) were calculated using the log transformed values which were normally distributed. Student's paired t test was used to compare the values between nedocromil and placebo results (11). Those subjects not recording a 15% decrease to mannitol were assigned a PD₁₅ value of the maximal dose of mannitol administered of 635 mg. A clinically significant difference in PD₁₅ was considered if response after nedocromil was beyond the range of repeatability of the mannitol challenge after placebo (i.e., 1.16 doubling doses using the Halermatic [n = 12], laboratory data). Percent protection to nedocromil was calculated using the percent fall in FEV₁ obtained at the dose of mannitol administered that was the highest cumulative dose common to both study days. Values for FEV₁ are expressed as a mean ± SD of the percentage of predicted normal FEV₁. Values for total dose of mannitol administered and percent fall in FEV₁ are also expressed as mean ± SD. Occurrence of a maximal response plateau was defined if the change in FEV₁, expressed as a percentage of baseline, varied by less than 5% at the highest doses (3).

	RESULTS

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Nedocromil significantly inhibited the response to inhaled mannitol. Fifty percent of subjects no longer responded to mannitol and failed to record a 15% fall in FEV₁ after the challenge (Figure 1). The remaining 50% still responded to mannitol following nedocromil, although a larger dose of mannitol was required to cause a similar response compared with placebo (Figure 1, Table 1). For seven of the 12 who remained positive to mannitol the shift in responsiveness to mannitol exceeded the repeatability of the response. The 12 subjects who remained responsive to mannitol after nedocromil all had a PD₁₅ less than 200 mg on both the control and placebo days. In this group the Gmean PD₁₅ was 88 mg (95% CI: 62-126) (p = 0.0008) after placebo and was significantly lower than the Gmean PD₁₅ of 275 mg (95% CI: 160-471) for the group that no longer responded to mannitol after nedocromil.

View larger version (24K): [in this window] [in a new window]   Figure 1.   Values for individual data showing an increase in the provoking dose of mannitol to cause a 15% reduction in FEV1 (PD15) after administration of nedocromil compared with placebo in 24 subjects. After nedocromil, half the subjects had a negative response to mannitol (with an assigned PD15 of 635 mg, open circle). The remaining subjects remained positive to mannitol after nedocromil but had a shift in responsiveness where a larger dose of mannitol was required to cause a similar response.

There was a significant reduction in the RDR after administration of nedocromil. The RDR after nedocromil was 0.02 (95% CI: 0.01-0.03) and placebo 0.09 (95% CI: 0.06-0.13) (p < 0.0001). All subjects, particularly those negative to mannitol after nedocromil had RDR values closely approaching that of healthy nonasthmatic subjects (8).

The percent protection afforded by nedocromil on the response to mannitol for the entire group was 55%. The total dose of mannitol administered was significantly greater after nedocromil (550 ± 157 mg) compared with placebo (315 ± 206 mg) (p < 0.001) even though the maximal percent fall in FEV₁ after nedocromil was significantly less (14.1 ± 8.6%) than placebo (21.5 ± 5.9%) (p < 0.0001).

For many subjects a plateau was observed in response to mannitol after administration of nedocromil. A plateau was recorded in 11 of 12 subjects who became negative to mannitol after nedocromil; however, it was also observed in three of 12 subjects who remained responsive to mannitol after nedocromil.

There was no significant difference in baseline FEV₁ values on the placebo (91.1 ± 6.8%) and nedocromil (91.2 ± 8.6%) (p = 0.09) study days and the variability between FEV₁ was less than 10.5% in all subjects. There was also no difference in FEV₁ after both treatments with 90.4 ± 7.2% predicted after placebo and 89.4 ± 7.7% predicted after nedocromil (p = 0.3). Thus, the FEV₁ recorded immediately before challenge was no different on the two test days.

Eight subjects had some protection afforded by the placebo. In six subjects this was observed to be greater than the repeatability of the response. Two subjects were no longer responsive to mannitol after placebo. In these eight subjects there was no significant difference in baseline FEV₁ values either before (p = 0.22) or after (p = 0.12) administration of placebo compared with the control day.

	DISCUSSION

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This study clearly demonstrates that, in asthmatic subjects, nedocromil sodium inhibits the airway response to inhaled mannitol when 8 mg is administered via a metered-dose inhaler 10 min before challenge. The protection afforded by a single dose of nedocromil against the airway narrowing provoked by mannitol is similar to that observed against other stimuli such as exercise (1), cold air hyperventilation (2), hypertonic saline (3), and distilled water (4). Airway narrowing to airway drying and osmotic stimuli is thought to act by raising the osmolarity of the airway surface leading to the release of inflammatory mediators from inflammatory cells and sensory nerves (5, 6). This finding supports the hypothesis that inhaled mannitol causes airway narrowing via a similar mechanism to exercise (7). The immediate onset of the protective effect afforded by nedocromil on the response to mannitol is similar to that observed with hyperpnea of dry air and known osmotic stimuli. We have previously suggested that the protective effect of nedocromil on these stimuli may be due to an effect on regulatory volume changes in response to cell shrinkage or swelling (12).

Those subjects remaining responsive to mannitol after nedocromil were the most responsive after placebo. This suggests that a more sensitive response to an osmotic challenge such as mannitol may be a result of more inflammation in the airways. This concept is supported by observations of a strong association between the response to hypertonic saline and the presence of eosinophils and mast cells in sputum of children (13). This suggests that responses to osmotic stimuli such as mannitol may provide an indirect index of airway inflammation.

The inhibitory effect of nedocromil was also demonstrated by changes in the RDR and the percent protection to the highest cumulative dose of mannitol common to both placebo and nedocromil days. There was a significant reduction in the RDR to mannitol after treatment with nedocromil. Two subjects had responses after nedocromil in the range of healthy nonasthmatics (8). The RDR is a useful index in that it allows comparisons to be made in the absence of recording a PD₁₅. Further, RDR can be used to compare responses before and after treatment and to compare responses with those measured in healthy, nonasthmatic subjects. When comparing the protective effect to the highest dose of mannitol commonly given on the placebo and nedocromil study days, the mean protection of 55% observed is similar to the protection obtained by nedocromil on exercise-induced asthma (EIA) when using an inhaled dose 4 to 8 mg immediately before exercise (1). Taking into account the repeatability of the response to mannitol, we did however observe a placebo effect in some subjects in this study. This could not simply be accounted for by change in baseline FEV₁ values or alteration in medication regimen. This was either a "true placebo" effect or the placebo's components may have some activity. Nedocromil was able to limit the extent to which the airways narrowed to mannitol. This confirms previous observations of the effects of nedocromil on challenge with hypertonic saline (3). As with hypertonic saline, a plateau was most likely to occur in subjects with milder bronchial responsiveness.

In conclusion, this study demonstrates that 8 mg of nedocromil sodium administered via a metered-dose inhaler is effective in inhibiting the response to inhaled mannitol in asthmatic subjects. The magnitude of the protection is similar to that observed with exercise, hyperventilation, hypertonic saline, and distilled water and further suggests that inhaled mannitol acts to cause airway narrowing via a similar mechanism. Inhaled mannitol has been shown to identify subjects with EIA (7) and may be a useful test to identify those who will be protected from EIA by nedocromil. Inhaled mannitol was initially developed to overcome the many practical disadvantages of other "indirect challenge" tests. Inhaled mannitol may not only permit point-of-need bronchial provocation testing but also serve to identify potentially useful drugs that can be used acutely to prevent attacks of asthma. Further studies are needed to investigate the association between mannitol responsiveness and airway inflammation and the effect of treatment with inhaled steroids. Such studies would serve to assess the role of inhaled mannitol in the monitoring of asthma therapy.