Fast effectiveness of a solubilized low‐dose budesonide nasal spray in allergic rhinitis

Abstract Background Budesonide, a poorly water‐soluble corticosteroid, is currently marketed as a suspension. Budesolv is a novel aqueous formulation containing dissolved budesonide showing increased local availability in preclinical models. Budesolv contains ~85% less corticosteroid than the marketed comparator. Objective The study (EudraCT:2018‐001324‐19) was designed to assess non‐inferiority of Budesolv compared to Rhinocort® Aqua 64 (RA) and early onset of action. Methods In a three‐way cross‐over double‐blinded randomized trial, Budesolv 10 was compared to RA and placebo in grass pollen allergic rhinoconjunctivitis volunteers (n = 83 (ITT); n = 75 (PP)). On day 1, participants entered the Vienna Challenge Chamber (VCC) for 6 hours; first treatment took place at 1:45 hours after entry. Participants treated themselves for further 6 days; on day 8, the last treatment was applied before entering the VCC. Subjective symptom scores, nasal airflow and nasal secretion were measured regularly during allergen challenge. Results Budesolv 10 was equally effective compared to RA with respect to TNSS and nasal airflow after eight days of treatment with a strongly reduced dose (more than 80% reduction). After first dose, only Budesolv 10 showed a significant reduction of nasal and respiratory symptoms starting 90 minutes (P < .05) and 15 minutes (P < .05) after application onwards, respectively, demonstrating an early onset of efficacy. A clinically significant 1 point reduction in nasal symptom score was reached at 195 minutes (P < .05) after application. Conclusions and clinical relevance The novel preservative‐free, aqueous low‐dose budesonide formulation is highly efficacious even after an initial single treatment. Thus, Budesolv 10 appears to be an effective acute treatment for allergic rhinitis as well as for AR comorbidities like mild asthma and conjunctivitis.


| INTRODUC TI ON
Allergic rhinitis (AR), either perennial or seasonal, is one manifestation of a type I hypersensitivity reaction caused by an immune reaction to otherwise innocuous agents such as pollen or house dust mites. In the past decades, a globally rising trend of AR has been observed with widely varying prevalence particularly in the developing countries. Up to one-quarter of the global population may be affected. 1 While allergic rhinitis refers to an inflammatory process of the nasal passages, symptoms involve the nose and may extend beyond to affect the eyes, ears, sinuses and bronchi. Commonly reported nasal symptoms include nasal itching and congestion, runny nose and sneezing. Often, AR will involve the conjunctivae; such patients may experience itching, tearing and red eyes. About 40% of AR patients also suffer from asthmatic symptoms like cough, wheeze and dyspnoea. 2 In fact, AR is considered an independent risk factor for subsequent asthma. 3 Apart from allergen avoidance and physical measures, current therapy of AR comprises two main treatment options: allergen immunotherapy or pharmaceuticals targeting the consequences of mast cell mediator release. While immunotherapy is the only treatment with a long-term sustained effect that is intended to result in a reduced reactivity to the respective allergen, pharmaceutical interventions either block histamine from binding to its receptor (anti-histamines), stabilize mast cells (eg cromoglycate), or reduce the release of pro-inflammatory mediators such as TNF-alpha. The latter is achieved with the topical application of corticosteroids such as budesonide, fluticasone propionate or others. Corticosteroids are synthetic analogues of cortisol and bind intracellular receptors resulting in the anti-inflammatory action. After binding to the drug, the receptor molecules are transferred into the nucleus of the cells and bind to promoter regions of target genes. In case of pro-inflammatory mediators, binding of a glucocorticoid receptor to the promoter induces a reduction of gene transcription into mRNA and thereby a reduction of synthesis and release of such mediators.
Corticosteroids are potent substances, but their intranasal application is hampered by the fact that new generation corticosteroids are highly hydrophobic and therefore generally applied as suspensions.
Budesonide is a steroid widely used in the indications of AR and allergic asthma. It is well established that the use of steroids is beneficial for patients suffering from AR, because adequate treatment provides disease control preventing the worsening of the underlying disease. While first-generation steroids, such as dexamethasone phosphate, are soluble in aqueous media including plasma and therefore exert systemic side effects, newer drugs including budesonide, fluticasone propionate and others are hardly soluble in water and therefore detected only in minimal amounts systemically. 4 In addition, budesonide is inactivated by the liver during the first passage by degradation via the cytochrome pathway. The low solubility, and the application of the drugs as suspensions as a consequence thereof, results in a lag time of up to several days until efficacy can be observed, a situation that is unsatisfactory for patients suffering from allergic symptoms. The poor bioavailability of suspended drugs is also illustrated by a study where different doses of budesonide in suspension (64 µg vs 256 µg per day) have been applied in patients suffering from AR: both doses were similarly effective suggesting that increased amounts of undissolved drug do not contribute efficiently to the clinical response. 5 Different combinations of corticosteroids and anti-histamines have been evaluated in clinical trials, showing superior efficacy compared to either of the components alone, 6-8 but for only one of these combinations a market authorization and real-world evidence exist yet. 9,10 The lag time is shortened, as the anti-histamines exert their function early on, while the corticosteroids address the inflammation later. The corticosteroid in these combinations is still presented as suspension, and thereby, most of the applied drug is not active in the nasal cavity but swallowed and transferred into the system. 4 Another pharmaceutical development strategy is increasing the solubility of corticosteroids. Actually, Captisol, a β-cyclodextrin, was used to solubilize budesonide in a nasal formulation. In two environmental challenge chamber studies, therapeutic equivalence of both nasal sprays, solubilized budesonide (640 µg/mL) nasal spray and marketed suspended nasal spray Rhinocort® aqua 64 was tested; in a pooled analysis, equivalence of dissolved and suspended budesonide in the same concentration was demonstrated. 11 Recently, a novel proprietary combination of known excipients was developed that allows the solubilization of budesonide. 12 The new budesonide formulation has been tested preclinically in vitro, ex vivo and in vivo with respect to permeation into cells and tissue, as well as effectivity in a lung inflammation model in comparison with the marketed product Rhinocort® Aqua 64 nasal spray (RA).
Although the concentration of budesonide in this new formulation is much lower than in RA, a significant decrease of inflammatory mediator release compared to placebo or marketed product could be shown in an acute lung inflammation model, if applied 24 hours, 18 hours or 3 hours before an LPS challenge [Nakowitsch et al, manuscript in preparation]. More importantly, the solubilized budesonide formulation also significantly reduced TNF-alpha in bronchoalveolar lavage, if applied 15 minutes after the challenge compared to placebo and marketed product, reflecting the fast availability of the drug in the target tissue. 12 The aim of the current study was to verify the promising preclinical results shown for Budesolv in a pivotal clinical trial. We hypothesize rapid onset of action of Budesolv with regard to individual K E Y W O R D S allergic rhinitis, budesonide, challenge chamber, early onset, inflammation, intranasal glucocorticoids, nasal spray, preservative-free symptom scores in patients with AR as well as equal efficacy in prolonged treatment compared to marketed product. Furthermore, they had a positive subjective response to a grass pollen challenge during a screening run. Detailed information on inclusion and exclusion criteria is given in the supplemental materials.

| Study design
This was a randomized, double-blind, placebo-controlled threeway cross-over study to evaluate a non-inferior therapeutic effect of Budesolv 10 compared to RA after 8 days of treatment.
A washout interval of 3 weeks was kept between treatment sequences. According to the patient information leaflet of RA, a dose of 256 µg/d is needed to reach maximal therapeutic effect after 7-14 days of treatment. Therefore, to demonstrate non-inferiority to RA, participants were treated for 8 days. Furthermore, an early onset of action was evaluated after application of the first dose.
The study was conducted between October 2018 and April 2019.
One to four weeks prior to first treatment period, each volunteer underwent a 6-hour screening grass pollen challenge session in the Vienna Challenge Chamber (VCC) to ensure a TNSS of at least 6 points out of 12 within the first two hours and a persistent response to the challenge. On day 1 of the first treatment period, eligible volunteers were randomized to one of the three treatment sequences (Placebo: Budesolv 10: RA = 1:1:1). For determination of onset of action, participants entered the VCC for 6 hours and received their first treatment 1 hour and 45 minutes after challenge start to assess the onset and the magnitude of symptom relief after first dose. The treatments were given as two actuations of 50 µL each per nostril and continued at home for further 6 days in the morning. On day 8 of each treatment period, participants applied their last treatment one hour prior to a 6-hour grass pollen allergy challenge ( Figure 1A).

| Total nasal symptom score (TNSS)
TNSS is the sum of the single symptom scores of nasal congestion, rhinorrhoea, nasal itching and sneezing. Each single symptom was scored on a four-point categorical scale from 0 to 3 (where 0 = absent symptoms, 1 = mild symptoms, 2 = moderate symptoms, and 3 = severe symptoms) giving a TNSS from 0 to 12.

| Total ocular symptom score (TOSS)
TOSS is the sum of the single symptom scores of conjunctival redness, itchiness and tearing. Each single symptom was scored on a four-point categorical scale from 0 to 3 (where 0 = absent symptoms, 1 = mild symptoms, 2 = moderate symptoms, and 3 = severe symptoms), giving a total ocular symptom score (TOSS) from 0 to 9.
The score represented an average for both eyes. F I G U R E 1 A, Study outline of the phase III clinical study on days 1 and 8 comparing Budesolv with Rhinocort® Aqua 64 and placebo. On day 1, volunteers were exposed to grass pollen in an environmental challenge chamber for six hours. After 1.45-2 hours, participants received the first treatment (2 puffs per nostril, ie 200 µL total) resulting in a residual observation period of 4.15 hours. On day 8, after eight days of treatment, participants were exposed to grass pollen for six hours. 1 hour prior exposure, the last dosing of the nasal spray was applied. At both days, subjective symptoms were recorded every 15 minutes (orange dots), rhinomanometry (grey dots), and nasal secretion was evaluated every hour or 30 minutes, respectively. B, Study populations. Ninety-nine subjects were screened resulting in 92 volunteers being eligible for the safety population. Eighty-three volunteers started the active phase of the study and finished at least one cycle of the trial comprising the intent-to-treat population (ITT). Seventy-five participants were eligible for the per-protocol population (PP)

| Total respiratory score (TRSS)
TRSS is the sum of the single symptom scores of cough, wheeze and dyspnoea. Each single symptom was scored on a four-point categorical scale from 0 to 3 (where 0 = absent symptoms, 1 = mild symptoms, 2 = moderate symptoms, and 3 = severe symptoms), giving a total respiratory symptom score (TRSS) from 0 to 9.

| Evaluation of nasal airflow with active anterior rhinomanometry
Nasal airflow was determined by using active anterior rhinomanometry at a pressure difference of 150 Pascal across the nasal passages (sum of the right and left nostril values) immediately before (baseline) and every 60 minutes during allergen exposure in the VCC, both on day 1 (baseline) and on day 8 of each study period (6 hours). On day 1 of each treatment period, 45 minutes after the first dose of medication a further assessment was done.

| Evaluation of nasal secretion weight
Pre-weight paper tissues were collected every 30 minutes, and nasal secretion weight was determined in grams.

| Safety
Adverse events (AEs) were monitored throughout the entire duration of the study starting from screening after signature of informed consent. Vital signs (heart rate and blood pressure) were measured at the screening visit, and on each allergen challenge day pre-dose and immediately after leaving the challenge chamber. Routine haematology, blood chemistry and urinalysis assessments, and single 12-lead ECGs were assessed at the screening and follow-up visit. FEV1 was measured on all allergen challenge study days pre-challenge, every 2 hours during the session and at the end of the allergen challenge.

| Statistical analysis
A sample size of at least 72 subjects was estimated based on the non-inferiority margin of 15 percent points for the TNSS assuming a standard deviation of 35%-40% and a power of at least 80%.
Considering a dropout rate of 10%-15%, up to 100 subjects were planned to be screened to randomize about 84 volunteers to get at least 72 evaluable participants for analysis.
The primary study end-point was the mean "Total Nasal Symptom Score" (TNSS), calculated as mean of TNSS measured at 17 time points (every 15 minutes) during the grass pollen allergen exposure challenge in the time window of 2 to 6 hours on day 8. The hypothesis to be tested was non-inferiority of Budesolv 10 in comparison with RA. The null hypothesis is defined as The alternative hypothesis is defined formally as Non-inferiority was concluded if the upper limit of the 95% confidence interval of the mean TNSS of Budesolv does not exceed 15% of the upper limit of the 95% confidence interval of the mean TNSS difference of Budesolv compared to the active comparator originating from the mean TNSS of the active comparator (ie the non-inferiority margin).
As secondary study end-point, the onset of action was calculated as mean difference of TNSS to pre-treatment baseline defined as the mean of the three last observations before treatment (ie at 1:15, 1:30 and 1:45 hours after chamber session start) during the first allergen challenge (day 1). Onset of action was assumed when the first time point of the mean TNSS difference between the active treatment and placebo is P < .05. Additional secondary efficacy end-points included mean nasal air flow, mean nasal secretion, mean TOSS and mean TRSS, respectively.
Intention-to-treat (ITT) and per-protocol (PP) populations were predefined. The per-protocol population will assess the effect of treatment on patients who are compliant with the protocol which comprises all patients with data obtained without major protocol deviations and will be the primary population of interest for the non-inferiority analyses.
Testing was carried out with type I error controlled at a twosided alpha level of 0.05 and point estimates with 95% CIs. There was no adjustment for multiplicity testing of the variables. Statistical analysis was performed with R 3.4. 14

| Patient population
A total of 99 grass pollen allergic patients underwent a screening procedure with 92 patients included in a six-hour grass pollen challenge (safety population). A total of 83 patients reached minimum scores of six points within 2 hours and were thereby eligible for the study. Five volunteers did not complete the cycles on day 1, and one additional participant did not complete the cycles on day 8, resulting in an evaluable ITT population of 78 (day 1) and 77 (day 8), respectively. Furthermore, two participants did not show any TNSS reduction in any cycle on day 8 and were therefore regarded as previously not discovered non-responders to steroid treatment and thus excluded from the PP population which then comprised 75 subjects for both, day 1 and day 8 ( Figure 1B).
Exclusion of the non-responding participants was decided before unblinding and was in accordance with the predefined exclusion criteria. Results were calculated for both study populations (ITT and PP), but data presented as figures for early onset of action (day 1) are presented for the ITT population, whereas data presented as figures for non-inferiority (day 8) are shown for the PP population (PP; Figure 1B). This approach is in accordance with FDA Guidance for Industry for non-inferiority clinical trials to establish effectiveness. 15 Demographic data of all randomized volunteers (N = 83) ( Table 1) show that 60% of the participants were female and 40% were male.
The median age was 31 years, and the median body mass index was 22.84 kg/m 2 .

| Budesolv is equally effective compared to the marketed comparator with respect to TNSS after eight days of treatment
In Figure 2A

| Nasal airflow and nasal secretion support the effectiveness of Budesolv
As an objective parameter to demonstrate non-inferiority between Budesolv and RA as well as superiority compared to placebo, an-

| Budesolv is superior compared to the marketed comparator and significantly reduces respiratory symptoms (TRSS)
In Figure 3A, left panel, the time course of the means of TRSS is shown for each study medication. The mean value between 2 hours and 6 hours determined for Budesolv was significantly lower compared to the mean value of placebo (P = .02 for both PP and ITT,   The blue-shaded area shows the time period applicable for the evaluation of the primary end-point (2-6 hours). *** means significant difference to placebo with P < .001

| Both active treatments significantly reduce ocular symptoms (TOSS)
As for TRSS, the time course of the mean values of TOSS is shown in Figure 3B, left panel. Both active treatments showed a significant reduction of mean TOSS between 2 hours and 6 hours (P = .01 and P = .04 (PP) and P = .02 and 0.04 (ITT), for Budesolv and RA, respectively; compare Figure 3B, right panel). There was no significant difference in mean TOSS between Budesolv and RA (P > .05).

| Safety
The safety population consisted of 92 subjects. The assessment of safety does not show any significant differences between the three treatment groups with respect to severity of AEs, drug relationship or action required (P > .05, compare Table 2). In the Budesolv 10 group, 3 possible related and 1 unlikely related AE occurred whereas in the RA group 1 possible related and 3 unlikely related AEs were found. In the placebo group, 1 possible related and 3 probable related AEs were seen (Table S1). hardly showed any short-term differences (within the first 12 hours after application), suggesting that the dissolved part of the formulation is relevant for the response. 5 In a comparison of 128 µg to 256 µg budesonide per day over four to six weeks, a slightly better response was observed for the higher dose; however, both concentrations resulted in a substantial or total control of the symptoms in 88.4% or 85.3% of treated patients, respectively. In the same study, fluticasone propionate was tested (200 µg per day), which showed a consistently lower responses than 256 µg/d

| D ISCUSS I ON
Budesonide. 16 This is of particular interest, as fluticasone has a Note: The number of AEs during screening and in the different treatment groups has been assessed with respect to severity, drug relationship and action required.
higher affinity to the glucocorticoid receptor and a stronger activation capacity to induce corticosteroid-dependent mRNA transcription than budesonide. 17,18 This higher activity, however, does not translate into higher activity in patients, most probably because fluticasone is even less bioavailable than budesonide. These data together suggest that the limiting factor for efficacy is solubility of the drug and the resulting bioavailability in the respective tissue on the one hand, as well as the availability of the respective receptor in the target tissue. shown that approximately 30% of the applied drug is found in the plasma. 18,19 As Budesolv only contains less than 16% of budesonide  rhinitis. AR is considered an independent risk factor for asthma 27 with an adjusted odds ratio of 3.21. 3 Some studies have shown that treatment for AR can reduce healthcare costs and lead to a better asthma control. 28,29 In the future, the challenge of AR care will be to optimize care pathways leading to a higher level of symptom control and prevent the progression towards asthma. Given the relationship between allergic rhinitis and asthma, it can be hypothesized that reducing inflammation in the upper airway with intranasal corticosteroid medications may have a positive effect on the onset of asthma. In fact, a systematic review of that subject found that intranasal corticosteroids improve some asthma-specific outcome measures in patients suffering from both allergic rhinitis and asthma. 30 After a treatment period of at least 4 weeks, the six pooled trials reported a significant improvement in respiratory symptom scores of 0.42 (95% CI; -0.53-0.03). In our study, we found a reduction in total respiratory symptom score of 0.25 (95% CI; -0.45-0.06) score points after an initial single treatment with Budesolv 10 when all participants were evaluated (reduction of 45% compared to placebo). If only participants showing any respiratory symptoms (ITT resp, n = 57) were considered, the respiratory symptom score was reduced by 0.51 score points (reduction of 47% compared to placebo); if only volunteers that showed a TRSS of more than 1 score point were included for analysis (n = 38), the TNSS reduction was as high as 0.9 score points. Our hypothesis for the somewhat unexpected finding of a fast and strong suppression of respiratory symptoms by the application of Budesolv 10 is the increased availability of budesonide in the pharyngeal and upper airway mucosa of the respiratory tract, where reduction in oedema and irritation could have led to reduced subjective respiratory symptoms. This hypothesis was supported by the fact that total ocular symptom score was not affected by Budesolv 10 treatment on day 1. The interaction between upper and lower airways has been discussed repeatedly in the literature, and several potential mechanisms can be considered accordingly: a nasobronchial reflex was described several decades ago already showing that nasal provocation induces an immediate bronchial reaction as well. 31,32 A comparable interaction could be proven for effector cell markers when a nasal provocation induced upregulation of ICAM-1, VCAM-1 and E-selectin also in the lower airways 33 or a segmental bronchoprovocation in non-asthmatic allergic rhinitis patients led to mast cell degranulation and increase of basophils in the nasal mucosa, too. 34 To conclude, Budesolv 10, the novel aqueous formulation of the corticosteroid budesonide, demonstrated non-inferiority to RA in this pivotal phase III study. The data showed that solubilization of budesonide allows sparing of more than 84% of drug relative to the originator product RA, while still resulting in a non-inferior clinical outcome with respect to TNSS on day 8, the primary end-point.
Consequently, the total exposure of patients to the drug was drastically reduced. The aqueous formulation is also free of preservatives such as benzalkonium chloride, paraben or phenyl-ethyl-alcohol, which reportedly have negative effects in the nasal cavity. 35,36 Moreover, treatment with Budesolv 10 led to a fast relief of allergic rhinitis symptoms, reported as TNSS. In addition, Budesolv 10 appears to be the first intranasal formulation of budesonide that reveals a significant effect on respiratory symptoms, exemplified by cough, wheezing and dyspnoea. Thus, Budesolv 10 can be a promising new option for the acute therapeutic treatment of seasonal and perennial allergic rhinitis with or without AR comorbidities like asthma and conjunctivitis.

CO N FLI C T O F I NTE R E S T
The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: PZ, RZ, RS, PL and MS declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. NU, AG, EP and SN are employees of Marinomed.