Crossover Trial to Assess Efficacy and Safety of Inhaled AQ001S Compared to a Budesonide Suspension in Mild Asthmatics: Complete Statistical Analysis

1. Trial at a Glance

This page covers the registry results for Crossover Trial to Assess Efficacy and Safety of Inhaled AQ001S Compared to a Budesonide Suspension in Mild Asthmatics (NCT04933383), a randomized phase 1, phase 2 study. The primary outcome is Change From Baseline in PC20 After Each Treatment Period Assessed by Methacholine (MCh) Challenge Test.

2. Clinical Question

3. Trial Design

This was a prospective, randomized, active-controlled, open-label, single-center, multiple-dose crossover trial. It was registered as Phase 1 and Phase 2. The record says 23 participants were enrolled and the trial is completed. It lists a two-period crossover: participants received AQ001S and budesonide inhalation suspension in separate treatment periods, with both treatments given by nebulization once daily.

The primary outcome description specifies a 29-day treatment period for each period. PC20 was assessed at baseline, at the end of Treatment Period 1 and at the end of Treatment Period 2. The primary outcome was the change from baseline after each period. The efficacy population required completion of both periods and availability of the PC20 measurement at baseline and after both periods.

A crossover design compares treatments within a participant across periods, rather than assigning each participant to only one treatment throughout the trial. In general, this can account for some stable differences between participants, but interpretation also depends on period effects, treatment sequence and any carryover between periods. The registry record does not report the treatment sequences, their allocation, a washout interval, or analyses of period or carryover effects. It identifies random allocation but does not describe its implementation.

The trial had no masking, so participants and trial personnel were not blinded according to the registry. The record does not report a sample-size justification, recruitment details beyond the enrollment figure, or a detailed protocol for handling departures from the planned treatment sequence. These details are important when judging how the trial was designed to address potential bias in a small crossover study.

4. Participant Flow and Accounting

The registry reports how participants moved through each stage of the study. All counts below are exactly as posted; missing counts are not inferred from other stages.

Overall Study

Analysis population for the primary outcome, as defined in the registry: The efficacy population encompasses all subjects that completed both treatment periods and for which the primary efficacy parameter PC20 was available at baseline (visit 1) and after both treatment periods (visit 2 and visit 4).

5. Baseline Characteristics

The table reproduces the baseline characteristics posted in the registry, which describe the participants analysed at baseline and help show whether the randomized groups were comparable.

Population: The efficacy population encompasses all subjects that completed both treatment periods and for which the primary efficacy parameters PC20 is available at baseline (visit 1) and after both treatment periods (visit 2 and visit 4). The safety population includes all randomized subjects who received at least one dose of the study medication (AQ001S or comparator).

6. Primary Endpoint

The registry lists the primary outcome as Change From Baseline in PC20 After Each Treatment Period Assessed by Methacholine (MCh) Challenge Test, measured over visit 1 (baseline) - visit 2 (at the end of Treatment Period 1) and visit 4 (end of Treatment Period 2).

The primary endpoint was change from baseline in PC20 after each treatment period, assessed by a methacholine challenge test. PC20 is the concentration of methacholine that provokes a 20% fall in FEV1. FEV1 is measured by spirometry. The registry specifies assessment at baseline and at the end of each of the 29-day treatment periods.

As a general explanation, a challenge test measures airway responsiveness under a controlled stimulus. PC20 expresses the concentration associated with a specified fall in FEV1, so changes in PC20 describe a change in the challenge-test response. The endpoint is not itself a direct measure of asthma symptoms, everyday activity or long-term disease course. The registry record does not report a minimum clinically important difference for PC20 or explain what size of change would be considered important to participants.

The use of change from baseline allows each treatment-period measurement to be interpreted relative to the participant’s starting measurement. In a crossover trial, both treatment periods contribute to the comparison, but the statistical interpretation depends on how within-participant measurements and period effects are handled. The record identifies an ANCOVA analysis, but its model specification is not reported in the registry record.

7. Why No Re-analysis Is Shown

8. Statistical Interpretation

9. Statistical Methods Explained

The posted primary outcome is continuous: it is a mean change in PC20, measured in mg/mL. The registry reports a mean and a 95% confidence interval for each treatment group. A mean summarizes the central value of the observations, while a confidence interval conveys uncertainty about that estimated mean under the analysis assumptions. These arm-level intervals should not be mistaken for a direct test of the difference between treatments.

What does the ANCOVA comparison represent?

The registry reports ANCOVA and describes its result as a difference/ratio of least square means. As a general explanation, analysis of covariance estimates a group contrast while accounting for covariates included in the model. Least square means are model-based estimates, rather than necessarily being identical to the raw arithmetic means. The registry record does not report which covariates were included, how baseline PC20 entered the model, how the crossover structure was represented, or whether treatment sequence and period were modeled.

The reported estimate is 180.6, with a 95% confidence interval from 106.42 to 306.48. The record does not clarify whether the result should be read as a ratio, a percentage-like scale, or another transformation of least square means. It is therefore important not to equate this estimate with a difference of 180.6 mg/mL or with the difference between the two arm-level mean changes. The registry’s reported metric and model output are the appropriate description of the comparison, but the missing scale details limit independent interpretation.

Why is a crossover model relevant?

As a general explanation, crossover trials measure participants under more than one treatment, so observations from the same participant are related. A valid analysis ordinarily needs to account for this within-participant structure. Period may also matter if measurements change over time regardless of treatment, and carryover may matter if effects from one treatment persist into another period. The registry identifies the trial as crossover, but does not report the model terms or checks used to account for these features. This is not evidence that they were ignored; their handling is not reported in the registry record.

How should the confidence intervals be read?

The 95% confidence intervals around the treatment-specific means express sampling uncertainty for each mean, conditional on the analysis. Their overlap or non-overlap is not a substitute for the registered treatment comparison, especially in a crossover design where the relevant estimates may depend on paired observations and model adjustments. The ANCOVA interval pertains to the comparison as reported, and it is the more direct inferential result for the trial’s primary question. General statistical interpretation still depends on the model and its assumptions being appropriate.

What does the p-value add?

The reported p-value is 0.001, which is less than 0.001. In general, a p-value measures the degree of compatibility between the observed data and a specified null model. It does not quantify the probability that the treatments are equivalent, the chance that the result will replicate, or the size of a clinically meaningful effect. It is also not a correction for design limitations or missing data. The registry labels the analysis as superiority, but does not report the full hypothesis statement or a detailed statistical analysis plan.

What can planning illustrations tell readers?

Sample-size illustrations based on an observed effect are planning calculations, not observed power and not a record of the trial’s original design rationale. The registry’s planning note describes assumptions of a two-sided significance level of 0.05, equal allocation, a normal approximation and an effect defined as a fraction of the observed effect. Such illustrations depend on assumptions and can be optimistic when based on an effect estimated from a small trial. The registry record does not report the trial’s own sample-size justification, so these illustrations should not be taken as evidence that the trial was planned with those assumptions.

Some values are unavailable or not estimable for a two-arm continuous-endpoint comparison. No recalculated treatment contrast should be inferred from the arm means alone. In general, recalculating a contrast for a crossover study requires information about within-participant variability or paired outcomes; marginal means and their confidence intervals do not provide all of that information. Where a sponsor’s reported analysis is available, it remains the trial’s own model-based result rather than an independently reconstructed contrast.

10. Analyses Reported by the Sponsor

The registry reports the sponsor’s ANCOVA analysis as a difference/ratio of least square means of 180.6, with a 95% confidence interval from 106.42 to 306.48 and p < 0.001. It labels the result as a superiority analysis. This is the reported inferential comparison for the primary outcome.

A separate unadjusted recalculation is not available for this continuous endpoint: the record marks values as missing or not estimable for a two-arm comparison. The arm-level mean changes and their confidence intervals cannot, by themselves, reproduce the ANCOVA result, particularly for a crossover analysis that may depend on paired measurements and model adjustments. There is therefore no independent recalculated estimate to compare numerically with the sponsor’s result.

The sponsor’s own analysis takes precedence when interpreting the reported trial comparison. However, the registry does not give enough detail to reproduce the model or fully establish whether the estimate is a ratio, a transformed value or another expression of least square means. The reported result should be described in the metric used by the registry, with that interpretive limitation made explicit. The statistical result does not, by itself, establish clinical importance.

11. Multiplicity, Missing Data and Analysis Populations

The registry lists 2 outcome measures and 1 analysis. It reports one primary endpoint analysis and no secondary outcomes posted. The record does not report a multiplicity adjustment, a hierarchy of hypotheses, or a plan for controlling the chance of false-positive findings across multiple outcomes or analyses. Absence of this information should not be read as evidence that no such considerations were made; they are not reported in the registry record.

The efficacy population was defined as participants who completed both treatment periods and had PC20 available at baseline and after each period. This is a complete-measurement requirement for the primary outcome. The record does not report the number screened, the number who started each treatment, participant flow through each period, reasons for missing measurements, or the method used to handle missing data. It also does not state whether any sensitivity analyses were performed.

As a general explanation, excluding participants without complete outcome measurements can affect interpretation if the reasons for missingness are related to treatment or outcome. Requiring completion of both crossover periods may also make the analyzed group differ from all enrolled participants. Without participant-level flow and missing-data methods, the extent or direction of any such influence cannot be determined.

The registry reports 23 enrolled participants and 22 in each arm-level efficacy summary. It does not report whether the same participants contributed to both arm summaries, how the crossover analysis treated participants with incomplete sequences, or whether all randomized participants were analyzed. The stated efficacy population definition gives a criterion for inclusion, but not a full accounting of analysis populations.

The record also does not report a prespecified interim-analysis plan, stopping rules, or a statistical analysis plan. It is consequently not possible to establish from the registry record whether the posted endpoint and analysis were selected before results were known. This is a limitation of the available reporting, not proof that prespecification did not occur.

12. Safety Results

The registry reports serious adverse events as affected participants divided by participants at risk.

Deaths

Other (non-serious) adverse events

Participants with at least one non-serious event at or above the registry's 0% reporting threshold.

Most frequent non-serious adverse event terms

The registry reports zero serious adverse events among 23 participants at risk for AQ001S and zero among 23 at risk for budesonide inhalation suspension. It also reports zero deaths in each group. For other adverse events, the record reports 4 affected participants for AQ001S and 2 for budesonide inhalation suspension.

These are participant counts, not a complete description of event types, timing, severity or duration. The list of top serious-event terms is empty. The registry does not report event-level detail, exposure-adjusted rates, a formal comparative safety analysis, or reasons for treatment discontinuation. A count of zero serious events in this trial does not establish that serious events cannot occur; it describes only what was reported in this study.

Because this was a crossover trial, participants could receive both treatments. The at-risk counts are stated separately by treatment, and should not be added to infer a total number of distinct participants. The record does not explain how adverse events were attributed to treatment periods or how events spanning periods were handled. These figures are descriptive and do not support a definitive comparative safety conclusion.

13. What the Result Does, and Does Not, Mean

The result describes a randomized, open-label crossover comparison of AQ001S and budesonide inhalation suspension in adults with mild asthma. The registered primary endpoint was change from baseline in PC20 after each treatment period. The registry reports mean changes for each treatment and an ANCOVA comparison labeled as superiority, with a p-value less than 0.001.

As a general explanation, statistical evidence against a null comparison does not establish that an observed difference is large enough to matter clinically. The registry does not report a threshold for clinically important change in PC20, and the scale of the reported difference/ratio of least square means is not fully explained. The results therefore support a description of the registered statistical finding, but do not independently resolve its clinical meaning.

The finding is specific to the measured challenge-test endpoint, the treatment regimens, the trial population and the treatment periods described in the record. PC20 is not a direct measure of symptoms, day-to-day control, quality of life or long-term outcomes. The trial record does not report results for those outcomes. It also does not establish how well findings generalize beyond adults with mild asthma enrolled in this single-center study.

The record reports 23 enrolled participants and 22 in each primary-outcome arm summary. That scale, together with the requirement for complete measurements across both periods, means the estimate should be read with attention to precision and analysis-population selection. The confidence interval for the ANCOVA result conveys uncertainty under the model, but the unavailable model details constrain independent assessment.

These findings do not determine how treatments should be used for an individual. They are a statistical account of the outcome and safety counts reported in the registry, with important uncertainties about model specification, missing data, clinical thresholds and the broader relevance of the measured endpoint.

14. Why This Trial Matters Statistically

This trial raises statistical issues that are characteristic of crossover studies. Each participant may contribute measurements under both treatments, making within-participant comparisons potentially informative. At the same time, those measurements are related, and the analysis needs to reflect the crossover structure. The registry identifies ANCOVA but does not provide enough model detail to assess how that structure, period effects or treatment sequence were handled.

The endpoint also illustrates the difference between a physiological measurement and a patient-centered outcome. PC20 is defined through a methacholine challenge and a specified fall in FEV1. A statistically persuasive comparison on this measure does not automatically establish a change in symptoms, daily functioning or long-term outcomes; those results are not reported in the registry record.

The record’s arm-level means and the sponsor’s model-based comparison answer related but distinct questions. The means describe treatment-period change summaries. The ANCOVA result is the registered inferential contrast, expressed as a difference/ratio of least square means. Because no recalculated contrast is available and the metric is not fully explained, readers must distinguish the reported model result from what can be derived from the arm summaries.

The efficacy population definition highlights how analysis populations matter. Requiring completion of both periods and available PC20 measurements can make the analyzed participants differ from everyone enrolled. The registry does not report flow or missing-data handling, leaving the effect of exclusions uncertain.

Finally, statistical significance and clinical importance are separate questions. The record reports a low p-value and a confidence interval for the ANCOVA comparison, but does not specify a clinically important PC20 threshold. Careful interpretation therefore depends not just on the statistical result, but on the endpoint’s meaning, analysis assumptions and reporting gaps.

15. Limitations

• The posted summary statistics are the only data these results rest on; patient-level data, full protocols and statistical analysis plans are not available.
• Recalculated results are simple, unadjusted calculations from posted summaries. They do not replace the sponsor's prespecified analysis.

The trial enrolled 23 participants and was conducted at a single center. The primary-outcome arm summaries each include 22 participants, while the record does not explain the difference between enrollment and the analysis counts. Participant flow, reasons for missing measurements and the handling of incomplete treatment periods are not reported in the registry record.

The study was open label. The registry does not describe any steps used to limit the influence of awareness of treatment assignment on outcome assessment or trial conduct. PC20 is assessed through a challenge test, but the record does not provide enough procedural detail to evaluate consistency of testing or assessment across periods.

The crossover design makes treatment sequence, period effects and carryover relevant considerations. The registry does not report the sequence assignments, a washout interval, or model details addressing these issues. It also does not provide the covariates used in ANCOVA or clarify the scale of the difference/ratio of least square means. This limits independent reproduction and interpretation of the reported comparison.

The registry does not report the trial’s sample-size justification, a statistical analysis plan, interim analyses, multiplicity procedures or sensitivity analyses. It does not provide a clinical-importance threshold for PC20. These absences prevent assessment of whether the trial was sized around a prespecified meaningful effect, how multiple analyses were handled, or how robust the result is to alternative assumptions.

The primary outcome is a challenge-test measure rather than a direct report of symptoms or longer-term outcomes. The record does not report secondary outcomes, and its safety information is limited to participant counts without event-level descriptions. Generalization beyond the enrolled adults with mild asthma and the single-center setting cannot be established from this record alone.

16. Related Tutorials

17. Related Statistical Calculators

18. Sources

• ClinicalTrials.gov: Crossover Trial to Assess Efficacy and Safety of Inhaled AQ001S Compared to a Budesonide Suspension in Mild Asthmatics — NCT04933383.
• Registry record last updated 2024-09-03; status last verified 2024-08.
• Results first posted 2024-09-03.
• Data retrieved from ClinicalTrials.gov on 2026-11-09.
• Group sizes, means, event counts and sponsor-reported p-values are reproduced as posted. Effect estimates, confidence intervals and p-values not attributed to the sponsor are independently calculated from the posted summary statistics.

19. Record Summary

NCT04933383 is a completed randomized, open-label, single-center, two-period crossover trial of AQ001S 0.125 mg/ml and budesonide 0.125 mg/ml inhalation suspension in adults with mild asthma. Both treatments were administered once daily by nebulization. The registry reports enrollment of 23 participants.

The primary endpoint was change from baseline in PC20 after each treatment period, measured through methacholine challenge. The reported mean changes were 3.7 mg/mL for AQ001S and 1.2 mg/mL for budesonide inhalation suspension. The registry reports an ANCOVA difference/ratio of least square means of 180.6, with a 95% confidence interval from 106.42 to 306.48 and p < 0.001.

There were zero serious adverse events and zero deaths reported in each treatment group. Other adverse events affected 4 participants for AQ001S and 2 for budesonide inhalation suspension. Details about the analysis model, missing data, treatment sequence and clinical importance are not reported in the registry record.