Phase I/II Study of PRO044 in Duchenne Muscular Dystrophy (DMD): Complete Statistical Analysis

1. Trial at a Glance

This page covers the registry results for Phase I/II Study of PRO044 in Duchenne Muscular Dystrophy (DMD) (NCT01037309), a non-randomized phase 1, phase 2 study. The primary outcome is Increase in Dystrophin Expression in the Muscle Biopsies by Immunofluorescence Analyses of Cross-sections and by Western Blot Analyses of Total Protein Extracts.

2. Clinical Question

These features frame the question as an early assessment of PRO044 across administration routes and dose levels, with dystrophin expression as the stated primary measure. The record also lists a pharmacokinetic secondary outcome: Cmax following subcutaneous administration. It does not give a randomized treatment contrast, so differences among cohorts cannot be read as unbiased comparisons of dose or route. The study description includes safety, tolerability and pharmacokinetics among its aims, but the posted information does not establish what conclusions were reached for those aims.

3. Trial Design

This completed study is titled as a Phase I/IIa, open-label, escalating-dose pilot of PRO044 in Duchenne muscular dystrophy. The registry lists phases I and II, non-randomized allocation, no masking and a parallel model. It records a start of 2009-12 and primary completion of 2013-05. The enrollment listed in the registry is 18.

Nine cohorts are described. Cohorts 1 through 6 received subcutaneous PRO044: cohort 1 received 0.5 mg/kg, and cohorts 2 through 6 received maximally 1.5 mg/kg, 5 mg/kg, 8 mg/kg, 10 mg/kg and 12 mg/kg, respectively. Cohorts 7 through 9 received intravenous PRO044, at maximally 1.5 mg/kg, 5 mg/kg and 8 mg/kg, respectively. The regimen descriptions specify injections on day 1, 8, 15, 22 and 29. The cohort descriptions do not establish that every participant received the maximum dose stated for a cohort.

Because allocation was non-randomized and masking was absent, the record does not support treating these cohorts as comparable randomized groups. Differences in participant characteristics, cohort conduct or other factors could contribute to differences between cohorts; details about such factors are not reported in the registry record. The dose-escalation format is suitable for describing an early pilot, but it does not itself provide a controlled estimate of a treatment effect. The registry record does not report a separate control arm, a randomization procedure or a masking procedure.

The recorded enrollment and cohort-level safety at-risk counts should be read as separately reported registry entries. The registry record does not explain their relationship. Further details about recruitment, participant flow and the reasons participants were included in particular cohorts are not reported in the registry record.

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.

Before assignment: 3 Subjects were recruited to each of cohorts 1-6 (total n=18) for SC administration PRO044 9 of these 18 subjects were recruited to cohorts 7-9 (n=3 in each cohort) for IV adminstration PRO044

Only the first 6 of 9 groups are shown.

Subcutaneous Cohort 1 - 6

Intravenous Period 7 - 9

Analysis population for the primary outcome, as defined in the registry: For some participants it was not possible to determine dystrophin expression in muscle biopsy

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.

Only the first 6 of 7 groups are shown.

6. Primary Endpoint

The registry lists the primary outcome as Increase in Dystrophin Expression in the Muscle Biopsies by Immunofluorescence Analyses of Cross-sections and by Western Blot Analyses of Total Protein Extracts, measured over Within 13 weeks after 5 weeks of treatment.

The primary endpoint concerns the number of participants with an increase in dystrophin expression in muscle biopsies. The registry describes assessment by immunofluorescence analyses of biopsy cross-sections and Western blot analyses of total protein extracts. The stated timeframe is within 13 weeks after 5 weeks of treatment.

This endpoint measures a change in the amount or presence of a protein in sampled muscle tissue, rather than a clinical function or symptom. In general, measuring the protein of interest can address whether an intervention is associated with a biological change relevant to the disease being studied. That makes the measure informative for an early study focused on dystrophin expression. It does not, by itself, establish whether participants experience a functional or clinical change.

The registry record notes that it was not possible to determine dystrophin expression in muscle biopsy for some participants. It does not report how many participants had an assessable biopsy, how an increase was defined, whether both laboratory methods had to agree, or how results from the methods were combined. A numerical count of participants meeting the endpoint and an estimate of uncertainty are not reported in the registry record. Without those details, the endpoint definition can be described, but its observed result and precision cannot be evaluated.

7. Why No Re-analysis Is Shown

8. Statistical Interpretation

9. Statistical Methods Explained

The registry describes the primary endpoint as a participant count: participants with an increase in dystrophin expression in muscle biopsies. This is a binary outcome at the participant level: each participant would be classified according to whether the endpoint was met, if the measurement was available and the endpoint rule was applied. The registry does not state the operational rule for an increase, so the threshold and any decision process cannot be assessed.

What can a participant count tell us?

In general, a count conveys how many observations fall into a defined category. To interpret that count as a rate, the relevant denominator must also be known: the number of participants whose outcome was assessed under the stated rule. A proportion can help describe how common the event was in the analyzed group, but it does not remove uncertainty, especially when the group is small. The registry record does not provide a primary endpoint count or an assessed denominator, so a primary endpoint proportion cannot be interpreted from the recorded information.

The primary measure uses muscle biopsy assessments by immunofluorescence and Western blot. These are different laboratory approaches to assessing dystrophin expression. In general, multiple measurement methods can provide complementary information, but a combined endpoint needs a clear rule for how results are classified when methods differ or a measurement is unavailable. That rule is not reported in the registry record. Nor does the record describe laboratory blinding, repeat testing, or how laboratory measurements were converted into the participant-level endpoint.

Why does the design matter for comparisons?

Randomization can help balance measured and unmeasured participant characteristics across treatment groups, while masking can reduce the influence of knowledge of treatment assignment on assessments or decisions. Those design features were not used according to the registry record: allocation was non-randomized and masking was none. This does not prevent a descriptive account of what was recorded in each cohort, but it limits the interpretation of differences between cohorts. A cohort contrast could reflect differences in participants, conduct or measurement as well as the treatment regimen.

The parallel model indicates that the record organizes participants into separate cohorts rather than describing a crossover comparison. The cohorts differ by route and dose. Because there is no stated randomized control comparison, a direct comparison between cohorts should not be treated as the effect of changing dose or route. General statistical adjustment can address measured differences only when the required participant-level information and a suitable analysis are available; such an analysis is not reported in the registry record.

What does an analysis add?

An inferential analysis typically combines an estimated effect with a measure of uncertainty, such as a confidence interval, and may also report a test of a specified statistical hypothesis. In general, a confidence interval communicates how precisely the data estimate an effect under the model and assumptions used. A p-value, when used, summarizes the compatibility of the data with a null hypothesis under those assumptions; it does not measure the probability that a treatment works. The registry record has no analysis records, so an estimated primary effect, confidence interval and p-value are not available to interpret.

The registry record lists three outcome measures, but does not report a statistical analysis plan. It is therefore not possible to establish which outcomes were designated for formal hypothesis testing, what estimands or contrasts were intended, or whether any model was prespecified. The record also does not describe adjustment for baseline characteristics, handling of repeated or incomplete measurements, or any interim analyses. These details are not reported in the registry record.

How should planning illustrations be read?

General sample-size planning uses assumptions about the effect to detect, the significance level, allocation, and the statistical model. The planning assumptions associated with the posted results specify a two-sided significance level of 0.05, equal allocation between two arms, a normal approximation, and an effect defined as a stated fraction of the observed effect. They are described as an illustration based on the observed effect, not as the trial's own design. The record does not report a sample-size justification for the study itself.

A planning calculation is not an observed result and does not tell readers whether the study detected an effect. It is sensitive to its assumptions, including the estimated effect used as a starting point. When the design is non-randomized and lacks a like-for-like control comparison, an observed difference may not represent a treatment effect, so an illustration based on such an effect requires particular caution. It should not be described as observed power or as proof that the study had adequate precision.

For binary outcomes, the observed count, its denominator, and the definition of the event are all needed for meaningful interpretation. For time-to-event outcomes, event counts and follow-up information are central; however, the primary endpoint in this study is a participant count, not a time-to-event measure. No time-to-event analysis is described in the registry record.

10. Exact Estimates for Each Arm

No two-arm comparison is shown for this trial, but each arm's result can still be summarized with exact interval estimates computed from the posted counts. The Clopper-Pearson interval is exact and conservative; the Wilson interval is a common closer-to-nominal alternative; the Bayesian credible interval uses a uniform Beta(1, 1) prior. For small groups the three can differ noticeably.

R code and output

arm    <- c("PRO044, Cohort 1", "PRO044, Cohort 2", "PRO044, Cohort 3", "PRO044, Cohort 4", "PRO044, Cohort 5", "PRO044, Cohort 6")
n      <- c(1, 1, 3, 3, 2, 3)
events <- c(1, 1, 0, 1, 1, 2)
for (i in seq_along(n)) {
  ci <- binom.test(events[i], n[i])$conf.int          # Clopper-Pearson exact interval
  cr <- qbeta(c(0.025, 0.975), 1 + events[i], 1 + n[i] - events[i])   # Bayesian, uniform Beta(1,1) prior
  wi <- (function(x, n) { p <- x / n; z <- qnorm(0.975); d <- 1 + z^2 / n
    c((p + z^2 / (2 * n) - z * sqrt(p * (1 - p) / n + z^2 / (4 * n^2))) / d, (p + z^2 / (2 * n) + z * sqrt(p * (1 - p) / n + z^2 / (4 * n^2))) / d) })(events[i], n[i])
  cat(sprintf("ARM|%s|%d|%d|%.1f|%.1f|%.1f|%.1f|%.1f|%.1f|%.1f\n", arm[i], n[i], events[i], 100 * events[i] / n[i],
              100 * ci[1], 100 * ci[2], 100 * wi[1], 100 * wi[2], 100 * cr[1], 100 * cr[2]))
}

ARM  PRO044, Cohort 1  1  1  100.0  2.5  100.0  20.7  100.0  15.8  98.7
ARM  PRO044, Cohort 2  1  1  100.0  2.5  100.0  20.7  100.0  15.8  98.7
ARM  PRO044, Cohort 3  3  0  0.0  0.0  70.8  0.0  56.1  0.6  60.2
ARM  PRO044, Cohort 4  3  1  33.3  0.8  90.6  6.1  79.2  6.8  80.6
ARM  PRO044, Cohort 5  2  1  50.0  1.3  98.7  9.5  90.5  9.4  90.6
ARM  PRO044, Cohort 6  3  2  66.7  9.4  99.2  20.8  93.9  19.4  93.2

11. Multiplicity, Missing Data and Analysis Populations

The registry lists three outcome measures, while the primary endpoint is the participant count for increased dystrophin expression in muscle biopsies. The record does not report how many outcomes were tested statistically, whether one analysis was designated as primary for inference, or whether any adjustment for multiple testing was planned. This is not reported in the registry record. Without an analysis plan, the possibility of selective emphasis among several outcomes cannot be evaluated from the endpoint list alone.

Missingness is directly relevant to the primary endpoint: the registry states that dystrophin expression could not be determined in muscle biopsy for some participants. It does not report the number affected, why measurement was not possible, whether missingness differed by cohort, or how unavailable results were handled. It also does not identify an analysis population, such as all enrolled participants or only those with an assessable biopsy. The result therefore cannot be reconstructed from the endpoint definition.

In general, excluding participants with unavailable outcomes can affect an estimate if the chance of missing measurement is related to participant characteristics or outcomes. Imputation or sensitivity analyses can sometimes examine the implications of missing data, but their appropriateness depends on the pattern and reasons for missingness. Neither an imputation approach nor a sensitivity analysis is reported in the registry record.

The record reports no analysis entries. Details about prespecified hypotheses, analysis populations, multiplicity control, model choice and missing-data procedures are not reported in the registry record. The absence of these details means the posted endpoint description should not be mistaken for a complete account of how primary and secondary outcomes were analyzed.

12. Safety Results

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

Other (non-serious) adverse events

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

Most frequent serious adverse event terms

Most frequent non-serious adverse event terms

Serious events with the highest arm-level rate: concussion (33.3%), international normalised ratio increased (33.3%), activated partial thromboplastin time prolonged (33.3%), orthostatic proteinuria (33.3%).

The serious adverse event entries are reported by cohort. Each cohort lists three participants at risk. The number recorded as affected by serious adverse events is 1 in cohort 1, 0 in cohort 2, 1 in cohort 3, and 0 in cohorts 4, 5, 6, 7, 8 and 9. The record lists other adverse events in three participants in cohorts 1, 2, 3, 4, 5, 6, 8 and 9, and in two participants in cohort 7.

The registry lists Concussion, International normalised ratio increased, activated partial thromboplastin time prolonged and orthostatic proteinuria as top serious-event terms, each with 33.3%. The registry record does not clarify the denominator or cohort distribution for these term percentages. These event counts and percentages are descriptive; the non-randomized cohorts are not a controlled comparison of safety. Death counts are not reported in the registry record, which should not be interpreted as a statement that no deaths occurred.

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

The record establishes that a completed, open-label, non-randomized pilot evaluated PRO044 in participants with Duchenne muscular dystrophy, using different subcutaneous and intravenous regimens across cohorts. It also specifies a primary biological endpoint: increased dystrophin expression in muscle biopsies, assessed by immunofluorescence and Western blot within the stated timeframe. These details describe the study question and measurement approach.

The posted information does not report a numerical result for the primary endpoint. It therefore cannot establish how many participants had an increase or how precise an estimate might be. The note that expression could not be determined for some participants adds an important qualification, but the number and handling of those cases are not reported. The endpoint definition is not itself evidence that expression increased.

The cohort-specific safety counts describe events recorded in the registry, but do not establish that PRO044 caused those events or that one regimen is safer than another. Non-randomized allocation and the lack of a control comparison leave potential differences between cohorts unaccounted for. The serious-event term percentages also lack a stated denominator in the registry record. These entries should therefore be read as descriptive reports, not comparative risk estimates.

More generally, a biological measure does not by itself establish a change in clinical function or patient experience. The primary endpoint focuses on dystrophin expression in tissue; the registry record does not report a numerical result linking that measure to a clinical outcome. The record also does not give the statistical plan, the analysis populations, or a sample-size justification. Consequently, the posted information supports describing the design and recorded outcome definitions, but not a quantitative conclusion about efficacy, precision, or comparative safety.

14. Why This Trial Matters Statistically

This study is statistically informative as an example of the limits and uses of an early, non-randomized dose-escalation design. Its structure allows the registry to document cohorts receiving different routes and dose levels. Such descriptive information can be useful for understanding how an early pilot was organized. It does not provide the same basis for estimating treatment effects as a randomized comparison with a suitable control group.

The primary endpoint also illustrates the distinction between a measurable biological outcome and a clinical outcome. Dystrophin expression in muscle tissue is the stated focus, and the record describes two laboratory approaches. For statistical interpretation, however, the endpoint needs an explicit classification rule, an assessable denominator and a reported result. The registry notes that some biopsies could not yield a determination, but does not report how many or how those observations were handled.

The safety entries show why counts need context. Cohort-specific counts and listed event terms can describe what was recorded, but their interpretation depends on denominators, follow-up and how events were collected. The registry record does not report enough detail to support a controlled comparison of safety across regimens. The absence of analysis records also means that readers cannot evaluate estimated effects or their uncertainty.

Finally, the distinction between study-design planning and retrospective planning illustrations matters. A calculation based on an observed effect is not the same as a prospective justification, and it is not evidence of observed power. In a non-randomized pilot, the interpretation of an observed effect is itself constrained by the absence of a randomized control comparison. These features make careful separation of endpoint definition, observed data and statistical inference especially important.

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 registry record describes a non-randomized, open-label study without a stated control arm. Cohorts differ in administration route and dose, so comparisons between them may reflect participant or study differences as well as regimen differences. The record does not report participant characteristics by cohort, allocation details, or procedures to address potential confounding.

The primary endpoint is not fully operationalized in the record. The criterion for an increase in dystrophin expression, the relationship between immunofluorescence and Western blot results, and procedures for discordant or unavailable measurements are not reported. The record says that expression could not be determined for some participants, but gives no count, reasons, or missing-data handling approach.

No numerical primary endpoint result or analysis record is included. Statistical methods, analysis populations, multiplicity procedures and sample-size justification are not reported in the registry record. The listed enrollment and cohort-level safety at-risk counts are both recorded, but their relationship is not explained. The registry also does not report a denominator for the listed serious-event term percentages or death counts.

These limitations restrict assessment of efficacy, precision, missing-data bias and comparative safety. They do not, by themselves, establish that the study measurements were invalid; rather, the registry information does not allow those questions to be resolved.

16. Related Tutorials

17. Related Statistical Calculators

18. Sources

• ClinicalTrials.gov: Phase I/II Study of PRO044 in Duchenne Muscular Dystrophy (DMD) — NCT01037309.
• Registry record last updated 2018-10-16; status last verified 2018-09.
• Results first posted 2015-04-09.
• 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

The registry record describes a completed Phase I/IIa open-label, non-randomized, parallel pilot study of PRO044 in Duchenne muscular dystrophy. It lists an enrollment of 18, a start of 2009-12 and primary completion of 2013-05. Nine cohorts are described, with subcutaneous dosing in cohorts 1 through 6 and intravenous dosing in cohorts 7 through 9.

The primary endpoint was the number of participants with increased dystrophin expression in muscle biopsies, assessed by immunofluorescence and Western blot within 13 weeks after 5 weeks of treatment. The record notes that expression could not be determined for some participants. A numerical primary endpoint result and analysis records are not reported in the registry record. Serious and other adverse event counts are recorded by cohort, but do not support a controlled comparison across regimens.