Safety and Immunogenicity of RNA-based Vaccines Against SARS-CoV-2 Variants in Healthy Participants: Complete Statistical Analysis

1. Trial at a Glance

This page covers the registry results for Safety and Immunogenicity of RNA-based Vaccines Against SARS-CoV-2 Variants in Healthy Participants (NCT05004181), a non-randomized phase 2 study. The primary outcome is All Parts - Percentage of Participants Reporting Local Reactions at the Injection Site (Pain, Tenderness, Erythema/Redness, Induration/Swelling).

2. Clinical Question

The trial considered safety and immune responses in several distinct settings rather than a single uniform comparison. In Parts A and B, cohorts received one or more doses of the multivalent vaccine, a monovalent variant vaccine, or the original vaccine. Part C evaluated a monovalent B.1.1.529.1 vaccine and the original vaccine in RNA COVID-19 vaccine-experienced participants with a history of Omicron infection, and included an observational cohort without vaccination within 3 months after Visit 1.

The registry summary also describes booster vaccination among participants who had received two doses of the parent BNT162b2 vaccine at 30 µg, at least 6 months after the second dose, as well as a three-dose multivalent regimen in participants without prior COVID-19 vaccination. Those distinct histories and schedules matter: observed results across cohorts would not automatically estimate the effect of changing only the vaccine formulation. The stated primary question is descriptive of reported local reactions after injection, not a direct efficacy comparison or a test of protection against infection.

3. Trial Design

This completed Phase II study was sponsored by BioNTech SE and used a non-randomized, unmasked, parallel design. The registry lists enrollment of 1380. The study started on 2021-08-25, with primary completion on 2023-08-16. It was organized into Parts A, B, and C, with cohorts receiving different vaccine formulations, dose schedules, or no vaccination during the specified period.

Parts A and B included cohorts receiving one dose of the multivalent BNT162b2 formulation, one dose of a monovalent formulation, or three doses of the multivalent formulation. Part A also included a cohort receiving two doses of the multivalent formulation and a cohort receiving one dose of the original vaccine. Part C included cohorts receiving one dose of BNT162b2 (B.1.1.529.1) or the original vaccine, plus a cohort receiving no vaccination within 3 months after Visit 1. The listed age ranges varied by cohort: Part A cohorts were described as ages 18 to 55 years, while Parts B and C cohorts were described as ages 18 to 85 years.

The trial’s summary describes additional differences in prior vaccination and infection history. Some participants received a booster after two doses of the parent vaccine, while another group was described as having no prior COVID-19 vaccination. In Part C, the vaccine assessment involved RNA COVID-19 vaccine-experienced participants with a history of Omicron infection, with natural immune response after infection also described as a point of contrast. The exact allocation procedures, cohort sizes for the primary endpoint, and a detailed protocol for these comparisons are not reported in the registry record.

Because allocation was not randomized and masking was absent, the cohorts should not be read as interchangeable arms in a controlled comparison. Differences in prior immunity, age range, vaccine history, infection history, and schedule may distinguish them. The registry record does not report a single common comparator or a design that isolates the effect of vaccine formulation across all parts.

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.

Only the first 6 of 12 groups are shown.

Overall Study

Reasons for not completing this period:

Analysis population for the primary outcome, as defined in the registry: Reactogenicity set, i.e., all participants included in the Safety Set with any reactogenicity data reported after IMP injection. Per protocol, participants in Part C - Cohort 9 did not receive IMP therefore local reactions were not collected for these participants.

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: Part A and B: Safety Set - All participants who received at least one dose in of IMP. 35 participants were randomized to Part C - Cohort 9. As per protocol, these participants did not receive treatment.

Only the first 6 of 13 groups are shown.

6. Primary Endpoint

The registry lists the primary outcome as All Parts - Percentage of Participants Reporting Local Reactions at the Injection Site (Pain, Tenderness, Erythema/Redness, Induration/Swelling), measured over from Day 1 to Day 7 after each IMP dose.

The primary endpoint is the percentage of participants reporting any-grade local reactions at the injection site: pain, tenderness, erythema or redness, and induration or swelling. Reactions were assessed from Day 1 to Day 7 after each investigational medicinal product dose. The registry description says grading was based on criteria from US FDA guidance for preventive vaccine clinical trials, with Grades 1 through 4 ranging from mild to potentially life-threatening. For redness and swelling, a reported size had to be at least 2.5 cm to count as a local reaction; smaller measurements were not included.

This endpoint captures short-term, participant-level reactogenicity after vaccination. Counting participants who experienced a reaction is different from counting every reaction episode: a participant may report more than one type of local reaction, but the endpoint is framed as the percentage of participants reporting reactions. The registry record defines the collection period and threshold but does not report cohort-level endpoint counts, percentages, or uncertainty intervals.

Part C Cohort 9 received no vaccination within 3 months after Visit 1 and was not included in this analysis because local reactions were not collected for participants who did not receive an injection. The stated population is the reactogenicity set: participants in the Safety Set with any reactogenicity data reported after injection. That restriction defines which participants contribute to the endpoint; it does not by itself show how much data were missing among those expected to contribute.

As a general statistical point, a short post-dose window makes the endpoint specific to reactions recorded during that period. It does not describe reactions outside the window or establish that a reported reaction was caused by a vaccine. The registry endpoint is a safety measure, not a measure of immune response or protection from disease.

7. Why No Re-analysis Is Shown

8. Statistical Interpretation

9. Statistical Methods Explained

The study has a binary primary outcome at the participant level: whether a participant reported one or more qualifying local reactions during the stated post-dose period. The registry describes the outcome as a percentage of participants, while its recorded outcome parameter is a participant count. No primary-endpoint result or analysis estimate is reported in the registry information.

What does a participant-level percentage represent?

In general, a participant-level percentage uses the number of participants meeting the endpoint as its numerator and the eligible participants assessed for that endpoint as its denominator. It is not the same as a count of symptoms or episodes. A participant reporting pain and redness may still contribute as one participant with a local reaction under an endpoint framed as participants reporting reactions. The exact aggregation across reaction types and doses is not described in further detail in the registry record.

The stated analysis population is the reactogenicity set: participants in the Safety Set with any reactogenicity data reported after injection. The endpoint excludes Part C Cohort 9 because those participants received no vaccination during the specified period and did not have local reactions collected. For a percentage to be interpretable, the denominator must correspond to participants eligible for and observed for the relevant post-dose assessment. The registry’s population definition helps identify that group, but the number analyzed and handling of incomplete diary data are not reported in the registry record.

Why do collection rules matter?

The collection window runs from Day 1 to Day 7 after each dose. A reaction outside that interval would not be part of the specified endpoint. Redness and swelling were counted only when the reported size was at least 2.5 cm; smaller sizes were excluded. Such operational rules make endpoint classification more reproducible, but they also define what the endpoint can capture. The endpoint therefore concerns reactions meeting the registry’s stated criteria, rather than every possible sensation or finding after injection.

Reaction grading used criteria based on US FDA guidance, with grades from mild through potentially life-threatening. The registry describes this grading framework but the primary outcome is the percentage reporting local reactions of any grade. No grade-specific results are reported in the information available for this analysis. The record also describes participant assessments collected in an electronic diary or mapped from an adverse event case report form. The detailed rules for reconciling those sources are not reported in the registry record.

How should cohort comparisons be understood?

As a general statistical concept, random allocation helps balance both measured and unmeasured participant characteristics between assigned groups, on average. This trial is recorded as non-randomized, so cohort membership may be associated with characteristics that also relate to outcomes. The listed cohorts differ in formulations, number of doses, age ranges, and vaccination or infection history. A difference in outcome between cohorts could therefore have multiple explanations. The unmasked design also means participants and study staff were not masked, as indicated by the registry record. The record does not report a statistical adjustment strategy for these potential differences.

As a general statistical concept, a confidence interval describes the statistical uncertainty around an estimate under the assumptions of the analysis. It is not a guarantee about the value in an individual participant or a measure of clinical importance. No primary-endpoint confidence interval is reported in the registry information. With no estimate or interval, neither the size nor the precision of the primary endpoint can be evaluated from these details.

What does the planning illustration say?

The registry’s sample-size assumptions describe a planning illustration using a two-sided significance level of 0.05, equal allocation between two arms, and a normal approximation. The effect to detect is described as a stated fraction of an observed effect. For non-time-to-event outcomes, the illustration is defined in participants per arm for 80% or 90% power; time-to-event illustrations are defined in total events using the Schoenfeld approach. These are general planning assumptions, not evidence that the study used those methods to set enrollment or that the trial achieved a particular power. The study-specific sample-size justification, observed effect used for the illustration, and resulting planning values are not reported in the registry record.

Power is a design calculation that depends on assumptions about the effect, variability, significance level, and analysis. As a general statistical point, a post hoc calculation based on an observed estimate is not observed power and does not replace an estimate with its uncertainty interval. The stated illustration should therefore be read as a planning aid, not as a summary of the evidence from participants in this trial.

Why might variability affect analysis?

General statistical explanation: when observations vary more than a model assumes, standard errors and uncertainty estimates can be too small if that extra variation is ignored. This issue is often called over-dispersion. The primary outcome is binary and reported at participant level, but the registry information does not state that over-dispersion was detected, nor does it report a model for this endpoint. It would be inappropriate to infer a particular model or variance correction.

The record lists 32 outcome measures and 3 analyses. A study with multiple outcomes or comparisons raises a general multiplicity concern: repeated testing can increase the chance of at least one apparently notable result when all null hypotheses are true. Whether that concern was addressed depends on prespecified hypotheses, testing order, adjustment procedures, and the role of each outcome. The multiplicity strategy and detailed analysis plan are not reported in the registry record. The listed number of outcomes and analyses alone does not establish how many formal statistical tests were performed or whether any adjustment was used.

10. Multiplicity, Missing Data and Analysis Populations

The registry lists 32 outcome measures and 3 analyses, but does not report a multiplicity strategy. The information does not specify which outcomes were treated as confirmatory, whether comparisons were prespecified in a hierarchy, or whether significance levels were adjusted across outcomes or cohorts. As a general statistical point, testing many hypotheses can increase the chance of a false-positive finding if results are interpreted without accounting for the testing structure. The number of listed outcomes does not by itself reveal how many formal tests were conducted.

The primary endpoint population is defined as participants in the Safety Set with any reactogenicity data reported after injection. Part C Cohort 9 is excluded because those participants did not receive an injection and had no local reactions collected. The registry record does not report the number included in the primary endpoint analysis, the amount of missing diary information, or the methods used to handle missing or incomplete assessments. It also does not specify whether participants with some post-dose data contributed to all relevant dose assessments.

Those details matter because missing observations can affect both the denominator and the interpretation of a participant-level percentage. General statistical explanation: an analysis restricted to observed assessments describes the participants with usable data, while assumptions about unobserved outcomes are needed to extend conclusions beyond that group. No missing-data assumptions or sensitivity analyses are reported in the registry record.

The record identifies safety and immunogenicity populations in broad terms, but a full accounting of analysis sets and deviations from the planned analyses is not reported. The protocol’s statistical analysis plan, interim-analysis procedures, and sample-size justification are also not reported in the registry record. Without those details, the prespecified status of individual analyses and their treatment of missing data or multiplicity cannot be determined.

11. 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 5% reporting threshold.

Most frequent serious adverse event terms

Most frequent non-serious adverse event terms

Serious events with the highest arm-level rate: obstructive pancreatitis (5.0%), hepatitis acute (5.0%), cholelithiasis (1.4%), fibula fracture (1.4%), lower limb fracture (1.4%).

The registry lists serious adverse events by cohort as counts of participants affected and participants at risk. For Part A Cohorts 1 through 6, the affected counts were 0 of 21, 1 of 20, 1 of 20, 0 of 20, 0 of 42, and 0 of 17, respectively. For Part B Cohorts 1, 4, and 6, the counts were 6 of 349, 9 of 352, and 13 of 361. For Part C Cohorts 7 and 8, the counts were 0 of 72 and 5 of 71. These are counts, not adjusted comparisons.

The record also lists deaths as 1 in Part B Cohort 1 and 2 in Part B Cohort 6; the other listed cohorts had 0 deaths. The listed serious adverse event terms include Obstructive pancreatitis at 5.0%, Hepatitis acute at 5.0%, Cholelithiasis at 1.4%, Fibula fracture at 1.4%, Lower limb fracture at 1.4%, and Tibia fracture at 1.4%. The record does not state in this information the denominator or additional event-level context for those term percentages.

As a general interpretation point, an adverse event occurring during a trial is not by itself evidence that the intervention caused it. These cohorts were non-randomized and differ in size and characteristics, so their counts should not be treated as a controlled comparison. Detailed event timing, relatedness assessments, and follow-up context are not reported in the registry record.

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

The registry describes a Phase II study designed to evaluate safety and immunogenicity across several vaccine formulations, dose schedules, and participant histories. It gives a clear primary endpoint definition for local reactions after injection and identifies the population intended for that endpoint. The registry information does not include the endpoint’s observed counts, percentages, or confidence intervals. Consequently, the magnitude and precision of local reaction results cannot be stated from this record description.

Even if cohort-level endpoint results are considered alongside the registry record, their interpretation would need to account for the non-randomized, unmasked design. Participants were grouped into cohorts with different formulations and schedules, and the summary identifies differences in prior vaccination and infection history. A numerical contrast between cohorts would not, by itself, establish that the vaccine formulation caused the difference. The record does not report an adjusted comparison that addresses these distinctions.

The endpoint measures short-term reporting of specified local reactions, with collection from Day 1 to Day 7 and a size criterion for redness and swelling. It is not an endpoint for disease prevention, nor does it directly measure immune response. The trial’s secondary outcomes include geometric mean titers, geometric mean fold rises, and percentages meeting a seroresponse definition, but the registry information does not provide their results.

The serious adverse event counts are useful as descriptive safety information, but they do not establish causation. Counts must be considered alongside the participants at risk, event context, and follow-up. The listed deaths and serious adverse event terms should not be interpreted as proof of a relationship to vaccination; causal attribution is not reported in the available information.

As a general statistical principle, an estimate is more informative when accompanied by an uncertainty interval and a transparent analysis population. Neither the primary endpoint estimate nor its uncertainty is available in the registry information. The trial description therefore supports understanding what was studied and how the primary outcome was defined, but not a conclusion about the size of that outcome or the comparative effects of the different regimens.

13. Why This Trial Matters Statistically

This trial is statistically informative as an example of a complex, multi-cohort design addressing several vaccine formulations and participant histories within one registered study. The three-part structure encompasses booster doses, different variant-targeted formulations, a three-dose regimen, and an observational cohort without vaccination within 3 months after Visit 1. Such breadth can address distinct descriptive questions, but it also means that the cohorts do not represent a single uniform comparison.

The non-randomized allocation is central to interpretation. When cohorts differ in prior vaccine exposure, infection history, age range, or dosing schedule, those factors can affect the outcomes being measured. A comparison of cohort outcomes therefore cannot isolate the contribution of one formulation unless the analysis and design support that interpretation. The registry record does not report a detailed adjustment strategy.

The primary endpoint also illustrates why operational definitions matter. The post-dose assessment window, reaction grading framework, and minimum size for redness or swelling determine what is counted. The endpoint’s reactogenicity population excludes the cohort without vaccination, making the analysis population distinct from total enrollment. Clear definitions help readers understand what an eventual estimate would represent, while missing endpoint results prevent assessment of its actual frequency and precision from this record description.

Finally, the record contains multiple outcome measures and analyses, yet does not describe the multiplicity plan, missing-data methods, or study-specific statistical analysis plan. These omissions limit assessment of how the analyses were selected and how uncertainty was handled. The trial is therefore a useful case for separating a study’s broad aims and endpoint definitions from the evidence needed to interpret specific estimates.

14. 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 design and reporting details impose important limits on interpretation. Allocation was non-randomized and masking was absent. Cohorts received different formulations and schedules, and the trial summary describes differing vaccination and infection histories. Those characteristics mean that cohort contrasts may reflect differences in participant history or other factors, as well as differences in the assigned vaccine regimen. A like-for-like randomized control comparison is not described.

The primary endpoint is defined in detail, but its results are not reported in the registry information. The number of participants contributing to the endpoint, cohort-specific estimates, and uncertainty intervals are not stated. The registry defines a reactogenicity set based on participants with data after injection, but the amount and pattern of missing data are not reported.

The record lists many outcome measures and analyses but does not report the multiplicity approach, missing-data assumptions, interim analyses, or a detailed statistical analysis plan. The trial’s sample-size justification is not reported. The separate planning illustration is described through assumptions, but its observed effect and calculated planning values are not stated in this information.

The registry summary spans Parts A, B, and C and describes different schedules, ages, and prior histories. These features make it difficult to reduce the study to one treatment-versus-control question. Part C’s unvaccinated observational cohort was not included in the local-reaction analysis, since reactions were not collected for participants without an injection. Finally, the adverse event counts are descriptive; detailed timing, relatedness, and follow-up information are not reported in the registry record. These limitations preclude causal interpretation of the listed safety counts.

15. Related Tutorials

16. Related Statistical Calculators

17. Sources

• ClinicalTrials.gov: Safety and Immunogenicity of RNA-based Vaccines Against SARS-CoV-2 Variants in Healthy Participants — NCT05004181.
• Registry record last updated 2024-11-22; status last verified 2024-11.
• Results first posted 2024-11-22.
• 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.

18. Record Summary

NCT05004181 is a completed, non-randomized, unmasked Phase II parallel trial sponsored by BioNTech SE, with listed enrollment of 1380. It evaluated monovalent and multivalent RNA-based SARS-CoV-2 vaccines in three parts, with cohorts receiving different formulations and dose schedules. The registry lists the study start as 2021-08-25 and primary completion as 2023-08-16.

The primary endpoint is the percentage of participants reporting qualifying local reactions after injection, assessed from Day 1 to Day 7 after each dose. Redness and swelling had to measure at least 2.5 cm to count. The endpoint results are not reported in the registry information. Serious adverse event counts and deaths are listed by cohort, but those descriptive counts do not establish causation. The detailed statistical plan, multiplicity approach, missing-data methods, and sample-size justification are not reported in the registry record.