ASCERTAIN: Assessment of Everolimus in Addition to Calcineurin Inhibitor Reduction in the Maintenance of Renal Transplant Recipients: Complete Statistical Analysis

1. Trial at a Glance

This page covers the registry results for ASCERTAIN: Assessment of Everolimus in Addition to Calcineurin Inhibitor Reduction in the Maintenance of Renal Transplant Recipients (NCT00170846), a randomized phase 4 study. The primary outcome is Renal Function Assessed by Measured GFR (mGFR).

2. Clinical Question

3. Trial Design

ASCERTAIN was a randomized, parallel-group, Phase 4 trial sponsored by Novartis Pharmaceuticals. The registry lists the study as completed, with enrollment of 394. Masking is recorded as none, so the trial was unmasked. The registry record does not report the randomization ratio or the original sample-size justification.

The three regimens differed in whether everolimus was used and how CNIs were managed. Group A continued CNI treatment, potentially alongside mycophenolate acid or azathioprine and steroids. Group B initiated everolimus and discontinued CNI treatment. Group C initiated everolimus and reduced CNI blood levels by 70-90%. The intervention descriptions give different initial everolimus doses and target concentration ranges for the two everolimus regimens.

The primary endpoint was mean measured GFR at 24 months. The acceptable measurement methods included Cr-EDTA, Tc-DTPA, iohexol clearance, inuline clearance, and iothalamate clearance; the method was to remain consistent for an individual patient across time points. The primary analysis population was modified ITT, defined in the registry as ITT patients with mGFR or calculated GFR at Month 24, including measurements taken after discontinuation of study medication.

The registry lists one secondary outcome, participants with safety parameters. It also records three outcome measures and two analyses. The study’s brief description says development of atherosclerosis in native arteries would also be explored, but the registry record does not give results for that exploration in the posted outcome details.

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

Reasons for not completing this period:

Analysis population for the primary outcome, as defined in the registry: The modified ITT population included all ITT patients who had mGFR or calculated GFR (cGFR) at Month 24 based on all values including those collected after discontinuation of study medication.

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.

6. Primary Endpoint

The registry lists the primary outcome as Renal Function Assessed by Measured GFR (mGFR), measured over 24 months.

The primary endpoint was renal function assessed by measured glomerular filtration rate (mGFR), expressed in mL/min/1.73m^2, at 24 months. GFR is used as an indicator of how well the kidneys filter blood. In this trial, the target organ was a transplanted kidney, so a measurement of renal function was relevant to the study’s stated interest in graft function and chronic allograft nephropathy.

The registry accepted several methods for measuring GFR: Cr-EDTA, Tc-DTPA, iohexol clearance, inuline clearance, and iothalamate clearance. It specified that a given patient should be measured using a consistent method at every time point. Consistency is important because different measurement procedures can yield results with different measurement properties; keeping the method consistent helps make within-patient changes easier to interpret. The registry also says the primary analysis included either mGFR or calculated GFR at Month 24.

The posted arm results are means with standard deviations. A mean describes the group’s average measurement, while a standard deviation describes variation among measurements within a group. Neither statistic alone describes every participant’s result. The endpoint is a measurement at the stated time point, not a direct count of graft failures or a measure of the trial’s broader atherosclerosis exploration.

7. Results Posted in the Registry

• Group B : CNI Withdrawal: mean 48.00 versus 46.02 on control; difference 1.98 (95% CI -3.96 to 7.92), standardized effect d = 0.09; z-test p = 0.514.
• Group C: CNI Reduction: mean 46.60 versus 46.02 on control; difference 0.58 (95% CI -5.00 to 6.16), standardized effect d = 0.03; z-test p = 0.839.

8. Statistical Interpretation

9. Statistical Methods Explained

General: A randomized parallel-group trial assigns participants to separate treatment regimens and follows the groups for comparison. Randomization is intended to make the groups comparable on average at the time of assignment, although it does not guarantee that every measured or unmeasured characteristic will be balanced. The registry records this study as randomized and parallel, but does not report the allocation ratio or further details of the randomization procedure. The study was unmasked.

What does a mean difference describe?

General: A mean difference is the average measurement in one group minus the average measurement in a comparator group, following the stated comparison direction. A positive estimate means the first named group’s average is higher; a negative estimate means it is lower. The recalculated comparisons use the posted group means: Group B’s mean was 48.0 versus 46.02 in Group A, and Group C’s was 46.6 versus 46.02. The resulting unadjusted differences are 1.98 and 0.58, respectively.

The recalculated intervals and p-values are based on unadjusted comparisons of the posted summary statistics. They do not account for any covariates. The recalculation uses the reported group sizes and standard deviations; the registry’s sponsor analyses instead use ANCOVA. The record does not indicate that the recalculated results replace the sponsor’s analysis.

What does ANCOVA contribute?

General: Analysis of covariance (ANCOVA) estimates a group contrast while adjusting for selected covariates. Adjustment can account for differences in measured baseline or other covariates included in the model, and the resulting least-squares means are model-based rather than simple raw group averages. The registry reports ANCOVA, differences in least-squares means, confidence intervals, and p-values for both comparisons with Group A. It does not identify the covariates, model specification, or any diagnostic checks, so the adjustment cannot be assessed in detail from the registry record.

The sponsor estimates and the recalculated estimates need not match exactly because they come from different methods and may target different adjusted quantities. Their directions are positive for both everolimus comparisons, and both sets of intervals include zero. That broad agreement does not make the methods interchangeable; the sponsor’s ANCOVA results remain the trial’s reported analyses, while the simple comparisons are unadjusted recalculations.

Why are standard deviations relevant?

General: A standard deviation describes how dispersed observations are around their group mean. It differs from a standard error, which describes uncertainty in an estimated mean. Greater variability generally makes a group mean less precise, especially when the number of observations is limited. The posted outcome summaries give standard deviations for each arm, and the recalculations use them to estimate uncertainty around the differences. The registry record does not provide individual participant data, so the summaries cannot show the shape of each group’s outcome distribution or identify unusual observations.

How should the analysis population be understood?

The registry defines the modified ITT population as ITT patients who had mGFR or calculated GFR at Month 24. It explicitly includes values obtained after study medication discontinuation. This definition preserves information from participants who stopped study medication, but it also means the endpoint analysis requires a recorded GFR measurement at the specified time point. The registry record does not describe imputation, other missing-data methods, or how many eligible participants lacked a Month 24 measure.

General: An analysis population definition tells readers which participants’ data contribute to a particular estimate. Modified ITT populations can differ from the full randomized population when eligibility for the analysis depends on availability of an outcome measure or another post-randomization condition. The posted description allows the reader to identify the key outcome requirement, but does not provide enough detail to evaluate the effects of missing measurements.

What does a p-value say?

General: A p-value describes how compatible data are with a specified null model, under the assumptions of the analysis. It is not the probability that the null hypothesis is true, and it does not measure the size or clinical importance of an effect. The unadjusted p-values are 0.514 and 0.839. The sponsor’s ANCOVA p-values are not reproduced because their reported precision exceeds the permitted display precision; the registry’s intervals and conclusions can still be described qualitatively.

Multiplicity arises when several outcomes or comparisons are examined. General: Testing multiple contrasts can increase the chance of at least one apparently unusual result if each test is considered in isolation. The registry reports two comparisons against Group A and one secondary outcome, but does not report whether multiplicity was addressed. It also does not report interim analyses or a complete statistical analysis plan. Those details are not reported in the registry record.

10. Confidence Intervals and Precision

The recalculated 95% confidence interval for Group B versus Group A extends from -3.961 to 7.921 mL/min/1.73m^2 around a mean difference of 1.98. For Group C versus Group A, it extends from -4.998 to 6.158 around a mean difference of 0.58. Both intervals include zero, so the posted recalculations do not rule out no average difference. They also include values in both directions, indicating uncertainty about the direction and size of the contrast.

General: A confidence interval gives a range of estimates compatible with the data under the method’s assumptions. It is not a guarantee that the true effect lies in the interval, nor does its width alone determine clinical importance. A narrower interval indicates greater statistical precision for the estimated contrast; a wider interval indicates more uncertainty. Interpreting the range also requires attention to the outcome’s units and to what differences would matter clinically, which the registry record does not specify.

The sponsor’s ANCOVA intervals also include zero for both comparisons. The registry reports adjusted estimates and intervals, but does not report the covariates or model details needed to assess how adjustment affected precision. The unadjusted and adjusted intervals should therefore be understood as results from different analyses rather than as competing versions of the same calculation.

The posted group standard deviations, 20.36 for Group A, 22.03 for Group B, and 21.08 for Group C, indicate substantial spread in the outcome within each group. The group means alone cannot capture this variation. The recalculated intervals communicate uncertainty in the average contrasts, not the range of individual participant outcomes.

11. Planning a Similar Trial: Sample Size

This is a planning illustration, not a re-evaluation of the original trial. It asks: if a new study wanted to detect a difference like the one observed, or a smaller one, how large would it need to be? It is deliberately not “observed power” of the finished trial, which is not a meaningful quantity.

The planning figures are participants per arm, not total event counts. They are illustrations calculated from the observed effect, rather than the trial’s original design assumptions. The registry record does not report the original sample-size justification. The listed calculation assumptions are a two-sided significance level of 0.05, equal allocation between the two arms, and a normal approximation; each scenario targets a stated fraction of the observed effect.

The 80% power column gives the number of participants per arm estimated to be needed for an 80% chance of detecting the specified effect under those assumptions, if the assumed effect is real and the model is appropriate. The 90% power column gives the corresponding planning number for a 90% chance. These are planning targets, not observed characteristics of the trial. This is not observed power.

For each comparison, the rows consider planning for 100%, 75%, and 50% of the observed effect. A smaller target effect requires more participants per arm to achieve the same planned power, because a smaller difference is harder to distinguish from ordinary sampling variation. The figures are especially large for the Group C comparison because its observed mean difference is small. This should not be read as proof that a future trial needs those exact numbers: the calculations rely on the stated planning assumptions and on the observed effect, which itself is uncertain.

The planning illustrations apply to two-arm comparisons. They do not incorporate information about the trial’s unreported original design rationale, any adjustment strategy, or multiplicity procedures. The sample-size figures are therefore useful for understanding the relationship between effect size and planning requirements, rather than as a retrospective judgment of whether the completed trial was adequately designed.

12. How the Sample Size Is Derived

Power is the probability that a study of a given size rejects the null hypothesis when a specific true effect exists. It depends on four things: the effect to detect, the variability of the outcome, the significance level (here a two-sided α of 0.05, so z_1−α/2 = 1.960) and the desired power (z_1−β = 0.842 for 80% and 1.282 for 90%).

Formula

n is the number of participants in each arm, Δ is the difference in means to detect and σ is the common standard deviation.

Worked calculation for this trial

1. Observed difference Δ = 1.980; pooled standard deviation σ = 21.173.
2. n = 2 × (1.96 + 0.842)² × 21.173² / 1.980² = 1,795.1, rounded up to 1,796 participants per arm for 80% power.

The smaller-effect rows in the table above repeat the same calculation with the effect multiplied by 0.75 and 0.5 (for ratios, applied to the logarithm of the ratio).

R code

alpha <- 0.05
z_a   <- qnorm(1 - alpha / 2)          # two-sided
delta <- 1.98                      # observed difference in means
sigma <- 21.1733807245724                     # pooled standard deviation
n_per_arm <- function(power, frac = 1) {
  ceiling(round(2 * (z_a + qnorm(power))^2 * sigma^2 / (delta * frac)^2, 6))
}
for (f in c(1, 0.75, 0.5))
  cat(sprintf("%3.0f%% of observed effect: n per arm = %s (80%% power), %s (90%% power)\n", 100 * f,
              format(n_per_arm(0.80, f), big.mark = ","), format(n_per_arm(0.90, f), big.mark = ",")))

# Check against R's built-in function (t-based, so very slightly larger)
cat("power.t.test:", round(power.t.test(delta = delta, sd = sigma, power = 0.80)$n, 1), "\n")

# Simulation: exact two-sample t-test using simulated means and variances
set.seed(170846)
n    <- n_per_arm(0.80)
reps <- 20000
m_diff <- rnorm(reps, delta, sigma * sqrt(2 / n))             # difference in sample means
s2     <- sigma^2 * rchisq(reps, 2 * (n - 1)) / (2 * (n - 1))   # pooled sample variance
tstat  <- m_diff / sqrt(s2 * 2 / n)
cat(sprintf("Simulated power at n = %s per arm: %.3f (%d replicates)\n", format(n, big.mark = ","), mean(abs(tstat) > qt(0.975, 2 * (n - 1))), reps))

Output from running the code

100% of observed effect: n per arm = 1,796 (80% power), 2,404 (90% power)
 75% of observed effect: n per arm = 3,192 (80% power), 4,273 (90% power)
 50% of observed effect: n per arm = 7,181 (80% power), 9,613 (90% power)
power.t.test: 1796.1 
Simulated power at n = 1,796 per arm: 0.802 (20000 replicates)

The simulation draws many sets of group means and variances at the planned sample size and counts how often the two-sample t-test rejects the null. It should land close to the target power; at very small required sample sizes it can fall below it, because the normal approximation is rough there.

13. Analyses Reported by the Sponsor

The registry’s reported analyses compare each everolimus group with Group A using ANCOVA, with differences in least-squares means as the measure of effect. The reported confidence intervals for both comparisons include zero, and the sponsor p-values do not indicate a statistically significant difference. The registry record does not report the covariates or other model details used for these adjusted estimates.

The unadjusted recalculations use posted arm means and standard deviations. They produce positive mean differences for both comparisons, and their confidence intervals also include zero. The recalculated estimates are not identical to the sponsor’s least-squares mean estimates because the analyses differ: one is an unadjusted comparison of group summaries, while the other is an ANCOVA-based adjusted comparison.

The registry record does not flag a discrepancy between the posted analyses and the recalculations. The sponsor’s analysis is the trial’s reported analysis; the recalculations are a complementary, simple comparison of the posted summary statistics. Neither analysis establishes a treatment effect merely from the direction of its estimate, and their intervals show uncertainty around the contrasts.

14. Multiplicity, Missing Data and Analysis Populations

The registry reports two primary-outcome comparisons against Group A: one for CNI withdrawal and one for CNI reduction. It also lists one secondary outcome, participants with safety parameters. The registry record does not report a multiplicity adjustment or explain how the two primary comparisons were handled as a family of tests. Without that information, the p-values should be interpreted as results for their stated analyses rather than as evidence that a particular family-wise error strategy was applied.

The registry reports ANCOVA for the sponsor comparisons, but does not provide the covariates, model specification, analysis diagnostics, or a complete statistical analysis plan. It also does not report whether interim analyses were planned or performed. These items are not reported in the registry record and should not be inferred from the posted results.

The modified ITT population included ITT patients with mGFR or calculated GFR at Month 24, including measurements collected after study medication discontinuation. This description clarifies that stopping medication did not itself exclude a participant if an eligible Month 24 measurement was available. However, the record does not report how many participants lacked that measurement, how missing measurements were handled, or whether any imputation or sensitivity analyses were used. It therefore does not permit a full assessment of missing-data effects.

General: Missing outcome data can affect an estimate when the availability of measurements is related to participant characteristics or outcomes. A population definition describes who was analyzed, but does not by itself show whether missingness could have biased the result. Likewise, reporting multiple outcomes or comparisons does not establish whether statistical multiplicity was addressed. The registry record leaves these aspects unresolved.

15. 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 5% reporting threshold.

Most frequent serious adverse event terms

Most frequent non-serious adverse event terms

Serious events with the highest arm-level rate: diarrhoea (8.3%), pyrexia (7.6%), urinary tract infection (6.9%), renal graft loss (6.2%), blood creatinine increased (5.7%).

The safety summary lists serious adverse events for each arm as participants affected among those at risk. Group A reports 52 affected among 123 at risk; Group B reports 72 among 127 at risk; Group C reports 78 among 144 at risk. These are descriptive counts, not adjusted comparisons. The registry record does not give a statistical comparison of these figures or identify whether the events were considered related to study treatment.

The most frequently listed serious adverse event terms are diarrhoea (8.3%), pyrexia (7.6%), urinary tract infection (6.9%), renal graft loss (6.2%), blood creatinine increased (5.7%), and gastroenteritis (4.2%). The denominator for these percentages is not reported in the registry record. The terms should be read as listed event categories, not as a complete account of all safety findings.

Deaths are not reported for the arms in the registry entries. The registry also lists participants affected by other adverse events, but the safety summary does not give the event definitions or a fuller account of severity and attribution. These figures alone do not establish comparative safety or explain why events occurred.

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

The posted results describe mean renal function at 24 months in three randomized groups. The unadjusted recalculations show small positive differences in mean mGFR for both everolimus regimens relative to the no-everolimus group. Their confidence intervals include zero and extend to values on either side of zero. The sponsor’s adjusted analyses likewise report intervals that include zero. Together, these results do not provide a precise estimate that isolates a difference between either experimental regimen and the comparator.

This interpretation is limited to the measured endpoint, analysis population, time point, and comparisons recorded in the registry. It does not establish whether the regimens improve or worsen health, and it does not provide a complete account of graft outcomes, other clinical endpoints, or the planned exploration of atherosclerosis in native arteries. The registry record does not report those results in the posted outcome details.

A non-significant result is not proof that groups are identical. General: Failure to distinguish a difference statistically may reflect a small effect, substantial variability, limited information, or a combination of factors. Conversely, a positive point estimate is not sufficient evidence of a meaningful effect. The confidence intervals are important because they show a range of contrasts compatible with the data under each analysis.

The registry’s modified ITT definition includes Month 24 measurements collected after study medication discontinuation, which is relevant to how the endpoint was assembled. But details about missing measurements, imputation, and sensitivity analyses are not reported in the registry record. The available summaries therefore cannot resolve how missing data may have influenced the findings.

The results should not be generalized beyond the studied maintenance renal transplant population without additional evidence. They also do not establish that any regimen is preferable for an individual patient. They are statistical summaries of the registry’s reported endpoint and comparisons, with uncertainty that should remain part of their interpretation.

17. Why This Trial Matters Statistically

This trial offers a useful statistical comparison of three strategies for managing calcineurin inhibitor use in maintenance renal transplant recipients: continuing CNI treatment without everolimus, discontinuing CNI treatment while initiating everolimus, and reducing CNI levels while initiating everolimus. The question is not captured by a single experimental-versus-control contrast because the two everolimus regimens alter CNI exposure in different ways.

The primary outcome is a continuous measure of renal function, and the registry reports group means and standard deviations alongside sponsor-adjusted comparisons. This makes it possible to contrast raw mean differences with ANCOVA estimates and to see why the choice of analysis matters. The adjusted and unadjusted results are not interchangeable, but both sets of intervals include zero, underscoring the uncertainty around the average contrasts.

The study also illustrates why estimates should be read together with their precision. The observed differences are small relative to the variation in mGFR, and the planning illustrations show that smaller target effects require larger sample sizes under the stated assumptions. Those calculations are not a verdict on the original design, whose sample-size justification is not reported; they demonstrate the general relationship between effect size, variability, and planning requirements.

Finally, the registry’s analysis-population definition, missing-data details, and multiplicity reporting affect how confidently the results can be evaluated. The record specifies which outcome measurements qualified for the modified ITT analysis, but does not provide a full account of missing outcomes or the statistical plan. Distinguishing what is reported from what remains unknown is essential when interpreting a completed trial.

18. 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 posted primary outcome summaries are aggregate statistics. They do not allow assessment of individual participant trajectories, the distribution or shape of mGFR values, or whether unusual observations influenced the means. The registry accepts several GFR measurement methods but does not report the method used for each participant or the distribution of methods across groups. It states that each participant’s method should be consistent across time points, but the extent of adherence is not reported.

The sponsor’s ANCOVA results are reported as adjusted estimates, yet the covariates and model specification are not reported in the registry record. That makes it difficult to evaluate the adjustment or compare the adjusted results with a fully specified model. The original sample-size justification, randomization ratio, and detailed statistical analysis plan are also not reported.

The modified ITT population required mGFR or calculated GFR at Month 24, while allowing values collected after study medication discontinuation. The registry record does not state how many participants lacked a qualifying value, how missing outcomes were handled, or whether sensitivity analyses assessed the effect of missingness. Multiplicity procedures and interim analyses are also not reported.

The safety summary gives counts of participants affected and a list of serious adverse event terms with percentages, but does not report the denominator for those percentages in the registry record. It does not provide a treatment-attribution analysis or a complete narrative of safety results. Death counts are not reported. These limitations constrain interpretation of the posted outcome and safety summaries and should not be filled in by assumptions.

19. Related Tutorials

20. Related Statistical Calculators

21. Sources

• ClinicalTrials.gov: ASCERTAIN: Assessment of Everolimus in Addition to Calcineurin Inhibitor Reduction in the Maintenance of Renal Transplant Recipients — NCT00170846.
• Registry record last updated 2015-01-27; status last verified 2011-03.
• Results first posted 2011-05-02.
• 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.

22. Record Summary

ASCERTAIN (NCT00170846) was a completed, randomized, unmasked, parallel-group Phase 4 study sponsored by Novartis Pharmaceuticals. It enrolled 394 participants with renal transplantation. The study compared a no-everolimus CNI regimen with everolimus initiated alongside either CNI withdrawal or CNI reduction.

The primary endpoint was mean measured GFR at 24 months in a modified ITT population with a qualifying Month 24 mGFR or calculated GFR value, including measurements after study medication discontinuation. The sponsor reported ANCOVA comparisons against the control group. Unadjusted recalculations produced positive mean differences for both experimental groups, but their confidence intervals included zero; the sponsor intervals also included zero.

The registry lists one secondary safety outcome. Details on missing-data methods, multiplicity, interim analyses, and the original sample-size justification are not reported in the registry record.