Study Populations and Associated Information
Data included in the analyses are de-identified ECG waveforms for moxifloxacin and placebo arms, as well as demographic information from 351 healthy individuals enrolled in seven TQT studies with crossover design. The studies were selected from the FDA warehouse based on willingness from pharmaceutical companies to share their data for research purpose. The pharmaceutical companies agreed on sharing their data with the Agency but requested to remain anonymous. Then, the data shared by the FDA enabled us to evaluate the placebo-controlled moxifloxacin-induced QT/QTc interval prolongation at expected peak concentration of the drug (2 hours post dosing) in six studies, and the time profile of the QTc interval within one study, the so-called study 7.
HL7 XML ECG files from the FDA ECG warehouse
The ECGs from the baseline and the placebo arms were recorded at the same time of the day than in the moxifloxacin arm in order to control for circadian effects. Intake time of the placebo was synchronized with intake time of moxifloxacin. The digital ECG files and the respective information (age, gender, and study treatment) were provided by the Center for Drug Evaluation and Research of the FDA to our organization. The HL7 XML ECGs files were sent to the University of Rochester by Mortara Instrument (Milwaukee, MN) following FDA request and were extracted from the ECG warehouse after FDA ensured that pharmaceutical companies agreed on having their ECG used for research purpose. Any information that could help identifying the source of the data or reveal the name of organizations involved in the study (pharmaceutical companies, ECG core laboratory, etc.) was removed from files send to University of Rochester. Information about the methods adopted by ECG core laboratory to measure the QT intervals were not available. However, the FDA (CG, ML) provided a code to identify the five different ECG core laboratories (ECL) involved in the QT measurements of these seven studies (ECL A to E).
RR and QT Interval Measurements
The descriptions of the methods used by ECL to measure QT interval were not available, consequently our HA-QT measurement method was compared to “current ECL methods” rather than a specific method. We expect that the ECLs involved in these seven TQT studies used semi-automatic QT method i.e. a computer algorithm identifies the end of the T-wave and a cardiologist or ECG technician visually assesses the end of T-wave and provides adjudication if needed. Most likely for each tracing, the QT measurements were based on three beats with adjudication based on on-screen tools such as the tangent method or on method using a global lead superimposition.
The HA-QT and RR intervals measurements were based on the technology developed at the University of Rochester Medical Center, NY. The COMPAS software provided the identification of the end of the T-wave based on a technique identifying the crossing-point between the baseline and the descending slope of the T-wave (least-squares technique).27
QT intervals were determined to be unreadable, preventing the measurements from being done, when the T-wave amplitude was inferior to 50 μV (flat T-wave). The QT interval measurements were computed in all available sinus beats. The median QT value from all measured cardiac beats was reported after applying rate correction formula described below.
The QT algorithm included a set of pre-processing steps such as: baseline estimation and removal based on spline interpolation, a low-pass filter with finite-impulse response of 20th order (FC=25 Hz) was designed. Finally, we developed a least-square method based on a moving-window seeking for the maximum fitting slope within the terminal part of the T-wave (independently from the identification of the end of the T-wave).
The QT measurements from University of Rochester were sent to the FDA where they were matched with the QT measurements submitted by pharmaceutical companies. If a missing QT measurement was found in one of the dataset (UR vs. FDA) then the corresponding data point was removed from the other dataset.
Quality Assessment and Adjudication of QT Measurements
A quality assessment process involving a two-level filtering process was designed to identify QT and RR interval measurements outside of their “expected” range. The identification of these outlying values was based on the following two criteria:
- Absolute thresholds: These first thresholds are used to identify failures of the computer methods leading to extreme values for QT and RR intervals. Based on clinical investigations related to short and long QT syndrome, we ensured that the QT criterion encompasses the normal QTc values: 350<QTc<440 msec (based on Fridericia correction formula)28. We broadened this range in order to focus on unlikely values in the type of study population of TQT studies (healthy individuals):
- Relative Threshold: We identified the ECG recordings associated with suspicious RR and QT interval values using a statistical tool (excluding ECGs meeting the above criteria). The QT interval values falling outside of the inter-quartile range computed from the overall set of available ECGs were labeled.
All ECGs associated with measurements fitting at least one of these two criteria were flagged and visually reviewed and then accepted, corrected or rejected in all studies but study 7.
Outlying measurements from the study 7 were identified using the criteria describe above (similar to other studies). However, we did not include manual adjudication of QT measurements rather we rejected the outlying measurements from the analysis when the end of the T-wave was flawed. We had to change this review process because of the limited resources we had to analyze this second set of 8,911 ECGs initially not included in our research plan. Data from the study 7 were sent by the FDA to the University of Rochester at later time in order to investigate the moxifloxacin-induced QT time profile.
Comparing QT Measurements
In summary, the QT measurements compared in this work are called the HA-QT, and the FDA-submitted QT. The HA-QT measurements are automatic QT measurement (based on COMPAS method) on which limited manual review based on quality metrics (described above) were done. All available beats in each ECGs are measured and median value from all beats is reported for each ECG tracing. The FDA-submitted QT measurements are expected to be based on automatic QT measurements with 100% manual review of the measurements. Most likely, different methods were used to measure automatically QT interval before manual review but these methods were not documented. The number of measurement manually adjudicated was not available either. Most likely, the QT measurements were based on three selected beats in each ECG tracing. Eventually, median value from the medians QT from the triplicate ECGs (when available) was computed.
Heart Rate Correction
Heart rate correction of the QT interval duration remains a challenging aspect of the assessment of QT-prolonging effect of drugs. First, it is known that QT/RR profiles are subject dependent and can be modified by drugs. Moxifloxacin does not significantly affect the heart rate in human; therefore, the issue of bias due to QT dependency to heart rate is not of strong concern in this analysis. Nevertheless, the Bazett’s formula is known to generate large biased and reduced QTc measurement precision so we have excluded this correction method from our analysis.29, 30
We opted for comparing QTc using Fridericia’s formula 31
QTc Prolongation, Peak Concentration, and Time Profile
The assay sensitivity for a QT measurement technique in thorough QT studies is demonstrated when two criteria are met: 1) the lower two-sided 90% interval of the baseline adjusted and placebo controlled population-based mean of QTc changes induced by moxifloxacin is above a 5 msec threshold (at the time of expected peak concentration) and 2) the QTc profile across time follows the expected plasma concentration of the drug (moxifloxacin).
In our study, the QT interval was measured in all available beats and the median value was computed to yield one value for each ECG. Since most individuals had triplicate ECGs, the median of the available replicates was calculated for each time point. Single time-matched delta QTc durations were computed as follow:
To account for the placebo effect, ΔQTcplacebo
was subtracted from ΔQTcmoxi
for each individual. The mean Δ ΔQTc was calculated for each time point. Importantly, the QTc at baseline was the time-match value when assessing the QTc prolongation at expected peak concentration whereas the QTc at baseline was measured from the pre-dose ECG when evaluating the QTc profile in study 7. These methods follow the recommended analytic strategies adopted by the FDA.32
The population-based mean Δ ΔQTc prolongations between methods at expected maximum concentration of moxifloxacin and their two-sided 90% confidence interval are reported. T-test were used to compare the population-based average of QT interval between genders, a level of statistical significance (p-value) was set to 0.05.
When comparing the curves describing the QT time profile between the QT methods (for study 7 exclusively), the primary analysis was based on a mixed-effects linear model for repeated measures of the differences from baseline in ΔQTc between the two measurement techniques. The model had no intercept and used an unstructured covariance structure as this model showed the best goodness of fit.
The QT measurements realized at University of Rochester were sent to the FDA (to ML and CG) to implement the comparative analysis between HA-QT and the measurements received by the Agency. Thus, most of the statistical analysis was implemented by ML and GC independently from the University of Rochester group.