ICH E14/S7B Implementation Working Group
Clinical and Nonclinical Evaluation of QT/QTc Interval Prolongation and Proarrhythmic
Potential
Questions and Answers

E14/S7B Q&As
Adopted on 21 February 2022

International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use
ICH, Route de Pré-Bois 20, P.O. Box 1894, 1215 Geneva, Switzerland
Telephone: +41 (22) 710 7480 - admin@ich.org, http://www.ich.org

E14/S7B Q&As

In order to facilitate the implementation of the E14 and S7B Guidelines,
the ICH E14/S7B Experts have developed a series of Q&As:

E14/S7B Q&As
Document History
Code
E14 Q&As
E14
Q&As (R1)E14
Q&As (R2)E14
Q&As (R3)
E14/S7B
Q&As

History
Approval by the ICH Steering Committee under Step 4
Approval by the ICH Steering Committee under Step 4 offour newly added Questions on: Sex Differences;
Incorporating New Technologies; Late Stage Monitoring;and Heart Rate Correction.
Approval by the ICH Steering Committee under Step 4 offour newly added Questions on: Concentration-Response
Relationships; Combination Products; Large Targeted
Proteins and Monoclonal Antibodies; and Special Cases.
Approval by the ICH Assembly under Step 4 of therevision of the Question #5.1 on: Use of Concentration
Response Modeling of QTc Data
Approval by the ICH Assembly under Step 4

Date
4 June 2008
5 April 2012

21 March2014

10 December
2015
21 February
2022

References
ICH E14

The Clinical Evaluation of QT/QTc Interval Prolongation and Proarrhythmic Potential for NonAntiarrhythmic Drugs

May 2005

ICH E14

Questions & Answers (R3)

Dec 2015

ICH S7B Nonclinical Evaluation of the Potential for Delayed Ventricular Repolarization (QT Interval Prolongation)
by Human Pharmaceuticals

May 2005

2

E14/S7B Q&As
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3E14/S7B Q&As

Table of Content
E14 Q&As .................................................................................................................................................................................................. 5
1. Electrocardiograms Methodology ...................................................................................................................................................... 5
2. Gender .............................................................................................................................................................................................. 11
3. Positive Control ................................................................................................................................................................................ 12
4. Study Design .................................................................................................................................................................................... 14
5. Use of Concentration Response Modeling of QTc Data .................................................................................................................. 15
6. Special Cases .................................................................................................................................................................................... 19
7. Electrocardiograms Monitoring in Late Stage Clinical Trials ......................................................................................................... 23
S7B Q&As ............................................................................................................................................................................................... 27
1. Integrated Risk Assessment ............................................................................................................................................................. 27
2. Best Practice Considerations for In vitro Studies............................................................................................................................. 31
3. Best Practice Considerations for the In vivo QT Studies ................................................................................................................. 39
4. Principles for Proarrhythmia Models ............................................................................................................................................... 434

E14/S7B Q&As

E14 Q&As
1. Electrocardiograms Methodology#
1.1

Date of
Questions
Approval
June
2008

Please discuss who should readelectrocardiograms(ECGs),
including the number and training ofreaders and the need for readers to beblinded.

Answers
The document recommends that the reader should be skilled but does not
identify specific training that is needed. A technician reading withcardiologist over-read would certainly be consistent with the guidance.
The attempt of the guidance to limit the number of readers represented an
attempt to increase consistency. The guidance asks for assessment of
intra- and inter-reader variability and suggests "a few skilled readers" (notnecessarily a single reader) to analyse a whole thorough QT study, since
many readers may increase variability. Training would be another way to
improve consistency.
It is recommended for the thorough QT Study that core ECG laboratories
blind subject, time and treatment in order to reduce potential bias. The T
wave analysis, which calls for all 12 leads, can be performed after the QT
analyses, and requires comparison to the baseline ECG; it can, however,be blinded as to treatment.

1.2

June
2008

What is the position of ICH
regarding the role of the following
reading methods in the thoroughQT/QTc study and other clinical
trials?


fully manualfully automated

manualadjudication(manualover-read,computer-assisted,semiautomated)

The techniques currently in use for the measurement of ECG intervals
can be classified into three broad categories: fully manual, fully
automated, and manual adjudication. Within each of these generalcategories, many different methodologies are subsumed that differ interms of lead selection, the conventions used for defining T wave offset,
and the criteria for the inclusion and exclusion of U waves.
ECG readings can be performed on the following waveformpresentations:

Raw waveforms: ECG waveforms recorded from a single lead;

Representative waveforms (median beats, reference cycles):Compositional waveforms constructed by a computer-based5E14/S7B Q&As

averaging process that involves aligning and combining datafrom all dominant, normally conducted raw ECG waveforms
from a single lead;Global Waveforms:
Composite representation of cardiac
electrical activity constructed by superimposing representative
waveforms from all or several simultaneously recorded leads toform a spatial-vector complex, by weighted averaging ofindividual representative complexes with low noise and long
duration, or by other methods.

Fully Manual
When using a fully manual reading technique, a human reader is
responsible for examining the ECG waveform and placing the fiducial
points to mark the beginning and the end of the intervals, without theassistance of a computer algorithm. Fully manual methods of fiducial
point placement can be applied to raw, representative, and global
waveforms. When fully manual measurements are made from the raw
ECG waveforms in a single lead, three or more cycles should be averaged
where available to produce the final determination of interval duration.
An advantage of this approach is that the reader will not be influenced byprior computer placement of the fiducial points, but a weakness can beinter- and intra-reader variability, especially when measurements are
performed over an extended time period (e.g., several months).
Laboratories using manual reading techniques should observe standard
operating procedures based on prospectively defined criteria for
determining where the fiducial points should be placed. All readers in
the laboratory should be trained in the consistent application of thesecriteria.
Fully AutomatedFully automated reading methods rely entirely upon a computer
algorithm for the placement of the fiducial points and the measurementof the ECG intervals. Automated ECG interval measurements can be6E14/S7B Q&As

performed on raw, representative, or global ECG waveforms. Most
digital electrocardiographs are equipped with algorithms that performmeasurements on global waveforms. Although automated methods havethe advantage of being consistent and reproducible, they can yield
misleading results in the presence of noise or when dealing withabnormal ECG rhythms, low amplitude P or T waves, or overlapping U
waves. The techniques used for construction and measurement ofrepresentative waveforms and global waveforms vary between different
computerized algorithms and between different software versions within
individual equipment manufacturers. As a result, between-algorithm andwithin-manufacturer variability of fully automated measurements canconfound serial comparisons when the equipment or algorithm is notconstant.
ManualAdjudication
Assisted/Semi-Automated)(ManualOver-Read/Computer-

The manual adjudication approach refers to reading methods in which acomputer algorithm is responsible for the initial placement of the fiducial
points on the ECG waveform. A human reader subsequently reviews thealgorithmic placement of the fiducial points, performing adjustments
wherever the computerized measurements are considered to beinaccurate. This approach can have the advantage of greater consistency
and reproducibility than fully manual readings, while providing an
opportunity to correct any mistakes made by the algorithmic readings.
Laboratories using manual adjudication techniques should observe
standard operating procedures based on prospectively defined criteria for
determining when fiducial points should be corrected. All readers in thelaboratory should be trained in the consistent application of these criteria.
The adjudication procedure should normally be performed on all
waveforms being used for interval determination. If an alternative
approach is used, such as adjudication limited to outlier intervals aboveand below a reference range, this methodology should be validated as
described in Question 1.3.
7

E14/S7B Q&As

The ICH E14 Guideline currently recommends either fully manual or
manual adjudication approaches for clinical trials in which the assessmentof ECG safety is an important objective, such as the thorough QT/QTc
study. When the thorough QT study is positive, fully manual or manual
adjudication methods are currently recommended for an adequate sampleof patients in late phase studies (see Section 2.3 in E14 document). Whenthe thorough QT/QTc study is negative, routine ECG safety assessments
in late phase clinical trials using fully automated reading methods would
be adequate.
1.3

June
2008

The ICH E14 Guideline contains thefollowing statement: “If wellcharacterized data validating the useof fully automated technologies
becomeavailable,
therecommendations in the guidance forthe measurement of ECG intervals
could be modified.” What would be
expected of a sponsor that wished tovalidate and apply an automatedreading method for regulatory
submissions?

Efforts to develop more sophisticated and reliable methods for automated
ECG readings for both QT interval and T wave morphology assessment
are encouraged. There are at present no large scale studies to validate theuse of fully automated reading methods in patients; however, there areexamples of thorough QT/QTc studies in healthy volunteers in which
automated methods have been used and validated for QT intervalmeasurements against manual methods.
QT Interval measurement
There are at present no clear and widely accepted criteria for validationof new semi-automated or automated methods, but it is expected thateach would be validated independently for its ability to detect the
QT/QTc prolongation effects of drugs that are near the threshold of
regulatory concern. Data supporting the validation of a new methodshould be submitted and could include descriptive statistics, BlandAltman plots of agreement, superimposed plots of the baseline- andplacebo- adjusted QTc and the RR as a function of time, together withdata from any trials that have employed the method.
T wave morphology assessment
The suitability of automated ECG reading techniques for the assessmentof morphological abnormalities has not yet been demonstrated. If asponsor intends to develop a fully automated approach, without visual8E14/S7B Q&As

assessment for morphological changes, validation studies should include
a demonstration that the automated method is capable of reading andinterpreting a test set of abnormal ECGs correctly (e.g., abnormalities ofT wave morphology, overlapping U waves). As with methods for QTinterval determination, there are at present no clear and widely accepted
criteria for validation of novel methods.
Because changes in morphology can affect interval measurement, fully
manual or manual adjudication (as defined in Question 1.2) techniquesshould be performed if treatment-emergent changes in morphology areobserved. If, on the other hand, no morphology changes are observed,
this would support the use of automated methodologies, provided they
have been validated.
1.4

April
2012

How does a sponsor incorporate newtechnology
orvalidatenewmethodology into the measurement
and/or analysis of the QT interval?

The ICH process is better suited to the determination of regulatory policy
once the science in a particular area has become more or less clear. In
general, it is not well-suited to the qualification or validation of newtechnology.
Sections 2.5.1 and 2.5.2 of the ICH E14 Guideline are rather
discouraging about methodology outside conventional carts and humandetermined measurements. Since ICH E14 was issued, 12-leadcontinuous recording devices have largely supplanted cart recorders inthorough QT studies without a formal validation process because of theirperformance in the context of a positive control. The impact of other
innovative technologies can be assessed in studies incorporating a
positive control. While some technologies could be assessed using othertechniques in the absence of a positive control, this is more complex andbeyond the scope of this Q&A.
12-lead continuous recording devices and other new technologies can beused in late phase clinical trials. Even though a positive control is not
used in late stage studies, the new technology could be validated in otherstudies (such as the thorough QT study). In cases where a thorough QT

9

E14/S7B Q&As

study is not done, a sponsor can provide alternative methods for validating
the technology.
1.5

April
2012

The ICH E14 Guideline states that
QT interval corrected by Fridericia’sand Bazett’s correction should besubmitted in all applications; is thisstill necessary?
Is there a
recommended approach to QT
correction that is different than thatspecified in ICH E14?

Changes in heart rate could variably influence a drug’s effect on
repolarisation (i.e., QT interval) and correction methods with different
characteristics are often applied. The principles set below would beapplicable in all clinical studies (thorough QT or other studies).
In adults, Bazett’s correction has been clearly shown to be an inferior
method of correcting for differences in heart rate among and within
subjects. Therefore, QT interval data corrected using Bazett’s
corrections is no longer warranted in all applications unless there is acompelling reason for a comparison to historical Bazett’s corrected QT
data. Presentation of data with a Fridericia’s correction is likely to beappropriate in most situations but other methods could be more
appropriate. There is no single recommended alternative (see Q&A 1.4
on Incorporating New Technologies), but the following are someconsiderations.
1. Analyses of the same data using different models for correcting QT
can generate discordant results. Therefore, it is important that themethod(s) of correction, criteria for the selection of the method ofcorrection, and rationale for the components of the method ofcorrection be specified prior to analysis to limit bias. Model selection
ought to be based on objective criteria and consider the uncertainty inparameter estimates. Alternative methods of correction should be
used only if the primary method fails the pre-specified criteria for
selection of the method of correction.
2. Corrections that are individualized to a subject’s unique heart rate QT
dynamic are not likely to work well when the data are sparse or when
the baseline data upon which the correction is based do not cover at
least the heart rate range observed on study drug.

10E14/S7B Q&As

2. Gender#

Date of
Questions
Approval

Answers

2.1

June
2008

There are recognized differences inthe baseline QTc between men andwomen. These were noted in earlyversions of the guidance. In E14,
however, it is recommended thatoutliers be categorized as >450, >480
and >500 ms, regardless of gender.
Can you say why there is no genderdifference in the recommendation?

The 450, 480, and 500 ms categories refer to the values the E14 document
suggests sponsors might use in characterizing outliers. The numbers
previously specified for males and females referred to “normal” QTcvalues, which may differ for men and women. This section was notincluded in the final document, however, and such considerations would
be largely irrelevant to larger durations (e.g., 480, 500 ms). As thethorough QT/QTc study is designed to examine the propensity of a drug
to prolong the QTc ierval, it is appropriate to perform the study in maleor female healthy volunteers.

2.2

April
2012

Should we enroll both sexes in athorough QT study, and does thestudy need to be powered forindependent conclusions about eachsex?

Post-pubertal males have lower heart-rate corrected QT intervals than do
pre-pubertal males or females generally. Women are generally smaller
than men, so their exposure to a given fixed dose of a drug will
generally be higher, and, if a drug prolongs QT, it can be expected toprolong it more in women because of the higher exposure. It is notsettled whether and how often there are sex differences in response to
QT-prolonging drugs not explained by exposure alone.
The thorough QT study is primarily intended to act as a clinicalpharmacology study in a healthy population using a conservative
primary objective defining the drug’s effect on QT. It is unlikely thatany of a variety of baseline demographic parameters would introduce alarge difference in QT response to a drug in subpopulations defined byfactors such as age, co-morbidity, and gender that is not explained byexposure.
It is encouraged, but not mandatory, to include both men and women in
the thorough QT study.
Analyses of Concentration-Response
Relationship (CRR) by sex can be helpful for studying the effect of the
drug on QT/QTc interval in cases where there is evidence or mechanistic11

E14/S7B Q&As

theory for a gender difference. However, the primary analysis of athorough QT study should be powered and conducted on the pooled
population. If the primary analysis is negative and if there is no otherevidence suggesting gender differences, subgroup analysis by sex is notexpected.

3. Positive Control#
3.1

Date of
Questions
Approval
June
2008

The ICH E14 Guideline emphasizes
the importance of assay sensitivity
and recommends the use of a positive
control. In order to accept a negative
thorough QT/QTc study, assay
sensitivity should be established in
the study by use of a positive control
with a known QT-prolonging effect.
Please clarify how to assess theadequacy of the positive control in
the thorough QT study.

Answers
The positive control in a study is used to test the study’s ability (its “assay
sensitivity”) to detect the study endpoint of interest, in this case QT
prolongation by about 5 ms. If the study is able to detect such QT
prolongation by the control, then a finding of no QT effect of that size forthe test drug will constitute evidence that the test drug does not in fact
prolong the QT interval by the amount of regulatory concern. There are
two conditions required for ensuring such assay sensitivity:
1. The positive control should show a significant increase in QTc; i.e., thelower bound of the one-sided 95% Confidence Interval (CI) must be
above 0 ms. This shows that the trial is capable of detecting an increase
in QTc, a conclusion that is essential to concluding that a negative
finding for the test drug is meaningful;
2. The study should be able to detect an effect of about 5 ms (the QTc
threshold of regulatory concern) if it is present. Therefore, the size ofthe effect of the positive control is of particular relevance. With thisaim, there are at least two approaches:a. To use a positive control showing an effect of greater than 5 ms
(i.e., lower bound of a one-sided 95% CI >5 ms). This approachhas been proven to be useful in many regulatory cases. However,if the positive control has too large an effect, the study’s ability to12

E14/S7B Q&As

detect a 5 ms QTc prolongation might be questioned. In thissituation, the effect of the positive control could be examined attimes other than the peak effect to determine whether an effect close
to the threshold of regulatory concern can be detected;b. To use a positive control with an effect close to 5 ms (point estimateof the maximum mean difference with placebo close to 5 ms, witha one-sided 95% CI lower bound >0). In using positive controls
with smaller effects it would be very important to have a reasonably
precise estimate of the drug’s usual effect.
Importantly, whatever approach is used, the effect of the positive control(magnitude of peak and time course) should be reasonably similar to its
usual effect. Data suggesting an underestimation of QTc might question
the assay sensitivity, thus jeopardizing the interpretability of the thoroughQT study results.
3.2

June
2008

Please clarify the need for blinding
the positive control in the thoroughQT study.

The use of a double-blinded positive control does not appear to be
essential, provided that the reading of ECGs is performed in a blindedmanner as described in Question 1.2 and the study is carefully designed
to ensure that specified study procedures are followed uniformly. Thismeans that the same protocol for administering the test drug and placebo,taking blood samples and collecting the ECG data should also be used
when giving the positive control. This does not mean that other aspectsof the study, such as the duration of treatment with the positive control
and the other treatment groups, would be identical. If blinding of thepositive control is performed, common methods include the use of
double-dummy techniques and over-encapsulation.

13E14/S7B Q&As

4. Study Design
#
4.1

4.2

Date of
Questions
Approval
June
2008

June
2008

Answers

In ICH E14, the recommended
metric to analyse for a cross-overstudy is the largest time-matchedmean difference between the drug
and placebo (baseline-adjusted) overthe collection period. Please discuss
the most appropriate metric to assess
a drug’s effect on QT/QTc interval
when the data are collected in aplacebo-controlled parallel design
study (i.e., when there is nocorresponding placebo value foreach patient).

Regardless of the study design, “the largest time-matched meandifference between drug and placebo (baseline-adjusted)” is determined
as follows: The mean QTc for the drug (i.e., averaged across the study
population) is compared to the mean QTc for placebo (averaged acrossthe study population) at each time point. The “largest time-matchedmean difference between drug and placebo” is the largest of thesedifferences at any time point.

Please discuss the need for baseline
measurements, and when needed,
how they should be collected, forcross-over and parallel design
thorough QT studies.

Adjustment for baseline measurements is potentially useful for several
purposes, including detection of carry-over effects, reducing theinfluence of inter-subject differences and accounting for diurnal effects
such as those due to food. There is no single best approach for baseline
adjustment, but all planned baseline computations should beprospectively defined in the clinical trial protocol. Two kinds of baseline
are commonly used: “time-matched” baseline (taken at exactly the sametime-points on the day prior to the beginning of treatment as on thetreatment day) and "pre-dose" baseline (taken shortly prior to dosing).
The “pre-dose” baseline is used for adjustment for inter-subject
differences but not for diurnal effects. The choice of baseline is
influenced by whether the study is parallel or cross-over.

The term “baseline-adjusted” in ICH E14 implies that the baseline data
are taken into account in the statistical analysis.
Differences in baseline assessment between cross-over and parallel design
studies are discussed in Question 4.2.

14

E14/S7B Q&As

For a parallel-group study a time-matched baseline allows the detectionof differences in diurnal patterns between subjects that would not be
detected by a predose baseline. In a parallel study a “time-matched”
baseline day, if performed, would ideally occur on the day before the startof the study.
In contrast, in a cross-over study a time-matched baseline is usually notnecessary because adjustments for subject- and study-specific diurnal
variation are implicit by design in the assessment of time-matched drugplacebo differences in QT/QTc effect. The “pre-dose” baseline istherefore usually adequate for cross-over studies.
Obtaining replicate ECG measurements (for example, the average of theparameters from about 3 ECGs) within several minutes of each nominal
time point at baseline and at subsequent times will increase the precisionof the estimated changes in QT/QTc effect.

5. Use of Concentration Response Modeling of QTc Data#
5.1

Date of
Questions
Approval
The ICH E14 Guideline states (inSection 3.2.3, page 10) that analysisof the relationship between drugconcentration and QT/QTc intervalchanges is under active investigation.
Has this investigation yielded areasonableapproachtoconcentration-responsemodeling
during drug development? How canassessment of the concentration-

Answers
Concentration-response analysis, in which all relevant data across all
doses are used to characterize the potential for a drug to influence QTc,
can serve as an alternative to the by-time-point analysis or intersectionunion test as the primary basis for decisions to classify the risk of a drug.
In either case this result is an important component of the totality ofevidence assessment of the risk of QT prolongation. The overall
assessment of risk of QT prolongation includes nonclinical data, the timecourse of QT prolongation, the magnitude of QT prolongation,categorical analyses of outliers, and certain adverse events in patients thatcan signal potential proarrhythmic effects. There are many different typesof models for the analysis of concentration-response data, including15

E14/S7B Q&As

response relationship guideinterpretation of QTc data?

the descriptive pharmacodynamic (PD) models (e.g., linear or Emax models),or empirical models that link pharmacokinetic (PK) models (doseconcentration-response) with PD models. It is recognized thatconcentration-response analyses of the same data using models withdifferent underlying assumptions can generate discordant results.
Therefore, it is important that the modeling methods and assumptions,criteria for model selection, rationale for model components, andpotential for pooling of data across studies be specified prior to analysis
to limit bias. Prospective specification of model characteristics (e.g.,
structural model, objective criteria, goodness of fit) based on knowledgeof the pharmacology is recommended whenever possible. On occasion,
the QT effect is not a direct function of plasma concentration. For
example, drugs that cause QT prolongation as a result of changes inprotein synthesis or trafficking or drugs with accumulation intomyocardial tissues might demonstrate hysteresis. Testing for model
assumptions, hysteresis (a plot of data by-time point and a hysteresis loopplot), and goodness of fit should be documented. Concentration-response
analysis can be challenging when more than one molecular entitymultiple drugs or parent plus metabolites-contributes to the QTc effect.
Dose and exposure definitions


Therapeutic dose: dose evaluated in Phase 3 trial orrecommended in product labeling
Clinical exposure: mean steady state maximum concentration(Cmax,ss) associated with the maximum therapeutic doseHigh clinical exposure: exposure (Cmax,ss) achieved when the
maximum therapeutic dose is administered in the presence of the
intrinsic or extrinsic factor (e.g. organ impairment, drug-druginteraction, food effect, etc.) that has the largest effect on
increasing Cmax,ss

16

E14/S7B Q&AsSupratherapeutic dose: dose that provides exposures (mean Cmax)
exceeding the high clinical scenario

Important considerations
Concentration-response data need not come from a dedicated QT study,
nor even a single study, but there are several new and importantconsiderations.
1. Data can be acquired from first-in-human (FIH) studies, multipleascending dose (MAD) studies, or other studies provided that theconcentrations that can be safely achieved are well above theexposure at the maximum therapeutic dose at steady-state, andreflect high clinical exposure scenario situations such as drug-drugand drug-food interactions, organ dysfunction, and/or genetically
impaired metabolism. Whenever possible, sponsors are encouraged
to explore a wide dose range in early phase studies to enablecharacterization of effects at high concentrations.
2. Efficient concentration-response analysis using data acquired instudies with other purposes requires as much quality control as isneeded for a dedicated study. This includes robust, high-quality
electrocardiogram (ECG) recording and analysis sufficient to support
a valid assay for ECG intervals (see ICH E14 and Q&A #1).
3. If there is an intention to pool ECG interval data from multiple
studies, it is important to test for heterogeneity. Pooling of studies thatwere not planned for this purpose can produce bias. This potential
should be critically discussed in the analysis plan.
4. A separate positive control would not be necessary if either of thefollowing conditions is met:a) There are data characterizing the response at a sufficient
multiple (commonly 2x) of the high clinical exposure (seeICH E14 Section 2.2.2);
17

E14/S7B Q&As

orb) If the high clinical exposure has been achieved in the clinical
ECG assessment, but a sufficient multiple has not beenobtained (e.g., for reasons of safety or tolerability, saturating
absorption, etc.), then a nonclinical integrated risk assessment
can be used as supplementary evidence. The reason higher
doses were not tested should be adequately justified. See ICH
S7B Q&A 1.1 for details; in summary, the nonclinical studies
should include (1) a hERG assay, following best practiceconsiderations (see ICH S7B Q&A 2), that shows low risk as
defined in ICH S7B Q&As 1.1-1.2 and (2) no evidence of QTc
prolongation in an in vivo assay conducted according to ICH
S7B at exposures that cover high clinical exposures (see ICH
S7B Q&As 1.1 and 3; note that some recommendations only
apply to decision making under ICH Q&A 6.1).
Decision-making
Both the intersection-union test and the concentration-response analysis
can estimate the maximum effect of a drug treatment on the QTc interval,
but they are not used to test the same hypothesis. As mentioned above,
inspection of the time course of QT prolongation is important. However,hypothesis testing based on a by-time point analysis (intersection-union
test or point estimate and confidence intervals) is inappropriate in studies
designed for a concentration-response analysis, if not powered to assess
the magnitude of QT prolongation for each time point. When using a
concentration-response analysis as the primary basis for decisions toclassify the risk of a drug, the upper bound of the two-sided 90%
confidence interval for the QTc effect of a drug treatment as estimated byexposure-response analysis should be <10 ms at the highest clinically
relevant exposure to conclude that an expanded ECG safety evaluation

18

E14/S7B Q&As

during later stages of drug development is not needed. (See ICH E14,
Section 2.2.4 and Q&A #7).
Other uses
In addition to serving as the basis for regulatory decision-making,concentration-response analysis has established its utility in several
settings enumerated below.
Providing insight into regimens not studied directly
An understanding of the concentration-response relationship can help
predict the QT effects of doses, dosing regimens, routes ofadministration, or formulations that were not studied directly.
Interpolation within the range of concentrations studied is more reliable
than extrapolation above the range.
Predicting QTc effects of intrinsic and extrinsic factors that affect PK
Understanding the concentration-response relationship can help predict
the effects of intrinsic (e.g., cytochrome P450 isoenzyme status) or
extrinsic (e.g., drug-drug PK interactions) factors, possibly affecting
inclusion criteria or dosing adjustments in later phase studies.

6. Special Cases#
6.1

Date of
Questions
Approval
The ICH E14 Guideline states that incertain cases a conventional thoroughQT study might not be feasible. In
such cases what other methods
should be used for evaluation ofAnswers
An integrated nonclinical and clinical QT/QTc risk assessment can beparticularly valuable when a thorough QT study or concentration-QTc
analysis meeting similar quality control as needed for a dedicated study
as described in 5.1 is not feasible. This situation can arise under scenarios
where a placebo-controlled comparison is not possible; safety19E14/S7B Q&As

QT/QTcpotential?

andproarrhythmic considerations preclude administering supratherapeutic doses to obtainhigh clinical exposures and/or safety or tolerability prohibit the use of the
product in healthy participants. The design elements that include placebo
and healthy participant dosing assist in decreasing variability, but theirabsence does not preclude interpretation.
The integrated nonclinical and clinical QT/QTc risk assessment should
include:1.

The hERG assay, an in vivo QT assay, and any follow-up
nonclinical studies, especially those selected to overcome thechallenges encountered in the clinical studies (see ICH S7B
Q&As 1.1 and 1.2); and2.

Alternative QT clinical study designs incorporating ECG
assessments with as many of the usual “thorough QT/QTc”
design features as possible (see ICH E14 Section 2.2 and Q&A
5.1).

In situations where it is not possible to evaluate the QT/QTc effects athigh clinical exposure, it is particularly important that the nonclinical invivo studies are conducted at exposures covering the high clinicalexposure (see ICH E14 Q&A 5.1 for definition of high clinical exposure).
An integrated QT/QTc risk assessment can also be particularly valuablefor drugs with confounding heart rate effects that could impact accurate
determination of the QTc. Advanced methodologies for controlling (e.g.,
pacing) or correcting for heart rate changes in the nonclinical in vivo
studies and/or conducting QTc assessments in the intended patient
population might be informative in this situation. If tolerance to thechronotropic effect develops with repeat dosing, upward titration
regimens can sometimes be employed to avoid or minimize the20E14/S7B Q&As

confounding effects of drug-induced heart rate changes on the QTc
assessment.
Decision-Making
A totality of evidence argument based on the results of an integrated
nonclinical and clinical QT/QTc assessment could be made at the time ofmarketing application.
A drug that meets the following criteria would be considered to have a
low likelihood of proarrhythmic effects due to delayed repolarization:
1. The nonclinical studies, following best practice considerations
for in vitro studies (see ICH S7B Q&A 2) and in vivo studies(see ICH S7B Q&A 3), show low risk as defined in ICH S7B
Q&A 1.1.
2. The high-quality ECG data (see ICH E14 and E14 Q&A #1)
collected in the alternative QT clinical assessment do notsuggest QT prolongation, generally defined under this Q&A as
an upper bound of the two-sided 90% confidence intervalaround the estimated maximal effect on ΔQTc less than 10 ms,
as computed by the concentration-response analysis or theintersection-union test. If applicable, there should be nonotable imbalances between treatment/dose arms in the
proportion of subjects exceeding outlier thresholds.
3. A cardiovascular safety database that does not suggest
increased rate of adverse events that signal potential forproarrhythmic effects (ICH E14 Section 4).
When justified, a totality of evidence argument for a drug to have a low
likelihood of proarrhythmic effects due to delayed repolarization could
still be made for a drug that has an upper bound of the two-sided 90%
confidence interval around the estimated maximal effect on ΔQTc of 10
21

E14/S7B Q&As

ms or more. The determination will depend on the quality and details of
the clinical data (e.g., estimated ΔQTc mean and upper bound values,
slope of any concentration-ΔQTc relationship) and nonclinical data (e.g.,difference between the hERG safety margin for the investigational drug
and the threshold for defining low risk).
If nonclinical studies do not show low risk (or are not performed), there
is reluctance to conclude a lack of an effect in an absence of a positive
control; however, if the upper bound of the two-sided 90% confidence
interval around the estimated maximal effect on ΔQTc is less than 10 ms,
the treatment is unlikely to have an actual mean effect as large as 20 ms.
6.2

6.3

March
2014

March
2014

The ICH E14 Guideline does notaddress the approach to QT
measurement
duringdrug
development in the case of
combination drug products. Is it
recommended that measurement of
QT prolongation be performed on
drug combinations?

In general, combinations of two or more drugs are unlikely to need athorough QT/QTc study or intensive late stage monitoring, if thecomponent drugs have been demonstrated to lack relevant effects inthorough QT/QTc studies as described in ICH E14.

Are sponsors expected to conduct
thorough QT studies as part of thedevelopment of large proteins andmonoclonal antibodies?

Large targeted proteins and monoclonal antibodies have a low likelihoodof direct ion channel interactions and a thorough QT/QTc study is notnecessary, unless the potential for proarrhythmic risk is suggested bymechanistic considerations or data from clinical or non-clinical studies.

If one or more of the component drugs have not been individuallycharacterised for effects on the QT/QTc interval, they may be evaluatedin combination or independently.

22E14/S7B Q&As

7. Electrocardiograms Monitoring in Late Stage Clinical Trials#
7.1

Date of
Questions
Approval

Answers

April
2012

Clarification of Approach to Evaluating QTc in Late Stage ClinicalDevelopment

The ICH E14 Guideline describes in
Section 2.3 (Clinical Trial Evaluation
After the “Thorough QT/QTc
Study”) that “adequate ECG
assessment to accomplish this[monitoring]
is
not
fully
established.” Is there now areasonable approach to evaluating
QTc in late stage clinicaldevelopment in the case of a findingof QT prolongation prior to latephase studies?

The purpose of a thorough QT study is to characterize the effect of the
drug on ventricular repolarization (QT interval). It is not the purpose of
the thorough QT study to assess the risk of Torsade de Pointes (TdP) in
the target population, but rather to determine whether further data arewarranted to assess risk. A finding of QT prolongation above theregulatory threshold of interest (a positive thorough QT study) might callfor further electrocardiographic follow up in late phase studies. The
extent of the follow up would be affected by the magnitude of theestimated prolongation at doses and concentrations at which this occurs.
If prolongation is substantial at concentrations expected to occur inclinical studies, it is important to protect patients in later trials and toobtain further information on the frequency of marked QT prolongation.
In some cases in which there is a large margin of safety between
therapeutic exposures and the exposures that result in significant ECG
interval changes, an intensive ECG follow-up strategy might not be
warranted.
The recommended intensity of the monitoring and assessment in latestage trials will depend on:
A. The magnitude of QTc prolongation seen in the thorough QTstudy or early clinical studies.
B. The circumstances in which substantial QT prolongation mightoccur (i.e., in ordinary use or only when drug concentrations aremarkedly increased [e.g., by renal or hepatic impairment,
concomitant medications]).

23

E14/S7B Q&As

C. Pharmacokinetic properties of the drug (e.g., high inter-individual
variability in plasma concentrations, metabolites).
D. Characteristics of the target patient population that would
increase the proarrhythmic risk (e.g., structural heart disease).
E. The presence of adverse effects that can increase proarrhythmic
risk (e.g., hypokalemia, bradycardia, heart block).
F. Other characteristics of the drug (e.g., pharmacodynamics, safety
pharmacology, toxicology, drug class, hysteresis).
The following examples delineate the scope of recommended ECG
investigations based on outcome of the thorough QT study or earlyclinical studies. These could be modified by other factors such as A-F
above.
Examples of ECG Monitoring in Late Stage:
1. The thorough QT study results in a negative finding as defined by theE14 criteria* at the therapeutic dose, but the supratherapeutic dose
(relative to Phase 3 dose) shows mean QTc effects between 10 and
20 ms. If there is reasonable assurance that the higher dose represents
drug exposures that are unlikely to be seen in the patient population,
only routine ECG monitoring is recommended in late phase trials.
This approach provides reassurance for safety because patients areunlikely to experience a clinically significant QTc effect.
2. The thorough QT study results in a positive finding as defined by theE14 criteria* at the therapeutic dose, with a mean prolongation <20
ms. For drugs with this magnitude of effect on the QTc interval,
intensive monitoring of Phase 3 patients is called for.
Intensive ECG monitoring in clinical trials has two main objectives.
One objective is to provide protection to patients who might havelarge worrisome QT intervals >500 ms. A second objective is24E14/S7B Q&As

identifying the frequency of marked QT increases (e.g., prolonged
QT >500 ms or increases in QTc >60 ms).
Given the limitations of collecting ECGs in late stage trials, the focusof the analysis is on outliers, not on central tendency. Other thandescriptive statistics, detailed statistical analysis is not expected.
This monitoring is intended to be performed locally, without the
involvement of a central core laboratory.
The timing of ECG collection should be based on the known
properties of the drug. All patients should receive baseline, steadystate, and periodic ECGs during the trial. In addition, ECGs should
be collected around Tmax at the first dose and/or around steady state
in a subgroup of patients or in dedicated studies. ECG collection ataround Tmax is not important for drugs with low fluctuations between
peak and trough concentrations. If the drug shows a delayed effectin QT prolongation, then the timing of ECG collection should reflect
this delay.
3. The thorough QT study results in a negative finding as defined by theE14 criteria* at the therapeutic dose, but the supratherapeutic doseshows a mean effect between 10 and 20 ms. If supratherapeutic
exposure is anticipated at the clinical dose only in a wellcharacterized subgroup, intensive monitoring as described in
Example 2 above could be carried out in this subset of the Phase 3
population. In this case, there should be reasonable assurance thatthe higher exposure is unlikely to be seen in the general patient
population. In contrast, if people in the general patient population(who can not be readily identified in advance) will in some cases
achieve this higher exposure, intensive ECG monitoring in the Phase
3 population is expected, as in Example 2.
4. The therapeutic dose results in a mean QTc prolongation of >20 ms.
For drugs with large QTc prolongation effects, intensive ECG
assessment would be appropriate in all patients in Phase 2/3. Becauseof the risk of TdP, another important use of ECG monitoring in late25E14/S7B Q&As

phase trials would be to assess any risk mitigation strategies (e.g.,
electrolyte monitoring, dose reduction strategies). Additional ECG
assessment over and above what is recommended earlier in the Q&A
might also be called for (e.g., 24 hour ECG recording, telemetry,multiple trough ECGs through steady state).
The sponsor is encouraged to discuss these approaches with the relevant
regulatory agency(ies) prior to initiation of the Phase 3 program.
*A negative study as defined by the ICH E14 criteria is an upper onesided 95% CI of QTc prolongation effect <10 ms.

26

E14/S7B Q&As

S7B Q&As
1. Integrated Risk Assessment#
1.1

Date of
Questions
Approval
What is the general strategy for useof nonclinical information as part ofan integrated risk assessment fordelayed ventricular repolarizationand torsade de pointes that can
inform the design of clinicalinvestigations and interpretation of
their results?

Answers
The ICH S7B guideline describes a nonclinical strategy for assessing riskof delayed ventricular repolarization and QT interval prolongation(Section 2.3). A mechanistic understanding of the development of torsadede pointes (TdP) and the emergence of new types of assays have made it
possible to obtain more information to assess TdP risk from nonclinicalassays.
The in vitro IKr/hERG assay and in vivo QT assay as well as optional
follow-up studies, as described by the ICH S7B guideline, are conducted
for hazard identification and risk assessment relevant to delayed
ventricular repolarization. It is generally accepted that drugs (note thatthe word “drug(s)” in the Q&As is used interchangeably with word“pharmaceutical(s)” in ICH S7B) that delay ventricular repolarization
may have increased risk of TdP.
In addition to supporting the planning and interpretation of First-inHuman clinical studies, nonclinical investigations can also contribute to
an integrated risk assessment for TdP in later stages of development whenclinical data are available. The following are points to consider whenusing in vitro IKr/hERG data and in vivo QT data in combination withclinical QT data as part of an integrated risk assessment for situations
described in ICH E14 Q&As 5.1 & 6.1.
1. To predict whether or not the hERG block poses a risk ofinterfering with ventricular repolarization or TdP, evaluation of
the hERG safety margin based on results of a best practice assay(see S7B Q&As 1.2 and 2.1) is recommended. ICH S7A Section27

E14/S7B Q&As

2.6 describes considerations for when human metabolite(s) should
be assessed with in vitro systems. In these cases, the metabolite’shERG safety margin should also be evaluated.
2. In the in vivo study, the effects on the QTc interval should beassessed at exposures that cover the anticipated high clinicalexposure scenario. The adequacy of exposure to any major humanmetabolites should be determined (see ICH S7A Sections 2.3.3.2& 2.6, and S7B Q&A 3.5). In addition, if the assay is to be usedas part of an integrated clinical and nonclinical risk assessment forsituations where a conventional thorough QT study is not feasible
as described in ICH E14 Q&A 6.1, the in vivo study should havesufficient sensitivity to detect a QTc prolongation effect of amagnitude similar to dedicated clinical QT studies (see ICH S7B
Q&A 3.4). This additional consideration (sensitivity similar to
dedicated clinical QT studies) does not apply to decision making
prior to First-in-Human studies or under ICH E14 Q&A 5.1.
A drug with low TdP risk would be expected to have (1) a hERG safetymargin higher than a threshold defined based on the safety margins
computed under the same experimental protocol for a series of drugs
known to cause TdP (see ICH S7B Q&A 1.2 for additional details); and(2) no QTc prolongation in an in vivo assay of sufficient sensitivity
conducted at exposures of parent compound and major human metabolites
that exceed clinical exposures. See ICH S7A Sec. 2.6 for considerationsof human metabolite(s) in the in vitro or in vivo nonclinical assays. If theseresults are used to support an integrated clinical and nonclinical risk
assessment strategy as described in ICH E14 Q&As 5.1 & 6.1, noadditional nonclinical studies are needed. When there are factors that canconfound or limit the interpretation of the nonclinical studies, such as
metabolites and heart rate changes, follow-up studies as described by ICH
S7B (Section 2.3.5) can be performed to address these specific issues.

28E14/S7B Q&As

If the hERG assay and/or the in vivo QT study suggest an effect at clinicalexposures, the drug has a risk of interfering with ventricularrepolarization. Under this scenario, the drug’s TdP risk could be affectedby various other factors, such as blocking of additional repolarization
currents (e.g., IKs), blocking of inward currents (e.g., sodium and L-type
calcium currents), effects on the trafficking of ion channel proteins fromcytoplasmic sites to the surface membrane, metabolites with ion channel
activities, and non-ion channel mediated QT prolongation. Follow-up
studies (ICH S7B Section 2.3.5) could be performed to further explore themechanisms and assess the TdP risk. If applicable, best practiceconsiderations should be followed for assessment of additional ion
channel currents (S7B Q&A 2.1), in vitro cardiomyocyte assays (S7B
Q&As 2.2–2.5), or in vivo studies (S7B Q&As 3.1–3.5). An appropriately
qualified proarrhythmia risk prediction model (see S7B Q&As 4.1–4.2)
could be used according to its context of use to assess the possibility of
TdP in humans. Use of in vitro and in silico models can reduce animal use
in follow-up studies in accordance with the 3R (reduce/refine/replace)principles. The assessment of TdP risk using these follow-up studies,
although optional, can be used together with other relevant nonclinicaland clinical information to contribute to the design of subsequent clinicalinvestigations and interpretation of their results.
1.2

What is the recommended method to A drug’s potency for hERG block, usually calculated as half-inhibitorycompute the hERG safety margin?
concentration (IC50), can be normalized to the drug’s estimated clinically
relevant exposures in patients to calculate the safety margin. As more
information is obtained during the clinical development, the estimatedvalues of clinical exposures can be refined. When estimating hERG
block potency, it is recommended to use standardized procedures and to
consider the principles described in S7B Q&A 2.1.
The free drug exposure is computed based on the drug’s total plasma
concentration and the fraction of protein binding. Because of uncertainties
29E14/S7B Q&As

in the protein binding measurements, the unbound (free) fraction inplasma should be set to 1% if experimentally determined to be < 1%. Thisapproach has been used in the regulatory assessment for the risk of drugdrug interactions. If protein binding values cannot be accurately assessed(e.g., questionable validation of the bioanalytical method, deviations frombest practices), safety margins should be calculated for both steady-state
free and total Cmax.
In terms of the exposure to use in the denominator of the safety margin
calculation for supporting decision making under ICH E14 Q&As 5.1 or
6.1, it is generally recommended to use the high clinical exposure as
defined in ICH E14 Q&A 5.1 (i.e., mean steady state maximum
concentration [Cmax,ss] when the maximum therapeutic dose isadministered in the presence of the intrinsic or extrinsic factor that has thelargest effect on increasing Cmax,ss).
To assess whether the hERG block poses a risk of delaying ventricularrepolarization or TdP, the resulting safety margin should be compared to
the range of safety margins computed under the same experimental
protocol for a series of reference drugs that have known clinical TdP risk
and cover diverse electrophysiological properties. Additional
pharmacological principles or modeling can be used to justify the use ofa certain safety margin threshold (e.g., relationship between hERG blockandQTc
prolongation
fromsemi-mechanistic
pharmacokinetic/pharmacodynamic models or systems pharmacology
models; see Leishman et al. J Pharmacol Toxicol Methods 2020
[https://doi.org/10.1016/j.vascn.2020.106900], for examples). Thisshould be supported by experimental data based on the principles in these
Q&As (e.g., same experimental protocol applied to a series of drugs withknown clinical TdP risk).
Data supporting the safety margin threshold to define a drug as having
low TdP risk should be supplied in or appended to the submitted study30

E14/S7B Q&As

report. If a recommended hERG margin threshold is published based onprinciples in these Q&As, a sponsor (or a contract laboratory) seeking to
use the same threshold should demonstrate that the inter-laboratory
variability of IC50s from a set of calibration drugs under the same
experimental protocol does not significantly decrease the sensitivity of thesafety margin threshold for detecting drugs that are not low risk for TdP.
Appropriate statistical methods should be applied to quantify
experimental IC50 variability and calculate uncertainty of the safetymargin as confidence/credible intervals.
2. Best Practice Considerations for In vitro Studies#
2.1

Date of
Questions
Approval
What are some “best practice”
considerations when evaluating drug
potency on affecting cardiac ioniccurrents using patch clamp method
and overexpression cell lines?

Answers
As outlined in ICH S7B, the in vitro IKr/hERG assay plays a critical rolein assessing the risk for delayed repolarization and QT interval
prolongation prior to first administration in humans. Nonclinicalinvestigations can also contribute to an integrated risk assessment in laterstages of development when clinical QT data are available. The
following “best practice” aspects should be considered when sponsors
are using IKr/hERG data to support interpretation of clinical QT data in
specific scenarios as described in S7B Q&As 1.1 & 1.2 and ICH E14
Q&As 5.1 & 6.1, and when using CaV1.2 and NaV1.5 to support aproarrhythmia assessment (ICH S7B Q&As 1.1). It is not the intent ofthese Q&As to make specific recommendations for a sponsor’s screening
activities or for all IKr/hERG assays to support first administration inhumans.
Several experimental factors are known to influence the potency of drug
effects on cardiac ionic currents. These include the voltage protocols
used to evoke specific ionic currents, experimental conditions (such as31

E14/S7B Q&As

recording temperature, composition of solutions, manual vs. automatedassay systems), data acceptance criteria, and data analysis methods
employed. Some recommended best practices are therefore provided toenhance reproducibility of in vitro results and the translation to clinicalfindings. These recommendations are generalizable to voltage clamp
experiments characterizing potency of drug inhibition (or potentiation)of cardiac currents.
1.

Recording temperature: The effects of some drugs aretemperature-sensitive and there is currently no method topredict which molecules exhibit temperature-dependent
effects or the magnitude of these effects. Thus, patch clamp
experiments on cells overexpressing cardiac ion channels,
including hERG, CaV1.2, and NaV1.5, should be performed atnear physiological temperature (35–37 °C).

2.

Voltage protocol: The voltage protocols used to evoke ioniccurrents should approximate the appropriate elements of aventricular action potential and be repeated at frequencies thatare sufficient to minimize the possibility of missing theeffects of a test drug at physiologically relevant heart rates.
For hERG, a stimulation frequency of 0.2 – 1 Hz is
recommended. For CaV1.2 and NaV1.5 currents, a stimulation
frequency of 0.2 Hz is recommended. The voltage protocol
should include steps that enable monitoring of cell health andconsistent electrophysiological recordings throughout theexperiment (i.e., estimation of input and series resistanceacross time). If high seal resistance is achieved, holding
current and input resistance (i.e., measures of passive
membrane properties at rest) can be used as indicators of cell
health and experimental stability. After application of the test
drug and if recording quality remains acceptable, a saturating32

E14/S7B Q&As

concentration of a selective blocker should be applied to cells
to determine residual background current in individual cells.
If prominent, background current should be factored into
potency determinations.
3.

Recording quality: Seal resistance should be high enough sothat the leak conductance at all voltages specified by thevoltage protocol and series resistance do not compromise
voltage control. The extent of series resistance compensation
applied to optimize voltage control should be noted. Stabilityof the ionic current should be demonstrated with baseline
recordings (prior to drug application) of sufficient duration tocharacterize drug-independent changes (such as current rundown). The time course of drug effects should be monitored
until steady state effect is obtained, and each cell can be
exposed to one or more drug concentrations as long as cell
health and recording quality remain stable.

4.

Primary endpoint measures: The primary derived endpoints
are inhibitory concentration such as the IC50 value (reported
in both micromolar and ng/mL units) and Hill coefficient. If
50% current inhibition could not be achieved, a justificationof the highest concentration tested should be provided
together with the relation of this concentration to therapeutic
free and total drug levels. Where necessary, to isolate thecurrent-of-interest, the background current remaining after ahigh concentration of selective blocker application should be
subtracted. If current inhibition with a selective blocker could
not be achieved, leak current can be calculated and subtracted
from the current traces. This approach assumes that only thecurrent-of-interest is voltage-dependent, hence evidence andjustification should be provided on why it was used.
33

E14/S7B Q&As

5.

Data summary: Inhibition at each drug concentration for eachcell should be provided, along with the mean values of IC50
and Hill coefficient (and appropriate measures of data
variability). To demonstrate recording quality, the study
report should also contain time-course plots of current
amplitude, input resistance, holding current for individual
cells in control condition followed by drug application, anddrug equilibration. If time-dependent changes such as current
run-up or run-down in baseline condition were corrected for
drug inhibition estimation, the correction method appliedshould be described.

6.

Concentration verification: The concentration of test
compound to which the cells were exposed should be verified
by applying a validated analytical method to the solution
collected from the cell chamber. Both nominal and measured
concentrations should be reported. If the nominal andmeasured concentrations differ significantly from each other,measured concentrations should be used to construct theconcentration-response relationship to estimate IC50 and Hillcoefficient.

7.

Positive and negative controls: The positive control drug
should be one of the “reference drugs” referred to in Q&A
1.2. The positive control drug should be tested using
sufficient replicates and two or more concentrations
achieving 20-80% block, to demonstrate consistency andreproducibility with the reference drug data. If positive
control data fall outside the range of expected values, then the
study is inconclusive, and it would not be recommended thatthe data be used to support the purposes outlined in ICH E14
Q&As 5.1 and 6.1. Vehicle (negative) controls should be34

E14/S7B Q&As

included in the experiments. The vehicle should include all
non-compound materials in the test article solution such assolubilizing agents and preservatives.
2.2

What are the relevant endpoints of an
informative
in
vitro
human
cardiomyocyte repolarization followup study?

As outlined in ICH S7B, follow-up studies (Section 2.3.5) can include invitro ventricular repolarization assays. Follow-up studies are not
performed for all submissions and are often designed to address specificissues. Since implementation of ICH S7B, new technologies havebecome available, including assays with human induced pluripotent stem
cell derived cardiomyocytes (hiPSC-CMs). S7B Q&As 2.2–2.5 outline
best practice considerations when in vitro cardiomyocyte assays are
performed as follow-up studies.
Drug-induced changes in the intracellular or extracellular action potential
waveforms recorded from hiPSC-CM preparations and acutely isolated
adult human ventricular myocytes reflect the integrated effect on
multiple ionic currents, exchangers, and carriers. Changes in cellular
repolarization recognized as markers of ventricular proarrhythmia
include delayed and abnormal repolarization (manifest as early
afterdepolarizations, triggered activity or irregular beating) and should
be noted.
Drug induced effects on cardiomyocyte calcium handling (calcium
transients, contractility, or calcium homeostasis) could affectelectrophysiological activity. These effects may manifest as abnormal
mechanical activity associated with proarrhythmic mechanisms (e.g.,
prolonged contractions, single or multiple premature contractions linked
to triggered electrical activity such as early afterdepolarizations). The
assessment of changes in calcium handling is useful for describing thepresence or absence of potential proarrhythmic activity. However, suchmeasures are inadequate to fully characterize the electrophysiologicaleffects of a drug on repolarization.
35

E14/S7B Q&As

2.3

What elements of the test system need It is important to describe the biological preparation and technology
to be considered for an in vitro human platform that define baseline electrophysiological characteristics andcardiomyocyte repolarization assay? drug responses.


Biological preparation: The origins of cells studied, and humandonor characteristics should be specified. If complex preparations
containing hiPSC-CMs are used (e.g., co-cultures, organoids,
engineered heart tissues), descriptions of the protocols used increating these preparations should be provided. For primary
human cardiomyocyte preparations, the tissue sources,
harvesting, isolation, and enrichment procedures followed should
be described. Acceptable morphological and functional inclusion
criteria for the preparations as well as electrophysiologic
characteristics (including baseline action potential/field potential
durations, spontaneous beat frequency and variability [ifapplicable], resting membrane potential, upstroke characteristics,
conduction patterns and/or velocity) should be clearly defined.
Estimates of the proportion of preparations fulfilling criteria
should be included.
Technology platform: The methodologies used (e.g.,transmembrane potential recordings [whole cell patch clamp,
sharp electrode, or voltage-sensing dye approaches], extracellular
recordings using field potentials, visual or impedance-based
motion approaches, or calcium-sensing dyes) should be clearly
described. The analysis package used for marking andinterpreting waveforms should be described, with representative
recordings (along with pertinent waveform markings) provided.
A description of the plates or chambers used (including presenceor absence of flow, substrate composition, recording electrodecharacteristics) should be provided.

36

E14/S7B Q&As

2.4

What are important considerations
when designing and implementing
experimental protocols for in vitro
cardiomyocyte repolarization studies?

Protocols should be designed to address a specific question (e.g.,concentration-dependent effects on repolarization). The rationale ofchoosing a single- or sequential-dose protocol should be provided. Bathtemperature should be stable at physiologic (35–37 ºC) temperature. The
sampling “window” for data collection should be clearly defined.
Deviations from protocols should be clearly described, along withexpected consequences.








For spontaneously beating preparations, changes in beating rate
influence repolarization independent of direct drug effects on
repolarizing currents. Spontaneous beat rates in the absence andpresence of drugs should be clearly indicated along with theextent of drug-induced rate changes. The choice and justificationof correction formula used when assessing repolarization effects
in such preparations should be provided. Due to limitations ofrate correction in spontaneously beating hiPSC-CMs,interpretation of potential repolarization changes may not be
possible when a drug causes a rate change.
For paced preparations, the pacing protocol (pattern and duration)
should be described and performed in the presence and absenceof test compound.
To demonstrate recording quality, the study report should containtime-course plots of primary endpoints (demonstrating drug
equilibration) and other parameters that can be used to infer
stability of the preparations and signal recordings.
Concentration-dependent repolarization effects can be derived
based on vehicle-corrected and/or baseline subtracted
comparisons of drug vs. vehicle treated preparations. For higher
throughput multi-well platforms, it is preferable to conduct
vehicle and test drug studies on the same plate. The number of
replicates (useful for evaluating reproducibility but not inferential37E14/S7B Q&As



2.5

statistical testing) should be reported. Power calculations arehelpful to establish statistical sensitivity to repolarization
endpoints.
It is important to characterize drug exposures during in vitro
cardiomyocyte repolarization studies. For well-based studies,drug exposures could be verified using media sampled from test
wells or from “satellite studies” (parallel studies using identical
protocols and study conditions conducted without measuring
electrophysiologic measurements).
With continuous flow
systems the sampling of effluent from test chambers is valuable
for assessing drug exposures. Exposures should be presented as
total drug concentration and free drug concentrations (if plasma
protein binding characteristics in the media used is known).

How does one define biological The electrophysiologic sensitivity of cardiomyocyte preparations should
sensitivity of a cardiomyocyte in vitro be calibrated with established positive controls to confirm their “fit for
repolarization assay?
purpose” role in defining pharmacological block of cardiac ionchannel(s). This is readily accomplished by constructing concentrationresponse curves with recognized and specific ion current blocking agents.
At minimum, it is important to characterize sensitivity to block of
the prominent outward repolarizing current IKr/hERG withspecific blocking agents (e.g., E-4031 or dofetilide) over relevant
concentration ranges.
Block of the inward L-type calcium current (ICaL) and late sodium
current (INaL) may mitigate delayed repolarization.
Demonstrating sensitivity to specific ICaL (e.g., nifedipine or
nisoldipine) and INaL (e.g., mexiletine or lidocaine) blocking
agents is helpful for clarifying integrated cellularelectrophysiological responses of multi-channel blocking drugs.

38E14/S7B Q&As

3. Best Practice Considerations for the In vivo QT Studies#
3.1

Date of
Questions
Approval
What are best practice considerations
for species selection and generaldesign of the (standard) in vivo QT
study?

Answers
The most appropriate species should be selected and justified (ICH S7B,
Section 3.1.3). It is preferable to use the same animal species in the safety
pharmacology and non-rodent toxicity studies to facilitate understandingof the possible relationship between adverse cardiovascularpharmacodynamic effects and structural effects on the heart, and toobtain complementary information on systemic exposure level(toxicokinetics).
While it is customary to use conscious freely moving telemeterizedanimals for the in vivo QT studies, the choice of alternative model
approaches (e.g., anesthetized or paced animals) might be justified incertain circumstances to achieve adequate exposures or to overcome
specificcompound-related
challenges(e.g., changes in heartrate, tolerability, or bioavailability limitations in conscious animals).

3.2

What should be considered for The ICH S7B guideline states that drug exposures should include andexposure assessment during the in exceed anticipated therapeutic concentrations. If the in vivo QT data arevivo QT study?
to be used as part of an integrated risk assessment for situations describedin ICH E14 Q&As 5.1 & 6.1, the exposure should cover the anticipatedhigh clinical exposure scenario (see S7B Q&A 1.1). An assessment of
exposure in the same animals used for the pharmacodynamic assessment
is encouraged. Sampling should take place at relevant timepoints and in amanner that limits interference with the pharmacodynamic effects. This
could be done by sampling complete pharmacokinetic profiles in the same
animals on a separate day after an adequate washout or different animals.
At least one pharmacokinetic sample should be obtained during thepharmacodynamic assessment day to demonstrate consistency with thefull pharmacokinetic profiles. In certain cases, the analysis of QTc interval
39E14/S7B Q&As

together with adequate pharmacokinetic sampling makes it possible toperform dedicated exposure-response modeling similar to concentrationQT analysis for clinical QT studies. This can be helpful when the study
should be powered to detect an effect similar to dedicated QT studies inhumans (e.g., when using in vivo QT data as part of an integrated
nonclinical and clinical risk assessment as described in ICH E14 Q&A
6.1) as it can reduce the number of animals in accordance with the 3R(reduce/refine/replace) principles. In addition, exposure-responsemodeling may be helpful in other circumstances when QT prolongation isobserved or anticipated based on hERG assay results.
3.3

What information is needed tosupport the choice of heart rate
correction method in an in vivo QT
assay?

Optimally, the sponsor should demonstrate the independence of QTc to
RR intervals observed in the study through QTc versus RR plots
accompanied by additional information (e.g., number of matched QTc-RR
pairs, correlation metric, 95% confidence intervals, p-values). QT-RR
interval relationship is also important. Justification of correction factors
used for QT measures should be provided when test drugs affect heart rate.
In certain cases, individual QT correction based on QT-RR relationship isa preferred method as it is more accurate and sensitive than the general
methods such as Bazett, Fridericia, or Van de Water when the test drugs
affect heart rate. The main reason for not using correction formulae basedon historical data is the fixed rate correction coefficients. Non-rodent
species show species-specific and individual differences in their QT-RR
relationships.

3.4

How should the sensitivity of the The test system used for an in vivo QT assay should provide a robust
assay be evaluated?
response. Assay sensitivity of relevant functional endpoints should be
evaluated and reported to enable data interpretation (in supporting
initiating first-in-human studies and/or an integrated nonclinical andclinical integrated risk assessment to be applied under the scenarios in ICH
E14 Q&As 5.1 or 6.1) and contextualization. Demonstration of assay
sensitivity can be achieved by defining minimum detectable differences40E14/S7B Q&As

and testing the effects of positive controls. Statistical power calculations
could also be provided from historical data from the same laboratory using
the identical protocol. If historical positive control data are utilized tojustify assay sensitivity or statistical power is calculated from historicalcontrol data, then the variance of the present data should be consistent
with that seen historically.
If study results are to be used to support an integrated nonclinical andclinical risk assessment described in ICH E14 Q&A 6.1, then the study
should have sensitivity to detect a QTc prolongation effect of a magnitude
similar to dedicated clinical QT studies, taking into consideration interspecies differences in the normal range of values for the QTc interval. The
overall sensitivity of the nonclinical assay in comparison to clinical QT
studies depends on both the electrocardiographic assessment and theexposure achieved in the in vivo assay relative to high clinical exposure.
This can help reduce the numbers of animals used in accordance with the
3R (reduce/refine/replace) principles. The following hypotheticalexample is offered for consideration with recognition that the QTc
threshold and exposure multiples selected for a particular study should bejustified by data obtained in the specific species tested, using recognizedreference compounds under conditions consistent with the best practicerecommendations set forth in these Q&As.


3.5

Hypothetical Example: The minimal detectable difference mightbe 5 milliseconds if drug exposure in the animal study only covers
the high clinical exposure but might be higher if a larger multipleof high clinical exposure is achieved (e.g., 10 milliseconds if 3X
high clinical exposure is achieved; or a higher QTc threshold if aneven larger multiple is achieved).

What
are
the
recommended To facilitate the regulatory review of an in vivo QT assay, the following
conventions for presenting the are general recommendations.
pharmacodynamic and41

E14/S7B Q&As

pharmacokinetic results of an in vivo
Pharmacodynamic Content
QT assay?
• Summary tables and figures showing absolute mean values, mean
percent change from baseline, confidence intervals, and p-values
for changes from baseline and vehicle control should be included.
• If study results are being used to support ICH E14 Q&A 6.1, thendata from positive controls should be included or appended. If
historical positive controls are used, then the variance of the present
data should be consistent with that seen historically, which can bedemonstrated by reporting minimal detectable differences with bytime analysis. A statement should be provided that the data for the
new drug and the historical data were collected according to thesame protocol and statistical analysis plan. If deviations are present,
they should be clearly justified. If concentration-QTc modeling is
performed, reporting should follow similar principles as for humanconcentration-QTc modeling (see ICH E14 Q&A 5.1).
Pharmacokinetic Content• Tabulations of summary statistics for Cmax, AUC, and Tmax for theparent drug and metabolites along with plasma concentration vs.
time plots (if sufficient samples have been collected to support theircalculation) should be provided.
Individual animal data should be provided.

42E14/S7B Q&As

4. Principles for Proarrhythmia Models#
4.1

Date of
Questions
Approval
The ICH S7B guideline (Section
3.1.4) states that directly assessing theproarrhythmicriskofpharmaceuticals that prolong the QTinterval would be a logicalundertaking and interested parties areencouraged to develop these models
and test their usefulness in predicting
risk in humans. What are generalprinciples to evaluate whether aproarrhythmic risk prediction model
could be used as part of an integrated
risk assessment strategy?

Answers
Different models, including in silico, in vitro, ex vivo and in vivo models,
have the potential to be used as part of an integrated risk assessment
strategy to evaluate the proarrhythmic risk of QT-prolonging
pharmaceuticals in humans. Use of in vitro and in silico models can also
reduce animal use in accordance with the 3R (reduce/refine/replace)principles. Because these models have a common feature of using
nonclinical experimental data as input and generating humanproarrhythmia risk prediction as output, they can generally be referred toas proarrhythmia risk prediction models. The model input can vary
among different models, for example, ion channel pharmacology data as
input to in silico models, drug-induced changes in cellular repolarization
and/or arrhythmia events as input to hiPSC-CM models, and druginduced electrocardiographic changes as input to ex vivo/in vivo models.
However, the model output (either discrete risk categories or continuous
risk scores) is similar among different models. Such a feature makes it
possible to develop generic principles for evaluating the predictivity ofproarrhythmia risk prediction models without specifying the type ofunderlying experimental data used as model input. The following
general principles should be applied to all proarrhythmia risk prediction
models intended to be used as part of an integrated risk assessment for
regulatory purposes. While the main focus of these principles is toevaluate a model’s predictivity of TdP risk, they are general enough to
guide the development of models predicting different types ofproarrhythmia.
1.

A defined endpoint consistent with the context of use of themodel. Examples of model endpoints (e.g., TdP risk vs QT

43

E14/S7B Q&As

prolongation risk) can be found in Li et al. Clin Pharmacol
Ther 2020 (https://doi.org/10.1002/cpt.1647).
2.

A fully disclosed algorithm to translate experimental
measurements (model input) to proarrhythmia risk (model
output), allowing independent reproduction of the model
development process using the associated training andvalidation datasets to evaluate the model performance.

3.

A defined domain of applicability/ scope and limitations of
the model. This includes the experimental protocols togenerate model input (experimental data capturing
pharmacological effects of drug), and the compounds tested
should have the same arrhythmic mechanisms covered by themodel.

4.

A prespecified analysis plan and criteria to assess model
predictivity. The analysis plan should include methods toseparate the training and validation steps. In the training step,a series of reference compounds is used to adjust the model.
In the validation step, another series of reference compounds
is used to evaluate the performance of the pre-specified
model. The reference compounds used for the training andvalidation steps should not overlap.

5.

A mechanistic interpretation of the model, which describes
the relationship between the model inputs and mechanism forthe arrhythmia.

6.

The uncertainty in the model inputs should be captured andpropagated to the model predictions. The experimental
variability associated with model input should be quantified
using appropriate statistical methods and then translated into
probabilities of the predicted risk.

After a proarrhythmia risk prediction model has been developed, aprocess should be followed to evaluate whether the model development
44E14/S7B Q&As

complied with these principles. Such a process would support that themodel is qualified for the intended context of use as part of an integrated
risk assessment for regulatory purposes. Some health authorities haveprocedures for the formal qualification of models that allow for a model
to be used within the qualified context of use without the regulatory
authority needing to reconsider and reconfirm its suitability. Modeldevelopers are encouraged to contact a regulatory agency about its
specific model qualification procedures. After a model has been
qualified, the use of such a model is not limited to the specific facility
that submitted the qualification package. However, if another facility
intends to use the qualified model, that facility should perform
laboratory-specific calibration and validation of the model using a subsetof the reference compounds that were originally used to develop themodel. An illustrative process of performing laboratory-specificcalibration and validation is given in Han et al. J Pharmacol Toxicol
Methods 2020 (https://doi.org/10.1016/j.vascn.2020.106890).
4.2

How can a sponsor use a model for Sponsors can use results from a qualified proarrhythmia model as one
regulatory submission and what are component in the totality of evidence approach to risk assessment under
the limitations?
the context of use for which the model was developed and qualified.
When a facility intends to use the model to produce data for regulatory
submission, a set of control compound(s) should be tested to assess theconsistency between the new data and the historical laboratory-specificvalidation data. The number and type of laboratory-specific calibrationand control compounds should be justified.
If a proarrhythmia model is included in a regulatory submission, proof of
qualification of the model under the guidance of the general principles inthis Q&A should be provided in an appendix to the study report.
Supportive documentation could include published papers, if the
included validation dataset is described in sufficient detail to allow an
independent assessment. Importantly, the general principles for model
qualification set forth in this Q&A only support the use of a45

E14/S7B Q&As

proarrhythmia risk prediction model as part of an integrated risk
assessment that incorporates all relevant nonclinical and clinicalinformation.

46