HERG assay and cardiac safety evaluation MCQs With Answer

Introduction:
This quiz collection focuses on hERG (human Ether-à-go-go-Related Gene) assays and cardiac safety evaluation — a core topic in Pharmacological & Toxicological Screening Methods-II for M.Pharm students. You will find practical and theory-driven multiple-choice questions that delve into hERG channel physiology, electrophysiological techniques (manual and automated patch-clamp), assay artefacts, interpretation of IC50 and safety margins, CiPA paradigms, in silico models, metabolite assessment, and ECG biomarkers relevant to proarrhythmic risk. These items are designed to deepen understanding of how preclinical data translate to clinical QT/Torsades risk assessment and regulatory expectations, while reinforcing critical experimental and analytical concepts used in modern cardiac safety pharmacology.

Q1. What best describes the hERG channel relevant to cardiac safety testing?

• Voltage-gated potassium channel responsible for the rapid delayed rectifier current (IKr)
• Voltage-gated sodium channel responsible for INa
• L-type calcium channel responsible for ICa-L
• Funny current (If) channel in sinoatrial node

Correct Answer: Voltage-gated potassium channel responsible for the rapid delayed rectifier current (IKr)

Q2. The primary clinical concern when a drug blocks hERG channels is:

• Prolongation of the QT interval and increased risk of Torsades de Pointes
• Immediate hypotension due to vascular K+ channel blockade
• Isolated increase in heart rate without repolarization changes
• Elevation of cardiac enzymes indicating myocardial necrosis

Correct Answer: Prolongation of the QT interval and increased risk of Torsades de Pointes

Q3. Which experimental technique provides the gold-standard measurement of hERG channel block and kinetics?

• Patch-clamp electrophysiology (manual voltage-clamp)
• Radioligand binding assay
• Western blot for hERG protein expression
• ELISA for extracellular hERG fragments

Correct Answer: Patch-clamp electrophysiology (manual voltage-clamp)

Q4. A commonly used heterologous expression system for hERG assays is:

• HEK293 cells stably expressing hERG
• HepG2 hepatocyte cell line
• Primary skeletal muscle myotubes
• Peripheral blood mononuclear cells

Correct Answer: HEK293 cells stably expressing hERG

Q5. In the context of hERG inhibition studies, IC50 refers to:

• The concentration of a drug that produces 50% inhibition of hERG current
• The concentration that activates 50% of hERG channels
• The half-life of channel recovery from inactivation
• The plasma concentration producing 50% of maximal therapeutic effect

Correct Answer: The concentration of a drug that produces 50% inhibition of hERG current

Q6. Regulatory safety margin for hERG is commonly calculated as which ratio?

• hERG IC50 divided by the unbound (free) plasma Cmax
• Therapeutic total Cmax divided by hERG IC50
• hERG IC50 divided by total (bound + unbound) plasma Cmax
• Unbound Cmax divided by IC50 of an unrelated ion channel

Correct Answer: hERG IC50 divided by the unbound (free) plasma Cmax

Q7. The CiPA (Comprehensive in vitro Proarrhythmia Assay) paradigm integrates which key elements?

• Multi-ion channel testing, in silico human ventricular models, human iPSC-derived cardiomyocytes, and ECG biomarker evaluation
• Only high-throughput hERG binding assays and animal telemetry ECG
• Hepatic clearance studies, genotoxicity assays, and hERG Western blots
• Single-channel recordings from native human atrial tissue only

Correct Answer: Multi-ion channel testing, in silico human ventricular models, human iPSC-derived cardiomyocytes, and ECG biomarker evaluation

Q8. Why is assay temperature important when measuring hERG block?

• Temperature affects hERG gating kinetics and drug binding; physiological temperature yields more clinically relevant kinetics
• Temperature only affects cell adhesion, not channel kinetics
• Assays at lower temperature always produce stronger block, which is preferred
• Temperature is irrelevant because hERG is temperature-insensitive

Correct Answer: Temperature affects hERG gating kinetics and drug binding; physiological temperature yields more clinically relevant kinetics

Q9. Which cardiac current is directly mediated by the hERG channel?

• IKr (rapid delayed rectifier potassium current)
• IKs (slow delayed rectifier potassium current)
• INa (fast sodium current)
• ICa-L (L-type calcium current)

Correct Answer: IKr (rapid delayed rectifier potassium current)

Q10. Regarding binding kinetics, which kinetic profile is generally associated with lower proarrhythmic risk for hERG blockers?

• Rapid association and rapid dissociation (fast on/off binding)
• Extremely tight, irreversible binding
• Very slow association with long residence time
• Binding only during diastole with no dissociation

Correct Answer: Rapid association and rapid dissociation (fast on/off binding)

Q11. A common source of false positives in fluorescence-based thallium flux hERG assays is:

• Compound intrinsic fluorescence or fluorescence quenching interfering with the readout
• Excessive ATP in the extracellular buffer
• Overexpression of IKs channels in the assay cell line
• Contamination with bacterial endotoxin

Correct Answer: Compound intrinsic fluorescence or fluorescence quenching interfering with the readout

Q12. The cellular mechanism most often underlying Torsades de Pointes in the setting of hERG block is:

• Early afterdepolarizations (EADs) due to prolonged action potential duration
• Delayed afterdepolarizations (DADs) from calcium overload only
• Pure conduction block in the His-Purkinje system
• Ischemic necrosis of ventricular myocytes

Correct Answer: Early afterdepolarizations (EADs) due to prolonged action potential duration

Q13. Compared with manual patch-clamp, automated patch-clamp platforms primarily offer:

• Much higher throughput suitable for screening multiple compounds and concentrations
• Substantially higher voltage resolution and better single-channel recordings
• Lower startup cost and no requirement for cell lines
• Guaranteed elimination of assay artefacts

Correct Answer: Much higher throughput suitable for screening multiple compounds and concentrations

Q14. A drug that impairs hERG trafficking typically causes cardiac risk by:

• Reducing surface expression of functional hERG channels, decreasing IKr and prolonging repolarization
• Increasing hERG mRNA translation leading to excessive IKr
• Enhancing trafficking of hERG to the membrane and shortening QT interval
• Causing immediate pore block without affecting channel number

Correct Answer: Reducing surface expression of functional hERG channels, decreasing IKr and prolonging repolarization

Q15. Why must major metabolites be evaluated for hERG activity during safety assessment?

• Metabolites can have equal or greater hERG affinity than the parent and contribute to clinical risk
• Metabolites are always inactive at ion channels and can be ignored
• Metabolites only affect hepatic clearance and not cardiac electrophysiology
• Regulatory guidelines prohibit testing metabolites

Correct Answer: Metabolites can have equal or greater hERG affinity than the parent and contribute to clinical risk

Q16. Block of which additional cardiac current can mitigate the proarrhythmic effect of hERG (IKr) inhibition?

• Late sodium current (INa-L) block can reduce action potential prolongation and mitigate risk
• Block of inward rectifier K+ (IK1) will counteract IKr block safely
• Block of fast sodium current (INa) at peak phase always increases proarrhythmia
• Block of acetylcholine-activated K+ current (IKACh) is the preferred mitigation strategy

Correct Answer: Late sodium current (INa-L) block can reduce action potential prolongation and mitigate risk

Q17. Which in silico ventricular action potential model is widely used in CiPA-style proarrhythmia prediction?

• O’Hara-Rudy human ventricular action potential model
• Hodgkin-Huxley squid axon model
• FitzHugh-Nagumo simplification for neurons
• Goldman-Hodgkin-Katz transporter model

Correct Answer: O’Hara-Rudy human ventricular action potential model

Q18. Reverse use-dependence of an hERG-blocking drug means:

• The degree of block is greater at slower heart rates (low pacing frequency)
• The drug blocks more at high heart rates than at low rates
• Block is independent of heart rate
• Block only occurs after multiple drug exposures (use-dependent potentiation)

Correct Answer: The degree of block is greater at slower heart rates (low pacing frequency)

Q19. In addition to QTc, which ECG-derived biomarker has been emphasized under CiPA to help discriminate proarrhythmic risk?

• J-Tpeak (JTp) interval corrected for heart rate
• PR interval prolongation
• QRS widening alone
• ST-segment amplitude increase

Correct Answer: J-Tpeak (JTp) interval corrected for heart rate

Q20. Per ICH and common cardiac safety practice, up to what multiple of unbound clinical Cmax is commonly tested in early hERG screens to define risk?

• At least 30-fold the unbound plasma Cmax (or up to solubility limits) in concentration-response testing
• Only at the therapeutic Cmax, no higher concentrations
• Exactly 2-fold the total plasma Cmax regardless of binding
• Up to the IC10 of unrelated ion channels

Correct Answer: At least 30-fold the unbound plasma Cmax (or up to solubility limits) in concentration-response testing

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