hKV4.3 assays

Preclinical cardiac safety

hKV4.3

Since the Kv4.3 channel encoding for Ito current, plays an important role in the early stage of cardiac action potential repolarisation, a pharmacological reduction of the Ito current may delay the repolarisation process and may lead to QT interval prolongation. Thus the Kv4.3 potassium channel represent a specific target for the cardiac safety assessment.

hKV4.3 assays & Preclinical safety studies

At the cellular level,  gain of the transient outward depolarizing current (Ito) conducted by the KCND3-encoded Kv4.3 α-subunit is hypothesized to result in an outward shift in current, heterogenous loss of the action potential dome, ST segment elevation on ECG, local re-excitation via phase II reentry, and the initiation of polymorphic ventricular tachycardia or ventricular fibrillation.

hKV4.3 assays & Preclinical safety studies

• Threshold concentration of hKV4.3 blockade or activation
• Leading to the determination of the safety range versus efficacious concentration
• You obtain your safety margin

Automated Patch Clamp

Technique

  • Chinese Hamster Ovary CHO cells
  • Experiment conducted at room temperature or at physiological temperature (35°C)
  • Up to 6 increasing concentrations of test compound or biologic.

Measured parameters

  • Amplitude of the peak current upon depolarization to +40mV (pA)
  • Amplitude of the base current at –80mV (pA)
  • Ion current amplitude measurement
  • Inhibition of Kv4.3 peak current amplitude (%)
  • IC50 value (at least 4 concentrations required)

Main adventages

  • High throughput screening
  • Cost effective
  • Fast process to delivery results
  • Perfect at earliest stages
  • Design protocol could be adapted

Manual Patch Clamp

Technique

  • Chinese Hamster Ovary CHO cells
  • Experiment conducted at room temperature or at physiological temperature (35°C)
  • Up to 5  increasing concentrations of test compound tested independently
  • At least 3 treated cells are recommended

Measured parameters

  • Amplitude of the peak current upon depolarization to +40mV (pA)
  • Amplitude of the base current at –80mV (pA)
  • Ion current amplitude measurement
  • Inhibition of Kv4.3 peak current amplitude (%)
  • IC50 value (at least 4 concentrations required)

Main adventages

  • Technically robust
  • Highly informative (more accurate IC50 value)
  • Strongly replicable
  • Design protocol could be adapted

Stimulation protocol

Potassium assays

Typical effects of flecainide effects on Ito current recorded from CHO cells.

Ref­er­ence com­pounds

Reference CompoundsIC₅₀

Flecainide19.6 µM

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