Calcium channel
Calcium channel

Calcium channel assays (CaV1.2) for preclinical drug studies

The voltage gated cardiac calcium ion channel Cav1.2, encoded by the CACNA1C gene, is a key component of the shape of the plateau phase and is highly implicated in the repolarization process of the action potential. When the Cav1.2 calcium channel cannot adequately conduct electrical current across the cell membrane, this may result in undesirable cardiovascular side effects leading to life threatening arrythmias called Torsades de Pointe.
Mutation LQT8: the gain-of-function mutation in the CACNA1C gene may result in a markedly prolonged action potential duration causing a rare congenital arrhythmia: the long QT syndrome LQT8 (Timothy syndrome).

CaV1.2 assay & preclinical studies

At a preclinical stage, Cav1.2 calcium channel assays aim to identify potential inhibitory effects early in the drug discovery process. A number of clinically successful drugs have had the tendency to inhibit the Cav1.2 calcium channel, entailing a concomitant risk of sudden death. As a result, a large number of drugs have been withdrawn from the market or during late stage clinical trials due to cardiotoxic effects.

What is shown?

A pharmacological reduction of the calcium current has been shown to shorten the repolarization process and decrease the contractility. This makes the human Cav1.2 calcium channel a major target for the cardiac safety assessment.

Technique

  • Conventional manual Patch Clamp
  • 3 or 4 increasing concentrations of assay compound tested independently

Study model

  • Human Embryonic Kidney HEK-293 cells
  • 5 treated cells per concentration tested and 5 control cells (as basic design protocol but can be adapted)

Measured parameters

  • Amplitude of the peak current upon depolarization to 0 mV (pA)
  • Amplitude of the base current at –80 mV (pA)
  • Inhibition of Cav1.2 peak current amplitude (%)
  • ion current inhibition measurement
  • IC50 value

Stimulation protocol

Cav 12 assay

Typical effects of verapamil on Cav1.2 calcium currents recorded from CHO cells.

Ref­er­ence com­pounds or pos­i­tive con­trols

Reference CompoundsIC₅₀

Ver­a­pamil3.4 µM

Nifedip­ine2.8 nM

Dil­ti­azem7.7 µM

Additional tests according to PhysioStim scientist’s expertise

Humans cardiomyocites

Human cardiomyocytes

PhysioStim Isolated Perfused Heart

Isolated perfused heart

Purkinje fiber assays

Isolated organ bath

Isolated organ baths

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