Cardiovascular Safety In Vitro Models
Many drugs have either been withdrawn from the market or placed on restricted use due to their (or their metabolites) undesirable ‘off-target’ effects on the heart. An increase in the cardiac action potential duration (APD), caused by a drug blocking the hERG (human ether a go-go) ion channel, can be defined by a prolongation of the QT interval as measured by body surface electrocardiogram (ECG) and may deteriorate into Torsades de Pointes and ventricular fibrillation.
Indeed, cisapride, a gastric prokinetic drug approved in the USA in 1993, had severe limitation placed on its use in 2000 after the FDA reported 341 incidences of arrhythmias, of which 80 resulted in a fatal outcome (Shah, 2002).
The ‘Holy Grail’ of cardiovascular safety in vitro models is to accurately predict the translational effect of an observation or measurement to an in vivo or clinical situation. At best, in vitro assays offer a highly predictive correlation and are usually one component of an integrated approach to cardiovascular safety assessment. Using a predictive in vitro model not only serves to reduce and eliminate the risk of adverse cardiac effects in man, but ethically reduces the number of animals used during development. Financial savings will be made as drug candidates are deselected earlier in the discovery cycle.
The cardiac action potential (AP) is comprised of a complex concert of ionic events across several classes of ion channels and associated sub-units (K+, Na+, Cl- and Ca2+). Currently it is impossible to screen against every ion channel and accessory sub-unit involved in the maintenance of cardiac rhythm, but several of the main candidates involved in the cardiac AP are commercially available.
These recombinant ion channels are used in electrophysiology recordings to measure the flow of ions across the membrane in response to exposure to a drug of interest. Any inhibition (or enhancement) to this flow can then be used to define a ‘safety margin’ and predict potential adverse cardiac effects in vivo based on the therapeutically relevant dose.
Many instruments are available to record electrophysiological changes from these cardiac cell lines. These can be screened using high-throughput electrophysiology instruments. The IonWorks Barracuda® records up to a 6000 data points per hour on a 384-well plate. PatchXpress®, a 16-well chip, offers lower throughput but with higher electrophysiology integrity. The traditional manual patch clamp offers the ‘gold standard’ for electrophysiology recording, but at the cost of a comparatively low throughput.
The recombinant hERG channel GLP assay is a regulatory requirement and must accompany any investigational new drug (IND) application. At Charles River, performing manual whole-cell patch clamp technique under GLP conditions, provides precise functional information on the in vitro hERG liability of novel drugs and, as a surrogate marker for QT prolongation, can be used to determine safety margins before testing in vivo or in the clinic. This assay is offered as part of the integrated safety pharmacology program at Charles River.
- Shah, R.R. 2002. The significance of QT interval in drug development. Br J Clin Pharmacol. 54, 188–202.