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The ICH S7B document, which provides guidance for the preclinical cardiovascular evaluation of pharmaceutical new chemical entities (NCE), is essentially focused on drug-induced QT lengthening, a biomarker for the proarrhythmic adverse drug reaction, torsades de pointes (TdP). In 2005, this guidance recommended the IKr assay and the in vivo QT telemetry study as mandatory assays for detecting potential torsades de pointes liability and relegated the cardiac action potential (AP) assay as a follow-up study. The IKr assay has become a mandatory screening tool in the early development and safety assessment process. Using only the IKr assay as a go/no go decision arbiter is regrettable since, due to the low specificity of the model (positives that are false for proarrhythmia liability, e.g. verapamil), promising, safe NCEs may be inadvertently discarded. Inclusion of additional medium throughput assays should be performed early to confirm or balance the putatively unfavourable IKr result with positive discovery model output (Pugsley et al., J Pharmacol Toxicol Methods 60:24–27, 2009). In the present chapter, the predictive value of in vitro guinea pig papillary muscle action potential assay will be discussed in terms of sensitivity and specificity and compared to currently available preclinical models such as IKr/hERG assay, dog Purkinje fibre action potential and in vivo QT measurements in dog and cynomolgus monkey.

Cardiac safety pharmacology is a continuously evolving discipline that uses the basic principles of pharmacology in a regulatory-driven process to generate data to inform risk/benefit assessment of a new chemical entity (NCE). The aim of cardiac safety pharmacology is to characterise the pharmacodynamic/pharmacokinetic (PK/PD) relationship of a drug’s adverse effects on the heart using continuously evolving methodology. Unlike Toxicology, safety pharmacology includes within its remit a regulatory requirement to predict the risk of rare cardiotoxic (potentially lethal) events such as torsades de pointes (TdP), which is statistically associated with drug-induced changes in the QT interval of the ECG due to blockade of I Kr or K v11.1 current encoded by hERG. This gives safety pharmacology its unique character. The key issues for the safety pharmacology assessment of a drug on the heart are detection of an adverse effect liability, projection of the data into safety margin calculation and clinical safety monitoring. This chapter will briefly review the current cardiac safety pharmacology paradigm outlined in the ICH S7A and ICH S7B guidance documents and the non-clinical models and methods used in the evaluation of new chemical entities in order to define the integrated risk assessment for submission to regulatory authorities. An overview of how the present cardiac paradigm was developed will be discussed, explaining how it was based upon marketing authorisation withdrawal of many non-cardiovascular compounds due to unanticipated proarrhythmic effects. The role of related biomarkers (of cardiac repolarisation, e.g. prolongation of the QT interval of the ECG) will be considered. We will also provide an overview of the ‘non-hERG-centric’ concepts utilised in the evolving comprehensive in vitro proarrhythmia assay (CIPA) that details conduct of the proposed ion channel battery test, use of human stem cells and application of in silico models to early cardiac safety assessment. The summary of our current understanding of the triggers of TdP will include the interplay between action potential (AP) prolongation, early and delayed afterdepolarisation and substrates for re-entry arrhythmias.

A number of marketed drugs have been withdrawn and numerous clinical and preclinical compounds discontinued due to prolongation of the time interval between Q and T waves of the electrocardiogram (long QT syndrome). Drug-induced QT interval prolongation, which may lead to life-threatening cardiac arrhythmia torsades de pointes (TdP), has been linked to blockade of a cardiac potassium channel, a product of the human ether-a-go-go-related gene (hERG). Consequently, drug discovery efforts across the pharmaceutical industry have been utilizing hERG in vitro assays to predict and minimize the risk of QT prolongation. This chapter discusses in silico, in vitro, ex vivo, and in vivo methods employed to measure blockade of hERG and QT prolongation, as well as regulatory recommendations, and medicinal chemistry strategies utilized to circumvent interactions with hERG.

Potassium (K+) channels play a central role in the electrical activity of excitable cells. Although there are variety of potassium channels, scientists have developed immense interest in human ether-a-go-go-related gene (hERG) potassium channels due to their involvement in life-threatening cardiac arrhythmia. hERG is a gene that encodes the pore-forming α-subunit of a voltage-gated potassium channel expressed in nervous and cardiac tissue including atrium, ventricles, purkinje fiber, SA node and AV node. Potassium flow through hERG channel plays an important role in action potential repolarization, particularly in ventricular muscle. Blockade of hERG potassium channel via pharmacological interventions or hereditary mutations of genes encoding the channel is associated with a prolongation of cardiac ventricular repolarization, that is long QT syndrome (LQTS), a disorder that predisposesindividuals to life-threatening arrhythmias and substantial risk of sudden death. Inherited or drug-induced mutations in hERG channel lead to disruption of delayed rectifier potassium current (IKr), increase in cardiac excitability subsequently torsades de pointes and sudden death. A large number of putative disease-causing mutations in hERG have been identified in affected families so far, yet mechanism behind these mutations is unspecified and undistinguished. Therefore, entire paradigm of drug discovery has shifted towards the safety of the new molecules to screen for potential cardiac arrhythmogenic effects. Non-clinical assays are not sensitive enough to accurately predict QT prolongation liabilities in humans. For this reason, International Conference on Harmonization (ICH) safety pharmacology S7B guidelines were proposed for new chemical entities. According to these guidelines, thorough studies (in vitro and in vivo) on QT are required for virtually all newly developed pharmaceutical agents. In this article, an overview on hERG channels, their functions and dysfunctions, therapeutic agents modulating these channels and associated QT prolongation, and assay have been discussed.

An effective approach to detect drug candidates that may exert unwanted effects on heart rate and rhythm is critical when characterizing the cardiovascular safety profile of novel drugs. As all cardiac channels represent potential off-target liabilities, numerous approaches have been developed to detect undesirable cardiac electrophysiologic effects during drug discovery. This chapter will describe strategies to assess potential proarrhythmic liabilities early in drug discovery using lessons learned from experiences with drugs that affect the hERG channel and cause acquired long QT syndrome.

A prolonged QT interval and increased risk of arrhythmia is a dangerous side effect of several antiarrhythmic agents and noncardiac medications. In most clinically relevant cases, drug-induced QT prolongation is caused by a block of the cardiac rapid delayed rectifier K+ current (I Kr), a current that is conducted by hERG1 channels and is a key determinant of phase 3 repolarization of ventricular cardiomyocytes. Some drugs do not block I Kr, but instead disrupt the trafficking of channels to the cell membrane. Because hERG1 channel block is a serious side effect that is associated with structurally diverse drugs, it is now common practice to screen for hERG1 channel block early in the drug development process. This screening process has also discovered compounds that activate hERG channels – these compounds could theoretically be used to treat long QT syndromes. The molecular determinants of hERG1 blockers and activators have been elucidated and these studies are the focus of this chapter.

Prolongation of ventricular repolarization, reflected by QT prolongation in the ECG, was introduced as a major antiarrhythmic mechanism, but soon became a surrogate for Torsade de Pointes (TdP) instead. However, drugs that prolong the QT interval range from potently torsadogenic activity to no proarrhythmic action and even antiarrhythmic effects. While block of hERG channels is commonly involved, this is neither complete nor correct: block and activation of other channels can also be torsadogenic, while block of hERG need not be torsadogenic per se. TdP is frequently caused by disturbances of repolarization, but it can also result from disturbances of conduction, even in the absence of prolongation of the action potential duration (APD). Most importantly, disturbances of λ-TRIaD are not only torsadogenic, as they can also lead to VT and VF [λ = CV × ERP; CV: conduction velocity; ERP: effective refractory period; T: triangulation; R: Reverse use dependence; I: instability and D: dispersion]. Thus, comprehensive proarrhythmic evaluation cannot focus only on QT prolongation; this will yield false positives (potentially depriving patients from much needed drugs) as well as false negatives (exposing patients to potentially lethal arrhythmias).

Structural and Functional Discrimination of Membrane.- Proteins.- Pharmacology of Hyperpolarization-Activated Cyclic Nucleotide-Gated (HCN) Channels.- Advanced Molecular Modeling Techniques Applied to Ion Channels Blockers.- Advances in Design and Development of Sodium Channel Blockers.- Potassium Channel Blockers as Anti-arrhythmic Agents.- hERG Potassium Channels in Drug Discovery and Development.- Prediction of hERG Channel Inhibition Using in silico Techniques.- Advances in Structure-Activity Relationship Studies on Potassium Channel Modulators.- Calcium Ion Channels and Their Blockers.- Therapeutic Potential of N-type Voltage Gated Ca2+ Channel.- Chloride Ion Channels: Structure, Functions, and Blockers.

Drug-induced long QT syndrome still represents a major risk for late-stage clinical failure of drug candidates. One of the main factors inducing long QT is inhibition of cardiac hERG channels. Early prediction of potential hERG inhibitory activity of hits and lead compounds thus is of major importance in the drug discovery and development process. Both structure- and ligand-based approaches were used to develop models, which shed light on the molecular basis of hERG channel inhibition. In this article, an overview on recent approaches for prediction of hERG channel blockers has been provided.

The myocardium is the target of toxicity for a number of drugs. Based on pharmacological evidence, cellular targets for drugs that produce adverse reactions can be categorized into a number of sites that include the cell membrane-bound receptors, the second messenger system, ionic channels, ionic pumps, and intracellular organelles. Additionally, interference with the neuronal input to the heart can also present a global site where adverse drug effects can manifest themselves. Simply, a drug can interfere with the normal cardiac action by modifying an ion channel function at the plasma membrane level leading to abnormal repolarization and/or depolarization of the heart cells thus precipitating a disruption in the rhythm and causing dysfunction in contractions and/or relaxations of myocytes. It is now recognized that toxic actions of drugs against the myocardium are not exclusive to the antitumor or the so-called cardiac drugs, and many other drugs with diverse chemical structures, such as antimicrobial, antimalarial, antihistamines, psychiatric, and gastrointestinal medications, seem to be capable of severely compromising myocardium function. At present, great emphasis in terms of drug safety is being placed on the interaction of many classes of drugs with the hERG potassium channel in cardiac tissue. The interest in the latter channel stems from the simplified view that drugs that block the hERG potassium channel cause prolongation of the QT interval, and this can cause life-threatening cardiac arrhythmias. Based on the evidence in the current literature, this concept does not seem to always hold true.

The great diversity of K+ channels and their wide distribution in many tissues are associated with important functions in cardiac and neuronal excitability that are now better understood thanks to the discovery of animal toxins. During the past few decades, sea anemones have provided a variety of toxins acting on voltage-sensitive sodium and, more recently, potassium channels. Currently there are three major structural groups of sea anemone K+ channel (SAK) toxins that have been characterized. Radioligand binding and electrophysiological experiments revealed that each group contains peptides displaying selective activities for different subfamilies of K+ channels. Short (35–37 amino acids) peptides in the group I display pore blocking effects on Kv1 channels. Molecular interactions of SAK-I toxins, important for activity and binding on Kv1 channels, implicate a spot of three conserved amino acid residues (Ser, Lys, Tyr) surrounded by other less conserved residues. Long (58–59 amino acids) SAK-II peptides display both enzymatic and K+ channel inhibitory activities. Medium size (42–43 amino acid) SAK-III peptides are gating modifiers which interact either with cardiac HERG or Kv3 channels by altering their voltage-dependent properties. SAK-III toxins bind to the S3C region in the outer vestibule of Kv channels. Sea anemones have proven to be a rich source of pharmacological tools, and some of the SAK toxins are now useful drugs for the diagnosis and treatment of autoimmune diseases.

Electrophysiological modeling of the heart enable quantitative description of electrical processes during normal and abnormal excitation. Cell models describe e.g. the properties of the cell membrane and the gating process of ionic channels. New measurement data is available for these channels for physiological and some pathological states. These data should be included in the models to enhance their features. In this work we describe a framework adapting ion channel models to measurement data by using a particle swarm optimization (PSO). As an example the data of whole-cell voltage-clamp measurements of the reggae mutation in hERG was integrated into a Markovian chain model describing I Kr.

The mutations to hERG (the human Ether-a-go-go Related Gene) that cause long QT syndromes produce effects on the rapid delayed rectifier K + current I Kr and, therefore, action potential duration (APD). These mutations can affect various properties that determine I Kr kinetics. We used computational models of human ventricular myocytes to identify which of these properties, when altered, cause profound changes to APD and transmural dispersion of repolarisation (TDR). Such increases in both APD and TDR is caused by a positive shift of activation V 0.5 , a negative shift of inactivation V 0.5 , or by reducing maximal conductance. The largest reduction in APD is achieved by a positive shift of inactivation V 0.5 . Altering the time constant of activation had relatively little effect. When two or more parameters were altered simultaneously, shifting inactivation V 0.5 had the dominant effect on APD, except for some extreme shifts of activation V 0.5 or moderate reductions of maximal conductance. HERG mutations observed clinically lie in the parameter range where maximal conductance has the dominant effect. Bifurcation analysis showed stable steady states (corresponding to physiological resting membrane potential) at all parameter values, and no APD alternans. We conclude that increased APD due to hERG mutations seen clinically are a combined effect of alterations to I Kr kinetic parameters that, in isolation, cause either shortening or prolongation of the AP. Therapeutics that alter I Kr conductance are potentially most beneficial.

Atrial fibrillation is the most common cardiac arrhythmia in humans. The precise cellular mechanisms underlying atrial fibrillation are still poorly understood. Recent studies have identified several genetic defects as predisposing factors for this pathology. One of the identified genetic defects is the mutation N588K, which affects the cardiac IKr channel. Genetic variants in this channel have been identified to modify ventricular repolarization. The aim of this work is to investigate the effect of this mutation on atrial repolarization and the predisposition to atrial fibrillation.

Atrial fibrillation (AF) is the most common cardiac arrhythmia in the western world. Genetic variants in the cardiac I Kr channel have been identified to influence ventricular repolarization. The aim of this work is to investigate the effect of the mutation N588K on atrial repolarization and the predisposition to AF. Experimental data of N588K mutated hERG channel were incorporated in an atrial ionic model using parameter fitting. The effects of the mutation were analyzed in cell and tissue. N588K showed a gain of function effect, causing a rapid repolarization and a shortening of the action potential duration. Computer simulations of a schematic right atrial geometry were used to investigate the excitation conduction properties. The effective refractory period of mutant cells were reduced from 317 to 233 ms at 1 Hz . The conduction velocity is not significantly influenced by the mutation. Nevertheless, the wavelength of mutant cells is for all frequencies smaller, indicating that the mutation N588K predisposes the initiation and perpetuation of AF.

With KvLQT1, the first partner of the IsK protein in the IKs channel complex has been identified following years of intensive searching. Several models have been proposed to explain the constitution of the functional IKs channel. A recent study demonstrated a direct interaction of the C-terminal end of the IsK protein with the pore region of KvLQT1 (Romey et al. 1997). These data support the assumption that the IsK protein participates in IKs channel gating and pore formation. Depending on the tissue of its expression, activation of IKs exerts distinct net effects. In the inner ear it represents a secretory pathway for K+, in the heart IKs participates in the repolarizing conductances of cardiocytes and in epithelial cells it may provide the driving force for Cl− secretion. Whether the IsK protein interacts with KvLQT1 in all tissues or with a homologous subunit has not been clarified. The relative significance of IKs can be strongly regulated by a number of second messengers. Particularly in heart, where IKs becomes the dominant repolarizing conductance at high frequency and β-adrenergic tonus, an inhibition of IKs may be beneficial in the treatment of certain arrhythmias. On the other hand, an increase of IKs with specific activators may be of therapeutic interest for patients with inherited long QT syndromes of the types LQT2 and LQT3. As for LQT2 and LQT3 syndromes, for which defects in HERG and SCN5A channels are responsible, a specific increase in IKs could possibly reduce the QT interval in these patients.