PHYSIOLOGY IN MEDICINE In collaboration with The American Physiological Society, Thomas E. Andreoli, MD, Editor

Molecular Biology of Kⴙ Channels and Their Role in Cardiac Arrhythmias Martin Tristani-Firouzi, MD, Jun Chen, MD, John S. Mitcheson, PhD, Michael C. Sanguinetti, PhD The configuration of cardiac action potentials varies considerably from one region of the heart to another. These differences are caused by differential cellular expression of several types of K⫹ channel genes. The channels encoded by these genes can be grouped into several classes depending on the stimulus that permits the channels to open and conduct potassium ions. K⫹ channels are activated by changes in transmembrane voltage or binding of ligands. Voltage-gated channels are normally the most important players in determining the shape and duration of action potentials and include the delayed rectifiers and the transient outward potassium channels. Ligand-gated channels include those that probably have only minor roles in shaping repolarization under normal conditions but, when activated by extracellular acetylcholine or a decrease in the intracellular concentration of ATP, can substantially shorten action potential

duration. Inward rectifier K⫹ channels are unique in that they are basically stuck in the open state but can be blocked in a voltage-dependent manner by intracellular Mg2⫹, Ca2⫹, and polyamines. Other K⫹ channels have been described that provide a small background leak conductance. Many of these cardiac K⫹ channels have been cloned in the past decade, permitting detailed studies of the molecular basis of their function and facilitating the discovery of the molecular basis of several forms of congenital arrhythmias. Drugs that block cardiac K⫹ channels and prolong action potential duration have been developed as antiarrhythmic agents. However, many of these same drugs, as well as other common medications that are structurally unrelated, can also cause long QT syndrome and induce ventricular arrhythmia. Am J Med. 2001;110:50 –59. 䉷2001 by Excerpta Medica, Inc.

PHYSIOLOGICAL ROLES AND MOLECULAR BASIS OF CARDIAC VOLTAGE-GATED Kⴙ CHANNELS

current (IKr) and the inward rectifier K⫹ current (IK1). These currents were discovered and characterized using voltage clamp techniques, which permit precise control of the membrane potential and measurement of ion currents under carefully controlled ionic conditions. Although voltage-clamp studies are useful in defining the properties of these currents, their molecular identities had to await the advancement of molecular cloning techniques. Starting with the cloning of the Shaker K⫹ channel from Drosophila in 1987 (1), many K⫹ channels have been cloned. These studies have shown that K⫹ channels are formed by coassembly of four subunits (2) and are sometimes associated with auxiliary beta subunits that can modify the gating properties of the heteromultimeric channel complex. With only a few exceptions, the molecular basis of cardiac K⫹ currents have been defined (Table 1). These channels can be grouped in many ways, most logically by amino acid sequence homology (3). Another classification scheme is based on biophysical characteristics of the currents determined by voltage clamp experiments. Based on function, cardiac K⫹ channels can be placed into one of four categories: transient outward, delayed rectifier, inward rectifier, and leak channels. Several excellent reviews of K⫹ channel diversity based on sequence and function are recommended for the reader

T

he initial upstroke of the cardiac action potential is determined by the opening and closing of Na⫹ channels. The configuration and rate of repolarization of action potentials are controlled by many types of K⫹ channel currents that differ with respect to their kinetics and density in the plasma membrane (Figure 1). Initial repolarization (phase 1) is mediated by the opening of transient outward K⫹ channels. This is followed by a plateau (phase 2) that is characterized by high membrane resistance resulting from the almost equal flow of outward currents through delayed rectifier K⫹ channels (IKr, IKs, IKur) and inward flow of current through L-type Ca2⫹ channels. The rate of terminal repolarization (phase 3) is enhanced after the plateau phase because of the increasing conductance of the rapid delayed rectifier K⫹

Am J Med. 2001;110:50 –59. From the Department of Medicine, Division of Cardiology, University of Utah, Salt Lake City, Utah. Requests for reprints should be addressed to Michael C. Sanguinetti, Department of Medicine, Division of Cardiology, University of Utah, 15 N 2030 E Room 4220, Salt Lake City, Utah 84112. 50

䉷2001 by Excerpta Medica, Inc. All rights reserved.

0002-9343/01/$–see front matter PII S0002-9343(00)00623-9

K⫹ Channels and Cardiac Arrhythmias/Tristani-Firouzi et al

Table 1. The Molecular Identity of Human Cardiac K⫹ Currents Cloned Channel Current Transient outward Ito

Alpha Subunit

Beta Subunit

Kv4.3 Kv1.4

Delayed rectifier IKur IKr IKs Inward rectifier IK1 IKATP IKACh Leak

Kv1.5 HERG KVLQT1

MiRP1 minK

Kir2 Kir6.2 Kir3.1

SUR1 Kir3.4 TWIK

Figure 1. K⫹ currents responsible for repolarization of a typical ventricular action potential. Ventricular action potential (top). Phase 0, rapid upstroke; phase 1, initial repolarization; phase 2, plateau; phase 3, terminal repolarization; phase 4, diastolic membrane potential. The rapid repolarization of phase 1 is the result of the contribution of the rapidly activating transient outward (Ito), the ultra-rapid delayed rectifier (IKur), and the leak (Ileak) currents (middle and bottom). During the plateau phase, the rapid (IKr) and slow (IKs) delayed rectifier K⫹ currents as well as IKur and Ileak counter the depolarizing influence of Ltype calcium current (not shown). IKr and the inward rectifier K⫹ current (IK1) provide repolarizing current during the terminal phase of the action potential [modified from (71). Reprinted with permission from the American Heart Association].

interested in a more comprehensive view of this subject (4 – 6).

Transient Outward K⫹ Channels

Transient outward K⫹ current (Ito) activates very rapidly in response to a rapid depolarization, such as occurs during the upstroke of the action potential. Soon after opening, these channels close, resulting in the transient nature of the net outward current (Figure 1). Ito is the sum of a Ca2⫹-dependent Cl⫺ current and a voltage-dependent K⫹ current. The K⫹ current component of Ito is conducted by channels formed by tetrameric assembly of Kv1.4, Kv4.2, and/or Kv4.3 subunits (6). A single functional channel forms in the endoplasmic reticulum by coassembly of four identical subunits. In human atrial myocytes, the Ito channel is formed by coassembly of four Kv4.3 protein subunits (7). Like other voltage-gated K⫹

channels, each subunit has six transmembrane domains (S1 to S6), including one domain (S4) that senses transmembrane voltage (Figure 2). The amino and carboxyl termini are located on the intracellular side of the membrane. Movement of the S4 domain in response to membrane depolarization is coupled to other regions of the protein that form the activation gate (8,9). When the activation gate is open, the channel conducts K⫹ in a direction that depends on the electrochemical gradient across the plasma membrane. Immediately after depolarization, repetitive opening (activation) and closing (deactivation) of the activation gate determines how long the channel is in a conducting state. However, soon (within 10 to 100s of msecs) a portion of the amino terminus binds to a specific site near the inside of the pore region and closes the channel, a process called inactivation. Unlike the deactivated state, the inactivated state is a long-lived closed state. The channel remains closed until the membrane is repolarized to the resting potential, where channels recover from the inactivated state and again become capable of opening in response to membrane depolarization.

Delayed Rectifier K⫹ Channels

The delayed rectifier K⫹ current, IK, is comprised of at least three distinct currents, IKur, IKr, and IKs that can be distinguished on the basis of kinetics of activation and pharmacologic properties (10 –15). IKur activates ultrarapidly (16), IKr activates rapidly, and IKs activates very slowly. IKur is blocked by 4-aminopyridine (17,18), IKr is blocked by several antiarrhythmic agents (eg, dofetilide) (19,20), and IKs is blocked by a compound called chromanol 293B (21). The amplitude of the delayed rectifier K⫹ currents varies during repolarization of the action potential because of changes in membrane potential, chemical driving January 2001

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Figure 2. Proposed topology of K⫹ channel subunits. A. Schematic representation of a voltage-gated K⫹ channel alpha subunit composed of six membrane-spanning alpha helices (S1 to S6). The fourth membrane-spanning unit (S4) contains positively charged residues at approximately every third position and is the voltage sensor. The residues between S5 and S6 (shown in orange) form the ion selective pore. Auxiliary beta subunits (shown in green) modify the gating properties and protein trafficking of the pore-forming alpha subunits. Kvbeta subunits are cytoplasmic proteins that bind to the N-terminus. MinK, a component of the IKs channel, is a membrane-spanning beta subunit. B. K⫹ channel alpha subunits coassemble to form a tetrameric channel composed of four identical subunits (homotetramer) or nonidentical subunits (heterotetramer). C. Inward rectifier K⫹ channels are formed by subunits containing two membrane-spanning alpha helices, separated by a pore domain. Like the six transmembrane voltage-gating ion channel, four subunits coassemble to form the inward rectifier K⫹ channel. TWIK channels are a unique class of channels formed by subunits containing four membrane-spanning domains and two pore loops.

force, and relative rates of activation and inactivation (Figure 1). IKur activates extremely rapidly and does not inactivate appreciably during the time course of the action potential. The magnitude of this current decreases 52

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during repolarization solely because of a decrease in electrochemical driving force. IKr amplitude increases during repolarization, reaching a peak at approximately ⫺30 mV, then decreases as the membrane potential reaches its

K⫹ Channels and Cardiac Arrhythmias/Tristani-Firouzi et al

resting level. This increase in current occurs in spite of a decrease in electrochemical driving force, because channels recover from inactivation to an open state in a voltage-dependent manner. Fast inactivation of IKr is not mediated by the amino-terminal region (22) but instead results from a mechanism believed to involve a slight constriction of the outer pore region of the channel (9). IKs activates extremely slowly and, therefore, increases in magnitude throughout the plateau phase. Only during phase 3 repolarization does IKs decrease in accordance with a decrease in electrochemical driving force for K⫹. The Kv1.5 gene encodes subunits (613 amino acids) that coassemble to form IKur channels in most species, including myocytes of the human atrium (23). HERG and KCNE2 genes encode subunits (HERG and MiRP1, respectively) that coassemble to form IKr channels (24 –26). KVLQT1 and KCNE1 encode subunits (KvLQT1 and minK, respectively) that coassemble to form IKs channels (27,28) (Table 1). HERG (1,159 amino acids) and KvLQT1 (676 amino acids) are alpha subunits that have an overall structure similar to the Kv1 and Kv4 subunits. MinK (129 amino acids) and MiRP1 (minK-related peptide number 1, 123 amino acids) are beta subunits that have a single transmembrane domain. It is presently unclear what regions of the alpha and beta subunits interact to stabilize the heteromultimeric complex, but it is clear that this association alters the gating of the tetrameric alpha subunit channel. The change in kinetics is especially remarkable for KvLQT1, where association with minK greatly slows the rate of activation and shifts the voltage dependence of the channel opening to a more positive membrane potential (27,28). HERG-MiRP1 channels have properties more similar to IKr in native myocytes than do channels formed by coassembly of HERG alone. MiRP1 decreases the single channel conductance and accelerates the rate of HERG channel deactivation (26). Alternative splicing of HERG (29 –31) produces a variant with a shortened amino terminus that also deactivates faster than full length HERG. Additional delayed rectifier K⫹ channels are expressed in the hearts of mammals other than humans. For example, Kv2.1 subunits coassemble to form a delayed rectifier K⫹ channel (IK,slow) in the mouse heart (32).

Inward Rectifier K⫹ Channels

In most cardiac cells, the inward rectifier K⫹ current, IK1, largely determines the resting membrane potential. The channels that conduct this current are open at all voltages. However, K⫹ is preferentially allowed to conduct in the inward direction (from the extracellular space to the cytosol). The conductance of outward current becomes progressively less as the membrane potential is made more positive than approximately ⫺95 mV, the equilibrium potential for K⫹ (Figure 1). A voltage-dependent block by cytosolic Mg2⫹ and polyamines of the inner

channel pore causes low outward conductance (33–35). The pacemaker cells of the sinoatrial and atrioventricular node do not express these inward rectifier K⫹ channels, and therefore the maximum diastolic potential is more depolarized than nonpacing cells of the atria or ventricles. Atrial pacemaker cells have another type of inward rectifier K⫹ current (IKACh) that is activated by binding of acetylcholine to m2 muscarinic receptors located on the surface of the cell membrane. M2 receptor binding activates a G protein that in turn increases the probability of K⫹ channel opening (36). Activation of IKACh slows the spontaneous firing rate of pacemaker cells and shortens action potential duration. Most cardiac myocytes also express an inward rectifier K⫹ channel that is inhibited by cytosolic ATP (37). The current conducted by these channels (IKATP) is activated under conditions of metabolic stress that reduce intracellular ATP and can lead to pronounced shortening of the action potential (38). Coassembly of Kir2.1, Kir2.2, Kir2.3, or Kir2.4 subunits form channels that underlie IK1 (5). These subunits are smaller than the voltage-gated transient outward or delayed rectifier K⫹ channel subunits. For example, human Kir2.1 subunits are composed of 427 amino acids. The proposed membrane topology of Kir2 channels is similar to all the other inward rectifier K⫹ channels, having two putative transmembrane domains linked by a single pore loop (39) (Figure 2). Analysis of the conduction properties of channels constructed by tandem multimers consisting of three or four Kir2.1 subunits suggest that subunits coassemble to form a tetrameric channel complex similar to the voltage-gated K⫹ channels. It is presently unclear if the channels form as homo- or heteromultimeric complexes. However, based on conductance measurements of single channel currents, there is ample evidence that multiple types of inward rectifier K⫹ channels are present in myocytes. KAch channels are formed by coassembly of two Kir3.1 and two 3.4 subunits into a tetrameric complex (40). Kir3.1 and 3.4 subunits have 501 and 419 amino acids, respectively. KATP channels are formed by coassembly of four Kir6.1 subunits and four sulfonylurea receptor (SUR) subunits (41). Human Kir6.1 and SUR1 subunits have 424 and 1,581 amino acids, respectively. Kir3 and Kir6 subunits have the same overall structure as Kir2 subunits, having two transmembrane domains flanking a pore region (Figure 2C).

Leak K⫹ Channels

Most cells have a very small background K⫹ conductance that contributes to maintenance of the resting potential and repolarization of the action potential. In the heart this conductance may be the result of a weakly inward rectifying K⫹ channel, TWIK-1 (42) or Kcnk3 (43). The TWIK channel is 336 amino acids in length with four transmembrane domains. This channel has an unusual structure, because it has two pore domains. Thus, TWIK January 2001

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Figure 3. The role of K⫹ channels in mediating phase 3 repolarization of the cardiac action potential. A. Yellow arrows represent K⫹ efflux through the rapid (IKr) and slow (IKs) delayed rectifier K⫹ channels. Outward movement of positively charged K⫹ hyperpolarizes the cell membrane and terminates the action potential. The surface electrocardiogram is depicted below. The QRS corresponds to the rapid upstroke of the action potential. The T wave represents the change in membrane potential associated with repolarization. B. Mutations in the genes encoding subunits of the IKr and IKs channels reduce the amount of repolarizing current available during the terminal phase of the cardiac action potential. Decreased repolarizing current prolongs the action potential, which is reflected on the surface electrocardiogram as prolongation of the QT interval.

is like two Kir channels that are connected in tandem (Figure 2).

LONG QT SYNDROME CAUSED BY MUTATIONS IN GENES ENCODING SUBUNITS OF CARDIAC RAPID AND SLOW DELAYED RECTIFIER Kⴙ CHANNELS Long QT syndrome (LQTS) is a disorder of ventricular repolarization that predisposes affected individuals to cardiac arrhythmias and sudden death. The most common form of LQTS is acquired, caused by medications that block cardiac K⫹ channels, such as certain antiarrhythmic drugs, antihistamines, and antibiotics, and is exacerbated by bradycardia and hypokalemia (44). LQTS can also be inherited as an autosomal dominant (Romano-Ward syndrome) or recessive (Jervell and Lange-Nielsen syndrome) disorder (45). The more severely affected individuals can have intermittent syncope caused by a self-terminating arrhythmia called torsades 54

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de pointes, characterized by a sinusoidal twisting of the QRS axis around the isoelectric line of the electrocardiogram (46). Spatial dispersion of ventricular repolarization and an alteration in the predominance of two ectopic foci have been postulated to be the underlying cause of torsades de pointes (47). Sudden cardiac death can occur if torsades de pointes arrhythmia degenerates into ventricular fibrillation. Mark Keating and colleagues used a positional cloning approach to discover one gene (KVLQT1) and a candidate gene approach to identify three other genes (SCN5A, HERG, KCNE1, KCNE2) that cause LQTS (48 –52). SCN5A encodes the cardiac sodium (INa) channel and so will not be discussed further in this review. As discussed above, the HERG and KCNE2 genes encode subunits that form the IKr channel, and KVLQT1 and KCNE1 genes encode subunits that form the IKs channel. Mutations in any of these K⫹ channel subunits cause a decreased outward K⫹ current during the plateau phase of the cardiac action potential, delayed ventricular repolarization, and an increased QT interval (Figure 3). It is clear that environmental and other genetic factors are likely to contrib-

K⫹ Channels and Cardiac Arrhythmias/Tristani-Firouzi et al

Table 2. Drugs That Prolong QT Interval and/or Cause Torsades de Pointes* Drug Class

Drug (Trade Name)

Class IA antiarrhythmic agents

Quinidine, procainamide (Procan, Procanbid, Pronestyl), disopyramide (Norpace) Flecainide (Tambocor) Amiodarone (Cordarone), sotalol (Betapace), ibutilide (Corvert), dofetilide (Tikosyn) Bepridil (Vascor), nicardipine (Cardene) Astemizole (Hismanol), terfenadine (Seldane), diphenhydramine (Benadryl), clemastine (Tavist) Amitriptyline (Elavil, Endep), desipramine (Norpramin), doxepin (Sinequan, Zonalon), fluoxetine (Prozac), imipramine (Tofranil), venlafaxine (Effexor) Chlorpromazine (Thorazine), haloperidol (Haldol), risoperidone (Risperdal), thioridazine (Mellaril) Erythromycin, clarithromycin (Biaxin), grepafloxacin (Raxar), moxifloxacin (Avelox), sparfloxacin (Zagam), trimethoprimsulfamethoxazole, amantidine, forscarnet (Foscavir), pentamidine (Pentacarinat, Pentam, NebuPent), fluconazole, ketoconazole, itraconazole, miconazole, halofantrine, chloroquine Felbamate (Felbatol), fosphenytoin (Cerebyx) Cisapride (Propulsid), droperidol (Inapsine), naratriptan (Amerge), pimozide (Orap), probucol (Lorelco), indapamide (Lozol), sumatriptan (Imetrex), tacrolimus (Prograf), tamoxifen (Nolvadex), zolmitriptan (Zomig)

Class IC antiarrhythmic agents Class III antiarrhythmic agents Calcium channel antagonists Antihistamines Antidepressants

Antipsychotics Antimicrobials

Anticonvulsants Miscellaneous agents

See website: http://www.dml.georgetown.edu/depts/pharmacology/torsades.html

ute to the pathology of this disorder, because many mutant gene carriers are asymptomatic. The different forms of LQTS are commonly referred to by their original loci assignment. Therefore, mutations in KVLQT1, HERG, KCNE1, and KCNE2 cause LQT1, LQT2, LQT5, and LQT6 forms of LQTS, respectively (53). Mutations of ion channel genes cause channel protein dysfunction by a variety of mechanisms. Most LQTS is the result of a dominant pattern of inheritance, where an offspring carries a single mutant gene from one parent and a normal gene from the other parent. The most common mutations are the result of a single base-pair change (missense mutation) that results in a change in a single amino acid. This type of mutation often causes subunit misfolding, which disrupts the coassembly of subunits and usually leads to early degradation of the channel complex. This “dominant-negative effect” causes a greater than 50% reduction in the number of functional channels. A dominant-negative effect can also occur if the missense mutation results in change in an amino acid with a function vital to the operation of the channel. An example is the G638S missense mutation in HERG, where the glycine (G) at amino acid position 638 is changed to a serine (S) residue. G638 is part of a highly conserved sequence of the channel that forms the K⫹-selective pore. Mutation of this amino acid has no effect on the folding of the channel complex or trafficking of the complete channel to the surface membrane (54), but evidently pre-

vents the channel from permitting the flow of K⫹ through the narrowest region of the pore. Coexpression of normal and G628S HERG subunits indicates that coassembly of even a single G628S subunit in the tetrameric channel results in loss of function, a “lethal” dominantnegative effect (55). Deletion of one or more nucleotides often leads to a premature stop codon and truncation of the resulting protein. If the truncated protein no longer contains a subunit association domain, then the mutant subunit cannot interact with normal subunits. The net effect is a reduction in the number of functional channel proteins by approximately 50%, a condition referred to as haploinsufficiency. In this case, the resulting phenotype would display reduced current amplitude without any change in biophysical properties. Other missense mutations result in subunits that can still coassemble with normal subunits but alter one or more properties of the channel. For example, specific mutations in HERG have been shown to shift the voltage dependence of gating associated with either activation or inactivation (56). Either a shift in the voltage dependence of inactivation to a more negative potential or a shift in the voltage dependence of activation to a more positive potential causes a reduction in IKr amplitude. Missense mutations in the amino terminus of HERG cause the channels to deactivate (close) much faster than normal (57). An increase in the rate of deactivation blunts the voltage-dependent increase in IKr that normally results January 2001

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Figure 4. Drug trapping within the K⫹ channel vestibule. Class III antiarrhythmic agents traverse the lipid bilayer as neutral molecules and equilibrate in the cytosol as positively charged molecules (top left). In the resting state, the activation gate (blue) of the K⫹ channel (green) remains closed. Upon depolarization, the activation gate opens and the drug enters the vestibule to block the channel (top right). The channel then enters a long-lasting closed state (inactivation), which increases the affinity of drug binding (bottom right). Upon repolarization, the activation gate closes (deactivation) and traps the drug within the channel vestibule (bottom left). On subsequent depolarizations, the channel is not available to conduct K⫹ because the drug remains bound.

from rapid recovery from inactivation and slow deactivation of channels during repolarization. Two different missense mutations in KCNE1 have also been shown to cause an increase in the rate of IKs deactivation and a reduction in magnitude (51). In all cases, these LQTSassociated mutations result in a decrease in the magnitude of delayed rectifier K⫹ current during the repolarization phase of ventricular action potentials, and the associated prolongation of the QT interval on the ECG. Dominant missense mutations in KvLQT1 cause loss of function and a dominant-negative effect (58 – 60). When expressed alone, the mutant subunits do not form functional channels. However, when coexpressed with normal subunits, they combine to form dysfunctional heterotetramers. These abnormally folded proteins usually undergo rapid degradation, thus reducing the magnitude of IKs. Ten mutations have been found in KCNE1, and all but one are missense mutations (61). Two mutations that are located in the putative cytoplasmic region of the protein (S74L, D76N) cause a shift in the voltage 56

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dependence of IKs activation to more positive potentials and increase the rate of deactivation; one (D76N) has a strong dominant-negative effect (51). These biophysical changes would also reduce IKs. Recessive mutations in KVLQT1 or KCNE1 cause a more rare and severe form of LQTS known as Jervell and Lange-Nielsen syndrome (50,62,63). This disorder is associated with sensorineural deafness and more severe arrhythmias than the dominantly inherited Romano-Ward syndrome. Deafness results from loss of IKs channel function in the inner ear. IKs normally conducts K⫹ into the inner ear, creating the potassium-rich fluid, endolymph. Recessive mutations of KVLQT1 or KCNE1 cause loss of IKs, inadequate endolymph production, and degeneration of the organ of Corti (64). According to a recent tabulation of all known novel mutations, there are presently 177 known mutations associated with LQTS (61). Mutations in HERG are the most common (45%), followed by KVLQT1 (42%), SCN5A (8%), KCNE1 (3%), and KCNE2 (2%). At the

K⫹ Channels and Cardiac Arrhythmias/Tristani-Firouzi et al

present rate of discovery, many more LQTS-associated mutations in these genes are likely to be described in the future. Moreover, because mutations were not found in some affected families, other genes that cause LQTS remain to be discovered.

INDUCTION OF VENTRICULAR ARRHYTHMIAS CAUSED BY PHARMACOLOGICAL BLOCK OF IKr Many commonly used medications, including antiarrhythmic, antihistamine, antipsychotic, and antibiotic agents, are associated with acquired LQTS (Table 2). The mechanism(s) responsible for drug-induced LQTS has not been definitively determined for all these agents, but for those that have been studied, a common effect has been identified. These drugs either block IKr or inhibit liver enzymes that are important for metabolic degradation of other drugs that block IKr. Thus, drug-induced LQTS is mechanistically linked to LQT2 caused by mutations in HERG (24). Although the factors that determine which patients are at greatest risk for developing druginduced LQTS and arrhythmia are not fully understood, it has been established that low serum K⫹ or Mg2⫹, or congenital LQTS are important risk factors (44,53). The most extensively characterized IKr blockers are the methanesulfonanilide class III antiarrhythmic agents (eg, E4031, MK-499, dofetilide, d-sotalol). These compounds were developed to prevent ventricular fibrillation and are quite effective in canine models of arrhythmia (65,66). Unfortunately, these drugs also have the side effect of inducing ventricular arrhythmias in some patients. The methanesulfonanilide compounds require HERG channels to open before they can gain access to the high affinity binding site located inside the channel vestibule (67). This requirement for channel opening indicates that the resting state of the channel is not blocked by drug and that the binding site is located behind the activation gate (Figure 4). Once inside the channel vestibule, closure of the activation gate (deactivation) traps the drug inside the channel and greatly slows recovery of the channel from the blocked state (68). Most experimental evidence also suggests that these drugs bind with greater affinity to channels in an inactivated state (69,70). Because druginduced LQTS is such a common observation, the development of all new drugs includes procedures to detect this unwanted side effect.

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