Multiple Mechanisms in the Long-QT Syndrome
Current Knowledge, Gaps, and Future Directions
Abstract The congenital long-QT syndrome (LQTS) is characterized by prolonged QT intervals, QT interval lability, and polymorphic ventricular tachycardia. The manifestations of the disease vary, with a high incidence of sudden death in some affected families but not in others. Mutations causing LQTS have been identified in three genes, each encoding a cardiac ion channel. In families linked to chromosome 3, mutations in SCN5A, the gene encoding the human cardiac sodium channel, cause the disease. Mutations in the human ether-a`-go-go–related gene (HERG), which encodes a delayed-rectifier potassium channel, cause the disease in families linked to chromosome 7. Among affected individuals in families linked to chromosome 11, mutations have been identified in KVLQT1, a newly cloned gene that appears to encode a potassium channel. The SCN5A mutations result in defective sodium channel inactivation, whereas HERG mutations result in decreased outward potassium current. Either mutation would decrease net outward current during repolarization and would thereby account for prolonged QT intervals on the surface ECG. Preliminary data suggest that the clinical presentation in LQTS may be determined in part by the gene affected and possibly even by the specific mutation. The identification of disease genes in LQTS not only represents a major milestone in understanding the mechanisms underlying this disease but also presents new opportunities for combined research at the molecular, cellular, and clinical levels to understand issues such as adrenergic regulation of cardiac electrophysiology and mechanisms of susceptibility to arrhythmias in LQTS and other settings.
The idiopathic or congenital LQTS1 is an inherited disease characterized by prolonged ventricular repolarization and a high risk for sudden cardiac death. Most of the life-threatening arrhythmias in LQTS occur during physical or emotional stress, although in some families, sudden death occurs during sleep. Thus, LQTS has been regarded as an important example of neurally mediated sudden cardiac death unrelated to coronary artery disease. Studies of the mechanisms underlying arrhythmias in LQTS have integrated clinical science, basic electrophysiology, and molecular genetics. Understanding of these mechanisms not only would contribute to improved therapy for patients with this intriguing, albeit uncommon, disease but also would provide information important for understanding neurally mediated arrhythmogenesis in other situations.
To define the state of the art in understanding LQTS, a multidisciplinary task force was convened in early 1995 by the Sudden Arrhythmia Death Syndromes Foundation. Since that time, important advances have occurred in studies of the molecular genetics and electrophysiology of LQTS. In this review, background information on the clinical and ECG features of LQTS and the probable underlying electrophysiological mechanisms will first be described. The molecular defects that have been linked to LQTS will then be presented, followed by a discussion of how these findings provide the groundwork for new lines of inquiry at the molecular, cellular, and clinical levels.
Clinical Features of LQTS
The inherited LQTS is characterized by recurrent syncope (often during exercise or emotional stress), QT prolongation and T- and U-wave abnormalities on the ECG, and sudden death. One form of LQTS, described in 1957 by Jervell and Lange-Nielsen,2 includes deafness and probable autosomal recessive inheritance.2 3 4 A similar condition, without deafness and with autosomal dominant transmission, was subsequently reported independently by Romano et al5 and Ward.6 Relatively few cases of the Jervell–Lange-Nielsen form are currently recognized, whereas the Romano-Ward variant is recognized with increasing frequency. Most of the information currently available regarding the syndrome relates to the autosomal dominant (Romano-Ward) form. There is considerable variability in the clinical presentation, and diagnosis is sometimes difficult. When the correct diagnosis is not made, syncopal events often are attributed incorrectly to a primary seizure disorder or to vasovagal events. To assist in diagnosis, criteria incorporating ECG, clinical, and family history findings have been developed (Table 1⇓).7 8
Syncope and Sudden Death
The cause of syncope in LQTS is the polymorphic ventricular tachycardia torsades de pointes. The term “torsades de pointes” (twisting of the points) was coined in 1966 by Dessertenne,9 who used it to describe the pause-dependent polymorphic ventricular tachycardias he observed in an elderly patient with syncope, heart block, and marked QT prolongation. Although others have used this term virtually interchangeably with all forms of polymorphic ventricular tachycardia, most authorities would confine its use to those polymorphic ventricular tachycardias with a distinctive time-dependent change in electrical axis (“twisting of the points”), along with at least some of the QTU abnormalities described here.
The onset of torsades de pointes in patients with LQTS can follow one of two patterns. In some patients, bradycardia with associated marked QT prolongation (which appears to be greater than what would be expected merely from rate slowing) is responsible. In other patients, adrenergic stress with sinus tachycardia is the rule. Nevertheless, even in the latter group, torsades de pointes often develops in a pause-dependent fashion, ie, immediately after a sinus beat after a postectopic pause (Fig 1A⇓). Most episodes of sudden death in LQTS almost certainly result from ventricular fibrillation triggered by torsades de pointes, but the mechanisms whereby this transition occurs are unknown. The frequencies of syncope and sudden death vary from family to family. These events are often precipitated by sudden autonomic stimuli such as anger, fear, or startle, although some sudden deaths occur during sleep or rest. These presentations tend to cluster in families and thus, as described below, may reflect different mutations in different families.
Equal numbers of males and females should be affected in an autosomal dominant condition. This has been shown for LQTS families in which affected individuals were identified by use of genetic markers.10 However, several studies in which the diagnosis was ascertained by ECG or by symptom criteria show a predominance of females.11 12 13 This observation raises the possibility of a contribution by the products of other genes (“modifier genes”) to prolongation of the QT interval in women or to its shortening in men. Interestingly, a similar female preponderance also has been described in drug-associated torsades de pointes,14 an observation that supports the link discussed below between the inherited and drug-induced arrhythmias.
In most patients with LQTS, the rate-corrected QT interval (QTc by Bazett's formula) is >0.46 seconds, but some affected individuals have QTc values in the normal to upper-normal range of 0.41 to 0.45 seconds.10 QT prolongation is absent in ≈5% to 10% of gene carriers, and it has also been reported that syncope or cardiac arrest can occur in ≈5% of LQTS family members who have an apparently normal QT interval.11 These findings suggest reduced penetrance or contributions by modifier genes or environmental factors (such as electrolyte abnormalities) to the clinical manifestations of LQTS.
T and U Waves
The ECG changes in LQTS include considerably more than simple prolongation of the QT interval. Most of the characteristic ECG findings are consistent with the concept that repolarization in LQTS displays substantial spatial and temporal heterogeneity.15 For example, QT dispersion, as assessed by the difference between the longest and shortest QT intervals on a 12-lead ECG, is increased in LQTS, indicating spatial heterogeneity in repolarization.16 The normal range for QT dispersion determined in this fashion is 46±18 ms, and in patients with LQTS, it is 133±21 ms. In addition, the T wave often has a notched or biphasic appearance.17 These findings, in turn, suggest that abnormal AP prolongation in LQTS may occur to a greater extent in some cells than in others. As described below, it is increasingly recognized that even under normal conditions, APs in the ventricle are highly heterogeneous, which suggests that different ion channel lesions might produce different and distinctive ECG patterns. Indeed, specific T-wave patterns have been associated with specific gene defects (Fig 1B⇑),18 although overlap among patterns may exist.
Another manifestation of spatially heterogeneous repolarization is a regional left ventricular wall motion abnormality,19 observed in approximately half the patients investigated; it is more prevalent among those with syncope and is abolished by verapamil.20 An unusual but diagnostic ECG manifestation of LQTS is T-wave alternans (Fig 1C⇑), in which the T wave is not only prolonged and morphologically bizarre (implying spatial heterogeneity of repolarization) but also varies on a beat-to-beat basis21 22 and with sympathetic activation,21 23 which indicates temporal variability in repolarization.
In addition to bifid T waves, U waves seem prominent in some patients with LQTS. Whereas U waves can occur in normal subjects with sinus bradycardia, U waves in some patients with LQTS are exaggerated, although objective data addressing this point are not available. Like T waves, U waves can also be labile and display alternans; in fact, marked prolongation of the QT interval in LQTS can represent a very prominent U wave (Fig 1C⇑).
Another important ECG manifestation of LQTS is sinus bradycardia and decreased heart rate with exercise. Again, the extent to which this is manifest in an individual subject is highly variable, which indicates that either certain gene defects are more likely to be associated with this manifestation of the disease than others or expression of modifier genes determines whether an individual subject develops sinus bradycardia.
Some ECG abnormalities, such as obvious QT prolongation and deformity with T-wave alternans, are diagnostic of LQTS. Conversely, borderline QT-interval prolongation is much more sensitive but far less specific. To circumvent difficulties in diagnosis, some have advocated the use of provocative tests, such as determination of the extent of QT-interval shortening with treadmill exercise or evaluation of the presence of U waves or arrhythmias with epinephrine or isoproterenol infusion.24 25 However, exercise under controlled conditions seldom provokes torsades de pointes, whereas spontaneous episodes often arise with the combination of exercise and emotional stress. Moreover, it is now apparent that the QT response to exercise may be determined to a large extent by the specific mutation causing LQTS in an individual. Thus, subjects with the sodium channel inactivation defect on chromosome 3 described below not only appear to have a distinctive ECG pattern at rest (Fig 1B⇑)18 but also are reported to display marked shortening of the QT interval during exercise.26 In these subjects, the onset of torsades de pointes is usually bradycardia dependent. Conversely, subjects with defects linked to a potassium channel gene defect on chromosome 7 seem to display a different QT pattern at rest, abnormally long QT intervals during exercise, and adrenergic-dependent onset of torsades de pointes. Further studies are seeking to verify and expand these preliminary observations. These genotype-phenotype correlations not only provide important clues to the mechanisms underlying arrhythmogenesis in LQTS but also may become important for clinicians as specific therapies for individual genetic subtypes of LQTS are identified and tested in prospective trials.
The mortality of untreated symptomatic patients with LQTS exceeds 20% in the year after their first syncopal episode and approaches 50% within 10 years.7 With therapy, this can be reduced to 3% to 4% in 5 years. Although no placebo-controlled, randomized, clinical trials have been undertaken, strong evidence supports the use of antiadrenergic interventions as mainstays of therapy. The trigger for many life-threatening events is emotional or physical stress, probably associated with a sudden increase in sympathetic activity. β-Adrenergic blockade prevents new syncopal episodes in ≈75% of patients. Among those with persistent syncope despite β-blockade, left cardiac sympathetic denervation appears to provide significant additional protection.27 28 In patients with evidence of pause- or bradycardia-dependent arrhythmias, cardiac pacing is a rational and effective adjunct to β-blockers.29 30 The acute therapy of torsades de pointes in LQTS includes cardioversion, elimination of any provoking factor such as hypokalemia or the concomitant use of QT-prolonging antiarrhythmic or other drugs, and β-blockade.
Rarely, torsades de pointes persists despite therapy with a combination of β-blockade, left cardiac sympathetic denervation, and pacing. The implantable cardioverter-defibrillator has been used successfully in this setting, although it is not first-line therapy because shocks from the device can precipitate further emotional distress and thus further arrhythmias. Other pharmacological interventions have been used sporadically or have been proposed on the basis of experiments in vitro or in animal models; these include α-blockers, calcium channel blockers, potassium channel openers, right cardiac sympathetic denervation, or sodium channel blockers.24 25 31 32 33 34 35 None of these therapies have been tested in sufficiently large numbers of patients that they can be viewed as anything but experimental in highly refractory patients.
With the identification of the precise molecular defect in some cases of LQTS, specific mechanism-based therapies may be developed and tested. For example, preliminary reports suggest that the sodium channel blocker mexiletine may be useful in shortening the QT interval in patients with a defect linked to the sodium channel gene on chromosome 3,26 and in vitro studies provide a rationale for this approach.36 37 38 39 This approach, however, remains highly speculative and obviously cannot be taken to include any sodium channel blocker, since some prolong the QT interval. Further studies in defined subsets of patients, with the end points being not QT-interval measurements but rather syncope and sudden death, will be required before specific recommendations for long-term, mechanism-based therapy can be formulated.
Mechanisms Underlying LQTS
Competing Hypotheses to Explain LQTS
The clinical features of LQTS led to the formulation of two hypotheses to explain its pathogenesis40 : (1) The striking adrenergic dependence of symptoms and the beneficial effect of antiadrenergic interventions led to the suggestion that there was an abnormality (eg, an imbalance) in the sympathetic innervation to the heart. Such a formulation would explain abnormal sinus rate, abnormal repolarization (possibly), and adrenergic dependence of arrhythmias and their correction by antiadrenergic drugs. (2) Many of the ECG manifestations of LQTS are indistinguishable from those in patients with “acquired” QT-interval prolongation and torsades de pointes, most often seen during therapy with drugs whose major effect is to alter cardiac ion currents. Thus, a second theory was that LQTS was due to an “intrinsic” abnormality in the mechanisms responsible for cardiac repolarization. Such an intrinsic abnormality would explain the long QT interval and could, as described below, also explain the arrhythmias and even their adrenergic dependence. At all three chromosomal loci at which mutations causing LQTS have been identified (accounting for >90% of cases), the mutated gene encodes a cardiac ion channel protein. Ion channels are structures that are responsible for currents that determine cardiac repolarization; thus, LQTS is due to an intrinsic abnormality in the vast majority of patients.
Ion Channels and Currents
Ion channels are individual proteins or complexes of proteins that reside in the cell membrane. The distinguishing characteristic of ion channel proteins is that, in response to such exogenous signals as a change in voltage across the cell membrane, they can form pores for the entry or egress of specific ions (sodium, potassium, calcium, etc). Thus, individual currents created by this movement of charged ions reflect the function of specific ion channel proteins. The application of whole-cell and patch-clamp techniques to cardiac cells has allowed electrophysiologists to identify multiple currents that flow across the cell membrane to determine cardiac repolarization. A key feature of the cardiac cell membrane during repolarization is that even very small changes in individual ion currents can markedly alter the delicate balance between inward and outward current flow during repolarization and thereby prolong or shorten the AP. This fundamental concept provides the basis for understanding how subtle abnormalities in individual ion channel genes can cause LQTS.
Many of the genes that encode ion channel proteins have been identified (Fig 2⇓, Table 2⇓). The features of a given ion current often can be replicated by expressing a single “α-subunit” gene in systems such as the Xenopus oocyte. However, for many ion channels, the presence of one or more ancillary proteins (often called β-subunits), the products of different genes, may be required for the faithful recapitulation of all the current's physiological features. Description of the relationship between individual ion currents identified by electrophysiological studies and individual ion channel gene products, whose function is commonly characterized in heterologous expression systems such as the Xenopus oocyte, is an area of active investigation.
Cellular Mechanisms of Arrhythmogenesis in LQTS
Since the QT interval is a surface manifestation of APD, any theory to explain arrhythmias in LQTS should include AP prolongation. Major advances in the understanding of the relationship between abnormally long QT intervals and arrhythmogenesis came in the early 1980s, when in vitro and in vivo studies linked AP prolongation to the development of early afterdepolarizations (EADs) in conducting system tissue (Purkinje fibers).41 42 43 EADs are oscillations or deformities in the repolarization phases (plateau [early] and phase 3 [late]) of the AP (Fig 3⇓). Some studies suggest that the mechanisms and possible clinical consequences of early and late EADs may differ. When EADs are observed in vitro, they often give rise to triggered APs. Such triggered activity would produce premature beat(s) if propagated in the whole heart. EADs are induced by interventions that decrease repolarizing (potassium) currents (eg, administration of cesium or antiarrhythmic drugs41 43 44 ) and/or that increase inward currents carried by calcium or sodium.45 46 47 These changes may produce EADs directly, or they may alter the trajectory of the AP to produce secondary changes in otherwise normal ion currents that then result in EADs (Fig 3⇓). The candidate currents may be inferred from Fig 2⇑; experimental and modeling studies suggest that increased inward current through L-type calcium channels or through sodium-calcium exchange is responsible.48 49 50 The triggered upstroke is most likely due to inward current through L-type calcium channels at positive potentials, whereas upstrokes occurring later during phase 3 (eg, see Fig 3⇓) might be due to inward current through sodium or T-type calcium channels. Triggered activity arising from EADs may initiate or help maintain torsades de pointes, as described below.
Multiple abnormalities in ion current flow across cell membranes could also explain T-wave alternans. The general principle, illustrated in Fig 4⇓, is that QT alternans reflects a failure of all ion channels to recover back to their rested state during diastole before the next AP. Alternans thus may be exaggerated when diastole is shortened (by fast rates or AP prolongation).51 The development of T-wave alternans in LQTS could thus reflect AP prolongation itself (with shorter diastolic times); abnormally slow deactivation of a potassium current (eg, see Fig 4⇓) or abnormal intracellular calcium handling may also play a role.51 52 53 54 55 The finding of alternans phenomena at both fast and slow rates in LQTS (Fig 1C⇑) suggests that multiple mechanisms may be involved. The morphology of the QT interval on the surface ECG is determined in part by the heterogeneity of the underlying APDs; thus, the presence of repolarization alternans provides further evidence for the temporal and spatial heterogeneity of repolarization in LQTS.
A problem in attributing the LQTS phenotype to EADs and EAD-related triggered activity in Purkinje fibers is that the specialized conducting system represents only a very small portion of the mass of the ventricle and that EADs are much more difficult to elicit in ventricular cells than in Purkinje fibers. Therefore, phenomena such as bifid T waves or U waves that presumably represent delayed repolarization of substantial portions of the ventricle are difficult to explain. An answer to this apparent paradox may lie with the identification in the midmyocardium of a large population of cells, called M cells, with distinctive conducting system–like electrophysiological properties.56 M cells display marked AP prolongation in response to stimuli such as hypokalemia, slow driving rates, or the presence of AP-prolonging drugs.57 In addition, EAD-related triggered activity is common in M cells with these stimuli. Some computer simulations have even suggested that apparent EADs recorded by use of monophasic AP methods may actually reflect marked prolongation of APs in M cells with normal APs in endocardium, in the absence of EADs in either tissue.58
In Vivo Mechanisms Underlying Torsades de Pointes
The mechanisms that underlie sustained episodes of torsades de pointes are not well understood. One possibility is raised by the observation that driving the heart rapidly at different rates from multiple foci produces a polymorphic ECG pattern.59 60 EAD-related triggered activity arising in multiple loci at different rates could similarly result in the distinctive ECG appearance. Triggered activity arising from EADs has also been reported in experimental animals, and it seems to produce many of the ECG features of torsades de pointes.42 46 61 A second possibility is that torsades de pointes result from reentrant excitation. In this scenario, the unidirectional block required for reentry is created by the M-cell layer with its abnormally prolonged APs. The reentrant pathway would vary in a relatively organized fashion from beat to beat as the site of reentry shifted along the functional barrier created by the M-cell region.62 63 A number of studies have identified heterogeneity of repolarization as an important risk factor for torsades de pointes in both LQTS and the drug-acquired forms.16 64 65 This lends support to the concept that reentrant excitation may be important in the maintenance of an episode of the arrhythmia. Finally, it is possible that torsades de pointes occurs as a result of EADs in the conducting system and the fact that propagation to the ventricle varies on a beat-to-beat basis, perhaps as a consequence of variable conduction block in the M-cell layer. Studies suggesting that the arrhythmia originates in the subendocardium support such a hypothesis.66
Chromosomal Defects in LQTS
The first chromosomal locus to which LQTS was linked was in the telomeric region of chromosome 11 (11p15.5) (LQT1; see Fig 5⇓). The initial analysis reported by Keating67 was performed in a very large family and demonstrated tight linkage, with a logarithm of the odds score >15, to the H-ras-1 locus in this region. In other words, there was a <1 in 1015 likelihood that the abnormality was not at this locus. This was an intriguing finding, because the H-ras locus encodes an oncogene that may play a role in modulation of cardiac potassium current by intracellular signaling pathways68 ; thus, a linkage to the adrenergic features of LQTS was suggested. However, subsequent studies excluded H-ras-1 as the disease gene, because no mutations were found on sequence analysis,69 70 recombination events between the disease phenotype and H-ras-1 were identified,69 70 and fine mapping of this region suggested that the disease gene was more centromeric.71 In early 1996, mutations in a newly cloned gene, KVLQT1, were identified as the cause of LQT1.72 No information is yet available on the function of KVLQT1, although its inferred partial amino acid sequence strongly suggests that it encodes a potassium channel. mRNA transcripts hybridizing to a KVLQT1 probe are present in the human heart, kidney, placenta, and lung, with greatest abundance in the heart.
Available evidence indicates that >50% of affected families link to KVLQT1,72 and all of the first seven families studied were linked to the chromosome 11p15.5 locus.73 However, Towbin and colleagues74 reported evidence of genetic heterogeneity in LQTS shortly after the initial reports, and subsequent studies identified families that failed to link to this locus, indicating that abnormalities in other chromosomal locations could cause LQTS.75 76 77 78 79 In late 1994, Jiang and colleagues80 provided evidence for linkage to chromosome 7q35-36 (LQT2) in nine families and linkage to chromosome 3p21-24 (LQT3) in three other families. A fourth locus has been identified at 4q25-27 (LQT4) in a single large French kindred81 ; this family is unusual because there is a high incidence of atrial fibrillation along with the LQT phenotype. In addition, several families are excluded from linkage at all four known regions, suggesting at least one more and possibly several more disease-causing genes for LQTS. The gene encoding the Jervell–Lange-Nielsen form associated with congenital deafness has not been identified. LQT1 and LQT2 seem to be the most common forms of the disease and LQT4 the rarest, with only the single kindred reported to date.
Once a chromosomal locus linked to a disease such as LQTS is identified, two general approaches can be used to identify the disease-causing gene. One is the “positional candidate gene” approach. In this method, once a chromosomal locus is identified, it is compared with that of genes whose dysfunction might reasonably be expected to result in the abnormal phenotype. In the case of LQTS, this would include genes that encode cardiac ion channels (Table 2⇑) as well as genes involved in regulation of sympathetic activity in the heart. This candidate gene approach was used successfully to identify mutations in two genes, HERG on chromosome 782 and SCN5A on chromosome 3,83 the causes of LQT2 and LQT3, respectively. In the second overall approach, a candidate gene is identified not on the basis of physiological considerations but rather through cloning of large portions of the genome in the region linked to the disease. Exons identified within these cloned regions are then analyzed to determine whether mutations segregating with the disease phenotype are present. This positional cloning and mutational analysis approach was used to identify mutations in KVLQT1 on chromosome 11,72 the cause of LQT1. As shown in Table 3⇓, mutations have been identified in multiple loci in different kindreds for each of the three LQTS disease genes.
Defects in Outward Current and LQTS
The first potassium channel genes to be cloned were isolated from the Shaker mutant (so called because it shakes when exposed to ether) of the fruit fly Drosophila melanogaster.84 Genes of the Shaker family (called Kv1.x) have also been isolated in humans, and some (eg, Kv1.5) are expressed in the heart.85 Subsequently, Shaker-like genes were cloned from a different Drosophila mutant, ether-a`-go-go (eag).86 Eag does not seem to be present in humans, but in 1994, Warmke and Ganetzky87 reported the cloning of a human eag-related gene (HERG) from a human hippocampal cDNA library. At the time HERG was cloned, its function was not established, although its inferred amino acid sequence did exhibit the six membrane-spanning segments motif seen in other potassium channels such as Shaker. HERG mRNA transcripts have been found in heart, and mutations in multiple regions of the HERG gene have been identified as the cause of LQT2.82 The coding sequence is unusually rich in cytosine-guanosine pairs, which may predispose this gene to mutagenesis. When HERG is expressed in Xenopus oocytes,88 89 most of the major physiological features of the rapidly activating cardiac delayed rectifier potassium current (IKr) are observed: inward rectification, relatively rapid activation, and block by drugs of the methanesulfonanilide class.90 91 In addition, IKr, unlike most other potassium currents, increases when extracellular potassium is increased.88 92 Lowered extracellular potassium has also been found to markedly potentiate drug block of IKr93 ; this finding may explain the relationship between hypokalemia and the development of torsades de pointes, and it is consistent with preliminary reports suggesting that elevation of extracellular potassium may increase IKr and shorten QT in some patients with LQTS on a congenital or drug-induced basis.93A Interestingly, despite the adrenergic features of LQTS and despite a cyclic nucleotide–binding domain on HERG (which is actually the site of one mutation), IKr is generally thought to be unaffected by β-adrenergic stimulation.94 95 It is likely that KVLQT1 also encodes a potassium channel, on the basis of its inferred amino acid sequence, which contains both the six membrane-spanning segments motif and an ion-conducting pore sequence common to potassium channel proteins.72
Other potassium channel proteins (eg, those of the Shaker family) are thought to coassemble in groups of four (tetramers),96 97 with or without ancillary β-subunits,98 99 100 to form functional channels. Since HERG proteins exhibit a topology similar to that of Shaker proteins, coassembly of a single abnormal HERG subunit with even three normal ones might be sufficient to disrupt IKr. This “dominant negative” effect would explain the autosomal dominant nature of LQT2. Two intragenic deletions of HERG have been identified (Table 3⇑; Fig 6A⇓). One results in a stop codon within the region encoding the S1 transmembrane domain; another results in a nine-amino-acid deletion within the S3 transmembrane domain. These mutant proteins do not form functional channels and do not interact with normal HERG channels when coexpressed in Xenopus oocytes.101 Thus, individuals with these mutations would be predicted to express half the normal number of channels carrying IKr. Three missense mutations that result in single amino acid substitutions in the S5 transmembrane region (A561V), pore region (G628S), and S2 transmembrane region (N470D) have also been evaluated.101 A561V and G628S mutant channels do not express detectable currents in Xenopus oocytes but do cause a dominant negative suppression of wild-type HERG function in oocytes injected with RNA encoding both wild-type and mutant channels (Fig 6B⇓). N470D mutant proteins form functional channels with altered kinetic properties, and coexpression of normal and N470D channels reveals a dominant negative effect. The order of severity in reduction of HERG function for mutations characterized to date is G628S (most severe) > A561V > N470D > intragenic deletions (least severe).101 It will be interesting to determine whether this spectrum of HERG channel dysfunction correlates with the severity of disease in patients with LQT2.
Pause-dependent torsades de pointes is a well-recognized complication102 of therapy with drugs that specifically block IKr (eg, dofetilide,103 almokalant104 ) or drugs that block IKr and also exert other electrophysiological effects (eg, quinidine,93 105 sotalol,90 106 ibutilide107 108 ). Thus, the identification of mutations in HERG as a cause of LQTS provides an important link between LQTS and drug-associated torsades de pointes. The [K+]o sensitivity of IKr may help explain why hypokalemia is such an important risk factor for torsades de pointes; in addition, drug-induced IKr block is relieved by raising [K+]o,93 and recent studies suggest marked QT shortening in LQT2.93A These data raise the possibility that drugs that interact with a potassium-binding site on HERG might be beneficial in LQT2. The bradycardia characteristic of both LQTS and the drug-induced form suggests that IKr can play an important role in sinus node function. Interestingly, although the onset of torsades de pointes is usually pause dependent in drug-related cases, a role for adrenergic stimulation has also been suggested by the observation that heart rate seems to increase just before the onset of the arrhythmia.109
Many other outward currents have been identified by electrophysiological studies in cardiac cells, and clones corresponding to some of these have been identified (Fig 2⇑). It is also increasingly clear that β-subunits may be important modulators of the function or level of expression of some of these channels. Defects in any of these channels or β-subunits could conceivably cause LQTS. Genes encoding K+ channels that lack the six membrane-spanning segment motif of Shaker and HERG have also been cloned. The inward rectifiers (IRK family) have a topology similar to that of Shaker and HERG, although they appear to represent truncated versions of these channels, with only two putative membrane-spanning segments.110 111 112 113 The minK gene encodes a short protein (130 amino acids).114 115 116 When minK is expressed in Xenopus oocytes, a current strongly resembling the slowly activating delayed rectifier (IKs) is recorded. However, there is considerable controversy as to whether minK encodes this current directly or acts as a regulator or activator of other genes that are responsible for IKs or other ion currents, including IKr.117 118 Other outward channels that control the fine balance between inward and outward current during repolarization include the transient outward current (Ito), a high-conductance plateau channel (IKp),119 and chloride channels.120 121 An important example of the latter is the cAMP-activated chloride current (ICl-cAMP) that appears to be encoded by the cystic fibrosis transmembrane conductance regulator, the gene whose dysfunction causes cystic fibrosis.122 Potassium channels that may play especially important roles in repolarization during pathological stress such as stretch123 or myocardial ischemia124 have also been identified; well-characterized examples include potassium channels regulated by ATP, acetylcholine, and intracellular sodium.125 126 The maintenance of normal intracellular ionic homeostasis requires normalization of intracellular calcium and sodium after each AP. Extrusion of sodium and calcium by ionic pumps is electrogenic (ie, results in a net transmembrane current that can be inward or outward depending on specific physiological conditions), and so dysfunction of these pumps could also underlie LQTS.
Defects in Inward Current and LQTS
Mutations in SCN5A, the gene encoding the cardiac sodium channel α-subunit, are the cause of LQT3.83 127 Three LQTS mutations in this large protein have been identified (Table 3⇑; Fig 7A⇓). Two are point mutations, and the third is a nine-nucleotide deletion that results in deletion of three amino acids—lysine (K), proline (P), and glutamine (Q)—in the intracytoplasmic loop linking domains III and IV of the channel protein. It has been known for several years that mutations in the III-IV linker can disrupt sodium channel inactivation.128 129 Moreover, toxins that inhibit sodium channel inactivation can cause EADs and triggered activity in vitro and long-QT–like arrhythmias in vivo.45 46 130 The ΔKPQ mutation results in a much more subtle change in the behavior of the sodium channel. Macroscopic activation and inactivation seem near normal. However, the wild-type channel generally inactivates completely during long depolarizing pulses, whereas the ΔKPQ mutant exhibits a small, sustained inward current, even with long depolarizing pulses (Fig 7B⇓).36 This small inward current is most likely sufficient to disrupt the normal balance between inward and outward currents during the plateau phase and hence prolong cardiac APs. At the single-channel level, the maintained inward current reflects a sporadic failure of the sodium channel to inactivate normally. Rather, in 3.5% of sweeps, it displays continuous opening and closing behavior and occasionally even remains open at the end of a very long depolarizing pulse. Presumably, this abnormal inactivating behavior reflects a failure of the III-IV linker to appropriately effect inactivation. The abnormal “late” inward current would be produced by even a single abnormal allele, and thus this “gain of function” explains the autosomal dominant nature of LQT3. The functional consequences of the two described point mutations (Table 3⇑; Fig 7A⇓) seem to be qualitatively similar but perhaps less severe than those for the ΔKPQ mutant.38 Thus, as in the case of the HERG mutations in LQT2, there may be a spectrum of severity of sodium channel dysfunction in LQT3 and thus a possible spectrum of disease severity among individuals with LQT3.
Whether the maintained inward sodium current completely accounts for the LQT3 phenotype is unknown. It is possible, for example, that increased intracellular sodium loading consequent to maintained inward sodium current could in turn activate electrogenic sodium-calcium exchange, thereby blunting the extent of AP prolongation while increasing intracellular calcium. Increased intracellular calcium, in turn, might account for sensitivity to adrenergic stimuli or might be directly arrhythmogenic.131 This scenario would be most prominent at rapid rates (when sodium loading is greatest) and thus is consistent with the report that LQT3 patients appear to display relatively normal (or even supernormal) QT shortening during exercise or other tachycardia-related sodium loading.26
Since the cardiac sodium channel is not thought to play a major role in sinus node function, it is not clear how the SCN5A defect explains the sinus bradycardia seen in LQTS, and specifically in LQT3.26 Perhaps sodium channels are more important in determining heart rate than previously appreciated; alternatively, variable contributions of modulator genes may be responsible. The ΔKPQ mutation also lies close to a putative protein kinase C phosphorylation site, so it is conceivable that the response of the mutant channel to adrenergic stimulation will prove to be abnormal.
Defects in other structural proteins encoding inwardly conducting ion channels could similarly alter inward current and thus cause other forms of LQTS. The L-type calcium channel has been cloned (Table 2⇑), but a T-type clone has not been identified. In addition, the syndrome could be attributable to defects in accessory subunits that can modify levels of expression and/or kinetics of these principal α-subunits. An accessory sodium channel β-subunit has been isolated; coexpression of this subunit with brain sodium channel α-subunit in Xenopus oocytes increases sodium current and accelerates its activation and inactivation.132 However, whether the β-subunit, which is detectable in heart, is necessary to faithfully recapitulate cardiac sodium channel function remains unclear.133 134 Transcripts for accessory calcium channel α2-δ and β-subunits have been identified in rabbit heart and probably are present in human heart.
Abnormalities That Might Underlie Other Forms of LQTS
Defects in KVLQT1, HERG, and SCN5A probably account for >90% of identified cases of LQTS. It seems reasonable that defects in other genes encoding cardiac ion channels will be identified as the cause of LQTS linked to other loci. These could be defects in channels or pumps responsible for inward current or those responsible for outward currents, as described above. It is also conceivable that abnormalities in genes involved in the regulation of ion channel expression might be defective; one possibility discussed below is defect(s) in sympathetically mediated expression of ion channel or other genes. Precedents in the study of other genetic diseases involving the cardiovascular system provide some direction in this regard. For example, defects in a number of genes encoding elements of the cardiac sarcomere can produce the hypertrophic cardiomyopathy phenotype.135 136 By analogy, mutations in any one of a number of proteins whose normal function determines the AP may produce the LQTS phenotype. In hypertrophic cardiomyopathy, it is now recognized that some mutations carry a much graver prognosis than others.137 This finding not only has clinical implications for management and genetic testing of patients but also provides a major lead for further studies of basic mechanisms; even the rudimentary current state of knowledge regarding LQTS suggests that similar genetically determined heterogeneity occurs. Another possibility is raised by studies of X-linked muscular dystrophy, in which a wide range of clinical manifestations can be observed with small changes in the mutated gene that encodes a protein called dystrophin.138 In Duchenne muscular dystrophy, severe skeletal myopathy (with or without dilated cardiomyopathy) is observed, whereas in Becker-type muscular dystrophy there is a milder skeletal myopathy (again with or without a dilated cardiomyopathy). A third defect linked to the dystrophin gene is the cardiospecific disorder called X-linked dilated cardiomyopathy. Interestingly, the cardiac defects in Duchenne and Becker-type muscular dystrophy are quite similar to each other, whereas the defect in X-linked dilated cardiomyopathy probably resides in a cardiospecific region of the dystrophin gene. Thus, subtly different mutations in a single gene may cause very dramatic differences in the clinical phenotype. Importantly, the function of dystrophin (which now appears to play a role in cytoskeletal support) was unknown when mutations in the gene were identified in muscular dystrophy.139 Consequently, it must be recognized that merely identifying the disease-causing gene, especially a novel gene, does not guarantee unlocking the secrets behind the mechanism of disease. In LQTS, therefore, it should not be completely surprising that, even after new gene defects are identified, the mechanisms producing the disease may still remain something of a mystery.
Importantly, not all of the clinical manifestations of LQTS need be attributable directly to the mutant gene product. As discussed above, in a highly integrated system such as the AP, abnormalities in any single element may then cause abnormal function of other (genetically normal) elements. For example, delayed repolarization by either defective inactivation of channels carrying inward current (eg, SCN5A in LQT3) or defective function of a potassium channel carrying outward current (eg, HERG in LQT2) may prolong APs sufficiently to allow normally inactivating currents such as ICa-L to reactivate and cause EADs and/or triggering.50 In addition, it is conceivable that compensatory changes in expression of other genes might come into play in the long-term control of the AP. For example, prolonged APs in other situations (eg, during treatment with AP-prolonging drugs) increase contractility, possibly by increasing intracellular calcium.140 Such increases, were they to be maintained, might not only promote arrhythmogenesis but also influence transcription of a variety of other genes to affect the phenotype observed clinically.
Adrenergic Dependence of LQTS
The adrenergic dependence of arrhythmias in LQTS must now be interpreted in the context of new molecular genetic information. Adrenergic stimulation can modify repolarization and produce arrhythmias in normal subjects and those with LQTS in a number of ways. First, β-adrenergic stimulation directly increases the magnitude of ICa-L, of the pacemaker current If, and of the repolarizing currents IKs and ICl-cAMP. The usual result is an increase in heart rate with AP shortening, presumably reflecting a greater increase in IKs and ICl-cAMP than in ICa-L. A defect in any of these important AP control mechanisms (eg, defective response of IKs or ICl-cAMP or enhanced response of ICa-L) might well account for the clinical feature that some patients with LQTS “fail to shorten” their QT interval appropriately with adrenergic stimulation, eg, during exercise. Alternatively, the response of mutant KVLQT1-, HERG-, or SCN5A-encoded channels to adrenergic stimulation may prove to be abnormal in LQTS. Second, in experimental situations in which cardiac APs are prolonged and EADs (but no triggering) are present, addition of sympathetic stimulation can be sufficient to elicit triggering.141 Thus, patients with LQTS may be particularly vulnerable to sympathetically mediated arrhythmias with even normal levels of sympathetic tone. Third, an increase in APD by a variety of mechanisms increases contractility, possibly by increasing intracellular calcium.140 A number of studies suggest that cells with increased intracellular calcium may be especially susceptible to the development of abnormal automaticity.131 142 143 Finally, it remains conceivable that as-yet-unidentified gene defects in LQTS could involve molecules important for cardiac sympathetic innervation or that the described ion channel defects result in secondary changes in the number or function of such molecules. In addition, abnormalities of distribution of the sympathetic nerves to the heart could contribute to alterations in ion channel function, since a number of studies now indicate that expression of some ion channel genes, including those encoding calcium and potassium channels, can be modulated by sympathetic activity,144 145 and abnormalities in cardiac sympathetic innervation in experimental animals can produce QT prolongation.146 147 148
Current Understanding of the Pathophysiology of LQTS and Unanswered Questions
Cardiac repolarization represents a delicate balance between inward and outward currents during the plateau phase of the cardiac AP. Either persistent inward current (eg, through sodium channels) or reduced outward current (eg, through the channels responsible for IKr) can cause the LQTS phenotype. AP prolongation then results in EADs that can induce the characteristic arrhythmia, torsades de pointes. The extent to which these cellular abnormalities can be attributed directly to changes in INa, IKr, or the channel encoded by KVLQT1 or indirectly to secondary changes in other currents, such as L-type calcium current or sodium-calcium exchange, requires further study, as do the mechanisms that confer sensitivity to adrenergic stimuli. The mechanisms underlying torsades de pointes are uncertain: possibilities include triggered activity arising from EADs or heterogeneity in repolarization (resulting from the presence of EADs in some regions of the heart and not in others) that facilitates reentrant excitation. Either mechanism appears most likely to result from EADs developing primarily in critical regions of the heart, such as the conducting system or M cells. Interestingly, IKr in M cells is similar to that in endocardium and epicardium, but IKs is reduced.149 It is increasingly recognized that ion currents display such regional heterogeneity even under physiological conditions.150 151 152 153 The resultant heterogeneity in APDs may play a key role in determining the ECG manifestations of individual gene defects and in the development of torsades de pointes and other arrhythmias.58
Since not all the molecular defects in LQTS have been identified, it is still conceivable that mutations in critical genes in the sympathetic receptor-effector apparatus may be responsible for adrenergic sensitivity in some subsets. Alternatively, the typical sensitivity to adrenergic stimuli displayed by patients with LQTS may reflect an abnormal response to normal sympathetic activity and innervation. Such an abnormal response to sympathetic activity may be related to increased intracellular calcium, probably a common finding in LQTS.
Many areas of research are opened up by the exciting new molecular genetic findings in LQTS. Examples include studies of the basic biophysics of the normal and mutant KVLQT1, SCN5A, and HERG gene products; studies to identify drug therapies that might correct the defects in these mutations; and studies to determine the relationships between clinical phenotypes and underlying genetic defects.18 26 36 37 38 88 101 The extent to which activation of intracellular signaling systems (as occurs during adrenergic stress) alters the function of these channels is unknown. Similarly, the mechanisms whereby abnormalities in SCN5A or HERG cause sinus bradycardia in LQTS remain to be explored. The HERG gene was originally cloned from a human hippocampal cDNA library, and mRNA transcripts encoding the murine homologue have been reported in extracardiac tissues such as brain and liver.154 It will be interesting to determine the role of HERG at these sites. Since it is now known that defects in sodium current or in IKr can produce abnormally long APs, it is reasonable to ask whether other diseases in which APs are abnormally prolonged (and in which abnormally long APs have been linked to arrhythmogenesis) might have a similar basis. The most obvious example is in patients with drug-associated LQTS: does a subset of such patients carry subclinical mutations in SCN5A or in HERG that are manifest upon only challenge with AP-prolonging drugs? Abnormally long APs have also been linked to arrhythmias in patients with dilated cardiomyopathy155 156 and hypertension.157 It will be interesting to determine the extent of the role of abnormalities in the regulatory or coding regions of KVLQT1, HERG, SCN5A, or other genes whose products are involved in repolarization. It seems likely that long-standing abnormalities in AP control in LQTS result in other changes in cellular homeostasis: for example, is the expression of other channels increased or are the control mechanisms for intracellular calcium abnormal? These questions may be best addressed in animal models in which mutant LQTS genes are expressed instead of or along with normal ones. The suggestion from in vitro studies that different mutations in the same gene may produce a spectrum of clinical phenotypes will need to be investigated. Preliminary data suggest a role for sodium channel blockers such as mexiletine in LQT3, linked to late-opening sodium channels.26 37 In patients with this defect, blockers of the late-opening sodium channels might entirely reverse all the manifestations of the syndrome. Use of sodium channel blockers in patients with other defects might theoretically also shorten the QT interval by reducing inward current during the plateau34 ; however, they would not be expected to reverse the ion channel defect underlying the syndrome. In patients with LQTS due to abnormal potassium channel function, therapies directed at altering the function of the abnormal channels to increase outward current (eg, potassium channel openers) might achieve the latter effect. While these are tantalizing suggestions for future therapy after evaluation in vitro and in controlled clinical trials, β-blockers still remain the mainstay of therapy.
The multiplicity of defects that have already been identified as causing LQTS and the fact that loci remain to be identified and characterized make routine genetic testing for the disease impractical at this time. At this point, presymptomatic diagnosis is possible for family members of those patients in whom the molecular basis of the defect has been characterized. If the identification of specific defects turns out to be important for therapy or for prognosis, then genetic testing may assume an important clinical role.
A major mechanism of action of many drugs that cause torsades de pointes is IKr block. On the basis of the identification of mutations in HERG as a cause of LQTS, one might well ask whether block of IKr is a suitable or desirable target for antiarrhythmic intervention. The effects of drugs that, at clinically useful dosages, reduce IKr by 50% would be predicted to be similar to those of HERG mutations that result in no functional IKr channels and that display no dominant negative effect (eg, Δ1261). Alternatively, drugs that rapidly produce IKr block during the plateau of the AP and whose block equally rapidly dissipates at the end of the plateau might still be antiarrhythmic, particularly in the diseased heart. Compounds that delay sodium channel inactivation have been developed as positive inotropic drugs.158 One mechanism is thought to be increased sodium loading, with resultant increased sodium-calcium exchange. On the basis of the finding that abnormalities in SCN5A cause LQTS, it will be interesting to see whether the beneficial effect of interfering with sodium channel inactivation might be outweighed by its proarrhythmic potential.
AP prolongation is well established in animal models as an effective antiarrhythmic intervention. A major liability is the development of torsades de pointes. One implication of the molecular data now available from studies of LQTS is that block of IKr or interference with inactivation of INa is not likely to be the most desirable strategy to achieve this effect. Rather, the identification of multiple ion channels active during repolarization raises the possibility that targets for drug action can be identified that achieve a desired antiarrhythmic effect without causing torsades de pointes.
The identification of lesions in KVLQT1, HERG, and SCN5A has already resulted in a dramatic increase in our understanding of the basic pathogenesis of LQTS and in the development of lines of research that will have important implications not only for LQTS but also for other disease states in which control of the cardiac AP is an important consideration. The identification of these and other mutations in LQTS therefore represents an exciting beginning to studies in which clinical arrhythmia syndromes will be related to their fundamental molecular and cellular mechanisms.
Selected Abbreviations and Acronyms
|APD||=||action potential duration|
|HERG||=||human ether-a`-go-go–related gene|
|ICl-cAMP||=||cAMP-activated chloride current|
|IKr||=||rapidly activating delayed rectifier potassium current|
|IKs||=||slowly activating delayed rectifier potassium current|
|Ito||=||transient outward current|
This article summarizes the outcome of a workshop sponsored and funded by the Sudden Arrhythmic Death Syndromes (SADS) Foundation, January 7-9, 1995. The need for the workshop was proposed by Peter J. Schwartz, MD (University of Pavia, Italy) and G. Michael Vincent, MD (LDS Hospital, Salt Lake City, Utah); Dr Schwartz acted as chairman. Other participants were Charles Antzelevitch, PhD (Masonic Medical Research Laboratory, Utica, NY); Arthur M. Brown, MD, PhD (MetroHealth Medical Center, Cleveland, Ohio); Thomas J. Colatsky, PhD (Wyeth-Ayerst Research, Princeton, NJ); Richard S. Crampton, MD (University of Virginia, Charlottesville, Va); Robert S. Kass, PhD (University of Rochester, Rochester, NY); Ralph Lazzara, MD (University of Oklahoma, Oklahoma City, Okla); Arthur J. Moss, MD (University of Rochester, Rochester, NY); Dan M. Roden, MD (Vanderbilt University, Nashville, Tenn); Michael R. Rosen, MD (Columbia University, New York City, NY); Michael C. Sanguinetti, PhD (University of Utah, Salt Lake City, Utah); Jeffrey A. Towbin, MD (Baylor College of Medicine, Houston, Tex); and Douglas P. Zipes, MD (University of Indiana, Indianapolis, Ind). Members of the writing committee were Drs Lazzara, Roden, Rosen, Schwartz, and Vincent. The final preparation and organization of the manuscript were the responsibility of Dr Roden.
Reprint requests to G. Michael Vincent, MD, Department of Medicine, LDS Hospital, Eighth Ave and C St, Salt Lake City, UT 84143-0001.
- Received January 29, 1996.
- Revision received April 19, 1996.
- Accepted April 23, 1996.
- Copyright © 1996 by American Heart Association
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