The Basis for Low-affinity hERG Potassium Channel Block by Sotalol

The human Ether-à-go-go-Related Gene (hERG) encodes potassium channels mediating the rapid delayed-rectifier K+ current, IKr, which is crucial for normal repolarization of the ventricles of the heart. hERG is established to be a pharmacological target for Class Ia and Class III antiarrhythmic drugs and for numerous noncardiac drugs associated with acquired long QT syndrome and torsades de pointes arrhythmia. Due to the pharmacological promiscuity of hERG channels, all novel pharmaceuticals must be tested for their propensity to inhibit hERG channel ionic current. The archetypal high-affinity hERG inhibitors come from the methanesulphonanilide drug family. This family includes dofetilide (Tikosyn), a drug licensed in the USA for the treatment of supraventricular arrhythmias and an experimental Class III drug, E-4031. Sotalol, which is used to treat both supraventricular arrhythmias and serious ventricular arrhythmias in structurally normal hearts, differs from other methanesulphonanilides in that it binds with low affinity to hERG/IKr channels. For example, a study of displacement of tritiated dofetilide by E-4031, dofetilide, and D-sotalol from guinea-pig ventricular myocytes yielded respective Ki values of 38 nM, 47 nM, and 100 μM. The structural basis for sotalol’s low-affinity hERG/IKr block has not yet been established. Binding determinants of E-4031 and dofetilide, as well as another high-affinity methanesulphonanilide, MK499, have been mapped to the S6- and pore-helices that form the inner cavity of the hERG channel; mutation of S6 aromatic residues (Y652 and F656) and of residues at the base of the pore helix (T623, S624, V625) markedly impaired the ability of these drugs to inhibit hERG current (IhERG). Comparable information is not currently available for sotalol, perhaps in part due to difficulties in obtaining pure D-sotalol (which lacks the marked β-adrenoceptor-blocking properties of the racemic mixture) and in part because the low potency of the drug makes it difficult to study at concentrations required to produce profound IhERG inhibition. Comparison of the structures of E-4031 and dofetilide with sotalol shows that sotalol is a smaller molecule than the other drugs. We have previously compared structurally similar hERG-blocking drugs of different sizes (ranolazine and lidocaine) and found that, despite structural similarities, the smaller drug (lidocaine) was less well able to interact with hERG pore side chains than the larger molecule. We hypothesized that a similar explanation accounts for relatively low-affinity IhERG block by sotalol. Accordingly, using a previously validated, MthK-based hERG pore model, we have docked D-sotalol in the hERG channel inner cavity. Low-energy-score binding poses show an ability for sotalol to interact with the aromatic canonical drug-binding residues Y652 and F656. The charged secondary aliphatic amino group and aromatic moiety of D-sotalol can make cation-π and π-stacking interactions, respectively, with the side chains of F656 or Y652. In these poses, D-sotalol does not make simultaneous interactions with pore-helical residues (T623, S624). This contrasts markedly with the situation for high-affinity methanesulphonanilides. To test this notion experimentally, we have performed experiments in which a D-sotalol concentration producing 50%–60% inhibition of wild-type (WT) channels was tested against alanine mutants of the S6 aromatic residue Y652 and pore-helical residue S624, expressed in HEK 293 cells. Recordings were made at physiological temperature using previously described conditions and protocols.