Mechanism of sodium channel block by local anesthetics, antiarrhythmics, and anticonvulsants

Abstract

Local anesthetics, antiarrhythmics, and anticonvulsants include both charged and electroneutral compounds that block voltage-gated sodium channels. Prior studies have revealed a common drug-binding region within the pore, but details about the binding sites and mechanism of block remain unclear. Here, we use the x-ray structure of a prokaryotic sodium channel, NavMs, to model a eukaryotic channel and dock representative ligands. These include lidocaine, QX-314, cocaine, quinidine, lamotrigine, carbamazepine (CMZ), phenytoin, lacosamide, sipatrigine, and bisphenol A. Preliminary calculations demonstrated that a sodium ion near the selectivity filter attracts electroneutral CMZ but repels cationic lidocaine. Therefore, we further docked electroneutral and cationic drugs with and without a sodium ion, respectively. In our models, all the drugs interact with a phenylalanine in helix IVS6. Electroneutral drugs trap a sodium ion in the proximity of the selectivity filter, and this same site attracts the charged group of cationic ligands. At this position, even small drugs can block the permeation pathway by an electrostatic or steric mechanism. Our study proposes a common pharmacophore for these diverse drugs. It includes a cationic moiety and an aromatic moiety, which are usually linked by four bonds.

Figure 1.

Ligand-sensing residues in the inner-pore region of Nav1.x channels. (A) Ligands considered in this study. (B) Sequences alignment of NavMs and Nav1.4. Key positions where mutations affect action of inner pore blockers (Mike and Lukacs, 2010) are highlighted. (C) The NavMs structure. Shown are C^α-C^β bonds in key ligand-sensing residues and Na_III. Repeats I, III, and IV are blue, green, and orange, respectively. Repeat II is not depicted for clarity. Red ovals mark approximate borders of the selectivity filter region (1), the inner pore region (2), the activation gate region (3), and two of the four repeat interfaces (4).

Figure 2.

Hands-free docking of lidocaine and CMZ in the Nav1.4 model with and without Na_III (yellow spheres). The oxygen atom of CMZ and the protonated nitrogen of lidocaine are shown as red and blue spheres, respectively. Distribution of ligands, as indicated by their key atoms, strongly depends on the presence of Na_III. In the presence of Na_III, low-energy positions of lidocaine and CMZ are scattered and clustered, respectively, and in the absence or presence of Na_III, the opposite distribution is observed. These results suggest that the Na_III site is a hotspot for ligands.

Figure 3.

Statistics of lidocaine-binding modes in the channel model without Na_III. Each binding mode is characterized by the Z-coordinates of the ammonium nitrogen, Z_N+, and the carbon atom in para-position of the aromatic ring, Z_Cp, and deviations of these atoms from the Z-axis, D_N+ and D_Cp. (A–D) Distributions of the characteristics of the ensemble of low-energy binding modes. (E–H) Correlation fields between the characteristics. Dots in E, which are close to the dashed line (Z_Cp = Z_N), correspond to horizontal binding modes of lidocaine. All coordinates and distances are given if angstroms.

Figure 4.

Examples of low-energy binding modes of lidocaine and CMZ. Side chains of F⁴ⁱ¹⁵ and Y⁴ⁱ²² are shown as sticks. Horizontal lines in B–G show levels of site Na_III and putative site Na_IV. (A) Location of ligand-binding region. (B–G) Representative structures from the ensembles of low-energy binding modes. (B) Horizontal binding mode of lidocaine. The aromatic group protrudes into the III/IV interface. (C and D) Vertical binding modes with the ammonium group at the levels of Na_III site and Na_IV site, respectively. (E) CMZ without tight contacts with Na_III. (F and G) CMZ bound to Na_III. The sodium ion is bound at its innate site seen in the NavMs structure (F) or shifted toward putative site Na_IV (G). Note that interactions of lidocaine and CMZ with F⁴ⁱ¹⁵ and Y⁴ⁱ²² depend on the ligand-binding modes.

Figure 5.

Statistics of CMZ and Na⁺ binding in the channel model. Each binding mode is characterized by the Z-coordinates of carbonyl oxygen, Z_O, and sodium ion, Z_Na, as well as deviations of these atoms from the Z-axis, D_O and D_Na. (A–E) Distributions of the characteristics of the ensemble of low-energy binding modes. (F–H) Correlation fields between the characteristics. All coordinates and distances are given in angstroms.

Figure 6.

Cationic and sodium-bound electroneutral ligands in the channel. (A) The triethylammonium group of QX-314 is close to the Na_III site, the aromatic ring binds between F⁴ⁱ¹⁵ and Y⁴ⁱ²², and the ligand also interacts with Q^1p49 and F^3p49 in agreement with mutational data (Yamagishi et al., 2009). (B) The bulky moiety of quinidine fits the hotspot at the Na_III site, whereas the hydroxyl group donates an H bond to ^3p48O=C. (C and D) Two binding modes of cocaine. In one binding mode (C), the cocaine amino group binds at the Na_III site. In another binding mode (D), the amino group binds close to the putative Na_IV site and forms cation-π contacts with F⁴ⁱ¹⁵, whereas the opposite end of cocaine reaches I³ⁱ²³. (E and F) The long sipatrigine molecule can bind in the horizontal (E) and vertical (F) modes, forming many contacts with the channel. In the vertical mode, sipatrigine extends from the Na_III site to the hydrophobic region at levels i22-i23. (G and H) Side and extracellular views of lamotrigine. In the side view (G), repeat II is removed for clarity. In the extracellular view (H), lamotrigine and backbone carbonyls ^p48O=C are space filled. Lamotrigine binds to Na_III, which is coordinated by ^3p48O=C and ^4p48O=C, donates an H bond to ^2p48O=C, interacts with ^1p48O=C, π-stacks with F⁴ⁱ¹⁵, and approaches Y⁴ⁱ²². (I and J) Side and extracellular views at two similar binding modes of bisphenol A. Both aromatic rings are involved in π-cation interactions with Na_III, two hydroxyls donate H bonds to carbonyls ^p48O=C, and methyl groups bind between F⁴ⁱ¹⁵ and Y⁴ⁱ²². (K and L) Side views of lacosamide and phenytoin. Both ligands interact with Na_III and form H bonds with Q^1p49, whereas their aromatic rings bind between F⁴ⁱ¹⁵ and Y⁴ⁱ²².

Figure 7.

Similarity of binding modes of charged and neutral blockers. (A) Cytoplasmic and side views at superposition of the channel-bound ligands. The charged nitrogens and ligand-bound sodium ions are blue and yellow, respectively. The carbon atoms at the opposite end of the ligands are gray. Note the clustering of blue/yellow atoms at the Na_III site and gray atoms at F⁴ⁱ¹⁵. The gray outlier belongs to bisphenol A, which lacks an aromatic ring protruding into the pore. (B) Scheme of ligand–channel interactions. The charged nitrogen or ligand-bound sodium ion interacts with the ring of carbonyls ^p48O=C (Na_III site) indicated by the red rectangle. The aromatic groups interact with hydrophobic residues in the inner pore (gray rectangle), among which F⁴ⁱ¹⁵ plays the major role. (C) Lidocaine and sodium-bound CMZ have a common pharmacophore.