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Molecular Insights into the Local Anesthetic Receptor within Voltage-Gated Sodium Channels Using Hydroxylated Analogs of Mexiletine

By Jean-François Desaphy, Antonella Dipalma, Teresa Costanza, Roberta Carbonara, Maria Maddalena Dinardo, Alessia Catalano, Alessia Carocci, Giovanni Lentini, Carlo Franchini and Diana Conte Camerino

Abstract

We previously showed that the β-adrenoceptor modulators, clenbuterol and propranolol, directly blocked voltage-gated sodium channels, whereas salbutamol and nadolol did not (Desaphy et al., 2003), suggesting the presence of two hydroxyl groups on the aromatic moiety of the drugs as a molecular requisite for impeding sodium channel block. To verify such an hypothesis, we synthesized five new mexiletine analogs by adding one or two hydroxyl groups to the aryloxy moiety of the sodium channel blocker and tested these compounds on hNav1.4 channels expressed in HEK293 cells. Concentration–response relationships were constructed using 25-ms-long depolarizing pulses at −30 mV applied from an holding potential of −120 mV at 0.1 Hz (tonic block) and 10 Hz (use-dependent block) stimulation frequencies. The half-maximum inhibitory concentrations (IC50) were linearly correlated to drug lipophilicity: the less lipophilic the drug, minor was the block. The same compounds were also tested on F1586C and Y1593C hNav1.4 channel mutants, to gain further information on the molecular interactions of mexiletine with its receptor within the sodium channel pore. In particular, replacement of Phe1586 and Tyr1593 by non-aromatic cysteine residues may help in the understanding of the role of π–π or π–cation interactions in mexiletine binding. Alteration of tonic block suggests that the aryloxy moiety of mexiletine may interact either directly or indirectly with Phe1586 in the closed sodium channel to produce low-affinity binding block, and that this interaction depends on the electrostatic potential of the drug aromatic tail. Alteration of use-dependent block suggests that addition of hydroxyl groups to the aryloxy moiety may modify high-affinity binding of the drug amine terminal to Phe1586 through cooperativity between the two pharmacophores, this effect being mainly related to drug lipophilicity. Mutation of Tyr1593 further impaired such cooperativity. In conclusion, these results confirm our former hypothesis by showing that the presence of hydroxyl groups to the aryloxy moiety of mexiletine greatly reduced sodium channel block, and provide molecular insights into the intimate interaction of local anesthetics with their receptor

Topics: Pharmacology
Publisher: Frontiers Research Foundation
OAI identifier: oai:pubmedcentral.nih.gov:3279704
Provided by: PubMed Central

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Citations

  1. (2007). (RS)-hydroxymethylmexiletine, one of the major metabolites of mexiletine.
  2. (1999). A molecular basis for the different local anesthetic affinities of resting versusopenandinactivatedstatesofthe sodiumchannel.Mol.Pharmacol.55,
  3. (2000). A point mutation in domain 4-segment 6 of the skeletal muscle sodium channel produces an atypicalinactivationstate.Biophys.J.
  4. (2008). Access and binding of local anesthetics in the closed sodium channel.
  5. and ConteCamerino,D.(2009).Involvement of voltage-gated sodium channels blockade in the analgesic effects of orphenadrine.
  6. andWang,G.K.(1999).Pointmutations at N434 in D1-S6 of m1 Na+ channels modulate binding affinity and stereoselectivity of local anesthetic enantiomers.
  7. (2005). anesthetic drug binding by voltagegated sodium channels.
  8. (2007). Charge at the lidocaine binding site residues Phe-1759 affects permeation in human cardiac voltagegated sodium channels.
  9. (2003). Closing and inactivation potentiate the cocaethylene inhibition of cardiac sodium channels by distinct mechanisms.
  10. (1996). Common molecular determinants of local anesthetic, antiarrhythmic, andanticonvulsantblockof voltagegated sodium channels.
  11. (2003). Different ability of clenbuterol and salbutamol to block sodium channels predicts their therapeutic use in muscle excitability disorders.
  12. (2004). Different flecainide sensitivity of hNav1.4 channels and myotonic mutants explained by state-dependent block.
  13. (2008). Electrostatic contributions of aromatic residues in the local anesthetic receptor of voltage-gated sodium channels.
  14. (2006). Evaluation of the pharmacological activity of the major mexiletine metabolites on skeletal muscle sodium currents.
  15. (2001). Gatingof myotonicNachannelmutants defines the response to mexiletine and a potent derivative.
  16. (2010). Hydroxylated analogs of mexiletine as tools for structural-requirements investigation of the sodium channel blocking activity.
  17. (2003). Inhibition of skeletal muscle sodium currents by mexiletine analogues: specific hydrophobic interactions rather than lipophilia per se accountfordrugtherapeuticprofile.
  18. (2007). Ion channel pharmacology.
  19. (1977). Local anesthetics: hydrophilic and hydrophobic pathways for the drug-receptor reaction.
  20. (1998). Lysine point mutat i o n si nN a + channel D4-S6 reduce inactivated channel block by local anesthetics.
  21. (2011). Molecular basis for class Ib anti-arrhythmic inhibition of cardiac sodium channels.
  22. (2000). Molecular determinants of mexiletine structure for potent and use-dependent block of skeletal muscle sodium channels.
  23. (2010). Molecular determinants of state-dependent blockof voltage-gatedsodiumchannelsbypilsicainide.Br.J.Pharmacol.
  24. (1994). Molecular determinants of statedependent block of Na+ channels by local anesthetics.
  25. (2001). Molecular determinants of voltagedependent gating and binding of pore-blocking drugs
  26. (2012). Molecular insights into the local anesthetic receptor within voltage-gated sodium channels using hydroxylated analogs of mexiletine.
  27. (2004). New potent mexiletine and tocainide analogues evaluated in vivo and in vitro as antimyotonic agents on myotonic ADR mouse.
  28. (2001). Point mutations in α-subunit of human cardiac Na+ channels alter Na+ current kinetics.
  29. (2000). Residues in Na+ channel D3-S6 segment modulate both batrachotoxin and local anesthetic affinities.
  30. (1995). Revealing the architecture of a K+ channel pore through mutant cycles withapeptideinhibitor.Science 268,
  31. (2002). Role of amino acid residues in transmembrane segments IS6 and IIS6 of the Na+ channel α subunit in voltage-dependent gating and drug block.
  32. (2010). Sodium channel molecular conformations and antiarrhythmic drug affinity. Trends Cardiovasc.
  33. (2011). Statement: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
  34. (2001). Structural and gating changes of the sodium channel induced by mutation of a residue in the upper third of IVS6, creating an external access path for local anesthetics.
  35. (2005). Systemic administration of local anesthetic agents to relieve neuropathic pain. Cochrane Database Syst.
  36. (2011). The sodium channel as a target of local anesthetic drugs.
  37. (2012). This is an open-access article distributed under the terms of the Creative Commons Attribution Non Commercial License, which permits noncommercial use, distribution, and reproduction in other forums, provided the original authors and source are credited.
  38. (2004). Tortorella,V.,De Luca,A.,De Bellis,M., Desaphy,J.-F.,and Conte Camerino,
  39. (2009). Using lidocaine and benzocaine to link sodium channel molecular conformations to state-dependent antiarrhythmicdrugaffinity.Circ.Res.105,
  40. V.,andDeLuca,A.(1999).Increased hindranceonthechiralcarbonatom of mexiletine enhances the block of ratskeletalmuscleNa+channelsina modelof myotoniainducedbyATX.

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