Inhibitory effect of eslicarbazepine acetate and S-licarbazepine on Na_v1.5 channels

Abstract

11Eslicarbazepine acetate (ESL) is a dibenzazepine anticonvulsant approved as adjunctive treatment for 12 partial-onset epileptic seizures. Following first pass hydrolysis of ESL, S-licarbazepine (S-Lic) 13represents around 95 % of circulating active metabolites. S-Lic is the main enantiomer responsible 14for anticonvulsant activity and this is proposed to be through the blockade of voltage-gated Na + 15 channels (VGSCs). ESL and S-Lic both have a voltage-dependent inhibitory effect on the Na + current 16 in N1E-115 neuroblastoma cells expressing neuronal VGSC subtypes including Nav1.1, Nav1.2, 17Nav1.3, Nav1.6 and Nav1.7. ESL has not been associated with cardiotoxicity in healthy volunteers, 18although a prolongation of the electrocardiographic PR interval has been observed, suggesting that 19ESL may also inhibit cardiac Nav1.5 isoform. However, this has not previously been studied. Here, 20we investigated the electrophysiological effects of ESL and S-Lic on Nav1.5 using whole-cell patch 21 clamp recording. We interrogated two model systems: (1) MDA-MB-231 metastatic breast 22 carcinoma cells, which endogenously express the 'neonatal' Nav1.5 splice variant, and (2) HEK-293 23 cells stably over-expressing the 'adult' Nav1.5 splice variant. We show that both ESL and S-Lic 24 inhibit transient and persistent Na + current, hyperpolarise the voltage-dependence of slow 25 inactivation, and slow the recovery from channel inactivation. These findings highlight, for the first 26 time, the potent inhibitory effects of ESL and S-Lic on the Nav1.5 isoform, suggesting a possible 27 explanation for the prolonged PR interval observed in patients on ESL treatment. Given that 28 numerous cancer cells have also been shown to express Nav1.5, and that VGSCs potentiate invasion 29 and metastasis, this study also paves the way for future investigations into ESL and S-Lic as potential 30 invasion inhibitors. 31

A novel Nav1.5-dependent feedback mechanism driving glycolytic acidification in breast cancer metastasis

“…The whole-cell patch clamp technique was used to record cell membrane currents from cells grown on glass coverslips (29). Series resistance was compensated by 40-60% and linear components of leak were subtracted using a P/6 protocol (30).…”

“…Filamented borosilicate capillary tubes were pulled and fire-polished to a resistance of ~5 MΩ for recording from cell lines and ~10 MΩ for recording from primary cells. The extracellular physiological saline solution (PSS) contained (in mM) NaCl 144, KCl 5.4, MgCl 2 1, CaCl 2 2.5, HEPES 5, D-glucose 5.6, and was adjusted to pH 7.2 (unless otherwise stated) using NaOH.The intracellular recording solution for measuring Na + currents contained (in mM) NaCl 5, CsCl 145, MgCl 2 2, CaCl 2 1, HEPES 10, EGTA 11 and was adjusted to pH 7.4 (unless otherwise stated) using CsOH(29). The intracellular recording solution for measuring K + currents contained (in mM) NaCl 5, KCl 145, MgCl 2 2, CaCl 2 1, HEPES 10, EGTA 11 and was adjusted to pH 7.4 using KOH.…”

“…For detection of small currents in primary cells, patient tumour slices and a panel of cell lines, signals were post-filtered at 1 kHz. For examination of pH dependency in MDA-MB-231 cells, currents were noise-corrected by subtracting half the peak-to-peak noise measured during the 10 ms period before depolarisation(29).…”

Sodium accumulation in breast cancer predicts malignancy and treatment response

“…We therefore investigated whether inhibition of either NHE1 (cariporide) or VGSCs (eslicarbazepine acetate, ESL) affected elevated [Na + ] within MDA-MB-231 xenograft tumours. ESL and its major active metabolite (S)-licarbazepine 49 inhibit inward Na + current in MDA-MB-231 cells 50 , and was selected due to its preferred use in the clinic compared to older VGSC-inhibiting antiepileptic medications such as phenytoin 51 . Moreover, cariporide was administered at a dose that previously exhibited efficacy in in vivo models of cancer 32,33,34 .…”