Inhibition of N-type calcium channels by fluorophenoxyanilide derivatives.

Ellen Gleeson,Richard Lewis,Kellie Tuck,Irina Vetter,Janease Graham,Peter Duggan,Sandro Spiller

doi:10.3390/md13042030

Abstract

A set of fluorophenoxyanilides, designed to be simplified analogues of previously reported ω-conotoxin GVIA mimetics, were prepared and tested for N-type calcium channel inhibition in a SH-SY5Y neuroblastoma FLIPR assay. N-type or Cav2.2 channel is a validated target for the treatment of refractory chronic pain. Despite being significantly less complex than the originally designed mimetics, up to a seven-fold improvement in activity was observed.

Highlights

Neuropathic pain, which results from nerve damage caused by surgery, trauma, infection or disease, often does not respond to existing therapies [1,2]
It has been shown that SH-SY5Y neuroblastoma cells endogenously expressing human Cav2.2 channels allow the rapid screening of potential channel blockers by means of a FLIPR assay [39]
SH-SY5Y cells were plated at a density of 30,000–50,000 cells/well on 384-well black-walled imaging plates and loaded for 30 min at 37 °C with Calcium 4 no-wash dye (Molecular Devices, Sunnyvale, CA, USA) diluted in physiological salt solution (PSS; composition: 140 mM NaCl, 11.5 mM glucose, 5.9 mM KCl, 1.4 mM MgCl2, 1.2 mM NaH2PO4, 5 mM NaHCO3, 1.8 mM CaCl2, 10 mM 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES), pH 7.4)

Summary

Introduction

Neuropathic pain, which results from nerve damage caused by surgery, trauma, infection or disease, often does not respond to existing therapies [1,2]. N-Type calcium channels (Cav2.2 channels) are strongly implicated in chronic and neuropathic pain and their inhibitors have been widely pursued [1,2,3,4] This approach has been best validated by Ziconotide (Prialt®), a synthetic version of the peptide ω-conotoxin MVIIA found in the venom of a fish-hunting marine cone snail Conus magnus. The most potent fluorinated analogue in this series was found to be the diguanidino compound (3) while the corresponding diamino compound (4) had comparatively weak activity This is consistent with previous results where truncation of the arginine side chain mimic to an amine in ω-conotoxin GVIA mimetics is typically detrimental to activity at the N-type channel [25,26,27,28,29]. The amino analogues and their corresponding monoguanidino analogues that were synthesised and tested in this study are shown in Figure 3, compounds 5a–c and 6a–c

Chemistry

Biology

General Experimental Procedures

Synthesis

Conclusions