2. Academic Validation
  3. Evaluation of the Spider ( Phlogiellus genus) Phlotoxin 1 and Synthetic Variants as Antinociceptive Drug Candidates

Evaluation of the Spider ( Phlogiellus genus) Phlotoxin 1 and Synthetic Variants as Antinociceptive Drug Candidates

  • Toxins (Basel). 2019 Aug 22;11(9):484. doi: 10.3390/toxins11090484.
Tânia C Gonçalves 1 2,  Pierre Lesport 3,  Sarah Kuylle 1,  Enrico Stura 1,  Justyna Ciolek 1,  Gilles Mourier 1,  Denis Servent 1,  Emmanuel Bourinet 3,  Evelyne Benoit 4 5,  Nicolas Gilles 6
Affiliations

Affiliations

  • 1 Service d'Ingénierie Moléculaire des Protéines (SIMOPRO), CEA, Université Paris-Saclay, F-91191 Gif sur Yvette, France.
  • 2 Sanofi R&D, Integrated Drug Discovery - High Content Biology, F-94440 Vitry-sur-Seine, France.
  • 3 Institut de Génomique Fonctionnelle (IGF), CNRS-UMR 5203, Inserm-U661, Université de Montpellier, Laboratories of Excellence - Ion Channel Science and Therapeutics, F-34094 Montpellier, France.
  • 4 Service d'Ingénierie Moléculaire des Protéines (SIMOPRO), CEA, Université Paris-Saclay, F-91191 Gif sur Yvette, France. [email protected].
  • 5 Institut des Neurosciences Paris-Saclay (Neuro-PSI), UMR CNRS/Université Paris-Sud 9197, Université Paris-Saclay, F-91198 Gif sur Yvette, France. [email protected].
  • 6 Service d'Ingénierie Moléculaire des Protéines (SIMOPRO), CEA, Université Paris-Saclay, F-91191 Gif sur Yvette, France. [email protected].
PMID: 31443554 DOI: 10.3390/toxins11090484
Abstract

Over the two last decades, venom toxins have been explored as alternatives to opioids to treat chronic debilitating pain. At present, approximately 20 potential analgesic toxins, mainly from spider venoms, are known to inhibit with high affinity the NaV1.7 subtype of voltage-gated sodium (NaV) channels, the most promising genetically validated antinociceptive target identified so far. The present study aimed to consolidate the development of phlotoxin 1 (PhlTx1), a 34-amino acid and 3-disulfide bridge peptide of a Phlogiellus genus spider, as an antinociceptive agent by improving its affinity and selectivity for the human (h) NaV1.7 subtype. The synthetic homologue of PhlTx1 was generated and equilibrated between two conformers on reverse-phase liquid chromatography and exhibited potent analgesic effects in a mouse model of NaV1.7-mediated pain. The effects of PhlTx1 and 8 successfully synthetized alanine-substituted variants were studied (by automated whole-cell patch-clamp electrophysiology) on cell lines stably overexpressing hNaV subtypes, as well as two cardiac targets, the hCaV1.2 and hKV11.1 subtypes of voltage-gated calcium (CaV) and potassium (KV) channels, respectively. PhlTx1 and D7A-PhlTx1 were shown to inhibit hNaV1.1-1.3 and 1.5-1.7 subtypes at hundred nanomolar concentrations, while their affinities for hNaV1.4 and 1.8, hCaV1.2 and hKV11.1 subtypes were over micromolar concentrations. Despite similar analgesic effects in the mouse model of NaV1.7-mediated pain and selectivity profiles, the affinity of D7A-PhlTx1 for the NaV1.7 subtype was at least five times higher than that of the wild-type peptide. Computational modelling was performed to deduce the 3D-structure of PhlTx1 and to suggest the Amino acids involved in the efficiency of the molecule. In conclusion, the present structure-activity relationship study of PhlTx1 results in a low improved affinity of the molecule for the NaV1.7 subtype, but without any marked change in the molecule selectivity against the other studied ion channel subtypes. Further experiments are therefore necessary before considering the development of PhlTx1 or synthetic variants as antinociceptive drug candidates.

Keywords

NaV1.7 channel subtype; Phlogiellus spider; human voltage-gated ion channel subtypes; mouse model of NaV1.7-mediated pain; phlotoxin 1.

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