title: Modulation of the conformational state of the SV2A protein by an allosteric mechanism as evidenced by ligand binding assays.
abstract: OBJECTIVE
  Synaptic vesicle protein 2A (SV2A) is the specific binding site of the anti-epileptic drug levetiracetam (LEV) and its higher affinity analogue UCB30889. Moreover, the protein has been well validated as a target for anticonvulsant therapy. Here, we report the identification of UCB1244283 acting as a SV2A positive allosteric modulator of UCB30889.
  
  
  METHODS
  UCB1244283 was characterized in vitro using radioligand binding assays with [(3)H]UCB30889 on recombinant SV2A expressed in HEK cells and on rat cortex. In vivo, the compound was tested in sound-sensitive mice.
  
  
  RESULTS
  Saturation binding experiments in the presence of UCB1244283 demonstrated a fivefold increase in the affinity of [(3)H]UCB30889 for human recombinant SV2A, combined with a twofold increase of the total number of binding sites. Similar results were obtained on rat cortex. In competition binding experiments, UCB1244283 potentiated the affinity of UCB30889 while the affinity of LEV remained unchanged. UCB1244283 significantly slowed down both the association and dissociation kinetics of [(3)H]UCB30889. Following i.c.v. administration in sound-sensitive mice, UCB1244283 showed a clear protective effect against both tonic and clonic convulsions.
  
  
  CONCLUSIONS
  These results indicate that UCB1244283 can modulate the conformation of SV2A, thereby inducing a higher affinity state for UCB30889. Our results also suggest that the conformation of SV2A per se might be an important determinant of its functioning, especially during epileptic seizures. Therefore, agents that act on the conformation of SV2A might hold great potential in the search for new SV2A-based anticonvulsant therapies.
authors.affiliation: NewMedicines, CNS Discovery Research, UCB Pharma, Braine-l'Alleud, Belgium.
authors.forename: V
authors.initials: V
authors.lastname: Daniels
authors.forename: M
authors.initials: M
authors.lastname: Wood
authors.forename: K
authors.initials: K
authors.lastname: Leclercq
authors.forename: R M
authors.initials: RM
authors.lastname: Kaminski
authors.forename: M
authors.initials: M
authors.lastname: Gillard
mesh_terms: D000494:Allosteric Regulation; D000813:Anilides / Q000494:pharmacology* / Q000627:therapeutic use; D000818:Animals; D000927:Anticonvulsants / Q000494:pharmacology* / Q000627:therapeutic use; D001386:Azides / Q000494:pharmacology*; D002540:Cerebral Cortex / Q000378:metabolism; D004827:Epilepsy; D005260:Female; D057809:HEK293 Cells; D006801:Humans; D008024:Ligands; D008297:Male; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000378:metabolism; D008811:Mice, Inbred DBA; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D011759:Pyrrolidines / Q000494:pharmacology*; D011869:Radioligand Assay; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000737:chemistry* / Q000378:metabolism
chemical_list: C487592:(2S)-2-(4-(3-azidophenyl)-2-oxopyrrolidin-1-yl)butanamide; C000596274:4-(3,5-dimethylphenyl)-N-(2-methoxyphenyl)-3-methylbutanamide; D000813:Anilides; D000927:Anticonvulsants; D001386:Azides; D008024:Ligands; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; D011759:Pyrrolidines; D011994:Recombinant Proteins; C489618:Sv2a protein, rat; C076414:SV2A protein, human