Toxicologic/transport properties of NCS-382, a γ-hydroxybutyrate (GHB) receptor ligand, in neuronal and epithelial cells: Therapeutic implications for SSADH deficiency, a GABA metabolic disorder

Toxicol In Vitro. 2018 Feb;46:203-212. doi: 10.1016/j.tiv.2017.10.015. Epub 2017 Oct 12.

Abstract

We report the in vitro assessment of pharmacotoxicity for the high-affinity GHB receptor ligand, NCS-382, using neuronal stem cells derived from mice with a targeted deletion of the aldehyde dehydrogenase 5a1 gene (succinic semialdehyde dehydrogenase(SSADH)-deficient mice). These animals represent a phenocopy of the human disorder of GABA metabolism, SSADH deficiency, that metabolically features accumulation of both GABA and the GABA-analog γ-hydroxybutyric acid in conjunction with a nonspecific neurological phenotype. We demonstrate for the first time using MDCK cells that NCS-382 is actively transported and capable of inhibiting GHB transport. Following these in vitro assays with in vivo studies in aldh5a1-/- mice, we found the ratio of brain/liver GHB to be unaffected by chronic NCS-382 administration (300mg/kg; 7 consecutive days). Employing a variety of cellular parameters (reactive oxygen and superoxide species, ATP production and decay, mitochondrial and lysosomal number, cellular viability and necrosis), we demonstrate that up to 1mM NCS-382 shows minimal evidence of pharmacotoxicity. As well, studies at the molecular level indicate that the effects of NCS-382 at 0.5mM are minimally toxic as evaluated using gene expression assay. The cumulative data provides increasing confidence that NCS-382 could eventually be considered in the therapeutic armament for heritable SSADH deficiency.

Keywords: GABA metabolism; NCS-382; Neuronal stem cells; SSADH deficiency (SSADHD); Succinic semialdehyde dehydrogenase (SSADH); γ-Hydroxybutyric acid (GHB).

MeSH terms

  • Amino Acid Metabolism, Inborn Errors
  • Animals
  • Anticonvulsants / metabolism
  • Anticonvulsants / toxicity
  • Benzocycloheptenes / metabolism*
  • Benzocycloheptenes / toxicity*
  • Biomarkers
  • Cell Survival
  • Developmental Disabilities
  • Epithelial Cells
  • Gene Expression Regulation / drug effects
  • Genotype
  • Humans
  • Mice
  • Mice, Knockout
  • Mitochondria / metabolism
  • Neural Stem Cells / metabolism
  • Neurons
  • Reactive Oxygen Species / metabolism
  • Receptors, Cell Surface
  • Succinate-Semialdehyde Dehydrogenase / deficiency
  • Superoxides / metabolism

Substances

  • 4-hydroxybutyric acid receptor
  • Anticonvulsants
  • Benzocycloheptenes
  • Biomarkers
  • Reactive Oxygen Species
  • Receptors, Cell Surface
  • Superoxides
  • NCS 382
  • Aldh5a1 protein, mouse
  • Succinate-Semialdehyde Dehydrogenase

Supplementary concepts

  • succinic semialdehyde dehydrogenase deficiency