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

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.

Determination of system level alterations in host transcriptome due to Zika virus (ZIKV) Infection in retinal pigment epithelium

“…Among the key regulators downregulated by ZIKV was ALDH5A1, the gene encoding a mitochondrial NAD(+) -dependent succinic semialdehyde dehydrogenase (SSADH) enzyme. In humans, SSADH deficiency is a rare autosomal recessive metabolic disorder affecting γ-aminobutyric acid degradation and is characterized by developmental delay, cognitive impairment, expressive language deficit, and mild ataxia 48 . Epilepsy is present in about half of affected individuals 49 .…”

Succinic semialdehyde dehydrogenase deficiency, a disorder of GABA metabolism: an update on pharmacological and enzyme‐replacement therapeutic strategies

“…Gene expression studies using NCS-382 in HepG2 cells revealed only a minor number of genes (of 370 tested) showing dysregulation (Table 1). Additionally, high-dose NCS-382 demonstrated only minimal pharmacotoxicity in neural stem cells (NSCs, or neuronal progenitor cells) derived from aldh5a1 −/− mice (e.g., aldh5a1 −/− NSCs) (Vogel et al, 2017e). These cells were developed as an in vitro model of SSADHD, showing increased GHB content in culture medium, enhanced biomarkers of oxidative stress and increased mitochondrial number and highlighting the utility of NSCs as a useful preclinical screening tool for evaluating therapeutics for SSADHD (Vogel et al, 2017f).…”

“…Our objective here was to investigate the potential of NCS-382 to block uptake of GHB. We demonstrated for the first time using Madin-Darby Canine Kidney (MDCK) cells that NCS-382 is actively transported and capable of inhibiting GHB transport (Vogel et al, 2017e). 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), which appeared paradoxical.…”

Transcranial Magnetic Stimulation in Succinic Semialdehyde Dehydrogenase Deficiency: A Measure of Maturational Trajectory of Cortical Excitability

“…Further studies using murine SSADHD models to correlate transcranial magnetic stimulation data with immunohistochemical and electrophysiologic data will be useful to better elucidate this inconsistency. 5,31,32…”