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Kara R Vogel - One of the best experts on this subject based on the ideXlab platform.
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succinic semialdehyde dehydrogenase deficiency a disorder of gaba metabolism an update on pharmacological and enzyme replacement therapeutic strategies
Journal of Inherited Metabolic Disease, 2018Co-Authors: Kara R Vogel, Garrett R Ainslie, Alice Mcconnell, Jeanbaptiste Roullet, Dana C Walters, Sameer C Dhamne, Alexander Rotenberg, Michael K GibsonAbstract:We present an update to the status of research on succinic semialdehyde dehydrogenase (SSADH) deficiency (SSADHD), a rare disorder of GABA metabolism. This is an unusual disorder featuring the accumulation of both GABA and its neuromodulatory analog, gamma-hydroxybutyric acid (GHB), and recent studies have advanced the potential clinical application of NCS-382, a putative GHB Receptor antagonist. Animal studies have provided proof-of-concept that enzyme replacement therapy could represent a long-term therapeutic option. The characterization of neuronal stem cells (NSCs) derived from aldehyde dehydrogenase 5a1−/− (aldh5a1−/−) mice, the murine model of SSADHD, has highlighted NSC utility as an in vitro system in which to study therapeutics and associated toxicological properties. Gene expression analyses have revealed that transcripts encoding GABAA Receptors are down-regulated and may remain largely immature in aldh5a1−/− brain, characterized by excitatory as opposed to inhibitory outputs, the latter being the expected action in the mature central nervous system. This indicates that agents altering chloride channel activity may be therapeutically relevant in SSADHD. The most recent therapeutic prospects include mTOR (mechanistic target of rapamycin) inhibitors, drugs that have received attention with the elucidation of the effects of elevated GABA on autophagy. The outlook for novel therapeutic trials in SSADHD continues to improve.
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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
Toxicology in Vitro, 2018Co-Authors: Kara R Vogel, Garrett R Ainslie, Alice Mcconnell, Jeanbaptiste Roullet, K M GibsonAbstract: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 (300 mg/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 1 mM NCS-382 shows minimal evidence of pharmacotoxicity. As well, studies at the molecular level indicate that the effects of NCS-382 at 0.5 mM 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.
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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
Toxicology in Vitro, 2017Co-Authors: Kara R Vogel, Garrett R Ainslie, Alice Mcconnell, Jeanbaptiste Roullet, K M GibsonAbstract: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 (300 mg/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 1 mM NCS-382 shows minimal evidence of pharmacotoxicity. As well, studies at the molecular level indicate that the effects of NCS-382 at 0.5 mM 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.
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In vitro toxicological evaluation of NCS-382, a high-affinity antagonist of γ-hydroxybutyrate (GHB) binding.
Toxicology in Vitro, 2017Co-Authors: Kara R Vogel, Garrett R Ainslie, Alice Mcconnell, Jeanbaptiste Roullet, K M GibsonAbstract:Abstract γ-Hydroxybutyric acid (GHB), a minor metabolite of the inhibitory neurotransmitter GABA, can accumulate to significant concentrations in the heritable disorder of GABA degradation, succinic semialdehyde dehydrogenase (SSADH) deficiency (SSADHD). Moreover, GHB may be employed in therapeutic settings (treatment of narcolepsy), as well as instances of illicit activity, including acquaintance sexual assault and the induction of euphoria. High-affinity binding sites for GHB in the brain have been identified, although the absolute identity of these Receptors remains unclear. Pharmacological antagonism of GHB binding may have multiple instances of therapeutic relevance. The high affinity GHB Receptor antagonist, NCS-382 (6,7,8,9-tetrahydro-5-hydroxy-5H-benzo-cyclohept-6-ylideneacetic acid) has not been piloted in humans. To address the potential clinical utility of NCS-382, we have piloted initial studies of its toxicology in HepG2 and primary hepatocyte cells. At high dose (0.5 mM), NCS-382 showed no capacity for inhibition of microsomal CYPs (CYP1A2, 2B6, 2C8, 2C9, 2C19, 2D6 and 3A4) and minimal potential for activation of xenobiotic nuclear Receptors. Additional cellular integrity and functional assays (viability, oxidative stress, apoptosis, ATP production) revealed little evidence for cytotoxicity, and a low degree of dysregulation of > 370 genes actively engaged in the mediation of cellular toxicity. In vitro testing indicates a low probability of cellular toxicity associated with NCS-382.
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a pharmacokinetic evaluation and metabolite identification of the GHB Receptor antagonist ncs 382 in mouse informs novel therapeutic strategies for the treatment of GHB intoxication
Pharmacology Research & Perspectives, 2016Co-Authors: Garrett R Ainslie, Michael K Gibson, Kara R VogelAbstract:Gamma‐aminobutyric acid (GABA) is an endogenous inhibitory neurotransmitter and precursor of gamma‐hydroxybutyric acid (GHB). NCS‐382 (6,7,8,9‐tetrahydro‐5‐hydroxy‐5H‐benzo‐cyclohept‐6‐ylideneacetic acid), a known GHB Receptor antagonist, has shown significant efficacy in a murine model of succinic semialdehyde dehydrogenase deficiency (SSADHD), a heritable neurological disorder featuring chronic elevation of GHB that blocks the final step of GABA degradation. NCS‐382 exposures and elimination pathways remain unknown; therefore, the goal of the present work was to obtain in vivo pharmacokinetic data in a murine model and to identify the NCS‐382 metabolites formed by mouse and human. NCS‐382 single‐dose mouse pharmacokinetics were established following an intraperitoneal injection (100, 300, and 500 mg/kg body weight) and metabolite identification was conducted using HPLC‐MS/MS. Kinetic enzyme assays employed mouse and human liver microsomes. Upon gaining an understanding of the NCS‐382 clearance mechanisms, a chemical inhibitor was used to increase NCS‐382 brain exposure in a pharmacokinetic/pharmacodynamic study. Two major metabolic pathways of NCS‐382 were identified as dehydrogenation and glucuronidation. The K m for the dehydrogenation pathway was determined in mouse (K m = 29.5 ± 10.0 μmol/L) and human (K m = 12.7 ± 4.8 μmol/L) liver microsomes. Comparable parameters for glucuronidation were >100 μmol/L in both species. Inhibition of NCS‐382 glucuronidation, in vivo, by diclofenac resulted in increased NCS‐382 brain concentrations and protective effects in gamma‐butyrolactone‐treated mice. These initial evaluations of NCS‐382 pharmacokinetics and metabolism inform the development of NCS‐382 as a potential therapy for conditions of GHB elevation (including acute intoxication & SSADHD).
Maharaj K Ticku - One of the best experts on this subject based on the ideXlab platform.
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characterization and pharmacology of the GHB Receptor
Annals of the New York Academy of Sciences, 2008Co-Authors: Maharaj K Ticku, Ashok K MehtaAbstract:: Radioligand binding using [(3)H]NCS-382, an antagonist of the GHB Receptor, revealed specific binding sites in the rat cerebrocortical and hippocampal membranes. Scatchard analysis of saturation isotherms revealed two different populations of binding sites. NCS-382 was about 10 times more potent than GHB in inhibiting [(3)H]NCS-382 binding. A variety of ligands for other Receptors did not affect [(3)H]NCS-382 binding. Quantitative autoradiographic analysis of [(3)H]NCS-382 binding revealed similar characteristics. Thus [(3)H]NCS-382, being more potent and selective, offers advantage over [(3)H]GHB as a radioligand. Unlike GHB, several analogues of GHB such as UMB68 (a tertiary alcohol analogue of GHB), UMB86 (4-hydroxy-4-napthylbutanoic acid, sodium salt), UMB72 [4-(3-phenylpropyloxy)butyric acid, sodium salt], UMB73 (4-benzyloxybutyric acid, sodium salt), UMB66 (3-chloropropanoic acid), gamma-hydroxyvaleric acid (that is, GHV, a 4-methyl-substituted analogue of GHB), 3-HPA (3-hydroxyphenylacetic acid), and ethers of 3-hydroxyphenylacetic acid (UMB108, UMB109, and UMB119) displaced [(3)H]NCS-382 without affecting [(3)H]GABA binding to GABA(B) Receptor. Thus these compounds offer an advantage over GHB as an experimental tool. Our study, aimed at exploring the potential involvement of the GHB Receptor in the pharmacology of ethanol, indicated that ethanol does not affect [(3)H]NCS-382 binding in the rat brain, thereby suggesting that ethanol does not interact directly with the GHB Receptor. Our study, aimed at exploring the involvement of the GHB Receptor in the pathology of succinate semialdehyde dehydrogenase deficiency, which is known to cause elevation of GHB levels, revealed no change in the affinity, Receptor density or displacement potency as determined by using [(3)H]NCS-382 as a radioligand in Aldh5a1(-/-) vs. Aldh5a1(+/+) mouse brain.
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Succinate semialdehyde dehydrogenase deficiency does not down-regulate γ-hydroxybutyric acid binding sites in the mouse brain
Molecular Genetics and Metabolism, 2006Co-Authors: Ashok K Mehta, Lawrence P Carter, Georgianna G. Gould, Maneesh Gupta, K. Michael Gibson, Maharaj K TickuAbstract:Abstract We investigated whether succinate semialdehyde dehydrogenase deficiency alters γ-hydroxybutyric acid (GHB) Receptor characteristics due to elevation of GHB levels in the mouse brain. The membrane homogenate binding and quantitative autoradiography using [ 3 H]NCS-382 revealed no significant changes in the affinity ( K d ), Receptor density ( B max ), or displacement potency (IC 50 ) in various brain regions of Aldh5a1 −/− vs. Aldh5a1 +/+ mice.
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ethanol does not alter the binding of the γ hydroxybutyric acid GHB Receptor ligand 3h ncs 382 in the rat brain
Drug and Alcohol Dependence, 2004Co-Authors: Ashok K Mehta, Nicole M Muschaweck, Maharaj K TickuAbstract:Abstract We investigated the effect of ethanol on the binding of the γ-hydroxybutyric acid (GHB) Receptor ligand [ 3 H ]NCS-382 in the rat cerebral cortex and hippocampus. Ethanol (50–100 mM) did not alter the binding of [ 3 H ]NCS-382. Furthermore, acute (3 g/kg, p.o.) as well as chronic (9–15 g/kg/day p.o. for 6 days) administration of ethanol also did not have any significant effect on the binding of [ 3 H ]NCS-382 in the rat cerebrocortical and hippocampal membranes. These observations suggest that ethanol does not interact directly with the GHB Receptor in vitro or in vivo, and GHB Receptor may not be involved in the pharmacological effects of ethanol.
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3-chloropropanoic acid (UMB66): a ligand for the gamma-hydroxybutyric acid Receptor lacking a 4-hydroxyl group.
Bioorganic & Medicinal Chemistry, 2004Co-Authors: Alba Macias, Maharaj K Ticku, Ashok K Mehta, R. Jason Hernandez, Alexander D. Mackerell, Andrew CoopAbstract:Gamma-Hydroxybutyric acid (GHB) has gained in notoriety in recent years due to its association with sexual assaults. GHB is an endogenous ligand for GHB Receptors, but its complete pharmacological mechanism of action in vivo remains unclear due to apparent GABAergic components. It has been proposed that the hydroxyl group in the 4-position acts as a hydrogen bond donor to the GHB Receptor. Herein we show that 3-chloropropanoic acid possesses significant affinity for the GHB Receptor, has no affinity for GABA Receptors, and cannot undergo metabolism to GABAergic compounds. UMB66 is thus a selective agent for the study of GHB in vivo. These results, in combination with data from quantum mechanical calculations, suggest that the hydroxyl group of GHB actually acts as a hydrogen bond acceptor in contrast to the currently accepted model. This finding is anticipated to facilitate the rational design of novel agents with selectivity for GHB Receptors that may be used to elucidate the mechanism of action of this common drug of abuse.
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short communication ethanol does not alter the binding of the hydroxybutyric acid GHB Receptor ligand 3 h ncs 382 in the rat brain
2004Co-Authors: Ashok K Mehta, Nicole M Muschaweck, Maharaj K TickuAbstract:We investigated the effect of ethanol on the binding of the -hydroxybutyric acid (GHB) Receptor ligand [ 3 H]NCS-382 in the rat cerebral cortex and hippocampus. Ethanol (50–100 mM) did not alter the binding of [ 3 H]NCS-382. Furthermore, acute (3 g/kg, p.o.) as well as chronic (9–15 g/kg/day p.o. for 6 days) administration of ethanol also did not have any significant effect on the binding of [ 3 H]NCS-382 in the rat cerebrocortical and hippocampal membranes. These observations suggest that ethanol does not interact directly with the GHB Receptor in vitro or in vivo, and GHB Receptor may not be involved in the pharmacological effects of ethanol. © 2004 Elsevier Ireland Ltd. All rights reserved.
Garrett R Ainslie - One of the best experts on this subject based on the ideXlab platform.
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succinic semialdehyde dehydrogenase deficiency a disorder of gaba metabolism an update on pharmacological and enzyme replacement therapeutic strategies
Journal of Inherited Metabolic Disease, 2018Co-Authors: Kara R Vogel, Garrett R Ainslie, Alice Mcconnell, Jeanbaptiste Roullet, Dana C Walters, Sameer C Dhamne, Alexander Rotenberg, Michael K GibsonAbstract:We present an update to the status of research on succinic semialdehyde dehydrogenase (SSADH) deficiency (SSADHD), a rare disorder of GABA metabolism. This is an unusual disorder featuring the accumulation of both GABA and its neuromodulatory analog, gamma-hydroxybutyric acid (GHB), and recent studies have advanced the potential clinical application of NCS-382, a putative GHB Receptor antagonist. Animal studies have provided proof-of-concept that enzyme replacement therapy could represent a long-term therapeutic option. The characterization of neuronal stem cells (NSCs) derived from aldehyde dehydrogenase 5a1−/− (aldh5a1−/−) mice, the murine model of SSADHD, has highlighted NSC utility as an in vitro system in which to study therapeutics and associated toxicological properties. Gene expression analyses have revealed that transcripts encoding GABAA Receptors are down-regulated and may remain largely immature in aldh5a1−/− brain, characterized by excitatory as opposed to inhibitory outputs, the latter being the expected action in the mature central nervous system. This indicates that agents altering chloride channel activity may be therapeutically relevant in SSADHD. The most recent therapeutic prospects include mTOR (mechanistic target of rapamycin) inhibitors, drugs that have received attention with the elucidation of the effects of elevated GABA on autophagy. The outlook for novel therapeutic trials in SSADHD continues to improve.
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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
Toxicology in Vitro, 2018Co-Authors: Kara R Vogel, Garrett R Ainslie, Alice Mcconnell, Jeanbaptiste Roullet, K M GibsonAbstract: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 (300 mg/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 1 mM NCS-382 shows minimal evidence of pharmacotoxicity. As well, studies at the molecular level indicate that the effects of NCS-382 at 0.5 mM 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.
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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
Toxicology in Vitro, 2017Co-Authors: Kara R Vogel, Garrett R Ainslie, Alice Mcconnell, Jeanbaptiste Roullet, K M GibsonAbstract: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 (300 mg/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 1 mM NCS-382 shows minimal evidence of pharmacotoxicity. As well, studies at the molecular level indicate that the effects of NCS-382 at 0.5 mM 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.
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In vitro toxicological evaluation of NCS-382, a high-affinity antagonist of γ-hydroxybutyrate (GHB) binding.
Toxicology in Vitro, 2017Co-Authors: Kara R Vogel, Garrett R Ainslie, Alice Mcconnell, Jeanbaptiste Roullet, K M GibsonAbstract:Abstract γ-Hydroxybutyric acid (GHB), a minor metabolite of the inhibitory neurotransmitter GABA, can accumulate to significant concentrations in the heritable disorder of GABA degradation, succinic semialdehyde dehydrogenase (SSADH) deficiency (SSADHD). Moreover, GHB may be employed in therapeutic settings (treatment of narcolepsy), as well as instances of illicit activity, including acquaintance sexual assault and the induction of euphoria. High-affinity binding sites for GHB in the brain have been identified, although the absolute identity of these Receptors remains unclear. Pharmacological antagonism of GHB binding may have multiple instances of therapeutic relevance. The high affinity GHB Receptor antagonist, NCS-382 (6,7,8,9-tetrahydro-5-hydroxy-5H-benzo-cyclohept-6-ylideneacetic acid) has not been piloted in humans. To address the potential clinical utility of NCS-382, we have piloted initial studies of its toxicology in HepG2 and primary hepatocyte cells. At high dose (0.5 mM), NCS-382 showed no capacity for inhibition of microsomal CYPs (CYP1A2, 2B6, 2C8, 2C9, 2C19, 2D6 and 3A4) and minimal potential for activation of xenobiotic nuclear Receptors. Additional cellular integrity and functional assays (viability, oxidative stress, apoptosis, ATP production) revealed little evidence for cytotoxicity, and a low degree of dysregulation of > 370 genes actively engaged in the mediation of cellular toxicity. In vitro testing indicates a low probability of cellular toxicity associated with NCS-382.
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a pharmacokinetic evaluation and metabolite identification of the GHB Receptor antagonist ncs 382 in mouse informs novel therapeutic strategies for the treatment of GHB intoxication
Pharmacology Research & Perspectives, 2016Co-Authors: Garrett R Ainslie, Michael K Gibson, Kara R VogelAbstract:Gamma‐aminobutyric acid (GABA) is an endogenous inhibitory neurotransmitter and precursor of gamma‐hydroxybutyric acid (GHB). NCS‐382 (6,7,8,9‐tetrahydro‐5‐hydroxy‐5H‐benzo‐cyclohept‐6‐ylideneacetic acid), a known GHB Receptor antagonist, has shown significant efficacy in a murine model of succinic semialdehyde dehydrogenase deficiency (SSADHD), a heritable neurological disorder featuring chronic elevation of GHB that blocks the final step of GABA degradation. NCS‐382 exposures and elimination pathways remain unknown; therefore, the goal of the present work was to obtain in vivo pharmacokinetic data in a murine model and to identify the NCS‐382 metabolites formed by mouse and human. NCS‐382 single‐dose mouse pharmacokinetics were established following an intraperitoneal injection (100, 300, and 500 mg/kg body weight) and metabolite identification was conducted using HPLC‐MS/MS. Kinetic enzyme assays employed mouse and human liver microsomes. Upon gaining an understanding of the NCS‐382 clearance mechanisms, a chemical inhibitor was used to increase NCS‐382 brain exposure in a pharmacokinetic/pharmacodynamic study. Two major metabolic pathways of NCS‐382 were identified as dehydrogenation and glucuronidation. The K m for the dehydrogenation pathway was determined in mouse (K m = 29.5 ± 10.0 μmol/L) and human (K m = 12.7 ± 4.8 μmol/L) liver microsomes. Comparable parameters for glucuronidation were >100 μmol/L in both species. Inhibition of NCS‐382 glucuronidation, in vivo, by diclofenac resulted in increased NCS‐382 brain concentrations and protective effects in gamma‐butyrolactone‐treated mice. These initial evaluations of NCS‐382 pharmacokinetics and metabolism inform the development of NCS‐382 as a potential therapy for conditions of GHB elevation (including acute intoxication & SSADHD).
Michael K Gibson - One of the best experts on this subject based on the ideXlab platform.
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succinic semialdehyde dehydrogenase deficiency a disorder of gaba metabolism an update on pharmacological and enzyme replacement therapeutic strategies
Journal of Inherited Metabolic Disease, 2018Co-Authors: Kara R Vogel, Garrett R Ainslie, Alice Mcconnell, Jeanbaptiste Roullet, Dana C Walters, Sameer C Dhamne, Alexander Rotenberg, Michael K GibsonAbstract:We present an update to the status of research on succinic semialdehyde dehydrogenase (SSADH) deficiency (SSADHD), a rare disorder of GABA metabolism. This is an unusual disorder featuring the accumulation of both GABA and its neuromodulatory analog, gamma-hydroxybutyric acid (GHB), and recent studies have advanced the potential clinical application of NCS-382, a putative GHB Receptor antagonist. Animal studies have provided proof-of-concept that enzyme replacement therapy could represent a long-term therapeutic option. The characterization of neuronal stem cells (NSCs) derived from aldehyde dehydrogenase 5a1−/− (aldh5a1−/−) mice, the murine model of SSADHD, has highlighted NSC utility as an in vitro system in which to study therapeutics and associated toxicological properties. Gene expression analyses have revealed that transcripts encoding GABAA Receptors are down-regulated and may remain largely immature in aldh5a1−/− brain, characterized by excitatory as opposed to inhibitory outputs, the latter being the expected action in the mature central nervous system. This indicates that agents altering chloride channel activity may be therapeutically relevant in SSADHD. The most recent therapeutic prospects include mTOR (mechanistic target of rapamycin) inhibitors, drugs that have received attention with the elucidation of the effects of elevated GABA on autophagy. The outlook for novel therapeutic trials in SSADHD continues to improve.
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a pharmacokinetic evaluation and metabolite identification of the GHB Receptor antagonist ncs 382 in mouse informs novel therapeutic strategies for the treatment of GHB intoxication
Pharmacology Research & Perspectives, 2016Co-Authors: Garrett R Ainslie, Michael K Gibson, Kara R VogelAbstract:Gamma‐aminobutyric acid (GABA) is an endogenous inhibitory neurotransmitter and precursor of gamma‐hydroxybutyric acid (GHB). NCS‐382 (6,7,8,9‐tetrahydro‐5‐hydroxy‐5H‐benzo‐cyclohept‐6‐ylideneacetic acid), a known GHB Receptor antagonist, has shown significant efficacy in a murine model of succinic semialdehyde dehydrogenase deficiency (SSADHD), a heritable neurological disorder featuring chronic elevation of GHB that blocks the final step of GABA degradation. NCS‐382 exposures and elimination pathways remain unknown; therefore, the goal of the present work was to obtain in vivo pharmacokinetic data in a murine model and to identify the NCS‐382 metabolites formed by mouse and human. NCS‐382 single‐dose mouse pharmacokinetics were established following an intraperitoneal injection (100, 300, and 500 mg/kg body weight) and metabolite identification was conducted using HPLC‐MS/MS. Kinetic enzyme assays employed mouse and human liver microsomes. Upon gaining an understanding of the NCS‐382 clearance mechanisms, a chemical inhibitor was used to increase NCS‐382 brain exposure in a pharmacokinetic/pharmacodynamic study. Two major metabolic pathways of NCS‐382 were identified as dehydrogenation and glucuronidation. The K m for the dehydrogenation pathway was determined in mouse (K m = 29.5 ± 10.0 μmol/L) and human (K m = 12.7 ± 4.8 μmol/L) liver microsomes. Comparable parameters for glucuronidation were >100 μmol/L in both species. Inhibition of NCS‐382 glucuronidation, in vivo, by diclofenac resulted in increased NCS‐382 brain concentrations and protective effects in gamma‐butyrolactone‐treated mice. These initial evaluations of NCS‐382 pharmacokinetics and metabolism inform the development of NCS‐382 as a potential therapy for conditions of GHB elevation (including acute intoxication & SSADHD).
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γ hydroxybutyric acid GHB and γ aminobutyric acidb Receptor gababr binding sites are distinctive from one another molecular evidence
Neuropharmacology, 2004Co-Authors: Ying Wu, Michael K Gibson, Gholamreza Ahmadian, Yu Tian Wang, Andrew R Calver, Joseph Francis, Menelas N Pangalos, Carter O SneadAbstract:Abstract γ-Hydroxybutyric Acid (GHB) is thought to be a weak partial agonist at the γ-aminobutyric acidB Receptor (GABABR), but the precise relationship of the GHB Receptor (GHBR) to the GABABR remains unclear. In order to test the hypothesis that the GHBR is not identical to the GABABR, we conducted two groups of experiments. First, GABABR subtype 1 (R1) and/or subtype 2 (R2) were over expressed in HEK 293 cells and membrane binding studies on the transfected cells done using [3H]GHB and [3H] (2E)-(5-hydroxy-5,7,8,9-tetrahydro-6H-benzo[a][7]annulen-6-ylidene) ethanoic acid ([3H]NCS-382). The latter is a specific antagonist at the GHB binding site. Second, [3H]GHB and [3H]NCS-382 autoradiographic binding studies were done on the brains of mice in which the gene for GABABR1a was deleted. Such mice do not have a functioning GABABR. There was no detectable specific [3H]GHB or [3H]NCS-382 binding in HEK 293 cells transfected with GABABR1, R2, or R1/R2. Binding to [3H]CGP54626A, a high affinity GABABR antagonist, was absent in GABABR1a−/− mice. There was no difference in [3H]NCS-382 binding observed in the brains of GABABR1a−/−, GABABR1a+/− or GABABR1a+/+ mice. Specific [3H]GHB binding was observed in the brain of GABABR1a−/− mice but was significantly lower than in wild type mice. These data support the hypothesis that the GHB binding site is separate and distinct from the GABABR.
K M Gibson - One of the best experts on this subject based on the ideXlab platform.
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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
Toxicology in Vitro, 2018Co-Authors: Kara R Vogel, Garrett R Ainslie, Alice Mcconnell, Jeanbaptiste Roullet, K M GibsonAbstract: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 (300 mg/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 1 mM NCS-382 shows minimal evidence of pharmacotoxicity. As well, studies at the molecular level indicate that the effects of NCS-382 at 0.5 mM 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.
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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
Toxicology in Vitro, 2017Co-Authors: Kara R Vogel, Garrett R Ainslie, Alice Mcconnell, Jeanbaptiste Roullet, K M GibsonAbstract: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 (300 mg/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 1 mM NCS-382 shows minimal evidence of pharmacotoxicity. As well, studies at the molecular level indicate that the effects of NCS-382 at 0.5 mM 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.
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In vitro toxicological evaluation of NCS-382, a high-affinity antagonist of γ-hydroxybutyrate (GHB) binding.
Toxicology in Vitro, 2017Co-Authors: Kara R Vogel, Garrett R Ainslie, Alice Mcconnell, Jeanbaptiste Roullet, K M GibsonAbstract:Abstract γ-Hydroxybutyric acid (GHB), a minor metabolite of the inhibitory neurotransmitter GABA, can accumulate to significant concentrations in the heritable disorder of GABA degradation, succinic semialdehyde dehydrogenase (SSADH) deficiency (SSADHD). Moreover, GHB may be employed in therapeutic settings (treatment of narcolepsy), as well as instances of illicit activity, including acquaintance sexual assault and the induction of euphoria. High-affinity binding sites for GHB in the brain have been identified, although the absolute identity of these Receptors remains unclear. Pharmacological antagonism of GHB binding may have multiple instances of therapeutic relevance. The high affinity GHB Receptor antagonist, NCS-382 (6,7,8,9-tetrahydro-5-hydroxy-5H-benzo-cyclohept-6-ylideneacetic acid) has not been piloted in humans. To address the potential clinical utility of NCS-382, we have piloted initial studies of its toxicology in HepG2 and primary hepatocyte cells. At high dose (0.5 mM), NCS-382 showed no capacity for inhibition of microsomal CYPs (CYP1A2, 2B6, 2C8, 2C9, 2C19, 2D6 and 3A4) and minimal potential for activation of xenobiotic nuclear Receptors. Additional cellular integrity and functional assays (viability, oxidative stress, apoptosis, ATP production) revealed little evidence for cytotoxicity, and a low degree of dysregulation of > 370 genes actively engaged in the mediation of cellular toxicity. In vitro testing indicates a low probability of cellular toxicity associated with NCS-382.