Nonketotic Hyperglycinemia Research Articles

Gldc is essential for embryonic development and kidney organogenesis

Glycine is a simple amino acid that is vital to the proper functioning of the body, but its roles during ontogeny are not well understood. Glycine levels are precisely regulated through the glycine cleavage system (GCS), a complex molecular machine that produces one-carbon units for later metabolism and folate production. Elevated glycine levels due to congenital mutations in GCS components, such as glycine dehydrogenase (gldc), cause devastating human birth defects and the rare disease nonketotic hyperglycinemia (NKH). The exact pathological implications of glycine accumulation are unknown, but NKH patients suffer from pleiotropic symptoms including seizures, apnea, lethargy, severe mental retardation, and early death. Animal models utilizing mice and zebrafish have shown increased ventriculomegaly and motor dysfunction in gldc deficient animals, but have not analyzed organogenesis. Here, we have utilized zebrafish as a genetic platform to explore the roles of gldc during early development. Whole mount in situ hybridization revealed that gldc transcripts were highly expressed in multiple tissues, including the central nervous system and embryonic kidney, recapitulating both mouse and human expression studies. gldc deficiency caused hydrocephalus and pericardial edema, suggesting defects in brain and renal ontogeny. Elevated apoptosis was detected in locations within the brain, kidney, and cloaca. Furthermore, formation of nephron cell populations in the kidney was disrupted in gldc deficient embryos, where the proximal straight tubule was increased at the expense of the distal tubule. Morphological and expression studies revealed severe alterations in brain patterning and morphogenesis as well. Taken together, these studies indicate that gldc has essential roles during early development and have revealed novel functions for pattern formation and cell survival in tissues such as the brain and kidney. Our evidence further emphasizes the zebrafish as a faithful model to examine NKH. Ongoing work with the zebrafish model is poised to expand our medical understanding about the developmental defects consequent to NKH and can be used to perform chemical screens to identify potential therapeutic interventions.

Stimulation of Dopamine Production by Sodium Benzoate, a Metabolite of Cinnamon and a Food Additive.

Background:Parkinson’s disease (PD) is one of the most important neurodegenerative disorders in human in which recovery of functions could be achieved by improving the survival and function of residual dopaminergic neurons in the substantia nigra pars compacta. Tyrosine hydroxylase (TH) is the rate-limiting enzyme in the dopamine (DA) biosynthesis pathway.Objective:Earlier our laboratory has shown that sodium benzoate (NaB), a metabolite of cinnamon and an FDA-approved drug against urea cycle disorders and glycine encephalopathy, increases neuroprotective molecules and protects dopaminergic neurons in a mouse model of PD. Here, we examined whether NaB could stimulate the production of DA in dopaminergic neurons.Methods:We employed PCR, real-time PCR, western blot, immunostaining, and HPLC to study the signature function of dopaminergic neurons. Locomotor functions were monitored in mice by open-field.Results:NaB increased the mRNA and protein expression of TH to produce DA in mouse MN9D dopaminergic neuronal cells. Accordingly, oral feeding of NaB increased the expression of TH in the nigra, upregulated striatal DA, and improved locomotor activities in striatum of normal C57/BL6 and aged A53T-α-syn transgenic mice. Rapid induction of cAMP response element binding (CREB) activation by NaB in dopaminergic neuronal cells and the abrogation of NaB-induced expression of TH by siRNA knockdown of CREB suggest that NaB stimulates the transcription of TH in dopaminergic neurons via CREB.Conclusion:These results indicate a new function of NaB in which it may be beneficial in PD via stimulation of DA production from residual dopaminergic neurons.

An Adult Case of Non-Ketotic Hyperglycinemia Due to Novel Compound Heterozygous Variants in GLDC (1656)

An Adult Case of Non-Ketotic Hyperglycinemia Due to Novel Compound Heterozygous Variants in GLDC (1656)

Genotypic and phenotypic features in Turkish patients with classic nonketotic hyperglycinemia.

Nonketotic hyperglycinemia is an autosomal recessive inborn error of glycine metabolism, characterized by deficient activity of the glycine cleavage enzyme system. Classic nonketotic hyperglycinemia is caused by mutations or genomic changes in genes that encode the protein components of the glycine cleavage enzyme system. We aimed to investigate clinical, biochemical, radiological findings and molecular genetic data in ten Turkish patients with classic nonketotic hyperglycinemia.Ten Turkish patients who were diagnosed with classic nonketotic hyperglycinemia in a single center from 2013 to 2019 were included in this study. Their clinical, radiological, electrophysiological and laboratory data were collected retrospectively.Sixty percent of the patients were in neonatal group, while 40 % of the patients were infantile. There were no late-onset patients. 90 % of the patients had the severe form. All patients had developmental delay and seizures. Mortality ratio was 30 % in all groups and 50 % in the neonatal group, while no mortality was seen in infantile group. Median (range) values of cerebrospinal fluid (CSF) glycine levels, plasma glycine levels and CSF/plasma glycine ratios were 148 (15-320) µmol/L, 896 (87-1910) µmol/L, 0.17 (0.09-0.21) respectively. Diffuse hypomyelination and corpus callosum anomaly were the most common cranial MRI findings and multifocal epileptic activity and burst supression pattern were the most common electroencephalographic findings. Six patients had variants in GLDC gene and four in AMT gene; five novel variants including AMT gene deletion were detected. Prognosis was poor and treatment was not effective, especially in the severe form.Classic nonketotic hyperglycinemia causes high morbidity and mortality. Neonatal-onset disease was more common and severe than infantile-onset disease. The ratio of AMT gene variants might be higher in Turkey than other countries. AMT gene deletion also plays a role in the etiology of classic nonketotic hyperglycinemia.

Alleviation of Huntington pathology in mice by oral administration of food additive glyceryl tribenzoate

Huntington's disease (HD) is a neurodegenerative disorder characterized by accumulation of mutant huntingtin protein and significant loss of neurons in striatum and cortex. Along with motor difficulties, the HD patients also manifest anxiety and loss of cognition. Unfortunately, the clinically approved drugs only offer symptomatic relief and are not free from side effects. This study underlines the importance of glyceryl tribenzoate (GTB), an FDA-approved food flavoring ingredient, in alleviating HD pathology in transgenic N171-82Q mouse model. Oral administration of GTB significantly reduced mutant huntingtin level in striatum, motor cortex as well as hippocampus and increased the integrity of viable neurons. Furthermore, we found the presence of sodium benzoate (NaB), a FDA-approved drug for urea cycle disorders and glycine encephalopathy, in the brain of GTB-fed HD mice. Accordingly, NaB administration also markedly decreased huntingtin level in striatum and cortex. Glial activation is found to coincide with neuronal death in affected regions of HD brains. Interestingly, both GTB and NaB treatment suppressed activation of glial cells and inflammation in the brain. Finally, neuroprotective effect of GTB and NaB resulted in improved motor performance of HD mice. Collectively, these results suggest that GTB and NaB may be repurposed for HD.

Elevated preoptic brain activity in zebrafish glial glycine transporter mutants is linked to lethargy-like behaviors and delayed emergence from anesthesia

Delayed emergence from anesthesia was previously reported in a case study of a child with Glycine Encephalopathy. To investigate the neural basis of this delayed emergence, we developed a zebrafish glial glycine transporter (glyt1 − / −) mutant model. We compared locomotor behaviors; dose–response curves for tricaine, ketamine, and 2,6-diisopropylphenol (propofol); time to emergence from these anesthetics; and time to emergence from propofol after craniotomy in glyt1−/− mutants and their siblings. To identify differentially active brain regions in glyt1−/− mutants, we used pERK immunohistochemistry as a proxy for brain-wide neuronal activity. We show that glyt1−/− mutants initiated normal bouts of movement less frequently indicating lethargy-like behaviors. Despite similar anesthesia dose–response curves, glyt1−/− mutants took over twice as long as their siblings to emerge from ketamine or propofol, mimicking findings from the human case study. Reducing glycine levels rescued timely emergence in glyt1−/− mutants, pointing to a causal role for elevated glycine. Brain-wide pERK staining showed elevated activity in hypnotic brain regions in glyt1−/− mutants under baseline conditions and a delay in sensorimotor integration during emergence from anesthesia. Our study links elevated activity in preoptic brain regions and reduced sensorimotor integration to lethargy-like behaviors and delayed emergence from propofol in glyt1−/− mutants.

Genomic analyses of glycine decarboxylase neurogenic mutations yield a large-scale prediction model for prenatal disease.

Hundreds of mutations in a single gene result in rare diseases, but why mutations induce severe or attenuated states remains poorly understood. Defect in glycine decarboxylase (GLDC) causes Non-ketotic Hyperglycinemia (NKH), a neurological disease associated with elevation of plasma glycine. We unified a human multiparametric NKH mutation scale that separates severe from attenuated neurological disease with new in silico tools for murine and human genome level-analyses, gathered in vivo evidence from mice engineered with top-ranking attenuated and a highly pathogenic mutation, and integrated the data in a model of pre- and post-natal disease outcomes, relevant for over a hundred major and minor neurogenic mutations. Our findings suggest that highly severe neurogenic mutations predict fatal, prenatal disease that can be remedied by metabolic supplementation of dams, without amelioration of persistent plasma glycine. The work also provides a systems approach to identify functional consequences of mutations across hundreds of genetic diseases. Our studies provide a new framework for a large scale understanding of mutation functions and the prediction that severity of a neurogenic mutation is a direct measure of pre-natal disease in neurometabolic NKH mouse models. This framework can be extended to analyses of hundreds of monogenetic rare disorders where the underlying genes are known but understanding of the vast majority of mutations and why and how they cause disease, has yet to be realized.

Glycine Cleavage System H Protein Is Essential for Embryonic Viability, Implying Additional Function Beyond the Glycine Cleavage System.

Glycine cleavage system H protein (GCSH) is a component of the glycine cleavage system (GCS), a conserved protein complex that acts to decarboxylate glycine. Mutation of AMT or GLDC, encoding the GCS components aminomethyltransferase and glycine decarboxylase, can cause malformations of the developing CNS (neural tube defects (NTDs) and ventriculomegaly) as well as a post-natal life-limiting neurometabolic disorder, Non-Ketotic Hyperglycinemia. In contrast, it is unclear whether mutation of GCSH contributes to these conditions and we therefore investigated GCSH loss of function in mice. Mice that were heterozygous for a Gcsh null allele were viable and did not exhibit elevated plasma glycine. Moreover, heterozygous mutation of Gcsh did not increase the frequency of NTDs in Gldc mutant embryos. Homozygous Gcsh null mice were not recovered at post-natal stages. Analysis of litters at E8.5-10.5, revealed the presence of homozygous null embryos which were much smaller than littermates and had failed to develop beyond early post-implantation stages with no visible somites or head-folds. Hence, unlike null mutations of Gldc or Amt, which are compatible with embryonic survival despite the presence of NTDs, loss of Gcsh causes embryonic death prior to mid-gestation. Maternal supplementation with formate did not restore embryonic development beyond E7.5, suggesting that the primary cause of lethality was not loss of glycine cleavage activity or suppression of folate one-carbon metabolism. These findings suggest that GCSH has additional roles beyond function in the glycine cleavage system. We hypothesize that GCSH potentially acts in lipoylation of 2-oxoacid dehydrogenase proteins, as reported in bacteria.

An Integrative Approach to Predict Phenotypic Severity in Nonketotic Hyperglycinemia

Background: Nonketotic hyperglycinemia (NKH) is a inherited neurometabolic disorder with variable clinical course and severity, in a spectrum ranging from infantile epileptic encephalopathy to psychiatric disorders. A precise phenotypic characterization and an evaluation of predictive approaches are needed. We report the results of the patient registry of the International Working Group on Neurotransmitter related Disorders (iNTD). Methods: The iNTD patient registry is a multicenter, uncontrolled, non-randomized, open, unblinded observational study (registered on DRKS, DRK7878). Longitudinal clinical and biochemical data of 25 individuals with NKH were studied with in silico analyses, pathogenicity scores and molecular modeling of GLDC and AMT variants. Findings: Age of onset (p=0 · 004) and diagnosis are earlier in life in severe NKH( p=0 · 005). Presenting symptoms affect the age at diagnosis. Psychiatric problems occur predominantly in attenuated NKH. Onset-age ≥3 months (66% specificity, 100% sensitivity, AUC = 0·87) and cerebrospinal fluid (CSF)/plasma glycine ratio ≤0 · 09 (57% specificity, 100% sensitivity, AUC = 0·88) are sensitive indicators for attenuated NKH while CSF glycine concentration ≥116 · 5 µmol/L (100% specificity, 93% sensitivity, AUC = 0·97) and CSF/plasma glycine ratio ≥0 · 15 15 (100% specificity, 64% sensitivity, AUC = 0·88) are specific for severe forms. A ratio threshold of 0 · 128 discriminates the overlapping range. In our study, we present ten new GLDC variants, two mild variants resulting in attenuated phenotype. Two severe variants and a combination of one mild and one severe variant lead to severe phenotype. Interpretation: This study widens the phenotypic spectrum of attenuated NKH and expands the number of pathogenic variants. The multiparametric approach provides a promising tool to predict disease severity, helping to improve clinical management and decision-making strategies. Trial Registration: This study was registered German Clinical Trials Register, https://www.drks.de, DRK7878). Funding: Ministry of Health of the Czech Republic (RVO-VFN 64165 GJIH-0599-00-7-846, ProgresQ26/LF1), FIS P118/00111 and PI19/00348 “Instituto de Salud Carlos III”, “Fondo Europeo de desarrollo regional (FEDER)”, and Dietmar Hopp Foundation. Declaration of Interest: A.G.C. has received teaching honorarium from PTC Therapeutics GT, Inc. G.F.H. receives teaching as well as consultancy honorarium from PTC Therapeutics GT, Inc. O.K.H. has received teaching honorarium from PTC Therapeutics GT, Inc. T.O. receives teaching honorarium and research support from PTC. GFH has received honoraria as a speaker from Takeda and consultancy honoraria from PTC Therapeutics International GT. T.S.P receives consulting honoraria from Teva Pharmaceuticals. The other authors declare no potential conflict of interest. Ethical Approval: iNTD registry study was approved by the Institutional Research Ethics Board (IRB) Heidelberg University Hospital (S-471/2014)

Genetic cause of epilepsy in a Greek cohort of children and young adults with heterogeneous epilepsy syndromes

We describe a cohort of 10 unrelated Greek patients (4 females, 6 males; median age 6.5 years, range 2-18 years) with heterogeneous epilepsy syndromes with a genetic basis. In these patients, causative genetic variants, including two novel ones, were identified in 9 known epilepsy-related genes through whole exome sequencing. A patient with glycine encephalopathy was a compound heterozygote for the p.Arg222Cys and the p.Ser77Leu AMT variant. A patient affected with Lafora disease carried the homozygous p.Arg171His EPM2A variant. A de novo heterozygous variant in the GABRG2 gene (p.Pro282Thr) was found in one patient and a pathogenic variant in the GRIN2B gene (p.Gly820Val) in another patient. Infantile-onset lactic acidosis with seizures was associated with the p.Arg446Ter PDHX gene variant in one patient. In two additional epilepsy patients, the p.Ala1662Val and the novel non-sense p.Phe1330Ter SCN1A gene variants were found. Finally, in 3 patients we observed a novel heterozygous missense variant in SCN2A (p.Ala1874Thr), a heterozygous splice site variant in SLC2A1 (c.517-2A>G), as a cause of Glut1 deficiency syndrome, and a pathogenic variant in STXBP1 (p.Arg292Leu), respectively. In half of our cases (patients with variants in the GRIN2B, SCN1A, SCN2A and SLC2A1 genes), a genetic cause with potential management implications was identified.

New Pathogenic Mutations on GLDC gene leading to Classic Nonketotic Hyperglycinemia

New Pathogenic Mutations on GLDC gene leading to Classic Nonketotic Hyperglycinemia

Attenuated form of Glycine Encephalopathy

Attenuated form of Glycine Encephalopathy

GLYT1 encephalopathy: Further delineation of disease phenotype and discussion of pathophysiological mechanisms.

GLYT1 encephalopathy is a form of glycine encephalopathy caused by disturbance of glycine transport. The phenotypic spectrum of the disease has not yet been completely described, as only four unrelated families with the disorder have been reported to date. Common features of affected patients include neonatal hypotonia, respiratory failure, encephalopathy, myoclonic jerks, dysmorphic features, and musculoeskeletal anomalies. All reported affected patients harbor biallelic genetic variants in SLC6A9. SNP array together with Sanger sequencing were performed in a newborn with arthrogryposis and severe neurological impairment. The novel genetic variant c.997delC in SLC6A9 was detected in homozygous state in the patient. At protein level, the predicted change is p.(Arg333Alafs*3), which most probably results in a loss of protein function. The variant cosegregated with the disease in the family. A subsequent pregnancy with ultrasound anomalies was also affected. The proband presented the core phenotypic features of GLYT1 encephalopathy, but also a burst suppression pattern on the electroencephalogram, a clinical feature not previously associated with the disorder. Our results suggest that the appearance of this pattern correlates with higher cerebrospinal fluid glycine levels and cerebrospinal fluid/plasma glycine ratios. A detailed discussion on the possible pathophysiological mechanisms of the disorder is also provided.

Can cinnamon spice down autoimmune diseases?

Autoimmune diseases are one of the dreadful group of human diseases that have always been of keen interest to researchers. Due to complex and broad-spectrum nature, scientists are not yet able to pinpoint the pathogenesis of and delineate effective therapy against this group of diseases. However, it is becoming clear that a decrease in number and function of T regulatory cells (Treg), an increase in autoreactive Th1/Th17 cells and associated immunomodulation and inflammation participate in the pathogenesis of many autoimmune diseases. Cinnamon (Cinnamonum verum or Cinnamonum cassia) is a widely used natural spice and flavoring ingredient and its metabolite sodium benzoate (NaB) is a food-additive and FDA-approved drug against nonketotic hyperglycinemia (NKH) and urea cycle disorders (UCD). Recent studies indicate that cinnamon either in powder or extract form and NaB are capable of modulating different autoimmune pathways as well as protecting animals from different autoimmune disorders. Here, we have made an honest attempt to delineate such pieces of evidence with available anti-autoimmune mechanisms and analyze whether cinnamon supplements could be used to control the fury of autoimmune disorders.

Epilepsy in inherited neurotransmitter disorders: Spotlights on pathophysiology and clinical management.

Inborn errors of neurotransmitter metabolism are ultrarare disorders affecting neurotransmitter biosynthesis, breakdown or transport or their essential cofactors. Neurotransmitter dysfunctions could also result from the impairment of neuronal receptors, intracellular signaling, vesicle release or other synaptic abnormalities. Epilepsy is the main clinical hallmark in some of these diseases (e.g. disorders of GABA metabolism, glycine encephalopathy) while it is infrequent in others (e.g. all the disorders of monoamine metabolism in exception for dihydropteridine reductase deficiency). This review analyzes the epileptogenic mechanisms, the epilepsy phenotypes and the principle for the clinical management of epilepsy in primary and secondary inherited disorders of neurotransmitter metabolism (disorders of GABA, serine and glycine metabolism, disorders of neurotransmitter receptors and secondary neurotransmitter diseases).

Mass spectrometry imaging reveals glycine distribution in the developing and adult mouse brain

Glycine is an important amino acid in the central nervous system. The aberrant conditions of glycine concentrations cause sever neurological disorders, such as nonketotic-hyperglycinemia (NKH), also known as glycine encephalopathy. Therefore, a better understanding of its relative abundance and distribution in the developing and adult brains would provide insights into the pathogeneses of this kind of disorders. Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) imaging has been used for direct molecular-specific compound detection, distribution mapping, and identifying molecular species in tissue sections. Although a few reports have already shown the imaging of glycine using MALDI-MS in the adult mouse brain, they lack detailed neuroanatomical and developmental evaluations. We, thus, investigated the detailed distribution and abundance of glycine not only in the adult mouse brain but also in the developing mouse brain using this technique. In both brains, we detected derivatized glycine throughout the mouse brain. Interestingly, in both brains, derivatized glycine was abundantly detected in the brain stem. The other areas showed relatively lower signal intensities. As many model mice are used for glycine-related diseases, MALDI-MS is a suitable technique to analyze the pathogenesis of these diseases.

Reduction of Lewy Body Pathology by Oral Cinnamon.

α-Synucleinopathies in a broader sense comprise of several neurodegenerative disorders that primarily include Parkinson's disease (PD), dementia with Lewy bodies (DLB) and multiple system atrophy (MSA). These disorders are well characterized by the accumulation of aggregated insoluble α-synuclein (α-syn) protein known as Lewy bodies. Till date no effective cure is available to reduce the burden of Lewy body. The present investigation underlines the importance of a naturally used spice and flavoring agent viz. cinnamon in reducing α-syn deposits in transgenic mice expressing mutant A53T human α-syn. Upon oral administration, cinnamon markedly reduced the level of insoluble α-syn in nigra, hippocampus and brain stem of A53T mice. We also demonstrated that sodium benzoate (NaB), a metabolite of cinnamon, a widely used food additive and a FDA-approved drug for glycine encephalopathy, was also capable of reducing α-syn deposits in A53T mice. In addition, both cinnamon and NaB treatments showed improvement in their motor and cognitive functions. Glial activation plays an important role in the pathogenesis of various neurodegenerative disorders including PD, DLB and MSA, and we found suppression of microglial and astroglial activation in the nigra of A53T mice upon cinnamon treatment. Moreover, neuroprotective proteins like DJ-1 and Parkin are known to reduce the formation of Lewy bodies in the CNS. Accordingly, we observed upregulation and/or normalization of DJ-1 and Parkin in the nigra of A53T mice by treatment with cinnamon and NaB. Together, these results highlight a new therapeutic property of cinnamon and suggest that cinnamon and NaB may be used to halt the progression of α-synucleinopathies.Graphical Abstract.

Nonketotic Hyperglycinemia in Tunisia. Report upon a Series of 69 Patients.

The aim of the study is to report on epidemiological, clinical, and biochemical characteristics of nonketotic hyperglycinemia (NKH) in Tunisia. Patients diagnosed with NKH in Laboratory of Biochemistry at Rabta hospital (Tunis, Tunisia) between 1999 and 2018 were included. Plasma and cerebrospinal fluid (CSF) free amino acids were assessed by ion exchange chromatography. Diagnosis was based on family history, patient's clinical presentation and course, and increased CSF to plasma glycine ratio. During 20 years, 69 patients were diagnosed with NKH, with 25 patients originating from Kairouan region. Estimated incidences were 1:55,641 in Tunisia and 1:9,684 in Kairouan. Consanguinity was found for 73.9% of the patients and 42% of the families have history of infantile death due to a disease of similar clinical course than the propositus. Clinical symptoms initiated within the first week of life in 75% of the patients and within the first 3 months in 95.7% ones. The phenotype was severe in 76.8% of the patients. Main symptoms were hypotonia, feeding difficulties, coma, apnea, and seizures. Most patients died within few days to months following diagnosis. CSF to plasma glycine ratio was increased in all patients. CSF and plasma glycine levels were negatively correlated with age of disease onset and severity. NKH is quite frequent in Tunisia. Kairouan region has the highest NKH incidence rate, worldwide. However, due to lack of confirmatory enzymatic and genetic tests, NKH diagnosis was based on first-line biochemical tests. Characterization of causal mutations is needed for accurate diagnosis and prenatal diagnosis of this devastating life-threatening disease.

Metabolic epilepsies amenable to ketogenic therapies: Indications, contraindications, and underlying mechanisms.

Metabolic epilepsies arise in the context of rare inborn errors of metabolism (IEM), notably glucose transporter type 1 deficiency syndrome, succinic semialdehyde dehydrogenase deficiency, pyruvate dehydrogenase complex deficiency, nonketotic hyperglycinemia, and mitochondrial cytopathies. A common feature of these disorders is impaired bioenergetics, which through incompletely defined mechanisms result in a wide spectrum of neurological symptoms, such as epileptic seizures, developmental delay, and movement disorders. The ketogenic diet (KD) has been successfully utilized to treat such conditions to varying degrees. While the mechanisms underlying the clinical efficacy of the KD in IEM remain unclear, it is likely that the proposed heterogeneous targets influenced by the KD work in concert to rectify or ameliorate the downstream negative consequences of genetic mutations affecting key metabolic enzymes and substrates-such as oxidative stress and cell death. These beneficial effects can be broadly grouped into restoration of impaired bioenergetics and synaptic dysfunction, improved redox homeostasis, anti-inflammatory, and epigenetic activity. Hence, it is conceivable that the KD might prove useful in other metabolic disorders that present with epileptic seizures. At the same time, however, there are notable contraindications to KD use, such as fatty acid oxidation disorders. Clearly, more research is needed to better characterize those metabolic epilepsies that would be amenable to ketogenic therapies, both experimentally and clinically. In the end, the expanded knowledge base will be critical to designing metabolism-based treatments that can afford greater clinical efficacy and tolerability compared to current KD approaches, and improved long-term outcomes for patients.

Regulation of glycine metabolism by the glycine cleavage system and conjugation pathway in mouse models of non-ketotic hyperglycinemia.

Glycine abundance is modulated in a tissue‐specific manner by use in biosynthetic reactions, catabolism by the glycine cleavage system (GCS), and excretion via glycine conjugation. Dysregulation of glycine metabolism is associated with multiple disorders including epilepsy, developmental delay, and birth defects. Mutation of the GCS component glycine decarboxylase (GLDC) in non‐ketotic hyperglycinemia (NKH) causes accumulation of glycine in body fluids, but there is a gap in our knowledge regarding the effects on glycine metabolism in tissues. Here, we analysed mice carrying mutations in Gldc that result in severe or mild elevations of plasma glycine and model NKH. Liver of Gldc‐deficient mice accumulated glycine and numerous glycine derivatives, including multiple acylglycines, indicating increased flux through reactions mediated by enzymes including glycine‐N‐acyltransferase and arginine: glycine amidinotransferase. Levels of dysregulated metabolites increased with age and were normalised by liver‐specific rescue of Gldc expression. Brain tissue exhibited increased abundance of glycine, as well as derivatives including guanidinoacetate, which may itself be epileptogenic. Elevation of brain tissue glycine occurred even in the presence of only mildly elevated plasma glycine in mice carrying a missense allele of Gldc. Treatment with benzoate enhanced hepatic glycine conjugation thereby lowering plasma and tissue glycine. Moreover, administration of a glycine conjugation pathway intermediate, cinnamate, similarly achieved normalisation of liver glycine derivatives and circulating glycine. Although exogenous benzoate and cinnamate impact glycine levels via activity of glycine‐N‐acyltransferase, that is not expressed in brain, they are sufficient to lower levels of glycine and derivatives in brain tissue of treated Gldc‐deficient mice.