Utilization of EEG microstates as a prospective biomarker for assessing the impact of ketogenic diet in GLUT1-DS.

OBJECTIVE: The aim of the study is to analyze microstate patterns in GLUT1-DS, both before and after the ketogenic diet (KD). METHODS: We conducted microstate analysis of a patient with GLUT-1 DS and 27 healthy controls. A systematic literature review and meta-analysis was done. We compared the parameters of the patients with those of healthy controls and the incorporating findings in literature. RESULTS: The durations of the patient were notably shorter, and the occurrence rates were longer than those of healthy controls and incorporating findings from the review. After 10 months of KD, the patient's microstate durations exhibited an increase from 53.05 ms, 57.17 ms, 61.80 ms, and 49.49 ms to 60.53 ms, 63.27 ms, 71.11 ms, and 66.55 ms. The occurrence rates changed from 4.0774 Hz, 4.9462 Hz, 4.8006 Hz, and 4.0579 Hz to 3.3354 Hz, 3.7893 Hz, 3.5956 Hz, and 4.1672 Hz. In healthy controls, the durations of microstate class A, B, C, and D were 61.86 ms, 63.58 ms, 70.57 ms, and 72.00 ms, respectively. CONCLUSIONS: Our findings suggest EEG microstates may be a promising biomarker for monitoring the effect of KD. Administration of KD may normalize the dysfunctional patterns of temporal parameters.

The use of ketogenic diets in children living with drug-resistant epilepsy, glucose transporter 1 deficiency syndrome and pyruvate dehydrogenase deficiency: A scoping review.

BACKGROUND: The ketogenic diet (KD) is a high fat, moderate protein and very low carbohydrate diet. It can be used as a medical treatment for drug-resistant epilepsy (DRE), glucose transporter 1 deficiency syndrome and pyruvate dehydrogenase deficiency. The aim of this scoping review was to map the KD literature, with a focus on epilepsy and associated metabolic conditions, to summarise the current evidence-base and identify any gaps. METHODS: This review was conducted using JBI scoping review methodological guidance and the PRISMA extension for scoping reviews reporting guidance. A comprehensive literature search was conducted in September 2021 and updated in February 2024 using MEDLINE, CINAHL, AMED, EmBASE, CAB Abstracts, Scopus and Food Science Source databases. RESULTS: The initial search yielded 2721 studies and ultimately, data were extracted from 320 studies that fulfilled inclusion criteria for the review. There were five qualitative studies, and the remainder were quantitative, including 23 randomised controlled trials (RCTs) and seven quasi-experimental studies. The USA published the highest number of KD studies followed by China, South Korea and the UK. Most studies focused on the classical KD and DRE. The studies key findings suggest that the KD is efficacious, safe and tolerable. CONCLUSIONS: There are opportunities available to expand the scope of future KD research, particularly to conduct high-quality RCTs and further qualitative research focused on the child's needs and family support to improve the effectiveness of KDs.

A randomized, double-blind trial of triheptanoin for drug-resistant epilepsy in glucose transporter 1 deficiency syndrome.

OBJECTIVE: This study was undertaken to evaluate efficacy and long-term safety of triheptanoin in patients >1 year old, not on a ketogenic diet, with drug-resistant seizures associated with glucose transporter 1 deficiency syndrome (Glut1DS). METHODS: UX007G-CL201 was a randomized, double-blind, placebo-controlled trial. Following a 6-week baseline period, eligible patients were randomized 3:1 to triheptanoin or placebo. Dosing was titrated to 35% of total daily calories over 2 weeks. After an 8-week placebo-controlled period, all patients received open-label triheptanoin through Week 52. RESULTS: The study included 36 patients (15 children, 13 adolescents, eight adults). A median 12.6% reduction in overall seizure frequency was observed in the triheptanoin arm relative to baseline, and a 13.5% difference was observed relative to placebo (p = .58). In patients with absence seizures only (n = 9), a median 62.2% reduction in seizure frequency was observed in the triheptanoin arm relative to baseline. Only one patient with absence seizures only was present in the control group, preventing comparison. No statistically significant differences in seizure frequency were observed. Common treatment-emergent adverse events included diarrhea, vomiting, abdominal pain, and nausea, mostly mild or moderate in severity. No serious adverse events were considered to be treatment related. One patient discontinued due to status epilepticus. SIGNIFICANCE: Triheptanoin did not significantly reduce seizure frequency in patients with Glut1DS not on the ketogenic diet. Treatment was associated with mild to moderate gastrointestinal treatment-related events; most resolved following dose reduction or interruption and/or medication for treatment. Triheptanoin was not associated with any long-term safety concerns when administered at dose levels up to 35% of total daily caloric intake for up to 1 year.

Mechanistic Insights into Protein Stability and Self-aggregation in GLUT1 Genetic Variants Causing GLUT1-Deficiency Syndrome.

Human sodium-independent glucose cotransporter 1 (hGLUT1) has been studied for its tetramerization and multimerization at the cell surface. Homozygous or compound heterozygous mutations in hGLUT1 elicit GLUT1-deficiency syndrome (GLUT1-DS), a metabolic disorder, which results in impaired glucose transport into the brain. The reduced cell surface expression or loss of function have been shown for some GLUT1 mutants. However, the mechanism by which deleterious mutations affect protein structure, conformational stability and GLUT1 oligomerization is not known and require investigation. In this review, we combined previous knowledge of GLUT1 mutations with hGLUT1 crystal structure to analyze native interactions and several natural single-point mutations. The modeling of native hGLUT1 structure confirmed the roles of native residues in forming a range of side-chain interactions. Interestingly, the modeled mutants pointed to the formation of a variety of non-native novel interactions, altering interaction networks and potentially eliciting protein misfolding. Self-aggregation of the last part of hGLUT1 was predicted using protein aggregation prediction tool. Furthermore, an increase in aggregation potential in the aggregation-prone regions was estimated for several mutants suggesting increased aggregation of misfolded protein. Protein stability change analysis predicted that GLUT1 mutant proteins are unstable. Combining GLUT1 oligomerization behavior with our modeling, aggregation prediction, and protein stability analyses, this work provides state-of-the-art view of GLUT1 genetic mutations that could destabilize native interactions, generate novel interactions, trigger protein misfolding, and enhance protein aggregation in a disease state.

Gene therapy for Glut1-deficient mouse using an adeno-associated virus vector with the human intrinsic GLUT1 promoter.

BACKGROUND: We generated an adeno-associated virus (AAV) vector in which the human SLC2A1 gene, encoding glucose transporter type 1 (GLUT1), was expressed under the human endogenous GLUT1 promoter (AAV-GLUT1). We examined whether AAV-GLUT1 administration could lead to functional improvement in GLUT1-deficient mice. METHODS: We extrapolated human endogenous GLUT1 promoter sequences from rat minimal Glut1 promoter sequences. We generated a tyrosine-mutant AAV9/3 vector in which human SLC2A1-myc-DDK was expressed under the human GLUT1 promoter (AAV-GLUT1). AAV-GLUT1 was administered to GLUT1-deficient mice (GLUT1+/- mice) via intracerebroventricular injection (1.85 × 1010 vg/mouse or 6.5 × 1010 vg/mouse). We analyzed exogenous GLUT1 mRNA and protein expression in the brain and other major organs. We also examined improvements of cerebral microvasculature, motor function using rota-rod and footprint tests, as well as blood and cerebrospinal fluid (CSF) glucose levels. Additionally, we confirmed exogenous GLUT1 protein distribution in the brain and other organs after intracardiac injection (7.8 × 1011 vg/mouse). RESULTS: Exogenous GLUT1 protein was strongly expressed in the cerebral cortex, hippocampus and thalamus. It was mainly expressed in endothelial cells, and partially expressed in neural cells and oligodendrocytes. Motor function and CSF glucose levels were significantly improved following intracerebroventricular injection. Exogenous GLUT1 expression was not detected in other organs after intracerebroventricular injection of AAV-GLUT1, whereas it was detected in the liver and muscle tissue after intracardiac injection. CONCLUSIONS: Exogenous GLUT1 expression after AAV-GLUT1 injection approximated that of physiological human GLUT1 expression. Local central nervous system administration of AAV-GLUT1 improved CSF glucose levels and motor function of GLUT1-deficient mice and minimized off-target effects.

The role of SLC2A1 mutations in myoclonic astatic epilepsy and absence epilepsy, and the estimated frequency of GLUT1 deficiency syndrome.

The first mutations identified in SLC2A1, encoding the glucose transporter type 1 (GLUT1) protein of the blood-brain barrier, were associated with severe epileptic encephalopathy. Recently, dominant SLC2A1 mutations were found in rare autosomal dominant families with various forms of epilepsy including early onset absence epilepsy (EOAE), myoclonic astatic epilepsy (MAE), and genetic generalized epilepsy (GGE). Our study aimed to investigate the possible role of SLC2A1 in various forms of epilepsy including MAE and absence epilepsy with early onset. We also aimed to estimate the frequency of GLUT1 deficiency syndrome in the Danish population. One hundred twenty patients with MAE, 50 patients with absence epilepsy, and 37 patients with unselected epilepsies, intellectual disability (ID), and/or various movement disorders were screened for mutations in SLC2A1. Mutations in SLC2A1 were detected in 5 (10%) of 50 patients with absence epilepsy, and in one (2.7%) of 37 patient with unselected epilepsies, ID, and/or various movement disorders. None of the 120 MAE patients harbored SLC2A1 mutations. We estimated the frequency of SLC2A1 mutations in the Danish population to be approximately 1:83,000. Our study confirmed the role of SLC2A1 mutations in absence epilepsy with early onset. However, our study failed to support the notion that SLC2A1 aberrations are a cause of MAE without associated features such as movement disorders.

Association between cerebrospinal fluid parameters and developmental and neurological status in glucose transporter 1 deficiency syndrome.

OBJECTIVE: In glucose transporter 1 deficiency syndrome (Glut1DS), cerebrospinal fluid glucose (CSFG) and CSFG to blood glucose ratio (CBGR) show significant differences among groups classified by phenotype or genotype. The purpose of this study was to investigate the association between these biochemical parameters and Glut1DS severity. METHODS: The medical records of 45 patients who visited Osaka University Hospital between March 2004 and December 2021 were retrospectively examined. Neurological status was determined using the developmental quotient (DQ), assessed using the Kyoto Scale of Psychological Development 2001, and the Scale for the Assessment and Rating of Ataxia (SARA). CSF parameters included CSFG, CBGR, and CSF lactate (CSFL). RESULTS: CSF was collected from 41 patients, and DQ and SARA were assessed in 24 and 27 patients, respectively. Simple regression analysis showed moderate associations between neurological status and biochemical parameters. CSFG resulted in a higher R2 than CBGR in these analyses. CSF parameters acquired during the first year of life were not comparable to those acquired later. CSFL was measured in 16 patients (DQ and SARA in 11 and 14 patients, respectively). Although simple regression analysis also showed moderate associations between neurological status and CSFG and CSFL, the multiple regression analysis for DQ and SARA resulted in strong associations through the use of a combination of CSFG and CSFL as explanatory variables. CONCLUSION: The severity of Glut1DS can be predicted from CSF parameters. Glucose and lactate are independent contributors to the developmental and neurological status in Glut1DS.

Establishment of a flow cytometry screening method for patients with glucose transporter 1 deficiency syndrome.

Objective: We assessed the usefulness of flow cytometry as a functional assay to measure glucose transporter 1 (GLUT1) levels on the surface of red blood cells (RBCs) from Japanese patients with glucose transporter 1 deficiency syndrome (Glut1DS). Methods: We recruited 13 genetically confirmed Glut1DS patients with a solute carrier family 2 member 1 (SLC2A1) mutation (eight missense, one frameshift, two nonsense, and two deletion) and one clinically suspected Glut1DS-like patient without an SLC2A1 mutation, and collected whole blood with informed consent. We stained pelleted RBCs (1 µL) from the patients with a Glut1.RBD ligand and anti-glycophorin A antibody, which recognizes a human RBC membrane protein, and analyzed the cells using flow cytometry. Results: Relative GLUT1 levels quantified by flow cytometry in 11 of 13 patients with definite Glut1DS were 90% below those of healthy controls. Relative GLUT1 levels were not reduced in two of 13 Glut1DS patients who had a missense mutation and no intellectual disability and one Glut1DS-like patient without an SLC2A1 mutation. Relative GLUT1 levels were significantly reduced in Glut1DS patients with an SLC2A1 mutation, more severe intellectual disability, and spasticity. Conclusions: This method to detect GLUT1 levels on RBCs is simple and appears to be an appropriate screening assay to identify severe Glut1DS patients in the early stage before the development of irreversible neurologic damage caused by chronic hypoglycorrhachia.

Head circumference in glucose transporter 1 deficiency syndrome: Normal for individuals, abnormal as a group.

In the literature, microcephaly is considered as part of the classical phenotype of glucose transporter 1 deficiency syndrome (GLUT1DS), and previous cohort studies reported a prevalence of microcephaly of around 50%. In our clinical experience, however, only very few patients with GLUT1DS appear to have microcephaly. Therefore, we conducted an observational study among a large cohort of Dutch patients with GLUT1DS to investigate the prevalence of microcephaly, defined as < 2 standard deviations (SD) below the mean. We analysed the head circumference of 54 patients and found a prevalence of microcephaly at last known measurement of 6.5%. Notably, none of the patients had a head circumference < -3 SD. However, we learned that 75.9% of the patients had a head circumference below 0 SD. This study shows that microcephaly occurs less often than previously thought in patients with GLUT1DS, and that primary or secondary microcephaly does not seem to be a sign for clinicians to suspect GLUT1DS. As a group, however, patients with GLUT1DS seem to have decreased head circumference compared to healthy individuals and as such, our study suggests that early brain development and brain growth may be compromised in GLUT1DS.

Quantitative Three-Dimensional Gait Evaluation in Patients With Glucose Transporter 1 Deficiency Syndrome.

BACKGROUND: Of the patients with glucose transporter 1 deficiency syndrome (GLUT1-DS), 90% have a pathologic gait. Ataxic-spastic and ataxic gaits are seen in 35% of patients each. A ketogenic diet and modified Atkins diet (MAD) are effective therapy in GLUT1-DS in terms of both the seizures and movement disorder. A three-dimensional gait analysis (3DGA) system can be used to evaluate gait quantitatively using spatiotemporal data and gait kinematics. We performed 3DGA in three ambulatory patients with GLUT1-DS to evaluate the characteristics of their gait pathology, and we compared the gait variables before and after enhancing the MAD in one patient. METHODS: After examination by pediatric neurologists and pediatric orthopedic surgeons, 3DGA was performed. We assessed walking speed, step length, step width, gait variability, Gait Deviation Index (GDI), Gait Profile Score (GPS), and Gait Variable Score (GVS). RESULTS: All three patients had a low GDI and high GPS, comprehensive indices of gait pathology. The unstable gait pattern featured a wide step width in one patient and high gait variability in two patients. In the sagittal plane, the patients had increased GVSs in the knee and ankle joints due to excessive knee flexion or extension and excessive ankle plantarflexion. In the horizontal plane, the patients had increased GVSs in the pelvis, hips, and foot due to excessive rotation during walking. After enhancing the MAD, GDI, GPS, and GVSs improved. CONCLUSIONS: 3DGA has potential for quantifying the characteristics of gait pathology and its improvement with dietary therapy in patients with GLUT1-DS.

The role of molecular analysis of SLC2A1 in the diagnostic workup of glucose transporter 1 deficiency syndrome.

The study aimed to investigate the role of molecular analysis of SLC2A1 in the diagnostic workup of glucose transporter 1 deficiency syndrome (Glut1DS). During 2006-2020, we received 100 requests for SLC2A1 variant analysis of patients clinically suspected for Glut1DS. Pathogenic variants were detected in 37 patients, among whom 11 were familial cases. Most patients presented with epilepsy (n = 31; 84%), movement disorders (MD) (n = 28; 76%), and intellectual disabilities (ID) (n = 29; 78%). Moreover, paroxysmal dyskinesias (PD) (n = 10; 27%) were more frequently seen in familial cases (55%) than in sporadic cases (15%) (p < .05). The Glut1DS patients with ID typically had either epilepsy or MD. The presence of MD, particularly when associated with epilepsy or ID, indicated Glut1DS (p < .05). The cerebrospinal fluid (CSF) glucose levels were at or below the 10th percentile in all 32 SLC2A1-positive patients but only in 16 of 52 (31%) SLC2A1-negative patients (p < .05). Thus, CSF analysis is an essential tool in the diagnostic workup of Glut1DS. SLC2A1 molecular analysis should be performed in patients with a family history of Glut1DS or with at least one of the following clinical features, such as epilepsy, MD, and PD with or without ID, and low CSF glucose level. This would help in precise molecular diagnosis of the disease and facilitate effective treatment and appropriate genetic counseling.

Developmental outcomes and prevalence of SLC2A1 variants in young infants with hypoglycorrhachia.

INTRODUCTION: The neurodevelopmental outcomes of young infants with hypoglycorrhachia that is comparable to glucose transporter 1 deficiency syndrome (GLUT1DS), i.e. cerebrospinal fluid (CSF) glucose ≤40 mg/dL and CSF lactate <2.2 mM without causes of secondary hypoglycorrhachia are unknown. This study investigated the developmental outcomes and possibility of GLUT1DS in infants with hypoglycorrhachia, or low CSF glucose concentration. MATERIAL AND METHODS: 1655 neurologically asymptomatic infants aged <4 months had CSF examinations for fever workup from 2006 to 2016. Among the infants with normal CSF cell counts and without isolated pathogens, there were hypoglycorrhachia group who had CSF glucose levels that were comparable to GLUT1DS, and age- and gender-matched non-hypoglycorrhachia group. Both groups were at a mean age of 5.9 ±â€¯2.4 years (ranged 1-10 years) at neurodevelopmental evaluation in 2017. Mutational analysis of solute-carrier-family 2, which facilitated the glucose transporter member 1 (SLC2A1) gene was performed. RESULTS: Among the 722 infants with normal CSF cell counts and without isolated pathogens, 30 (4.2%) had hypoglycorrhachia that was comparable to GLUT1DS. In the 25 infants with hypoglycorrhachia available for follow-up, 4 (16%) had abnormal outcomes, of which 3 (12%) had the history of mixed-type developmental delay before age 6 and 1 (4%) had type 1 diabetes mellitus. In the non-hypoglycorrhachia control group (n = 50), 2 patients (4%) showed abnormal outcomes, both with the history of pure speech delay. The hypoglycorrhachia group had a higher rate of the history of mixed-type of developmental delay than the control group (12% vs. 0%, P = 0.034). No SLC2A1 pathogenic variants were observed in the hypoglycorrhachia group. CONCLUSION: Hypoglycorrhachia may be a potential biomarker for neurodevelopmental delay instead of for GLUT1DS in neurologically asymptomatic young infants.

Food and Food Products on the Italian Market for Ketogenic Dietary Treatment of Neurological Diseases.

The ketogenic diet (KD) is the first line intervention for glucose transporter 1 deficiency syndrome and pyruvate dehydrogenase deficiency, and is recommended for refractory epilepsy. It is a normo-caloric, high-fat, adequate-protein, and low-carbohydrate diet aimed at switching the brain metabolism from glucose dependence to the utilization of ketone bodies. Several variants of KD are currently available. Depending on the variant, KDs require the almost total exclusion, or a limited consumption of carbohydrates. Thus, there is total avoidance, or a limited consumption of cereal-based foods, and a reduction in fruit and vegetable intake. KDs, especially the more restrictive variants, are characterized by low variability, palatability, and tolerability, as well as by side-effects, like gastrointestinal disorders, nephrolithiasis, growth retardation, hyperlipidemia, and mineral and vitamin deficiency. In recent years, in an effort to improve the quality of life of patients on KDs, food companies have started to develop, and commercialize, several food products specific for such patients. This review summarizes the foods themselves, including sweeteners, and food products currently available for the ketogenic dietary treatment of neurological diseases. It describes the nutritional characteristics and gives indications for the use of the different products, taking into account their metabolic and health effects.

Short-term impact of a classical ketogenic diet on gut microbiota in GLUT1 Deficiency Syndrome: A 3-month prospective observational study.

BACKGROUND&AIMS: The classical ketogenic diet (KD) is a high-fat, very low-carbohydrate normocaloric diet used for drug-resistant epilepsy and Glucose Transporter 1 Deficiency Syndrome (GLUT1 DS). In animal models, high fat diet induces large alterations in microbiota producing deleterious effects on gut health. We carried out a pilot study on patients treated with KD comparing their microbiota composition before and after three months on the diet. METHODS: Six patients affected by GLUT1 DS were asked to collect fecal samples before and after three months on the diet. RT - PCR analysis was performed in order to quantify Firmicutes, Bacteroidetes, Bifidobacterium spp., Lactobacillus spp., Clostridium perfringens, Enterobacteriaceae, Clostridium cluster XIV, Desulfovibrio spp. and Faecalibacterium prausnitzii. RESULTS: Compared with baseline, there were no statistically significant differences at 3 months in Firmicutes and Bacteroidetes. However fecal microbial profiles revealed a statistically significant increase in Desulfovibrio spp. (p = 0.025), a bacterial group supposed to be involved in the exacerbation of the inflammatory condition of the gut mucosa associated to the consumption of fats of animal origin. CONCLUSIONS: A future prospective study on the changes in gut microbiota of all children with epilepsy started on a KD is warranted. In patients with dysbiosis demonstrated by fecal samples, it my be reasonable to consider an empiric trial of pre or probiotics to potentially restore the «ecological balance¼ of intestinal microbiota.

Novel mutation in a patient with late onset GLUT1 deficiency syndrome.

Glucose transporter 1 deficiency syndrome (GLUT1-DS) is an inborn error of metabolism caused by impaired glucose transport through blood brain barrier due to mutation in SLC2A1 gene, encoding transporter protein. Clinical spectrum includes various signs and symptoms, ranging from severe epileptic encephalopathy to movement disorders. The diagnosis of GLUT1-DS requires hypoglycorrhachia in the presence of normoglycaemia with a reduced cerebrospinal fluid (CSF):plasma glucose ratio. The absence of pathogenic mutation in SLC2A1 gene does not exclude the diagnosis. This case report describes a patient with late onset GLUT1-DS with a novel sporadic mutation c.539T>A, p.Met180Lys in exon 5 of the SLC2A1 gene. The dominating clinical features were epilepsy and paroxysmal dyskinesias provoked by infection, emotional stress and fasting. The ictal EEG was characterized by generalized paroxysmal 3-3.5Hz spike-slow wave complexes (absences). Treatment with ketogenic diet showed clinical improvement with the reduction of paroxysmal dyskinesias.

Long-term effects of a ketogenic diet on body composition and bone mineralization in GLUT-1 deficiency syndrome: a case series.

OBJECTIVE: The only known treatment of glucose transporter 1 deficiency syndrome (GLUT-1 DS) is a ketogenic diet (KD), which provides the brain with an alternative fuel. Studies in children with intractable epilepsy have shown that a prolonged KD can induce a progressive loss of bone mineral content associated with poor bone health status, probably as a consequence of a chronic acidic environment. The aim of this study is to determine the long-term effects of a KD on body composition and bone mineral status of patients with GLUT-1 DS, is currently unknown. METHODS: In this case series, we report the changes in body composition and bone mineral status observed in three adult patients with GLUT-1 DS who have been treated with a KD for more than 5 y. RESULTS: A long-term KD did not produce appreciable changes in weight and body composition of adults with GLUT-1 DS. Moreover, we found no evidence of potential adverse effects of a KD on bone health. In summary, this case series contributes to a small but growing body of literature that investigated the potential long-term effects of a KD on bone health. CONCLUSIONS: Our data suggest that maintaining a KD for more than 5 y does not pose any major negative effects on body composition, bone mineral content, and bone mineral density in adults with GLUT-1 DS, a finding that is at variance with previous reports focusing on children with intractable epilepsy. Further studies with larger sizes are needed to confirm and expand our findings.

Reversible white matter lesions during ketogenic diet therapy in glucose transporter 1 deficiency syndrome.

BACKGROUND: Glucose transporter type 1 deficiency syndrome is caused by brain energy failure resulting from a disturbance in glucose transport. PATIENTS: We describe a 4-year-old boy with classical type glucose transporter type 1 deficiency syndrome with a heterozygous splice acceptor site mutation (c.517-2A>G) in the SLCA2A1 gene. RESULTS: We initiated a ketogenic diet at 4 months of age. However, even though his condition was good during ketogenic diet therapy, multiple cerebral white matter and right cerebellum lesions appeared at 9 months of age. The lesions in the cerebral white matter subsequently disappeared, indicating that white matter lesions during diet therapy may be reversible and independent of the ketogenic diet. CONCLUSIONS: This is the first report of reversible white matter lesions during ketogenic diet therapy in glucose transporter type 1 deficiency syndrome.