Glycine Amidinotransferase Deficiency Research Articles

Creatine Deficiency Disorders: Phenotypes, Genotypes, Diagnosis, and Treatment Outcomes

Creatine is synthetized from arginine and glycine. There are two enzymes in the synthesis: l-arginine:glycine amidinotransferase and guanidinoacetate methyltransferase. After the synthesis, it is taken up by high-energy-requiring organs using creatine transporter. Biallelic pathogenic variants in GAMT result in guanidinoacetate methyltransferase deficiency and biallelic pathogenic variants in GATM result in l-arginine:glycine amidinotransferase deficiency. Hemizygous pathogenic variant in males and heterozygous pathogenic variant in females in SLC6A8 result in creatine transporter deficiency. Patients with these disorders present with a wide range of symptoms, including developmental delay, seizures, movement disorder, behavioral problems, and hypotonia. The diagnosis can be suspected by elevated guanidinoacetate and low creatine levels in body fluids in guanidinoacetate methyltransferase deficiency, low guanidinoacetate and low creatine levels in body fluids in l-arginine:glycine amidinotransferase deficiency, and elevated creatine-to-creatinine ratio in urine in creatine transporter deficiency in males as well as absent or significantly decreased creatine level in brain proton magnetic resonance spectroscopy. Genetic investigations such as targeted next-generation sequencing panel or exome sequencing can also identify these disorders; however, metabolite measurements and creatine in proton magnetic resonance spectroscopy are crucial to confirm the diagnosis. While all 3 disorders are currently treated with creatine supplementation, guanidinoacetate methyltransferase deficiency is also treated with ornithine supplementation and a protein- or arginine-restricted diet, and creatine transporter deficiency is treated with arginine and glycine supplementation (with no proven improvements).

P241 CARDIAC INVOLVEMENT IN CREATINE DEFICIENCY SYNDROME

Abstract Background Previous reports have demonstrated electrocardiographic and echocardiographic abnormalities consistent with early–stage cardiomyopathy, in patients with creatine transporter deficiency (CRT–D), but cardiac involvement has never been accurately characterized. Our aim was to widen the current knowledge of the cardiac phenotype associated with creatine deficiency in a murine transgenic model and in patients with creatine deficiency syndrome (CDS). Methods Wild type (n = 6) and transgenic CRT–/y mice (n = 6) were evaluated in vivo by electrocardiogram (EKG) and echocardiography for six months, and ex vivo by histological and biochemical analyses. Nine patients with CDS (4 with L–arginine:glycine amidinotransferase deficiency – AGAT–D, 5 with CRT–D) underwent blood samples, including cardiac biomarkers, EKG, Holter monitoring, echocardiography and cardiac magnetic resonance (CMR). Results Compared to wild–type, CRT–/y mice showed a prolongation of QT interval (p = 0.008) (Fig. 1), as well as a trend of reduction of mitochondrial function (Fig. 2). No differences were dected in the left ventricular systolic function, in terms of ejection fraction and global longitudinal strain, at different times. Aldosterone levels were also not different between the two groups. Three patients with CDS also showed a prolonged QTc interval (median QTc 515 ms), while other 3 had a borderline QTc at Holter monitoring, without tachyarrhythmias. No subject had abnormal systolic and diastolic function. At CMR, an increase of native T1 value was reported in 4 subjects (median 1076 ms), 1 with an increase of extracellular volume matching with the detection of late gadolinium enhancement. Finally, 7 subjects with CDS presented with biohumoral signs of neurohormonal activation (2 with elevated renin, 1 with increased aldosterone and 4 with increased norepinephrine levels). Familiar carriers (3 for AGAT–D and 1 for CRT–D) showed no notable cardiac functional alterations, except for QTc prolongation in 2 AGAT–D carriers. Conclusion Prolonged QT interval is observed in a murine model of CRT–D deficiency and in patients with CDS, in absence of an arrhythmic burden. No alterations in cardiac function were detected in the two setting, except for an increase of native T1 in patients with CDS suggesting an interstitial and replacement fibrosis.

Increased creatine demand during pregnancy in Arginine: Glycine Amidino-Transferase deficiency: a case report

BackgroundCreatine (Cr), an amino acid derivative, is one of the most important sources of energy acting as both a spatial and temporal energy buffer through its phosphorylated analogue phosphocreatine (PCr) and creatine kinase (CK). Maternal Cr biosynthesis and metabolism seem to play an important role in pregnancy, as shown in preclinical and in healthy human pregnancy studies. Patients with Arginine:Glycine Amidino-Transferase deficiency (AGAT-d), due to the deficit of the first enzyme involved in Cr synthesis, are at a disadvantage due to their failure to synthesize Cr and their dependence on external intake, in contrast to normal subjects, where changes in Cr biosynthesis supply their needs.We report the outcomes of a pregnancy in an AGAT-d woman, and the challenge we faced in managing her treatment with oral Cr to ensure optimal conditions for her fetus.Case presentationA 22-year-old AGAT-d woman referred to our Institute for the management of her first conception at 11 weeks of fetal gestational age. Sonographic monitoring at 20 w GA indicated a reduction of fetal growth, in particular of the head circumference that was below the 3rd centile. Biochemical monitoring of Cr in biological fluids of the mother revealed a decline of the Cr concentrations, in particular in the urine sample, requiring prompt correction of the Cr dose. At 35 weeks of gestation the patient delivered a male infant, heterozygous for GATM mutation, with normal brain Cr levels; at one year the baby achieved typical developmental milestones.ConclusionsThis rare pregnancy demonstrates that Cr levels in the blood and urine of the mother with AGAT-d decreased since the first months of gestation. The increase of the Cr daily dose administered to the mother seems to have produced beneficial effects also on the fetus.

Homoarginine- and Creatine-Dependent Gene Regulation in Murine Brains with l-Arginine:Glycine Amidinotransferase Deficiency.

l-arginine:glycine amidinotransferase (AGAT) and its metabolites homoarginine (hArg) and creatine have been linked to stroke pathology in both human and mouse studies. However, a comprehensive understanding of the underlying molecular mechanism is lacking. To investigate transcriptional changes in cerebral AGAT metabolism, we applied a transcriptome analysis in brains of wild-type (WT) mice compared to untreated AGAT-deficient (AGAT−/−) mice and AGAT−/− mice with creatine or hArg supplementation. We identified significantly regulated genes between AGAT−/− and WT mice in two independent cohorts of mice which can be linked to amino acid metabolism (Ivd, Lcmt2), creatine metabolism (Slc6a8), cerebral myelination (Bcas1) and neuronal excitability (Kcnip3). While Ivd and Kcnip3 showed regulation by hArg supplementation, Bcas1 and Slc6a8 were creatine dependent. Additional regulated genes such as Pla2g4e and Exd1 need further evaluation of their influence on cerebral function. Experimental stroke models showed a significant regulation of Bcas1 and Slc6a8. Together, these results reveal that AGAT deficiency, hArg and creatine regulate gene expression in the brain, which may be critical in stroke pathology.

LC-MS/MS measurements of urinary guanidinoacetic acid and creatine: Method optimization by deleting derivatization step

BackgroundCerebral Creatine deficiency syndromes (CCDS) include three hereditary diseases affecting the metabolism of creatine (Cr): arginine glycine amidinotransferase deficiency, guanidinoacetate methyltransferase deficiency and disorders of creatine transporter. These pathologies cause a brain creatine deficiency responsible of non-specific neurological impairments with mental retardation. LC-MS/MS measurements of guanidinoacetic acid (GAA) and creatine in urine and plasma are an important screening test to identify the deficit. Analysis of this polar and basic molecules not hold on standard column requires a derivatization step to butyl-esters. To overcome this long and fastidious derivatization, an ion pairing (IP) method was chosen in this study. MethodIP method was validated using Comité francais d'accréditation (COFRAC) recommendations. Then, urine GAA and creatine of 15 patients with a CDS deficiency suspected were tested y LC-MS/MS using IP technique, and performances were assessed with reference laboratory method (butylation method). Moreover, references values were suggested y the study of 100 urines samples of healthy patients. ResultsThe method developed provided a good accuracy and precision with intra and inter-day coefficients of variation (CVs) <15%. The curve was linear for the biological and pathological concentrations. The comparison with the reference method did not reveal any significant difference for analytical performances but showed a simplification of the preparation of samples. ConclusionThe use of IP technique that we have developed demonstrated a good correlation with the butylation method. Moreover, this new method not only allows a simplification of the technique, but also decreases in run time.

Fifteen-year follow-up of Italian families affected by arginine glycine amidinotransferase deficiency

BackgroundArginine:glycine amidinotransferase deficiency (AGAT-d) is a very rare inborn error of creatine synthesis mainly characterized by absence of brain Creatine (Cr) peak, intellectual disability, severe language impairment and behavioural disorder and susceptible to supplementary Cr treatment per os. Serial examinations by magnetic resonance spectroscopy are required to evaluate Cr recovery in brain during treatment of high doses of Cr per os, which have been proved beneficial and effective in treating main clinical symptoms.A long term study with detailed reports on clinical, neurochemical and neuropsychological outcomes of the first Italian patients affected by AGAT-d here reported can represent a landmark in management of this disorder thus enhancing medical knowledge and clinical practice.ResultsWe have evaluated the long term effects of Cr supplementation management in four Italian patients affected by AGAT-d, correlating specific treatments with serial clinical, biochemical and magnetic resonance spectroscopy examinations as well as the neuropsychological outcome by standardized developmental scales. Consecutive MRS examinations have confirmed that Cr depletion in AGAT-d patients is reversible under Cr supplementation. Cr treatment is considered safe and well tolerated but side effects, including weight gain and kidney stones, have been reported.ConclusionsEarly treatment prevents adverse developmental outcome, while patients diagnosed and treated at an older age showed partial but significant cognitive recovery with clear improvements in adaptive functioning.

L-arginine:glycine amidinotransferase deficiency protects from metabolic syndrome

L-arginine:glycine amidinotransferase deficiency protects from metabolic syndrome

L-arginine:glycine amidinotransferase deficiency protects from metabolic syndrome

Phosphorylated creatine (Cr) serves as an energy buffer for ATP replenishment in organs with highly fluctuating energy demand. The central role of Cr in the brain and muscle is emphasized by severe neurometabolic disorders caused by Cr deficiency. Common symptoms of inborn errors of creatine synthesis or distribution include mental retardation and muscular weakness. Human mutations in l-arginine:glycine amidinotransferase (AGAT), the first enzyme of Cr synthesis, lead to severely reduced Cr and guanidinoacetate (GuA) levels. Here, we report the generation and metabolic characterization of AGAT-deficient mice that are devoid of Cr and its precursor GuA. AGAT-deficient mice exhibited decreased fat deposition, attenuated gluconeogenesis, reduced cholesterol levels and enhanced glucose tolerance. Furthermore, Cr deficiency completely protected from the development of metabolic syndrome caused by diet-induced obesity. Biochemical analyses revealed the chronic Cr-dependent activation of AMP-activated protein kinase (AMPK), which stimulates catabolic pathways in metabolically relevant tissues such as the brain, skeletal muscle, adipose tissue and liver, suggesting a mechanism underlying the metabolic phenotype. In summary, our results show marked metabolic effects of Cr deficiency via the chronic activation of AMPK in a first animal model of AGAT deficiency. In addition to insights into metabolic changes in Cr deficiency syndromes, our genetic model reveals a novel mechanism as a potential treatment option for obesity and type 2 diabetes mellitus.

Creatine metabolism in urea cycle defects

Creatine (Cr) and phosphocreatine play an essential role in energy storage and transmission. Maintenance of creatine pool is provided by the diet and by de novo synthesis, which utilizes arginine, glycine and s-adenosylmethionine as substrates. Three primary Cr deficiencies exists: arginine:glycine amidinotransferase deficiency, guanidinoacetate methyltransferase deficiency and the defect of Cr transporter SLC6A8. Secondary Cr deficiency is characteristic of ornithine-aminotransferase deficiency, whereas non-uniform Cr abnormalities have anecdotally been reported in patients with urea cycle defects (UCDs), a disease category related to arginine metabolism in which Cr must be acquired by de novo synthesis because of low dietary intake. To evaluate the relationships between ureagenesis and Cr synthesis, we systematically measured plasma Cr in a large series of UCD patients (i.e., OTC, ASS, ASL deficiencies, HHH syndrome and lysinuric protein intolerance). Plasma Cr concentrations in UCDs followed two different trends: patients with OTC and ASS deficiencies and HHH syndrome presented a significant Cr decrease, whereas in ASL deficiency and lysinuric protein intolerance Cr levels were significantly increased (23.5 vs. 82.6 μmol/L; p < 0.0001). This trend distribution appears to be regulated upon cellular arginine availability, highlighting its crucial role for both ureagenesis and Cr synthesis. Although decreased Cr contributes to the neurological symptoms in primary Cr deficiencies, still remains to be explored if an altered Cr metabolism may participate to CNS dysfunction also in patients with UCDs. Since arginine in most UCDs becomes a semi-essential aminoacid, measuring plasma Cr concentrations might be of help to optimize the dose of arginine substitution.

Developmental progress and creatine restoration upon long-term creatine supplementation of a patient with arginine:glycine amidinotransferase deficiency

BackgroundArginine:glycineamidinotransferase (AGAT/GATM) deficiency has been described in 9 patients across 4 families. Here we describe the clinical outcome and response to creatine supplementation in a patient of the second family affected with AGAT deficiency—a 9-year-old girl. Patient and methodsDelayed motor milestones were noticed from 4months of age and at 14months moderate hypotonia, developmental delay and failure to thrive. Laboratory studies revealed low plasma creatine as well as extremely low levels of guanidinoacetic acid in urine and plasma. Proton magnetic resonance spectroscopy (MRS) of the brain showed absence of creatine. DNA sequence analysis revealed a homozygous mutation (c.484+1G>T) in the AGAT/GATM gene. AGAT activity was not detectable in lymphoblasts and RNA analysis revealed a truncated mRNA (r.289_484del196) that is degraded via Nonsense Mediated Decay. At 16months, Bayley's Infant Development Scale (BIDS) showed functioning at 43% of chronologic age. Oral creatine supplementation (up to 800mg/kg/day) was begun. ResultsAt age 9years she demonstrated advanced academic performance. Partial recovery of cerebral creatine levels was demonstrated on MRS at 25months of age. Brain MRS at 40months of age revealed a creatine/NAA ratio of about 80% of that in age-matched controls. Conclusions8years post initiation of oral creatine supplementation, patient demonstrates superior nonverbal and academic abilities, with average verbal skills. We emphasize that early diagnosis combined with early treatment onset of AGAT deficiency may lead to improvement of developmental outcome.

Arginine:glycine amidinotransferase (AGAT) deficiency in a newborn: Early treatment can prevent phenotypic expression of the disease

Arginine:glycine amidinotransferase deficiency is a treatable inborn error of creatine synthesis, characterized by mental retardation, language impairment, and behavioral disorders. We describe a patient in whom arginine:glycine amidinotransferase was diagnosed at birth and treated at 4 months with creatine supplementation. In contrast with his 2 older sisters, he had normal psychomotor development at 18 months.

Arginine:Glycine Amidinotransferase Deficiency: The Third Inborn Error of Creatine Metabolism in Humans

Arginine:glycine amidinotransferase (AGAT) catalyzes the first step of creatine synthesis, resulting in the formation of guanidinoacetate, which is a substrate for creatine formation. In two female siblings with mental retardation who had brain creatine deficiency that was reversible by means of oral creatine supplementation and had low urinary guanidinoacetate concentrations, AGAT deficiency was identified as a new genetic defect in creatine metabolism. A homozygous G-A transition at nucleotide position 9297, converting a tryptophan codon (TGG) to a stop codon (TAG) at residue 149 (T149X), resulted in undetectable cDNA, as investigated by reverse-transcription PCR, as well as in undetectable AGAT activity, as investigated radiochemically in cultivated skin fibroblasts and in virus-transformed lymphoblasts of the patients. The parents were heterozygous for the mutant allele, with intermediate residual AGAT activities. Recognition and treatment with oral creatine supplements may prevent neurological sequelae in affected patients.