Urinary Orotic Acid Research Articles

Mutation delTTCT 1471 de l’intolérance aux protéines dibasiques : Caractéristiques cliniques d’une série pédiatrique tunisienne.

Lysinuric protein intolerance (LPI) is a rare inherited metabolic disease. It is caused by a deficiency in cationic amino acid transport caused by mutations in SLC7A7 gene. To identify the clinical, diagnostic and therapeutic features of lysnuric protein intolerance. This was a retrospective study conducted in the pediatric department of La Rabta Hospital over a period of 30 years (1992 to 2022). We included patients with clinical signs suggestive of lysinuric protein intolerance and orotic acid in the urine. We enrolled seven patients. The median age at disease onset was nine months. The median age at positive diagnosis was 21 months. Growth retardation, hepatosplenomegaly and haematological abnormalities were the main features of the disease. Hyperammonia and increased urinary orotic acid were present in all patients. Molecular biology revealed the del TTCT 1471 mutation in five patients. All patients were prescribed a low protein diet and citrulline supplementation. Complications of the disease were growth retardation (n=7), psychomotor or intellectual retardation (n=5), haemophagocytic lymphohistiocytosis (n=4) and osteoporosis (n=3). After a median follow-up of 11 years, six of our patients are still alive. One patient died from acute hyperammonemic encephalopathy. In this paediatric series, delays in diagnosis and treatment of LPI were responsible for long-term sequelae, particularly bone and neurological. The delTTCT1471 mutation appears to be the mutation of paediatric-onset forms in Tunisia. This mutation was not associated with pulmonary involvement, which is a prognostic factor and the main cause of death.

Validating urinary orotic acid through high-pressure liquid chromatography (HPLC): Insights from a specialized biochemical genetics laboratory in Pakistan

Validating urinary orotic acid through high-pressure liquid chromatography (HPLC): Insights from a specialized biochemical genetics laboratory in Pakistan

The efficacy of Carbamylglutamate impacts the nutritional management of patients with N-Acetylglutamate synthase deficiency

BackgroundThe autosomal recessive disorder N-acetylglutamate synthase (NAGS) deficiency is the rarest defect of the urea cycle, with an incidence of less than one in 2,000,000 live births. Hyperammonemic crises can be avoided in individuals with NAGS deficiency by the administration of carbamylglutamate (also known as carglumic acid), which activates carbamoyl phosphatase synthetase 1 (CPS1). The aim of this case series was to introduce additional cases of NAGS deficiency to the literature as well as to assess the role of nutrition management in conjunction with carbamylglutamate therapy across new and existing cases.MethodsWe conducted retrospective chart reviews of seven cases of NAGS deficiency in the US and Canada, focusing on presentation, diagnosis, medication management, nutrition management, and outcomes.ResultsFive new and two previously published cases were included. Presenting symptoms were consistent with previous reports. Diagnostic confirmation via molecular testing varied in protocol across cases, with consecutive single gene tests leading to long delays in diagnosis in some cases. All patients responded well to carbamylglutamate therapy, as indicated by normalization of plasma ammonia and citrulline, as well as urine orotic acid in patients with abnormal levels at baseline. Although protein restriction was not prescribed in any cases after carbamylglutamate initiation, two patients continued to self-restrict protein intake. One patient experienced two episodes of hyperammonemia that resulted in poor long-term outcomes. Both episodes occurred after a disruption in access to carbamylglutamate, once due to insurance prior authorization requirements and language barriers and once due to seizure activity limiting the family’s ability to administer carbamylglutamate.ConclusionsFollow-up of patients with NAGS deficiency should include plans for illness and for disruption of carbamylglutamate access, including nutrition management strategies such as protein restriction. Carbamylglutamate can help patients with NAGS deficiency to liberalize their diets, but the maximum safe level of protein intake to prevent hyperammonemia is not yet known. Patients using this medication should still monitor their diet closely and be prepared for any disruptions in medication access, which might require immediate dietary adjustments or medical intervention to prevent hyperammonemia.

Genetic analysis and prenatal diagnosis for a Chinese pedigree affected with co-morbid Ornithine carbamoyl transferase deficiency and MECP2 duplication syndrome

To explore the genetic basis for a Chinese pedigree affected with co-morbid Ornithine carbamoyl transferase deficiency (OTCD) and MECP2 duplication syndrome. A proband who was admitted to the Neonatal Intensive Care Unit of Gansu Provincial Maternal and Child Health Care Hospital on December 19, 2017 was selected as the study subject. High-throughput sequencing and multiplex ligation-dependent probe amplification (MLPA) were carried out for her pedigree, and short tandem repeat-based linkage analysis and chromosome copy number variation sequencing (CNV-seq) were used for the prenatal diagnosis. The proband, a 3-day-old female, was found to harbor heterozygous deletion of exons 7-9 of the OTC gene. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the variant was classified as likely pathogenic (PVS1+PM2_Supporting+PP4). The proband was diagnosed with OTCD , which was in keeping with her acute encephalopathy and metabolic abnormalities (manifesting as hyperammonemia, decreased blood citrulline, and increased urine orotic acid). Prenatal diagnosis was carried out for the subsequent pregnancy. The fetus did not harbor the exons 7-9 deletion of the OTC gene, but was found to carry a duplication in Xq28 region (which encompassed the whole region of MECP2 duplication syndrome) and was positive for the SRY sequence. The same duplication was also found in the proband and her mother. Considering the possible existence of X-chromosome inactivation, the proband was diagnosed with two X-linked recessive disorders including OTCD and MECP2 duplication syndrome, and the fetus was determined as a male affected with the MECP2 duplication syndrome. Discoveries of the pathogenic variants underlying the OTCD and MECP2 duplication syndrome have enabled clinical intervention, treatment, genetic counseling and prenatal diagnosis for this pedigree.

The Strange Case of Orotic Acid: The Different Expression of Pyrimidines Biosynthesis in Healthy Males and Females.

Orotic acid (OA) is an intermediate metabolite of pyrimidine nucleotide biosynthesis and represents a minor diet constituent. The measurement of urinary orotic acid is useful in confirming the diagnosis of hereditary metabolic diseases. Moreover, it could be of interest to know how the physiological concentration of this metabolite changes in relation to different conditions of clinical normality. The purpose of this study was to determine the orotic acid concentration in the urine of healthy patients, to observe normal oroticuria and to evaluate if the expression of pyrimidine intermediate biosynthesis differs between healthy males and females. The orotic acid concentration in urine was performed via the ICH M10-validated analytical method. Unexpectedly, females showed a greater oroticuria than males in pediatric age (0-10); conversely, we did not find significant differences until 70 years of age. The LC-MS/MS method was suitable for use in the differential diagnosis of hereditary metabolic disease and metabolic monitoring of anticancer drug-induced toxicity. The analytical protocol was found to be rapid and ideal, and was used in the routine analysis of a clinical chemistry laboratory. The biochemical aspects related to the expression of pyrimidine biosynthesis should be further investigated in light of the obtained results.

Abstract 1017: A novel blood diagnostic method for fibrolamellar hepatocellular carcinoma

Abstract Fibrolamellar hepatocellular carcinoma (FLC) is an often-lethal liver cancer affecting primarily children and young adults. The disease presents with vague symptoms, and, as a result, it is usually diagnosed at an advanced stage. Thus, there is a high recurrence rate among those who undergo surgical resection. Diagnostic tests based on scans are differentially interpreted by pathologists and radiological, and scans often miss small tumors. We are developing tests for FLC based on metabolites and nucleic acids in the blood and in the urine. We used samples from 29 FLC patients and 9 age/gender-matched healthy donors. A clinical testing lab was used to quantify the plasma for Vitamin B12, alpha-fetoprotein (AFP), estrogen (E2), and liver panel (albumin, alkaline phosphatase, alanine transaminase, aspartate transferase, creatine kinase, direct bilirubin, total bilirubin, total protein, globulin). We found no difference between FLC vs. non-FLC subjects in the liver panel tests. In addition, there is no difference between the AFP and E2 tests. Vitamin B12 levels were about 8.5 times higher (p<0.05) in FLC patients compared to healthy donors. We are currently validating other protein and metabolite markers in the blood and urine, and C-reactive protein in blood and orotic acid in urine. In nucleic acid assays, we found specific miRNAs increased in the circulation. These included: miR-122-5p, miR-514a-5p, miR-192-5p, miR-615-5p, miR-4532, miR-4322, miR-214-3p, and miR-122-3p. They were upregulated 8- to 21-fold in FLC (n= 24) vs. non-FLC patients (n=16). These results identified biomarkers and microRNAs associated with FLC tumors. The tests are currently being evaluated as part of a larger study to determine whether they can detect recurrence of disease and help assess therapeutic responses in FLC patients. Citation Format: Ruisi Wang, Arlene M. Hurley, Sanford M. Simon. A novel blood diagnostic method for fibrolamellar hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1017.

Abstract 6729: Urea cycle metabolism is disturbed in Fibrolamellar carcinoma

Abstract In fibrolamellar hepatocellular carcinoma (FLC), hyperammonemic encephalopathy is a common occurrence and occasionally causes death. Using mass spectrometry, we quantitatively analyzed the proteomes of FLC patient’s tumor and adjacent normal, to find pathways that are changed in FLC. These data identified multiple proteins that were altered in the proteome, among these, enzymes involved in metabolism of ammonia. These results were confirmed with immunofluorescence demonstrating that these alterations occur in all tumor cells. These results suggest that FLC cells have defects in the two primary ammonia detoxification pathways in the liver, which are responsible for detoxification of 70% of the ammonia in the body: 1) consumption of ammonia by glutamine synthetase (GLUL), and 2) addition of ammonia by ornithine carbamoyltransferase (OTC) to the urea cycle. Additionally, they also generate extra ammonia because of upregulation of glutaminase (GLS). This was tested with a targeted metabolomics of the reactants and products of these enzymes. The results were consistent with both a loss of the two pathways for consumption of ammonia activation of a pathway for generating ammonia. This production of ammonia is consistent with the observation that surgical resection of fibrolamellar reduces the ammonia in patients. All FLC patients with hyperammonemic encephalopathy and documented urine test results showed increased urinary orotic acid, evidence of blockage of the OTC pathway. This study implies that hyperammonemic encephalopathy in FLC may require alternatives to commonly used therapies. Citation Format: Mahsa Shirani, Solomon Levin, Michael D. Tomasini, James Knox, Bassem Shebl, David Requena, Jackson Clark, Søren Heissel, Hanan Alwaseem, Rodrigo Surjan, Ron Lahasky, Henrik Molina, Barbara Lyons, Rachael D. Migler, Philip Coffino, Sanford M. Simon. Urea cycle metabolism is disturbed in Fibrolamellar carcinoma. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6729.

A curious case of confusion

A curious case of confusion

Hyperammonemia in a pregnant woman with citrullinemia type I: a case report and literature review

BackgroundCitrullinemia type I (CTLN1) is a rare urea cycle disorder (UCD) with few adult cases described so far. Diagnosis of late-onset CTLN1 is difficult, and delayed treatment may increase the risk of severe hyperammonemia. Pregnancy is an important risk factor for women with CTLN1. However, the clinical manifestations of CTLN1 in a pregnant woman may be mistaken for pregnancy side effects and ultimately delay a timely diagnosis.Case presentationA 34-year-old woman developed vomiting and disturbance of consciousness after 12 weeks of gestation. A blood test showed hyperammonemia (454 μg/dL) with normal liver function tests. She fell into a deep coma, and her serum ammonia level increased to 800 μg/dL. Continuous renal replacement therapy (CRRT) was administered as a diagnostic treatment for UCD and serum ammonia. This patient’s case was complicated by co-infection; her dependents decided to withdraw life support and the patient died. She was diagnosed with CTLN1 by analyses of plasma amino acids, urinary orotic acid, and second-generation gene sequencing.Discussion and conclusionWhen a patient displays symptoms of emesis and disturbance of consciousness in early pregnancy, blood ammonia should be monitored, and UCD should be considered, particularly for patients with hyperammonemia in the absence of severe liver function abnormalities.

314: EXTRACORPOREAL LIVER ASSIST DEVICE FOR ACUTE LIVER INJURY DUE TO LIKELY PARTIAL UREA CYCLE DISORDER

Introduction: Extracorporeal liver assist device (ELAD) is used to bridge patients in acute liver failure to transplant or in severe acute liver injury until spontaneous recovery. ELAD employs an albumin circuit to dialyze albumin-bound toxins from the blood stream. Urea is an albumin bound molecule formed in the liver as a result of nitrogen breakdown. Urea Cycle Disorders (UCDs) are a rare group of genetic enzymatic deficiencies that typically manifest early in life but partial enzymatic deficiencies can present later in life in times of metabolic stress. Description: A 38-year-old female presented with progressive encephalopathy 2 months after being diagnosed with alcoholic hepatitis at which time she had stopped drinking alcohol. She had no prior liver disease. Initial labs showed an elevated ammonia level of 87 mcmol/L, despite decrease in total bilirubin from 33 mg/dL to 3.5mg/dL, and INR of 1.9 to 1.4 since the peak of her alcoholic hepatitis. She had no imaging evidence of cirrhosis and had normal aminotransferases, platelets, and renal function. Initial head CT, infectious evaluation, and work-up for alternate etiologies of acute liver injury were unremarkable. She was intubated for progressive hepatic encephalopathy on hospital day 9. Her ammonia level significantly rose to 203 mcmol/L and a repeat head CT showed diffuse cerebral edema. Due to a rising ammonia level to 247 mcmol/L and CT evidence of progressive cerebral edema despite mannitol, therapeutic hyperventilation, and sedation. She was started on MARS (TM) therapy on hospital day 11. After one 7-hour session, her ammonia level decreased to 101 mcmol/L, mental status improved and head CT showed improved cerebral edema. A urinary orotic acid level was elevated at 1.8 mmol per mol of creatinine. Discussion: A partial UCD was suspected due to atypical time course of encephalopathy and hyperammonemia in the context of improving liver synthetic function and absence of cirrhosis. Genetic testing is pending however, a partial UCD is likely given the elevated urinary orotic acid. A single case report exists of confirmed acquired glutamine synthetase deficiency after orthotopic liver transplant that also improved after a single MARS session. Our report adds to the scarce literature on this topic.

Дефіцит орнітин-транскарбамілази - мало відоме орфанне захворювання в дитячій гепатології (клінічний випадок)

The article reviews literature and demonstrates personal observation of a preschool patient with ornithine transcarbamylase deficiency. This disease is characterized by an excessive accumulation of ammonia in the body, which leads to damage of the nervous system, gastrointestinal tract, and impaired liver function. It has a genetically determined mechanism of development associated with mutations in the OTC gene. Purpose - to study specific aspects of manifestation, clinical picture, course and diagnosis of ornithine transcarbamylase deficiency. Clinical case. Girl U. was admitted to the pediatric hepatology department with complaints of abdominal pain, periodic vomiting, accompanied by high ketones in urine test, irritability, tearfulness, emotional lability, and increased liver enzyme levels. Physical exam revealed hepatosplenomegaly. Bloodwork series showed transaminase levels 4-20 times ULN, hyperammonemia, and increased orotic acid in urine. A molecular genetic study was performed, and identified a variant of uncertain significance in the OTC gene associated with X-linked ornithine transcarbamylase deficiency. Metabolic therapy and protein-restricted nutrition were prescribed to prevent hyperammonemia. Conclusions. Transcarbamylase deficiency is a rare genetic disease characterized by excessive accumulation of ammonia in the body with typical clinical and laboratory symptoms. Detection of ammonia concentration and molecular genetic testing are crucial diagnostic components in children with liver diseases associated with other organs and systems damage. Untimely diagnosis and treatment correlates with the severity of the disease, the development of irreversible changes, disability and possible death in young age. The study was performed according to the principles of the Declaration of Helsinki. The informed consent of the child's parents was obtained for the conducted research. The authors declare no conflict of interest.

Proximal urea cycle defects are challenging to detect with newborn screening: Results of a prospective pilot study using post-analytical tools.

The purpose of this pilot project was to evaluate the efficacy of the Collaborative Integrated Laboratory Reports (CLIR) postanalytical tools from Mayo Clinic for detection of newborns with proximal urea cycle disorders (PUCD) in the Georgia newborn screening program that uses the underivatized Neobase2 kit (Perkin Elmer). We evaluated 138,560 newborn screening (NBS) samples (between 125,000 and 130,000 children) and used the CLIR result interpretation guidelines to stratify results. Children at higher risk of having a PUCD received follow-up services including confirmatory lab testing (ammonia, plasma amino acids, urine orotic acid) or a repeat NBS sample. We made multiple adjustments to our CLIR PUCD tool and to our follow-up algorithms in order to reduce false positives. Regardless, a high number of NBS samples resulted with false positives in part due to the glutamine peak also containing lysine. No children were diagnosed with a PUCD during our study period, and the Emory Genetics Metabolic Center is unaware of any children diagnosed outside of the NBS system during that time. Based on our experience, PUCD is not suitable for statewide NBS using Neobase2 and CLIR. Other methodologies that can separate glutamine from other amino acids may have better performance.

Organic Acidemias.

Organic Acidemias.

Hyperargininemia Due to Arginase 1 Deficiency: Variability in Clinical and Biochemical Presentations in Malaysian children.

Objective:Hyperargininemia due to Arginase 1 deficiency is a rare inborn error of the urea cycle with an incidence estimated at 1:950 000. It has typical severe and progressive abnormal neurological features with biochemical findings of hyperargininemia and hyperexcretion of orotic acid. The aim of our study is to review the clinical and biochemical presentations of 4 children diagnosed with Arginase 1 deficiency in Malaysia and compare with the literature review.Design and Methods:We retrospectively reviewed the medical records of 4 patients with molecularly confirmed Arginase 1 deficiency. Patients were identified from a selective high-risk screening of 51 682 symptomatic patients from January 2006 to December 2020.Results:Our patients exhibited heterogeneous clinical presentations with acute and progressive neurological abnormalities and varying degrees of plasma arginine and urine orotic acid excretions. Interestingly, an unusual hyperexcretion of homocitrulline was found in 3 patients.Conclusions:Hyperargininemia due to Arginase 1 deficiency can present acutely and hyperexcretion of homocitrulline can be an additional biochemical feature of Arginase 1 deficiency.

HYPERAMMONEMIC COMA IN A PATIENT WITH ORNITHINE TRANSCARBAMYLASE DEFICIENCY

TOPIC: Critical Care TYPE: Medical Student/Resident Case Reports INTRODUCTION: Ornithine transcarbamylase (OTC) deficiency is a rare, X-linked inherited disorder of the urea cycle affecting 1 in 80,000 live births. It is characterized by the inability to break down nitrogen to urea. The resulting hyperammonemia leads to lethargy, hypotonia, seizures and developmental delay if the illness manifests in infancy, and migraines, ataxia, hyperammonemic coma in adults. Early recognition and treatment of hyperammonemia is the key to prevent neurological disability in infancy and hyperammonemic coma leading to death in adults. CASE PRESENTATION: 46-year-old female with past medical history of partial OTC deficiency and complex partial seizures presented to the hospital with chief complaint of altered mental status. In the emergency department, vitals were remarkable for tachycardia. Exam was significant for altered mental status and a Glasgow Coma Scale of 8. Initial labs were significant for hyperammonemia (234 mcmol/L), hypernatremia (160 mmol/L) and abnormal urinary analysis with signs of infection. Patient was admitted to the intensive care unit followed by initiation of lactulose and rifaximin for treatment of hyperammonemia. She was also started on broad spectrum antibiotics for suspicion of urinary tract infection. Electrolyte abnormalities were gradually corrected. However, her condition kept on deteriorating to the point that she was not responding to verbal, tactile or painful stimuli. She was subsequently intubated on the second day of hospitalization due to failure to protect airway. Serial ammonia levels were monitored that kept on increasing. Electroencephalogram showed severe metabolic encephalopathy without seizures. Computerized tomography scan of the brain did not show acute intracranial process. At this point, nephrology was consulted, and patient underwent continuous renal replacement therapy with dramatic improvement of serum ammonia level and her neurological status. This was followed by successful extubation and a gradual return to baseline mental status. She was continued on lactulose, rifaximin and advised to remain on low protein diet. DISCUSSION: This case highlights the rare complication of hyperammonemic coma in an adult patient with history of OTC deficiency. It is an X-linked defect with variable onset and presentation depending on the sex and activity of the enzyme. Patients classically present with lethargy, seizures and hyperammonemia leading to metabolic encephalopathy, coma and death. Diagnosis involves serum amino acid quantitative analysis and urinary orotic acid concentration in urine. Low serum citrulline and elevated urinary orotic acid levels confirm the OTC deficiency. DNA sequencing-based mutation analysis can also be used as first line for diagnosis. Treatment involves rapidly decreasing serum ammonia with lactulose, rifaximin or dialysis. Prognosis depends on the age of presentation and duration of hyperammonemia. CONCLUSIONS: XXX. REFERENCE #1: Summar, Marshall. "Current strategies for the management of neonatal urea cycle disorders." The Journal of pediatrics 138, no. 1 (2001): S30-S39. DISCLOSURES: No relevant relationships by Fatima Ayub, source=Web Response No relevant relationships by Muhammad Hasib Khalil, source=Web Response

Hepatic Manifestations of Urea Cycle Disorders.

Content available: Author Interview and Audio Recording.

Characterization and Treatment Responsiveness of Genetically Engineered Ornithine Transcarbamylase-Deficient Pig.

To develop novel medical technologies, pig disease models are invaluable especially in the final stages of translational research. Recently, we established a genetically engineered ornithine transcarbamylase-deficient (OTCD) pig strain. Here, we report its characterization and treatment responsiveness. OTCD pigs were obtained by mating an OTCD carrier female (OTC-Xc.186_190delXWT) with a wild-type male. Due to the X-linked recessive mode of inheritance, the disease phenotype emerged only in males. Medication with nitrogen-scavenging agents was based on a clinical protocol. OTCD pigs were born smaller than their wild-type and carrier littermates, showing anemia and faltering. Biochemically, high levels of urinary orotic acid and loss of OTC activity were observed. The natural life course of OTCD pigs was characterized by a decrease in arterial percentage saturation of oxygen and body temperature, as well as an increase in blood ammonia levels; the pigs died in 24.0 ± 5.0 h (mean ± SD, n = 6). The established standard medication composed with nitrogen-scavenging agents and transfusion nearly doubled the survival time to 42.4 ± 13.7 h (n = 6). Our OTCD pig model appropriately mimicked the human pathology. Along with established protocols in handling and medication, this is a first step in developing a large animal disease model that is useful for translational research into novel medical technologies, such as cell transplantation and gene therapy, as well as in relation to urea cycle disorder.

Ornithine Transcarbamylase Deficiency Presenting with Symptoms of Mania in a Young Adult Male

The purpose of this case study is to review the clinical presentation and medical workup of a young adult male presenting with acute behavior changes in the setting of undiagnosed ornithine transcarbamylase deficiency (OTCD). This case study involves a 19-year-old male with a psychiatric history of depression and one previous suicide attempt, who presented to a large midwestern university hospital emergency department after being found by police naked in a neighbor's yard. He displayed manic signs and symptoms, including euphoria, lack of sleep for five days, and attempting to purchase a new car and three large screen TVs. Family reported the patient uncharacteristically announced three weeks earlier that he was vegetarian and stopped eating his frequent customary cheeseburgers. Due to increased anxiety and inability to sleep, the patient received lorazepam 2 mg in the emergency department. Upon transfer to the psychiatric unit, therapy was initiated with aripiprazole 5 mg daily and valproate 1000 mg nightly on Day 1 of treatment. The patient refused medications on hospital Day 2, then received this combination again on Day 3. The next morning, the patient complained of lethargy, headache, nausea, and vomiting. The patient's ammonia level was found to be 204 micromol/L with ALT and AST of 714 and 647 IU/L respectively. Tests for infectious hepatitis were negative. Medical consultation recommended discontinuation of current medications, vigorous hydration, and further work up. On further investigation, the patient was found to have low plasma citrulline level of 8 micromol/L, undetectable plasma arginine, and high urinary orotic acid. The laboratory data showed a biochemical phenotype consistent with a diagnosis of partial OTCD, an X-linked urea cycle disorder resulting in toxic hyperammonemia. The patient was treated with a low protein diet modification as well as a combination of sodium benzoate and sodium phenylbutyrate to reduce serum ammonia concentration. With treatment the patient's laboratory values normalized, and mental status improved. In conclusion, partial ornithine transcarbamylase deficiency may manifest with psychiatric symptoms in early adulthood. In young patients with elevated ammonia and mental status change, OTCD is an important diagnosis to consider, as it is the most common inherited cause of hyperammonemia.

Correction of a urea cycle defect after ex vivo gene editing of human hepatocytes

Ornithine transcarbamylase deficiency (OTCD) is a monogenic disease of ammonia metabolism in hepatocytes. Severe disease is frequently treated by orthotopic liver transplantation. An attractive approach is the correction of a patient’s own cells to regenerate the liver with gene-repaired hepatocytes. This study investigates the efficacy and safety of ex vivo correction of primary human hepatocytes. Hepatocytes isolated from an OTCD patient were genetically corrected ex vivo, through the deletion of a mutant intronic splicing site achieving editing efficiencies >60% and the restoration of the urea cycle in vitro. The corrected hepatocytes were transplanted into the liver of FRGN mice and repopulated to high levels (>80%). Animals transplanted and liver repopulated with genetically edited patient hepatocytes displayed normal ammonia, enhanced clearance of an ammonia challenge and OTC enzyme activity, as well as lower urinary orotic acid when compared to mice repopulated with unedited patient hepatocytes. Gene expression was shown to be similar between mice transplanted with unedited or edited patient hepatocytes. Finally, a genome-wide screening by performing CIRCLE-seq and deep sequencing of >70 potential off-targets revealed no unspecific editing. Overall analysis of disease phenotype, gene expression, and possible off-target editing indicated that the gene editing of a severe genetic liver disease was safe and effective.

Humanized liver mouse model with transplanted human hepatocytes from patients with ornithine transcarbamylase deficiency.

Ornithine transcarbamylase deficiency (OTCD) is a metabolic and genetic disease caused by dysfunction of the hepatocytic urea cycle. To develop new drugs or therapies for OTCD, it is ideal to use models that are more closely related to human metabolism and pathology. Primary human hepatocytes (HHs) isolated from two patients (a 6‐month‐old boy and a 5‐year‐old girl) and a healthy donor were transplanted into host mice (hemi‐, hetero‐OTCD mice, and control mice, respectively). HHs were isolated from these mice and used for serial transplantation into the next host mouse or for in vitro experiments. Histological, biochemical, and enzyme activity analyses were performed. Cultured HHs were treated with ammonium chloride or therapeutic drugs. Replacement rates exceeded 80% after serial transplantation in both OTCD mice. These highly humanized OTCD mice showed characteristics similar to OTCD patients that included increased blood ammonia levels and urine orotic acid levels enhanced by allopurinol. Hemi‐OTCD mice showed defects in OTC expression and significantly low enzymatic activities, while hetero‐OTCD mice showed residual OTC expression and activities. A reduction in ammonium metabolism was observed in cultured HHs from OTCD mice, and treatment with the therapeutic drug reduced the ammonia levels in the culture medium. In conclusion, we established in vivo OTC mouse models with hemi‐ and hetero‐patient HHs. HHs isolated from the mice were useful as an in vitro model of OTCD. These OTC models could be a source of valuable patient‐derived hepatocytes that would enable large scale and reproducible experiments using the same donor.