Casein Glycomacropeptide: An Alternative Protein Substitute in Tyrosinemia Type I

Tyrosinemia type I (HTI) is treated with nitisinone, a tyrosine (Tyr) and phenylalanine (Phe)-restricted diet, and supplemented with a Tyr/Phe-free protein substitute (PS). Casein glycomacropeptide (CGMP), a bioactive peptide, is an alternative protein source to traditional amino acids (L-AA). CGMP contains residual Tyr and Phe and requires supplementation with tryptophan, histidine, methionine, leucine, cysteine and arginine. Aims: a 2-part study assessed: (1) the tolerance and acceptability of a low Tyr/Phe CGMP-based PS over 28 days, and (2) its long-term impact on metabolic control and growth over 12 months. Methods: 11 children with HTI were recruited and given a low Tyr/Phe CGMP to supply all or part of their PS intake. At enrolment, weeks 1 and 4, caregivers completed a questionnaire on gastrointestinal symptoms, acceptability and ease of PS use. In study part 1, blood Tyr and Phe were assessed weekly; in part 2, weekly to fortnightly. In parts 1 and 2, weight and height were assessed at the study start and end. Results: Nine of eleven children (82%), median age 15 years (range 8.6–17.7), took low Tyr/Phe CGMP PS over 28 days; it was continued for 12 months in n = 5 children. It was well accepted by 67% (n = 6/9), tolerated by 100% (n = 9/9) and improved gastrointestinal symptoms in 2 children. The median daily dose of protein equivalent from protein substitute was 60 g/day (range 45–60 g) with a median of 20 g/day (range 15 to 30 g) from natural protein. In part 2 (n = 5), a trend for improved blood Tyr was observed: 12 months pre-study, median Tyr was 490 μmol/L (range 200–600) and Phe 50 μmol/L (range 30–100); in the 12 months taking low Tyr/Phe CGMP PS, median Tyr was 430 μmol/L (range 270–940) and Phe 40 μmol/L (range 20–70). Normal height, weight and BMI z scores were maintained over 12 months. Conclusions: In HTI children, CGMP was well tolerated, with no deterioration in metabolic control or growth when studied over 12 months. The efficacy of CGMP in HTI needs further investigation to evaluate the longer-term impact on blood Phe concentrations and its potential influence on gut microflora

Therapeutic Targeting of Fumaryl Acetoacetate Hydrolase in Hereditary Tyrosinemia Type I

Fumarylacetoacetate hydrolase (FAH) is the fifth enzyme in the tyrosine catabolism pathway. A deficiency in human FAH leads to hereditary tyrosinemia type I (HT1), an autosomal recessive disorder that results in the accumulation of toxic metabolites such as succinylacetone, maleylacetoacetate, and fumarylacetoacetate in the liver and kidney, among other tissues. The disease is severe and, when untreated, it can lead to death. A low tyrosine diet combined with the herbicidal nitisinone constitutes the only available therapy, but this treatment is not devoid of secondary effects and long-term complications. In this study, we targeted FAH for the first-time to discover new chemical modulators that act as pharmacological chaperones, directly associating with this enzyme. After screening several thousand compounds and subsequent chemical redesign, we found a set of reversible inhibitors that associate with FAH close to the active site and stabilize the (active) dimeric species, as demonstrated by NMR spectroscopy. Importantly, the inhibitors are also able to partially restore the normal phenotype in a newly developed cellular model of HT1.

The Importance of Succinylacetone: Tyrosinemia Type I Presenting with Hyperinsulinism and Multiorgan Failure Following Normal Newborn Screening

Tyrosinemia type I (TT1) is an inborn error of tyrosine metabolism with features including liver dysfunction, cirrhosis, and hepatocellular carcinoma; renal dysfunction that may lead to failure to thrive and bone disease; and porphyric crises. Once fatal in most infantile-onset cases, pre-symptomatic diagnosis through newborn screening (NBS) protocols, dietary management, and pharmacotherapy with nitisinone have improved outcomes. Succinylacetone provides a sensitive and specific marker for the detection of TT1 but is not universally utilized in screening protocols for the disease. Here, we report an infant transferred to our facility for evaluation and management of hyperinsulinism who subsequently developed acute-onset liver, respiratory, and renal failure around one month of life. She was found to have TT1 caused by novel pathogenic variant in fumarylacetoacetate hydrolase (c.1014 delC, p.Cys 338 Ter). Her NBS, which utilized tyrosine as a primary marker, had been reported as normal, with a tyrosine level of 151 µmol/L (reference: <280 µmol/L). Retrospective analysis of dried blood spot samples via tandem mass spectrometry showed detectable succinylacetone ranging 4.65–10.34 µmol/L. To our knowledge, this is the first patient with TT1 whose initial presenting symptom was hyperinsulinemic hypoglycemia. The case highlights the importance of maintaining a high suspicion for metabolic disease in critically ill children, despite normal NBS. We also use the case to advocate for NBS for TT1 using succinylacetone quantitation.

A Compensatory U1snRNA Partially Rescues FAH Splicing and Protein Expression in a Splicing-Defective Mouse Model of Tyrosinemia Type I

The elucidation of aberrant splicing mechanisms, frequently associated with disease has led to the development of RNA therapeutics based on the U1snRNA, which is involved in 5′ splice site (5′ss) recognition. Studies in cellular models have demonstrated that engineered U1snRNAs can rescue different splicing mutation types. However, the assessment of their correction potential in vivo is limited by the scarcity of animal models with the targetable splicing defects. Here, we challenged the U1snRNA in the FAH5961SB mouse model of hepatic fumarylacetoacetate hydrolase (FAH) deficiency (Hereditary Tyrosinemia type I, HT1) due to the FAH c.706G>A splicing mutation. Through minigene expression studies we selected a compensatory U1snRNA (U1F) that was able to rescue this mutation. Intriguingly, adeno-associated virus-mediated delivery of U1F (AAV8-U1F), but not of U1wt, partially rescued FAH splicing in mouse hepatocytes. Consistently, FAH protein was detectable only in the liver of AAV8-U1F treated mice, which displayed a slightly prolonged survival. Moreover, RNA sequencing revealed the negligible impact of the U1F on the splicing profile and overall gene expression, thus pointing toward gene specificity. These data provide early in vivo proof-of-principle of the correction potential of compensatory U1snRNAs in HTI and encourage further optimization on a therapeutic perspective, and translation to other splicing-defective forms of metabolic diseases.