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A Case Series and Functional Study of Two Commonly Reported Pathogenic Variants in OTC Supports a Hypomorphic Classification

Biochemical/Metabolic and Therapeutics
  • Primary Categories:
    • Metabolic Genetics
  • Secondary Categories:
    • Metabolic Genetics
Introduction:
Ornithine transcarbamylase deficiency (OTCD), the most common urea cycle disorder, is an X-linked condition that typically manifests as neonatal onset hyperammonemia in males and with variable severity in females. Due to the risk for life-threatening hyperammonemia and the availability of therapies to prevent this complication, OTCD is included in the ACMG list of secondary findings to be reported in clinical exome and genome sequencing and in many larger carrier screening panels. Thus, incidentally, apparently asymptomatic individuals with pathogenic variants in OTC are increasingly being identified. At our institution, we identified 12 patients with either the NM_000531.6:c.118C>T (p.Arg40Cys) or NM_000531.6:c.1061T>G (p.Phe354Cys) OTC variant. These two variants have been reported in the literature in males presenting with acute hyperammonemia as well as in apparently asymptomatic males who were ascertained through carrier screening, family testing, or as secondary findings and are classified as pathogenic by most laboratories. Thus, to understand the phenotypic consequences associated with these variants and to better inform counseling, we conducted a clinical study and functional studies of OTC enzyme activity in a yeast model-system.

 

Methods:
First, we performed a retrospective chart review of individuals who were heterozygous or hemizygous for p.Arg40Cys and p.Phe354Cys variants and who were evaluated in the Texas Children’s Hospital’s Metabolic Genetics Clinic from 1995-2024. We then conducted a search of the Urea Cycle Disorders Consortium’s database and queried the All of Us restricted tier dataset v7 to create cohorts of individuals with either of these two variants. Lastly, we performed a systematic literature review for publications or abstracts containing these two variants using PubMed, Google Scholar, Mastermind, and LitVar2. We used a quantitative yeast-based assay to measure the impact of these variants on the activity of human OTC.

 

Results:
We identified 13 individuals (5 females, 8 males) with the p.Arg40Cys variant. Six males were identified because of symptomatic hyperammonemia with a median age of onset of 25.5 years (range 9 to 66 years); three died from their first hyperammonemic crisis. There were no instances of neonatal hyperammonemia in males. Two males without known hyperammonemia were 3 months and 40 years old at last follow-up. None of the  females had hyperammonemia.

Similarly, we identified 14 individuals (5 females, 9 males) with the p.Phe354Cys variant. Two males were identified because of symptomatic hyperammonemia with median age of first metabolic decompensation of 14 years (range 13 to 68 years); one died from his first episode of hyperammonemia. One male who initially was identified by cascade testing, developed symptomatic hyperammonemia at age of 68 years.  There were no instances of neonatal hyperammonemia in males. Six males did not have history of hyperammonemia (age range 3 years to 55 years). None of the females had hyperammonemia.

In a validated assay for human OTC protein function, yeast cells harboring the codon-harmonized version of human OTC, yOTC, grow robustly in medium lacking arginine, whereas yeast cells harboring the gene deletion arg3del do not. Normalized growth estimates for the two OTC variants in this assay were 0.276 (SE = 0.0047) for p.Arg40Cys and 0.361 (SE = 0.0050) for p.Phe354Cys. These findings place both variants in the hypomorphic range (5%-90% growth) as previously defined by Lo et al. 2023.

 

Conclusion:
Our data supports the classification of both p.Arg40Cys and p.Phe354Cys variants as hypomorphic variants causative of late-onset disease in males. Although the presence of more subtle signs and symptoms of chronic hyperammonemia could not be elucidated in our study, these findings have implications for clinical management and genetic counseling.  

 

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