Invitae Elevated Citrulline Panel

• argininosuccinate lyase (ASL) deficiency
• argininosuccinate synthase (ASS1) deficiency – also known as citrullinemia type 1 (CTLN1)
• citrin (SLC25A13) deficiency – also known as citrullinemia type 2 (CLTN2)
• pyruvate carboxylase (PC) deficiency

Elevated citrulline detected during newborn screening may be associated with argininosuccinate synthetase (ASS1) deficiency, argininosuccinate lyase (ASL) deficiency, and the neonatal form of citrin (SLC25A13) deficiency.

ASL
Argininosuccinate Lyase (ASL) deficiency can present either as a severe neonatal-onset form with hyperammonemia within the first few days of life, or as a late-onset form with episodic hyperammonemia or long-term complications, including liver dysfunction, neurocognitive deficits, and hypertension. These long-term complications can occur in the absence of hyperammonemic episodes. The biochemical diagnosis of ASL deficiency is typically established with elevation of plasma citrulline together with elevated argininosuccinic acid in the plasma or urine.

ASS1
ASS1 deficiency causes citrullinemia type 1 (CTLN1). CTLN1 presents as a clinical spectrum with a severe neonatal form, a milder later-onset form, and some individuals who are asymptomatic. Increased intracranial pressure can occur secondarily to hyperammonemia, resulting in increased neuromuscular tone, spasticity, and ankle clonus. Initial plasma ammonia concentration may be 1000–3000 µmol/L (normal: 40–50 µmol/L). In the milder form, symptoms manifest as recurrent lethargy and somnolence, intellectual disability, and chronic or recurrent hyperammonemia. A lower plasma ammonia concentration may be seen than in the classic form (adult upper limit of normal: <35 µmol/L). A rare form of CTLN1 occurs during and after pregnancy, with affected women experiencing vomiting, lethargy, seizures, confusion, hallucinations, behavioral changes (e.g., manic episodes, psychosis), and swelling of the brain.

SLC25A13
The SLC25A13 gene encodes the aspartate/glutamate transporter called citrin. Citrin deficiency impairs the shuttling of aspartate and glutamate across the mitochondria, causing a mild hyperammonemia and citrullinemia (also known as citrullinemia type II [CTLN2]). The clinical presentation in adults with citrin deficiency is milder than that of CTLN1, and asymptomatic or presymptomatic individuals with citrin deficiency do not always show biochemical abnormalities. For this reason, definitive diagnosis of at-risk relatives may be dependent on the molecular genetic findings of SLC25A13 in the index case.

Citrin deficiency can manifest in newborns as neonatal intrahepatic cholestasis (NICCD), in older children as failure to thrive and lipid abnormalities, and in adults as recurrent hyperammonemia with neuropsychiatric symptoms.

PC
Pyruvate carboxylase deficiency has a broad phenotypic spectrum with a severe neonatal-onset form (Type A), infantile form (Type B) to an intermittent or benign form (Type C). Type A generally presents with lactic acidemia, severe developmental delay, failure to thrive, hypotonia and other neurologic findings such as ataxia, nystagmus and seizures. Premature death typically occurs in infancy or early childhood. Type B presents in the neonatal period with seizures, hypotonia, anorexia, abnormal movements, ocular movement abnormalities, motor delays, hypoglycemia, hyperammonemia and hypernatremia. Most affected individuals do not survive past the first few months of life. Type C is very rare and onset typically occurs within the first year of life. Symptoms typically include episodic metabolic acidosis with lactic acidemia and ketoacidosis precipitated by metabolic stress. Neurologic development can be normal or mildly affected.

Analysis of the genes on this panel will identify pathogenic variants in 90% of patients who are diagnosed with CTLN1 or CTLN2. ASL is the only gene in which pathogenic variants are known to cause argininosuccinate lyase deficiency. PC is the only gene in which pathogenic variants are known to cause pyruvate carboxylase deficiency.

All genetic causes of elevated citrulline are inherited in an autosomal recessive manner.

The prevalence of CTLN1 is 1 in 57,000 births in the US, 1 in 22,150 births in Korea, 1 in 118,543 births in Taiwan, and 1 in 77,811 births in Austria. The prevalence of CTLN2 is unknown, though most identified patients are Japanese. The prevalence of ASL deficiency is approximately 1 in 70,000 live births. In the majority of populations the incidence of PC deficiency is 1:250,000. Type A PC deficiency has an increased incidence in native North Americans of the Ojibwa, Cree and Micmac tribes of the Algonquin-speaking peoples. Type B PC deficiency has an increased incidence in Europe.

For considerations for testing please refer to:

• American College of Medical Genetics. NBS ACT Sheet. Amino Aciduria/Urea Cycle Disorder. https://www.acmg.net/StaticContent/ACT/Citrullinemia.pdf Accessed February 2016.

For management guidelines please refer to:

• Batshaw, ML, et al. Alternative pathway therapy for urea cycle disorders: twenty years later. J. Pediatr. 2001; 138(1 Suppl):S46-54; discussion S54-5.
• New England Consortium of Metabolic Progams. Neonate/Infant/Child with Hyperammonemia. http://newenglandconsortium.org/for-professionals/acute-illness-protocols/urea-cycle-disorders/neonate-infant-child-with-hyperammonemia/ Accessed February 2016.
• Summar, M. Current strategies for the management of neonatal urea cycle disorders. J. Pediatr. 2001; 138(1 Suppl):S30-9.