Identification of novel variants in carbamoyl phosphate synthetase 1 gene and comparative pathogenicity assessments of CPS1 missense variants following ACMG/AMP-ClinGen recommendation for computational tools

Abstract

Carbamoyl phosphate synthetase I (CPS1) deficiency is a rare autosomal recessive metabolic abnormality cause by dysfunctionality of CPS1 and often result in unfavorable outcome. In this study, we presented the detailed laboratory features and genetic analysis of two patients with heterozygous variants of CPS1, c.1927 A > G (p.Asn643Asp), c.2375 T > G (p.Met792Arg), c.3443 T > A (p.Met1148Lys) in patient 1; c.3784C > T (p.Arg1262Ter), c.3734 T > A (p.Leu1245His) in patient 2, respectively. c.1927 A > G (p.Asn643Asp) and c.2375 T > G (p.Met792Arg) are novel out of 5 variants and classified as variants of uncertain significance (VUS) under the guidelines of ACMG/AMP-ClinGen. Structure-based analysis of 4 missense variants indicates deleterious alterations to the protein. Since the employment of genetic testing as a clinical diagnostic tool, distinguishing pathogenic from polymorphic changes poses significant problems for geneticists. As recommendation for PP3/BP4, the computational tools for missense variant have been published, we performed a comparative evaluation for pathogenicity interpretation in our patients and in ClinVar database regarding CPS1 missense variants under the updated guidelines of ACMG/AMP-ClinGen. The application of computational tools under the ACMG/AMP-ClinGen criteria revealed an increased sensitivity for pathogenicity evaluation, from variants of uncertain significance (VUS) to likely pathogenic (LP) in previously reported cases; while for variants without clinic information in the ClinVar database, the pathogenicity assessment of VUS remained, and shows a more optimistic and reliable clinical application in molecular diagnosis.

Keywords: Carbamoyl phosphate synthetase I deficiency; Missense variant; Pathogenicity interpretation; Urea cycle disorder.

Fig. 1

Sanger sequencing of patients with CPS1 variants. A. P1 has three heterozygous variants of c.1927 A > G (p.Asn643Asp), c.2375 T > G (p.Met792Arg) and c.3443 T > A (p.Met1148Lys). B. P2 has two heterozygous variants of c.3784C > T (p.Arg1262Ter), c.3734 T > A (p.Leu1245His). The red arrows refer corresponding variants.

Fig. 2

The non-covalent interactions between wild-type and mutant structure, non-covalent bonds are represented by dotted lines. The ADP molecules are colored in red, magnesium ion is shown as a green spheres and the potassium ion is shown as a violet sphere. A. Asn643Asp result in non-covalent interaction changes and might lead to a reduction in the ability of the enzyme in ligand binding in process of bicarbonate phosphorylation. S2 and L1 domain are colored in yellow and green respectively. B. Met792Arg result in in non-covalent interaction changes and might lead to a reduction in the ability of the enzyme in ligand binding in process of bicarbonate phosphorylation. L1 and L3 domain are colored in cyan and violet respectively. C. Met1148Lys result in non-covalent interaction changes might lead to a reduction in the ability of the enzyme in ligand binding in process of carbamate phosphorylation. L3 domain are colored in yellow. D. Leu1245His result in non-covalent interaction changes might lead to a reduction in the ability of the enzyme in ligand binding in process of carbamate phosphorylation. L3 domain are colored in cyan.

Fig. 3

Distribution and flow changes of CPS1 pathogenicity classification in ClinVar. A. Left: 95.12 % (312 variants) are classified as VUS, 4.57 % (15 variants) are classified as LP and 0.3 % (1 variant) are classified as B in ClinVar Database before PP3 criterion application; Right: 89.63 % (294 variants) are classified as VUS, 9.45 % (31 variants) are classified as LP and 0.61 % (2 variants) are classified as P and 0.3 % (1 variant) are classified as B in ClinVar Database after PP3 criterion application. B. Reported variants group; Left: 67.57 % (25 variants) are classified as VUS and 32.43 % (12 variants) are classified as LP before PP3 criterion application; Right: 18.92 % (7 variants) are classified as VUS, 75.68 % (28 variants) are classified as LP and 5.41 % (2 variants) are classified as P after PP3 criterion application. C. Unreported variants group; Left: 98.63 % (287 variants) are classified as VUS, 1.03 % (3 variants) are classified as LP and 0.34 % (1 variants) are classified as B before PP3 criterion application; Right: 98.63 % (287 variants) are classified as VUS, 1.03 % (3 variants) are classified as LP and 0.34 % (1 variants) are classified as B after PP3 criterion application.