Variant summary: MCCC2 c.1065A>T (p.Leu355Phe) results in a non-conservative amino acid change located in the Acetyl-coenzyme A carboxyltransferase, C-terminal domain (IPR011763) of the encoded protein sequence. Five of five in-silico tools predict a damaging effect of the variant on protein function. The variant allele was found at a frequency of 0.00036 in 251280 control chromosomes. This frequency is not significantly higher than expected for a pathogenic variant in MCCC2 causing Methylcrotonyl-CoA Carboxylase Deficiency (0.00036 vs 0.0042), allowing no conclusion about variant significance. c.1065A>T has been reported in the literature as a homozygous genotype in multiple individuals affected with Methylcrotonyl-CoA Carboxylase Deficiency with characteristic biochemical profiles, namely, elevated excretion of 3-hydroxyisovaleric acid (3-HIVA), 3-methylcrotonylglycine (3-MCG) and also subsequently cited by others (example, Nguyen_2011, Grunert_2012, Adhikari_2020, Shepard_2015, Fonseca_2016). These data indicate that the variant is very likely to be associated with disease. At least one publication reports experimental evidence evaluating an impact on protein function. The most pronounced variant effect results in 10%-<30% of normal MCC enzyme activity in a homozygous individual. Four clinical diagnostic laboratories have submitted clinical-significance assessments for this variant to ClinVar after 2014 without evidence for independent evaluation. All laboratories classified the variant as pathogenic/likely pathogenic. Based on the evidence outlined above, the variant was classified as pathogenic.

The p.Leu355Phe variant in MCCC2 has been reported in the homozygous state in at least 9 individuals and in the compound heterozygote state with another pathogenic variant in MCCC2 in 2 individuals with 3-methylcrotonyl-CoA (MCC) deficiency or a positive newborn screen (Nguyen 2011 PMID: 21071250, Shepard 2015 PMID: 25356967, Forsyth 2016 PMID: 27033733, GeneDx pers. comm., Invitae pers. comm.). Most of these individuals were typically asymptomatic at the time of testing, however had MCC deficiency determined by biochemical analyses. This variant has also been reported by other clinical laboratories in ClinVar (Variation ID 203806) and has been identified in 0.085% (13/15282) of Latino chromosomes by gnomAD (http://gnomad.broadinstitute.org, v.3.1.2). Computational prediction tools and conservation analyses suggest that this variant may impact the protein. In summary, although additional studies are required to fully establish its clinical significance, this variant meets criteria to be classified as likely pathogenic for autosomal recessive MCC deficiency. ACMG/AMP Criteria applied: PM3_Strong, PM2_Supporting, PP3, PP4.

PS3_Moderate, PM3_Strong

The c.1065A>T (p.L355F) alteration is located in exon 11 (coding exon 11) of the MCCC2 gene. This alteration results from a A to T substitution at nucleotide position 1065, causing the leucine (L) at amino acid position 355 to be replaced by a phenylalanine (F). Based on data from gnomAD, the T allele has an overall frequency of 0.036% (90/251280) total alleles studied. The highest observed frequency was 0.257% (89/34574) of Latino alleles. This alteration has been detected in the homozygous state and/or in conjunction with other MCCC2 variant(s) in multiple unrelated individuals with 3-methylcrotonyl-CoA carboxylase deficiency (Nguyen, 2011; Forsyth, 2016; Cook, 2024) This amino acid position is highly conserved in available vertebrate species. This alteration is predicted to be deleterious by in silico analysis. Based on the available evidence, this alteration is classified as pathogenic.

This sequence change replaces leucine, which is neutral and non-polar, with phenylalanine, which is neutral and non-polar, at codon 355 of the MCCC2 protein (p.Leu355Phe). This variant is present in population databases (rs757052602, gnomAD 0.3%). This missense change has been observed in individual(s) with 3-methylcrotonylglycinuria (PMID: 21071250, 25356967; internal data). ClinVar contains an entry for this variant (Variation ID: 203806). Invitae Evidence Modeling of protein sequence and biophysical properties (such as structural, functional, and spatial information, amino acid conservation, physicochemical variation, residue mobility, and thermodynamic stability) indicates that this missense variant is expected to disrupt MCCC2 protein function with a positive predictive value of 80%. For these reasons, this variant has been classified as Pathogenic.

In silico analysis supports that this missense variant has a deleterious effect on protein structure/function; This variant is associated with the following publications: (PMID: 25356967, 22642865, 27033733, 21071250)

The c.1065A>T variant in MCCC2 is a missense variant predicted to cause substitution of leucine to phenylalanine at amino acid 355. This variant is rare in the general population with a frequency below the threshold expected for the associated phenotype(s). This variant has been observed in one or more individuals affected with the associated recessive disease, as either homozygous or compound heterozygous with a second variant (PMID: 25356967, 21071250, 27033733). This variant has been identified in one or more affected individual with a phenotype highly consistent with the associated gene (PMID: 21071250). Computational prediction algorithms indicate this variant is likely to affect gene or protein function. Given the available evidence, this variant is classified as Likely Pathogenic.

The MCCC2 c.1065A>T variant is predicted to result in the amino acid substitution p.Leu355Phe. This variant has been reported in the homozygous state in multiple apparently unrelated individuals with enzymatically confirmed 3-methylcrotonyl-CoA carboxylase deficiency, though it should be noted that two of these patients were asymptomatic adults (Nguyen et al. 2011. PubMed ID: 21071250; Shepard et al. 2015. PubMed ID: 25356967). It has also been reported in the heterozygous state with a second heterozygous likely pathogenic MCCC2 variant in a patient who was reportedly clinically “normal” during the neonatal period (Forsyth et al. 2016. PubMed ID: 27033733). This variant is reported in 0.26% of alleles in individuals of Latino descent in gnomAD. This variant is interpreted as likely pathogenic or pathogenic by multiple independent submitters to ClinVar (https://www.ncbi.nlm.nih.gov/clinvar/variation/203806). Taken together, this variant is interpreted as likely pathogenic.