ClinVar Genomic variation as it relates to human health

The SLC22A5 c.248G>T; p.Arg83Leu variant (rs72552726) is reported in the literature in numerous individuals affected with primary carnitine deficiency (PCD), both in the homozygous state and in individuals with a second pathogenic SLC22A5 variant (El-Hattab 2010, Han 2014, Li 2010, Makhseed 2004, Rose 2012). This variant is reported as pathogenic by multiple laboratories in ClinVar (Variation ID: 25361) and has been observed to co-segregate with disease in affected families (El-Hattab 2010, Makhseed 2004). The p.Arg83Leu variant is found in the South Asian population with an overall allele frequency of 0.19% (51/27338 alleles) in the Genome Aggregation Database, though the prevalence of PCD in the general population may be confounded by asymptomatic individuals (Magoulas 2012). The arginine at codon 83 is moderately conserved, and computational analyses (SIFT, PolyPhen-2) predict that this variant is deleterious. Functional analyses indicate that the p.Arg83Leu variant has negligible carnitine transport activity (Amat di San Filippo 2006, Amat di San Filippo 2011, Frigeni 2017, Makhseed 2004, Rose 2012), and it is poorly expressed and aberrantly localized to the cytoplasm, possibly due to loss of glycosylation (Amat di San Filippo 2011). Based on available information, this variant is considered to be pathogenic. References: Amat di San Filippo C et al. Glycosylation of the OCTN2 carnitine transporter: study of natural mutations identified in patients with primary carnitine deficiency. Biochim Biophys Acta. 2011 Mar;1812(3):312-20. Amat di San Filippo C et al. Pharmacological rescue of carnitine transport in primary carnitine deficiency. Hum Mutat. 2006 Jun;27(6):513-23. El-Hattab A et al. Maternal systemic primary carnitine deficiency uncovered by newborn screening: clinical, biochemical, and molecular aspects. Genet Med. 2010; 12(1):19-24. Frigeni M et al. Functional and molecular studies in primary carnitine deficiency. Hum Mutat. 2017 Dec;38(12):1684-1699. Han L et al. Analysis of genetic mutations in Chinese patients with systemic primary carnitine deficiency. Eur J Med Genet. 2014 Oct;57(10):571-5. Li FY et al. Molecular spectrum of SLC22A5 (OCTN2) gene mutations detected in 143 subjects evaluated for systemic carnitine deficiency. Hum Mutat. 2010 Aug;31(8):E1632-51. Magoulas P et al. Systemic primary carnitine deficiency: an overview of clinical manifestations, diagnosis, and management. Orphanet J Rare Dis. 2012; 7:68. Makhseed N et al. Carnitine transporter defect due to a novel mutation in the SLC22A5 gene presenting with peripheral neuropathy. J Inherit Metab Dis. 2004;27(6):778-80. Rose E et al. Genotype-phenotype correlation in primary carnitine deficiency. Hum Mutat. 2012; 33(1):118-23.

Reported previously in association with primary/systemic carnitine deficiency (PCD) in patients who were homozygous for the R83L variant, as well as in both an infant who was identified who had low free carnitine levels by newborn screening and in the infant's affected mother (Makhseed et al., 2004; Li et al., 2010; El-Hattab et al., 2010); Reported with a second pathogenic variant in the asymptomatic mother of an infant identified with low free carnitine levels by newborn screening (Rose et al., 2012); In functional studies R83L impaired maturation of the transporter to the plasma membrane (Filippo et al., 2011), and reduced carnitine transport to less than 1% in CHO cell expression studies (Frigeni et al., 2017); 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: 15714519, 23430858, 28841266, 20027113, 26589311, 16652335, 21126579, 16602102, 15617188, 26828774, 20574985, 21922592, 25132046)

The missense variant c.248G>T(p.Arg83Leu) in SLC22A5 gene has been reported previously in homozygous state and compound heterozygous state in multiple individuals with Primary carnitine deficiency. Experimental studies have shown that this missense change affects SLC22A5 function (Filippo CA, et al., 2011). The variant has 0.03% allele frequency in gnomAD Exomes and is novel (not in any individuals) in 1000 Genomes. This variant has been reported to the ClinVar database as Uncertain Significance/Likely Pathogenic/ Pathogenic (multiple submissions). The amino acid Arginine at position 83 is changed to a Leucine changing protein sequence and it might alter its composition and physico-chemical properties. The variant is predicted to be damaging by SIFT. The amino acid change p.Arg83Leu in SLC22A5 is predicted as conserved by GERP++ and PhyloP across 100 vertebrates. For these reasons, this variant has been classified as Pathogenic.

This sequence change replaces arginine, which is basic and polar, with leucine, which is neutral and non-polar, at codon 83 of the SLC22A5 protein (p.Arg83Leu). This variant is present in population databases (rs72552726, gnomAD 0.2%). This missense change has been observed in individuals with primary carnitine deficiency (PMID: 15617188, 20574985, 21126579, 21922592; Invitae). It has also been observed to segregate with disease in related individuals. ClinVar contains an entry for this variant (Variation ID: 25361). Advanced modeling of protein sequence and biophysical properties (such as structural, functional, and spatial information, amino acid conservation, physicochemical variation, residue mobility, and thermodynamic stability) performed at Invitae indicates that this missense variant is expected to disrupt SLC22A5 protein function with a positive predictive value of 95%. Experimental studies have shown that this missense change affects SLC22A5 function (PMID: 15617188, 16652335, 21126579). For these reasons, this variant has been classified as Pathogenic.

Variant summary: The SLC22A5 c.248G>T (p.Arg83Leu) variant is located in an extracellular loop close to putative glycosylation sites of SLC22A5 (via Filippo_2011) involves the alteration of a conserved nucleotide, which 4/4 in silico tools (SNPs&GO not captured due to low reliability index) predict a damaging outcome. The variant of interest was observed in the large, broad control population, ExAC, with an allele frequency of 22/50892 (1/2313), predominantly in the South Asian cohort, 20/11664 (1/583), which does not exceed the estimated maximal expected allele frequency for a pathogenic SLC22A5 variant of 1/219. Multiple publications have cited the variant in affected homozygous and compound heterozygous individuals, and the variant has also been reported in asymptomatic individuals with serum carnitine deficiency. In addition, a functional study (Filippo_2011) indicates the variant impaired glycosylation and maturation of SLC22A5 to the plasma membrane. Furthermore, multiple clinical diagnostic laboratories/reputable databases classified this variant as pathogenic. Taken together, this variant is classified as pathogenic.

NM_003060.3(SLC22A5):c.248G>T(R83L) is classified as likely pathogenic in the context of primary carnitine deficiency. Sources cited for classification include the following: PMID 24516753, 25132046, 20574985, 16652335, 21126579, 21922592 and 15617188. Classification of NM_003060.3(SLC22A5):c.248G>T(R83L) is based on the following criteria: There is strong evidence of association with the variant and the relevant disease and there is functional data showing deficient protein function. Please note: this variant was assessed in the context of healthy population screening.