ClinVar Genomic variation as it relates to human health

The p.Ser113Leu variant in CPT2 is the most common variant associated with late- onset carnitine palmitoyltransferase deficiency (myopathic form); most of the pa tients reported to date (>14) were homozygous (Taroni 1993, Bonnefont 1996, Mart in 1999). The p.Ser113Leu variant has been identified in 0.2% (134/66568) of Eur opean chromosomes by the Exome Aggregation Consortium (ExAC, http://exac.broadin stitute.org; dbSNP rs74315294). Although this variant has been seen in the gener al population, its frequency is low enough to be consistent with a recessive car rier frequency. Biochemical characterization and in vitro functional studies als o provide evidence that the p.Ser113Leu variant may impact protein function (Tar oni 1993, Bonnefont 1996). In summary, this variant meets criteria to be classif ied as pathogenic for carnitine palmitoyltransferase II deficiency in an autosom al recessive manner. It should be noted that a few heterozygous carriers of this variant have been reported with mild symptoms (Taggert 1999, Kaufmann 1997).

NM_000098.2(CPT2):c.338C>T(S113L) is classified as pathogenic in the context of carnitine palmitoyltransferase II deficiency. Please note that the S113L variant is associated with the myopathic form of carnitine palmitoyltransferase II deficiency. Disease phenotype is dependent on, but not necessarily predicted by, the combination of variants inherited. Sources cited for classification include the following: PMID 21913903, 16996287, 15642848 and 835844. Classification of NM_000098.2(CPT2):c.338C>T(S113L) is based on the following criteria: This is a well-established pathogenic variant in the literature that has been observed more frequently in patients with clinical diagnoses than in healthy populations. Please note: this variant was assessed in the context of healthy population screening.

The CPT2 c.338C>T; p.Ser113Leu variant (rs74315294), is reported in the homozygous and compound heterozygous state in the literature as the most common variant in individuals affected with the adult myopathic form of carnitine palmitoyltransferase II deficiency (Avila-Smirnow 2018, Balasubramanian 2018, Edmondson 2017, Fontaine 2018, Joshi 2012, Kottlors 2001, Taroni 1993, Vavlukis 2014, Vivante 2017, Wataya 1998). This variant is reported as pathogenic by multiple laboratories in ClinVar (Variation ID: 8953), and is found in the general population with an overall allele frequency of 0.14% (393/282,834 alleles, including 4 homozygotes) in the Genome Aggregation Database. The serine at codon 338 is highly conserved, and computational analyses predict that this variant is deleterious (REVEL: 0.887). Functional analyses of the variant protein shows a reduction in stability leading to decreased enzyme activity (Motlagh 2016a, Motlagh 2016b, Taroni 1993). Based on available information, the p.Ser113Leu variant is considered to be pathogenic. References: Avila-Smirnow D et al. Carnitine palmitoyltransferase type 2 deficiency: novel mutation in a Native South American family with whole-body muscle magnetic resonance imaging findings: two case reports. J Med Case Rep. 2018 Aug 28;12(1):249. PMID: 30149802. Balasubramanian M et al. Recurrent rhabdomyolysis caused by carnitine palmitoyltransferase II deficiency, common but under-recognised: Lessons to be learnt. Mol Genet Metab Rep. 2018 Mar 6;15:69-70. PMID: 29744303. Edmondson AC et al. Missed Newborn Screening Case of Carnitine Palmitoyltransferase-II Deficiency. JIMD Rep. 2017;33:93-97. PMID: 27067077. Fontaine M et al. Fluxomic assay-assisted diagnosis orientation in a cohort of 11 patients with myopathic form of CPT2 deficiency. Mol Genet Metab. 2018 Apr;123(4):441-448. PMID: 29478820. Joshi PR et al. Clinically symptomatic heterozygous carnitine palmitoyltransferase II (CPT II) deficiency. Wien Klin Wochenschr. 2012 Dec;124(23-24):851-4. PMID: 23184072. Kottlors M et al. Valproic acid triggers acute rhabdomyolysis in a patient with carnitine palmitoyltransferase type II deficiency. Neuromuscul Disord. 2001 Nov;11(8):757-9. PMID: 11595519. Motlagh L et al. Malony-CoA inhibits the S113L variant of carnitine-palmitoyltransferase II. Biochim Biophys Acta. 2016 Jan;1861(1):34-40. PMID: 26477380. Motlagh L et al. Stabilization of the thermolabile variant S113L of carnitine palmitoyltransferase II. Neurol Genet. 2016 Feb 25;2(2):e53. PMID: 27123472. Taroni F et al. Identification of a common mutation in the carnitine palmitoyltransferase II gene in familial recurrent myoglobinuria patients. Nat Genet. 1993 Jul;4(3):314-20. PMID: 8358442. Vavlukis M et al. Rhabdomyolysis and Cardiomyopathy in a 20-Year-Old Patient with CPT II Deficiency. Case Rep Genet. 2014;2014:496410. PMID: 24563797. Vivante A et al. Exome sequencing in Jewish and Arab patients with rhabdomyolysis reveals single-gene etiology in 43% of cases. Pediatr Nephrol. 2017 Dec;32(12):2273-2282. PMID: 28779239. Wataya K et al. Two CPT2 mutations in three Japanese patients with carnitine palmitoyltransferase II deficiency: functional analysis and association with polymorphic haplotypes and two clinical phenotypes. Hum Mutat. 1998;11(5):377-86. PMID: 9600456.

The variant is observed at an extremely low frequency in the gnomAD v2.1.1 dataset (total allele frequency: 0.139%). Protein truncation variants are a common disease-causing mechanism. Functional studies provide supporting evidence of the variant having a damaging effect on the gene or gene product (PMID: 26477380, 27123472, 8358442). In silico tool predictions suggest damaging effect of the variant on gene or gene product (REVEL: 0.89; 3Cnet: 0.34). Same nucleotide change resulting in same amino acid change has been previously reported as pathogenic/likely pathogenic with strong evidence (ClinVar ID: VCV000008953 / PMID: 8358442). The variant has been reported to be in trans with a pathogenic variant as either compound heterozygous or homozygous in at least 2 similarly affected unrelated individuals (PMID: 11994355, 14615409, 20810031). Therefore, this variant is classified as Pathogenic according to the recommendation of ACMG/AMP guideline.

The CPT2 c.338C>T (p.Ser113Leu) missense variant is the most commonly occurring pathogenic variant in the CPT2 gene, accounting for approximately 60% of disease alleles (Wieser et al. 2014). Across a small selection of the available literature, the p.Ser113Leu variant was identified in a total of 77 individuals with carnitine palmitoyltransferase II deficiency, including 38 homozygotes, 18 compound heterozygotes, and 21 heterozygotes in whom a second variant was not reported (Taroni et al. 1993; Bonnefont et al. 1996; Martin et al. 1999; Anichini et al. 2011; Fanin et al. 2012; Shima et al. 2016). The variant was also found in nine unaffected heterozygous carriers (Martin et al. 1999). The p.Ser113Leu variant was absent from 64 controls but is reported at a frequency of 0.00201 in the European (non-Finnish) population of the Exome Aggregation Consortium. Taroni et al. (1993) demonstrated that the p.Ser113Leu variant displayed significantly reduced CPT II activity but similar enzyme synthesis as compared to wild type when transiently expressed in COS1 cells. Additionally, steady-state levels of CPT II from lymphoblasts from an individual homozygous for the p.Ser113Leu variant were significantly decreased as compared to control cells, though normal biosynthesis of CPT II was observed. Bonnefont et al. (1996) showed a reduction to 20% of wild type of residual CPT II activity in fibroblasts from an individual homozygous for the p.Ser113Leu variant. Based on the collective evidence, the p.Ser113Leu variant is classified as pathogenic for carnitine palmitoyltransferase II deficiency. This variant was observed by ICSL as part of a predisposition screen in an ostensibly healthy population.

The best available variant frequency is 1-3 times higher than the disease allele frequency. Found in at least one patient with expected phenotype for this gene. Predicted to have a damaging effect on the protein. This variant is statistically more frequent in affected individuals than in the general population and/or healthy controls. In multiple individuals, this variant has been seen with a single recessive pathogenic variant in the same gene, suggesting this variant may also be pathogenic. Assessment of experimental evidence suggests this variant results in abnormal protein function. This variant appears to segregate with disease in at least one family.

This CPT2 variant (rs74315294) has been identified in a large population dataset and the minor allele frequency is neither low enough to consider the variant rare (<0.1%) nor high enough to consider it a population polymorphism (>1%) within the European (non-Finnish) subpopulation (gnomAD: 238/129144 alleles; 0.18%, 1 homozygote). However, the minor allele frequency is low enough to be consistent with a recessive carrier frequency. This variant, c.338C>T (p.Ser113Leu), has been reported in ClinVar, and is the most common pathogenic variant associated with late-onset (myopathic) carnitine palmitoyltransferase deficiency in individuals of European ancestry; most of the patients reported to date were homozygous for this variant. Experimental studies demonstrate that this missense variant significantly reduces enzyme activity. We consider this variant to be pathogenic.

This mutation has been previously reported as disease-causing and was found once in our laboratory in trans with a frameshift mutation in a 1-year-old female with IUGR, global delays, spastic tetraparesis, brain calcifications and leukomalacia, and a similarly affected brother (who was a single heterozygote; did not carry the frameshift mutation). Heterozygotes are expected to be asymptomatic carriers.

ACMG classification criteria: PS3 supporting, PM3 very strong, PP3 supporting

PS3, PS4, PM2, PP5

This sequence change replaces serine, which is neutral and polar, with leucine, which is neutral and non-polar, at codon 113 of the CPT2 protein (p.Ser113Leu). This variant is present in population databases (rs74315294, gnomAD 0.7%), and has an allele count higher than expected for a pathogenic variant. This missense change has been observed in individuals with carnitine palmitoyltransferase II deficiency (PMID: 8786066, 9309694, 10090476, 10398215, 12673791, 12707442, 15642848, 15776096, 17936304, 21913903). It is commonly reported in individuals of European ancestry (PMID: 8786066, 9309694, 10090476, 10398215, 12673791, 12707442, 15642848, 15776096, 17936304, 21913903). ClinVar contains an entry for this variant (Variation ID: 8953). An algorithm developed to predict the effect of missense changes on protein structure and function (PolyPhen-2) suggests that this variant is likely to be disruptive. Experimental studies have shown that this missense change affects CPT2 function (PMID: 2647738, 8358442, 27123472). For these reasons, this variant has been classified as Pathogenic.

This sequence change is predicted to replace serine with leucine at codon 113 of the CPT2 protein (p.(Ser113Leu)). The serine residue is highly conserved (100 vertebrates, UCSC), and is located in the mitochondrial matrix region of carnitine O-palmitoyltransferase 2 (UniProt). There is a large physicochemical difference between serine and leucine. The variant is present in a large population cohort at a frequency of 0.14% (rs74315294, 393/282,834 alleles, 4 homozygotes in gnomAD v2.1). The variant is the most commonly occurring pathogenic variant in CPT2 associated with the myopathic form CPT II deficiency (PMID: 20301431). It has been identified in the homozygous state and with a second pathogenic allele in multiple individuals diagnosed with recurrent myoglobinuria/rhabdomyolysis (PM3_VeryStrong; for example PMID: 8358442, 12707442, 20810031). Segregation of the homozygous variant has been reported in at least two families with the phenotype ranging from reduced tolerance to exercise to recurrent rhabdomyolysis (PP1_Strong; PMID: 8786066, 20810031). Homozygous cases are reported with significantly reduced CPT II enzyme activity and slightly reduced or normal long-chain fatty acid oxidation levels in tissues, and markedly reduced CPT II activity is also demonstrated in in vitro expression studies (PP4, PS3_Supporting; PMID: 8358442, 8651281). Multiple lines of computational evidence predict a deleterious effect for the missense substitution (PP3; 6/6 algorithms). Based on the classification scheme RMH ACMG Guidelines v1.2.1, this variant is classified as PATHOGENIC. Following criteria are met: PM3_VeryStrong, PP1_Strong, PS3_Supporting, PP3, PP4.

The c.338C>T (p.S113L) alteration is located in exon 3 (coding exon 3) of the CPT2 gene. This alteration results from a C to T substitution at nucleotide position 338, causing the serine (S) at amino acid position 113 to be replaced by a leucine (L). Based on data from the Genome Aggregation Database (gnomAD) database, the CPT2 c.338C>T alteration was observed in 0.14% (393/282834) of total alleles studied, with a frequency of 0.75% (78/10370) in the Ashkenazi Jewish subpopulation. This variant is a common mutation for the myopathy form of carnitine palmitoyltransferase 2 (CPTII) deficiency and has been reported in numerous homozygous and compound heterozygous individuals (Vladutiu, 2002; Taroni, 1993; Mart&iacute;n, 2000; Taggart, 1999; Handig, 1996). This amino acid position is well conserved in available vertebrate species. COS1 cells with this variant demonstrated reduced activity compared to wild type (Taroni, 1993). The p.S113L alteration is predicted to be deleterious by in silico analysis. Based on the available evidence, this alteration is classified as pathogenic.

The CPT2 c.338C>T (p.Ser113Leu) variant is the most common variant in individuals affected with carnintine palmitoyl transferase II deficiency, the mild myopathic form and is reported to segregate with disease. Of those individuals, approximately 18 were compound heterozygous and 38 homozygous. At least 21 heterozygous carriers have also been described with a subset that are clinically symptomatic with muscle CPT activity 39-45% of normal reference (Joshi PR et al., PMID: 23184072; Kaufmann P et al., PMID: 9309694; Taggart RT et al., PMID: 10090476; Taroni F et al., PMID: 8358442). Functional studies show abnormal regulation of the enzyme and thermal instability, indicating that this variant impacts protein function (Taroni F et al., PMID: 8358442; Motlagh L et al., PMID: 26477380). The highest population minor allele frequency in the population database genome aggregation database (v.2.1.1) is 0.75% in the Ashkenazi Jewish population. Computational predictors indicate that the variant is damaging, evidence that correlates with impact to CPT2 function. This variant has been reported in the ClinVar database as a pathogenic variant by 35 submitters and likely pathogenic by 3 submitters. Based on available information and the ACMG/AMP guidelines for variant interpretation (Richards S et al., PMID: 25741868), this variant is classified as likely pathogenic.

Most common CPT2 pathogenic variant found in approximately 60% of mutant alleles in European patients with the adult myopathic form of CPT2 deficiency (PMID: 15363638); Children who are compound heterozygotes with S113L have also been reported, presenting with myopathic features or an intermediate phenotype (PMID: 14615409); Published functional studies demonstrate a damaging effect (PMID: 27123472, 8358442, 26477380); 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: 26477380, 35460704, 36964972, 36473042, 36478999, 15776096, 17936304, 30149802, 30609409, 30957255, 15363638, 8358442, 9600456, 23184072, 11595519, 21228398, 22975760, 24563797, 25333069, 27034144, 28054946, 15642848, 12707442, 26990548, 29744303, 27067077, 29478820, 28779239, 31191612, 31517061, 31407473, 32272925, 31980526, 34426522, 16225172, 31589614, 8786066, 9309694, 32528171, 32140910, 33815142, 35314707, 34495297, 27123472, 14615409)

CPT2: PM3, PS4:Moderate, PM2:Supporting, PP1, PS3:Supporting

Based on the classification scheme VCGS_Germline_v1.3.4, this variant is classified as Pathogenic. Following criteria are met: 0102 - Loss of function is a known mechanism of disease in this gene and is associated with neonatal, infantile and stress-induced myopathic CPT II deficiency. (I) 0106 - This gene is associated with autosomal recessive disease. This gene is generally considered to be associated with an autosomal recessive condition, but some cases of manifesting carriers have been reported for the myopathic form of CPT II deficiency (PMID: 32295037). (I) 0115 - Variants in this gene are known to have variable expressivity. The myopathic form of this condition can manifest from infancy to adulthood and variable onset, frequency, and severity of symptoms have been reported (PMID: 32295037). (I) 0200 - Variant is predicted to result in a missense amino acid change from serine to leucine. (I) 0251 - This variant is heterozygous. (I) 0304 - Variant is present in gnomAD <0.01 for a recessive condition (v2) (385 heterozygotes, 4 homozygotes) and is enriched in the Ashkenazi Jewish sub-population. (SP) 0501 - Missense variant consistently predicted to be damaging by multiple in silico tools or highly conserved with a major amino acid change. (SP) 0600 - Variant is located in the annotated Choline/Carnitine o-acyltransferase domain (NCBI domain). (I) 0801 - This variant has strong previous evidence of pathogenicity in unrelated individuals. This variant has been reported as the most common cause of the myopathic form of CPT II deficiency in individuals of European ancestry (ClinVar, PMID: 8358442, PMID: 21913903, PMID: 32295037). (SP) 1208 - Inheritance information for this variant is not currently available in this individual. (I) Legend: (SP) - Supporting pathogenic, (I) - Information, (SB) - Supporting benign

NM_000098.2:c.338C>T in the CPT2 gene has an allele frequency of 0.008 in Ashkenazi Jewish subpopulation in the gnomAD database. The p.Ser113Leu (NM_000098.2:c.338C>T) variant has been observed to segregate with disease in one family with three affected individuals and two unaffected individuals (PMID: 8786066). Martin MA et al performed molecular analysis in a group of 14 Spanish patients with CPT II deficiency from ten unrelated families. The S113L mutation was observed in 8 of the 14 patients studied. Seven patients were homozygous for the mutation (PMID: 10398215). In addition, Anichini A et al reported that p.Ser113Leu was identified in five homozygote patients and in compound heterozygosity in five cases: c.338C>T (p.Ser113Leu)/c.1724T>C (p.Leu575Pro); c.338C>T (p.Ser113Leu)/c.1547T>C (p.Phe516Ser); c.338C>T (p.Ser113Leu) c.1348A>T (p.Arg450Stop); c.338C>T (p.Ser113Leu)/c.1932insA (p.E645RfsX4); c.338C>T (p.Ser113Leu)/c.1239G>A (p.Gln413Gln) (PMID: 20810031). Functional analysis demonstrates that the c.338C>T variant is associated with a significant reduction in enzyme activity (PMID:8358442). Pathogenic computational verdict because pathogenic predictions from DANN, DEOGEN2, EIGEN, FATHMM-MKL, M-CAP, MVP, MutationAssessor, MutationTaster, PrimateAI, REVEL and SIFT. Taken together, we interprete this variant as Pathogenic/Likely pathogenic variant. ACMG/AMP criteria applied: PS3; PM3_Strong; PP1_Moderate; PP3.

Variant interpretted as Pathogenic and reported most recently on 11-25-2015 by Lab or GTR ID 500068. The variant was also interpretted as Pathogenic and reported on 11/17/2015 by GTR ID 26957. GenomeConnect assertions are reported exactly as they appear on the patient-provided report from the testing laboratory. GenomeConnect staff make no attempt to reinterpret the clinical significance of the variant.