ClinVar Genomic variation as it relates to human health

The NM_001114753.3: c.991G>A variant in ENG is a missense variant predicted to cause substitution of glycine by serine at amino acid 331 (p.Gly331Ser). This variant is absent from gnomAD v2.1.1 (PM2_Supporting). This variant has been reported in >10 probands with a phenotype consistent with HHT (PS4; PMID: 19767588, 34872578, 16690726, 34880085, 15517393, 25970827, 29171923, 32573726, 21158752, Internal lab contributors). At least one patient's phenotype meets Curacao Criteria for HHT, and sequencing and large deletion/duplication analysis was performed for ENG and ACVRL1, which is highly specific for HHT (PP4_Moderate; PMID: 19767588, 21158752). The variant has been reported to segregate with HHT in 3 affected meioses from 2 families (PP1; PMID: 19767588, 34872578). The computational predictor REVEL gives a score of 0.202, which is neither above nor below the thresholds predicting a damaging or benign impact on ENG function. However, RT-PCR analysis of cDNA demonstrated that the variant impacts splicing by leading to exon 7 skipping (PS3; PMID: 16690726). In summary, this variant meets the criteria to be classified as pathogenic for autosomal dominant hereditary hemorrhagic telangiectasia based on the ACMG/AMP criteria applied, as specified by the ClinGen Hereditary Hemorrhagic Telangiectasia Variant Curation Expert Panel: PS3, PS4, PP4_Moderate, PP1, PM2_Supporting (specification version 1.0.0; 1/4/2024).

PM2+PP3+PP4+PP5

The ENG c.991G>A; p.Gly331Ser variant (rs1060501410) is published in the literature in individuals and families diagnosed with HHT (Bossler 2006, Gedge 2007, Letteboer 2005, McDonald 2009, Nishida 2012), and is reported as pathogenic in ClinVar (Variation ID: 407115). This variant is absent from general population databases (Exome Variant Server, Genome Aggregation Database), indicating it is not a common polymorphism. This variant occurs in the last nucleotide of the exon, which is a highly conserved nucleotide, and computational algorithms (Alamut v.2.11) predict this variant results in altered mRNA splicing. In support of these predictions, a functional study shows this alteration leads to exon 7 skipping (Prigoda 2006). Based on available information, this variant is considered to be pathogenic. REFERENCES Bossler AD et al. Novel mutations in ENG and ACVRL1 identified in a series of 200 individuals undergoing clinical genetic testing for hereditary hemorrhagic telangiectasia (HHT): correlation of genotype with phenotype. Hum Mutat. 2006 Jul;27(7):667-75. Gedge F et al. Clinical and analytical sensitivities in hereditary hemorrhagic telangiectasia testing and a report of de novo mutations. J Mol Diagn. 2007 Apr;9(2):258-65. Letteboer TG et al. Hereditary hemorrhagic telangiectasia: ENG and ALK-1 mutations in Dutch patients. Hum Genet. 2005 Jan;116(1-2):8-16. McDonald J et al. Multiple sequence variants in hereditary hemorrhagic telangiectasia cases: illustration of complexity in molecular diagnostic interpretation. J Mol Diagn. 2009 Nov;11(6):569-75. Nishida T et al. Brain arteriovenous malformations associated with hereditary hemorrhagic telangiectasia: gene-phenotype correlations. Am J Med Genet A. 2012 Nov;158A(11):2829-34. Prigoda NL et al. Hereditary haemorrhagic telangiectasia: mutation detection, test sensitivity and novel mutations. J Med Genet. 2006 Sep;43(9):722-8.

Based on the classification scheme VCGS_Germline_v1.3.3, this variant is classified as Pathogenic. Following criteria are met: 0103 - Dominant negative and loss of function are known mechanisms of disease in this gene and are associated with Hereditary Hemorrhagic Telangiectasia type 1 (MIM#187300). Loss-of-function variants have been reported, while missense variants have been demonstrated to have both loss of function and dominant negative effects on protein activity (PMID: 25312062). (I). 0107 - This gene is associated with autosomal dominant disease. (I) 0200 - Variant is predicted to result in a missense amino acid change from glycine to serine. (I) 0212 - Non-canonical splice site variant without proven consequence on splicing (no functional evidence available). This variant affects the last nucleotide of exon 7 and it was said that RT-PCR of a patient’s RNA showed splicing defect leading to exon skipping. However, data was not shown in the manuscript for independent confirmation (PMID: 16690726). (SP) 0251 - This variant is heterozygous. (I) 0301 - Variant is absent from gnomAD (both v2 and v3). (SP) 0503 - Missense variant consistently predicted to be tolerated by multiple in silico tools or not conserved in placental mammals with a minor amino acid change. (SB) 0505 - Abnormal splicing is predicted by in silico tools but affected nucleotide is moderately conserved. (I) 0604 - Variant is not located in an established domain, motif, hotspot or informative constraint region. (I) 0801 - This variant has strong previous evidence of pathogenicity in at least 10 unrelated individuals with Hereditary Hemorrhagic Telangiectasia type 1 (MIM#187300) (ClinVar; PMIDs: 15517393, 17384219, 19767588, 24196379, 25970827, 29171923). (SP) 1208 - Inheritance information for this variant is not currently available in this individual. (I) Legend: (SP) - Supporting pathogenic, (I) - Information, (SB) - Supporting benign

Not observed in large population cohorts (gnomAD); Functional studies and in silico splice prediction algorithms suggest that the c.991 G>A variant, which affects the last nucleotide of exon 7, damages the natural splice donor site for intron 7 and leads to skipping of exon 7 (Prigoda et al., 2006); 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: 24196379, 19767588, 15517393, 25970827, 16752392, 17384219, 29171923, 22991266, 16690726, 26582918, 34880085, 34872578, 32573726)

This sequence change replaces glycine, which is neutral and non-polar, with serine, which is neutral and polar, at codon 331 of the ENG protein (p.Gly331Ser). This variant also falls at the last nucleotide of exon 7, which is part of the consensus splice site for this exon. This variant is not present in population databases (gnomAD no frequency). This missense change has been observed in individuals with hereditary hemorrhagic telangiectasia (HHT) and brain arteriovenous malformations (BAVM) (PMID: 15517393, 16690726, 19767588, 22991266, 24196379, 25970827). ClinVar contains an entry for this variant (Variation ID: 407115). 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. Variants that disrupt the consensus splice site are a relatively common cause of aberrant splicing (PMID: 17576681, 9536098). For these reasons, this variant has been classified as Pathogenic.

The c.991G>A pathogenic mutation (also known as p.G331S), located in coding exon 7 of the ENG gene, results from a G to A substitution at nucleotide position 991. The glycine at codon 331 is replaced by serine, an amino acid with similar properties. However, this change occurs in the last base pair of coding exon 7, which makes it likely to have some effect on normal mRNA splicing. A functional study demonstrated that this mutation results in the skipping of coding exon 7 (Prigoda NL et al. J. Med. Genet., 2006 Sep;43:722-8). This mutation has been reported in multiple individuals with a clinical diagnosis of hereditary hemorrhagic telangiectasia (Letteboer TG et al. Hum. Genet., 2005 Jan;116:8-16; McDonald J et al. J Mol Diagn, 2009 Nov;11:569-75). This variant is considered to be rare based on population cohorts in the Genome Aggregation Database (gnomAD). This nucleotide position is highly conserved in available vertebrate species. Based on the supporting evidence, this alteration is interpreted as a disease-causing mutation.

PP3, PM2, PS3_moderate, PS4