ClinVar Genomic variation as it relates to human health

The NM_001114753.3: c.-9G>A variant is located in the 5' UTR of ENG. Because the variant is located in the 5' UTR, is it not expected to alter the amino acid sequence. The highest population minor allele frequency in gnomAD v2.1.1 is 0.0007942 (76/95698 alleles) in the European (non-Finnish) population. This variant has been observed in patients with an alternate molecular basis for disease (patients also carry a likely pathogenic/pathogenic ACVRL1 variant) (BP5; Internal lab contributors). Expression studies showed this variant causes a small reduction (approximately 18.4%) of endoglin compared to wild type (PS3_Supporting, PMID: 22192717). Due to conflicting evidence, this variant is classified as a variant of uncertain significance 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_Supporting, BP5 (specification version 1.0.0; 1/4/2024).

This variant was observed as part of a predisposition screen in an ostensibly healthy population. A literature search was performed for the gene, cDNA change, and amino acid change (where applicable). Publications were found based on this search. However, the evidence from the literature, in combination with allele frequency data from public databases where available, was not sufficient to rule this variant in or out of causing disease. Therefore, this variant is classified as a variant of unknown significance.

The ENG c.-9G>A variant (rs368423516 ) has been described in the literature in families with hereditary hemorrhagic telangiectasia or pulmonary arterial hypertension and has been reported as a hypomorphic allele causing reduced functionality (Best 2017, Kim 2011, McDonald 2011, McDonald 2020). Additionally, this variant has been shown to cause reduced translation (Damjanovich 2011). The variant is listed in the ClinVar database (Variation ID: 414302) and in the general population with an allele frequency of 0.05% (108/222760 alleles) in the Genome Aggregation Database. The nucleotide at the -9 position is moderately conserved across mammals and creates a novel methionine translational start, leading to the inclusion of additional amino acids. Due to its description in the literature as a hypomorphic allele with reduced translation, we consider the c.-9G>A variant to be pathogenic with mild clinical outcomes. References: Best DH et al. EIF2AK4 Mutations in Patients Diagnosed With Pulmonary Arterial Hypertension. Chest. 2017 Apr;151(4):821-828 Damjanovich K et al. 5'UTR mutations of ENG cause hereditary hemorrhagic telangiectasia. Orphanet J Rare Dis. 2011 Dec 22;6:85. Kim MJ et al. Clinical and genetic analyses of three Korean families with hereditary hemorrhagic telangiectasia. BMC Med Genet. 2011 Oct 3;12:130. McDonald J et al. Curacao diagnostic criteria for hereditary hemorrhagic telangiectasia is highly predictive of a pathogenic variant in ENG or ACVRL1 (HHT1 and HHT2). Genet Med. 2020 Jul;22(7):1201-1205. McDonald J et al. Molecular diagnosis in hereditary hemorrhagic telangiectasia: findings in a series tested simultaneously by sequencing and deletion/duplication analysis. Clin Genet. 2011 Apr;79(4):335-44.

This substitution variant occurs in the 5'UTR of the ENG gene and it introduces a premature start codon 9bp prior to the existing start codon, maintaining the reading frame but elongating the protein by three additional amino acid residues. This variant has been observed in gnomAD with a total MAF of 0.0549% across multiple subpopulations, but occurring with the highest MAF of 0.0921% in the European population; these frequencies are inconsistent with the disease frequency. This position is not conserved. This variant has been identified in the literature in both families and individuals with hereditary hemorrhagic telangiectasia, both in the heterozygous state and homozygous state (HHT) (PMID: 21158752, 22192717); segregation of the variant within affected families has been inconsistent or incompletely assessed (PMID: 22192717). A single in vitro protein expression study suggested that this variant may be hypomorphic after it demonstrated that this variant causes decreased expression of the protein, to only ~80% of the wild-type, and it was suggested that this may be a mild variant when heterozygous and cause a more classic HHT presentation when homozygous, where expression dropped to ~60% of the wild-type (PMID: 22192717). In the literature, this variant has also been called benign or a polymorphism due to its frequency in a population of individuals with suspected or possible HHT (PMID: 17384219). Considering this variant occurs relatively frequently, despite having been observed in some individuals and families who are affected, the impact of this variant and segregation with disease require further investigations to elucidate pathogenicity; thus this variant is classified as a Variant of Uncertain Significance.

This variant occurs in a non-coding region of the ENG gene. It does not change the encoded amino acid sequence of the ENG protein. This variant is present in population databases (rs368423516, gnomAD 0.09%). This variant has been observed in individual(s) with clinical features of hereditary hemhorrhagic telangiectasia (PMID: 21158752, 22192717, 32300199; Invitae). It has also been observed to segregate with disease in related individuals. ClinVar contains an entry for this variant (Variation ID: 414302). Studies have shown that this variant alters ENG gene expression (PMID: 22192717). In summary, the available evidence is currently insufficient to determine the role of this variant in disease. Therefore, it has been classified as a Variant of Uncertain Significance.

Variant summary: ENG c.-9G>A is located in the untranslated mRNA region upstream of the initiation codon, predicted to introduce a premature start codon. The variant allele was found at a frequency of 0.00052 in 191406 control chromosomes. The observed variant frequency is approximately 12.41 fold of the estimated maximal expected allele frequency for a pathogenic variant in ENG causing Hereditary Hemorrhagic Telangiectasia phenotype (4.2e-05), these frequencies are inconsistent with disease frequencies. c.-9G>A has been reported in the literature in heterozygous and homozygous individuals affected with symptoms of Hereditary Hemorrhagic Telangiectasia (Damjanovich_2011, Gedge_2007, McDonald_2011). Segregation was observed in some families. Functional studies of this variant showed that expression of the mutant protein was reduced approximately 20% compared to the wild type protein (Damjanovich_2011). Five clinical diagnostic laboratories have submitted clinical-significance assessments for this variant to ClinVar after 2014 and classified as VUS (n=3), Likely Pathogenic (n=1) and Pathogenic (n=1). Based on the evidence outlined above, the variant was classified as uncertain significance.

BS1, PP1, PM4, PS3_supporting, PS4_moderate

This sequence change in ENG is located in the 5' untranslated region. It introduces an alternative initiation codon three residues upstream from the existing initiation codon. In vitro protein expression analyses showed the variant decreases protein expression by around 20% (PMID: 22192717). The highest population minor allele frequency in the population database gnomAD v2.1 is 0.08% (76/95,698 alleles) in the European (non-Finnish) population. This variant has been described as a hypomorphic allele and reported in at least three probands (one homozygous) with epistaxis and telangiectasia suggestive of hereditary haemorrhagic telangiectasia but without visceral arteriovenous malformations (PMID: 22192717). It has also been reported in a proband with idiopathic primary arterial hypertension (PMID: 27884767). The variant has been reported to segregate with epistaxis and telangiectasia in multiple individuals from two families (PMID: 22192717). The variant has conflicting pathogenicity interpretations in ClinVar (ID: 414302). Based on the classification scheme RMH Modified ACMG/AMP Guidelines v1.6.1, this variant is classified as a VARIANT OF UNCERTAIN SIGNIFICANCE. Following criteria are met: PP1.

The c.-9G>A alteration is located in the 5' untranslated region (5'UTR) of the ENG gene. This alteration consists of a G to A substitution 9 nucleotides upstream from the first translated codon. Based on insufficient or conflicting evidence, the clinical significance of this alteration remains unclear.

This variant is classified as likely benign based on ACMG/AMP sequence variant interpretation guidelines (Richards et al. 2015 PMID: 25741868, with internal and published modifications).