ClinVar Genomic variation as it relates to human health

The c.571G>A (p.Glu191Lys) variant in the ALPL gene is observed at a minor allele frequency (MAF) of 1.6% in the European (Finnish) population, including 3 homozygous individuals (https://gnomad.broadinstitute.org/variant/1-21890632-G-A). This variant has been reported in multiple individuals affected with autosomal recessive hypophosphatasia and observed to segregate with the disease in family studies (PMID: 12357339, PMID: 24569605). In vitro functional studies have suggested this variant mildly reduced the alkaline phosphatase (ALP) activity (PMID: 24569605).

Published functional studies demonstrate that E191K has delayed membrane anchoring (Brun-Heath et al., 2007) and relatively high residual alkaline phosphatase activity (56% and 88% of wildtype), though no dominant negative effect was observed (Zurutuza et al., 1999; Fauvert et al., 2009; Hofmann et al., 2014); Observed in homozygous state in large population cohorts (gnomAD) and in a clinically unaffected adult relative of an individual referred for genetic testing at GeneDx, supporting that this mild variant leads to disease only when a more severe variant is present on the opposite allele (in trans); In silico analysis, which includes protein predictors and evolutionary conservation, supports that this variant does not alter protein structure/function; This variant is associated with the following publications: (PMID: 24569605, 20739387, 1409720, 19500388, 11855933, 10332035, 18328985, 17719863, 11438998, 12357339, 29659871, 10679946, 19232125, 27920814, 10737975, 29236161, 15671102, 32160374, 31589614, 33083013, 8606878, 33101980, 34213743, 33093890, 33549410)

This sequence change replaces glutamic acid, which is acidic and polar, with lysine, which is basic and polar, at codon 191 of the ALPL protein (p.Glu191Lys). This variant is present in population databases (rs121918007, gnomAD 1.7%), and has an allele count higher than expected for a pathogenic variant. This missense change has been observed in individuals with hypophosphatasia (PMID: 12357339, 15671102, 24569605). It has also been observed to segregate with disease in related individuals. This variant is also known as p.Glu174Lys. ClinVar contains an entry for this variant (Variation ID: 13670). 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 ALPL protein function with a positive predictive value of 95%. Experimental studies have shown that this missense change affects ALPL function (PMID: 1409720, 20739387, 24569605). For these reasons, this variant has been classified as Pathogenic.

The ALPL c.571G>A (p.Glu191Lys) missense variant, also known as p.Glu174Lys, is a well-documented pathogenic variant. It was first identified by Henthorn et al. (1992) in seven out of 46 patients with variable clinical presentation. At least four of these patients were compound heterozygotes for the p.Glu191Lys variant and a second variant, while zygosity was not confirmed in other patients. These patients represented three clinical forms of hypophosphatasia: perinatal (lethal), childhood, and adult and showed autosomal recessive inheritance. Hérasse et al. (2002) reported that the p.Glu191Lys variant is a recurrent variant found in approximately 7% of affected Caucasian chromosomes. In this patient population, the p.Glu191Lys variant was seen in 31% of patients with mild hypophosphatasia (child, adult, and odonto forms). The p.Glu191Lys variant is found at a frequency of 0.01995 in the European (Finnish) population of the Exome Aggregation Consortium. In vitro functional studies performed by Fauvert et al. (2009) showed that the p.Glu191Lys variant has a residual activity of 56% compared to wild type. Based on the collective evidence, the p.Glu191Lys variant is classified as pathogenic for hypophosphatasia. This variant was observed by ICSL as part of a predisposition screen in an ostensibly healthy population.

NM_000478.4(ALPL):c.571G>A(E191K, aka E174K) is classified as pathogenic in the context of hypophosphatasia. Sources cited for classification include the following: PMID 25731960, 11438998, 19232125, 10679946, 11855933 and 10332035. Classification of NM_000478.4(ALPL):c.571G>A(E191K, aka E174K) 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 c.571G>A (p.E191K) alteration is located in exon 6 (coding exon 5) of the ALPL gene. This alteration results from a G to A substitution at nucleotide position 571, causing the glutamic acid (E) at amino acid position 191 to be replaced by a lysine (K). Based on data from gnomAD, the A allele has an overall frequency of 0.25% (699/282574) total alleles studied. The highest observed frequency was 1.65% (414/25032) of European (Finnish) alleles. This variant, also described as E174K in literature, is a common founder mutation accounting for up to approximately 10% of disease-causing mutations in the ALPL gene (Hérasse, 2002; Mornet, 2021). This variant has been reported in the heterozygous (Hofmann, 2014; Taillandier, 2018; Mornet, 2021; Ambry internal data) and compound heterozygous (Henthorn, 1992; Schalin-Jäntti, 2010; Hofmann, 2014; Zurutuza, 1999; Mornet, 2021; Sperelakis-Beedham, 2021) states in individuals with hypophosphatasia and low alkaline phosphatase, with the clinical severity correlating with degree of loss of function. This amino acid position is not well conserved in available vertebrate species. Multiple in vitro functional studies using transient expression in cells show slightly reduced activity consistent with a mild loss of function allele based on comparison to wild type and pathogenic controls (Hofmann, 2014; Zurutuza, 1999; Del Angel, 2020). This alteration is predicted to be deleterious by in silico analysis. Based on the available evidence, this alteration is classified as pathogenic.

ACMG categories: PS3,PM1,PP3,PP5,BS1

ALPL: PM3:Very Strong, PM5, PM2:Supporting, PP3, PS3:Supporting

Based on the classification scheme VCGS_Germline_v1.3.4, 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. Later-onset/mild disease is associated with monoallelic dominant negative variants or biallelic loss of function variants that retain residual enzyme activity, while early-onset/severe disease is caused by more complete loss of function variants (PMIDs: 20301329, 19500388). (I) 0108 - This gene is associated with both recessive and dominant disease. Severe forms of hypophosphatasia (perinatal and infantile) are generally associated with autosomal recessive disease, while the milder forms of hypophosphatasia (childhood, adult and odontohypophosphatasia) have been associated with both autosomal dominant and recessive disease (PMID: 19500388, 23688511). (I) 0112 - The condition associated with this gene has incomplete penetrance (PMID: 20301329). (I) 0115 - Variants in this gene are known to have variable expressivity (PMID: 20301329). (I) 0200 - Variant is predicted to result in a missense amino acid change from glutamic acid to lysine. (I) 0251 - This variant is heterozygous. (I) 0305 - Variant is present in gnomAD v2 >=0.01 and <0.03 for a recessive condition (689 heterozygotes, 5 homozygotes). (I) 0309 - An alternative amino acid change at the same position has been observed in gnomAD (v2) (1 heterozygote, 0 homozygotes). (I) 0502 - Missense variant with conflicting in silico predictions and uninformative conservation. (I) 0600 - Variant is located in the annotated alkaline phosphatase domain (DECIPHER). (I) 0801 - This variant has strong previous evidence of pathogenicity in unrelated individuals. This is a well-established pathogenic variant and has been observed in many individuals with hypophophatasia (ClinVar, PMIDs: 32973344, 32811521) . (SP) 1002 - This variant has moderate functional evidence supporting abnormal protein function. Site directed mutagenesis studies have shown this variant has residual activity of 56% compared to wild type and is thought to be a moderate allele (PMID: 19500388). (SP) 1101 - Very strong and specific phenotype match for this individual. (SP) 1206 - This variant has been shown to be paternally inherited (by segregation analysis, VCGS #22G000749). (I) Legend: (SP) - Supporting pathogenic, (I) - Information, (SB) - Supporting benign

PP3, PM3_very_strong, PS3, PS4

NM_000478.4:c.571G>A in the ALPL gene has an allele frequency of 0.017 in European(Finnish) subpopulation in the gnomAD database. Functional studies demonstrate that c.571G>A has reduced alkaline phosphatase activity due to structural disturbances and delay in membrane anchoring (PMID:17719863). It was detected in multiple individuals with autosomal recessive hypophosphatasia, compound heterozygous with p.Gly334Asp (PMID: 24569605), c.186G>C (p.M45I) (PMID: 15671102). The patient's phenotype is highly specific for ALPL gene (PMID: 15671102). Co-segregation evidence in a pedigree,one patient was affected and two sister unaffected (PMID:15671102). Pathogenic computational verdict because pathogenic predictions from DANN, DEOGEN2, EIGEN, FATHMM-MKL, MVP, MutationTaster, PrimateAI and REVEL. Phenotype date Taken together, we interprete this variant as Pathogenic/Likely pathogenic variant. ACMG/AMP criteria applied: PS3; PM3_Strong; PP1; PP4; PP3.

The ALPL c.571G>A variant is predicted to result in the amino acid substitution p.Glu191Lys. This variant, alternatively referred to as p.Glu174Lys using legacy nomenclature, has been reported in the compound heterozygous state in numerous individuals with hypophosphatasia (HPP), predominantly childhood, adult and odontohypophosphatasia forms (see, for example, Henthorn et al. 1992. PubMed ID: 1409720, reported as 747A (p.Glu174Lys); Fauvert et al. 2009. PubMed ID: 19500388; Hofmann et al. 2014. PubMed ID: 24569605). It has also been reported in the heterozygous state in individuals with odontohypophosphatasia (Fauvert et al. 2009. PubMed ID: 19500388). This variant was also shown to segregate with HPP in two families (Henthorn et al. 1992. PubMed ID: 1409720; Hofmann et al. 2014. PubMed ID: 24569605). Functional studies using COS7 and HEK293 cells show mild reduction in tissue non-specific alkaline phosphatase (TNSALP) activity and also indicate this variant does not result in a dominant-negative effect. It is speculated that phenotypic variability is related to either an unidentified second variant or varying alkaline phosphatase tolerance thresholds (Fauvert et al. 2009. PubMed ID: 19500388; Hofmann et al. 2014. PubMed ID: 24569605). An alternate nucleotide change affecting the same amino acid, p.Glu191Gly (p.Glu174Gly using legacy nomenclature), has been reported in a patient with HPP (Goseki-Sone et al. 1998. PubMed ID: 9452105, reported as p.Glu174Gly; Michigami et al. 2019. PubMed ID: 31707452). This variant is reported in 1.7% of alleles in individuals of European (Finnish) descent in gnomAD, including 5 homozygotes. Reduced penetrance and expressivity is known in ALPL-related disease. This variant is interpreted as pathogenic.