ClinVar Genomic variation as it relates to human health

The NM_000527.5(LDLR):c.1784G>A (p.Arg595Gln) variant is classified as Pathogenic for Familial Hypercholesterolemia by applying evidence codes (PM2, PP3, PM5, PS3_supporting, PS4, PP4, PM3) as defined by the ClinGen Familial Hypercholesterolemia Expert Panel LDLR-specific variant curation guidelines (https://doi.org/10.1016/j.gim.2021.09.012). The supporting evidence is as follows: PM2 - PopMax MAF = 0.0002389 (0.02389%) in European (Finnish) exomes+genomes (gnomAD v2.1.1), since this is a founder effect variant in Finnish population (PMID: 11585102) we can consider PopMax MAF = 0.00005012 (0.005012%) in East Asian exomes+genomes (gnomAD v2.1.1), so PM2 is Met. PP3 - REVEL = 0.956. It is above 0.75, so PP3 is Met. PM5 - 2 other missense variants in the same codon: - NM_000527.5(LDLR):c.1783C>T (p.Arg595Trp) (ClinVar ID: 161290) - Pathogenic by these guidelines - NM_000527.5(LDLR):c.1784G>T (p.Arg595Leu) (ClinVar ID: 252029) - Likely pathogenic by these guidelines There is 1 variant in the same codon classified as Pathogenic by these guidelines, so PM5 is Met. PS3_supporting - Level 3 assay: PMID 11585102: Heterozygous patient cells, 125I-LDL assays - result - 70% LDL-LDLR binding, 54-49% LDL-LDLR uptake and degradation. ---- functional study is consistent with damaging effect, so PS3_Supporting is Met. PS4 - Variant meets PM2 and is identified in 11 unrelated index cases: 1 index case with Simon Broome definite criteria from Molecular Genetics Laboratory (Centre for Cardiovascular Surgery and Transplantation), 4 index cases with DLCN criteria for FH (LDL > 95th percentile and presence of tendon xanthomas) from Finland (PMID: 11585102), 1 index case with FH criteria suggested by the Japan Atherosclerosis Society (TC > 260 mg/dL with tendon xanthomas OR TC > 260 mg/dL and TC > 260 mg/dL with tendon xanthomas in first or second degree relatives) from Japan ( PMID: 18718593), 1 index case with DLCN criteria for FH (score=8) from Sweden (PMID: 29974534), 1 index case with DLCN criteria for FH (score>=6) from Argentina (PMID: 30270055), 2 index cases with DLCN criteria for FH (score>=6) from Russia (PMID: 33418990), 1 index case with homozygous FH phenotype (LDL = 16 mmol/L) from The Netherlands (PMID: 24585268), so PS4 is Met. PP4 - Variant meets PM2 and is identified in 11 unrelated index cases as described before, so PP4 is Met. PM3 - Variant meets PM2 and is identified in an index case with homozygous FH phenotype (LDL = 16 mmol/L) and LDLR variant c.(1705+1_1706-1)_(*2514_?)del (described as 16kb deletion exon 12-18) (PMID: 24585268), classified as Pathogenic by these guidelines, in trans, so PM3 is Met.

The variant has been reported in individuals with familial hypercholesterolemia and one healthy control in the published literature (PMID: 27497240 (2016), 25487149 (2015), 25437892 (2014), 18718593 (2009), 16250003 (2005), 15359125 (2004), 15256764 (2004)). One functional study indicated that this variant does not have an impact on LDL uptake in vitro (PMID: 25647241 (2015)). Analysis of this variant using bioinformatics tools for the prediction of the effect of amino acid changes on protein structure and function yielded predictions that this variant is disease causing and damaging. A different variant affecting the same amino acid position has been described as pathogenic. Based on the available information, the variant is predicted to be likely pathogenic.

DNA sequence analysis of the LDLR gene demonstrated a sequence change, c.1784G>A, in exon 12 that results in an amino acid change, p.Arg595Gln. The p.Arg595Gln change affects a highly conserved amino acid residue located in a domain of the LDLR protein that is known to be functional. The p.Arg595Gln substitution appears to be deleterious using several in-silico pathogenicity prediction tools (SIFT, PolyPhen2, Align GVGD, REVEL, CADD). This particular amino acid change has been reported in multiple individuals with familial hypercholesterolemia (Hooper et al., 2012; PMIDs: 15359125, 16250003, 15256764). Additionally, a different pathogenic sequence change affecting the same amino acid residue (p.Arg595Trp) has been described in patients with LDLR-related hypercholesterolemia (PMIDs: 25461735, 27784735). This sequence change has been described in the gnomAD database with a low population frequency of 0.004% (dbSNP rs201102492).

In silico analysis supports that this missense variant has a deleterious effect on protein structure/function; Published functional studies suggest a damaging effect as there is a reduction in LDLR binding, uptake, and degredation (Vuorio et al., 2001); This variant is associated with the following publications: (PMID: 26332594, 16250003, 9399845, 15256764, 15359125, 33740630, 34037665, 25487149, 25647241, 11585102, 18718593, 24585268, 30270055, 33418990, 29974534)

This sequence change replaces arginine, which is basic and polar, with glutamine, which is neutral and polar, at codon 595 of the LDLR protein (p.Arg595Gln). This variant is present in population databases (rs201102492, gnomAD 0.02%). This missense change has been observed in individuals with familial hypercholesterolemia and/or myocardial infarction (PMID: 15256764, 15359125, 16250003, 18718593, 25437892, 25487149, 27497240). This variant is also known as p.Arg574Gln and FH-Pogosta. ClinVar contains an entry for this variant (Variation ID: 183126). 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 LDLR protein function with a positive predictive value of 95%. Experimental studies have shown that this missense change does not substantially affect LDLR function (PMID: 25647241). This variant disrupts the p.Arg595 amino acid residue in LDLR. Other variant(s) that disrupt this residue have been determined to be pathogenic (PMID: 11737238, 16250003, 20538126, 25461735, 27784735). This suggests that this residue is clinically significant, and that variants that disrupt this residue are likely to be disease-causing. For these reasons, this variant has been classified as Pathogenic.

This missense variant replaces arginine with glutamine at codon 595 of the LDLR protein. This variant is also known as p.Arg574Gln in the mature protein, and FH-Pogosta in the literature. This variant alters a conserved arginine residue in the LDLR type B repeat 5 of the LDLR protein (a.a. 572-615), where pathogenic missense variants are found enriched (ClinVar-LDLR). Computational prediction tools indicate that this variant has a deleterious impact on protein structure and function. An experimental study using fibroblasts derived from a heterozygous carrier individual has shown that this variant causes a significant defect in LDL binding, internalization and degradation (PMID: 11585102). Another in vitro high throughput functional study using transfected HeLa cells has shown that this variant causes normal LDL uptake (PMID: 25647241). This LDLR variant has been reported in over 10 heterozygous individuals affected with familial hypercholesterolemia (PMID: 11585102, 15256764, 15359125, 16250003, 18718593, 27497240, 28161202, 29974534, 30270055, 33418990, 37848354). This variant has also been observed in compound heterozygous state with a known pathogenic LDLR variant in one individual affected with severe homozygous familial hypercholesterolemia, a phenotype expected of having two deleterious LDLR variants (PMID: 24585268). This variant has also been reported in two individuals affected with early-onset myocardial infarction (PMID: 25487149). This variant has been identified in 11/282862 chromosomes in the general population by the Genome Aggregation Database (gnomAD). A different variant affecting the same codon, p.Arg595Trp, is considered to be disease-causing (ClinVar variation ID: 183126), suggesting that arginine at this position is important for LDLR protein function. Based on the available evidence, this variant is classified as Pathogenic.

PS4_moderate, PM1, PM2, PM3, PP3, PP4

This missense variant (also known as p.Arg574Gln in the mature protein and as FH-Pogosta) is located in the LDLR type B repeat 5 of the EGF precursor homology domain of the LDLR protein. Computational prediction tools and conservation analyses suggest that this variant may have deleterious impact on the protein function. An experimental study has shown that this variant causes significant defect in LDL binding, internalization and degradation in cells from a heterozygous carrier (PMID: 11585102). Another study has shown normal LDL uptake in a high throughput screening assay in vitro (PMID: 25647241). This variant has been identified in over 10 individuals affected with familial hypercholesterolemia and myocardial infarction, and reported to be a prevalent mutation in Finland (PMID: 11585102, 15359125, 15256764, 16250003, 18718593, 25437892, 25487149, 27497240). This variant has been identified in 11/282862 chromosomes in the general population by the Genome Aggregation Database (gnomAD). A different missense variant p.Arg595Trp occurring at the same codon is associated with familial hypercholesterolemia (Clinvar), suggesting that arginine at this codon is important for the protein function. Based on available evidence, this variant is classified as Likely Pathogenic.

The c.1784G>A (p.R595Q) alteration is located in exon 12 (coding exon 12) of the LDLR gene. This alteration results from a G to A substitution at nucleotide position 1784, causing the arginine (R) at amino acid position 595 to be replaced by a glutamine (Q). Based on data from gnomAD, the A allele has an overall frequency of 0.004% (11/282862) total alleles studied. The highest observed frequency was 0.024% (6/25120) of European (Finnish) alleles. This alteration (also described as R574Q and FH-Pogosta) has been detected in multiple individuals who met clinical criteria for familial hypercholesterolemia (FH) and has been described as a Finnish founder mutation (Vuorio, 2001; Kim, 2004; Maurer, 2016; Corral, 2018; Junna, 2023). This amino acid position is highly conserved in available vertebrate species. This alteration is predicted to be deleterious by in silico analysis. Based on the available evidence, this alteration is classified as pathogenic.

PM2, PP3, PM5, PS4