19-11102788-T-C
Variant summary
Our verdict is Pathogenic. Variant got 17 ACMG points: 17P and 0B. PP1_ModeratePM2PM3PS3_ModeratePP4PVS1_StrongPS4
This summary comes from the ClinGen Evidence Repository: The NM_000527.5(LDLR):c.313+2T>C variant is classified as Pathogenic for Familial Hypercholesterolemia by applying evidence codes PM2, PS3_moderate, PVS1_strong, PP1_moderate, PS4, PM3, PP4 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 = 0.00002941 (0.002941%) in European (non-Finnish) exomes (gnomAD v3.1.2), so PM2 is Met.PS3_moderate - Level 2 assays: PMID 19026292:Homozygous patient cells, 125I-LDL assays - result - 10% LDLR activity.---- functional study is consistent with damaging effect, so PS3_Moderate is Met.PVS1_strong - Variant is in +2 intronic position, but there are no studies on effect predicted, so give PVS1_Strong to be conservative.PP1_moderate - Variant segregates with FH phenotype in 4 informative meiosis from 2 families: 2 relatives with the phenotype and the variant from Robarts Research Institute, and 2 affected family members with the variant from Germany (PMID:26036859), so PP1_Moderate is Met.PS4 - Variant meets PM2 and is identified in 19 unrelated index cases from different labs (4 index cases with DLCN>6 from Robarts Research Institute; 4 index cases with DLCN>6 from Color Health, Inc; 1 index case who fulfills the following criteria for FH: i) total cholesterol >= 9.5 mmol/l, ii) triglycerides < 2 mmol/l, iii) presence of tendon xanthomas; or iv) occurrence of hypercholesterolemia or early onset of coronary artery disease in first degree relatives, from The Netherlands (PMID:7616128); 3 index cases with DLCN>6 (LDL between 7,98 - 9,50 mmol/l and hypercholesterolemia in 1st degree relative) from Germany (PMID:11462246); 1 index case who fulfills the following criteria for FH: (1) LDL > 95th percentile and triglycerides below 2 mmol/l, and (2) the presence of xanthoma or CHD in the proband or 1st degree relative with type IIa hypercholesterolemia, xanthoma, or CVD, from France (PMID:12436241); 1 index case with SB criteria for FH (Total cholesterol >290 mg/dL and LDL >200 mg/dL and severe hypercholesterolemia in 1st degree relatives), from Germany (PMID:14974088); 1 index case with definite FH according to SB criteria from Northern Ireland (PMID:16159606); 3 index cases fulfilling the following criteria for FH: i) total cholesterol > 8.0 mmol/l and LDL > 6 mmol/l; (ii) premature coronary or vascular disease, or a family history of cardiovascular disorder; and (iii) the presence of tendon xanthoma, from Denmark (PMID:16542394); 1 index case with definite FH according to SB criteria (total cholesterol = 658 mg/dl and presence of xanthomas, and primary hypercholesterolemia in the probands’ parents or 1st degree relatives), from USA (PMID:19026292)), so PS4 is Met.PM3 - Variant meets PM2 and is identified in a true homozygous with total cholesterol = 658 mg/dl (PMID:19026292), so PM3 is Met.PP4 - Variant meets PM2 and is identified in 19 unrelated index cases from different labs as described, so PP4 is Met. LINK:https://erepo.genome.network/evrepo/ui/classification/CA023690/MONDO:0007750/013
Frequency
Consequence
NM_000527.5 splice_donor, intron
Scores
Clinical Significance
Conservation
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ACMG classification
Verdict is Pathogenic. Variant got 17 ACMG points.
Transcripts
RefSeq
| Gene | Transcript | HGVSc | HGVSp | Effect | Exon rank | MANE | Protein | UniProt |
|---|---|---|---|---|---|---|---|---|
| LDLR | NM_000527.5 | c.313+2T>C | splice_donor_variant, intron_variant | Intron 3 of 17 | ENST00000558518.6 | NP_000518.1 |
Ensembl
Frequencies
GnomAD3 genomes 0.0000132 AC: 2AN: 152084Hom.: 0 Cov.: 31
GnomAD4 exome AF: 0.00000274 AC: 4AN: 1460906Hom.: 0 Cov.: 31 AF XY: 0.00000138 AC XY: 1AN XY: 726762
GnomAD4 genome 0.0000132 AC: 2AN: 152084Hom.: 0 Cov.: 31 AF XY: 0.00 AC XY: 0AN XY: 74288
ClinVar
Submissions by phenotype
Hypercholesterolemia, familial, 1 Pathogenic:15Benign:1
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This variant was determined to be pathogenic according to ACMG Guidelines, 2015 [PMID:25741868]. -
_x000D_ Criteria applied: PVS1_STR, PS4, PS3_MOD, PM3, PP1_MOD, PM2_SUP, PP4 -
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The NM_000527.5(LDLR):c.313+2T>C variant is classified as Pathogenic for Familial Hypercholesterolemia by applying evidence codes PM2, PS3_moderate, PVS1_strong, PP1_moderate, PS4, PM3, PP4 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 = 0.00002941 (0.002941%) in European (non-Finnish) exomes (gnomAD v3.1.2), so PM2 is Met. PS3_moderate - Level 2 assays: PMID 19026292: Homozygous patient cells, 125I-LDL assays - result - 10% LDLR activity. ---- functional study is consistent with damaging effect, so PS3_Moderate is Met. PVS1_strong - Variant is in +2 intronic position, but there are no studies on effect predicted, so give PVS1_Strong to be conservative. PP1_moderate - Variant segregates with FH phenotype in 4 informative meiosis from 2 families: 2 relatives with the phenotype and the variant from Robarts Research Institute, and 2 affected family members with the variant from Germany (PMID: 26036859), so PP1_Moderate is Met. PS4 - Variant meets PM2 and is identified in 19 unrelated index cases from different labs (4 index cases with DLCN>6 from Robarts Research Institute; 4 index cases with DLCN>6 from Color Health, Inc; 1 index case who fulfills the following criteria for FH: i) total cholesterol >= 9.5 mmol/l, ii) triglycerides < 2 mmol/l, iii) presence of tendon xanthomas; or iv) occurrence of hypercholesterolemia or early onset of coronary artery disease in first degree relatives, from The Netherlands (PMID: 7616128); 3 index cases with DLCN>6 (LDL between 7,98 - 9,50 mmol/l and hypercholesterolemia in 1st degree relative) from Germany (PMID: 11462246); 1 index case who fulfills the following criteria for FH: (1) LDL > 95th percentile and triglycerides below 2 mmol/l, and (2) the presence of xanthoma or CHD in the proband or 1st degree relative with type IIa hypercholesterolemia, xanthoma, or CVD, from France (PMID: 12436241); 1 index case with SB criteria for FH (Total cholesterol >290 mg/dL and LDL >200 mg/dL and severe hypercholesterolemia in 1st degree relatives), from Germany (PMID: 14974088); 1 index case with definite FH according to SB criteria from Northern Ireland (PMID: 16159606); 3 index cases fulfilling the following criteria for FH: i) total cholesterol > 8.0 mmol/l and LDL > 6 mmol/l; (ii) premature coronary or vascular disease, or a family history of cardiovascular disorder; and (iii) the presence of tendon xanthoma, from Denmark (PMID: 16542394); 1 index case with definite FH according to SB criteria (total cholesterol = 658 mg/dl and presence of xanthomas, and primary hypercholesterolemia in the probands’ parents or 1st degree relatives), from USA (PMID: 19026292)), so PS4 is Met. PM3 - Variant meets PM2 and is identified in a true homozygous with total cholesterol = 658 mg/dl (PMID: 19026292), so PM3 is Met. PP4 - Variant meets PM2 and is identified in 19 unrelated index cases from different labs as described, so PP4 is Met. -
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This variant changes a single nucleotide in the intron 3 canonical splice donor site of the LDLR gene and is predicted to cause aberrant splicing. Although RNA studies have not been performed for this variant, other variants affecting the same splice donor site have been shown to cause skipping of exon 3, resulting in an in-frame deletion of part of the ligand binding domain of the LDLR protein (PMID: 27821657). Functional studies in a homozygous patient fibroblast culture showed 10% LDLR activity compared to wild type cells (PMID: 19026292). This variant has been reported in numerous individuals affected with familial hypercholesterolemia (PMID: 7616128, 11462246, 14974088, 16542394, 19026292, 21475731, 28008010, 37119068). This variant has not been identified in the general population by the Genome Aggregation Database (gnomAD). Loss of LDLR function is a known mechanism of disease. Based on available evidence, this variant is classified as Pathogenic. -
PM2_Supporting+PVS1_Strong+PS3_Moderate+PS4+PP4 -
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not provided Pathogenic:3
PP1, PP4, PM2, PM3, PS3_moderate, PS4, PVS1_strong -
Deletions involving coding exons in this gene are frequently reported as pathogenic, regardless of frame prediction (Stenson et al., 2014); Canonical splice site variant expected to result in aberrant splicing, although in the absence of functional evidence the actual effect of this sequence change is unknown.; Reported as pathogenic or likely pathogenic in ClinVar by other clinical laboratories (ClinVar Variant ID# 189296; Landrum et al., 2016); This variant is associated with the following publications: (PMID: 22390909, 25525159, 14974088, 11462246, 12436241, 15199436, 16159606, 16542394, 26036859, 23936638, 21475731, 21935675, 15556092, 31447099, 32041611, 33303402, 32770674, 33740630, 34037665, 28008010, 7616128) -
LDLR: PS4, PVS1:Strong, PM2 -
Familial hypercholesterolemia Pathogenic:3
This sequence change affects a donor splice site in intron 3 of the LDLR gene. It is expected to disrupt RNA splicing. Variants that disrupt the donor or acceptor splice site typically lead to a loss of protein function (PMID: 16199547), and loss-of-function variants in LDLR are known to be pathogenic (PMID: 20809525, 28645073). This variant is not present in population databases (gnomAD no frequency). Disruption of this splice site has been observed in individuals with familial hypercholesterolemia (PMID: 7616128, 11462246, 14974088, 16542394, 21475731). Invitae Evidence Modeling of clinical and family history, age, sex, and reported ancestry of multiple individuals with this LDLR variant has been performed. This variant is expected to be pathogenic with a positive predictive value of at least 99%. This is a validated machine learning model that incorporates the clinical features of 363,995 individuals referred to our laboratory for LDLR testing. ClinVar contains an entry for this variant (Variation ID: 189296). Algorithms developed to predict the effect of sequence changes on RNA splicing suggest that this variant may disrupt the consensus splice site. For these reasons, this variant has been classified as Pathogenic. -
This variant causes a T>C nucleotide substitution at the +2 position of intron 3 of the LDLR gene. Splice site prediction tools suggest that this variant may have a significant impact on RNA splicing. Although this prediction has not been confirmed in published RNA studies, this variant is expected to result in an absent or disrupted protein product. Other variants affecting the same splice donor site have been shown to cause skipping of exon 3, resulting in an in-frame deletion of part of the ligand binding domain of the LDLR protein (PMID: 27821657). Functional studies in a homozygous patient fibroblast culture showed 10% LDLR activity compared to wild type cells (PMID: 19026292). This variant has been reported in numerous individuals affected with familial hypercholesterolemia (PMID: 7616128, 11462246, 14974088, 16542394, 19026292, 21475731, 28008010, 37119068). This variant has not been identified in the general population by the Genome Aggregation Database (gnomAD). Loss of LDLR function is a known mechanism of disease. Based on available evidence, this variant is classified as Pathogenic. This variant creates an alternative donor site beginning with a GC dinucleotide. Some GC donor sites have been shown to generate variable levels of wild-type transcript (PMID: 31131953). Hence, this variant could be less penetrant than a conventional splice donor site loss variant. -
Variant summary: LDLR c.313+2T>C is located in a canonical splice-site and is predicted to affect mRNA splicing resulting in a significantly altered protein due to either exon skipping, shortening, or inclusion of intronic material. Several computational tools predict a significant impact on normal splicing: Three predict the variant abolishes a 5 splicing donor site. However, these predictions have yet to be confirmed by functional studies. The variant was absent in 251436 control chromosomes (gnomAD). c.313+2T>C has been reported in the literature in multiple individuals affected with Familial Hypercholesterolemia (FH), including at least one homozygote (Brusgaard_2006, Kolansky_2008, Kusters_2011). In addition, Kusters_2011 reported this variant was one of most prevalent FH gene mutations and 6.4% of total FH subjects. These data indicate that the variant is very likely to be associated with disease. Kolansky_2008 showed this variant results in 10% LDLR activity of WT in skin fibroblast culture from one homozygous patient. 15 ClinVar submitters (evaluation after 2014) cite this variant as pathogenic (n=13), likely pathogenic (n=1) and benign (n=1), including one expert panel (ClinGen) classified as pathogenic. Based on the evidence outlined above, the variant was classified as pathogenic. -
Homozygous familial hypercholesterolemia Pathogenic:1
The c.313+2T>C variant in LDLR has been reported in 10 individuals with familial hypercholesterolemia (Lombardi 1995, Nauck 2001, Amsellem 2002, Graham 2005, Br usgaard 2006, Braenne 2015). This variant reportedly did not segregate with elev ated LDL cholesterol levels in 2 relatives from 1 family although the authors li st a second variant in both affected family members (Braenne 2015). This variant has also been reported by other clinical laboratories in ClinVar (Variation ID 189296) and was absent from large population studies. The c.313+2T>C variant occ urs in the invariant region (+/- 1,2) of the splice consensus sequence and is pr edicted to cause altered splicing leading to an abnormal or absent protein. Hete rozygous loss of LDLR function is an established disease mechanism in familial h ypercholesterolemia. Of note, 2 other prevalent, pathogenic variants have been reported in association with FH at this splice site (c.313+1G>A and c.313+1G>C) in our laboratory, supporting pathogenicity of the c.313+2T>C variant. In summar y, this variant meets criteria to be classified as pathogenic for familial hyper cholesterolemia in an autosomal dominant manner based upon the predicted impact to the protein, presence in multiple affected individuals and absence from the g eneral population. -
Cardiovascular phenotype Pathogenic:1
The c.313+2T>C intronic pathogenic mutation results from a T to C substitution two nucleotides after coding exon 3 in the LDLR gene. This alteration has been reported in association with familial hypercholesterolemia (Lombardi P, J. Lipid Res. 1995 Apr; 36(4):860-7; Dedoussis GV, Hum. Mutat. 2004 Mar; 23(3):285-6; Graham CA, Atherosclerosis 2005 Oct; 182(2):331-40; Huijgen R, Eur. Heart J. 2012 Sep; 33(18):2325-30). This alteration was also reported in a homozygous state in an Indian female with a total cholesterol level of 658 mg/dl (Kolansky DM, Am. J. Cardiol. 2008 Dec; 102(11):1438-43). In addition to the clinical data presented in the literature, alterations that disrupt the canonical splice site are expected to cause aberrant splicing, resulting in an abnormal protein or a transcript that is subject to nonsense-mediated mRNA decay. As such, this alteration is classified as a disease-causing mutation. -
Computational scores
Source:
Splicing
Find out detailed SpliceAI scores and Pangolin per-transcript scores at