Low Density Lipoprotein Receptor Activity Research Articles

Evinacumab for Homozygous Familial Hypercholesterolemia

BackgroundHomozygous familial hypercholesterolemia is characterized by premature cardiovascular disease caused by markedly elevated levels of low-density lipoprotein (LDL) cholesterol. This disorder is associated with genetic variants that result in virtually absent (null–null) or impaired (non-null) LDL-receptor activity. Loss-of-function variants in the gene encoding angiopoietin-like 3 (ANGPTL3) are associated with hypolipidemia and protection against atherosclerotic cardiovascular disease. Evinacumab, a monoclonal antibody against ANGPTL3, has shown potential benefit in patients with homozygous familial hypercholesterolemia.MethodsIn this double-blind, placebo-controlled, phase 3 trial, we randomly assigned in a 2:1 ratio 65 patients with homozygous familial hypercholesterolemia who were receiving stable lipid-lowering therapy to receive an intravenous infusion of evinacumab (at a dose of 15 mg per kilogram of body weight) every 4 weeks or placebo. The primary outcome was the percent change from baseline in the LDL cholesterol level at week 24.ResultsThe mean baseline LDL cholesterol level in the two groups was 255.1 mg per deciliter, despite the receipt of maximum doses of background lipid-lowering therapy. At week 24, patients in the evinacumab group had a relative reduction from baseline in the LDL cholesterol level of 47.1%, as compared with an increase of 1.9% in the placebo group, for a between-group least-squares mean difference of –49.0 percentage points (95% confidence interval [CI], –65.0 to –33.1; P<0.001); the between-group least-squares mean absolute difference in the LDL cholesterol level was –132.1 mg per deciliter (95% CI, –175.3 to –88.9; P<0.001). The LDL cholesterol level was lower in the evinacumab group than in the placebo group in patients with null–null variants (–43.4% vs. +16.2%) and in those with non-null variants (–49.1% vs. –3.8%). Adverse events were similar in the two groups.ConclusionsIn patients with homozygous familial hypercholesterolemia receiving maximum doses of lipid-lowering therapy, the reduction from baseline in the LDL cholesterol level in the evinacumab group, as compared with the small increase in the placebo group, resulted in a between-group difference of 49.0 percentage points at 24 weeks. (Funded by Regeneron Pharmaceuticals; ELIPSE HoFH ClinicalTrials.gov number, NCT03399786.)

Bempedoic Acid: A Novel LDL Cholesterol-Lowering Agent.

Hypercholesterolemia is commonly associated with diabetes. In combination with diabetes, it puts patients at an increased risk for premature coronary heart disease and atherosclerosis (1). Lipoproteins are developed in one of two ways, via the exogenous or endogenous pathways. LDL cholesterol is a product of VLDL metabolism via the endogenous pathway. Levels of LDL in the plasma are determined by the rate of LDL production and clearance. The LDL production rate from VLDL depends on LDL receptor activity. High receptor activity leads to a decrease in LDL production. Alternatively, low LDL receptor activity leads to an increase in LDL production. These levels of receptors in the liver are regulated by the cholesterol content of the hepatocytes. If the content is low, LDL receptor activity increases to allow for the increased uptake of cholesterol. If the content is higher, LDL receptor activity is decreased, and the LDL uptake is diminished (2). Many medications are available today to manage hypercholesterolemia. The most commonly used are statins, fibrates, bile acid sequestrants, niacin, and dietary cholesterol uptake inhibitors. Less commonly used therapies for cholesterol management include PCSK9 (proprotein convertase subtilisin/kexin type 9) inhibitors, MTP (microsomal triglyceride transfer protein) inhibitors, CETP (cholesteryl ester transfer protein) inhibitors, and apolipoprotein B-100 antisense oligonucleotides. None of these therapies have the same mechanism of action as the novel therapy bempedoic acid (Esperion Therapeutics). Bempedoic acid has been shown to significantly lower LDL cholesterol levels when added to maximally tolerated statin therapies compared with ???

The Efficacy and Safety of Evinacumab in Homozygous Familial Hypercholesterolemia Patients with Little to No Low-Density Lipoprotein Receptor Activity

Robert Rosenson has received honoraria from Amgen, Kowa, Pfizer, and Regeneron; consultancy/advisory board fees from Amgen, C5, Corvidia, CVS Caremark, Medicines Company, and Regeneron; and research funding from Akcea, Amgen, AstraZeneca, Medicines Compan.

Homozygous familial hypercholesterolemia: what treatments are on the horizon?

Homozygous familial hypercholesterolemia (HoFH) is a rare disorder associated with early atherosclerotic disease due to impairment of the LDL receptor (LDLR) pathway. Because of their molecular defect, current treatment options have limited success in bringing HoFH patient to LDL-C target and morbidity and mortality remain high. We review current and upcoming therapies directed at HoFH, including gene therapy. Recent real-world studies have confirmed the strength in lomitapide as a treatment adjunct to statins and other lipid-lowering therapies in HoFH patients. The approval of proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitor monoclonal antibodies has also been a welcome addition to the treatment armamentarium offering an additional average reduction in LDL-C levels of 24% when added to background lipid-lowering therapies in this population. Although achieving adequate LDL-C levels in this population is difficult, there are several therapies on the horizon that may help more patients reach goal. Evinacumab, a monoclonal antibody against ANGPTL3, has been shown to substantially reduce LDL-C of an average of 49%, independently of residual LDLR activity. RNA interference targeting PCSK9 and ANGPTL3 shows promise in clinical trials. Adeno-associated virus-mediated gene transfer and gene editing techniques are in early clinical and preclinical development. LDL-C lowering in HoFH patients remains very challenging. However, novel treatment options are emerging. Upcoming therapies directed at PCSK9 and ANPTL3 may offer additional LDL-C reduction, to help patients achieve adequate LDL-C levels. Gene therapy and gene editing techniques, if proven effective, may offer a unique opportunity to treat patients with a one-time treatment.

The LDL-Receptor and its Molecular Properties: From Theory to Novel Biochemical and Pharmacological Approaches in Reducing LDL-cholesterol.

The Low-Density Lipoprotein (LDL) Receptor (LDL-R) is a transmembrane protein playing a crucial role in effective lipid homeostasis. Various therapeutic agents have been used in the management of dyslipidemias, however, the outcome of therapeutic target is debated. The aim of this review is to summarize and fully understand the current concept regarding LDL-R and its molecular properties, metabolic pathway, factors affecting LDL-R activity and all available pharmacological interventions. Additionally, non-lipid related properties of LDL-R are also referred. Literature from the PubMed database was extracted to identify papers between 1984 to 2017 regarding LDL-R and therapeutic agents on dyslipidemia management. We analyzed basic data regarding agents associated with LDL-R (Sterol Regulating Element-Binding Proteins - SREBPs, Protein ARH, IDOL, Thyroid Hormones, Haematologic Disorders, Protein convertase subtilisin kexintype 9 - PCSK-9, ApoC-III) as well as non-lipid related properties of LDL-R, while all relevant (common and novel) pharmacological interventions (statins, fibrates, cholesterol absorption inhibitors, bile acid sequestrants and PCSK- 9) are also referred. LDL-R and its molecular properties are involved in lipid homeostasis, so potentially sets the therapeutic goals in cardiovascular patients, which is usually debated. Further research is needed in order to fully understand its properties, as well as to find the potential pharmacological interventions that could be beneficial in cholesterol homeostasis and various morbidities in order to reach the most appropriate therapeutic goal.

Oxysterol-binding protein-related protein 1 variants have opposing cholesterol transport activities from the endolysosomes.

OSBPL1 encodes the full-length oxysterol-binding protein-related protein ORP1L, which transports LDL-derived cholesterol at membrane contacts between the late endosomes/lysosomes (LEL) and the endoplasmic reticulum (ER). OSBPL1 also encodes the truncated variant ORP1S that contains only the C-terminal lipid binding domain. HeLa cells in which both variants were knocked out (ORP1-null) were used to determine the functional relationship between ORP1L and ORP1S with respect to cellular cholesterol localization and regulation. ORP1-null cells accumulated cholesterol in LEL and had reduced plasma membrane (PM) cholesterol. PM cholesterol was restored by expression of wild-type ORP1S or a phosphatidylinositol phosphate-binding mutant but not by a sterol-binding mutant. Expression of ORP2, another truncated variant, also restored PM cholesterol in ORP1-null cells. Consistent with a LEL-to-PM cholesterol transport activity, a small fraction of ORP1S was detected on the PM. As a consequence of reduced delivery of cholesterol to the PM in ORP1-null cells, cholesterol was diverted to the ER resulting in normalization of de novo cholesterol synthesis. The deficiency in PM cholesterol also reduced ABCA1-dependent cholesterol efflux and LDL receptor activity in ORP1-null cells. We conclude that ORP1S, which lacks discrete membrane-targeting motifs, transports cholesterol from LEL to the PM.

Lomitapide treatment in a female with homozygous familial hypercholesterolaemia: a case report.

BackgroundHomozygous familial hypercholesterolaemia (FH) is an autosomal-dominant inherited disease presenting with highly elevated low-density lipoprotein cholesterol (LDL-C) levels. Untreated, the patient can develop atherosclerosis and cardiovascular disease already in adolescence. Treatment with statins and ezetimibe is usually not sufficient and LDL apheresis is often required. Lomitapide, an inhibitor of the microsomal triglyceride transfer protein, reduces LDL-C and triglyceride levels and can be used alone or in combination with other therapies in homozygous FH. However, experience with this agent is still limited.Case summaryWe present a young female who was diagnosed with homozygous FH at 6 years of age. She shows a complete lack of normal LDL receptor activity and no cholesterol-lowering effect from statins. The patient was treated with LDL apheresis from 7 years of age. When LDL apheresis treatment extended to twice a week, she began to experience adverse effects, including catheter-related complications, infections, and hospital admissions. When lomitapide treatment was initiated, the frequency of apheresis reduced, the LDL-C levels improved and she has not had any further hospital admissions since. Initially, she suffered from gastrointestinal disturbances. However, after 3 years of treatment with lomitapide 20 mg/day, the patient has not experienced any adverse effects.DiscussionIn this female with homozygous FH adding lomitapide treatment to LDL apheresis has contributed to improved LDL-C levels, a reduction in LDL apheresis sessions and enhanced quality of life. No adverse effects have been reported. These findings suggest that lomitapide can be a drug of choice in patients with homozygous FH.

Preparation of a Functional Rat LDL Receptor Minigene

The majority of the low-density lipoprotein (LDL) receptors present in the body are expressed in the liver. Therefore, plasma LDL levels significantly correlate with changes in the activity of the hepatic LDL receptor. Based on this, there is a need to understand the regulatory mechanisms that control the hepatic expression of the low-density lipoprotein (LDL) receptor. Herein, we have prepared a functional rat LDL receptor minigene construct that can produce mRNA after splicing. Sequence analysis suggests that this construct has the potential to code for a truncated version of LDL receptor protein. This minigene could be used as a research tool to identify small molecules, natural products, and regulators of the LDL receptor gene that could be developed into LDL receptor-specific activators for therapeutic use.

GPR146 is a potential new therapeutic target for lipid lowering.

Deficiency in the orphan G protein-coupled receptor 146 (GPR146) decreases blood lipid levels and protects against atherosclerosis in mice independently of LDL receptor activity, according to a new study.

Cascade screening for familial hypercholesterolemia-identification of the C308Y mutation in multiple family members and relatives for the first time in mainland China

BackgroundFamilial hypercholesterolemia (FH), an autosomal dominant genetic disorder, is underdiagnosed and undertreated. The majority of FH cases are caused by low density lipoprotein receptor (LDL-R) gene mutations. The C308Y mutation in LDL-R results in approximately 70% loss of LDL-R activity, leading to the elevation of low density lipoprotein-cholesterol (LDL-C) and an increased risk of premature coronary heart disease (CHD). The aim of this study was to identify FH cases by cascade screening in family members and relatives of a 37-year old male with premature CHD and hypercholesterolemia.MethodsClinical exam, blood lipid profiling and genomic DNA sequencing of all exons of LDL-R were performed for the proband and his 14 family members and relatives. FH diagnosis was carried out using the Dutch Lipid Clinic Network (DLCN) criteria.ResultsLipid profiling showed that 9 individuals, including the proband, had hypercholesterolemia. All these 9 subjects had a G > A substitution at nucleotide 986 in exon 7 resulting in the C308Y mutation as determined by DNA sequencing, and all those carrying the mutation were diagnosed as having definite FH under the DLCN criteria. However, most (7/9) did not have suggestive clinical manifestations of CHD.ConclusionsThe C308Y mutation was discovered in multiple family members and relatives for the first time in mainland China. Cascade screening is key for the confirmatory diagnosis of FH. Our hypothesis that the C308Y is a common variant in the population of Southern China origin warrants further validation by screening for the C308Y mutation in a large population.

Functional analysis of six uncharacterised mutations in LDLR gene

Background and aimsFamilial hypercholesterolemia (FH) is a primary hyperlipemia. It is an autosomal dominant genetic disorder of lipoproteins metabolism mainly caused by mutations in the low density lipoprotein receptor gene (LDLR). We aimed to investigate the functional impact on the low density lipoprotein receptor (LDLR) activity of six uncharacterised variants located in the coding region of the LDLR gene, namely c.428G > T, c.640T > C, c.1708C > T, c.1736A > T, c.1981C > G and c.2114C > G (NM_000527.4) and to attempt to define their clinical status. MethodsFunctional studies were carried out using site-directed mutagenesis techniques and expression of LDLR protein in vitro. Results were correlated with clinical data and in silico analyses in order to assess the physiopathological role of these variants. ResultsThis work provides functional information about 6 uncharacterised mutations in LDLR. ConclusionsThe six variants studied here appeared to affect the LDLR function in vitro to different degrees, ranging from receptors with normal to slightly reduced activity to receptors exhibiting less than 10% of the wild-type activity. According to these studies and The American College of Medical Genetics and Genomics (ACMG) Standards and Guidelines, two variants could be classified as “Likely Benign” (p.(Ala705Gly) and p.(Leu570Phe)), three variants as “Pathogenic” (p.(Asp579Val), p.(Cys143Phe) and p.(Trp214Arg)) and one variant as “Likely Pathogenic” (p.(Pro661Ala)).

Structure-Function Relationships of LDL Receptor Missense Mutations Using Homology Modeling.

Mutations in the low-density lipoprotein receptor (LDLR), which cause familial hypercholesterolemia (FH), present a variable clinical FH phenotype. To date, over 1600 FH-causing mutations have been found worldwide. The aim of this study was to investigate the structure-function relationships of LDLR mutations by using homology modeling. Structural analysis of 36 missense mutations of known receptor activity (33 severe, 1 mild, and 2 non-pathogenic phenotypes) using sequence comparison and homology modeling was performed. Severe phenotypes had less than 2% to 32% of residual LDLR activity. Mild phenotypes had 76-92% of residual LDLR activity. Finally, non-pathogenic phenotypes had normal residual LDLR activity. Sequence comparisons showed that most of the severe phenotypes were located within the fully conserved residues of LDLR, while most of the mild and non-pathogenic phenotypes were located within the poorly conserved residues. Homology modeling demonstrated several phenomena for severe phenotypes: disruption of disulfide bond formation, disturbance of the calcium binding sites, and perturbation of LDLR hydrophobic conserved packing. In contrast, mild and non-pathogenic phenotypes did not disturb the critical region of LDLR. In addition, the root mean square deviation (RMSD) values of severe phenotype tended to be higher than the mild and non-pathogenic phenotypes, and the mean of solvent accessible surface area (ASA) of the residues in wild type structure for the severe phenotype was lower than mild and non-pathogenic phenotypes. These findings provide a better understanding in the structure-function relationships of LDLR mutations and may be useful in predicting FH severity based on future genotyping.

Hypolipidaemia among patients with PMM2-CDG is associated with low circulating PCSK9 levels: a case report followed by observational and experimental studies

BackgroundProprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitors are novel therapeutics for reducing low-density lipoprotein cholesterol (LDLc). While serious side-effects have not been observed in short-term clinical trials, there remain concerns...

Control of LDL Uptake in Human Cells by Targeting the LDLR Regulatory Long Non-coding RNA BM450697

Hypercholesterolemia is a condition that is characterized by very high levels of cholesterol in the blood and is a major correlating factor with heart disease. Indeed, high levels of the low-density lipoprotein (LDL) have been causally linked to the development of atherosclerotic cardiovascular disease (ASCVD). A method to specifically reduce cholesterol in the blood in a long-term, stable manner could prove therapeutically relevant. Cholesterol is removed from the blood by the LDL receptor (LDLR) in the liver. Others and we have discovered that a long non-coding RNA (lncRNA; BM450697) functions as an endogenous epigenetic regulator of LDLR and that the repression of this lncRNA by the action of small interfering RNAs (siRNAs) results in the activation of LDLR. We found here, through the interrogation of two siRNAs that can target this lncRNA, both in a transcriptional and post-transcriptional manner, that BM450697 functions as a local scaffold for modulating LDLR transcription. Moreover, we found that conjugation of α-N-acetylgalactosamine (GalNAc) with two lncRNA-directed siRNAs allows for direct liver cell targeting of this lncRNA and functional enhanced uptake of cholesterol. Collectively, these data suggest that targeting the BM450697 lncRNA regulator of LDLR may result in a more specific, long-term, targeted approach to regulating cholesterol in the blood.

The emerging concept of "individualized cholesterol-lowering therapy": A change in paradigm.

High LDL-cholesterol concentrations constitute a risk for atherosclerotic cardiovascular disease. By consensus, cholesterol-lowering therapy is initiated with a statin that reduces endogenous cholesterol synthesis, upregulates hepatic LDL receptor activity, increases LDL clearance and lowers LDL-cholesterol concentrations in the bloodstream. The efficacy of statin treatment is dose dependent and achieves a risk reduction of up to 50%. However, a substantial body of evidence suggests that a quarter of statin-treated patients do not respond adequately as a result of low endogenous cholesterol synthesis. In humans fractional cholesterol absorption varies from 20% to 80%. High cholesterol absorbers, which are characterized by a low-to-normal cholesterol synthesis, exhibit poor responsiveness to statin treatment. On the other hand, the cholesterol absorption inhibitor ezetimibe effectively reduces serum cholesterol levels in these patients. On this background, we suggest to "get personal" and individualize cholesterol-lowering therapies, according to the individual's status of cholesterol synthesis and absorption.

Inhibiteurs de PCSK9 : quelle place dans la prise en charge actuelle des dyslipidémies ?

PCSK9 protein is a key regulator of LDL receptor activity. Gain-of-function mutations in PCSK9 are one of the genetic causes of familial hypercholesterolemia. Conversely, loss-of-function mutations are associated with lower levels of LDL cholesterol and reduced coronary heart disease. Monoclonal antibodies targeting PCSK9 are highly efficacious in lowering LDL-C levels, with a good tolerability and safety profile. Two PCSK9 inhibitors, alirocumab and evolocumab, have demonstrated a cardiovascular benefit in addition to statin therapy in patients with established cardiovascular disease. A recent European consensus has defined the candidates for PCSK9 inhibitors, e.g., patients with established cardiovascular disease and patients with familial hypercholesterolemia in primary prevention, with substantially elevated LDL-C levels despite maximally tolerated statin with or without ezetimibe therapy.

Low density lipoprotein mimics insulin action on autophagy and glucose uptake in endothelial cells

Elevated plasma low density lipoprotein (LDL) is an established risk factor for cardiovascular disease. In addition to being able to cross the endothelial barrier to become accumulated in subendothelial space and thereby initiate atherosclerosis, LDL may exert a direct effect on vascular endothelial cells through activation of LDL receptor and its downstream signaling. Whether LDL can modulate the signaling for autophagy in endothelial cells is not clear. The present study firstly demonstrated that LDL can suppress endothelial autophagy through activation of the PI3K/Akt/mTOR signaling pathway and can promote glucose uptake by translocating glucose transporter 1 (GLUT1) from cytoplasm to cell membrane, actions similar to those of insulin. A co-immunoprecipitation assay found that LDL receptor (LDLR) and insulin receptor (IR) formed a complex in HUVECs. Knock down of the insulin receptor by small interfering RNA blocked the suppression of autophagy by LDL, as well as the signaling pathway involved. We conclude that LDL may mimic the action of insulin in endothelial cells, which might partly explain the increased incidence of diabetes in patients receiving some LDL-lowering therapy.

A new variant (c.1A>G) in LDLRAP1 causing autosomal recessive hypercholesterolemia: Characterization of the defect and response to PCSK9 inhibition

Background and aimsAutosomal recessive hypercholesterolemia (ARH) is a rare disorder caused by mutations in LDLRAP1, which impairs internalization of hepatic LDL receptor (LDLR). ARH patients respond relatively well to statins or the combination of statins and Ezetimibe, but scarce and variable data on treatment with PCSK9 inhibitors is available. We aimed to identify and characterize the defect in a hypercholesterolemic patient with premature cardiovascular disease and determine the response to lipid-lowering treatment. Methods and resultsGene sequencing revealed a homozygous c.1A > G:p.? variant in LDLRAP1. Primary lymphocytes were isolated from the ARH patient, one control and two LDLR-defective subjects, one LDLR:p.(Cys352Ser) heterozygote and one LDLR:p.(Asn825Lys) homozygote. The patient had undetectable full-length ARH protein by Western blotting, but expressed a lower-than-normal molecular weight peptide. LDLR activity was measured by flow cytometry, which showed that LDL binding and uptake were reduced in lymphocytes from the ARH patient as compared to control lymphocytes, but were slightly higher than in those from the LDLR:p.(Cys352Ser) heterozygote. Despite the analogous internalization defect predicted in ARH and homozygous LDLR:p.(Asn825Lys) lymphocytes, LDL uptake was higher in the former than in the latter. LDL-cholesterol levels were markedly reduced by the successive therapy with Atorvastatin and Atorvastatin plus Ezetimibe, and the addition of Evolocumab biweekly decreased LDL-cholesterol by a further 39%. ConclusionsThe LDLRAP1:c.1A > G variant is associated with the appearance of an N-terminal truncated ARH protein and to reduced, although still significant, LDLR activity in lymphocytes. Residual LDLR activity may be relevant for the substantial response of the patient to Evolocumab.

Triciribine increases LDLR expression and LDL uptake through stabilization of LDLR mRNA

Low-density lipoprotein receptor (LDLR) is a key regulator of the metabolism of plasma low-density lipoprotein cholesterol (LDL-C), the elevated levels of which are associated with an increased risk of cardiovascular disease. Therefore, enhancing LDLR expression represents a potent treatment strategy for hypercholesterolemia. Here, we report that in cultured human hepatoma cells, triciribine, a highly selective AKT inhibitor, increases the stability of LDLR mRNA, an event that translates into upregulation of cell-surface LDLR levels and induction of cellular LDL uptake. This effect of triciribine requires ERK activity and is partially dependent on the intervening sequence between the AU-rich elements ARE3 and ARE4 in LDLR 3′UTR. We also show that triciribine downregulates the expression of PCSK9 mRNA and blunts the secretion of its protein. Notably, triciribine was found to potentiate the effect of mevastatin on LDLR protein levels and activity. We also show that primary human hepatocytes respond to triciribine by increasing the expression of LDLR. Furthermore, a pilot experiment with mice revealed that a two-weeks treatment with triciribine significantly induced the hepatic expression of LDLR protein. These results identify triciribine as a novel LDLR-elevating agent and warrant further examination of its potential as a hypocholesterolemic drug either as monotherapy or in combination with statins.

Replacement of cysteine at position 46 in the first cysteine-rich repeat of the LDL receptor impairs apolipoprotein recognition.

Background and aimsPathogenic mutations in the Low Density Lipoprotein Receptor gene (LDLR) cause Familial Hypercholesterolemia (FH), one of the most common genetic disorders with a prevalence as high as 1 in 200 in some populations. FH is an autosomal dominant disorder of lipoprotein metabolism characterized by high blood cholesterol levels, deposits of cholesterol in peripheral tissues such as tendon xanthomas and accelerated atherosclerosis. To date, 2500 LDLR variants have been identified in the LDLR gene; however, only a minority of them have been experimentally characterized and proven to be pathogenic. Here we investigated the role of Cys46 located in the first repeat of the LDL receptor binding domain in recognition of apolipoproteins.MethodsActivity of the p.(Cys46Gly) LDLR variant was assessed by immunoblotting and flow cytometry in CHO-ldlA7 expressing the receptor variant. Affinity of p.(Cys46Gly) for LDL and VLDL was determined by solid-phase immunoassays and in silico analysis was used to predict mutation effects.Results and conclusionFunctional characterization of p.(Cys46Gly) LDLR variant showed impaired LDL and VLDL binding and uptake activity. Consistent with this, solid-phase immunoassays showed the p.(Cys46Gly) LDLR variant has decreased binding affinity for apolipoproteins. These results indicate the important role of Cys46 in LDL receptor activity and highlight the role of LR1 in LDLr activity modulation. This study reinforces the significance of in vitro functional characterization of LDL receptor activity in developing an accurate approach to FH genetic diagnosis. This is of particular importance because it enables clinicians to tailor personalized treatments for patients’ mutation profile.