ANGPTL3 Mutations Research Articles

ANGPTL3 Deficiency and Risk of Hepatic Steatosis.

ANGPTL3 (angiopoietin-like 3) is a therapeutic target for reducing plasma levels of triglycerides and low-density lipoprotein cholesterol. A recent trial with vupanorsen, an antisense oligonucleotide targeting hepatic production of ANGPTL3, reported a dose-dependent increase in hepatic fat. It is unclear whether this adverse effect is due to an on-target effect of inhibiting hepatic ANGPTL3. We recruited participants with ANGPTL3 deficiency related to ANGPTL3 loss-of-function (LoF) mutations, along with wild-type (WT) participants from 2 previously characterized cohorts located in Campodimele, Italy, and St. Louis, MO. Magnetic resonance spectroscopy and magnetic resonance proton density fat fraction were performed to measure hepatic fat fraction and the distribution of extrahepatic fat. To estimate the causal relationship between ANGPTL3 and hepatic fat, we generated a genetic instrument of plasma ANGPTL3 levels as a surrogate for hepatic protein synthesis and performed Mendelian randomization analyses with hepatic fat in the UK Biobank study. We recruited participants with complete (n=6) or partial (n=32) ANGPTL3 deficiency related to ANGPTL3 LoF mutations, as well as WT participants (n=92) without LoF mutations. Participants with ANGPTL3 deficiency exhibited significantly lower total cholesterol (complete deficiency, 78.5 mg/dL; partial deficiency, 172 mg/dL; WT, 188 mg/dL; P<0.05 for both deficiency groups compared with WT), along with plasma triglycerides (complete deficiency, 26 mg/dL; partial deficiency, 79 mg/dL; WT, 88 mg/dL; P<0.05 for both deficiency groups compared with WT) without any significant difference in hepatic fat (complete deficiency, 9.8%; partial deficiency, 10.1%; WT, 9.9%; P>0.05 for both deficiency groups compared with WT) or severity of hepatic steatosis as assessed by magnetic resonance imaging. In addition, ANGPTL3 deficiency did not alter the distribution of extrahepatic fat. Results from Mendelian randomization analyses in 36 703 participants from the UK Biobank demonstrated that genetically determined ANGPTL3 plasma protein levels were causally associated with low-density lipoprotein cholesterol (P=1.7×10-17) and triglycerides (P=3.2×10-18) but not with hepatic fat (P=0.22). ANGPTL3 deficiency related to LoF mutations in ANGPTL3, as well as genetically determined reduction of plasma ANGPTL3 levels, is not associated with hepatic steatosis. Therapeutic approaches to inhibit ANGPTL3 production in hepatocytes are not necessarily expected to result in the increased risk for hepatic steatosis that was observed with vupanorsen.

Evinacumab for treatment of familial hypercholesterolemia

Introduction Familial hypercholesterolemia (FH) is characterized by lifelong elevation of low-density lipoprotein cholesterol (LDL-C), early onset coronary atherosclerosis and premature death. FH is underdiagnosed and undertreated, but requires aggressive LDL-C-lowering to prevent complications. Current treatment strategies, lifestyle modification and numerous LDL-C-lowering medications, are often insufficient to achieve lipid goals in FH. Areas covered Angiopoietin-like 3 protein (ANGPTL3) is intricately involved in lipid metabolism. Loss-of-function mutations in ANGPTL3 are associated with panhypolipidemia and reduced coronary atherosclerosis. Evinacumab, a fully human monoclonal antibody, inhibits ANGPTL3 and reduces multiple lipoprotein fractions ~50%, including LDL-C. The use of evinacumab within the FH population is described, as well as its regulatory journey to an approved therapeutic. Expert opinion Evinacumab, with its capacity to lower multiple lipoprotein fractions, particularly LDL-C, independently of LDLR function has potential to revolutionize treatment for FH patients. Current FDA-approval is only for homozygous FH (HoFH), arguably the most impactful indication, but use in other lipid disorders is under investigation. Short-term tolerability of evinacumab is very good, with infrequent, mild, and transient adverse events, however, long-term safety data are needed. The high cost and requirement for intravenous administration may limit adoption of evinacumab, but dramatic LDL-C-lowering and need for new therapeutic options for HoFH will drive interest.

Antisense oligonucleotide–mediated inhibition of angiopoietin-like protein 3 increases reverse cholesterol transport in mice

Supported by an abundance of experimental and genetic evidence, angiopoietin-like protein 3 (ANGPTL3) has emerged as a promising therapeutic target for cardiovascular disease. ANGPTL3 is primarily produced by the liver and is a potent modulator of plasma lipids and lipoproteins. Experimental models and subjects with loss-of-function Angptl3 mutations typically present with lower levels of HDL-C than noncarriers. The effect of ANGPTL3 on HDL-C is typically attributed to its function as an inhibitor of the enzyme endothelial lipase. The ability to facilitate reverse cholesterol transport (RCT), the transport of cholesterol from peripheral tissues back to the liver, is a proposed antiatherogenic property of HDL. However, the effect of ANGPTL3 inhibition on RCT remains unclear. Here, we performed a series of dose-response and RCT studies using an Angptl3 antisense oligonucleotide (ASO) in mouse models with varying plasma lipid profiles ranging from moderately to severely hyperlipidemic. Angptl3 ASO-mediated reduction in HDL-C was limited to the model with moderate lipidemia, where the majority of plasma cholesterol was associated with HDL. Surprisingly, regardless of the effect on HDL-C, treatment with the Angptl3 ASO enhanced RCT in all models tested. The observations from the RCT assays were confirmed in HDL clearance studies, where mice treated with the Angptl3 ASO displayed increased plasma clearance and hepatic uptake of labeled HDL. The results from our studies suggest that inhibition of ANGPTL3 not only reduces levels of proatherogenic lipids but also improves HDL-mediated RCT.

The impact of ANGPTL3 deficiency on hepatic steatosis: Observations from carriers of loss-of-function mutations

Background and Aims: Loss-of-function (LOF) mutations in ANGPTL3 cause familial combined hypolipidemia (FHBL2) characterized by very low levels of all three major lipoprotein fractions (LDL-C, TGs and HDL-C). Targeting ANGPTL3 has emerged as a new therapeutic opportunity to lower two causal risk factors (LDL-C and TG) for coronary heart disease (CHD) with potentially favorable metabolic effects. It is unknown, however, if inhibiting ANGPTL3 will result in adverse consequences. The aim of this study was to evaluate if genetic loss of ANGPTL3 leads to hepatic fat accumulation.

ANGPTL3 deficiency alters the lipid profile and metabolism of cultured hepatocytes and human lipoproteins

Loss-of-function (LOF) mutations in ANGPTL3, an inhibitor of lipoprotein lipase (LPL), cause a drastic reduction of serum lipoproteins and protect against the development of atherosclerotic cardiovascular disease. Therefore, ANGPTL3 is a promising therapy target. We characterized the impacts of ANGPTL3 depletion on the immortalized human hepatocyte (IHH) transcriptome, lipidome and human plasma lipoprotein lipidome. The transcriptome of ANGPTL3 knock-down (KD) cells showed altered expression of several pathways related to lipid metabolism. Accordingly, ANGPTL3 depleted IHH displayed changes in cellular overall fatty acid (FA) composition and in the lipid species composition of several lipid classes, characterized by abundant n-6 and n-3 polyunsaturated FAs (PUFAs). This PUFA increase coincided with an elevation of lipid mediators, among which there were species relevant for resolution of inflammation, protection from lipotoxic and hypoxia-induced ER stress, hepatic steatosis and insulin resistance or for the recovery from cardiovascular events. Cholesterol esters were markedly reduced in ANGPTL3 KD IHH, coinciding with suppression of the SOAT1 mRNA and protein. ANGPTL3 LOF caused alterations in plasma lipoprotein FA and lipid species composition. All lipoprotein fractions of the ANGPTL3 LOF subjects displayed a marked drop of 18:2n-6, while several highly unsaturated triacylglycerol (TAG) species were enriched. The present work reveals distinct impacts of ANGPTL3 depletion on the hepatocellular lipidome, transcriptome and lipid mediators, as well as on the lipidome of lipoproteins isolated from plasma of ANGPTL3-deficient human subjects. It is important to consider these lipidomics and transcriptomics findings when targeting ANGPTL3 for therapy and translating it to the human context.

Statins Reduce Hepatic Angptl3 Expression And Secretion, Likely By Interfering With Lxr-Mediated Transcription

Background and Aims: Loss of function mutations in ANGPTL3 cause low plasma lipid concentrations. Statins supposedly suppress hepatic ANGPTL3 mRNA expression in healthy subjects (J. Intern. Med. 2011: 269: 333). How statins regulate Angptl3 and whether statins reduce plasma Angptl3 concentrations in patients with elevated LDL cholesterol levels, e.g., familial hypercholesterolemia (FH), is unknown.

Angiopoietin-Like 3 Protein Inhibition: A New Frontier in Lipid-Lowering Treatment.

Angiopoietin-like 3 protein (ANGPTL3) is an inhibitor of both lipoprotein lipase and endothelial lipase in humans. Population studies indicate a relationship between loss of function mutations in ANGPTL3 and favorable reductions in triglycerides and non- high-density lipoprotein cholesterol. In addition, loss of function mutations is associated with a reduced risk of coronary artery disease. Whereas ANGPTL3's role in human lipid metabolism has yet to be fully clarified, it is unlikely that ANGPTL3 impacts cholesterol uptake via the low-density lipoprotein-receptor, unlike the proprotein convertase subtilisin/kexin9 inhibitors. In contrast to other forms of lipid-lowering therapy, ANGPTL3 inhibition may improve insulin sensitivity. The promise of this new therapy, particularly its independence from the low-density lipoprotein-receptor, has prompted the creation of a monoclonal antibody inhibitor; evinacumab. Evinacumab has shown favorable lipid-lowering action in both human and mouse models. Efficacy trials are currently ongoing and will be completed in the near future. In addition, ANGPTL3 inhibition via an antisense oligonucleotide was performed in healthy human subjects, which resulted in a dose-dependent reduction in circulating ANGPTL3 levels and an antiatherogenic lipid profile. When tested in mouse models, administration of the antisense oligonucleotide caused a reduction in progression of atherosclerosis. Further investigation is required to evaluate the efficacy, safety and net benefit of clinical ANGPTL3 inhibition before it can be accepted into clinical practice.

ANGPTL3 Mutations in Unrelated Chinese Han Patients with Familial Hypercholesterolemia.

Familial hypercholesterolemia (FH) is a severe genetic hyperlipidemia characterized by increased levels of low-density lipoprotein cholesterol (LDL-C), leading to premature atherosclerosis. Angiopoietin-like protein (ANGPTL3) is a hepatocyte-specific protein that can be used to lower LDL in FH. However, it was unknown whether ANGPTL3 variants are present in FH patients. This study was performed to identify ANGPTL3 variants in unrelated Chinese Han patients with FH. We screened 80 patients with FH (total cholesterol >7.8mmol/L, LDL-cholesterol >4.9mmol/L) and 77 controls using targeted next-generation sequencing (NGS) of six FH candidate genes (LDLR, ApoB100, PCSK9, ABCG5, ABCG8, and ANGPTL3). Candidate pathogenic variants identified by NGS were validated by Sanger sequencing. Mutant and wild-type plasmids containing the variant sequence were constructed and verified by Sanger sequencing. The gene expression profile was analyzed by an expression profile chip in transfected HepG2 cells using quantitative real-time (qRT)-PCR. We identified 41 variants in 28 FH patients, including two ANGPTL3 mutations: one exonic (c.A956G: p.K319R) and one in the untranslated region (c.*249G>A). Gene ontology analyses found that the cholesterol metabolic process and ANGPTL3 expression were significantly up-regulated in the ANGPTL3 K319R mutation group compared with the wild-type group. qRT-PCR findings were consistent with the expression profile analysis. Rare ANGPTL3 variants were identified in Chinese patients with FH, including ANGPTL3: p.(Lys319Arg) which affected the expression of ANGPTL3 and the cholesterol metabolic process as determined by bioinformatics analysis. Chinese Clinical Trial Registration (ChiCTR-ROC-17011027) http://www.chictr.org.cn/listbycreater.aspx.

Angiopoietin-like proteins - their role in lipoprotein metabolism and association with atherogenic dyslipidemia

Scientists have been trying to find the best parameter for laboratory assessment of the risk of cardiovascular diseases (CVD) for decades. Initially, the results of many studies indicated that the analysis of the lipid profile was sufficient to evaluate the risk of CVD. Further studies revealed that more precise laboratory prediction of cardiovascular risk requires quantification of atherogenic lipoproteins. Recently, angiopoietin-like proteins 3, 4 and 8 (ANGPTLs) have been described as important regulators of plasma lipoprotein metabolism and triglyceride homeostasis. Mutations in ANGPTL3 leading to loss of its function have been linked to decreased risk of CVD in humans. Among potential new targets for the management of dyslipidemia, ANGPTL3 may become a considerably promising one.

Angiopoietin-like 3 in lipoprotein metabolism.

Triglycerides and cholesterol circulate in the bloodstream as part of various lipoprotein particles. Three members of the angiopoietin-like (ANGPTL) protein family - ANGPTL3, ANGPTL4 and ANGPTL8 - have emerged as important regulators of plasma lipoprotein levels by inhibiting the enzyme lipoprotein lipase. Here, I review the role of ANGPTL3 in lipoprotein metabolism. In contrast to ANGPTL4 and ANGPTL8, ANGPTL3 is exclusively produced in the liver and can therefore be classified as a true hepatokine. ANGPTL3 cooperates with ANGPTL8 to inhibit lipoprotein lipase and is mostly active after feeding, whereas ANGPTL4 is mostly active after fasting. Inactivation of ANGPTL3 in mice reduces plasma triglyceride and free fatty acid levels and suppresses atherosclerosis. In humans, homozygous loss-of-function mutations in ANGPTL3 lead to low plasma levels of low-density lipoproteins, high-density lipoproteins and triglycerides, a condition referred to as familial combined hypolipidaemia. Heterozygous carriers of loss-of-function mutations in ANGPTL3 have a lower risk of coronary artery disease than non-carriers. At present, researchers are investigating antisense oligonucleotide and monoclonal antibody-based inactivation of ANGPTL3 in human clinical trials for the therapeutic management of dyslipidaemia and atherosclerosis. Thus, ANGPTL3 is an important liver-derived regulator of lipoprotein metabolism that holds considerable promise as a target for atherosclerosis.

Heart disease: Putative medicines that mimic mutations.

Molecules that block the activity or production of the protein ANGPTL3 have now been found to lower blood levels of lipoproteins and cholesterol in mice and healthy humans, mimicking the protective effects of genetic mutations in ANGPTL3.

Using human genetics to discover new therapeutic targets for plasma lipids.

Genetic variation arises through multiple different alleles that vary in frequency and severity of effect. Mutations that give rise to Mendelian disorders, such as the LDL receptor (LDLR) mutations that result in familial hypercholesterolaemia, are efficiently winnowed from the population by purifying selection and are almost inevitably rare. Conversely, alleles that are common in the population (such that homozygotes for the minor allele are present even in modest sample sizes) typically have very modest phenotypic effects. Mutations in the gene for proprotein convertase subtilisin/kexin type 9 (PCSK9) represent an unusual but informative exception in that they are relatively common but have large effects on phenotype. Loss-of-function mutations in PCSK9 occur in ~2.5% of African Americans and are associated with large reductions in coronary heart disease (CHD) risk. The development of agents to inhibit PCSK9 demonstrates the utility of translating genetics into clinical therapeutics. Attempts to identify genes responsible for hypercholesterolaemia have used traditional linkage analysis, which requires samples collected from multiple families with defects in the same gene, or genome-wide association, which requires thousands of samples from the population. More recently, whole-exome sequencing studies have revealed loss-of-function mutations in ANGPTL3 associated with pan-hypolipidemia, and in APOC3 that confer protection against CHD. The application of whole-exome sequencing to large populations or to carefully selected patients can streamline the discovery of causal genetic mutations.

Abstract 27: Therapeutic Targeting of Human Lipid Genes With in vivo CRISPR-Cas9 Genome Editing

Individuals with naturally occurring loss-of-function PCSK9, APOC3, and ANGPTL3 mutations experience reduced blood low-density lipoprotein cholesterol (LDL-C) levels, reduced blood triglyceride levels, and/or protection against cardiovascular disease. We recently established that genome editing using a Streptococcus pyogenes clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) system, delivered by adenovirus, can efficiently introduce loss-of-function mutations into the endogenous mouse Pcsk9 gene in vivo, resulting in near-complete knockout of Pcsk9 and a 35%-40% reduction in the blood cholesterol level. Thus, the in vivo genome-editing approach may have therapeutic potential for the prevention of cardiovascular disease in humans. To better evaluate the feasibility of the approach in humans, we are performing preclinical studies using a liver-humanized mouse model in which the endogenous mouse hepatocytes have been replaced with human-derived hepatocytes and in which the plasma lipoprotein profile better reflects human physiology, particularly with respect to LDL-C. One disadvantage of the S. pyogenes CRISPR-Cas9 system is that its Cas9 gene is too large to fit in adeno-associated virus (AAV) vectors, which offer significant therapeutic advantages over adenoviruses. We have adapted the smaller Staphylococcus aureus CRISPR-Cas9 system and demonstrated efficient genome editing of the human PCSK9, APOC3, and ANGPTL3 genes in vitro. Additionally, using a minimal liver-specific promoter and a novel human-hepatocyte-optimized AAV capsid protein, we are employing AAV vectors to knock out each of the three genes in both primary human hepatocytes and liver-humanized mice. This is allowing us to interrogate both the efficacy (lowering of LDL-C and/or triglycerides) and safety (degree of off-target mutagenesis) of genome editing in human hepatocytes in vitro and in vivo. These translational studies are providing critical information on the viability of an approach that could ultimately yield a one-shot, long-term therapy that permanently reduces a person’s blood LDL-C and triglyceride levels, thus serving as a “vaccination” against cardiovascular disease.

Abstract 13133: Silencing of Angiopoietin-like Protein 3 ( ANGPTL3 ) in IHH- Hepatocytes Results in Increased Insulin Sensitivity and Reduced Triglyceride-rich VLDL Secretion

AIMS: Homozygosity of loss-of-function mutations in ANGPTL3 -gene results in familial combined hypolipidemia (FHBL2,OMIM #605019) characterized by reduction of all major plasma lipoprotein classes VLDL, LDL, HDL and low circulating free fatty acids, glucose and insulin levels. Thus complete lack of ANGPTL3 in humans not only affects lipid metabolism but also whole-body insulin and glucose balance. Our aim was to investigate the function of ANGPTL3, a hepatic secretory protein, in promoting hypolipidemia and hepatic insulin sensitivity. METHODS: We used wild type and ANGPTL3-silenced human immortalized hepatocytes (IHH) to investigate the effect of ANGPTL3-silencing on hepatic VLDL secretion and glucose uptake. RESULTS: We demonstrate that insulin downregulates hepatic secretion of ANGPTL3 and triglyceride-enriched VLDL1-type particles in a dose dependent manner. Similar effect on VLDL secretion was demonstrated with a treatment of PPAR[[Unable to Display Character: &amp;#436;]] agonist rosiglitazone. We further show that ANGPTL3-silenced cells display a more pronounced shift from the secretion of TG-enriched VLDL1-type particles to a secretion of lipid poor VLDL2-type particles during insulin stimulation. Silencing of ANGPTL3 improved hepatic glucose uptake by 20-50 % depending on the glucose and insulin concentration, resulted in a trend towards increased AKT/PKB phosphorylation upon insulin stimulation and downregulated fasting induced transcription factor PGC1α and its downstream targets. CONCLUSION: Our results indicate a similar function of both insulin and rosiglitazone regarding regulation of ANGPTL3 and VLDL secretion in hepatocytes, and suggest that insulin and PPAR[[Unable to Display Character: &amp;#436;]] might mediate some of their functions via ANGPTL3. Our results give more insight into the liver specific role of ANGPTL3 and links silencing of ANGPTL3 with Insulin sensitivity. Since humans with elevated levels of ANGPTL3 display hyperlipidemia and insulin resistance it might be beneficial to target ANGPTL3 silencing in the liver, the major site of ANGPTL3 expression, to balance lipid and glucose homeostasis and lower risk for cardiovascular diseases.

Hypobetalipoproteinemia and abetalipoproteinemia.

Several mutations in the apoB, proprotein convertase subtilisin/kexin type 9 (PCSK9), and MTP genes result in low or absent levels of apoB and LDL-cholesterol in plasma, which cause familial hypobetalipoproteinemia and abetalipoproteinemia. Mutations in the ANGPTL3 gene cause familial combined hypolipidemia. Clinical manifestations range from none to severe, debilitating, and life-threatening disorders. This review summarizes recent genetic, metabolic, and clinical findings and presents an update on management strategies. Cases of cirrhosis and hepatocellular carcinoma have now been identified in heterozygous familial hypobetalipoproteinemia probably because of decreased triglyceride transport capacity from the liver. ANGPTL3 mutations cause low levels of LDL-cholesterol and low HDL-cholesterol in compound heterozygotes and homozygous individuals, decrease reverse cholesterol transport, and lower glucose levels. The effect on atherosclerosis is unknown; however, severe fatty liver has been identified. Loss-of-function mutations in PCSK9 cause familial hypobetalipoproteinemia, which appears to lower risk for coronary artery disease and has no adverse sequelae. Phase III clinical trials are now underway examining the effect of PCSK9 inhibitors on cardiovascular events in combination with statin drugs. Mutations causing low LDL-cholesterol and apoB have provided insight into lipid metabolism, disease associations, and the basis for drug development to lower LDL-cholesterol in disorders causing high levels of cholesterol. Early diagnosis and treatment are necessary to prevent adverse sequelae from familial hypobetalipoproteinemia and abetalipoproteinemia.

Recent developments in the genetics of LDL deficiency

Inherited diseases of lipoprotein metabolism may give rise to marked hypocholesterolemia with low or absent levels of LDL, depending on the gene involved and mode of inheritance of the condition, together with the severity of the mutation or mutations present. In this review, we discuss the recent developments in the genetics of LDL deficiency. Carriers of a single loss-of-function variant in ANGPTL3 have reduced LDL-cholesterol and triglyceride concentrations, whereas homozygotes have markedly reduced LDL-cholesterol, triglyceride and HDL-cholesterol concentrations, a recessive form of hypocholesterolemia designated as familial combined hypolipidemia. The identification of loss-of-function ANGPTL3 mutations as a cause of familial combined hypolipidemia suggests a new mechanism for the regulation of LDL metabolism in humans, thereby making ANGPTL3 an attractive protein to target for therapeutics.

Identification of a novel mutation in the ANGPTL3 gene in two families diagnosed of familial hypobetalipoproteinemia without APOB mutation

BackgroundFamilial hypobetalipoproteinemia (FHBL), characterized by extremely low levels of plasma apolipoprotein (apo) B and cholesterol associated with low-density lipoproteins (LDLc), is considered to be an autosomal co-dominant disorder of heterogeneous origin. The main genetic disorder associated with FHBL consists of mutations in the APOB gene, while other less frequent forms are associated with mutations in NPC1L1, PCSK9, a still unidentified gene in 3p21.1-22 and, more recently, in ANGPTL3. MethodsWe scanned for ANGPTL3 mutations in 4 unrelated Spanish families with FHBL criteria but negative for mutations in APOB. The entire coding region and intron–exon boundaries of the ANGPTL3 gene were amplified and sequenced. ResultsTwo probands were positive for the same frameshift mutation, a deletion of 5bp in codon 121 in ANGPTL3, which produces a truncated protein of 122 residues. This mutation in homozygosis was associated in both families with combined hypolipidemia, characterized by low plasma apoB, low total, LDL and HDL cholesterol and low triglycerides. ConclusionWe confirm the existence of a new phenotype of FHBL, denominated familial combined hypolipidemia, which consist of a biochemical phenotype of low LDLc, low apoB, low TG and, unlike APOB mutations, low HDL cholesterol, due to a loss-of-function mutation in ANGPTL3.

Familial hypobetalipoproteinemia in a hospital survey: genetics, metabolism and non-alcoholic fatty liver disease

Introduction. Familial hypobetalipoproteinemia (FHBL) is an autosomal dominant disease characterized by abnormally low levels of apolipoprotein-B (apoB) containing lipoproteins. FHBL is caused by APOB, PCSK9 or ANGPTL3 mutations or is associated with loci located in chromosomes 10 and 3p21. However, other genes should be involved. This study describes the kinetic parameters of the apoB containing lipoproteins and sequence abnormalities of the APOB and PCSK9 genes of FHBL patients identified in a large hospital based survey.Material and methods. Cases with primary or secondary causes of hypobetalipoproteinemia were identified. ApoB kinetics were measured in cases with primary forms in whom truncated forms of apoB were not present in VLDL (n = 4). A primed constant infusion of [13C] leucine was administered, VLDL and LDL apoB production and catabolic rates measured by a multicompartmental model and compared to normolipemic controls. In addition, these subjects had an abdominal ultrasound and direct sequencing was carried out for the PCSK9 and apoB genes.Results. Three individuals had normal apoB production with increased catabolic rate; the remaining had reduced synthetic and catabolic rates. Various polymorphisms, some of them previously unreported (*), in the PCSK9 gene (R46L, A53V, I474V, D480N*, E498K*) and in the apoB gene (N441D*, Y1395C, P2712L, D2285E*, I2286V, T3540S*, T3799M*) were found in the FHBL patients. We found hepatic ultrasound changes of hepatic steatosis in only one of the four probands. Conclusion. FHBL without truncated apoB is a heterogeneous disease from a metabolic and a genetic perspective. Hypobetalipoproteinemia is a risk factor but not an obligate cause of steatosis.

Exome Sequencing Links Gene Mutation in Angiopoietin-Like Protein 3 With Low-Density Lipoprotein Cholesterol

1. Musunuru K, Pirruccello JP, Do R, Peloso GM, Guiducci C, Sougnez C, Garimella KV, Fisher S, Abreu J, Barry AJ, Fennell T, Banks E, Ambrogio L, Cibulskis K, Kernytsky A, Gonzalez E, Rudzicz N, Engert JC, DePristo MA, Daly MJ, Cohen JC, Hobbs HH, Altshuler D, Schonfeld G, Gabriel SB, Yue P, Kathiresan S. Exome sequencing, ANGPTL3 mutations, and familial combined hypolipidemia. N Engl J Med. 2010;363(23):2220–2227. PMID: 20942659. ### Study Hypothesis Familial hypobetalipoproteinemia is an inherited disorder of lipid metabolism defined by extremely low levels of plasma apolipoprotein B (apoB) and low-density lipoprotein cholesterol (LDL-C). Some cases have been linked to the apoB gene, but in other families, efforts to identify the gene defects associated with this disorder have been unsuccessful. The authors hypothesized that next-generation exome sequencing (sequencing of all protein-coding regions of the genome) of select cases of familial hypobetalipoproteinemia not linked to apoB would allow for identification of the causative gene. ### How Was the Hypothesis Tested? A family of European descent, some of whose members had hypobetalipoproteinemia, was used for the study. Linkage analysis had previously ruled out apoB as the …

Exome Sequencing,ANGPTL3Mutations, and Familial Combined Hypolipidemia

We sequenced all protein-coding regions of the genome (the "exome") in two family members with combined hypolipidemia, marked by extremely low plasma levels of low-density lipoprotein (LDL) cholesterol, high-density lipoprotein (HDL) cholesterol, and triglycerides. These two participants were compound heterozygotes for two distinct nonsense mutations in ANGPTL3 (encoding the angiopoietin-like 3 protein). ANGPTL3 has been reported to inhibit lipoprotein lipase and endothelial lipase, thereby increasing plasma triglyceride and HDL cholesterol levels in rodents. Our finding of ANGPTL3 mutations highlights a role for the gene in LDL cholesterol metabolism in humans and shows the usefulness of exome sequencing for identification of novel genetic causes of inherited disorders. (Funded by the National Human Genome Research Institute and others.).