A High-Throughput NMR Method for Lipoprotein-X Quantification.

Lipoprotein X (LP-X) is an abnormal cholesterol-rich lipoprotein particle that accumulates in patients with cholestatic liver disease and familial lecithin-cholesterol acyltransferase deficiency (FLD). Because there are no high-throughput diagnostic tests for its detection, a proton nuclear magnetic resonance (NMR) spectroscopy-based method was developed for use on a clinical NMR analyzer commonly used for the quantification of lipoproteins and other cardiovascular biomarkers. The LP-X assay was linear from 89 to 1615 mg/dL (cholesterol units) and had a functional sensitivity of 44 mg/dL.

Cell-free, high-density lipoprotein-specific phospholipid efflux assay predicts incident cardiovascular disease.

BACKGROUNDCellular cholesterol efflux capacity (CEC) is a better predictor of cardiovascular disease (CVD) events than HDL-cholesterol (HDL-C) but is not suitable as a routine clinical assay.METHODSWe developed an HDL-specific phospholipid efflux (HDL-SPE) assay to assess HDL functionality based on whole plasma HDL apolipoprotein-mediated solubilization of fluorescent phosphatidylethanolamine from artificial lipid donor particles. We first assessed the association of HDL-SPE with prevalent coronary artery disease (CAD): study I included NIH severe-CAD (n = 50) and non-CAD (n = 50) participants, who were frequency matched for sex, BMI, type 2 diabetes mellitus, and smoking; study II included Japanese CAD (n = 70) and non-CAD (n = 154) participants.

CD49d Expression Identifies a Biologically Distinct Subtype of Chronic Lymphocytic Leukemia with Inferior Progression-Free Survival on BTK Inhibitor Therapy.

Purpose: To determine the role of CD49d for response to Bruton's tyrosine kinase inhibitors (BTKi) in patients with chronic lymphocytic leukemia (CLL).

Patients And Methods: In patients treated with acalabrutinib (n = 48), CD49d expression, VLA-4 integrin activation, and tumor transcriptomes of CLL cells were assessed. Clinical responses to BTKis were investigated in acalabrutinib- (n = 48; NCT02337829) and ibrutinib-treated (n = 73; NCT01500733) patients.

A Simple Fluorescent Cholesterol Labeling Method to Cryoprotect and Detect Plasma Lipoprotein-X.

Lipoprotein-X (LpX) are abnormal nephrotoxic lipoprotein particles enriched in free cholesterol and phospholipids. LpX with distinctive lipid compositions are formed in patients afflicted with either familial LCAT deficiency (FLD) or biliary cholestasis. LpX is difficult to detect by standard lipid stains due to the absence of a neutral lipid core and because it is unstable upon storage, particularly when frozen.

Transgelin: a new gene involved in LDL endocytosis identified by a genome-wide CRISPR-Cas9 screen.

A significant proportion of patients with elevated LDL and a clinical presentation of familial hypercholesterolemia do not carry known genetic mutations associated with hypercholesterolemia, such as defects in the LDL receptor. To identify new genes involved in the cellular uptake of LDL, we developed a novel whole-genome clustered regularly interspaced short palindromic repeat-Cas9 KO screen in HepG2 cells. We identified transgelin (TAGLN), an actin-binding protein, as a potentially new gene involved in LDL endocytosis.

A novel loop-mediated isothermal amplification-based genotyping method and its application for identifying proprotein convertase subtilisin/kexin type 9 variants in familial hypercholesterolemia.

Background: Proprotein convertase subtilisin/kexin type 9 (PCSK9) plays a key role in regulating low-density lipoprotein levels in plasma. While PCSK9 variants are causatively associated with familial hypercholesterolemia (FH), additional genotyping methods for FH targeting PCSK9 variants are required in a clinical setting. Loop-mediated isothermal amplification (LAMP) is a unique amplification method that amplifies a target gene under isothermal conditions (60-65 °C).

COVID-19-Associated dyslipidemia: Implications for mechanism of impaired resolution and novel therapeutic approaches.

The current coronavirus disease 2019 (COVID-19) pandemic presents a global challenge for managing acutely ill patients and complications from viral infection. Systemic inflammation accompanied by a "cytokine storm," hemostasis alterations and severe vasculitis have all been reported to occur with COVID-19, and emerging evidence suggests that dysregulation of lipid transport may contribute to some of these complications. Here, we aim to summarize the current understanding of the potential mechanisms related to COVID-19 dyslipidemia and propose possible adjunctive type therapeutic approaches that modulate lipids and lipoproteins.

Incorporation of α-methylated amino acids into Apolipoprotein A-I mimetic peptides improves their helicity and cholesterol efflux potential.

Apolipoprotein A-I (ApoA-I) mimetic peptides are potential therapeutic agents for promoting the efflux of excess cellular cholesterol, which is dependent upon the presence of an amphipathic helix. Since α-methylated Ala enhances peptide helicity, we hypothesized that incorporating other types of α-methylated amino acids into ApoA-I mimetic peptides may also increase their helicity and cholesterol efflux potential. The last helix of apoA-I, peptide 'A' (VLESFKVSFLSALEEYTKKLNT), was used to design peptides containing a single type of α-methylated amino acid substitution (Ala/A, Glu/D, Lys/K, Leu/L), as well as a peptide containing both α-methylated Lys and Leu (6).

A dual apolipoprotein C-II mimetic-apolipoprotein C-III antagonist peptide lowers plasma triglycerides.

Recent genetic studies have established that hypertriglyceridemia (HTG) is causally related to cardiovascular disease, making it an active area for drug development. We describe a strategy for lowering triglycerides (TGs) with an apolipoprotein C-II (apoC-II) mimetic peptide called D6PV that activates lipoprotein lipase (LPL), the main plasma TG-hydrolyzing enzyme, and antagonizes the TG-raising effect of apoC-III. The design of D6PV was motivated by a combination of all-atom molecular dynamics simulation of apoC-II on the Anton 2 supercomputer, structural prediction programs, and biophysical techniques.

LCAT protects against Lipoprotein-X formation in a murine model of drug-induced intrahepatic cholestasis.

Familial lecithin:cholesterol acyltransferase (LCAT) deficiency (FLD) is a rare genetic disease characterized by low HDL-C levels, low plasma cholesterol esterification, and the formation of Lipoprotein-X (Lp-X), an abnormal cholesterol-rich lipoprotein particle. LCAT deficiency causes corneal opacities, normochromic normocytic anemia, and progressive renal disease due to Lp-X deposition in the glomeruli. Recombinant LCAT is being investigated as a potential therapy for this disorder.

Interleukin 10 promotes macrophage uptake of HDL and LDL by stimulating fluid-phase endocytosis.

Objective: Highly elevated plasma levels of interleukin-10 (IL-10) are causally associated with "Disappearing HDL Syndrome" and low plasma LDL-cholesterol, but the underlying mechanism is poorly understood. Fluid-phase endocytosis, a process highly dependent on actin dynamics, enables cells to internalize relatively high amounts of extracellular fluids and solutes. We sought to investigate whether IL-10 induces lipoprotein uptake by fluid-phase endocytosis in macrophages.

Plasma lipoprotein-X quantification on filipin-stained gels: monitoring recombinant LCAT treatment ex vivo.

Familial LCAT deficiency (FLD) patients accumulate lipoprotein-X (LP-X), an abnormal nephrotoxic lipoprotein enriched in free cholesterol (FC). The low neutral lipid content of LP-X limits the ability to detect it after separation by lipoprotein electrophoresis and staining with Sudan Black or other neutral lipid stains. A sensitive and accurate method for quantitating LP-X would be useful to examine the relationship between plasma LP-X and renal disease progression in FLD patients and could also serve as a biomarker for monitoring recombinant human LCAT (rhLCAT) therapy.

Molecular basis for activation of lecithin:cholesterol acyltransferase by a compound that increases HDL cholesterol.

Lecithin:cholesterol acyltransferase (LCAT) and LCAT-activating compounds are being investigated as treatments for coronary heart disease (CHD) and familial LCAT deficiency (FLD). Herein we report the crystal structure of human LCAT in complex with a potent piperidinylpyrazolopyridine activator and an acyl intermediate-like inhibitor, revealing LCAT in an active conformation. Unlike other LCAT activators, the piperidinylpyrazolopyridine activator binds exclusively to the membrane-binding domain (MBD).

Structural properties of apolipoprotein A-I mimetic peptides that promote ABCA1-dependent cholesterol efflux.

Peptides mimicking the major protein of highdensity lipoprotein (HDL), apolipoprotein A-I (apoA-I), are promising therapeutics for cardiovascular diseases. Similar to apoA-I, their atheroprotective property is attributed to their ability to form discoidal HDL-like particles by extracting cellular cholesterol and phospholipids from lipid microdomains created by the ABCA1 transporter in a process called cholesterol efflux. The structural features of peptides that enable cholesterol efflux are not well understood.

Apolipoprotein C-II: New findings related to genetics, biochemistry, and role in triglyceride metabolism.

Apolipoprotein C-II (apoC-II) is a small exchangeable apolipoprotein found on triglyceride-rich lipoproteins (TRL), such as chylomicrons (CM) and very low-density lipoproteins (VLDL), and on high-density lipoproteins (HDL), particularly during fasting. ApoC-II plays a critical role in TRL metabolism by acting as a cofactor of lipoprotein lipase (LPL), the main enzyme that hydrolyses plasma triglycerides (TG) on TRL. Here, we present an overview of the role of apoC-II in TG metabolism, emphasizing recent novel findings regarding its transcriptional regulation and biochemistry.

Development of a novel fluorescent activity assay for lecithin:cholesterol acyltransferase.

Background Lecithin:cholesterol acyltransferase (LCAT) is a plasma enzyme that esterifies cholesterol. Recombinant human LCAT (rhLCAT) is now being developed as an enzyme replacement therapy for familial LCAT deficiency and as a possible treatment for acute coronary syndrome. The current 'gold standard' assay for LCAT activity involves the use of radioisotopes, thus making it difficult for routine clinical use.

Lecithin:Cholesterol Acyltransferase Activation by Sulfhydryl-Reactive Small Molecules: Role of Cysteine-31.

Lecithin:cholesterol acyltransferase (LCAT) catalyzes plasma cholesteryl ester formation and is defective in familial lecithin:cholesterol acyltransferase deficiency (FLD), an autosomal recessive disorder characterized by low high-density lipoprotein, anemia, and renal disease. This study aimed to investigate the mechanism by which compound A [3-(5-(ethylthio)-1,3,4-thiadiazol-2-ylthio)pyrazine-2-carbonitrile], a small heterocyclic amine, activates LCAT. The effect of compound A on LCAT was tested in human plasma and with recombinant LCAT.

A Novel APOC2 Missense Mutation Causing Apolipoprotein C-II Deficiency With Severe Triglyceridemia and Pancreatitis.

Context: Familial chylomicronemia syndrome (FCS) is a rare heritable disorder associated with severe hypertriglyceridemia and recurrent pancreatitis. Lipoprotein lipase deficiency and apolipoprotein C-II deficiency are two well-characterized autosomal recessive causes of FCS, and three other genes have been described to cause FCS. Because therapeutic approaches can vary according to the underlying etiology, it is important to establish the molecular etiology of FCS.

Dietary α-cyclodextrin reduces atherosclerosis and modifies gut flora in apolipoprotein E-deficient mice.

Scope: α-Cyclodextrin (α-CD), a cyclic polymer of glucose, has been shown to lower plasma cholesterol in animals and humans; however, its effect on atherosclerosis has not been previously described.

Methods And Results: apoE-knockout mice were fed either low-fat diet (LFD; 5.2% fat, w/w), or Western high fat diet (21.

The Changing Face of HDL and the Best Way to Measure It.

Background: HDL cholesterol (HDL-C) is a commonly used lipid biomarker for assessing cardiovascular health. While a central focus has been placed on the role of HDL in the reverse cholesterol transport (RCT) process, our appreciation for the other cardioprotective properties of HDL continues to expand with further investigation into the structure and function of HDL and its specific subfractions. The development of novel assays is empowering the research community to assess different aspects of HDL function, which at some point may evolve into new diagnostic tests.

High-density lipoprotein reduces inflammation from cholesterol crystals by inhibiting inflammasome activation.

Interleukin-1β (IL-1β), a potent pro-inflammatory cytokine, has been implicated in many diseases, including atherosclerosis. Activation of IL-1β is controlled by a multi-protein complex, the inflammasome. The exact initiating event in atherosclerosis is unknown, but recent work has demonstrated that cholesterol crystals (CC) may promote atherosclerosis development by activation of the inflammasome.

Familial lecithin:cholesterol acyltransferase deficiency: First-in-human treatment with enzyme replacement.

Background: Humans with familial lecithin:cholesterol acyltransferase (LCAT) deficiency (FLD) have extremely low or undetectable high-density lipoprotein cholesterol (HDL-C) levels and by early adulthood develop many manifestations of the disorder, including corneal opacities, anemia, and renal disease.

Objective: To determine if infusions of recombinant human LCAT (rhLCAT) could reverse the anemia, halt progression of renal disease, and normalize HDL in FLD.

Methods: rhLCAT (ACP-501) was infused intravenously over 1 hour on 3 occasions in a dose optimization phase (0.