Molecular Dynamics Simulation and Experimental Studies of Gold Nanoparticle Templated HDL-like Nanoparticles for Cholesterol Metabolism Therapeutics

High-density lipoprotein (HDL) plays an important role in the transport and metabolism of cholesterol. Mimics of HDL are being explored as potentially powerful therapeutic agents for removing excess cholesterol from arterial plaques. Gold nanoparticles (AuNPs) functionalized with apolipoprotein A-I and with the lipids 1,2-dipalmitoyl-<i>sn</i>-glycero-3-phosphocholine and 1,2-dipalmitoyl-<i>sn</i>-glycero-3-phospho­ethanolamine-N-[3-(2-pyridyldithio)­propionate] have been demonstrated to be robust acceptors of cellular cholesterol. However, detailed structural information about this functionalized HDL AuNP is still lacking. In this study, we have used X-ray photoelectron spectroscopy and lecithin/cholesterol acyltransferase activation experiments together with coarse-grained and all-atom molecular dynamics simulations to model the structure and cholesterol uptake properties of the HDL AuNP construct. By simulating different apolipoprotein-loaded AuNPs, we find that lipids are oriented differently in regions with and without apoA-I. We also show that in this functionalized HDL AuNP, the distribution of cholesteryl ester maintains a reverse concentration gradient that is similar to the gradient found in native HDL

Thrombin-Targeted Liposomes Establish a Sustained Localized Anticlotting Barrier against Acute Thrombosis

The goal of the present work was to design and test an acute-use nanoparticle-based antithrombotic agent that exhibits sustained local inhibition of thrombin without requiring a systemic anticoagulant effect to function against acute arterial thrombosis. To demonstrate proof of concept, we functionalized the surface of liposomes with multiple copies of the direct thrombin inhibitor, d-phenylalanyl-l-prolyl-l-arginyl-chloromethyl ketone (PPACK), which exhibits high affinity for thrombin as a free agent but manifests too rapid clearance <i>in vivo</i> to be effective alone. The PPACK-liposomes were formulated as single unilamellar vesicles, with a diameter of 170.78 ± 10.59 nm and a near neutral charge. <i>In vitro</i> models confirmed the inhibitory activity of PPACK-liposomes, demonstrating a <i>K</i>_<i>I</i>′ of 172.6 nM. In experimental clots <i>in vitro</i>, treatment of formed clots completely abrogated any further clotting upon exposure to human plasma. The liposomes were evaluated <i>in vivo</i> in a model of photochemical-induced carotid artery injury, resulting in significantly prolonged arterial occlusion time over that of controls (69.06 ± 5.65 min for saline treatment, <i>N</i> = 6, 71.33 ± 9.46 min for free PPACK treated; <i>N</i> = 4, 85.75 ± 18.24 min for precursor liposomes; <i>N</i> = 4, 139.75 ± 20.46 min for PPACK-liposomes; <i>P</i> = 0.0049, <i>N</i> = 6). Systemic anticoagulant profiles revealed a rapid return to control levels within 50 min, while still maintaining antithrombin activity at the injury site. The establishment of a potent and long-acting anticoagulant surface over a newly forming clot with the use of thrombin targeted nanoparticles that do not require systemic anticoagulation to be effective offers an alternative site-targeted approach to the management of acute thrombosis