Abstract 448: The Association of 5A Peptide With Sphingomyelin Increases Its Ability to Efflux Cholesterol Both in vitro and in vivo

Introduction 5A (DWLKAFYDKVAEKLKEAF-P-DWAKAAYDKAAEKAKEAA) is an apolipoprotein A-I mimetic peptide that is specific for promoting cholesterol efflux by the ABCA1 transporter and has been shown to reduce atherosclerosis and inflammation in animal models. Hypothesis In this study, we complexed the 5A peptide with sphingomyelin (SM) or 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) to compare the cholesterol efflux capacity of the complex and its affect on atherosclerosis. Results The optimal peptide to lipid ratio was found to be 1 to 7 (mole/mole) for both lipids. The average particle size measured by light scattering technique was 15.9 ± 2.2 nm for 5A-SM complex and 14.4 ± 3.5 nm for 5A-POPC complex. 5A-SM exhibited higher physical binding of fluorescently labeled cholesterol relative to 5A-POPC with Bmax of 182.6 ± 14.4 versus 124.8 ± 8.2, respectively. In the cholesterol efflux assay (18h), both complexes showed near equal efflux capacity through ABCA-1 transporter (25.4 ± 1.2 efflux for 5A-SM and 23.0 ± 0.2 % efflux for 5A-POPC). Interestingly, 5A-SM showed significantly higher cholesterol efflux in ABCG-1 and SR-B1 transfected cell lines compere to 5A-POPC (42.7 ± 1.8% vs. 29.1 ± 0.6 and 29.0 ± 2.4% vs. 13.3 ± 0.2%, respectively). Lipid free 5A peptide showed lower cholesterol efflux through ABCA-1 transporter (15.8 ± 1.2%) and showed no efflux through ABCG-1 or SR-B1. The abilities of the peptide-lipid complexes to mobilize cholesterol in vivo were tested on wild type rats. Two hours after intravenous injection of 100 mg/kg 5A-SM plasma cholesterol increased about 3-fold above baseline with a peak at 6 hours and returned to the baseline by 24 hours. 5A-POPC increased plasma cholesterol 2-fold above and returned to baseline by 24 hours. Conclusions The 5A complexed with SM showed similar ability to efflux cholesterol by ABCA1 than the 5A-POPC complex but improved cholesterol efflux capacity by ABCG1 and SR-BI. A greater rise in HDL-C was observed in vivo after treatment of rats with 5A-SM versus 5A-POPC, suggesting that the 5A-SM complex may show superior ability in increasing Reverse Cholesterol Transport and in reducing atherosclerosis, which will be examined in future studies.