PROJECT SUMMARY/ABSTRACT Atherosclerosis is the leading cause of human mortality worldwide. There has been a great interest in developing novel therapeutics for this disease that function via an orthogonal mechanism of action to currently available drugs. One promising strategy is to enhance the function of high-density lipoproteins (HDLs). HDLs facilitate reverse cholesterol transport (RCT) to transfer excess cholesterol from peripheral tissues to the liver for elimination. The proposed research, supported by strong recent progress, seeks to advance a novel supramolecular strategy for improving HDL function in vivo to combat atherosclerosis. The proposed studies build on the successes of our research program, funded by an NHLBI R01 grant over the past eight years. Despite substantial progress in the field of apolipoprotein mimetic peptides, including evaluation of several candidates in phase 1 and 2 human clinical trials, the inherent pharmacological shortcomings typically associated with linear peptides have been a major impediment to advancing HDL- modulating peptides through the clinic. Our proposed research seeks to develop a novel class of chemotypes that can recapitulate the functional attributes of helical lipoprotein mimetics, but without their inherent limitations. We have established that appropriately designed eight-residue self-assembling cyclic D,L-a-peptides are effective HDL modulating agents that can remodel human and mouse plasma HDLs and enhance cholesterol efflux from macrophage cells. These peptides specifically increase the level of pre-beta HDL particles (subspecies of HDLs considered to be the most anti-atherogenic), reduce LDL-cholesterol and triglyceride levels, and promote anti-inflammatory effects with concomitant prevention of atherosclerosis in mouse disease models. The abiotic structure of cyclic D,L-a-peptides overcomes many of the shortcomings typically associated with linear peptides, such as low serum/plasma stability and high production costs, among others. Specific aims 1 and 2 of the proposed research involve i) implementing a series of mechanism-based functional bioassays and biophysical assays to optimize and better characterize the mode of action of the lead compounds, and ii) establishing in vivo pharmacology, toxicology, and efficacy of selected cyclic peptides in two leading mouse models of atherosclerosis. Specific aim 3 of the proposed research is based on an unexpected, remarkable observation made in the course of our recent studies. We have discovered that trifluoroacetate (TFA), administered orally or parenterally, causes dose-dependent reductions in plasma cholesterol levels and prevents the development of atherosclerosis in vivo. The implications of this discovery are potentially broad because nearly all synthetic peptides, employed across diverse biological settings, contain TFA counterions resulting from purification by preparative HPLC. As such, the effects of TFA on cellular metabolism in ...