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Beta-carotene and cholesterol absorption in wild type and scavenger receptor class B type I (SR-BI) knockout mice and the uptake of these lipids in vitro were measured using brush border membrane (BBM) vesicles. The analysis shows that ezetimibe, a representative of a new class of drugs that inhibit intestinal cholesterol absorption, blocks SR-BI- and CD36-facilitated uptake of cholesterol into COS-7 cells.

Research has been conducted on the long-chain fatty acids. The kinetics of the fatty acid transfer from the different donor systems to the unilamellar egg phosphatidylcholine vesicles which contain pH-sensitive fluorophore pyranine has been investigated and the results are reported.

Research demonstrates that the scavenger receptor, cluster determinant 36, of human and rabbit brush border membrane mediates free cholesterol uptake whereas esterified cholesterol uptake is through scavenger receptor class B type 1. However, both receptors are involved in the uptake of free cholesterol.

Research has been conducted on the lipid uptake inhibition mechanism. Results indicate that such inhibitors as serum apolipoproteins, serum lipoproteins and synthetic alpha-helical peptides are the ligands of the scavenger receptor class B type I.

Research has been conducted on the sterol carrier protein 2 known as nonspecific lipid transfer protein. Results indicate that it functions as a fatty acid and sterol carrier.

The presence of scavenger receptor class B type I in the small intestine brush border membrane was investigated. Experimental results confirmed this presence and showed that scavenger class B type I facilitates the uptake of dietary cholesterol from either bile salt micelles or phospholipid vesicles. The receptor was also found to be capable of functioning as a port for several additional classes of lipids such as phospholipids, cholesteryl esters and triacylglycerols.

Reconstitution of the cholesterol transport activity of rabbit brush border membrane (BBM) artificial membrane system was done to establish the effect of hydrolysis of lipids to monoacylglycerols and fatty acid in the uptake of triacylglycerols by the brush border membrane. Results have shown that sterol transport protein was integrated into the lipid bilayer as evidenced by density gradient centrifugation on a Ficoll gradient.

The differences between cholesterol and plant sterol sitosterol uptake in the brush border was investigated through the utilization of comparative study of sterol uptake by brush border membrane vesicles and sterol absorption by Caco-2 cells. Results have shown that detection of possible inhibitory effects on cholesterol uptake can occur only at the presence of large amounts of sitosterol contents in the donor. The mechanism of energy-independent uptake of sitosterol mimics that of the cholesterol.

The protein present in the lumenal side of the brush border membrane vesicles catalyzes the chain cholesteryl ester and ether absorption. The protein-mediated absorption of cholesteryl ester follows a second-order reaction in the first phase. Autoproteolysis induces the release of water-soluble form of the protein. The free and esterified cholesterol exchange between the two populations of small unilamellar phospholipid vesicles is catalyzed by the protein.

We have utilized our recently developed in vitro assay to address two key questions in the design of small-molecule cholesterol absorption inhibitors using ezetimibe, the only drug yet approved for the inhibition of cholesterol absorption in the small intestine, as a starting point: (1) the role of glycosylation and (2) the importance of the beta-lactam scaffold of ezetimibe for inhibitory activity. A wide range of nonhydrolyzable phenolic glycosides of ezetimibe were synthesized and demonstrated to be active inhibitors of cholesterol absorption using the brush border membrane vesicle assay. The analogous azetidines provided access to a variety of inhibitors in vitro, suggesting that the beta-lactam of ezetimibe merely serves as a ring scaffold to appropriately position the required substituents. Our findings highlight several promising strategies for the design of alternative small-molecule cholesterol absorption inhibitors that could ultimately be useful in preventing cardiovascular disease by lowering blood cholesterol levels.

[reaction: see text] The use of carbohydrate sulfonyl chlorides is introduced as a new, facile glycoconjugation method which could find broad applications. We demonstrate the approach by synthesizing a number of glycosylated cholesterol absorption inhibitors which display high inhibitory efficacies in our recently established in vitro assay. Furthermore, we highlight an advantage of the electron-withdrawing nature of the sulfonyl linkage which allowed the synthesis of otherwise unstable azetidine conjugates.