Irreversible inhibitors of cholesterol esterase

胰腺胆固醇酯酶（CE）在膳食胆固醇的吸收中具有双重功能。首先，CE催化胆固醇酯水解，释放胆固醇供吸收；其次，CE将胆固醇从胶束转运到肠细胞表面，在那里发生吸收。也有证据表明，CE在肠细胞内的作用是在导致乳糜微粒形成的途径中重新酯化胆固醇。我们的假设是，抑制CE的任何这些功能将通过限制膳食胆固醇的生物利用度为治疗高胆固醇血症提供一种新的方法。本课题将重点研究不可逆CE抑制剂的开发，以防止胆固醇酯的水解。CE是一种丝氨酸酯酶，其催化机制与丝氨酸蛋白酶相似。我们建议开发选择性不可逆CE抑制剂。我们的具体目标是：(1)开发合成卤烯醇内酯的通用方案，如取代的6-氯吡咯酮作为胆固醇酯酶的潜在不可逆抑制剂；(2)使用分子建模和动力学研究来减少结构-活性关系，以开发选择性CE抑制剂。我们的初步工作证明了我们开发的综合方案的多功能性。选择性将在酶水平上确定，通过筛选化合物与蛋白酶凝乳胰蛋白酶、胰蛋白酶和弹性蛋白酶的CE失活率进行比较，这些蛋白酶都是在肠道内起作用的丝氨酸蛋白酶。

Pancreatic cholesterol esterase (CE) has a dual function in the absorption of dietary cholesterol. First, CE catalyzes the hydrolysis of cholesterol esters to liberate cholesterol for absorption; second, CE transports cholesterol from micelles to the surface of the enterocyte where absorption takes place. There is also evidence that CE functions within the enterocyte to re-esterify cholesterol in the pathway leading to the formation of chylomicrons. It is our hypothesis that inhibition of any of these functions of CE would provide a new approach to the treatment of hypercholesterolemia through limiting the bioavailability of dietary cholesterol. This proposal will focus on the development of irreversible inhibitors of CE for prevention of the hydrolysis of cholesterol ester. CE is a serine esterase with a catalytic mechanism that is similar to that of serine proteases. We propose to develop selective irreversible inhibitors of CE. Our specific aims are: (1) to develop versatile schemes for the synthesis of haloenol lactones such as substituted 6-chloropyrones as potential irreversible inhibitors of cholesterol esterase, and (2) to use molecular modeling and kinetic studies to reduce structure-activity relationships for the development of selective inhibitors of CE. Our preliminary work demonstrates the versatility of the synthetic schemes that we have developed. Selectivity will be determined at the enzyme level by screening compounds for their rates of inactivation of CE compared with the proteases chymotrypsin, trypsin and elastase, all of which are serine proteases that function within the intestine.