Targeting the serine protease PCSK9 via covalent complementarity

描述（由申请人提供）：Proprotein convertase subtilisin/ keexin type 9 （PCSK9）是一种丝氨酸蛋白酶，可调节LDL胆固醇水平，进而调节动脉粥样硬化的发展。流行病学和遗传学研究提供了强有力的证据，表明PCSK9突变与心血管疾病有关。有趣的是，PCSK9的催化活性并不是PCSK9/ ldl -受体相互作用的直接要求。因此，该蛋白已成为开发阻断PCSK9/LDL-R相互作用的治疗方法的一个令人兴奋的靶点。本研究拟从生化角度评价PCSK9催化活性与PCSK9/LDL-R相互作用之间的关系。在Aim 1中，该研究将设计具有合理设计活性位点修饰的“亲电敏感”PCSK9类似物

DESCRIPTION (provided by applicant): Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a serine protease which regulates LDL cholesterol levels and, by extension, the development of atherosclerosis. Epidemiologic and genetic studies have provided strong evidence that mutations in PCSK9 are related to cardiovascular outcomes. Interestingly, the catalytic activity of the PCSK9 is not directly required for the PCSK9/LDL-receptor interaction. As such, this protein has become an exciting target for development of therapeutics to block this PCSK9/LDL-R interaction. This study proposes to evaluate the relationship between PCSK9 catalytic activity and the PCSK9/LDL-R interaction on a biochemical basis. In Aim 1, the study will engineer "electrophile-sensitive" PCSK9 analogs with rationally designed active site modifications that will allow for specific and covalent binding of small molecule inhibitors with electrophilic handles. The catalytic activity of the ES-PCSK9 analogs, along with the ability of the small molecules to achieve specific inhibition, will be assessed in vitro. The ES-PCSK9 will be introduced in a cellular system, and the effects on LDL processing will be assessed. In Aim 2, the structure activity relationship of the specific inhibitors will be assessed. The study will test the hypothess that binding of inhibitor to the PCSK9 active site will cause allosteric inhibition of the PCSK9/LDLR interaction. In Aim 3, the role of intracellular and extracelullar PCSK9 to LDL regulation will be evaluated. The mechanisms of clinically significant polymorphisms will be assessed by introducing GOF and LOF mutations into ES-PCSK9. The possibility of dominant negative and dominant positive effects will be evaluated. In addition to the training in basic science and translational research through the conduct of the study, the applicant will undertake didactic instruction in synthetic chemistry and chemical biology techniques through auditing graduate courses offered at the University of California, San Francisco. PUBLIC HEALTH RELEVANCE: Atherosclerosis is the underlying disease entity causing the majority of heart disease in Western countries. PCSK9 is a recently identified protease which has a major biochemical role in modulating LDL cholesterol, a major risk factor in the development of atherosclerosis. This proposal develops selective inhibitors of engineered PCSK9 analogs to further define the nature and possibility for therapeutic inhibition of the PCSK9/LDL receptor.