Tools for AD Research: Design of BACE2 Ligands

描述（由申请人提供）：本R21探索性提案采用一种综合的、靶向的方法来合理开发BACE2抑制剂-化学工具，可用于探索BACE2的基本生物学，并提高目前发现的活性BACE1抑制剂的选择性，这些抑制剂目前正在开发作为阿尔茨海默病的药物。阿尔茨海默病研究界普遍认为，成功治疗阿尔茨海默病的关键在于特异性抑制BACE1。BACE1对于淀粉样蛋白片段的释放至关重要，这些淀粉样蛋白片段随后形成细胞外神经性淀粉样斑块，这是AD发病的一个因素。大多数已知的BACE1抑制剂是相对非选择性的，并且靶向其他天冬氨酸蛋白酶，包括高度同源的BACE2。研究表明，BACE2可能作为一种替代α -分泌酶，这是另一种在淀粉样蛋白前体蛋白（APR）降解中起重要作用的关键酶，抑制它是非常不可取的。为了更好地了解BACE2在人体生理和APP加工中的作用，我们认为需要选择这种酶的抑制剂作为研究工具。我们假设，利用基于计算机辅助药物设计、药物化学和生物学的综合多学科方法，设计和合成BACE2抑制剂应该是可能的。在低纳摩尔范围内显示BACE2活性的化合物将在全细胞水平上进行筛选，以确定其改变APP处理的能力。这一迭代过程有望产生几种低纳摩尔、铅或药物样的选择性BACE2抑制剂。为了实现这一目标，我们的具体目标如下：(1)利用计算机辅助药物设计，包括从头/合理药物设计，虚拟聚焦组合文库（vFCL）生成，以及vFCL的计算机筛选，确定BACE2的铅和药物样低分子量选择性抑制剂。从从头/理性设计和vFCL方法中综合最佳候选方案；(2)测定候选配体对分离酶BACE1和BACE2的抑制作用。有效的、选择性的BACE2抑制剂将通过反复的分子建模、化学合成和生物学测试来进一步优化配体的活性和选择性；(3)对于最佳配体，在基于细胞的检测中探索其对APP加工的影响。根据活性、细胞毒性和由此产生的选择性指数对抑制剂进行优先排序。

DESCRIPTION (provided by applicant): This R21 exploratory proposal uses an integrated, target-oriented approach in the rational development of BACE2 inhibitors - chemical tools that can be used to explore the basic biology of BACE2 and improve the selectivity of presently discovered active BACE1 inhibitors that are currently being developed as medications for Alzheimer's disease. There exists a broad consensus in the AD research community that the key to successful treatment of AD lies in the specific inhibition of BACE1. BACE1 is crucial for release of the amyloidogenic fragments that later form the extracellular neuritic amyloid plaques - one of factors in the pathogenesis of AD. Most of the known BACE1 inhibitors are relatively non-selective and target other aspartic proteases including the highly homologous BACE2. It has been shown that BACE2 may serve as an alternative alpha-secretase, another crucial enzyme that plays an important role in degradation of the amyloid precursor protein (APR), inhibition of which is highly undesirable. To better understand the role of BACE2 in human physiology and APP processing, we believe that selective inhibitors of this enzyme are needed as research tools. We hypothesize that it should be possible to design and synthesize inhibitors for BACE2 using an integrated multidisciplinary approach based on the combination of the computer-aided drug design, medicinal chemistry, and biology. Those compounds demonstrating BACE2 activity in the low nanomolar range will be screened at the whole cell level for their ability to alter APP processing. This iterative process is expected to yield several low nanomolar, lead- or drug-like selective inhibitors of BACE2. To achieve this goal, our specific aims are as follow: (1) Identify lead- and drug-like low molecular weight selective inhibitors of BACE2 using computer-aided drug design including de novo/rational drug design, virtual focused combinatorial library (vFCL) generation, and in silico screening of vFCL. To synthesize the best candidates from the de novo/rational design and vFCL approaches; (2) Assay candidate ligands for their inhibition of BACE1 and BACE2 in isolated enzyme assays. Potent, selective BACE2 inhibitors will be used for further optimization of ligand activity and selectivity through iterative molecular modeling, chemical synthesis, and biological tests; (3) For the best ligands, explore their effects on APP processing in cell-based assays. Prioritize inhibitors on the basis of activity, cytotoxicity, and resultant selectivity indices.