Structure-based Identification and Functional Characterization of SSH1 Inhibitors

 描述(申请人提供)：由于阿尔茨海默病(AD)大脑的主要定义特征是被称为Aβ和Tau的有毒蛋白质的过度积累，了解Aβ与tau功能障碍的生物学机制对于设计有效的AD治疗方法至关重要。我们已经确定SSH1，一种激活Cofilin的磷酸酶，作为AD的一个有希望的治疗靶点，因为SSH1影响Aβ和Tau。我们还鉴定了几种SSH1抑制剂化合物，它们可以有效地降低Aβ诱导的神经毒性和Tau过度磷酸化，同时减少Aβ的产生。在提案中，我们将对SSH1抑制剂化合物进行化学修饰，以验证对SSH1抑制至关重要的官能团，并建立一个铅优化平台。我们将优化并进行SSH1和其他双特异性磷酸酶的体外磷酸酶活性测定。我们还将使用现有的和化学修饰的化合物进行AD相关的基于细胞的分析。复杂的X射线晶体结构将使用现有的和修饰的化合物与SSH1的催化结构域进行物理接触来确定，以研究它们相互作用的分子基础。双扰动实验将利用修饰化合物和SSH1的定点突变来探索特定的抑制剂官能团和蛋白质残基对配体结合的贡献。基于修正的结构模型，将进行虚拟对接和基于结构的设计，以提高现有的配体效率，并将进行新的虚拟筛选，以确定更多的SSH1抑制剂化学类型。最后，基于新的基于结构的配基效率模型，我们将合成一种修饰的化合物，以证明这种修饰的化合物在体外具有更好的SSH1抑制活性和ADME特性。因此，这一R21提案的结果有望在SSH1与现有化合物和新化合物之间产生有价值的结构功能关系，并为未来R01提案中广泛的先导优化和药代动力学研究建立一个基于结构的平台。

 DESCRIPTION (provided by applicant): As the major defining characteristic of Alzheimer's disease (AD) brains is the excessive accumulation of toxic proteins called Aβ and Tau, understanding the biological mechanisms by which Aβ connects to tau dysfunction are critical for designing effective therapeutic treatments for AD. We have identified SSH1, a phosphatase that activates Cofilin, as a promising therapeutic target for AD, as SSH1 impacts both Aβ and Tau. We have also identified several SSH1 inhibitor compounds that effectively decrease both Aβ-induced neurotoxicity and Tau hyperphosphorylation while decreasing Aβ production. In the proposal, we will perform chemical modifications of SSH1 inhibitor compounds to verify the functional groups critical for SSH1 inhibition and establish a platform for lead optimization. We will optimize and carry out in vitro phosphatase activity assays for SSH1 and other dual specificity phosphatases. We will also perform AD-relevant cell-based assays using existing and chemically modified compounds. Complex X-ray crystal structures will be determined using existing and modified compounds in physical contact with the catalytic domain of SSH1, to investigate the molecular basis of their interactions. Double perturbation experiments will be used to probe the contribution of specific inhibitor functional groups and protein residues to ligand binding using modified compounds and site-specific mutagenesis of SSH1. Based on a revised structural model, virtual docking and structure-based design will be performed to improve existing ligand efficiency, and new virtual screening will be carried out to identify additional SSH1 inhibitor chemotypes. Finally, based on the new structure-based ligand efficiency model, we will perform synthesis of a modified compound to demonstrate the proof-of-concept that such modified compound exhibits improved SSH1 inhibitory activity and ADME characteristics in vitro. Therefore, results of this R21 proposal are expected to yield valuable structure function relationship between SSH1 and existing and new compounds as well as establish a structure-based platform for extensive lead optimization and pharmacokinetic studies in a future R01 proposal.