Probing the Proteinopathy Component of Light Chain Amyloidosis Pharmacologically

从药理学角度探讨轻链淀粉样变性的蛋白病成分

基本信息

批准号：
10625486
负责人：
JEFFERY W KELLY
金额：
$ 45.25万
依托单位：
SCRIPPS RESEARCH INSTITUTE, THE
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-07-01 至 2025-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10625486
关键词：
Albumins American Amino Acids Amyloidosis Aniline Binding Binding Sites Biological Assay Buffers Cardiac Cell secretion Chemistry Chemotherapy-Oncologic Procedure Chromatography Clinical Trials Clonal Expansion Computer software Coumarins Coupled Data Degenerative Disorder Deterioration Diagnosis Dissociation Disulfides Docking Endopeptidase K Family Fluorescence Polarization Human Kinetics Label Lead Learning Length Light Light-Chain Immunoglobulins Link Malignant Neoplasms Medical Metabolic Molecular Conformation Multiple Myeloma Organ Pathology Patients Peptide Hydrolases Pharmaceutical Chemistry Pharmaceutical Preparations Plasma Plasma Cells Plasma Proteins Population Positioning Attribute Process Publishing Structure Structure-Activity Relationship Therapeutic Time Variant X-Ray Crystallography analog base cancer cell computerized tools design dimer drug candidate experimental study improved lead candidate molecular sequence database monomer pharmacologic preservation prevent primary amyloidosis of light chain type scaffold screening small molecule standard of care

项目摘要

Project Summary Immunoglobulin light chain amyloidosis (AL) is a degenerative disease that putatively arises from light chain (LC) misfolding and aggregation. Full-length (FL) LCs are secreted from a clonally expanded plasma cell population in AL. Thus, AL patients suffer from both a plasma cell cancer and a LC misfolding and aggregation-associated proteinopathy that appears to compromise organ function, leading to progressive organ deterioration. Currently, AL is treated by repurposed multiple myeloma drugs that kill the clonal plasma cells secreting FL LCs. Mechanistically distinct therapeutic approaches are needed, particularly for patients with cardiac involvement who cannot tolerate the currently available chemotherapy regimens. In this proposal, we seek to produce lead candidates that ultimately would enable a clinical trial on a FL LC kinetic stabilizer targeting the proteinopathy component of AL. In Specific Aim 1, we hypothesize that the LC kinetic stabilizers to be synthesized in our hit-to-lead medicinal chemistry efforts can be dissected into four substructures—the “anchor substructure”, the “aromatic core”, the “linker module” and the “terminal aromatic component”. This hypothesis is based on 11 (FL LC)2•kinetic stabilizer crystal structures solved to date as well as the structure- activity relationship data resulting from the synthesis of over 300 FL LC2 kinetic stabilizers during the past 18 months. We will continue to use structure-based design principles that we learned developing the drug tafamidis, in addition to computational tools, especially the Schrödinger LiveDesign software, to replace the metabolically unstable and potentially toxic coumarin aromatic core and the diethyl aniline anchor substructure in our LC kinetic stabilizers, while introducing functionality to reduce albumin binding. The LiveDesign software utilizes a weighted combination of the predicted LogP values and docking scores to accurately predict albumin binding. In Specific Aim 2, we will develop FL LC2 plasma binding selectivity assays to validate our computational efforts to diminish albumin binding. First, we will add fluorescently labeled FL amyloidogenic LCs to pooled healthy donor plasma, along with proteinase K and a candidate kinetic stabilizer, and follow proteinase K endoproteolysis linked to FL LC2 conformational excursions chromatographically over time. If the kinetic stabilizer candidate selectively binds the amyloidogenic FL LC2 over all the other plasma proteins, including albumin, FL LC2 proteinase K endoproteolysis will be prevented–this assay is largely developed. Next, a subunit exchange assay will be developed to quantify pharmacologic FL LC2 kinetic stabilization in human plasma. This assay also quantifies kinetic stabilizer binding to albumin. A fluorescently labeled Cys214Ser FL LC2 variant facilitates subunit exchange experiments that afford the KD of kinetic stabilizer binding to the FL LC2 variants added to healthy plasma, as well as the kinetic stabilizer KD for binding to endogenous human albumin in plasma. These KDs will be used to generate binding selectivity ratios, i.e., KD of kinetic stabilizer binding to albumin / KD of kinetic stabilizer binding to the FL LC2, which we will maximize.

项目概要免疫球蛋白轻链淀粉样变性 (AL) 是一种由轻链引起的退行性疾病 (LC) 错误折叠和聚集全长 (FL) LC 由克隆扩增的浆细胞分泌。因此，AL 患者同时患有浆细胞癌和 LC 错误折叠。聚集相关的蛋白质病似乎会损害器官功能，导致进行性目前，AL 通过改变用途的多发性骨髓瘤药物来治疗，这些药物可杀死克隆血浆。需要机制不同的细胞分泌 FL LC，特别是对于患者。患有心脏受累的患者不能耐受目前可用的化疗方案。我们寻求生产最终能够对 FL LC 动力学稳定剂进行临床试验的主要候选药物针对 AL 的蛋白病成分在具体目标 1 中，我们采用了 LC 动力学稳定剂。在我们的先导药物化学工作中合成的可以分为四个子结构—— “锚定子结构”、“芳香核心”、“连接模块”和“末端芳香成分”。假设基于迄今为止已解决的 11 (FL LC)2•动力学稳定剂晶体结构以及结构- 过去 18 年间合成超过 300 种 FL LC2 动力学稳定剂的活性关系数据几个月后，我们将继续使用我们在开发药物时学到的基于结构的设计原则。 tafamidis，除了计算工具，特别是薛定谔LiveDesign软件，以取代代谢不稳定且具有潜在毒性的香豆素芳香核心和二乙基苯胺锚子结构在我们的 LC 动力学稳定剂中，同时引入了减少白蛋白结合的功能。利用预测LogP值和对接分数的加权组合来准确预测白蛋白在具体目标 2 中，我们将开发 FL LC2 血浆结合选择性测定法来验证我们的研究。首先，我们将添加荧光标记的 FL 淀粉样蛋白生成。将 LC 加入到汇集的健康供体血浆中，以及蛋白酶 K 和候选动力学稳定剂，然后遵循蛋白酶 K 内切蛋白水解与 FL LC2 构象偏移随时间的色谱变化有关。与所有其他血浆蛋白相比，候选动力学稳定剂选择性地结合淀粉样蛋白 FL LC2，包括白蛋白在内，FL LC2 蛋白酶 K 内切蛋白水解作用将被阻止——该测定法已得到广泛开发。接下来，将开发亚基交换测定来量化 FL LC2 的药理学动力学稳定性该测定还定量了与荧光标记的白蛋白 A 的结合。 Cys214Ser FL LC2 变体促进亚基交换实验，提供动力学稳定剂的 KD 结合添加到健康血浆中的 FL LC2 变体，以及用于结合的动力学稳定剂 KD 这些 KD 将用于生成结合选择性比，即 KD 。与白蛋白结合的动力学稳定剂/与 FL LC2 结合的动力学稳定剂的 KD，我们将其最大化。