Identification of new LAT-1 transporter substrates for drug delivery

用于药物输送的新型 LAT-1 转运蛋白底物的鉴定

• 批准号: 9377441
• 负责人: Allen Thomas
• 金额: $ 39.91万
• 依托单位: UNIVERSITY OF NEBRASKA KEARNEY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-18 至 2022-03-17
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9377441
• 关键词: Active Biological Transport; Alzheimer' s Disease; Amino Acid Transporter; Amino Acids; Antineoplastic Agents; Binding; Binding Sites; Biological; Biological Assay; Biology; Blood - brain barrier anatomy; Brain; Brain Diseases; Carboxylic Acids; Carrier Proteins; Cells; Central Nervous System Agents; Chemicals; Computer Simulation; Data; Development; Disease; Drug Delivery Systems; Drug Design; Drug Industry; Drug Kinetics; Drug Targeting; Drug Transport; Drug vehicle; Glucose; Hormones; Knowledge; Legal patent; Levodopa; Link; Literature; Malignant Neoplasms; Malignant neoplasm of brain; Mass Spectrum Analysis; Melphalan; Modeling; Neuraxis; Neutral Amino Acids; Organic Chemistry; Parkinson Disease; Peer Review; Pharmaceutical Preparations; Phenylalanine; Prodrugs; Proteins; Publications; Research; Role; Route; SN-38; Scientist; Structure; Structure-Activity Relationship; Surface; Techniques; Testing; Thyroid Hormones; Time; Tissues; Topoisomerase-I Inhibitor; Work; analog; base; beta-site APP cleaving enzyme 1; cancer cell; cancer clinical trial; cancer type; chemical property; chemical substitution; clinical toxicology; cost; design; experience; experimental study; gabapentin; improved; inhibitor/antagonist; irinotecan; mimetics; novel; pre-clinical; predictive modeling; prevent; scaffold; targeted agent; targeted delivery; tumor; uptake

The blood-brain barrier (BBB) prevents entry of most drug molecules into the brain, an obstacle which adds time and cost to the discovery and development of central nervous system (CNS) drugs. The BBB possesses transporter proteins that allow biological molecules including amino acids, glucose, and hormones to cross as needed. One of these transporter proteins, the L-type amino acid transporter (LAT-1) transports specific amino acids (e.g. phenylalanine) as well as compounds that resemble them (e.g. the drug L-DOPA). This transporter offers a potential route for targeted drug delivery to the brain; moreover, it has been demonstrated that drugs chemically linked to amino acids can cross the BBB via LAT-1. It is known that some molecules are transported (substrates), while others only bind to the transporter surface (inhibitors); however, the structural requirements for either type of activity is poorly understood. We propose mapping the structure-activity relationship (SAR) for molecules that interact with LAT-1 with the aspiration of making this transporter a practical and widely used vehicle for drug delivery. Our approach involves using a computational model of the LAT-1 binding site to guide the synthesis of compounds similar in structure to natural amino acids but with unique chemical properties that could be used to design improved drug delivery agents. Our specific aims are three-fold. First, we will synthesize amino acids chemically substituted with groups that our model predicts will form beneficial interactions with the transporter. Second, we will use cell-based assays to test these compounds. To determine whether they are transported, we will use cis-inhibition and trans-stimulation cell assays which have successfully identified previously unknown substrates. SAR from these experiments will be used to refine our computational model in order to generate hypotheses to guide further substrate optimization. The intracellular concentration of selected compounds will be analyzed by LC-MS/MS to verify our hypothesis that LAT-1 activity increases cell uptake. A third aim of this project will involve the synthesis and testing of amino acid prodrugs of a topoisomerase I inhibitor for cancer (SN-38) to assess whether the SAR derived from optimization of LAT-1 substrates in specific aims 1 and 2 can be applied to improve prodrug cellular uptake. Additionally, the effect of the linker joining SN-38 and the amino acid promoiety, which may have a significant impact on prodrug pharmacokinetics and efficacy, will be examined. This project has broad applications for treating many different diseases both in the brain and other tissues where LAT-1 is heavily expressed, including cancer.

血脑屏障（BBB）阻止大多数药物分子进入大脑，这一障碍增加了时间 以及中枢神经系统（CNS）药物的发现和开发的成本。BBB拥有 转运蛋白，允许生物分子，包括氨基酸，葡萄糖和激素， needed. L型氨基酸转运蛋白（LAT-1）是这些转运蛋白中的一种，它转运特定的氨基酸， 酸（例如苯丙氨酸）以及类似于它们的化合物（例如药物L-DOPA）。这种转运蛋白 提供了一个潜在的途径，靶向药物输送到大脑;此外，已经证明，药物 化学连接的氨基酸可以通过LAT-1穿过BBB。我们知道有些分子 （底物），而其他仅结合到转运蛋白表面（抑制剂）;然而，结构要求 对这两种活动都知之甚少。我们建议映射的构效关系（SAR） 与LAT-1相互作用的分子，希望使这种转运蛋白成为一种实用和广泛使用的 用于药物递送的载体。我们的方法涉及使用LAT-1结合位点的计算模型来指导 合成结构与天然氨基酸相似但具有独特化学性质的化合物， 可以用来设计改进的药物输送剂。我们的具体目标有三个方面。首先，我们将综合 我们的模型预测，用化学基团取代的氨基酸将与 传送器。其次，我们将使用基于细胞的测定来测试这些化合物。以确定它们是否 运输，我们将使用顺式抑制和反式刺激细胞测定，已成功地确定 以前未知的基板。来自这些实验的SAR将用于改进我们的计算模型， 以产生假设来指导进一步的衬底优化。选择的细胞内浓度 将通过LC-MS/MS分析化合物以验证我们的假设，即LAT-1活性增加细胞摄取。一 本项目的第三个目标是合成和测试拓扑异构酶I抑制剂的氨基酸前药 对于癌症（SN-38），评估SAR是否源自特定目标1中LAT-1底物的优化 和2可用于改善前药的细胞摄取。此外，连接SN-38和SN-38的接头的作用也被证实。 可能对前药药代动力学和功效具有显著影响氨基酸前体部分将 接受检查。该项目具有广泛的应用，用于治疗许多不同的疾病，无论是在大脑和其他 LAT-1高表达的组织，包括癌症。