Prenyldiphosphate Synthase Inhibitors: Novel Anti-Infective Agents

描述(由申请人提供)：这项工作的总体目标是开发新的抗感染药物，通过结合使用X射线结晶学、核磁共振光谱、量热法、酶和细胞生长抑制分析、定量结构-活性关系(QSAR)技术、量子化学、合成、药物释放和动物试验，针对原生动物和细菌中异戊二烯的生物合成途径。第一个具体目标是使用X射线结晶学和固体核磁共振波谱来研究新的抑制剂的结构，主要是来自克氏锥虫(美国锥虫病或恰加斯病的病原体)、布鲁氏锥虫(非洲昏睡病的病原体)、大肠杆菌和金黄色葡萄球菌的法尼基二磷酸合成酶(FP PS)，以及人FP PS(用于治疗骨吸收疾病的双膦酸类药物的靶标，在设计抑制剂选择性的背景下)。固体核磁共振将被用来补充结晶学结果，提供动力学(来自2HNMR)和质子化状态信息(来自13C，15N，31P化学位移)，用于QSAR研究。第二个具体目标是研究结构与功能的关系。我们将使用等温滴定热法和差示扫描量热法以及经典的酶抑制技术来研究缓蚀剂的结合，以推导出目标1中所研究的体系的配体结合常数(Kb)和Kis以及结合信息的热力学。此外，还将使用层析和核磁共振技术研究缓蚀剂与人骨和骨矿模型的结合。然后将使用QSAR方法对这些结果进行分析，包括使用不同的QSAR方法(以优化寄生虫/细菌抑制与人类FPPS抑制，并将骨吸收降至最低)，以及使用非传统的QM描述符和2H核磁共振序参数，以提高这些方法的预测实用价值。第三个也是最后一个特定目标是使用目标1和目标2的结果来设计并合成异戊二烯生物合成途径的特定抑制剂。这些抑制剂将包括新的吡啶、硫和膦物种，并将针对在原生动物和细菌酶的活性部位发现的独特的极性残基。然后，这些抑制剂将以生物可用形式配制，用于动物试验。我们还将重点研究协同或联合治疗方法，使用两到三种化合物，每一种化合物都针对异戊二烯生物合成途径：已经观察到这种强烈的协同作用，现在需要使用新型抑制剂进行优化。通俗摘要：提出的这项研究旨在引导治疗各种传染病的新疗法。在美国，这些疾病主要是细菌性的，是对公共健康的主要威胁，而在世界其他地方，传染病主要是由原生动物引起的。这项工作旨在开发治疗这两种疾病的药物。

DESCRIPTION (provided by applicant): The broad, overall objective of this work is to develop novel anti-infective agents which target the isoprene biosynthesis pathways in protozoa and bacteria by using a combination of x-ray crystallography, NMR spectroscopy, calorimetry, enzyme and cell growth inhibition assays, QSAR (quantitative structure-activity relationship) techniques, quantum chemistry, synthesis, drug delivery and animal testing. The First Specific Aim is to use x-ray crystallography and solid-state NMR spectroscopy to investigate the structures of novel inhibitors bound to, primarily, farnesyl diphosphate synthase (FPPS) from Trypanosoma cruzi (the causative agent of American trypanosomiasis or Chagas disease), Trypanosoma brucei (the causative agent of African sleeping sickness), E. coli and Staphylococcus aureus, as well as human FPPS (the target for the bisphosphonate drugs used in treating bone resorption diseases, of interest in the context of designing inhibitor selectivity). Solid-state NMR will be used to complement the crystallographic results by providing dynamics (from 2H NMR) and protonation state information (from 13C, 15N, 31P chemical shifts) for use in the QSAR investigations. The Second Specific Aim is to investigate structure-function relationships. We will investigate inhibitor binding by using isothermal titration calorimetry and differential scanning calorimetry, as well as by using classical enzyme inhibition techniques, to deduce ligand binding constants (KB) and Kis together with thermodynamics of binding information for the systems studied in Aim 1. In addition, inhibitor binding to human bone and bone mineral models will be investigated using chromatographic and NMR techniques. These results will all then be analyzed by using QSAR methods, including the use of differential QSAR methods (to optimize parasite/bacterial inhibition versus human FPPS inhibition, and to minimize bone adsorption), together with the use of non-conventional QM descriptors and 2H NMR order parameters, to enhance the predictive utility of these methods. The Third and Final Specific Aim is to use the results from Aims 1 and 2 to design and then synthesize specific inhibitors of the isoprene biosynthesis pathway. These inhibitors will include novel pyridium, sulfonium and phosphonium species and will be designed to target unique polar residues found in the active site of the protozoal and bacterial enzymes. These inhibitors will then be formulated in bioavailable forms for use in animal testing. We will also focus on synergistic or combination therapy approaches by using two or three compounds, each of which target the isoprene biosynthesis pathway: such strongly synergistic interactions have already been observed and now need to be optimized using the novel inhibitors. Lay Summary: The research proposed is designed to lead to new therapeutic approaches to treat a variety of infectious diseases. In the US, these diseases are primarily bacterial and are a major public health threat while in the rest of the world, infectious diseases are mainly caused by protozoa. This work seeks to develop drugs to treat both sorts of disease.