Novel inhibitors of 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR)

描述(申请人提供)：本提案的总体目标是利用传统药物化学和基于计算的、基于结构的药物设计相结合的方法来开发新型小分子1-脱氧-D-木酮糖-5-磷酸还原异构酶(DXR)抑制剂，并测试其对病原菌和寄生虫的体外生物活性。异戊二烯的生物合成是所有生物所必需的。人类利用甲氧戊酸途径产生异戊二烯基二磷酸(IPP)和二甲基烯丙基二磷酸(DMAPP)，这是所有异戊二烯生物合成的两个共同前体；然而，在大多数致病细菌，如铜绿假单胞菌和结核分枝杆菌，以及顶复形体寄生虫，如恶性疟原虫和弓形虫，非甲戊酸途径用于产生IPP和DMAPP。由于人类缺乏非甲伐他酸途径的所有7种酶，它已成为抗感染药物开发的一个有吸引力的目标。在最近的临床试验中，磷霉素被发现是这一途径的唯一有效的抑制剂，它阻断了第二种酶DXR，并对许多革兰氏阴性菌具有抗菌活性和抗疟疾活性。然而，革兰氏阳性菌(如结核分枝杆菌)和一些革兰氏阴性菌以及某些致病寄生虫(如弓形虫)对磷霉素有抗药性。此外，它的药代动力学曲线很差，在血浆中的半衰期为0.5-1.5小时。鉴于目前面临快速增长的耐药性和新的抗感染药物短缺的毁灭性形势，迫切需要寻找治疗传染病的新武器。第一个具体目标是利用药物化学和计算、基于结构的药物设计相结合的方法来开发DXR的新型抑制剂。基于合理的、基于结构的设计，我们发现了新型的类药物铅抑制剂，其对重组大肠杆菌DXR酶的KI低至310 nM。我们的对接研究表明，它们可以与DXR以不同于磷霉素类的方式结合。这些类药物化合物应该有很大的发展潜力。我们建议1)利用药物化学的方法，在先导抑制剂骨架的基础上建立几个系列的化合物文库，以寻找活性更高的化合物；2)对这些化合物进行QSAR研究；3)获得DXR与我们的新型抑制剂络合物的X射线晶体结构；4)使用计算和结晶学研究的结果来指导我们进一步的药物设计和合成。第二个具体目标是测试我们的抑制剂对广泛的细菌和顶面复合体寄生虫及其重组DXR酶的体外生物活性。最后，我们还将测试我们强大的DXR抑制剂对人类细胞株的细胞毒性，以评估它们的潜在毒性。 公共卫生相关性：提出的这项研究旨在引导新的潜在疗法来治疗耐药传染病。我们将专注于发现和开发新的化合物，这些化合物可以阻断细菌和疟疾寄生虫中唯一存在的基本生物靶标。

DESCRIPTION (provided by applicant): The overall objectives of this proposal are to use a combination of traditional medicinal chemistry and computational, structure based drug design to develop novel small molecule inhibitors of 1-deoxy-D-xylulose-5- phosphate reductoisomerase (DXR) and test their in vitro biological activities on pathogenic bacteria and parasites. Isoprene biosynthesis is essential to all organisms. Humans use the mevalonate pathway to produce isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), two common precursors for all isoprenoid biosynthesis; however, in most pathogenic bacteria, such as P. aeruginosa and M. tuberculosis, as well as apicomplexan parasites, such as P. falciparum and T. gondii, the non-mevalonate pathway is used to make IPP and DMAPP. Since humans lack all the 7 enzymes in the non-mevalonate pathway, it has become an attractive target for anti-infective drug discovery. Fosmidomycin has been found to be the only potent inhibitor of this pathway, blocking DXR, the 2nd enzyme, and has antibacterial activity against many Gram- negative bacteria and antimalarial activity in recent clinical trials. However, Gram-positive bacteria (e.g., M. tuberculosis) and some Gram-negative bacteria as well as certain pathogenic parasites (e.g., T. gondii) are resistant to fosmidomycin. In addition, it has a poor pharmacokinetic profile with a half-life in plasma of 0.5-1.5 h. Given the current devastating situation facing quickly rising drug resistance as well as shortage of new anti- infective drugs, there is a pressing need to find new weaponry for infectious diseases. The first Specific Aim is to use a combination of medicinal chemistry and computational, structure based drug design to develop novel inhibitors of DXR. Based on rational, structure based design, we have found novel, drug-like lead inhibitors with Kis as low as 310 nM against a recombinant E. coli DXR enzyme. Our docking studies showed that they could bind to DXR in different modes from that of fosmidomycin. These drug-like compounds should have great potential for further development. We propose 1) to use medicinal chemistry to make several series of compound libraries based on the scaffolds of the lead inhibitors, in order to find compounds with improved activity; 2) to carry out QSAR studies of these compounds; 3) to obtain x-ray crystal structures of DXR in complex with our novel inhibitors; and 4) to use the results from the computational and crystallographic studies to guide our further drug design and synthesis. The second Specific Aim is to test in vitro biological activity of our inhibitors on a broad range of bacteria and apicomplexan parasites as well as their recombinant DXR enzymes. Finally, we will also test the cytotoxicity of our potent DXR inhibitors on human cell lines to evaluate their potential toxicity. PUBLIC HEALTH RELEVANCE: The research proposed is designed to lead to new potential therapeutics to treat drug-resistant infectious diseases. We will focus on the discovery and development of novel compounds that block essential biological targets that are exclusively found in bacteria and malaria parasites.