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

描述（由申请人提供）：本申请的总体目标是利用传统药物化学和基于计算结构的药物设计相结合，开发1-脱氧-d -木醛糖-5-磷酸还原异构酶（DXR）的新型小分子抑制剂，并测试其对致病菌和寄生虫的体外生物活性。异戊二烯的生物合成对所有生物都是必不可少的。人类使用甲羟戊酸途径产生二磷酸异戊烯基（IPP）和二磷酸二甲基烯基（DMAPP），这是所有类异戊二烯生物合成的两种常见前体；然而，在大多数致病菌中，如铜绿假单胞菌和结核分枝杆菌，以及顶复合体寄生虫，如恶性疟原虫和弓形虫，非甲羟戊酸途径被用来制造IPP和DMAPP。由于人类缺乏非甲羟戊酸途径中的所有7种酶，因此它已成为抗感染药物发现的一个有吸引力的靶点。在最近的临床试验中，Fosmidomycin已被发现是该途径的唯一有效抑制剂，可阻断第二酶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性寄生虫（如弓形虫）对fosmidomycin具有耐药性。此外，它的药代动力学特征较差，在血浆中的半衰期为0.5-1.5 h。鉴于目前面临的耐药性迅速上升的破坏性局面以及新的抗感染药物的短缺，迫切需要寻找新的武器来治疗传染病。第一个具体目标是结合药物化学和计算，基于结构的药物设计来开发新的DXR抑制剂。基于合理的结构设计，我们发现了新的药物样先导抑制剂，其Kis低至310 nM，可抑制重组大肠杆菌DXR酶。我们的对接研究表明，它们可以以不同于fosmidomycin的模式结合DXR。这些类药物化合物应该具有进一步开发的巨大潜力。我们建议：1)利用药物化学的方法，以先导抑制剂的支架为基础，构建多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)得到DXR与新型抑制剂配合物的x射线晶体结构；4)利用计算和晶体学研究的结果来指导我们进一步的药物设计和合成。第二个特定目标是测试我们的抑制剂对广泛的细菌和顶复合体寄生虫及其重组DXR酶的体外生物活性。最后，我们还将测试我们的强效DXR抑制剂对人类细胞系的细胞毒性，以评估其潜在毒性。

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.