Novel Structure-Based Rifamycins for Drug-resistant TB and HIV Co-infection

 描述（由申请人提供）：结核病是最重要的世界卫生问题之一，2012年造成45万人死亡。耐多药结核病和广泛耐药结核病发病率的增加已成为一个日益严重的健康危机。显然，开发针对M的新型抗生素存在关键的未满足的需求。结核病（MTB），以克服目前的治疗耐药性。然而，在过去的40年里，只有一种具有新靶点的新药（贝达喹啉）被批准用于治疗结核病。在抑制新靶点方面缺乏成功，这表明重新研究以前有效药物的靶点可能是一种更好的方法。MTB RNA聚合酶（RNAP）是一个被证明的和有吸引力的目标，因为它是细菌生存所必需的，并且在原核和真核RNAP之间存在低相似性。利福霉素（RIF）是MTB RNAP的非常有效的抑制剂;然而，这些药物通过靶RNAP的突变和由RIF激活人黑曲霉素X受体（hPXR）引起的药物-药物相互作用而产生耐药性（RIFR）（特别是在TB/HIV合并感染中存在问题）。我们提出了一个多学科和综合性的计划，涉及基于结构的类似物合成，高通量筛选（HTS），体外RNAP抑制评价，抑制剂 *RNAP复合物的X射线晶体结构测定，以及研究，以最大限度地减少hPXR激活，以发现新的药物，以解决当前MTB治疗的局限性。我们组建了一支在RNA和细菌RNA聚合酶的生物化学和酶学方面具有技能和经验的团队，数十年的合成药物化学和制药行业的分子建模，开创性的细菌RNA聚合酶结构生物学，以及国际公认的M.结核我们团队最近的出版物（四篇论文，一篇综述和一项专利申请）证明了我们方法的原理。我们的第一组RIF类似物表现出增强的针对RIFR RNAP的活性，在一种情况下hPXR活化减少，并以设计的模式结合RNAP，如我们的X射线晶体结构所示。这些X射线晶体结构现在为我们开发改进的RIF提供了概念框架。Ebright及其同事最近的研究（Zhang et al.，（2014）eLife 3 e02450，3994528）提供了进一步的原理证明，即利福霉素核心的制备可以增强针对RIFR MTB的活性。我们还开发了一种有效的体外RNAP检测方法，可用于HTS，并将其用作发现新型RNAP抑制剂的补充方法。我们期望，本申请中描述的充分阐述的活动将产生用于药物开发的新型候选药物，并最终改善结核病的治疗，特别是在艾滋病毒-结核病合并感染和耐药结核病患者中。

 DESCRIPTION (provided by applicant): Tuberculosis is one of the most important world health problems, responsible for 450,000 deaths in 2012. Increases in the occurrence of multidrug-resistant TB (MDR-TB) and extensively drug-resistant TB (XDR-TB) have become an increasingly problematic health crisis. Clearly there is a critical unmet need for the development of novel antibiotics against M. tuberculosis (MTB) to overcome resistance to current therapeutics. However, only one new drug (bedaquiline) with a novel target, the mycobacterial ATP synthase, has been approved for the treatment of tuberculosis in the last 40 years. The lack of success in inhibiting novel targets suggests that the reinvestigation of targets of previously effective drugs may be a better approach. The MTB RNA polymerase (RNAP) is a proven and attractive target because it is essential for bacterial survival, and there is low similarity between prokaryotic and eukaryotic RNAPs. The rifamycins (RIFs) are very potent inhibitors of MTB RNAP; however, these agents suffer from resistance (RIFR) via mutation of the target RNAP and drug-drug interactions that result from RIF activation of the human pregnane X receptor (hPXR) (particularly problematic in TB/HIV co-infection). We propose a multi-disciplinary and comprehensive program that involves structure-based analogue synthesis, high-throughput screening (HTS), in vitro RNAP inhibition evaluation, X-ray crystal structure determinations of inhibitor*RNAP complexes, and studies to minimize hPXR activation towards uncovering novel agents to address current MTB treatment limitations. We have assembled a team with skills and experience in biochemistry and enzymology of RNA and bacterial RNA polymerase, decades of synthetic medicinal chemistry and molecular modeling in the pharmaceutical industry, groundbreaking structural biology of bacterial RNA polymerase, and internationally-recognized expertise in the microbiology of M. tuberculosis. The recent publications (four papers, one review, and one patent application) of our team demonstrate the proof of principle of our approach. Our very first set of RIF analogues exhibit enhanced activity against RIFR RNAP, in one case a reduction in hPXR activation, and bind to the RNAP in the designed mode as shown by our X-ray crystal structures. These X-ray crystal structures now provide a conceptual framework for our development of improved RIFs. Recent studies by Ebright and co-workers (Zhang et al., (2014) eLife 3 e02450, 3994528) provide further proof of principle that elaboration of the rifamycin core can yield enhanced activity against RIFR MTB. We have also developed an efficient in vitro RNAP assay, scalable for HTS, and will use this as a complementary approach for discovering novel RNAP inhibitors. It is our expectation that the fully articulated campaign described in this application will yield novel candidates for drug development and ultimately improve treatment for tuberculosis, especially in patients with HIV-TB co-infection and drug-resistant tuberculosis.