Optimization of novel pyranopyridine efflux pump inhibitors

摘要 目前治疗耐多药革兰氏阴性病原菌感染的选择不足 高发病率和高死亡率及其伴随的治疗费用。这项提案的目标是 是开发RND系列外排泵的一系列吡喃并吡啶(PyPy)抑制剂，这是 将革兰氏阴性病原体的多药耐药表型转化为由优化的外排泵组成的辅助治疗 抑制剂(EPI)和米诺环素(MIN)。辅助疗法将用于治疗由血液引起的感染。 由肠杆菌科多药耐药(MDR)病原体，包括被指定为紧急 和疾病预防控制中心的严重威胁，即碳青霉烯耐药和超广谱β-内酰胺酶 产生肠杆菌科细菌。PyPy Epis是主要的RND外排泵(AcrB)的有效抑制剂。 大肠杆菌和其他肠杆菌科细菌，它们从周质中分泌出不同类别的抗生素。 空间到单元格的外部。RND外排泵的过度表达在MDR中起重要作用 相当一部分临床分离株的表型，这种表型可以被PyPy Epis逆转。系统的优化设计 通过SAR驱动和基于结构的药物设计，PyPy系列产生了类似物，增强了 几种AcrB底物抗生素在亚微摩尔浓度下的体外抗菌活性。 我们目前的铅，MBX-4191，可溶于水溶液(≥100微米)，显示出有限的细胞毒性(CC5047 µM)，以及良好的体外ADME图谱。MBX-4191在小鼠中达到高水平暴露(AUC=26,000 HR*ng/mL，静脉注射10 mg/kg)，在单次静脉给药(MTD≥200 mg/kg)后耐受性良好， 多次静脉注射(50 mg/kg，2次，4d)。MBX-4191在4微克/毫升时解救MIN活性，即断点 在体外试验中，对耐MIN的大肠埃希菌和肺炎克雷伯菌的浓度。值得注意的是，MBX- 4191挽救MIN(50毫克/公斤，BID)对耐MIN的Kpc+肺炎克雷伯氏菌株的活性 (最小抑菌浓度=32微克/毫升)在小鼠败血症感染模型中。在这个项目中，我们将从化学上优化 吡喃吡啶类EPI先导序列在SAR和结构驱动的理性药物发现工作中的重点 改善小鼠的类药物特性、药代动力学和疗效，同时保持多年的效力 1和2.在第二年，我们将为初始IND启用(GLP)选择一名临床前候选人和一名后备候选人 大鼠药代动力学、毒理学和安全性药理学研究。为了实现临床前研究，我们将启动 制造非GMP批次(1公斤)的药品。 为实现拟议项目的目标，我们将完成以下具体目标：目标1.优化领导 系列通过合成孔径雷达驱动的模拟生成。(1-2年级)。目标2：通过体外实验优先考虑铅系列类似物 生物学和ADME评估(1-2年级)。目标3：选择临床前候选对象并根据体内情况进行支持 物业(1-2年级)。目标4：开展支持IND的药代动力学、毒理学和安全药理学 学习(三年级)。目标5：化学、制造和控制。(二至三年级)。

ABSTRACT The inadequacy of current treatment options to cure infections caused by MDR Gram-negative pathogens results in high rates of morbidity and mortality along with their concomitant treatment costs. The goal of this proposal is to develop a series of pyranopyridine (PyPy) inhibitors of RND family efflux pumps, a major component of the MDR phenotype of Gram-negative pathogens, into an adjunctive therapy consisting of an optimized efflux pump inhibitor (EPI) and minocycline (MIN). The adjunctive therapy will be used to treat bloodstream infections caused by multidrug resistant (MDR) pathogens of the Enterobacteriaceae, including organisms designated as urgent and serious threats by the CDC, namely Carbapenem-Resistant and Extended Spectrum Beta Lactamase producing Enterobacteriaceae. PyPy EPIs are potent inhibitors of the major RND efflux pump (AcrB) in Escherichia coli and other Enterobacteriaceae, which extrude diverse classes of antibiotics from the periplasmic space to the exterior of the cell. Overexpression of RND efflux pumps plays an important factor in the MDR phenotype of a significant fraction of clinical isolates, which can be reversed by PyPy EPIs. Optimization of the PyPy series through SAR-driven and structure-based drug design has generated analogs that potentiate the antibacterial activity of several antibiotics which are substrates of AcrB at submicromolar concentrations in vitro. Our current lead, MBX-4191, is soluble in aqueous solutions (≥ 100 µM), exhibits limited cytotoxicity (CC50 47 µM), and a favorable in vitro ADME profile. MBX-4191 achieves high levels of exposure in mice (AUC = 26,000 hr*ng/mL at 10 mg/kg IV) and is well-tolerated after a single intravenous dose (MTD ≥200 mg/kg), and after multiple IV doses (50 mg/kg, bid, 4d). MBX-4191 rescues the activity of MIN at 4 µg/ml, the breakpoint concentration, against MIN-resistant strains of E. coli and K. pneumoniae in in vitro assays. Significantly, MBX- 4191 rescues the activity of MIN (50 mg/kg, BID) against a MIN-resistant, KPC+ strain of Klebsiella pneumoniae (MIN MIC = 32 µg/ml) in a murine sepsis model of infection. In this project, we will chemically optimize the pyranopyridine EPI lead series in an SAR and structure-driven rational drug discovery effort focused on improving drug-like properties, pharmacokinetics, and efficacy in mice, while maintaining potency during years 1 and 2. In year 2, we will select a preclinical candidate and a back-up candidate for initial IND-enabling (GLP) pharmacokinetic, toxicology and safety pharmacology studies in rats. To enable preclinical studies, we will initiate manufacturing of a non-GMP lot (1 kg) of drug product. To achieve the goal of the proposed project, we will complete the following specific aims: AIM 1. Optimize lead series through SAR-driven analog generation. (years 1-2). AIM 2: Prioritize lead series analogs through in vitro biology and ADME evaluations (years 1-2). AIM 3: Select a preclinical candidate and backup based on in vivo properties (years 1-2). AIM 4: Conduct IND-enabling pharmacokinetic, toxicology and safety pharmacology studies (year 3). AIM 5: Chemistry, Manufacturing and Controls. (years 2-3).