Membrane-active quinoline and quinazoline antibacterials that target Gram positive pathogens

摘要 由耐药的革兰氏+细菌引起的感染正在上升，可能是由于各种因素，包括较长的时间 住院时间、更频繁的侵入性手术和普遍的抗生素治疗。复利 问题是在许多革兰氏阳性病原体(MRSA、S. 表皮葡萄球菌、肠球菌和肺炎链球菌)。尽管抗生素管理和感染控制，新的 迫切需要针对这些革兰氏阳性病原体的药物。在筛选了~6万个预选的 通过化合物文库，我们获得了具有抗菌活性的4-羟基喹啉衍生物DNAC-2 抗MRSA和肠球菌。我们随后合成了3个系列的类似物，涉及50多个 化合物。第二系列中的两个类似物，JRS-3-56(化合物1)和JRS-4-32(化合物2)， 对耐甲氧西林金黄色葡萄球菌、表皮葡萄球菌、粪肠球菌和粪肠球菌均有杀灭作用，最低抑菌浓度(≤)为0.2μg/ml。 2具有较高的cLogP(分别为7.7和6.0)，预测水溶解度较差。转换为 将喹啉改为喹唑啉可提高cLogP(从6.06提高到5.08)，但导致MIC略有增加(0.25 至2微克/毫升)。在最新的系列中，我们在C-4上引入了羰基，在C-1上引入了C到N的取代 位置，产生具有低cLogP和非常低MIC(对于USA300为0.06微克/毫升)的化合物3和4， 伴随着一个更紧密的SAR。利用大分子合成分析，膜特异染料FM4- 和电子显微镜的研究，我们有证据表明，1和2靶向革兰氏+膜(3和4 也导致膜缺陷，如FM4-染料检测到的)，但不是革兰氏或真核膜， 从而暗示了某种程度的特异性。然而，这些化合物的确切目标可能有所不同 是否来自达托霉素，目前尚不清楚。在本申请中，我们试图定义这些组件的作用机制 喹啉/喹诺酮衍生物，并进一步探索控制体外和体内活性的SAR和 药物处置属性。相应地，我们有以下具体目标：1)设计和合成 喹啉/喹诺酮类衍生物，通过定义管理对主要革兰氏阳性病原体和 药物处置特性(最低抑菌浓度、溶解性、显性毒性和血清结合等)；2)阐明其作用机制 利用遗传、生物化学和生物物理手段研究喹啉/喹诺酮类化合物的作用；3) 将对候选化合物进行药物评价的药代动力学和疗效研究 确保安全性和选择性的处置性能，然后选择用于完全PK的“铅”化合物 用两种动物模型进行评价和疗效研究。这些研究的目标是确定“可用药” 膜活性化合物“具有广泛的革兰氏+活性。如果成功，我们相信这些化合物将 代表了一类新的膜活性化合物，在药物开发方面取得了重大进展。

Abstract Infections due to resistant Gram+ organisms are on the rise, likely due to a variety of factors including longer hospital stay, increased frequency of invasive procedures and pervasive antibiotic therapy. Compounding the problem is the emergence of multi-drug resistance (MDR) among many Gram+ pathogens (MRSA, S. epidermidis, Enterococcus and S. pneumoniae). Despite antibiotic stewardship and infection control, new agents against these Gram+ pathogens are urgently needed. After screening a ~60,000 preselected compound library, we obtained DNAC-2, a 4-hydroxyquinoline derivative, that exhibited antibacterial activities against MRSA and Enterococcus. We subsequently synthesized 3 series of analogues involving over 50 compounds. Two of these analogues in the 2th series, JRS-3-56 (compound 1) and JRS-4-32 (compound 2), were cidal against MRSA, S. epidermidis, E. faecalis and E. faecium, with MIC ≤0.2 μg/ml. However, both 1 and 2 have poor predicted aqueous solubility with high cLogP (7.7 and 6.0, respectively). Conversion of quinoline to quinazoline for 1 improved the cLogP (from 6.06 to 5.08) but led to a slight increase in MIC (0.25 to 2 µg/ml). In the latest series, we introduced a carbonyl group at C-4 and a C to N substitution at the C-1 position, yielding compounds 3 and 4 with low cLogPs and very low MIC (0.06 µg/ml for USA300), accompanied by a much tighter SAR. Using macromolecular synthesis assays, membrane-specific dye FM4- 64 and electron microscopy studies, we have evidence that 1 and 2 target the Gram+ membrane (3 and 4 also resulted in membrane defect as detected by the FM4-64 dye), but not Gram- or eukaryotic membrane, thus implying some degree of specificity. However, the exact target of these compounds which likely differs from daptomycin, is not known. In this application, we seek to define the mechanism of action of these quinoline/quinolone derivatives and further explore the SAR that governs in vitro and in vivo activities and drug disposition properties. Accordingly, we have the following specific aims: 1) design and synthesize quinoline/quinolone derivates by defining the SAR that governs activity against major Gram+ pathogens and drug disposition properties (MIC, solubility, overt toxicity and serum binding etc.); 2) delineate the mechanism of action of the quinoline/quinolone derivatives with genetic, biochemical and biophysical tools; 3) pharmacokinetic and efficacy studies where candidates compounds will be evaluated for their drug disposition properties to ensure safety and selectivity followed by selection of “lead” compounds for full PK evaluation and efficacy studies with two animal models. The goal of these studies is to identify “druggable membrane-active compounds” with broad Gram+ activity. If successful, we believe these compounds will represent a new class of membrane-active compounds that offer a significant advance in drug development.