Nutrient-Mediated Delivery of Antimicrobials

目前，抗菌素耐药性问题非常严重，以至于世界卫生组织（World Health organization）将其描述为一个至关重要的问题。一些权威人士认为，人类正在进入后抗生素时代。环丙沙星等氟喹诺酮类药物被广泛用于治疗一系列感染，然而，随着氟喹诺酮类药物使用的增加，几乎所有使用氟喹诺酮类药物治疗的细菌都出现了耐药性。挑战在于化学修饰FQs以增加细胞内浓度并克服渗透性和外排介导的阻力。我们已经探索了利用细菌营养转运将抗生素输送到细菌细胞（特洛伊木马方法）。到目前为止，我们已经研究了铁载体（铁载体-柠檬酸盐，葡萄铁蛋白A）和碳水化合物通过非生物连接连接到环丙沙星的哌嗪基氮上。针对我们已建立的细菌菌株面板的筛选证实，环丙沙星，当结合时，保留抗菌活性，但其效力降低。随后的DNA旋切酶抑制试验显示，活性下降主要是由于环丙沙星缀合物抑制旋切酶（药物的靶标）的能力受损，而不是细菌摄取不足。我们认为共轭部分的存在限制了与DNA的结合。这表明，“前药物方法”，即营养物质被切割，与细胞内氟喹诺酮释放将改善项目目标：本研究的主要目的是通过细胞内，还原激活，生物稳定的连接物将FQ与关键营养物质结合，以逃避FQ渗透性阻力并增加细胞内积累。研究流程为：(1)合成一种环丙沙星偶联物。连接体的设计是这样的，在主动运输后，细胞内还原（通过细胞内含巯基的物质，如谷胱甘肽）将在细菌细胞质中释放游离环丙沙星。最初，缀合物将使用附着在FQ COOH上的柠檬酸盐单元（我们的模型铁载体）。该结合物将对野生型大肠杆菌进行筛选。获得的最低抑菌浓度/最低杀菌浓度将与母体药物环丙沙星和具有不稳定连接剂的相应类似物的浓度进行比较。(2)我们提出探索的铁载体成分是受到最近才发现的一类新的铁载体盐螯素的结构的启发。这些糖基化的铁载体的主要优点是它们能够逃避哺乳动物宿主的免疫反应。此外，它们不易受血清白蛋白结合和膜分配的影响，这两个因素都会降低血清中常规铁载体的浓度。

BackgroundThe problem of antimicrobial resistance is now so severe that it has been described as critical by the World Health Organisation and some authorities believe that mankind is now entering a post-antibiotic era. Fluoroquinolones (FQ) such as ciprofloxacin are widely used for the treatment of a range of infections, however, with their increased use the incidence of resistance in almost all bacteria that fluoroquinolones are used against has emerged.The challenge is to chemically modify FQs to increase intracellular concentrations and overcome permeability and efflux mediated resistance.We have explored the use of bacterial nutrient transport to deliver antibiotics into the bacterial cell (Trojan horse approach). To date we have investigated iron carriers (siderophores-citrate, Staphyloferrin A) and carbohydrates attached to the piperazinyl nitrogen of ciprofloxacin via a non-biolabile linker. Screens against our established panel of bacterial strains confirmed that ciprofloxacin, when conjugated, retains antibacterial activity, however, its potency is reduced. Subsequent DNA gyrase inhibition assays revealed that the decrease in activity is mainly due to an impaired ability of the ciprofloxacin conjugate to inhibit gyrase, the target of the drug, rather than lack of bacterial uptake. We believe that the presence of the conjugated moiety limit binding to DNA gyrase5-7. This suggests that a 'prodrug approach', were the nutrient is cleaved, with intracellular release of the fluoroquinolone will improve Project Aims: The primary aim of this research is to conjugate FQs to key nutrients via an intracellular, reductively activated, biolabile linker in order to evade FQ permeability resistance and increase intracellular accumulation. The research workflow is: (1) To synthesise a ciprofloxacin conjugates with a self immolative linker. The design of the linker is such that, after active transport, intracellular reduction (via intracellular sulfhydryl-containing species such as glutathione) will release free ciprofloxacin in the bacterial cytoplasm. Initially the conjugate will use a citrate unit (our model siderophore) attached to FQ COOH. The conjugate will be screened against wild-type E. coli. The minimum inhibitory concentrations/minimum bactericidal concentrations obtained will be compared to those of the parent drug ciprofloxacin and a corresponding analog with non-labile linker. (2) The siderophore components that we propose to explore are inspired by the structure of the salmochelins, a new class of siderophores that has only recently discovered. The main advantage of these glycosylated siderophores is that they are able to evade the immune response of the mammalian host. In addition, they are less susceptible to serum albumin binding and membrane partitioning, both factors that reduce the concentration of conventional siderophores in the serum.