DNA GYRASE AND QUINOLONE RESISTANCE IN TUBERCULOSIS

描述(改编自申请人的摘要)：抗药性 是抗结核化疗的一个主要问题，也是 氟喹诺酮类化合物不是第一线的主要原因 探员们。使用各种细菌的两项最新进展鼓励 氟喹诺酮类药物的精制。首先，阻力是逐步产生的 DNA旋转酶和DNA拓扑异构酶两个独立靶点的突变 因此，可以利用第一步抗性旋转酶突变体来筛选新的 氟喹诺酮类药物需要两个突变才能产生 抵抗。由于双突变发生频率低于单突变 多个数量级的突变，对新化合物的抗药性 应该很少出现。第二，双重突变的概念是 实验验证：C8-甲氧基减少了获得 阻力至少增加了三个数量级。有两种方法 建议界定这些原则如何适用于氟喹诺酮类药物 结核分枝杆菌的耐药性。一种是氟喹诺酮类药物 将检查结构变体是否有能力防止 抵抗。抗药性的预防将与贪婪相关 生长条件下纯培养中抗病菌株的杀灭 以及2)在培养的人巨噬细胞内。 结论的一般性将用各种各样的方法进行检验。 结核分枝杆菌临床分离株。在另一种方法中， 将通过检查基因的核苷酸序列来研究抗药性 编码旋转酶和拓扑异构酶IV。 对各种氟喹诺酮同系物的耐药性，其中一些是 已经知道会引发不同的等位基因。选定的突变体将是 测试1)交叉抗性，2)适应变化的环境， (3)人巨噬细胞侵袭。已经推出了许多新的 化合物，两种新的检测方法(抗病突变体和预防 抗药性)，人类巨噬细胞感染系统，以及一个大型 收集结核分枝杆菌临床分离株。意义重大 在寻找氟喹诺酮类药物方面有望取得进展，以防止 结核分枝杆菌产生的耐药性。

DESCRIPTION (Adapted from the applicant's abstract): Drug resistance is a major problem for anti-tuberculous chemotherapy and is one of the main reasons why compounds such as fluoroquinolones are not first line agents. Two recent advances, using a variety of bacteria, encourage refinements of fluoroquinolones. First, resistance arises stepwise from mutations in two independent targets, DNA gyrase and DNA topoisomerase IV. Thus a first step resistant gyrase mutant can be used to screen new fluoroquinolones for derivatives that require two mutations to confer resistance. Since double mutations occur less frequently that single mutations by many orders of magnitude, resistance to the new compounds should arise very rarely. Second, the double mutant concept was experimentally validated: a C8-methoxyl group reduced the acquisition of resistance by at least three orders of magnitude. Two approaches are proposed to define how these principles apply to fluoroquinolone resistance in Mycobacterium tuberculosis. In one, fluoroquinolone structural variants will be examined for their ability to prevent resistance. Prevention of resistance will be correlated with avid killing of moderately resistant strains 1) in pure culture under growing and non-growing conditions, and 2) inside cultured human macrophages. The generality of the conclusions will be tested using a wide variety of clinical isolates of M. tuberculosis. In the other approach, resistance will be studied by examining nucleotide sequences of genes encoding gyrase and topoisomerase IV. Mutants will be selected for resistance to various fluoroquinolone congeners, some of which are already known to elicit different alleles. Selected mutants will be tested for 1)cross-resistance, 2) adaptation to changing environment, and 3) invasion of human macrophages. Already available are many new compounds, two novel assays (killing resistant mutants and prevention of resistance), a human macrophage infection system, and a large collection of clinical isolates of M. tuberculosis. Significant progress is expected toward finding fluoroquinolones that will prevent resistance from arising in M. tuberculosis.