替加环素联合氨基糖苷类抑制耐碳青霉烯肺炎克雷伯菌耐药发生的作用及机制研究

Carbapenem-resistant Klebsiella pneumoniae (CR-KP) has become an important pathogenic bacteria of nosocomial infections. It is resistant to nearly all currently used antimicrobial agents in clinical practice, and the mortality rate of CR-KP infections is over 40%. Tigecycline is the "last resort" for the treatment of CR-KP infections, but the problem of rapid in vivo emergence of resistance during treatment is quite worthy of attention. Therefore, it is extremely important to adopt effective strategies to inhibit the further occurrence of tigecycline resistance. Our previous in vitro studies have found that tigecycline combined with aminoglycosides can not only significantly improve the antibacterial activity, but also significantly reduce or even close the drug-resistant mutant selection window, thus inhibiting the emergence of resistance. However, whether this combination can effectively inhibit the emergence of tigecycline resistance in vivo, and the mechanisms by which this combination inhibits the development of resistance are not clear. This study will combine antimicrobial pharmacokinetic/pharmacodynamic theory with drug-resistant mutant selection window theory, and establish a rat tissue-cage infection model to evaluate the ability of combination therapy to restrict the emergence of tigecycline resistance in vivo. This study will also evaluate the effects of combination therapy on the function of efflux pumps and on the genes which encodes and regulates the expression of efflux pumps, and analyze of the relationship between drug mutations in the different drug sites and the inhibition of drug resistance in combination, trying to illuminate the possible mechanisms of combination therapy on restricting the emergence of resistance in CR-KP. It provides a valuable theoretical basis for guiding clinical use of tigecycline combination therapy for treating CR-KP infections and preventing further resistance emergence.

耐碳青霉烯肺炎克雷伯菌（CR-KP）已成为医院内感染的重要病原菌，对绝大部分抗菌药物耐药，感染病死率超过40%。替加环素为治疗CR-KP感染的“最后防线”，但临床使用后体内很快出现耐药的问题值得关注，故采取有效策略抑制耐药发生有极为重要的意义。我们前期体外研究发现替加环素联合氨基糖苷类不仅能明显提高抗菌活性，还可显著缩小甚至关闭耐药突变选择窗，抑制耐药发生，但联合用药能否有效抑制体内耐药发生及该组合抑制耐药发生的机制尚不明确。本研究拟将药动学/药效学理论与耐药突变选择窗理论相结合，建立大鼠组织笼感染模型，评价联合用药抑制替加环素体内耐药发生的能力；研究联合用药对耐药相关的细菌外排泵功能及其编码、调控基因表达的影响，并分析药物作用位点基因突变与联合用药抑制耐药发生的关系，以此探讨联合用药抑制替加环素耐药发生的可能机制，为指导临床治疗CR-KP感染和预防替加环素耐药发生提供有价值的理论依据。

耐碳青霉烯肺炎克雷伯菌（CR-KP）已成为医院内感染的重要病原菌，对绝大部分抗菌药物耐药，感染病死率超过40%。替加环素为治疗CR-KP感染的“最后防线”之一，但临床使用后体内很快出现耐药的问题值得关注，故采取有效策略抑制耐药发生有极为重要的意义。.本项目在前期体外研究发现替加环素联合氨基糖苷类明显提高抗菌活性，并且显著缩小甚至关闭耐药突变选择窗，抑制耐药发生的基础上，进一步探究联合用药的体内外抗菌活性及抑制耐药发生能力，并探究其潜在机制。.研究发现：①替加环素联合氨基糖苷类药物对CR-KP具有体内外协同杀菌作用，并有助于抑制体内外的耐药发生；②蛋白质组学研究表明，CR-KP对替加环素与氨基糖苷类药物具有不同的蛋白应答反应，联合用药时细菌蛋白变化与替加环素单药类似。联合用药可能通过降低CR-KP适应性调节能力发挥协同作用；③氨基糖苷类与四环素类耐药菌株之间存在“侧枝敏感性”，其中CR-KP对氨基糖苷类的适应性耐药涉及细胞膜电位变化，通过影响外排泵功能增加对四环素类药物的敏感性。CR-KP对四环素类的耐药机制主要涉及AcrAB外排泵，而四环素耐药菌株中不平衡的氧化还原过程可能增加细菌对氨基糖苷类药物敏感性。体内外研究证实基于侧枝敏感性的该药物组合可有效抑制CR-KP耐药进化，此外抗生素轮替治疗可使CR-KP对先前耐药的抗菌药物重新敏感。.这些研究结果为指导临床治疗CR-KP感染和预防替加环素耐药发生提供了有价值的理论依据。