Dealing with antibiotic resistance: antisense technology

项目摘要/摘要 细菌感染是导致死亡、危害健康和残疾的主要原因。不幸的是，我们是 目前见证了多重耐药感染的增加和新的 抗菌剂。因此，治疗成本在增加，越来越多的患者 屈从于这些感染。此外，难以治疗甚至无法治疗的细菌的增加也 妥协医疗程序，如癌症和其他慢性病的治疗，外科手术，器官 移植、牙科治疗和早产儿护理。雪上加霜的是，由于SARS-CoV-2病毒 与多重耐药细菌的混合感染已经被记录在案，新冠肺炎大流行可能 通过增加患者对抗菌素的暴露来加速抗生素耐药性的上升。一种解决方案 抗生素耐药性问题可能是新的抗菌药类别不断发展所致。然而， 这条路线缓慢且成本高昂，需要与其他战略相补充。这项提议回应了 这就需要并集中于寻找延长现有药物使用寿命的策略。这个 氨基糖苷6‘-N-乙酰转移酶Ib型[AAc(6’)-Ib]是大多数耐药病例的原因 革兰氏阴性杆菌中的阿米卡星(AK)和其他氨基糖苷类。AAC(6‘)-Ib的传播 这些病原体之间的基因会侵蚀这些抗生素的疗效，这些抗生素是 预防严重感染的医疗设备。这项研究的长期目标是开发化合物 这降低了AK对敏感性水平的耐药性，并可用作治疗耐AK感染的佐剂。 本项目的具体目标1提出优化结合到细胞穿透肽(CPP)的结构 寡核苷酸类似物，称为外部引导序列(ESS)，它结合了 Aac(6‘)-Ib mRNA，并形成RNaseP的底物，它切割阻止翻译的mRNA。这个 计划中的实验包括设计最大限度地内化和测试的抗蛋白酶CPP 由脱氧核糖核苷酸和最新一代桥联核酸组成的嵌合低聚物。 特定目标2将利用组合文库鉴定AAC(6‘)-Ib的小分子抑制剂并优化 对它们进行构效关系分析。这一具体目的还提出了设计水溶性 与锌离子形成络合物的离子载体是AK酶失活的强烈抑制物。特定目标 3包括测试AK与下列不同化合物的组合相关的效果 以往针对大肠埃希菌、肺炎克雷伯菌和鲍曼不动杆菌模型的研究 使用三维棋盘试验、时间消磨试验和治疗感染的菌株 梅隆格氏杆菌感染模型。最有希望的组合将在大约100个品种上进行测试 基因分型明确的肺炎克雷伯菌、鲍曼氏杆菌、铜绿假单胞菌和阴沟肠杆菌 临床分离株。

Project Summary/Abstract Bacterial infections are a leading cause of death, compromised health, and disability. Unfortunately, we are currently witnessing an increase in multiresistant infections and a decrease in the development of new antimicrobials. Consequently, the treatment costs are increasing, and a growing number of patients are succumbing to these infections. Furthermore, the increase in hard-to-treat or even untreatable bacteria also compromises medical procedures such as treatment of cancer and other chronic diseases, surgery, organ transplants, dental work, and care for premature infants. Compounding the problem, since SARS-CoV-2 coinfection with multidrug resistant bacteria has already been documented, the COVID-19 pandemic could accelerate the rise in antibiotic resistance by increasing patient exposure to antimicrobials. A solution to the antibiotic resistance problem could be the continuous development of new classes of antimicrobials. However, this route is slow and costly and needs to be complemented with other strategies. This proposal responds to this need and concentrates on searching strategies to extend the useful life of currently available drugs. The aminoglycoside 6′-N-acetyltransferase type Ib [AAC(6′)-Ib] is responsible for most cases of resistance to amikacin (Ak) and other aminoglycosides in Gram-negative pathogens. The dissemination of the aac(6′)-Ib gene among these pathogens erodes the efficacy of these antibiotics, which are an important component of the armamentarium against severe infections. The long-term goal of this research is to develop compounds that reduce Ak resistance to susceptibility levels and can be used as adjuvants to treat Ak-resistant infections. Specific aim 1 of this project proposes to optimize the structure of cell-penetrating peptides (CPP) bound to oligonucleotide analogs, known as external guide sequences (EGSs), that bind a complementary region of the aac(6′)-Ib mRNA and form a substrate for RNase P, which cleaves the mRNA preventing translation. The planned experiments consist of designing protease-resistant CPPs that maximize internalization and testing chimeric oligomers composed of deoxyribonucleotides and the newest generation of bridge nucleic acids. Specific Aim 2 will identify small molecule inhibitors of the AAC(6′)-Ib using combinatorial libraries and optimize them by structure-activity relationship analysis. This Specific aim also proposes to design water-soluble ionophores that in complex with zinc ions are strong inhibitors of the enzymatic inactivation of Ak. Specific aim 3 consists of testing the effect of Ak in association with combinations of the different compounds identified in the previous specific aims on Escherichia coli, Klebsiella pneumoniae, and Acinetobacter baumannii model strains using three-dimensional checkerboard assays, time-kill assays, and treatment of infections in the Galleria mellonella infection model. The most promising combinations will then be tested on about 100 genotypically well-defined K. pneumoniae, A. baumannii, Pseudomonas aeruginosa, and Enterobacter cloacae clinical isolates.