Carbapenem Antibiotic Resistance in Enterobacteriaceae: Understanding Interactions of KPC Carbapenemases with Substrates and Inhibitors

肠杆菌科碳青霉烯类抗生素耐药性：了解 KPC 碳青霉烯酶与底物和抑制剂的相互作用

• 批准号: MR/T016035/1
• 负责人: James Spencer
• 金额: $ 86.54万
• 依托单位: University of Bristol
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2020
• 资助国家: 英国
• 起止时间: 2020 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FT016035%2F1
• 关键词: Carbapenem; Antibiotic; Resistance; Enterobacteriaceae; Understanding

beta-lactams (BLs, penicillin and its relatives) are the most used antibiotics worldwide. Carbapenems are the newest and most potent BLs, and particularly important for treating infections by so-called opportunistic Gram-negative bacteria (GNB). These are organisms, either normally present in the human body or the natural environment (soil, water), that are not considered harmful to healthy individuals but can cause infections, possibly severe and even life-threatening, in patients whose immune defences are compromised. Risk factors for such infections include wounds (surgery, burns, injury), use of medical devices (catheters, ventilators), and conditions (HIV) or treatments (cancer chemotherapy or drugs that prevent transplant rejection) that affect immune defences. Growing numbers of patients fall into these categories. GNB are a particular treatment problem as their cell structure prevents many antibiotics that kill other types of bacteria from reaching their targets; efforts to discover new antibiotics effective against GNB have been largely unsuccessful.Until recently, carbapenems were regarded as "last resort" drugs for infections by GNB unresponsive to other treatments. However, growing resistance to other antibiotics makes carbapenems increasingly a first choice when infection by a GNB is suspected. When carbapenems fail alternatives are limited and often toxic, hence carbapenem resistance is regarded as a major public health challenge. In GNB carbapenem resistance is largely due to proteins called carbapenemases that bind to and degrade carbapenems, removing their ability to kill bacteria. Carbapenemases are part of a larger group of proteins (beta-lactamases) that destroy other types of BL antibiotics, however most beta-lactamases cannot break down carbapenems. beta-lactamases can be countered by a second group of drugs (beta-lactamase inhibitors) that block their activity and enable BL antibiotics to be used to treat bacteria carrying beta-lactamases, but not all beta-lactamases can be blocked by this route and some can mutate or evolve to escape the action of inhibitors.This proposal investigates how one carbapememase from the GNB Klebsiella pneumoniae, KPC (Klebsiella pneumoniae carbapenemase) degrades carbapenems and other BL antibiotics, and interacts with one specific class of inhibitors (diazabicyclooctanes, DBOs) and how these activities are affected by mutations in KPC. Klebsiella pneumoniae is an important cause of infections (urinary and respiratory infections, sepsis) associated with healthcare, and KPC is one of the main causes of carbapenem resistance worldwide.We recently described, for the first time, how KPC binds carbapenems and other BLs (ceftazidime, an antibiotic used for healthcare-associated infections) during a key stage in their breakdown; and how KPC binds DBOs. Based on this information we will use state-of-the-art computational methods to construct detailed models of the reaction of KPC with each of these three classes of molecules, in order to identify the most likely route by which each reaction occurs. The accuracy of these models will be tested by comparing the speed predicted for each reaction with actual values measured in experiments for a range of antibiotics used in patient treatment. We will then investigate how these reactions are affected by specific alterations in KPC, seeking to understand how such changes now identified in bacteria from human patients can improve the ability of KPC to break down antibiotics and reduce the ability of DBOs to block KPC action. Finally we will use this information to design and test, in computer models and experiments, new carbapenems that resist breakdown by KPC and new DBOs that are more effective KPC inhibitors; and that in each case are not affected by KPC mutations. This provides a route by which understanding of KPC can be exploited to design new treatments effective against an important group of antibiotic resistant bacteria.

β-内酰胺类抗生素（BLs，青霉素及其相关物）是全球使用最多的抗生素。碳青霉烯类是最新和最有效的BL，对于治疗所谓的机会性革兰氏阴性菌（GNB）感染特别重要。这些生物通常存在于人体或自然环境（土壤，水）中，被认为对健康个体无害，但可能导致感染，可能严重甚至危及生命，在免疫防御受损的患者中。此类感染的风险因素包括伤口（手术，烧伤，损伤），使用医疗器械（导管，呼吸器）和影响免疫防御的条件（HIV）或治疗（癌症化疗或预防移植排斥的药物）。越来越多的患者属于这些类别。GNB是一个特殊的治疗问题，因为它们的细胞结构阻止了许多杀死其他类型细菌的抗生素到达它们的目标;发现新的抗生素对GNB有效的努力在很大程度上是不成功的。直到最近，碳青霉烯类被认为是治疗对其他治疗无效的GNB感染的“最后手段”药物。然而，对其他抗生素的耐药性不断增加，使得碳青霉烯类抗生素越来越多地成为怀疑GNB感染时的首选。当碳青霉烯类失败时，替代品有限且通常有毒，因此碳青霉烯类耐药性被视为主要的公共卫生挑战。在GNB中，碳青霉烯类耐药性主要是由于称为碳青霉烯酶的蛋白质结合并降解碳青霉烯类，从而消除了它们杀死细菌的能力。碳青霉烯酶是破坏其他类型BL抗生素的较大蛋白质组（β-内酰胺酶）的一部分，但大多数β-内酰胺酶不能分解碳青霉烯。β-内酰胺酶可以被第二组药物所抵消（β-内酰胺酶抑制剂）阻断其活性并使BL抗生素能够用于治疗携带β-内酰胺酶的细菌，但并非所有的β-内酰胺酶都可以通过该途径阻断，并且一些β-内酰胺酶可以突变或进化以逃避抑制剂的作用。KPC（肺炎克雷伯菌碳青霉烯酶）降解碳青霉烯类和其他BL抗生素，并与一类特定的抑制剂（二氮杂双环辛烷，DBO）相互作用，以及KPC突变如何影响这些活性。肺炎克雷伯菌是引起医疗相关感染（泌尿道和呼吸道感染、败血症）的重要原因，而KPC是全球碳青霉烯类耐药的主要原因之一。我们最近首次描述了KPC如何在碳青霉烯类和其他BL（头孢他啶，一种用于医疗相关感染的抗生素）分解的关键阶段结合它们;以及KPC如何结合DBO。基于这些信息，我们将使用最先进的计算方法来构建KPC与这三类分子中的每一类反应的详细模型，以确定每个反应发生的最可能途径。这些模型的准确性将通过比较每个反应的预测速度与在患者治疗中使用的一系列抗生素的实验中测量的实际值来测试。然后，我们将研究这些反应如何受到KPC特定改变的影响，试图了解现在在人类患者细菌中发现的这些变化如何提高KPC分解抗生素的能力并降低DBO阻断KPC作用的能力。最后，我们将使用这些信息来设计和测试，在计算机模型和实验中，新的碳青霉烯类，抵抗由KPC和新的DBO，更有效的KPC抑制剂的故障;在每种情况下，不受KPC突变的影响。这提供了一种途径，通过这种途径，可以利用对KPC的理解来设计有效对抗一组重要的抗生素耐药细菌的新治疗方法。

项目成果

A multiscale approach to predict the binding mode of metallo beta-lactamase inhibitors.

• DOI: 10.1002/prot.26227
• 发表时间: 2022-03
• 期刊: Proteins
• 影响因子: 2.9
• 作者: Gervasoni S;Spencer J;Hinchliffe P;Pedretti A;Vairoletti F;Mahler G;Mulholland AJ
• 通讯作者: Mulholland AJ

QM/MM Simulations Reveal the Determinants of Carbapenemase Activity in Class A ß-lactamases

QM/MM 模拟揭示了 A 类 - 内酰胺酶中碳青霉烯酶活性的决定因素

• DOI: 10.26434/chemrxiv-2022-4jdc5
• 发表时间: 2022
• 作者: Chudyk E