High-throughput development of bacterial PROTACs

抗菌素耐药性（AMR）是对全球健康的最严重威胁之一，可能使手术和化疗等挽救生命的医学进步变得如此危险，以至于不切实际。如果抗菌素耐药性影响医疗程序并缩短预期寿命，即使是神经变性等老年疾病也可能成为不那么重要的疾病。抗生素耐药性的挑战由于缺乏新的抗生素渠道而加剧，人们普遍认识到迫切需要具有新的作用机制的新分子。双功能分子对人类疾病的研究兴趣激增；然而，缺乏在抗菌素耐药性中的应用。靶向蛋白水解嵌合体（Proteolysis-targeting chimeras, PROTACs）将靶蛋白招募到蛋白酶体中进行降解。与传统抑制剂相比，PROTACs具有优势：活性不需要完全占据靶标，这意味着一个双功能分子可以降解多个靶标蛋白分子；由于需要靶合成来恢复功能，可以实现较长的作用时间。我们最近首次证明大肠杆菌周围质蛋白酶DegP可以降解功能性β -内酰胺酶TEM116，并合成了DegP和β -内酰胺酶的配体。此外，我们还建立了PROTAC自动化合成平台，快速生成细菌PROTAC以探索结构-活性关系。因此，本项目的目的是开发抗生素抗性酶的细菌PROTAC降解物。这将通过以下目标来实现：DegP和β -内酰胺酶配体的SAR优化细菌PROTACs连接体长度对细菌裂解物中TEM116降解的影响，3。优化细胞渗透，分析其他β -内酰胺酶的降解，以及证明TEM116在完整细菌中的降解。该项目的潜在应用和好处是广泛的。首先，这将提供一系列具有新的作用机制的分子，使耐药细菌对β -内酰胺类抗生素敏感，为继续翻译做好准备。其次，这开辟了一个新的研究领域，现有的抗生素可能转化为细菌PROTACs，与新的或现有的抗生素合作以延长寿命，并且与耐药酶抑制剂相比，目标降解可能产生增强的效果。第三，高通量PROTAC生成的自动化合成平台可能会加速其他疾病PROTAC的开发。

Antimicrobial resistance (AMR) is one of the most serious threats to global health, potentially making life-saving medical advances such as surgery and chemotherapy so dangerous as to be impractical. Even diseases of old age such as neurodegeneration may become a lessening priority if AMR compromises medical procedures and curtails life expectancy. The challenge of AMR is exacerbated by a scarce pipeline of new antibiotics and it is widely appreciated that new molecules with novel mechanisms of action are urgently needed.Bifunctional molecules have seen an explosion of research interest in human diseases; however, use in AMR is lacking. Proteolysis-targeting chimeras (PROTACs) recruit target proteins to the proteasome machinery for degradation. PROTACs have advantages over traditional inhibitors: activity does not require complete target occupancy meaning one bifunctional molecule can degrade multiple target protein molecules; and a longer duration of action can be achieved as target synthesis is required to recover function. We have recently shown for the first time that the E. coli periplasmic protease DegP can degrade functional beta-lactamase enzyme TEM116, and have synthesised ligands for both DegP and beta-lactamases. In addition, we have established a platform for automated PROTAC synthesis to rapidly generate bacterial PROTACs to explore structure-activity relationships.The aim of this project is therefore to develop bacterial PROTAC degraders of antibiotic resistance enzymes. This will be achieved through the following objectives:1. SAR optimisation of DegP and bet-lactamase ligands2. Modification of bacterial PROTACs linker lengths for degradation of TEM116 in bacterial lysates,3. Optimisation of cell penetration, profiling for degradation of other beta-lactamases, and demonstration of proof-of-concept degradation of TEM116 in intact bacteriaThe potential applications and benefits of this project are broad. Firstly, this will provide a new series of molecules with a novel mechanism of action to sensitize resistant bacteria to beta-lactam antibiotics, primed for continued translation. Secondly, this opens a new area of research where existing antibiotics may be converted to bacterial PROTACs to partner with new or existing antibiotics to prolong lifetime, and where target degradation may produce enhanced effects compared to resistance enzyme inhibitors. Thirdly, the automated synthesis platform for high-throughput PROTAC generation may accelerate development of PROTACs for other diseases.