Title: Defining the chemical space for small molecule accumulation in bacteria
Funder: The Academy of Medical Sciences

Abstract: The aim of this Springboard Award is to develop a high-throughput assay to measure accumulation of small molecules in bacteria and screen for molecular characteristics that lead to a compound preferentially accumulating in a bacterial cell. This information will rationally guide development of DNA repair inhibitors that can penetrate challenging Gram-negative bacteria as part of my on-going research vision, whilst also accelerating future antibiotic discovery efforts. This aim will be achieved by a PDRA employed to conduct the following objectives: O1. Establish assay to measure cellular accumulation of compounds O2. Optimisation to high-throughput screening-compatible format O3. Screening of compound libraries to determine cell-penetrant chemical space These objectives will be met by the following steps: O1. An initial LC-MS assay protocol will be established by the PDRA in the Lanyon-Hogg lab based on literature methodology for compound accumulation in bacterial cells, with analysis performed in the McCullagh lab. Compounds tested will include known accumulators (e.g. siderophores) and putative accumulators (e.g. siderophore-DNA repair inhibitor conjugates). O2. The LC-MS accumulation assay will be adapted to a high-throughput screening format using 384-well filter plates that allow separation of cells and washing on a vacuum manifold to facilitate analysis. This key objective is built on my extensive experience in successfully developing novel assay formats for screening. O3. A screen of ~4,000 compounds representative of the chemical space of >140,000 compound library will be conducted. Identification of the chemical space that is preferentially accumulated will guide synthesis of DNA-repair inhibitors or new antibiotic molecules.

Lay abstract: Antimicrobial resistance (AMR) is one of the most serious threats to health globally, potentially making life-saving medical advances such as surgery or chemotherapy so dangerous as to be impractical. AMR has the potential to cause >10m deaths/year by 2050 and an economic toll >$100 trillion, making it arguably the greatest economic and healthcare issue post-COVID-19. Indeed, the COVID-19 pandemic should have taught us that we need to act pre-emptively to address these emerging threats. To address this challenge, I have focused on identifying new molecules which can be administered alongside existing antibiotics to overcome the challenges of AMR. These new molecules inhibit the bacteria's ability to repair damage to its DNA and can make resistant bacteria sensitive to antibiotics, as well as decreasing the rate of resistance emergence from mutations during DNA repair. These molecules have proven highly promising in bacteria such as MRSA, but in other bacteria such as E. coli they show no effect. This is because before a molecule can block DNA repair, it actually has to get inside the bacterial cell. This Springboard Award will develop a method to screen thousands of molecules in order to identify patterns in the type of molecules that bacterial cells like to let in. These patterns of molecular characteristics can then be used to design new DNA-repair inhibitors that are preferentially let into bacterial cells, such as E. coli. This will also provide a method for studying other bacteria and accelerate the design of new cell-penetrant antibiotics in future.

Research activity codes: 2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT;5.1 PHARMACEUTICALS
Health categories: INFECTION