Novel antibiotics targeting bacterial gene expression

Lead Research Organisation: Newcastle University
Department Name: Inst for Cell and Molecular Biosciences

Abstract

The threat of multi-drug-resistant infections is now the most serious since the golden era of antibiotic discovery. The rate of resistant strains appearance has been increasing in the last few decades putting us in an alarming situation of fighting against bugs. There is a vital need for new antibiotics with new modes of action and binding sites to avoid cross resistance.
RNA polymerase is the enzyme that performs transcription, the first step of gene expression, and is a very attractive target for antibiotic discovery. The only two antibiotics in clinical use targeting RNA polymerase are Rifamycins and Fidaxomycin, the latter approved only a few years ago. Rifampicin remains a first-line antibiotic against lethal infections like tuberculosis, though, rifampicin resistance is growing. However, the discovery of more inhibitors of RNA polymerase depicts its potential as a target for antibiotic discovery. In addition to this clear clinical importance, inhibitors of transcription have also proved to be powerful tools for investigation of the molecular details of the process. Therefore, analysis of the mode of action of new inhibitors will have a strong fundamental aspect.
To discover new antibiotic leads targeting bacterial transcription, we are going to combine our world-leading expertise in studying inhibitors of transcription with a unique library of novel potential antibiotics developed by the collaborating company. The study is directly connected to areas of antibiotic resistance and public health, satisfying the mission of MRC: "to improve human health through world-class medical research". It lies perfectly within the science areas of Molecular and Cellular Medicine and Infections and Immunity Research Boards, and thus is well within the MRC's remit.
We have a proven track record for uncovering the mode of action of RNA polymerase inhibitors, including sorangicin, microcin J25, streptolydigin and tagetitoxin, which allowed us to discover new functions of RNA polymerases. These projects led to a collaboration with a local biotech company who possess a library of about 10,000 highly diverse and well dereplicated actinomycete bacteria, which are prolific producers of antibiotics. They also have equipment and expertise in screening, purifying and analysing novel antibiotics targeting transcription. In a pilot study we have already found a new molecule that is able to kill rifampicin-resistant bacteria and inhibit rifampicin-resistant RNA polymerases.
This project will focus on investigation of inhibitors of RNAP that presently have unknown modes of action. The central goals will be (i) to understand the mechanisms of inhibition of transcription by these compounds; (ii) search for new inhibitors of transcription. By better understanding both the enzyme and the mode of action of the inhibitors, we hope to identify molecules with potential therapeutic value and to facilitate the development of the new molecules as potential antibiotics and/or antifungals. Our collaborators can also provide a future route to market for inhibitors that turn out to have commercial potential.
The work will involves a wide range of biochemical, molecular biological and microbiological techniques provided by both academic and commercial collaborators, as well as providing experience of biotech company environment. The academic supervisor's laboratory is based in the Centre for Bacterial Cell Biology, which brings together leading scientists in the field of bacterial cell biology, and is fitted with state of the art equipment. Commercial partner Demuris is a biotech SME, which is well set up for antibiotic discovery, with excellent facilities for large scale fermentation, natural product purification and chemical characterization. The company will also provide experience of a business environment, especially the operational procedures and commercial discipline that distinguish the company from the academic lab.

Publications

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Title Novel In vitro transcription assays 
Description We purify the transcription apparatus from the bacteria to then use in an in vitro system in which we dissect the mechanism of drug action by using unique DNA templates and RNAP constructs that resemble states of the transcription apparatus at different point in the transcription cycle. 
Type Of Material Model of mechanisms or symptoms - in vitro 
Year Produced 2018 
Provided To Others? Yes  
Impact It has allowed elucidation of the mechanism of action of ureidothiophene that will hopefully be published in 2019 
 
Description Structural basis of inhibitor mechanism of action 
Organisation University of Pennsylvania
Department Department of Cell and Developmental Biology
Country United States 
Sector Academic/University 
PI Contribution We deduced the biochemical basis of transcription inhibition by the antibiotic kanglemycin A and ureidothiophene
Collaborator Contribution Our collaborators in the Lab of Katsuhiko Murakami in Penn state university, USA deduced the structural basis of Kanglemycin A inhibition at both E.coli and T.thermophillus RNA Polymerase. They are presently attempting to elucidate the structural basis of ureidothiophene inhibition at RNAP.
Impact We published our communal work in a molecular cell publication in 2018. (see publications). Our collaborators used x-ray crystallography to discern the structural basis for KglA inhibition.
Start Year 2018
 
Description Upstream promoter elements of genes unregulated in Streptococcus in response to sub-inhibitory concentrations of rifampicin. 
Organisation University of Lausanne
Country Switzerland 
Sector Academic/University 
PI Contribution I have recently begun a collaboration with Jan-Willem Veening's lab of the University of Lausanne investigating the role of upstream promoter elements of genes upregulated in Streptococcus in response to sub-inhibitory concentrations of rifampicin.
Collaborator Contribution -
Impact No data has yet been published. This collaboration is in its infancy
Start Year 2019
 
Description RNAP Workshop 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact I have gave 2 poster presentation on my work at the UK RNAP workshop. I will be giving an oral presenation in 2019
Year(s) Of Engagement Activity 2017,2018,2019
 
Description School Visit (organised for late 2019) 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact I am to hold a talk at Durham school for there science society in which i will talk about our publication in molecular cell and my own carer in science
Year(s) Of Engagement Activity 2019