Public Funding for Demuris Limited

As the death toll from the COVID-19 pandemic passes 1 million and healthcare systems around the globe are struggling to cope there is another problem on the horizon, of Antimicrobial Resistance (AMR). The COVID-19 pandemic has increased the usage of antibiotics due to being administered to patients for the prevention of secondary bacterial infections. The pandemic has highlighted how unprepared the world is for significant healthcare disruption and the need for investment in medicines to combat these disruptions. AMR was already a global problem, each year the US Centers for Disease Control and Prevention estimates that 2.8 million Americans acquire serious infections caused by antibiotic-resistant bacteria, and 35,000 of them die as a result. AMR is a growing problem with vast societal and economic consequences. Pathogenic bacteria are becoming increasingly resistant to antibiotics and some strains are now resistant to all clinically used antibiotics. At the same time, the rate of discovery and deployment of new antibiotics has declined. Unfortunately, most major pharmaceutical companies have curtailed their antimicrobial and anti-infective discovery programs in favour of more profitable, lower-risk programs. Demuris is a SME that through screening a large collection of actinomycete bacteria has identified a novel ansamycin antibiotic. It has good potency towards multi-drug-resistant Gram-negative bacteria ,which are greatest threat to the public. This project will enable the production of the compound at a scale which will enable the evaluation of any human toxicity and the potency towards a panel of hospital pathogens.

Mycobacterium tuberculosis (M.tb) is the causative agent of tuberculosis (TB). The World Health Organisation (WHO) estimates that someone dies from TB every 20 seconds, which is around 1.7 million people per year. There is a growing emergence of M.tb strains which are resistant to first and second line antibiotic treatments. The bacterium can hide in the body remaining dormant for long periods of time, and treatments can last up to one year. New antibiotics that are effective against drug resistant strains are urgently needed. Demuris, in collaboration with Newcastle University, has identified novel antibiotics that are similar to the first-line treatment for TB but, crucially, are active against multi-drug resistant strains. This project will allow us to make a series of closely related compounds, and test these for the range of properties needed for development as an anti-TB drug.

New antibiotics are urgently needed to combat the rise of Multi-Drug Resistance (MDR), and by far the most pressing is the identification of compounds to inhibit MDR-Gram-negative pathogens, where the treatment options for several pathogens have become severely limited. Demuris is an SME with a world-leading collection of actinomycete bacteria which are both extremely diverse and almost devoid of replicates. The collection is being used to identify and develop antibiotic and antifungal agents to combat the rise of antimicrobial resistance (AMR). Through screening >7000 diverse and dereplicated actinomycetes from the collection, Demuris has identified over 100 strains with broad-spectrum antibacterial activity that inhibit a panel of multiply antibiotic resistant E. coli strains. In this project Demuris will aim to sequence the genomes of all of these broad spectrum antibiotic producer strains. This will be performed rapidly and cheaply using long read, single molecule sequencing from Oxford-Nanopore. We will create an in-house technology platform to sequence and then bioinformatically screen the genomes of antibiotic producing bacteria at low cost using existing software. Strains bioinformatically predicted to produce known broad spectrum antibiotic compounds will be removed from the discovery pipeline, dramatically speeding up the dereplication process and zooming in rapidly on strains producing novel antibiotic compounds. Novel antibiotics will be structurally characterised and a data package consisting of microbiology and cytotoxicity will be generated. These data will be sufficient to enable evaluation for clinical development. This "data first" based approach promises to improve efficiency and will allow all resources to be prioritised on the compounds with the best chance of clinical success.

"Actinobacterial Natural Products are an important source of commercially used compounds, including many clinically used antibiotic, antifungal and anticancer compounds. The genes required to produce these compounds are encoded by Biosynthetic Gene Clusters (BGCs). Genomic sequence data have revealed the presence of a large fraction of putatively silent biosynthetic gene clusters in the genomes of actinomycetes that encode for secondary metabolites, which are not detected under standard fermentation conditions. We have developed a system that can boost the weak production from these clusters and has application for the discovery of new antibiotics and antifungal compounds. This project will allow the methods to be optimised and applied to other BGCs of known or unknown function. In addition we will further develop this method and combine the system with a transposon which can then be inserted randomly into the genomes of actinomycetes boosting the production of natural products in this area. The system has application for boosting compound titre from known BGCs and will be used for the identification of otherwise undetectable antibiotic and antifungal compounds. The system will be used on a variety of rare actinomycetes and induced novel antibiotic and antifungal compounds will be characterised."

Antibiotic discovery from natural sources is beset by the re-isolation of known compounds and the difficulties in working with wild-type strains. Demuris and TGAC will transform this approach and use genome sequencing to identify and dereplicate known antibiotic gene clusters from a set of high value actinomycete strains that produce broad-spectrum antibiotics though presently unknown. Known and novel gene clusters will be identified bioinformatically and software developed to allow the integration data facilitating cluster prioritisation. To confirm the bioinformatic predictions the masses of the most promising novel antibiotics will be identifed and the gene clusters cloned and heterologously expressed using an optimised host. This "data first" based approach promises to reinvigorate the natural products sector, and this is desperately needed if new and novel antibiotics are to be developed.

New antibiotics are urgently needed to replace those that are lost to increasing antibiotic resistance. Ourtechniques focus on using Synthetic Biology to transfer the biosynthetic gene clusters for antibiotics frompoorly- and un-characterised environmental species into optimised SuperHosts. Traditional approaches for thisfocus on constructing bacterial or phage-derived artificial chromosomes; our approach provides a step-changeto the protracted traditional methods. We propose new methods that will also accelerate discovery andexploitation of previously unseen antibiotics from existing libraries.

New antibiotics are urgently needed to replace those that are lost to increasing antibiotic resistance. Demuris has been using synthetic biology approaches to access these silent or cyptic gene clusters by heterologous production. We have been working to extract the biosynthetic genes from poor- or non-producing donor strains and introducing them to a Superhost, optimised for antibiotic production and its own growth characteristics. We propose new methods that will accelerate discovery and exploitation of previously unseen antibiotics from existing libraries.

New antibiotics are urgently needed to replace and supplement those eroded by bacteria resistance. Synthetic biology approaches have vast potential to aid the discovery and development of antibiotics and other medicines. Benefits may include overcoming common problems associated with antibiotic discovery from natural sources such as poor growth characteristics, reproducibility and poor yield. Demuris has identified a promising broad spectrum antibiotic but it is produced in low quantity. Bioinformatics and Synthetic Biology tools will be used to identify and refactor the gene cluster for optimum production and allow the creation of antibiotics with improved properties. Developing these Synthetic Biology methods may give scientists new weapons to be used in the battle against bacterial resistance.

Demuris has a pipeline of novel antibiotics produced by a variety of different actinobacteria. The leading molecule is presently made by an organism that is not ideal for large scale fermentation, and the compound structure may not yet be optimal in terms of pharmacological properties. We wish to identify the antibiotic producer gene cluster and re-engineer it for production in an optimized chassis for production of actinobacterial natural products based on Streptomyces venezuelae (Sven). Pathway engineering will be used to generate analogues of the lead antibiotic, which will be used to facilitate drug optimization and IP protection.

Antimicrobial resistance is regarded by the World Health Organisation (WHO) as one of the greatest threats to the human race. Several multi-drug-resistant strains are becoming untreatable, and due to the long development times there is an urgent need to accelerate research into new therapies. Unfortunately, many pharmaceutical companies have abandoned their antibiotic programmes, and the number of new antibiotic compounds in development is diminishing. Demuris has screened several thousand rare and diverse antibiotic producing bacteria and several novel antibiotics have been identified which target problem multi-drug resistant pathogens. Demuris must now obtain enough antibiotic material for structure characterisation and perform the following experiments to generate a data package which will identify any potential toxicity. This project will show whether these compounds have the key properties needed for antibiotics of the future.

Infectious diseases remain a leading cause of global deaths. New diseases and the continuous rise of multi-drug resistant bacteria suggest that there will be a continual need for novel antibacterial therapies with superior activity and better tolerance. However the number of antibiotic compounds in development is worryingly small. Demuris has screened a unique collection of bacterial isolates and has identified three potentially novel antibiotics. This project has developed these compounds from the early stages of discovery through to a point where the compounds can be evaluated for viability of commercialisation and pre-clinical development. The National Industrial Biotechnology Facility has provided the resources not currently available to Demuris and enabled the production of sufficient quantities of purified compounds for testing and development purposes.