A pipeline technology for discovery of new antibiotics from Streptomyces

It is inescapable that modern medicine is faced with a severe challenge in the form of 'Superbugs' - disease-causing bacteria that are resistant to front-line antibiotics. The antibiotic era in medicine has spanned the last 70 years with the first drugs, penicillin and streptomycin, being introduced in the late 1940's. A dramatic reduction in fatalities associated with bacterial infections followed, as did the discovery of many new antibiotics, reaching a peak in the 1980's. Since then, there has been a dramatic decline in new antibiotic discovery, and an increase both in the incidence of resistance to front-line antibiotics and in patient fatalities due to infections by these resistant bacteria. The majority of antibiotics are produced by soil bacteria called Streptomyces. Recent genome sequencing projects with representative species of these bacteria have revealed that they possess many 'cryptic' antibiotic biosynthetic pathways; that is they have the genetic potential to produce many more antibiotics than previously realised. An outcome of a European collaborative research project, ActinoGEN, coordinated by Dyson, indicates that these cryptic pathways are not genetic relics but can be activated to direct production of new antibiotics. Consequently we can now interpret the last 70 years as an initial period during which 'low-hanging fruit', our current front-line antibiotics, were discovered. Thereafter, the success in discovery has declined. A current challenge is now to harvest the plentiful higher-hanging fruit, the products of the cryptic pathways, and extend the range of antibiotics that can be used in medicine. The ways to do this are at present very time-consuming, dependent on first obtaining and analysing the genome sequences of Streptomyces species, before genetic engineering to activate a cryptic pathway. This proposal addresses a technological gap by devising a means of inexpensive strain manipulation coupled with high-throughput screening of extensive Streptomyces strain libraries to discover new antibiotic products of cryptic pathways. The strain manipulations involve rapid and synergistic approaches to activate and over-produce new antibiotics. To demonstrate a proof-of-principle, having obtained manipulated strains over-producing a novel antibiotic, we will then analyse the antibiotic and determine the cryptic biosynthetic pathway that directs its synthesis. Successful demonstration of the technology will subsequently lead to its adoption to discover new antibiotics produced in large strain collections, in collaboration with industrial partners from the pharmaceutical/biotech sector.

Streptomyces genome sequencing projects have revealed many 'cryptic' potential antibiotic biosynthetic gene clusters in representative species. The EU ActinoGEN project has demonstrated that for two representative cryptic pathways, genetic manipulation of pathway-specific regulators can activate production of novel antibiotics. We can now interpret the antibiotic era, spanning the last 70 years, as an initial period of discovery of 'low-hanging fruit' - our current front-line antibiotics - and a subsequent decline in success of discovery. A technology gap exists in our ability to harvest the many higher-hanging fruit, the products of cryptic pathways, as the existing approaches are dependent on a combination of genome sequencing, genome mining and strain manipulation, all of which do not lend themselves to high-throughput screening of extensive strain libraries. This proposal addresses the technology gap by combining rapid, synergistic empirical strain manipulation with high-throughput screening to permit the discovery of new antibiotic products of cryptic pathways. To demonstrate a proof-of-principle, we will apply ribosome/RNA polymerase engineering with interspecific protoplast fusion, random mutagenesis and genome shuffling to obtain recombinant strains over-producing novel antibiotics. We will analyse new antibiotics obtained and, at least for one, identify the corresponding cryptic biosynthetic gene cluster directing its synthesis by targeted mutation. Although the proof-of-principle will be demonstrated with species for which we have genome sequence information, the technology is designed for future application with extensive libraries of non-sequenced species.

A current BBSRC research priority is 'Ageing research: lifelong health and wellbeing'. Given that the efficacy of front-line antibiotic treatments has been compromised, with the elderly being conspicuously susceptible to bacterial infections, this technology will significantly impact on the research priority. Who will benefit from this research? The immediate non-academic beneficiaries include companies in the pharma/biotech sector involved in drug screening and development. The technology is designed to improve the success rate for future drug discovery and hence increase the profitability of companies in the sector. Long-term beneficiaries are patients with life-threatening infectious diseases who do not respond to treatment from current front-line antibiotics. These will include patients in developed countries who have contracted resistant community-acquired or nosocomial infections. Effective short treatments will greatly help reduce health service budgets committed to treatment of these patients. In addition, with respect to tuberculosis, the potential of new anti-tubercular drugs will benefit millions of patients in both developed and under-developed nations. How will they benefit from this research? The high throughput screening technology available on license to companies in the pharma/biotech sector will permit identification of valuable new antibiotic leads. Long-term patient beneficiaries will be treated with potential life-saving new antibiotics, with consequent reductions in mortality and savings to health service budgets. Communications and Engagement Research data generated during the course of this project will be communicated to the scientific community through presentation at national and international scientific meetings and publication in peer-reviewed scientific journals in a timely fashion. Publication of peer-reviewed outputs will take place as soon as possible during the course of the research project or at the end of the funded period. In addition we will communicate with our University Publicity departments and BBSRC to ensure that where possible, significant findings are also communicated more generally. The PI has good contacts with several UK/EU companies engaged in the sector, and once IP has been protected, he will seek collaborations with these companies. Collaboration The pilot project involves no collaboration. Subsequent to establishing a proof-of-principle, extensive collaborations will be sought with companies in the pharma/biotech sector. Further exploitation and application We will seek to protect the IP derived from the project and then engage with industry to exploit this technology with established industrial strain collections. Capability Primary responsibility for impact activities will rest with the PI, Prof Paul Dyson, who has good experience of interactions with industrial partners.