Exploration of the TPP riboswitch as a new target for antibiotics

The antibiotics we are currently using are loosing effectiveness due to the emergence of resistant bacteria. Therefore, there is an urgent need to develop new antibiotics. In this project, we aim to develop ligands for an RNA element (a so called riboswitch), that controls the expression of essential bacterial genes. These ligands can then serve as starting points for drug discovery for future antibiotics. To reach this goal, we have assembled a team of highly skilled researchers from Norway, the Czech Republic, Latvia, Germany, and Canada, who are experts in different disciplines which are essential for this project. In the part of the project funded by the NFR, in vitro transcription of the riboswitch has been established. Further, with our collaborators in Canada and the Czech Republic, we have established an assay that can be used to screen thousands of compounds for riboswitch binding and this assay has now been used to screen the library available at CZ-Openscreen. This has resulted in a number of hits using the primary assay. Validation with alternative assays is ongoing. Up now, the validation results are inconclusive, therefore, alternative assays are being setup.

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In this project, we will explore the TPP riboswitch as a new drug target for antibiotics for key ESKAPE pathogens (E. coli, K. pneumoniae, A. baumannii, P. aeruginosa, S. aureus) and Streptococcus pneumoniae. The TPP riboswitch has already been validated as a drug target, however, potent and drug-like ligands with antibiotic activity are needed as starting points to develop novel strategies for anti-infective treatments. The goal of this proposal is to deliver such compounds. To reach this goal, we have assembled a team of highly skilled researchers from Norway, the Czech Republic, Latvia, Germany, and Canada, who are experts in riboswitch biology, microbiology, compound screening, structure-based drug design, organic synthesis and medicinal chemistry. Using an innovative assay technology, we will develop a high-throughput assay that monitors simultaneously transcription efficiency and the regulatory activity of the riboswitch, which is crucial for its action, and use this assay to screen the CZ- and EU-OPENSCREEN libraries of lead-like compounds. The hits obtained will be thoroughly validated and the most promising hits will be optimized to improve their affinity. The advanced compounds will be evaluated for antibiotic activity against the key ESKAPE pathogens and Streptococcus pneumoniae. We will also assess the broad-spectrum potential of the compounds and carry out mode of action studies to ensure that the compounds act on target. If the TPP riboswitch holds up to its high promises, this project will pave the way for urgently needed new antibiotics.