METABACT

This project involves chemists, bacteriologists and physicists, for the study of metallodrugs with antibacterial activities. Usually, most of the compounds are already active in vitro, but display limited activities in cellulo because their internal concentration is low since the bacterial membrane has a low permeability, especially in Gram-negative bacterial species. The aim of this project is to find new integrated strategies to improve drugs penetration, evaluate biological activities and relate the antibacterial activities to the concentration and location of the drugs inside the cells. The project will focuse : i) on peptide deformylase (PDF-inhibs) and methionine aminopeptidases inhibitors (MetAP-inhibs), since these proteins are relevant targets for antibiotics ii) on Mn glycocomplexes with antisuperoxide activity (Mn-SOD-like), as relevant compounds to produce inside the bacteria a cascade of killing agents such as H2O2 upon reaction with O2•-, and then HO• (Haber Weiss réaction) or HOCl (réaction of H2O2 with myeloperoxidase). To improve drugs penetration, permeabilizers such as cationic peptides (related to antimicrobial peptides but shorter), oligosaccharides and aminoglycosides will be used either in combination with the free PDFinhibs or MetAPinhibs and Mn-SOD-like (binary sytems) or grafted on the ligand of the (Mn-SOD-like) metallodrug or on the ancillary ligand of a metal-chaperone constructed with a PDFinhib or a MetAPinhib (bioconjugates) and used as carrier of these inhibitors. Bioconjugates based on Mn-SOD-like that are able to deliver inside the bacteria two active species, a compound with a SOD activity and a PDF or MetAP inhibitor, will be prepared. The biological activities of the binary systems and bioconjugates will be evaluated in diverse bacterial backgrounds and conditions including strains devoid of efflux pump activity and on resistant strains overexpressing them. These metallodrugs whose activities are directly related to the metal oxidation state will be characterized by EXAFS and XANES in order to validate their molecular and chemical integrity in media related to cell culture media. The final goal of the project is to relate the biological activities of the drugs as binary systems or bioconjugates to their cellular uptake and concentration. This will be achieved by cell imaging experiments using micro-fluorescence VUV at DISCO at SOLEIL. For the most efficient compounds some prospective experiments will be performed at the nano range (µ-SXRF) at ESRF and on nanoscopium beamline at SOLEIL when available within 3 years. In these experiments, the probe used will be a heavy atom such as Br that is présent on the inhibitor, or on the glycoligand or on the ancillary ligand of the metallodrugs, and/or Mn, Co or Fe metal cations. The strength of the mapping experiments is that they use non invasive techniques with no need for additional markers that may lead to side-effects or to mis-localization (membranes versus soluble compartments). Overall, the project may allow to determine key structural and chemical parameters associated to metallodrugs (pharmacophors) and grafted permeabilizers, that play a key role in the diffusion through the bacterial membranes. The characterization of these systems will provide new tools to improve the antibacterial efficiency of compounds known to be active towards bacterial targets in vitro.