A-051. Characterization of the Physiological Response of Staphylococcus aureus to the Bactericidal Pyrrolobenzodiazepine Dimer ELB-21

M. Doyle1, K. Bailey2, D. E. Thurston1,3, P. W. Taylor1;
1Sch. of Pharmacy, London, UNITED KINGDOM, 2Queens Med. Ctr., Nottingham, UNITED KINGDOM, 3Spirogen Ltd, Sch. of Pharmacy, London, UNITED KINGDOM.

Staphylococcus aureus causes a broad spectrum of infectious conditions, ranging from superficial skin abscesses to severe systemic infections such as septicemia and osteomyelitis. It is a leading cause of nosocomial and community-acquired infections worldwide and is of increasing concern due its rapid ability to acquire antibiotic resistance genes. The pathogenicity of Staphylococcus aureus is dependent on the coordinated expression of a plethora of extracellular and cell-wall-anchored proteins. The production of these virulence determinants is growth-phase-dependent and is tightly controlled by a network of regulatory proteins. Recently, we have shown that the pyrrolobenzodiazepine dimer, ELB-21, has potent anti-staphylococcal activity due to efficient cell penetration and sequence-selective cross-linking of the bacterial genome. In this study, we used differential proteomics to determine the physiological response of methicillin-resistant Staphylococcus aureus to ELB-21 exposure. Bacterial cultures were grown to exponential or stationary phase of growth and exposed to subinhibitory concentrations of ELB-21 for 1 or 5 h. Total cytoplasmic and extracellular protein extracts were separated by two-dimensional gel electrophoresis and differentially expressed protein spots identified by mass spectrometry. ELB-21 DNA sequence-binding sites within methicillin-resistant Staphylococcus aureus genomes were identified in silico using the DNA viewer and annotation tool Artemis. Cytoplasmic and extracellular proteins involved in a variety of key cellular functions, including fatty acid biosynthesis, folate biosynthesis, beta-lactam resistance and systemic infection were shown to be downregulated. In addition, ELB-21 DNA sequence-binding sites were found near or within the open reading frame of the genes coding for these proteins. These results strongly indicate that ELB-21 cross-links genomic DNA in a sequence-selective manner and subsequently inhibits translation of specific proteins required for staphylococcal pathogenesis.