ABSTRACT
Intrinsic resistance in Pseudomonas aeruginosa, defined by chromosomally encoded low outer membrane permeability and constitutively over-expressed efflux pumps, is a major reason why the pathogen is refractory to many antibiotics. Herein, we report that heterodimeric rifampicin-tobramycin conjugates break this intrinsic resistance and sensitize multidrug and extensively drug-resistant P. aeruginosa to doxycycline and chloramphenicol in vitro and in vivo. Tetracyclines and chloramphenicol are model compounds for bacteriostatic effects, but when combined with rifampicin-tobramycin adjuvants, their effects became bactericidal at sub MIC levels. Potentiation of tetracyclines correlates with the SAR of this class of drugs and is consistent with outer membrane permeabilization and efflux pump inhibition. Overall, this strategy finds new uses for old drugs and presents an avenue to expand the therapeutic utility of legacy antibiotics to recalcitrant pathogens such as P. aeruginosa.
Subject(s)
Anti-Bacterial Agents/pharmacology , Chloramphenicol/pharmacology , Doxycycline/pharmacology , Pseudomonas aeruginosa/drug effects , Rifampin/pharmacology , Tobramycin/pharmacology , Animals , Anti-Bacterial Agents/chemical synthesis , Anti-Bacterial Agents/chemistry , Anti-Bacterial Agents/toxicity , Bacterial Outer Membrane Proteins/metabolism , Drug Resistance, Multiple, Bacterial/drug effects , Drug Synergism , HEK293 Cells , Hep G2 Cells , Humans , Membrane Transport Proteins/metabolism , Microbial Sensitivity Tests , Molecular Structure , Moths , Rifampin/analogs & derivatives , Rifampin/chemical synthesis , Rifampin/toxicity , Swine , Tobramycin/analogs & derivatives , Tobramycin/chemical synthesis , Tobramycin/toxicityABSTRACT
Molecular recognition of small molecule ligands by the nucleic acid aptamers for tobramycin, ATP, and FMN has been examined using electrospray ionization mass spectrometry (ESI-MS). Mass spectrometric data for binding stoichiometry and relative binding affinity correlated well with solution data for tobramycin aptamer complexes, in which aptamer/ligand interactions are mediated by hydrogen bonds. For the ATP and FMN aptamers, where ligand interactions involve both hydrogen bonding and significant pi-stacking, the relative binding affinities determined by MS did not fully correlate with results obtained from solution experiments. Some high-affinity aptamer/ligand complexes appeared to be destabilized in the gas phase by internal Coulombic repulsion. In CAD experiments, complexes with a greater number of intermolecular hydrogen bonds exhibited greater gas-phase stability even in cases when solution binding affinities were equivalent. These results indicate that in at least some cases, mass spectrometric data on aptamer/ligand binding affinities should be used in conjunction with complementary techniques to fully assess aptamer molecular recognition properties.