Multidrug antibiotic resistance is a global public health crisis that leads to thousands of people's deaths every year. One of the primary causes of resistance in Gram-negative bacteria such as Escherichia coli is the overexpression of multicomponent molecular machines called multidrug efflux pumps. Efflux pump inhibitors (EPIs)are a promising alternative approach to combat antibiotic resistance. We previously identified a diaminoquinoline acrylamide, NSC-33353, as an active EPI of E. coli efflux pump.

This report will describe a series of cinnamoyl compounds that are analogs of NSC-33353 showing significant activity against the AcrAB-TolC efflux pump of E. coli. To determine the antibacterial properties and propensity of these analogs to act as efflux substrates, we analyzed bacterial growth inhibitory activities of the compounds using efflux proficient and efflux deficient E.coli cells. Surprisingly, the results show that a large number of the analogs possess an antibacterial activity, although their original hit was only a weak antibacterial agent. In the presence of antibiotics novobiocin and erythromycin compounds have a significant potentiation activity. The surface plasmon resonance data show that compounds bind with high affinities both AcrA, a membrane fusion protein, and AcrB, an efflux transporter. The fluorescence-based accumulation assay showed that these compounds inhibit the efflux of fluorescent probes. Taken together, these results show that this series of compounds are promising EPIs.

In summary, we have identified compounds that bind to AcrA, AcrB and potentiate the antibacterial properties of novobiocin and erythromycin in E. coli. We report a new series of EPIs that inhibit the activity of AcrAB-TolC efflux pump.