Strategies for bypassing the membrane barrier in multidrug resistant Gram-negative bacteria.

In Gram-negative bacteria, the envelope is a sophisticated barrier protecting the cell against external toxic compounds. Membrane transporters, e.g., porins or efflux pumps, are main filters regulating the internal accumulation of various hydrophilic molecules. Regarding bacterial susceptibility towards antibacterial agents, membrane permeability is part of the early bacterial defense. The bacterium manages the translocation process, influx and efflux, to control the intracellular concentration of various molecules. Antibiotics and biocides are substrates of these mechanisms and the continuing emergence of multidrug resistant isolates is a growing worldwide health concern. Different strategies could be proposed to bypass the bacterial membrane barrier, comprising influx and efflux mechanisms, in order to restore the activity of antibiotics against resistant bacteria.

Efflux pumps are involved in the defense of Gram-negative bacteria against the natural products isobavachalcone and diospyrone.

The activities of two naturally occurring compounds, isobavachalcone and diospyrone, against documented strains and multidrug-resistant (MDR) Gram-negative bacterial isolates were evaluated. The results indicated that the two compounds exhibited intrinsic antibacterial activity against several Gram-negative bacteria, and their activities were significantly improved in the presence of an efflux pump inhibitor (MIC values decreased to below 10 microg/ml). In addition, the activities of isobavachalcone and diospyrone against various strains exhibiting deletions of the major efflux pump components (AcrAB, TolC) were significantly increased. The overall results indicate that isobavachalcone and diospyrone could be candidates for the development of new drugs against MDR strains and that their use in combination with efflux pump inhibitors reinforces their activity.

Efflux mechanism, an attractive target to combat multidrug resistant Plasmodium falciparum and Pseudomonas aeruginosa.

Chemoresistance is a general health problem concerning infectious diseases and cancer treatments. In this context, the worldwide dissemination of << pandrug >> and << multidrug>> resistant pathogens has severely compromised the efficacy of our antimicrobial weapons and dramatically increased the occurence of therapeutic failure. To efficiently combat multi-resistant pathogens, it is necessary to clearly define the molecular basis of the general resistance mechanism associated with the expression of active efflux pumps, which strongly restrict the intracellular concentration of antimicrobial drugs. Several families of efflux systems capable of multiple drug extrusion have been described. The activity of some efflux systems requires ATP hydrolysis for drug transport while others require a sodium or proton antiport. In this review we focus on two important human pathogens, Plasmodium falciparum and Pseudomonas aeruginosa, which exhibit a high level of antimicrobial resistance associated with the expression of efflux mechanisms. The efflux mechanisms and the development of efflux pump inhibitors (EPIs) are discussed regarding these two pathogens.

Antibiotic efflux pumps in Gram-negative bacteria: the inhibitor response strategy.

After several decades of continuously successful antibiotic therapy against bacterial infections, we are now facing a worrying prospect: the accelerated evolution of antibiotic resistance to important human pathogens and the scarcity of new anti-infective drug families under development. Efflux is a general mechanism responsible for bacterial resistance to antibiotics. This active drug transport is involved in low intrinsic susceptibility, cross-resistance to chemically unrelated classes of molecules, and selection/acquisition of additional mechanisms of resistance. Thus, inhibition of bacterial efflux mechanisms appears to be a promising target in order to (i) increase the intracellular concentration of antibiotics that are expelled by efflux pumps, (ii) restore the drug susceptibility of resistant clinical strains, and (iii) reduce the capability for acquired additional resistance. Structurally unrelated classes of efflux pump inhibitors (EPIs) have been described and tested in the last decade, including some analogues of antibiotic substrates and new chemical molecules. Among the current collection of EPIs, only a few compounds have been studied taking into account the structure-activity relationships and the spectrum of activity in terms of antibiotics, pumps and bacteria. While large efforts have characterized an increasing number of bacterial efflux pumps and generated several potentially active EPIs, they have not elucidated the molecular basis of efflux transport and inhibition. Recent studies of pump-substrate complexes, the 3D resolution of the efflux pumps, the synthesis of novel compounds and molecular dynamic studies may generate new clues to decipher and select novel targets inside the efflux mechanisms and, finally, may result in a clinically useful molecule.

Alkylaminoquinolines inhibit the bacterial antibiotic efflux pump in multidrug-resistant clinical isolates.

Over the last decade, MDR (multidrug resistance) has increased worldwide in microbial pathogens by efflux mechanisms, leading to treatment failures in human infections. Several Gram-negative bacteria efflux pumps have been described. These proteinaceous channels are capable of expelling structurally different drugs across the envelope and conferring antibiotic resistance in various bacterial pathogens. Combating antibiotic resistance is an urgency and the blocking of efflux pumps is an attractive response to the emergence of MDR phenotypes in infectious bacteria. In the present study, various alkylaminoquinolines were tested as potential inhibitors of drug transporters. We showed that alkylaminoquinolines are capable of restoring susceptibilities to structurally unrelated antibiotics in clinical isolates of MDR Gram-negative bacteria. Antibiotic efflux studies indicated that 7-nitro-8-methyl-4-[2'-(piperidino)ethyl]aminoquinoline acts as an inhibitor of the AcrAB-TolC efflux pump and restores a high level of intracellular drug concentration. Inhibitory activity of this alkylaminoquinoline is observed on clinical isolates showing different resistance phenotypes.

In vitro reversal of chloroquine resistance in Plasmodium falciparum with dihydroethanoanthracene derivatives.

The effects of combining four dihydroethanoanthracenic (DEA) derivatives and chloroquine were assessed in vitro against Plasmodium falciparum chloroquine resistant parasites W2, Palo Alto, FCR3, and Bres1. Like verapamil or promethazine, the four dihydroethanoanthracenic derivatives tested can be added to the growing list of agents that show capability in enhancing the activity of chloroquine against resistant parasites. The structurally related tricyclic antihistaminic compounds examined in this study exerted different intrinsic antimalarial activity, but the same chloroquine-potentiating activity as verapamil or promethazine. They may act both on the rate of chloroquine accumulation and on its access to ferriprotoporphyrin IX. The reversal mechanism would be assumed to result from competition between DEA derivatives and chloroquine for efflux translocation sites, thus causing an increase in steady-state accumulation of chloroquine and a return to susceptibility. Restoration of therapeutic efficacy of chloroquine against resistant parasites by the administration of an additional drug available at relatively low cost may be a more effective strategy than the introduction of another antimalarial drug at the national level.