Unpriming Bacterial Pumps to Block Antibiotic Resistance

    When a bacterial machine called the efflux pump is in working order, it expels antibiotics, contributing to antibiotic resistance. The pump, however, cannot work if it lacks all of its molecule-sized parts. Keeping these parts out of stock, then, is a promising strategy in preserving the effectiveness of antibiotics that qualify as “last line of defense” drugs. The strategy could even restore the effectiveness of antibiotics that bacteria have come to tolerate.

    To realize the pump-defeating strategy, scientists based at the University of Texas Southwestern Medical Center (UTSW) decided to interfere with the synthesis of certain proteins that are essential pump components. Interestingly, the scientists were able to make use of the genomic blueprints for these parts. Specifically, the scientists developed a short antisense oligomer designed to inhibit the expression of a pump part coded by the acrA gene. The antisense oligomer—more properly, the peptide-conjugated phosphorodiamidate morpholino oligomer (PPMO)—was used to increase antibiotic susceptibility in Escherichia coli.

    Details of the work appeared September 15 in the journal PLoS Biology, in an article entitled, “Sequence-Specific Targeting of Bacterial Resistance Genes Increases Antibiotic Efficacy.” This article explained that the PPMO compound itself didn’t kill bacteria. Instead, the PPMO worked by mimicking DNA or RNA and binding to messenger RNA like the other half of a zipper, preventing the synthesis of the acrA-coded protein, which is essential to the AcrAB-TolC efflux system.

    “By employing this strategy, we can inhibit E. coli growth using 2- to 40-fold lower antibiotic doses, depending on the antibiotic compound utilized,” wrote the article’s authors. “The sensitizing effect of the antisense oligomer is highly specific to the targeted gene’s sequence, which is conserved in several bacterial genera, and the oligomer does not have any detectable toxicity against human cells.”

    In fact, some of the antibiotics evaluated in the current study are never used against E. coli because they are thought to be ineffective. Yet these antibiotics were able to kill bacteria when used in conjunction with the AcrA-PPMO.

    “This is just a different strategy,” said David Greenberg, M.D., associate professor of internal medicine and microbiology at UTSW and a co-senior author of the study. “There is a lot of interest in trying to develop new antibiotics, or antibiotics that act in new ways. The other way of thinking about this challenge is to try to make a resistant organism sensitive.”

    The current work builds on earlier work by Erdal Toprak, Ph.D., assistant professor of pharmacology and a co-senior author of the PLoS Biology article. About a year ago, his research team identified a mutation in E. coli that increased drug sensitivity by blocking the AcrAB-TolC efflux pump complex.

    The AcrA-PPMO also was effective against the human pathogens Klebsiella pneumoniae and Salmonella enterica, as those bacteria contain the same efflux pump with a matching gene sequence, Dr. Greenberg noted.

    The next step planned by the researchers will be to study the effect of the AcrA-PPMO in animal models. The researchers also will study whether a PPMO could be effective against other extremely antibiotic-resistant strains of bacteria.