The quest for compounds capable of circumventing antimicrobial resistance is important and urgent. Current research has been focusing not only in the search for new antibiotics, but also for “helper” compounds, capable of blocking resistance mechanisms and, therefore, regaining the activity of currently used antibacterial drugs. In this scope, bacterial efflux pump inhibitors arise as interesting compounds, as they can block this resistance mechanism and lead to increased efficacy of antibiotics.

Our group has been studying the potential of steroid derivatives, and has already established their potential as antifouling agents, also capable of inhibiting the growth of biofilm-forming marine bacteria. One amide derivative was also found to display promising activity in antibacterial and in synergy assays, as well as in the efflux pump inhibition assays. Therefore, further amide derivatives were synthesized, using coupling agents such as COMU and HOBt.

A screening for antimicrobial activity has been performed in Gram-positive bacteria (Staphylococcus aureus ATCC 29213, Enterococcus faecalis ATCC 29212 and S. aureus MRSA 272123) and Gram-negative bacteria (Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853 and a mutant strain of Salmonella enterica serovar Typhimurium expressing the AcrAB-TolC efflux system with the acrA gene inactivated). Then, they were tested for their capability to modulate ethidium bromide accumulation, a known efflux pump substrate, whose accumulation leads to an increase in fluorescence. The derivatives tested were able to increase the accumulation of ethidium bromide, which translates into efflux pump inhibition.

Further studies will focus on their ability to interact with other resistance mechanisms.