New amphiphilic neamine conjugates bearing a metal binding motif active against MDR E. aerogenes Gram-negative bacteria.

Structure of bacterial envelope is one of the major factors contributing to Gram negative bacterial resistance. To develop new agents that target the bacterial membranes, we synthesized, by analogy with our previous peptide conjugates, new amphiphilic 3',4',6-trinaphthylmethylene neamines functionalized at position 5 through a short spacer by a chelating group, tris(2-pyridylmethyl)amine (TPA) and di-(picolyl)amine (DPA) and tetraazacyclotetradecane (Cyclam). ESI mass spectrometry analyses showed that neither Zn(II)(NeaDPA) nor Cu(II)(NeaCyclam) were stable in the Mueller Hinton (MH) medium used for antibacterial assays. In contrast Zn(NeaTPA) was stable in the MH medium. Interestingly, in MH, the free ligand NeaTPA was found bound to zinc, the zinc salt being the most abundant salt in this medium. Thus, the antibacterial activities of all compounds were evaluated as free ligands against E. coli strains, wild type AG100 and E. aerogenes EA289 (a clinical MDR strain that overexpresses AcrAB-TolC efflux pump), as well as AG100A an AcrAB- E. coli strain and EA298 a TolC- derivative. NeaCyclam and Zn(NeaTPA) were by far the most efficient compounds active against resistant isolate EA289 with MICs in the range 16-4 and 4 μM, respectively, while usual antibiotics such as β-lactams and phenicols were inactive (MICs > 128) and ciprofloxacin was at 64 μM. Zn(NeaTPA) and NeaCyclam were shown to target and permeabilize the outer membrane of EA289 by promoting the cleavage of nitrocefin by periplasmic β-lactamase. Moreover, all the neamine conjugates were able to block the efflux of 1,2'-dinaphthylamine in EA289, by acting on the efflux transporter located in the inner membrane. These membranotropic properties contribute to explain the activities of these neamine conjugates toward the MDR EA289 strain.