Exploring metallophore-antibiotic conjugates as a novel antimicrobial strategy that exploits Pseudomonas aeruginosa zinc dependence

Pseudomonas aeruginosa (PA) is a ubiquitous Gram-negative bacterium. Pulmonary infections caused by PA represent a serious threat, especially to patients with cystic fibrosis (CF). This risk is further exacerbated by the increasing spread of bacterial strains that show antibiotic resistance1. The ease with which PA can colonize the respiratory tract depends on its ability to proliferate even in environments where essential metals (such as Zn) are present in low concentration. In particular, zinc plays a fundamental role, as it is present in many bacterial proteins crucial for virulence. A peculiar tactic of PA to obtain zinc involves the secretion of pseudopaline (Fig. 1), that is a low molecular weight zinc chelating molecule, capable of binding zinc present in the external environment, extracting it also from proteins present in the host, and deliver it inside the bacterial cell2. The mechanism by which the zinc pseudopaline complex is transported into the bacterial cell has not yet been completely defined. The use of pseudopaline to chelate zinc represents a potential vulnerability of the pathogen, which can be exploited to allow antibiotics to enter the cell via the Trojan horse approach3. The aim of this study is to synthesize simplified analogs of pseudopaline, to evaluate the minimal structural requirements needed to participate in the zinc transport process of PA. These simplified zincophore compounds, simpler to synthesize than pseudopaline, can be more easily conjugated to various antibiotics that are still in use and active against these PA (such as aztreonam or ampicillin), to facilitate their entry into the bacterial cell through a preferential pathway. The antibacterial activity of these conjugates is evaluated against different PA strains to confirm the involvement of the zinc recovery pathway and demonstrate the proof of concept of the Trojan horse approach.