New antimicrobial strategies against bacterial infections

Abstract

Bacterial infections are a major health concern worldwide due to the mortality and comorbidity associated levels. Besides the development of drug-resistance mechanisms, most recalcitrant infections are caused by bacteria forming biofilms, which are bacterial communities that grow within an extracellular polymeric substance (EPS) matrix that protects bacterial cells from the action of antimicrobials and immune mechanisms. The frequent appearance and spreading of bacterial drug-resistant strains, together with the recalcitrance of infections caused by biofilms, has pointed out the urgent need to develop novel antibacterial agents that target essential bacterial processes and biofilm-forming bacteria. This thesis investigated the development of new antibacterial strategies by both targeting bacterial essential processes and biofilm-forming bacteria. Ribonucleotide reductase (RNR) are essential enzymes required by any cellular organism, since they catalyze the synthesis of deoxyribonucleotides, critical for both DNA synthesis and repair processes. Due to its key role in DNA replication, several RNR inhibitors have been developed as antiproliferative drugs in cancer and infectious diseases. Amongst RNR inhibitors, radical scavenger molecules have proved for long its inhibitory activity against the RNR enzyme, including some cytotoxic hydroxylamine derivatives such as hydroxyurea (HU). Here, the use of hydroxylamine derivative compounds as antibacterial agents specifically targeting the bacterial RNR enzyme was investigated. The results provided show that N-methyl-hydroxylamine (M-HA) molecule and some newly synthesized Nhydroxylamine derivative molecules (N-HA) inhibit bacterial RNR, displaying a wide range of antibacterial activity against different bacterial pathogens, together with low cytotoxicity to eukaryotic cells. Also, M-HA molecule shows intracellular antimycobacterial activity during macrophages infection, together with Pseudomonas aeruginosa in vitro antibiofilm activity. Several of the N-HA molecules display antibiofilm activity against P. aeruginosa, Staphylococcus aureus, and Escherichia coli biofilms, and we demonstrate the ability of such molecules to inhibit bacterial growth by radical scavenging of the RNR enzyme. Further, this thesis explores the use of a drug delivery system based on poly(lactic-co-glycolic) (PLGA) nanoparticles (NPs) to remove P. aeruginosa biofilms by combining the action of the antibiotic ciprofloxacin and the DNA-hydrolytic activity of the deoxyribonuclease I enzyme (DNase I). The synthesized biodegradable PLGA NPs, loaded with ciprofloxacin and functionalized with DNase I through poly(lysine) (PL) coating, synergistically improve the ciprofloxacin antibacterial efficacy by disassembling the extracellular DNA, one of the main components of the EPS matrix.