Combating multidrug-resistant Gram-negative bacteria with structurally nanoengineered antimicrobial peptide polymers.

With the recent emergence of reports on resistant Gram-negative 'superbugs', infections caused by multidrug-resistant (MDR) Gram-negative bacteria have been named as one of the most urgent global health threats due to the lack of effective and biocompatible drugs. Here, we show that a class of antimicrobial agents, termed 'structurally nanoengineered antimicrobial peptide polymers' (SNAPPs) exhibit sub-µM activity against all Gram-negative bacteria tested, including ESKAPE and colistin-resistant and MDR (CMDR) pathogens, while demonstrating low toxicity. SNAPPs are highly effective in combating CMDR Acinetobacter baumannii infections in vivo, the first example of a synthetic antimicrobial polymer with CMDR Gram-negative pathogen efficacy. Furthermore, we did not observe any resistance acquisition by A. baumannii (including the CMDR strain) to SNAPPs. Comprehensive analyses using a range of microscopy and (bio)assay techniques revealed that the antimicrobial activity of SNAPPs proceeds via a multimodal mechanism of bacterial cell death by outer membrane destabilization, unregulated ion movement across the cytoplasmic membrane and induction of the apoptotic-like death pathway, possibly accounting for why we did not observe resistance to SNAPPs in CMDR bacteria. Overall, SNAPPs show great promise as low-cost and effective antimicrobial agents and may represent a weapon in combating the growing threat of MDR Gram-negative bacteria.

Physicochemical and immunological assessment of engineered pure protein particles with different redox states.

The development of subunit antigen delivery formulations has become an important research endeavor, especially in cases where a whole cell vaccine approach has significant biosafety issues. Particle-based systems have shown particular efficacy due to their inherent immunogenicity. In some cases, fabrication techniques can lead to changes in the redox states of encapsulated protein antigens. By employing a uniform, well-characterized, single-protein system, it is possible to elucidate how the molecular details of particle-based protein antigens affect their induced immune responses. Using mesoporous silica-templated, amide bond-stabilized ovalbumin particles, three types of particles were fabricated from native, reduced, and oxidized ovalbumin, resulting in particles with different physicochemical properties and immunogenicity. Phagocytosis, transcription factor activation, and cytokine secretion by a mouse macrophage cell line did not reveal significant differences between the three types of particles. Oxidation of the ovalbumin, however, was shown to inhibit the intracellular degradation of the particles compared with native and reduced ovalbumin particles. Slow intracellular degradation of the oxidized particles was correlated with inefficient antigen presentation and insignificant levels of T cell priming and antibody production in vivo. In contrast, particles fabricated from native and reduced ovalbumin were rapidly degraded after internalization by macrophages in vitro and resulted in significant T cell and B cell immune responses in vivo. Taken together, the current study demonstrates how the redox state of a protein antigen significantly impacts the immunogenicity of the particulate vaccine formulations.