Nanofilm: a new drug delivery system to counteract Multi Drug Resistant Gram negative bacteria infections

Introduction: Evolution of resistance to last-resort antibiotics, such as colistin (Col), in multidrug–resistant (MDR) Gram-negative bacteria (GNB), including Pseudomonas aeruginosa, Acinetobacter baumannii and Klebsiella pneumoniae, causes a relevant problem in the treatment of different nosocomial infections as the skin and soft tissue infections (SSTI). Nanocarriers have been proposed for antibiotic delivery as a promising strategy to counteract resistant infections increasing drug efficacy and penetration. This work aimed at developing a topical formulation of colistin-loaded albumin nanoparticles for the treatment of MDR GNB SSTI infections. Materials and methods: We developed a formulation of chitosan-coated human albumin nanoparticles for the Col delivery (Col/haNPs) and evaluated the antimicrobial and antibiofilm activity as well as the biocompatibility. Starting from this preparation it was developed a topical formulation based on a film forming spray (consisting of glycerol, etilacetate and the copolymer Plastoid® B) and the Col/haNPs, called “NANOFILM”. The formulations were characterized and biocompatibility was performed by MTT assay on human fibroblasts and human skin and by haemolytic activity determined spethrophotometrically on red blood cells. The broth microdilution test was performed according to EUCAST guidelines to evaluate MIC on different MDR GNB (A. baumannii, P. aeruginosa and K. pneumoniae) and the effect on biofilm formation was assessed by crystal violet staining method. An ex vivo skin infection model was performed to evaluate the antibacterial effect of the topical formulation. Results: the Col/haNPs showed sizes lower than 200 nm, an high encapsulation efficiency and a prolonged in vitro release of Col. The safety of the nanoformulations was demonstrated by no cytotoxicity on epithelial cells and human skin and by no hemolytic activity. Both Col/haNPs and the NANOFILM can reduce the MIC values respect to free Col, in both MDR Col R and Col S strains. Moreover the Col/haNPs displayed a potent biofilm inhibition and significantly reduced the biofilm also at 1/2 MIC. By contrast Col free is able to reduce biofilm only at higher concentrations. The antimicrobial effect was also demonstrated in an ex vivo skin infection model using A. baumannii and the NANOFILM carrying the nanoparticles. Discussion and Conclusions: SSTI caused by MDR GNB are becoming increasingly prevalent and constitute a global problem because they are difficult to treat and are associated with high morbidity and mortality rates in patients with underlying immunodeficiency, as well as burn or trauma-related injuries. Our findings suggest that Col/haNPs represent a promising nanocarrier for Col topical delivery with high antimicrobial activity on MDR GNB.