PROJECT SUMMARY Infectious diseases are a growing threat to public health owing to increasing antimicrobial resistance (AMR) and stagnation in new antibiotic development. Left unchecked, the annual number of deaths attributable to AMR is estimated to reach 10 million by 2050, exceeding deaths due to cancers and diabetes. Thus, there is an urgent need to develop innovative approaches to tackle this serious global crisis. We aim to develop a new class of dual-stimuli responsive polysaccharide-coated nanoparticles (NP) capable of encapsulating a wide range of FDA-approved antibiotics to effectively treat multidrug-resistant (MDR) bacterial infections. The polysaccharide NP shell ensures good stability and long blood circulation time, thus leading to high NP accumulation in the infected tissues via the enhanced permeation and retention effect. Furthermore, polysaccharides enable the NP to physically bind the pathogens due to multivalent affinity for bacterial lectins. The uniquely engineered NP is activated by high levels of ROS and/or low pH in the inflammatory microenvironment to release both cationic antimicrobial polymers and antibiotics that show a strong synergy to combat MDR pathogens. The cationic polymers can induce pores on the bacterial cell membrane, and significantly diminish the intrinsic resistance of the pathogens by enhancing the transport of antibiotics into the bacteria and allowing them to bypass the efflux pump. The cationic polymers released in the infected tissues can also agglomerate the pathogens and shape a microenvironment entrapping a high level of antimicrobial materials, thus leading to high antimicrobial efficacy. Moreover, the NP can penetrate through bacterial biofilms, and enhance the uptake of antibiotics by macro- phages, thereby effectively eliminating notoriously challenging biofilm and intracellular infections, respectively. Finally, the cationic polymer contains GSH-cleavable bonds in its main chain, which can be readily degraded in the cytosol of mammalian cells, thereby sidestepping the problem of dose-limiting toxicity with other cationic polymers. Following on our successful pilot studies, we will systematically optimize and characterize NPs tailored to treat four different MDR pathogens. In Aim 1, we will determine the optimal polysaccharide NP shell, antibiotics, and NP formulation for each of the four MDR pathogens. In Aim 2, we will study the candidate NPs’ antimicrobial and antibiofilm efficacy, drug resistance development profile, and biocompatibility to gain a fundamental understanding of the design rules for efficacious and safe antimicrobial NP against pathogens of interest. In Aim 3, we will determine the maximum tolerated dose, systemic toxicity, immunological consequences, in vivo biodistribution, pharmacokinetics, and antimicrobial efficacy of the selected NPs in healthy mice and three clinically relevant animal infection models. Altogether, this study will lead to a new class of antimicrobial ...