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 innovative, highly efficient, and biocompatible pH- or ROS-responsive antimicrobial polymer- drug (i.e., antibiotics) conjugates (PDCs), which can effectively treat serious infectious diseases and overcome AMR while ensuring high biocompatibility. We will accomplish this goal utilizing existing FDA-approved antibiotics, disease-specific stimuli, and a uniquely engineered biocompatible cationic polymer. Cationic polymers can be effective antibiotic carriers as they can induce pores on the bacterial wall/membrane, thus significantly enhancing the transport of antibiotics into the bacteria and allowing them to bypass the efflux pump in the bacterial membrane. Cationic PDCs can also (1) stick to the bacteria’s surface, thereby serving as a drug reservoir to release drug locally, and (2) effectively infiltrate bacterial biofilms, thereby leading to deeper antibiotic penetration. The strong synergistic effects between cationic polymers and antibiotics diminish the intrinsic resistance of the pathogens, thus leading to significantly enhanced antimicrobial efficacy, especially for AMR pathogens. Antibiotics will be conjugated onto the cationic polymer via pH- or ROS-responsive linkers as the inflammatory microenvironment in infected tissues have low pH levels and high levels of reactive oxygen species (ROS). Furthermore, we engineered a GSH-cleavable and charge-reversal cationic polymer that can greatly reduce its systemic toxicity as well as cellular toxicity for mammalian cells. Lastly, PDC capable of stimuli (disease-specific)- controlled drug release can accumulate preferentially at the infected tissues due to the enhanced permeation and retention (EPR) effect, thereby further reducing systemic toxicity while achieving high antimicrobial efficacy. In Aim 1, we will design, synthesize and characterize pH- and ROS-responsive PDCs. We will first investigate the synergy between a number of free (i.e., before conjugation) FDA-approved antibiotics and our uniquely designed stimuli-responsive and charge-reversal biocompatible cationic polymer. In Aim 2, the antimicrobial and antibiofilm efficacies, drug resistance development profiles, and biocompatibilities of the resulting stimuli- responsive PDCs will be evaluated in multiple bacteria species. In Aim 3, we will systematically determine the maximum tolerated dose, in vivo biodistribution, antimicrobial efficacy, and potential systemic toxicity of the selected PDCs in three clinically relevant bacterial infection mouse models. This study will create a new class of PDCs bas...