Project Summary Multi-drug resistant (MDR) Gram-negative bacterial infections are an ongoing challenge to public health. Indeed, 4 of the 6 “ESKAPE” pathogens – recently highlighted as responsible for the majority of hospital acquired infections – are Gram-negative pathogens. Although it is clear that novel antibiotics for Gram-negative infections are desperately needed, there has been minimal progress in this regard, and it has been over five decades since a new class of drugs have been introduced for Gram-negative bacteria. The development of new antibiotics to treat these pathogens is complicated by the fact that Gram-negative bacteria have an impenetrable membrane that confers intrinsic resistance to antimicrobial agents. The broad objective of this program is to study glycoconjugates to combat Gram-negative pathogens. We have made two major discoveries that suggests human milk oligosaccharides (HMOs) may be transformative in this regard. First, HMOs cause major changes to the behavior of bacteria, with strong effects on growth and the formation of biofilms, architectures that aid in bacterial survival. We have also observed that HMOs function as potent adjuvants, potentiating the activity of intracellular-targeting antibiotics by increasing cell permeability. These two discoveries form the foundation of the projects proposed in this application. In Project 1 we seek to characterize the impact of HMOs on Gram-negative causes of microbiome imbalance. In Project 2 we will explore HMOs in combination therapies against A. baumannii, an important Gram- negative pathogen. In Project 3 we investigate the chemistry and biochemistry of the mollemycin glycopeptides, a rare glycopeptide with antimicrobial activity against Gram-negative pathogens. While not sourced from milk, we plan to leverage our experience in human milk science to study the biochemistry of the mollemycins. A significant output of this work is a mechanistic understanding of the types of compounds that can enter Gram-negative bacteria.

Project Narrative Drug resistance presents a significant threat to public health and encompasses all major microbial pathogens and antimicrobial agents. Some pathogens have acquired resistance to multiple antibiotics and cause infections that are nearly untreatable. Gram-negative bacterial infections are the most difficult organisms to treat due to difficulty for drug candidates to penetrate and accumulate inside their cells. Accordingly, the development of novel approaches to treat Gram-negative pathogens is an area of serious medical need. The goal of this project is to use chemical synthesis and microbiology to discover novel compounds with potent antimicrobial activity.