Novel antimicrobials in fighting carbapenem-resistant Klebsiella pneumoniae

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Project Summary/Abstract: Klebsiella is a common Gram-negative pathogen causing community-acquired bacterial pneumonia, and K. pneumoniae pneumonia is considered the most common cause of hospital-acquired pneumonia in the United States. K. pneumoniae is a difficult infection to treat because of the organism's thick capsule that is usually best treated with the last line antibiotic therapy such as carbapenems. However, carbapenem-resistant K. pneumoniae (CRKP), one of the carbapenem-resistant Enterobacteriaceae (CRE), is an emerging cause of antibiotic-resistant nosocomial infections associated with high rates of morbidity and mortality. New therapies in controlling CRKP-induced infections are urgently needed. Using recombinant protein and genetically modified mouse knockout models, we have demonstrated that the short palate, lung, nasal epithelium clone (PLUNC) 1 (SPLUNC1) contributes to pulmonary host defense against K. pneumoniae induced infection. A novel SPLUNC1- derived peptide from the antimicrobial motif of the SPLUNC1 protein, α4-Short, demonstrated more potent antibacterial properties than the full-length recombinant SPLUNC1 protein and in vivo efficacy in a murine model of respiratory infection. Based on additional modifications of α4-Short, we have recently developed a series of rationally engineered antimicrobial peptides (AMPs) that rapidly kill their microbial targets by permeabilizing bacterial membrane regardless of the specific metabolic state of the bacteria. One of our lead AMPs, A4-153, has demonstrated potent bactericidal activity against diverse difficult-to-treat multidrug resistant (MDR) pathogens, including CRKP. Exciting, A4-153 is active against many CRKP that have developed resistance to other membrane-active compounds, such as the natural AMP LL37 and colistin, an antibiotic of last resort. In addition, we have found a substantially lower tendency for bacteria to develop resistance to A4-153 compared to standard antibiotic agents and natural AMPs. Importantly, we found that similar to natural AMP LL37, A4-153 displayed no detectable hemolysis and could be safely administered to mouse lungs with very high concentrations. We propose in this SBIR application to explore the feasibility of using the newly developed A4- 153 to prevent CRKP-induced pneumonia by killing CRKP and eradicating the CRKP biofilm in the abiotic and biotic system using in vitro and in vivo models. The successful completion of the proposed aims in this Phase I application will prepare us for IND-enabling studies to be presented in a subsequent Phase II application, including multidose MTD, GLP toxicity in rodents and large animals, and initiation of GMP manufacturing.