DEVELOPMENT OF A PREDICTIVE MODERATE THROUGHPUT ASSAY TO SCREEN NOVEL DESIGNER PROLINE-RICH ANTIMICROBIAL PEPTIDE CHAPERONE PROTEIN INHIBITORS (DPCS) AGAINST MULTI-DRUG RESISTANT PATHOGEN

Multi-drug resistant (MDR) bacterial infections represent a significant threat to public health. It is estimated thatthe impact of antibiotic resistance will continue to grow resulting in a global economic burden of $100 trillion andcausing 10 million deaths annually. Many of the therapeutics currently under development represent traditionalantibiotic approaches or next generation antibiotics that will likely suffer a limited market lifespan due to the rapidemergence of resistance. To effectively address antibiotic resistance and have therapeutic options that areeffective, approaches that utilize new mechanisms of action must be explored.Arrevus is developing a novel approach to addressing MDR bacterial infections using Designer Proline-richantimicrobial peptide Chaperone protein inhibitors (DPCs), derived from insects and selectively modified, actingas inhibitors to one of the critical bacterial proteins responsible for bacterial protein folding, DnaK. Preliminarystudies have demonstrated the potential of DPCs as antibiotic potentiating agents against MDR gram-positiveand gram-negative bacterial pathogens. Our efforts have shown that DPCs: 1) enhance antibiotic activity; 2)reduce bacterial burden in systemic infection models; 3) have favorable preliminary safety profiles; and 4) providean enhancement to antimicrobial agents through a novel mechanism of action. While early efforts to developDPCs has been successful, the pace of the programs and expansion into additional indications has beenhindered due to the lack of reliable in vitro screening assays that correlate with in vivo efficacy. DPCs that havedisplayed potency using minimum inhibitory assays have failed in subsequent in vivo efficacy studies.The objective of the proposed Phase I program is to develop and validate an in vitro assay for DPC activity. InAim 1, a scratch test assay using human alveolar and keratinocyte cells will be characterized for severalendpoints, including scratch closure, cytokine production, and apoptosis in the presence of well-characterizedDPCs and antibiotics to develop an assay that is predictive of in vivo DPC activity. In Aim 2, the assay developedin Aim 1 will then be used to inform a medical chemistry program that is designed to identify DPCs with activityagainst methicillin-resistant Staphylococcus aureus. Two lead DPCs identified using the using the assay will thenbe evaluated for in vivo efficacy using a standard thigh infection model. Successful completion of the proposedprogram will provide proof-of-concept for the use of the developed in vitro assay for detecting DPC activity thatis predictive of in vivo activity and will support a Phase II program that will expand the assay to use with gram-negative pathogens and evaluate lead DPCs in indication-specific models.