A rapid non-traditional approach for developing biologicals against drug- resistant bacteria and yeasts.

Abstract

Development of effective, long-lasting antibiotics is a challenge due to the rate at which the pathogens acquire resistance, prompting the major pharma players to omit them from their portfolio. We present a non-traditional approach for rapidly developing anti-infective biologicals that is resilient to resistance. Antibody-drug conjugates were developed against three different pathogens Pseudomonas aeruginosa (ABG 14), Staphylococcus aureus (ABG 16) and Candida. albicans (ABG 07) that were extremely specific and effective in neutralising them irrespective of their drug resistance profiles. Neutralising camelid antibody fragment (VHH) were isolated from immunised camel libraries by phage display that effectively neutralised the pathogens at a MIC 99 of 125 {micro}g mL-1 in vitro. Antimicrobial peptides (AMP) were then conjugated to them by pathogen-specific cleavable linkers and the resultant molecules were 10 -20 times more effective due to a dual mode of action-the inhibitory action of the VHH on surface transporters and enzymes and the activity of the AMPs released by the pathogen surface proteases. Called AbTids, these molecules were extremely specific, stable in plasma and were activated in the presence of 104 to 105 CFU mL-1 of the pathogens and had an efficacy in the sub-micromolar range (6.25-12.5 {micro}g mL-1, 250 nM) inhibiting their growth within 2 hrs. of administration. They were economically and efficiently produced in microbial production system as a single chain fusion protein. One of the molecules ABG 14 was characterised further and found to be pathogen specific with negligible frequency of resistance, was non-toxic, had the capability to destroy biofilms, and cleared a systemic infection in a mouse with a carbapenem resistant strain of P. aeruginosa at a dose of 5 mg kg-1. This strategy can be used to generate new hits against medically important pathogens in a matter of weeks by simply shuffling the components, using different VHHs and AMPs, bypassing years of expensive drug development efforts that can potentially rejuvenate drug discovery efforts against emerging superbugs.