Multivalent artificial opsonin for the recognition and phagocytosis of Gram-positive bacteria by human phagocytes

Abstract

Hospital-acquired infections (HAIs) remain a leading cause of death in the United States. Unfortunately, treatment of HAIs is complicated by the emergence of antibiotic-resistant bacterial strains. In an effort to enhance the body’s natural immune response to infection, we have developed an artificial opsonin to promote the recognition, phagocytosis, and destruction of pathogenic bacteria by human phagocytes. The artificial opsonin is constructed from multivalent conjugates of poly( l-lysine)- graft-poly(ethylene glycol) with vancomycin and human IgG-Fc. Our approach utilizes vancomycin’s inherent ability to bind to d-Ala- d-Ala terminated peptides present in the cell wall of Gram-positive bacteria. Here, we show that conjugation of vancomycin to PLL- g-PEG prevents its action as an antibiotic and allows vancomycin to function solely as a recognition molecule. Human IgG-Fc antibody fragment serves as a phagocyte recognition molecule and is recognized by the Fcγ cell surface receptors expressed on professional human phagocytes. Using flow cytometry, we found that a polysaccharide-encapsulated, methicillin-resistant strain of Staphylococcus epidermidis is efficiently recognized by the artificial opsonin (nearly 100% of cells were opsonized) and that opsonin binding is specific since it can be inhibited by the soluble cell wall peptide analog acetyl-Lys- d-Ala- d-Ala. Opsonization of S. epidermidis resulted in an approximate 2-fold increase in phagocytosis by a human neutrophil cell line. Notably, Enterococcus faecalis VanB, a bacterial strain with inducible vancomycin resistance, was used to show that the artificial opsonin does not unintentionally induce antibiotic resistance mechanisms.