Improved Pharmacokinetics and Reduced Antibody Reactivity of Lysostaphin Conjugated to Polyethylene Glycol

Lysostaphin is a 27-kDa endopeptidase that enzymatically disrupts the cell wall of Staphylococcus aureus and is a promising candidate for treating S. aureus blood-borne infections. It would be extremely useful to define conditions that would both increase lysostaphin's in vivo half-life to allow for more effective tissue distribution and reduce its immunogenicity. Conjugation of polyethylene glycol (PEG) to lysostaphin (PEGylation) was investigated as a means to accomplish these goals. Rather than using linear forms of PEG, branched PEGs were chosen as the initial candidates because their large spatial volumes prevent entry of the polymer into the enzyme's active sites, which could potentially reduce enzymatic function. Enzymatic activity for most PEGylated lysostaphins was reduced, but these compounds were still considerably active compared to unconjugated lysostaphin, with conjugates that had lower degrees of PEG modification having greater activity than those with higher degrees. PEGylated lysostaphin injected intravenously had a serum drug half-life of up to 24 h and resulted in much higher plasma drug concentrations than an equal dose of unconjugated lysostaphin, which had a half-life of less than 1 h. Finally, reduced binding affinity was shown for PEGylated lysostaphin in an antilysostaphin capture enzyme-linked immunosorbent assay, with some PEG-lysostaphin conjugates having binding affinities that were reduced more than 10-fold compared to unconjugated lysostaphin. These findings demonstrate that PEGylation of lysostaphin, while diminishing its S. aureus killing activity, results in prolonged serum drug persistence and reduced antibody binding. These features should significantly enhance lysostaphin's therapeutic value as an intravenous "antibiotic" against S. aureus.