Engineering recombinant lubricin to combat orthopedic infection

项目摘要/摘要

PROJECT SUMMARY / ABSTRACT This application seeks funding to support a NIAMS K08 recipient during her transition to research independence. The broad objective is to investigate how lubricin attenuates orthopedic biofilms and to determine whether distinct lubricin O-glycans mediate these anti-biofilm properties. The ability of many orthopedic implant-associated bacteria to form biofilms renders these organisms resistant to antimicrobial therapy. Once established, implant- associated biofilms typically require debridement, implant exchange or implant removal and can lead to severe functional deterioration necessitating arthrodesis or even amputation. Therefore, prevention and treatment of orthopedic-associated biofilms has been identified as a critical research priority in musculoskeletal infection. Recent work has identified mucin biopolymers as potent antagonists of biofilm assembly and virulence gene expression, raising the possibility that recombinant lubricin therapeutics could be an effective strategy for clinically relevant biofilms. Our preliminary data demonstrate that the recombinant lubricin developed as part of the applicant’s K08 research not only inhibits biofilm formation but also dissipates established P. aeruginosa biofilms in vitro. In the current proposal, we will investigate the hypothesis that the dual properties of surface adhesion (N- and C-terminal domains) and anti-adhesion (mucin O-glycan domains) make lubricin an ideal anti- biofilm agent. In Aim 1, we will investigate the mechanisms by which lubricin inhibits biofilm formation and induces dispersion of established biofilms in growth media, in synovial fluid, and on titanium alloy (Ti6Al4V). In Aim 2, we will determine whether distinct O-glycans mediate these anti-biofilm functions and whether lubricin functionality could be optimized through glycoengineering. As part of Aim 2, we will manipulate the expression of key glycosyltransferase enzymes to produce recombinant lubricin with altered compositions of Core-1, Core- 2 and sialylated O-glycan structures. This proposal builds upon the applicant’s K08 research, in which lubricin has been primarily investigated as a tribological agent for the treatment of post-traumatic osteoarthritis. By investigating a new, anti-biofilm application for lubricin therapy, this award will enable Dr. Reesink to branch into a new sub-discipline of orthopedic research with the potential to integrate with the applicant’s clinical veterinary expertise in small and large animal models of orthopedic infection. Dr. Reesink’s proposal is supported by an exceptional team, including experts in glycoengineering, molecular characterization of bacteria-surface interactions, and in vivo models of periprosthetic joint infection and clinical translation.