Pharmacokinetics and delivery of proteins modified with FcRn binding ligands
- Author(s): Sockolosky, Jonathan
- Advisor(s): Szoka, Jr., Francis C
- et al.
The importance of therapeutic recombinant proteins in medicine has led to a variety of tactics to increase their circulation time or to enable routes of administration other than injection. One clinically successful tactic to improve protein circulation and delivery is to fuse the Fc-domain of immunoglobulin G (IgG) to therapeutic proteins so that the resulting fusion proteins interact with the neonatal Fc receptor (FcRn). Although successful, Fc-fusion proteins significantly increase molecular weight thereby restricting tissue penetration, decrease protein function, and are limited to mammalian expression systems. As an alternative to grafting the high molecular weight Fc-domain to therapeutic proteins, we have modified their N- and/or C-terminus with a short peptide sequence that interacts with FcRn. Our strategy was motivated by results from Mezo and coworkers [Mezo et al. (2008) PNAS 105:2337-42] who identified peptides that compete with human IgG for FcRn. The small size and simple structure of the FcRn binding peptide (FcBP) allows for expression of FcBP fusion proteins in E. coli and results in their pH-dependent binding to FcRn with an affinity comparable to that of hIgG1. The FcBP fusion proteins are internalized, recycled and transcytosed across cell monolayers that express FcRn. Although FcBP fusion proteins mimic the human IgG1 interaction with human FcRn in vitro, the half-life of FcBP fusion proteins in wild type C57BL/6J and human FcRn transgenic mice is independent of FcRn binding, whereas the half-life of human and mouse IgG1 correlate with their species matched FcRn-binding affinity. These results promoted a detailed investigation into potential factors that may contribute to the lack of correlation between the in vitro and in vivo results including: serum stability, serum competition, renal clearance, and the function of the hybrid human-mouse FcRn/B2m receptor; however, no single variable explains the FcRn-independent half-life of FcBP fusion proteins. We speculate that an additional component(s), in collaboration with FcRn, regulates IgG homeostasis in vivo and is not replicated by FcBP fusion. If such a novel FcRn regulatory component exists, it would have a significant impact on FcRn biology and provide new opportunities to engineer IgG or alternative FcRn-targeted molecules for enhanced serum persistence.