FcRn-enabling strategies for improved thrombolytic therapy

项目总结/文摘

PROJECT SUMMARY/ABSTRACT While clinical use of therapeutic proteins has grown exponentially in recent decades, utility is often limited by unfavorable pharmacokinetics (PK), mediated by rapid elimination. One approach to overcome this limitation is half-life extension (HLE), achieved by attachment of biotherapeutics to polymers (PEG), plasma proteins (albumin, IgG), and blood cells. There is a paucity of data describing the impact of properties of cargo proteins and HLE-conferring ligands on PK of proteins tested using these strategies. One area where HLE has the potential to advance clinical therapy is acute, life-threatening, thrombosis (pulmonary embolism, ischemic stroke, etc.). The only pharmacologic treatment available for treatment of this condition is infusion of plasminogen activators, which have an unfavorable pharmacologic profile, mediated by a half-life of minutes, rapid inactivation in plasma, and severe adverse effects (e.g. hemorrhagic transformation). Pilot data demonstrates that conjugation of a derivative of urokinase selectively activated in thrombin-rich pathological thrombi (scuPA-T) to an albumin-binding nanobody (Nb) leads to PK that is identical to RBC-binding scuPA-T, an approach pioneered by Drs. Muzykantov and Cines, albeit with distribution in the plasma rather than the cellular fraction of blood. The central hypothesis of this proposal is that coupling of scuPA-T to albumin-binding ligands will provide prolonged circulation, mediated by the neonatal Fc receptor (FcRn), and selective thrombolysis of pathological thrombi, sparing hemostatic plugs from lysis. In the mentored stage, the impact of properties of the cargo drug on FcRn-enabled HLE will be identified, using FcRn knockout mice to directly elucidate the FcRn-mediated component of HLE conferred by albumin binding (Aim 1; K99). Based on pilot data, safety and efficacy of albumin-binding scuPA-T will be studied in a mouse model of pulmonary embolism. These results will be used to develop a predictive modeling platform that will be used to for further engineering and PK optimization of HLE-scuPA-T constructs (Aim 2; K99). With an eye on mechanistic and translational advancement of this strategy, the role of albumin-binding affinity on the PK of scuPA-T will be defined, using a newly developed library of nanobodies (Aim 3; R00). Additionally, thrombin-cleavable HLE ligands will be devised, permitting selective release of scuPA-T in thrombi, improving diffusion into clots and lysis (Aim 3; R00). Overarching themes of this proposal include identification of critical features of albumin-mediated HLE and mechanism-based modeling to guide optimization and reengineering of protein therapeutics. A mentoring team has been identified with expertise spanning the areas of research in this grant, namely, thrombosis and hemostasis, antibody engineering, and mathematical modeling of biological systems. Mentored research will be conducted at the University of Pennsylvania, which has a highly collegial and collaborative faculty and extensive resources available to conduct the proposed research. This proposal is geared towards gaining the expertise necessary to be successful as a tenured faculty member conducting high quality biomedical research.