title: Targeting and blocking B7 costimulatory molecules on antigen-presenting cells using CTLA4Ig-conjugated liposomes: in vitro characterization and in vivo factors affecting biodistribution.
abstract: OBJECTIVE
  CTLA4Ig, a fusion protein of CTLA-4 and Fc of immunoglobulin (Ig) heavy chain, inhibits the essential costimulatory signal for full T cell activation via blocking the interaction between CD28 and B7 molecules and renders T cell nonresponsiveness. CTLA4Ig has been used to control deleterious T cell activation in many experimental systems. We hypothesized that by conjugating CTLA4Ig to liposomes the efficacy of CTLA4Ig could be enhanced through multivalent ligand effect, superior targetability, and modification of the fate of ligated costimulatory molecules.
  
  
  RESULTS
  Consistent with this hypothesis, liposome-conjugated CTLA4Ig bound to B7 and blocked their binding sites more efficiently than free CTLA4Ig, lowering the half maximal dose for B7 blocking by an order of the magnitude. These results were similar both in B7-1 expressing p815 cells and in activated macrophages. Moreover, CTLA4Ig-liposomes underwent rapid internalization upon cell surface binding through B7 molecules. In allogenic mixed lymphocyte reaction assays, the CTLA4Ig-liposomes were tested to show effective inhibition of T cell proliferation. In vivo, however, when CTLA4Ig-liposomes were injected into mice, a significant fraction was localized to the reticuloendothelial system (RES), presumably because of its binding to Fc receptors expressed on tissue macrophages. The Fc receptor-mediated uptake could be alleviated by coinjection of anti-FcR monoclonal antibody. In the mouse engrafted with pancreatic islets of Langerhans underneath the capsule of one kidney, despite the increased localization in RES, enhanced accumulation of CTLA4Ig-conjugated liposome was observed in the engrafted kidney compared to the contralateral kidney.
  
  
  CONCLUSIONS
  We show that the conjugation of CTLA4Ig to liposome could increase the efficiency of the targeting by increasing the binding avidity at cellular level and by increasing the concentration at the target site in in vivo system. The biodistribution and circulation time data suggested that the CTLA4Ig-liposomes could be improved upon minimizing the FcR-mediated uptake by Fc receptor-bearing cells. Thus, the strategy of conjugating CTLA4Ig to liposomes could be exploited for immune intervention in transplantation and autoimmune diseases for the efficient blocking of costimulation.
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, University of Michigan, Ann Arbor, Michigan 48109-1065, USA.
authors.forename: Chung-Gyu
authors.initials: CG
authors.lastname: Park
authors.forename: Natalie W
authors.initials: NW
authors.lastname: Thiex
authors.forename: Kyung-Mi
authors.initials: KM
authors.lastname: Lee
authors.forename: Gregory L
authors.initials: GL
authors.lastname: Szot
authors.forename: Jeffery A
authors.initials: JA
authors.lastname: Bluestone
authors.forename: Kyung-Dall
authors.initials: KD
authors.lastname: Lee
mesh_terms: D000069594:Abatacept; D000818:Animals; D000938:Antigen-Presenting Cells / Q000378:metabolism*; D018122:B7-1 Antigen / Q000378:metabolism*; D001854:Bone Marrow Cells / Q000378:metabolism; D002478:Cells, Cultured; D015536:Down-Regulation; D018796:Immunoconjugates / Q000008:administration & dosage / Q000493:pharmacokinetics*; D066298:In Vitro Techniques; D007274:Injections, Intraperitoneal; D016381:Islets of Langerhans Transplantation; D008081:Liposomes; D008264:Macrophages / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D013997:Time Factors; D014018:Tissue Distribution
chemical_list: D018122:B7-1 Antigen; D018796:Immunoconjugates; D008081:Liposomes; D000069594:Abatacept