title: Tolerance induction via a B-cell delivered gene therapy-based protocol: optimization and role of the Ig scaffold.
abstract: Our previous studies indicated that antigen-specific tolerance could be achieved by the injection of LPS-activated B-cell blasts that were retrovirally gene-transferred with an IgG-antigen fusion construct. This system was shown to be effective for tolerance induction with a variety of inserted antigens ranging in size from a single peptide to a large multi-epitope protein in a variety of mouse strains. Moreover, it was shown to be effective in four animal models for human disease. To optimize the existing protocol, establish the role of the IgG H chain scaffold, and provide baseline for potential clinical applications, we examined the effects of different B-cell activators, including lipopolysaccharide (LPS), anti-CD40, CpG oligodeoxynucleotide (CpG-ODN), and anti-IgM plus IL-4, on B-cell proliferation, GFP transduction efficiency, and tolerance induction in vivo. The results show that all activators except CpG-ODN have similar effects on retroviral gene transfer and peptide-IgG-induced tolerance. Furthermore, dose-response analyses showed that T-cell tolerance could be induced with 10(5) peptide-IgG LPS B-cell blasts, but that 10(6) transduced B-cells were needed for humoral unresponsiveness. Transduced anti-IgM-induced blasts were tolerogenic at 10(6) cells, but no dose of transduced CpG blasts was tolerogenic. Finally, to examine the role of IgG scaffold, a retroviral construct encoding lambda repressor p1-102 and signal peptide of murine IgG heavy chain was engineered to allow secretion of the p1-102 domain in the same manner as that of p1-102-IgG fusion protein. The results demonstrate that not only is IgG scaffold important in tolerance induction and maintenance of the long-lasting immune hyporesponsiveness, but assembly of the IgG heterodimer may be required for the efficacy of this system.
authors.affiliation: Department of Surgery and of Microbiology and Immunology, University of Maryland School of Medicine, Rockville, MD 20855, USA.
authors.forename: Tie Chi
authors.initials: TC
authors.lastname: Lei
authors.forename: Yan
authors.initials: Y
authors.lastname: Su
authors.forename: David W
authors.initials: DW
authors.lastname: Scott
mesh_terms: D019264:Adoptive Transfer; D000818:Animals; D001402:B-Lymphocytes / Q000276:immunology / Q000637:transplantation*; D015316:Genetic Therapy; D005822:Genetic Vectors; D007108:Immune Tolerance / Q000235:genetics*; D007074:Immunoglobulin G / Q000235:genetics* / Q000502:physiology; D007143:Immunoglobulin Heavy Chains / Q000235:genetics* / Q000502:physiology; D008213:Lymphocyte Activation / Q000235:genetics; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008979:Moloney murine leukemia virus / Q000235:genetics / Q000276:immunology; D010455:Peptides / Q000235:genetics / Q000276:immunology; D014161:Transduction, Genetic
chemical_list: D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D010455:Peptides