Cloning and characterization of recombinant rhesus macaque IL-10/Fc(ala-ala) fusion protein: a potential adjunct for tolerance induction strategies

Cytokine. 2007 Dec;40(3):183-92. doi: 10.1016/j.cyto.2007.09.008. Epub 2007 Nov 5.

Abstract

The powerful anti-inflammatory and immunosuppressive activities of IL-10 make it attractive for supplemental therapy in translational tolerance induction protocols. This is bolstered by reports of IL-10-mediated inhibition of innate immunity, association of human stem cell and nonhuman primate (NHP) islet allograft tolerance with elevated serum IL-10, and evidence that systemic IL-10 therapy enhanced pig islets survival in mice. IL-10 has not been examined as adjunctive immunosuppression in NHP. To enable such studies, we cloned and expressed rhesus macaque (RM) IL-10 fused to a mutated hinge region of human IgG1 Fc to generate IL-10/Fc(ala-ala). RM IL-10/Fc(ala-ala) was purified to approximately 98% homogeneity by affinity chromatography and shown to be endotoxin-free (<0.008 EU/microg protein). The biological activity of IL-10/Fc(ala-ala) was demonstrated by (1) costimulation of the mouse mast cell line, MC/9 proliferation in a dose-dependent fashion, (2) suppression of LPS-induced septic shock in mice and (3) abrogation of LPS-induced secretion of proinflammatory cytokines/chemokines in vitro and in vivo in NHP. Notably, RM IL-10/Fc(ala-ala) had significantly greater potency than human IL-10/Fc(ala-ala) and exhibited a circulating half-life of approximately 14 days. The availability of this reagent will facilitate definitive studies to determine whether supplemental therapy with RM IL-10/Fc(ala-ala) can influence tolerance outcomes in NHP.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Line
  • Cell Proliferation / drug effects*
  • Cloning, Molecular
  • Dose-Response Relationship, Drug
  • Graft Survival / drug effects
  • Graft Survival / immunology
  • Humans
  • Immunoglobulin Fc Fragments / genetics
  • Immunoglobulin Fc Fragments / immunology
  • Immunoglobulin Fc Fragments / pharmacology*
  • Immunoglobulin G / genetics
  • Immunoglobulin G / immunology
  • Immunoglobulin G / pharmacology
  • Insulin-Secreting Cells / immunology
  • Insulin-Secreting Cells / transplantation
  • Interleukin-10 / genetics
  • Interleukin-10 / immunology
  • Interleukin-10 / pharmacokinetics
  • Interleukin-10 / pharmacology*
  • Lipopolysaccharides / toxicity
  • Macaca mulatta
  • Mast Cells / immunology*
  • Mice
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / immunology
  • Recombinant Fusion Proteins / pharmacokinetics
  • Recombinant Fusion Proteins / pharmacology*
  • Shock, Septic / chemically induced
  • Shock, Septic / drug therapy*
  • Shock, Septic / immunology
  • Swine
  • Transplantation Tolerance / drug effects*
  • Transplantation, Heterologous

Substances

  • Immunoglobulin Fc Fragments
  • Immunoglobulin G
  • Lipopolysaccharides
  • Recombinant Fusion Proteins
  • Interleukin-10