A Fusion Protein Consisting of IL-16 and the Encephalitogenic Peptide of Myelin Basic Protein Constitutes an Antigen-Specific Tolerogenic Vaccine That Inhibits Experimental Autoimmune Encephalomyelitis

Abstract

To test a novel concept for the generation of tolerogenic vaccines, fusion proteins were constructed encompassing a tolerogenic or biasing cytokine and the major encephalitogenic peptide of guinea pig myelin basic protein (GPMBP; i.e., neuroantigen or NAg). The cytokine domain was predicted to condition APC while simultaneously targeting the covalently linked encephalitogenic peptide to the MHC class II Ag processing pathway of those conditioned APC. Rats were given three s.c. injections of cytokine-NAg in saline 1–2 wk apart and then at least 1 wk later were challenged with NAg in CFA. The rank order of tolerogenic activity in the Lewis rat model of EAE was NAgIL16 > IL2NAg > IL1RA-NAg, IL13NAg ≥ IL10NAg, GPMBP, GP69–88, and saline. NAgIL16 was also an effective inhibitor of experimental autoimmune encephalomyelitis when administered after an encephalitogenic challenge during the onset of clinical signs. Covalent linkage of the NAg and IL-16 was required for inhibition of experimental autoimmune encephalomyelitis. These data identify IL-16 as an optimal cytokine partner for the generation of tolerogenic vaccines and indicate that such vaccines may serve as Ag-specific tolerogens for the treatment of autoimmune disease.

Experimental Autoimmune Encephalomyelitis in Lewis rats: IFN-β Acts As a Tolerogenic Adjuvant for Induction of Neuroantigen-Dependent Tolerance

“…2D) or the synthetic gp69 -88 peptide (not shown). These data provide evidence that the cytokine domain may facilitate presentation of the NAg by professional APC, as was described for the IL4-NAg, IL2-NAg, and NAg-IL16 fusion proteins (25)(26)(27). Although the antigenic potency of IFN␤-NAg appeared greater than GPMBP as measured by the concentration eliciting a half-maximal response, the peak proliferative response stimulated by IFN␤-NAg was a Rats were pretreated with saline (first row), with 1 nmol of NAg (synthetic peptide gp69 -88) in saline (second row), or with 1 nmol of IFN␤-NAg in saline (third row) on days Ϫ21, Ϫ14, and Ϫ7 and were challenged with 50 g of DHFR-NAg in CFA on day 0.…”

“…Several Ag-specific therapies are being developed, including those based on altered peptide ligands, DNA vaccines, and mucosal Ag delivery. Cytokine-Ag fusion proteins were originally developed for vaccination against cancer and infectious agents but have also been explored as tolerogenic vaccines based on the use of inhibitory, antiinflammatory, or tolerogenic cytokines as the cytokine fusion partner (25,26). Two cytokine-Ag fusion proteins in which the IL-2 or IL-16 cytokines were fused to the encephalitogenic deterEAE.…”

“…EAE was induced in Lewis rats by injection of an emulsion containing 50 g of dihydrofolate reductase (DHFR)-NAg (25,26) in CFA (200 g of Mycobacterium tuberculosis) in a total volume of 0.1 ml. DHFR-NAg was comprised of the mouse DHFR as the N-terminal domain and the encephalitogenic gp69 -87 peptide of GPMBP as the C-terminal domain.…”

“…Previous studies of IL2-NAg and NAg-IL16 fusion proteins showed that covalent linkage of the cytokine and NAg domains was necessary for inhibition of EAE (25,26). That is, administration of a fused cytokine-NAg protein was tolerogenic, whereas administration of cytokine and NAg as separate molecules to the same inoculation site was without activity.…”

GM-CSF–Based Fusion Cytokines as Ligands for Immune Modulation

“…With regard to the IL-16/MBP fusion, IL-16 was previously found to inhibit IL-2 production by CD4 + T cells by interfering with CD4's interaction with MHC class II (12, 13). It is unclear how this could induce Ag-specific tolerance, but the investigators speculated that APCs may be involved, because the IL-16/MBP fusion was effective at having APCs present MBP through MHC class II to Rsl.11 T cells (11). Blanchfield and Mannie (14) further explored how cytokines could be fused to more effectively deliver MBP to APCs by combining it with GM-CSF, finding that this fusion was also capable of significantly reducing the severity of EAE in rats.…”