Fusion of the Human Cytomegalovirus pp65 Antigen with Both Ubiquitin and Ornithine Decarboxylase Additively Enhances Antigen Presentation to CD8⁺T Cells in Human Dendritic Cells

Abstract

Antigenic molecules are modified for targeting to the proteasome by ubiquitin (Ub) or by a Ub-independent system such as ornithine decarboxylase (ODC) to be presented by MHC class I molecules. In this study, we compared the immunogenicity of human cytomegalovirus pp65 antigen fused with Ub and/or ODC, using RNA electroporation of human dendritic cells. Among the C-terminal mutants of Ub (G76, A76, and V76), Ub(G) showed the best ability to enhance the degradation of a target protein and stimulate T cells. The pp65 antigens fused with either Ub(G) or ODC enhanced the stimulation to CD8(+) T cells, and the effects of Ub(G) and ODC were similar. Furthermore, the fusion of both Ub and ODC additively increased immunogenicity compared with the single-fusion proteins. The fusion of Ub(G) and ODC enhanced primarily the stimulation of CD8(+) rather than CD4(+) T cells and more efficiently induced pp65-specific T cells in vitro. These additive effects of Ub and ODC in antigen processing may provide improved strategies to stimulate CD8(+) T cells for the development of immunotherapies against the variety of viral diseases and cancers.

GM-CSF Promotes the Expansion and Differentiation of Cord Blood Myeloid-Derived Suppressor Cells, Which Attenuate Xenogeneic Graft-vs.-Host Disease

“…Therefore, we used mononuclear cells from a healthy volunteer donor that has been shown to have a high T cell immune response to the HCMV pp65 antigen. It has been established in our previous study that DCs electroporated with mRNA encoding HCMV pp65 antigens present antigen to T cells through HLA class I and class II molecules after antigens processing, so T cell responses to whole antigens are measured rather than specific HLA-restricted antigen epitopes (28).…”

Simultaneousin vitrogeneration of CD8 and CD4 T cells specific to three universal tumor associated antigens of WT1, survivin and TERT and adoptive T cell transfer for the treatment of acute myeloid leukemia

“…LAA-specific CTLs were generated using autologous DCs as previously described [ 54 ]. Briefly, purified CD8 and Th1-polarized CD4 T cells were stimulated with WT1 , WT1 - survivin or TERT IVT mRNA-electroporated DCs.…”

Engineering Dendritic Cells to Enhance Cancer Immunotherapy

“…Moreover, when the antigen was linked to both ubiquitin and ornithine decarboxylase, immunogenicity was further increased. 13 Recently, we have observed that the immunogenicity of a given TAA can be greatly enhanced by deleting its nuclear localization signal (D. Benteyn, S. Anguille, A.M.T. Van Nuffel, C. Heirman, J. Corthals and W. Waelput, unpublished results), demonstrating that further manipulation of the TAAencoding sequence can result in favorable induction of potent antitumor immune responses.…”