Immunization of mice with human dopachrome tautomerase (hDCT) provides greater protection against melanoma than immunization with the murine homologue (mDCT). We mapped the CD8 + and CD4 + T-cell epitopes in both proteins to better understand the mechanisms of the enhanced protection. The dominant CD8 + T-cell epitopes were fully conserved between both proteins, yet immunization with hDCT produced frequencies of CD8 + T cells that were 5-to 10-fold higher than immunization with mDCT. This difference was not intrinsic to the two proteins because comparable frequencies of CD8 + T cells were elicited by both antigens in DCT-deficient mice. Strikingly, only hDCT elicited a significant level of specific CD4 + T cells in wild-type (WT) mice. The murine protein was not devoid of CD4 + T-cell epitopes because immunization of DCT-deficient mice with mDCT resulted in robust CD4 + T-cell immunity directed against two epitopes that were not identified in WT mice. These results suggested that the reduced immunogenicity of mDCT in WT mice may be a function of insufficient CD4 + T-cell help. To address this possibility, the dominant CD4 +

We have investigated the therapeutic potential of a prototypic melanoma vaccine based on recombinant adenovirus expressing human dopachrome tautomerase in the B16F10 murine melanoma model. We found that in the presence of a tumor, the magnitude of T-cell immunity evoked by the vaccine was significantly reduced. This impairment was compounded by defects in cytokine production and degranulation within the tumor-infiltrating lymphocytes (TILs). We showed that the combination of vaccination with high-dose cyclophosphamide was able to skew the response toward the target antigen and enhanced both the quantity and quality of antigen-specific CD8 þ and CD4 þ T-cell responses in tumor-bearing mice, which resulted in the inhibition of tumor growth. Furthermore, when tumor-specific antigens were targeted by the vaccine, the combination therapy could actually produce tumor regression, which appeared to result from the high frequency of antigen-specific T cells. These data show that recombinant adenovirus vaccines are compatible with conventional high-dose chemotherapy and that the combined treatment results in improved therapeutic outcomes relative to either agent individually.

<div>Abstract<p>Immunization of mice with human dopachrome tautomerase (hDCT) provides greater protection against melanoma than immunization with the murine homologue (mDCT). We mapped the CD8⁺ and CD4⁺ T-cell epitopes in both proteins to better understand the mechanisms of the enhanced protection. The dominant CD8⁺ T-cell epitopes were fully conserved between both proteins, yet immunization with hDCT produced frequencies of CD8⁺ T cells that were 5- to 10-fold higher than immunization with mDCT. This difference was not intrinsic to the two proteins because comparable frequencies of CD8⁺ T cells were elicited by both antigens in DCT-deficient mice. Strikingly, only hDCT elicited a significant level of specific CD4⁺ T cells in wild-type (WT) mice. The murine protein was not devoid of CD4⁺ T-cell epitopes because immunization of DCT-deficient mice with mDCT resulted in robust CD4⁺ T-cell immunity directed against two epitopes that were not identified in WT mice. These results suggested that the reduced immunogenicity of mDCT in WT mice may be a function of insufficient CD4⁺ T-cell help. To address this possibility, the dominant CD4⁺ T-cell epitope from hDCT was introduced into mDCT. Immunization with the mutated mDCT evoked CD8⁺ T-cell frequencies and protective immunity comparable with hDCT. These results reveal a novel mechanism by which xenoantigens overcome tolerance. Our data also suggest that immunologic tolerance is more stringent for CD4⁺ T cells than CD8⁺ T cells, providing a mechanism of peripheral tolerance where autoreactive CD8⁺ T cells fail to be activated due to a lack of autoreactive CD4⁺ T cells specific for the same antigen. [Cancer Res 2007;67(13):6459–67]</p></div>

Supplementary Figures 1-2 from Elevated Frequencies of Self-reactive CD8⁺ T Cells following Immunization with a Xenoantigen Are Due to the Presence of a Heteroclitic CD4⁺ T-Cell Helper Epitope

Supplementary Figures 1-2 from Elevated Frequencies of Self-reactive CD8⁺ T Cells following Immunization with a Xenoantigen Are Due to the Presence of a Heteroclitic CD4⁺ T-Cell Helper Epitope