Supplementary Figure 1. Construction of two conventional WT1 DNA vaccines and generation of WT1-expressing AB1 malignant mesothelioma cell line. Supplementary Figure 2. Construction of HIV-1 GAG-expressing AB1 malignant mesothelioma cell line. Supplementary Figure 3. Importance of inducing p24-specific cellular responses for AB1-GAG tumor rejection. Supplementary Figure 4. Anti-tumor effect from CD8+ T cells was cross-confirmed in a B16F10 melanoma mouse model. Supplementary Figure 5. Secreted cytokines from CD8+ T cells significantly arrested AB1-GAG tumor cells at G1 phase. Supplementary Figure 6. Gating strategies for flow cytometric scatter plots. Supplementary Figure 7. Analysis of CD8+ T cell subsets in spleen and tumor. Supplementary Figure 8. Analysis of Th2 and exhaustion markers on CD8+ T cells in spleen and tumor. Supplementary Figure 9. Rapid increase of tumor-induced PD-1+CD8+ T, Tim-3+CD8+ T, MDSCs and CD4+ Treg cells. Supplementary Figure 10. Splenic and tumor-infiltrating CD8+ T cells exhibit distinct phenotypic and functional characteristics. Supplementary Figure 11. Correlation analyses of vaccine-elicited CD8+ T cells in therapeutic exp. Supplementary Figure 12. Correlation analyses in adoptive transfer experiments for inhibitory CD8+ T cells and their cytokine function. eriments.
ARTICLE ABSTRACT
Eradicating malignant tumors by vaccine-elicited host immunity remains a major medical challenge. To date, correlates of immune protection remain unknown for malignant mesothelioma. In this study, we demonstrated that antigen-specific CD8+ T-cell immune response correlates with the elimination of malignant mesothelioma by a model PD-1–based DNA vaccine. Unlike the nonprotective tumor antigen WT1-based DNA vaccines, the model vaccine showed complete and long-lasting protection against lethal mesothelioma challenge in immunocompetent BALB/c mice. Furthermore, it remained highly immunogenic in tumor-bearing animals and led to therapeutic cure of preexisting mesothelioma. T-cell depletion and adoptive transfer experiments revealed that vaccine-elicited CD8+ T cells conferred to the protective efficacy in a dose-dependent way. Also, these CD8+ T cells functioned by releasing inflammatory IFNγ and TNFα in the vicinity of target cells as well as by initiating TRAIL-directed tumor cell apoptosis. Importantly, repeated DNA vaccinations, a major advantage over live-vectored vaccines with issues of preexisting immunity, achieve an active functional state, not only preventing the rise of exhausted PD-1+ and Tim-3+ CD8+ T cells but also suppressing tumor-induced myeloid-derived suppressive cells and Treg cells, with the frequency of antigen-specific CD8+ T cells inversely correlating with tumor mass. Our results provide new insights into quantitative and qualitative requirements of vaccine-elicited functional CD8+ T cells in cancer prevention and immunotherapy. Cancer Res; 74(21); 6010–21. ©2014 AACR.
