This project tests an improved dendritic cell (DC) vaccine, which is designed to promote in vivo cross presentation and determinant spreading. Based on results from our previous trials, we have made several important improvements: engineering the DC with 3 full-length, defined, tumor antigens to activate multiple CD8+ and CD4+ T cell clones (reducing the concern for antigen loss variants); providing antigen presentation for the life of the DC; providing cognate CD4+ T cell help to the CD8+ T cells (“helped” CTL); using a matured DC (a cocktail specifically matched to adenovirus (AdV) transduction signals); activating innate immunity via natural killer (NK) cell migration and activation; and boosting with systemic IFNα (for endogenous DC type 1 skewing, improved cross-priming and direct effects on T cells). Together, this vaccine strategy should more potently activate a polyclonal anti-tumor response incorporating multiple adaptive and innate effectors which we predict will lead to a higher frequency of patients who not only activate vaccine-encoded antigen-specific T cell responses, but also develop determinant spreading and a significant clinical response.
Specific Aim 1: Completion of the AdVTMM2/DC +/- IFNα clinical trial. We will fully assess the clinical and immunologic impact of the vaccine and IFNα boost, and test: A) Clinical outcomes in the advanced stage patients; B) Immunologic outcomes (from blood and tumor infiltrating lymphocytes (TIL); baseline, post-vaccine and post-IFNα), including CD8+ and CD4+ T cells specific for vaccine antigens (tyrosinase, MART-1 and MAGE-A6), NK cell activation and tumor infiltration (TIL and tumor), T cell response to “spreading” antigens (including gp100, NY-ESO-1 and vascular antigens), T cell responses to autologous tumor, antibody responses to melanoma antigens, and regulatory responses (Treg, MDSC); and C) AdV-specific responses, including CD4+ and CD8+ T cells specific for total viral proteins, total anti-AdV antibodies, and anti-AdV neutralizing antibodies.
Specific Aim 2: Mechanism and biomarkers of clinical response. To understand patient differences in tumor biology, in DC vaccines and in candidate biomarkers, we will examine: A) DC vaccine transcriptional profiling (+/- IFNγ and LPS maturation, +/- AdVTMM2) and compare to DC vaccine phenotype, cytokine production and antigen expression; B) Tumor transcriptional profiling (baseline, post-vaccine and post-IFNα) to examine tumor modulation by DC vaccines and IFNα therapy; C) Peripheral blood signaling induced by IFNα (phospho-STAT-1); D) Serum profiling (autoimmunity antibodies, LDH, CRP, cytokines) to examine previously identified candidate serum biomarkers; E) Molecular mimicry with mycoplasma (impact of baseline memory to mycoplasma); and F) Germline DNA SNP analysis with a focus on identified immunologic melanoma SNPs (IL-12p40, Calreticulin, and CRP). These studies will identify predictive and prognostic biomarkers of immunologic and clinical response.
Specific Aim 3: Mechanism of NK cell “activation” in anti-melanoma immunity. NK cell activity may also be critical for clinical response. We will perform in vitro studies to further our recent findings in AdV/DC-NK cell cross-talk utilizing banked melanoma patient blood and tumor to examine: A) Direct NK cell interactions with melanoma tumor cells (cytotoxicity, cytokines); B) Helper/type 1 skewing by NK cells to shape adaptive CD8+ and CD4+ T cell responses; and C) Melanoma tumor-derived inhibitory impact on NK cell function. These studies will identify the mechanisms of innate NK cell anti-tumor activity and any potential suppression by tumor.
Completion of the AdVTMM2/DC +/- IFNα trial and these three aims will allow us to test novel hypotheses. We will determine: 1) Vaccine-induced clinical and immune response, correlation with subsequent induction of determinant spreading, and the impact of adding IFNα to the antigen-engineered DC vaccine; 2) Layers of potential mechanisms of response and biomarkers of immunologic and clinical response; and 3) Innate-adaptive cross-talk between DC vaccines, NK cells, vaccine-activated T cells, and systemic IFNα.
We will test a potent vaccine combination therapy which may promote a higher frequency and duration of clinical response. The examination of several predictive biomarkers (tumor infiltration and gene expression, pathogen memory, SNPs, NK cell activation) and prognostic biomarkers (adaptive cellular and humoral immunity, determinant spreading, NK cell activation, serum biomarkers, autoimmunity) may identify patients able to benefit from immune-based approaches, as well as the mechanisms of that benefit.