Multiple Antigen-Engineered DC Immunization and IFNalpha-2b Boost for Metastatic Melanoma

该项目测试了一种改进的DC疫苗，其旨在促进体内交叉呈递和决定簇传播。基于我们以前的试验结果，我们已经做出了几项重要的改进：用3种全长、确定的肿瘤抗原工程化DC以激活多个CD 8+和CD 4 + T细胞克隆（减少对抗原丢失变体的关注）;为DC的生命提供抗原呈递;为CD 8 + T细胞提供同源CD 4 + T细胞帮助（“帮助的”CTL）;使用成熟DC（与腺病毒（AdV）转导信号特异性匹配的混合物）;通过NK细胞迁移和激活激活先天免疫;以及用系统性IFNα加强（对于内源性DC 1型偏斜，改善交叉引发和对T细胞的直接作用）。总之，这种疫苗策略应该更有效地激活包含多种适应性和先天性效应物的多克隆抗肿瘤应答，我们预测这将导致更高频率的患者不仅激活疫苗编码的抗原特异性T细胞应答，而且还产生决定性T细胞应答。 扩散和显著的临床反应。 三个目标是：目标1：完成AdVTM 2/DC +/- IFNα临床试验1.a。IV期患者的临床结局; 1.b.免疫学结局（来自血液和TIL;基线、疫苗接种后和IFN α后）; 1.c. AdV特异性细胞和体液反应。 目的2：临床反应的机制和生物标志物：2.a. DC疫苗转录谱分析（+/-成熟，+/-AdVTMM 2）; 2. b.肿瘤转录谱（基线、疫苗接种后和IFN α后）; 2.c. IFNα诱导的外周血信号传导（STAT）; 2.d.血清分析（自身免疫抗体、LDH、CRP细胞因子）; 2.e.支原体的分子模拟（基线记忆对支原体的影响）; 2.f.种系DNA SNP分析 目的3：NK细胞在抗黑色素瘤免疫中的“激活”机制 体外研究以扩展我们最近在AdV/DC-NK细胞与库存的黑素瘤患者血液和肿瘤（扩展至细胞系的原发性肿瘤）的串扰中的发现：NK细胞与黑素瘤肿瘤细胞的直接相互作用（细胞毒性、细胞因子）; 3b：辅助/1型在形成适应性CD 8+和CD 4 + T细胞应答中的偏斜作用; 3c：黑素瘤肿瘤对NK细胞功能的影响。

This project tests an improved 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 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. The three aims are: Aim 1: Completion of the AdVTMM2/DC +/- IFNα clinical trial 1.a. Clinical outcomes in the stage IV patients; 1.b. Immunologic outcomes (from blood and TIL; baseline, post-vaccine and post-IFNα); 1.c. AdV-specific cellular and humoral responses. Aim 2: Mechanism and biomarkers of clinical response: 2.a. DC Vaccine Transcriptional Profiling (+/- maturation, +/- AdVTMM2); 2.b. Tumor Transcriptional Profiling (baseline, post-vaccine and post-IFNα); 2.c. Peripheral blood signaling induced by IFNα (STATs); 2.d. Serum profiling (autoimmunity antibodies, LDH, CRP cytokines); 2.e. Molecular mimicry with mycoplasma (impact of baseline memory to mycoplasma); 2.f. Germline DNA SNP analysis Aim 3: Mechanism of NK cell "activation" in anti-melanoma immunity In vitro studies to expand our recent findings in AdV/DC-NK cell cross-talk with banked melanoma patient blood and tumor (primary tumor expanded to cell lines): 3.a. Direct NK cell interactions with melanoma tumor cells (cytotoxicity, cytokines); 3b: Helper/type 1 skewing role in shaping adaptive CD8+ and CD4+ T cell responses; 3c: Melanoma tumor impact on NK cell function.