Dendritic Cell-Based Genetic Immunotherapy for Melanoma

这是对ROI CA79976《基于树突状细胞的黑色素瘤遗传免疫疗法》的竞争性续订，我们在其中要求 对05-08年的支持。我们在上一个资助期取得的成就包括：一、确定了免疫事件 在小鼠黑色素瘤模型中，使用树突状细胞(DC)与已定义的肿瘤抗原Makt-1，2进行工程。 接受MAKT-127.35肽冲击DC的黑色素瘤患者的L/II期临床试验和3.开启一项使用 腺病毒(Adv)制备L转导的DC。基于这一进展，我们建议继续我们对遗传学的翻译研究 人类黑色素瘤的免疫治疗围绕着三个特定的目标。 目的I：CDS缺乏环境中的遗传免疫治疗。我们已经做出了非凡和独到的观察， 用AdVMART1转导的DC免疫的CD8或I类基因敲除小鼠对B16肿瘤具有更好的保护水平 而不是野生型(Wt)小鼠。由于缺乏CD8细胞的wt小鼠不能产生保护性免疫，CD8KO小鼠 开发了一种补偿机制，可产生对DC疫苗的强大肿瘤免疫力。我们呈现的是初步的 有证据表明这种抗肿瘤免疫是由先天效应细胞(NK样细胞)和获得性效应细胞之间的协作所介导的 (Cd4)免疫系统的手臂。我们建议对潜在的机制进行描述。 目的2：AdVMART1/DC免疫的人T细胞免疫应答的生物学研究 黑色素瘤。用AdVMAKT1/DC免疫IV期MAKT-L阳性黑色素瘤患者的临床试验， 提供了一个独特的机会来定义由基因免疫触发的免疫学事件，以定义已定义的“出售”肿瘤抗原。 这种小分子蛋白只有两个表位被描述，即人类白细胞抗原A2.1限制性的Makt-LzT.35和HLADK4限制性的Makt。 15T-73.利用ELISPOT和四聚体分析这些I类和II类表位，我们将定量、分离和研究MART-L-反应 免疫后患者CD8和CIM T细胞的变化我们还将研究决定簇传播和交叉呈现在临床中的作用。 树突状细胞用于疫苗接种的生物学反应及天然免疫在树突状细胞基础上的可能参与 免疫疗法。 目的3：CTLA4阻断在临床树突状细胞免疫治疗中的应用。基于DC的免疫疗法偶尔会产生 但在临床抗肿瘤方面有显著的反应。我们仔细研究了超市管理的一个主题--I/DC 疫苗之后是CTLA4封闭抗体。免疫学分析表明，抗肿瘤免疫反应 DC疫苗的启动由CTLA4阻断维持。为了验证这一假设，我们设计了一个随机的第二阶段 试验的主要目的是检测MART-127.3Y‘DC+CTLA4阻断对黑色素瘤抗原频率的影响。 ELISPOT法检测特异性活化T细胞。这项试验将使我们深入了解管理 以DC为基础的免疫化疗的活性。 总之，我们建议继续我们在遗传免疫治疗中的翻译研究，重点放在免疫机制上。 和临床假设检验。

This is a competitive renewal for ROI CA79976 "Dendritic cell-based genetic immunotherapy for melanoma" in which we request support for years 05-08. Our accomplishments in the previous funding period include: I. defining the immunological events taking place in a murine melanoma model using dendritic cell (DC) engineered with a defined tumor antigen MAKT-1, 2. completing a phase l/II clinical trial in melanoma patients receiving MAKT-127.35 peptide pulsed DC and 3. opening a gane therapy trial using adenovims (AdV) MAKT-l-transduced DC. Based on this pr6gress, we propose to continue our translational studies of genetic immunotherapy of human melanoma centered around three specific aims. Aim I: Genetic Immunotherapy in a CDS-Deficient Environment. We have made the remarkable and original observations that CD8 or Class I knock out mice immonized with AdVMARTl-transduced DC have superior levels of protection to B16 nu_lanoma than wild type (wt) mice. Since wt mice depleted of CD8 cells are unable to generate protective immunity, CD8 KO mice have developed a compensatory mechanism fi'om generating robust tumor immunity to DC vaccination. We present preliminary evidence that this antitumor immunity is mediated by a collaboration between effector cells of the innate (NK-like) and adaptive (CD4) arms of the immune systems. We propose to characterize the underlying mechanism. Aim 2: The Biology of Class I and Class II-Restritted T Cell Responses in AdVMART1/DC Immunized Patients with Melanoma. This clinical trial, in which patients with stage IV MAKT-l-positive melanoma are immunized with AdVMAKT1/DC, provides a unique opportunity to define immunological events triggered by genetic immunization to a defined "sell" tumor antigen. Only two epitopes have been described for this small protein- HLA-A2.1-restricted MAKT-lzT.35 and HLA-DK4 restricted MAKT- 15t-73.Using ELISPOT and tetramer assays for these class I and II epitopes, we will quantitate, isolate and study MART-l-reactive CD8 and CIM T cell in immunized patients. We will also study the role of determinant spreading and cross-presentation in clinical response, the biology of DC used for vaccination and the possible participation of innate (NK) immunity in DC-based immunotherapy. Aim 3:CTLA4 Blockade in Clinical Dendritic Cell-Based Immunotherapy. DC-based immunotherapy generates occasional but dramatic clinical antitumor responses. We have closely studied one subject in whom the administration of MART-I/DC vaccines was followed by a CTLA4 blocking antibody. Immunological analysis suggests that the antitumor imm,ne response initiated by the DC vaccines was maintained by CTLA4 blockade. To test this hypothesis, we have designed a phase II randomized trial with the primary goal of detecting the effect of MART-127.3y'DC + CTLA4 blockade on the frequency of melanoma antigen- specific activated T cells using ELISPOT assays. This trial will provide insight in the autoregulat0ry mechanisms that govern the activity ofDC-based imm_unotherapy. In summary, we propose to continue our translational prosram in genetic immunotherapy with an emphasis on immune mechanism and clinical hypothesis-testing.