Dendritic Cell-Based Genetic Immunotherapy for Melanoma

描述（由申请人提供）：这是ROI CA79976“基于树突状细胞的黑色素瘤遗传免疫疗法”的竞争性续期，我们要求支持05-08年。我们在上一个资助期内取得的成就包括：1。利用树突状细胞（DC）修饰特定肿瘤抗原mart - 1,2来定义小鼠黑色素瘤模型中发生的免疫事件。完成黑色素瘤患者接受MART-1/27-35肽脉冲DC和3的i /II期临床试验。开启一项使用腺病毒（AdV） mart -1转导DC的基因治疗试验。基于这一进展，我们建议围绕三个具体目标继续进行人类黑色素瘤遗传免疫治疗的转化研究。目的1:cdss缺陷环境中的基因免疫治疗。我们已经做出了显著的和原始的观察，用advmart1转导的DC免疫CD8或I类敲除小鼠对B16黑色素瘤的保护水平优于野生型（wt）小鼠。由于CD8细胞缺失的wt小鼠无法产生保护性免疫，CD8 KO小鼠已经发展出一种对DC疫苗产生强大肿瘤免疫的代偿机制。我们提出的初步证据表明，这种抗肿瘤免疫是由免疫系统的先天（nk样）和适应性（CD4）臂的效应细胞之间的协同作用介导的。我们建议描述其潜在机制。目的2:AdVMART1/DC免疫黑色素瘤患者I类和ii类限制性T细胞反应的生物学特性。在这项临床试验中，IV期mart -1阳性黑色素瘤患者接种了AdVMART1/DC，提供了一个独特的机会来定义由基因免疫对定义的“自我”肿瘤抗原触发的免疫事件。只有两个表位被描述为这种小蛋白- hla - a2.1限制性的MART-1/27-35和HLA-DK4限制性的MART-1/51-73。使用ELISPOT和四聚体检测这类I和II表位，我们将定量、分离和研究免疫患者的mart -1反应性CD8和CD4 T细胞。我们还将研究决定扩散和交叉呈现在临床反应中的作用，DC用于疫苗接种的生物学以及先天（NK）免疫在DC免疫治疗中的可能参与。目的3:CTLA4阻断在临床树突状细胞免疫治疗中的应用。以dc为基础的免疫疗法偶尔会产生显著的临床抗肿瘤反应。我们仔细研究了一名受试者，该受试者在接种了MART-1/DC疫苗后接种了CTLA4阻断抗体。免疫学分析表明，DC疫苗启动的抗肿瘤免疫应答通过CTLA4阻断得以维持。为了验证这一假设，我们设计了一项II期随机试验，主要目的是使用ELISPOT检测MART-1/27-35/DC + CTLA4阻断剂对黑色素瘤抗原特异性活化T细胞频率的影响。该试验将为控制dc免疫疗法活性的自身调节机制提供见解。总之，我们建议继续我们的基因免疫治疗的翻译项目，重点是免疫机制和临床假设检验。

DESCRIPTION (provided by applicant): 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: 1. defining the immunological events taking place in a murine melanoma model using dendritic cell (DC) engineered with a defined tumor antigen MART-1, 2. completing a phase l/II clinical trial in melanoma patients receiving MART-1/27-35 peptide pulsed DC and 3. opening a gene therapy trial using adenovirus (AdV) MART-1-transduced DC. Based on this progress, we propose to continue our translational studies of genetic immunotherapy of human melanoma centered around three specific aims. Aim 1: Genetic Immunotherapy in a CDS-Deficient Environment. We have made the remarkable and original observations that CD8 or Class I knock out mice immunized with AdVMART1-transduced DC have superior levels of protection to B16 melanoma 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 from generating robust tumor immunity to DC vaccination. We present preliminary evidence that this antitumor immunity is mediated by 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-Restricted T Cell Responses in AdVMART1/DC Immunized Patients with Melanoma. This clinical trial, in which patients with stage IV MART-1-positive melanoma are immunized with AdVMART1/DC, provides a unique opportunity to define immunological events triggered by genetic immunization to a defined "self" tumor antigen. Only two epitopes have been described for this small protein-HLA-A2.1-restricted MART-1/27-35 and HLA-DK4 restricted MART-1/51-73. Using ELISPOT and tetramer assays for this class I and II epitopes, we will quantitate, isolate and study MART-1-reactive CD8 and CD4 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-1/DC vaccines was followed by a CTLA4 blocking antibody. Immunological analysis suggests that the antitumor immune 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-1/27-35/DC + CTLA4 blockade on the frequency of melanoma antigen-specific activated T cells using ELISPOT assays. This trial will provide insight in the autoregulatory mechanisms that govern the activity of DC-based immunotherapy. In summary, we propose to continue our translational program in genetic immunotherapy with an emphasis on immune mechanism and clinical hypothesis-testing.