Telomerase RNA Transfected Dendritic Cell Vaccines

描述(申请人提供)：我们的长期目标是通过诱导对人类端粒酶的蛋白质亚单位端粒酶逆转录酶(TERT)的免疫，为广泛的癌症患者开发一种临床有效和广泛适用的疫苗策略。由于端粒酶在正常组织中不表达，但在包括前列腺癌在内的大多数人类实体肿瘤中重新激活和过度表达，端粒酶是一个具有广泛表达的肿瘤排斥抗原的有吸引力的候选者。我们已经进行了广泛的临床前研究，表明TERT RNA转基因的自体DC在体外刺激癌症患者PBMC的TERT特异性T细胞反应是一种显着有效的策略。这些反应主要是由CD8+T细胞介导的，这是因为转基因DC表达的抗原将优先进入内源性I类递呈途径。鉴于CD4+T细胞在诱导和维持抗肿瘤反应中起着至关重要的作用，我们已经证明，含有溶酶体靶向信号灯(LAMP-TERT)的修饰TERT转录本通过将TERT蛋白重定向到II类递呈途径，从而增加对TERT特异性CD4+细胞的刺激。在这里，我们建议通过进行一项I期临床试验来将这些临床前发现转化为临床环境，该试验旨在评估TERT RNA和LAMP-TERT RNA转基因DC的安全性和生物活性，以刺激转移性前列腺癌患者的潜在治疗性免疫反应(AIM I)。在这一应用的目标2中，我们建议通过确定接种前后TERT特异性CD8+和CD4+T细胞反应的存在、大小和持续时间来分析这些T细胞反应，方法是：(A)通过自动ELISPOT分析来分析治疗后激活的T细胞的细胞因子谱的变化；(B)通过四聚体分析来测量TERT表位特异性CTL亚群的诱导；(C)测量体内产生的CTL特异性识别和溶解TERT表达细胞靶点的功能能力；d)通过传统的增殖分析或新型的简化的全血细胞检测，使用细胞因子流式细胞术确定CD4+T细胞反应的存在。在这项资助的目标3中，我们试图通过使用针对不变链表达的反义寡核苷酸介导的抑制方法来进一步改善LAMP-TERT对CD4+T细胞反应的刺激。拟议的项目将为科学有效的第二阶段研究奠定基础，以评估用转TERT RNA的DC接种前列腺癌患者的临床疗效，并评估使用TERT作为抗原治疗大量过度表达TERT的癌症的效用。

DESCRIPTION (provided by applicant): Our long-term objective is to develop a clinically effective and broadly applicable vaccine strategy for a wide range of cancer patients by inducing immunity against the protein subunit of human telomerase, telomerase reverse transcriptase (TERT). Telomerase is an attractive candidate for a broadly expressed tumor rejection antigen since telomerase is silent in normal tissues but reactivated and overexpressed in the majority of human solid tumors including prostate cancers. We have performed extensive preclinical studies demonstrating that autologous DC transfected with TERT RNA are a remarkable effective strategy to stimulate TERT-specific T cell responses from the PBMC of cancer patients in vitro. These responses were predominantly CD8+ T cell mediated due to the fact that antigen expressed from mRNA transfected DC will be channeled preferentially into the endogenous class I presentation pathway. In view of the fact that CD4+ T cells play a critically important role in the induction and maintenance of an antitumor response, we have shown that modified TERT transcripts containing the lysosomal targeting signal LAMP (LAMP-TERT) lead to increased stimulation of TERT-specific CD4+ cells by redirecting the TERT protein into the class II presentation pathway. Here we propose to translate these preclinical findings into a clinical setting by conducting a phase I clinical trial designed to evaluate the safety and bioactivity of TERT RNA and LAMP-TERT RNA transfected DC to stimulate potentially therapeutic immune responses in metastatic prostate cancer patients (Aim I). In Aim 2 of this application, we propose to analyze these T cell responses by determining the presence, magnitude and duration of TERT-specific CD8+ and CD4+ T cell responses prior to and following vaccination by (a) analyzing changes in the post treatment cytokine profiles of activated T cells as assessed by an automated ELISPOT assay; (b) measuring the induction of TERT epitope-specific CTL subsets by tetramer analysis, (c) measure the functional capability of the in vivo generated CTL to specifically recognize and lyse TERT expressing cellular targets, d) determine the presence of CD4+ T cell responses by conventional proliferation assays or by novel, simplified whole blood assays using cytokine flow-cytometry. In Aim 3 of this grant we seek to further improve the LAMP-TERT-driven stimulation of CD4+ T cell responses by using antisense oligonucleotide mediated inhibition methods directed against invariant chain expression. The proposed project will set the stage for scientifically valid phase II studies to assess the clinical efficacy of vaccinating prostate cancer patients with TERT RNA transfected DC and evaluate the utility of using TERT as antigen to treat a wide range of cancers, which overexpress TERT.