Telomerase RNA Transfected Dendritic Cell Vaccines

描述（由申请人提供）：我们的长期目标是通过诱导针对人端粒酶蛋白亚单位端粒酶逆转录酶（TERT）的免疫力，为广泛的癌症患者开发临床有效且广泛适用的疫苗策略。端粒酶是广泛表达的肿瘤排斥抗原的有吸引力的候选者，因为端粒酶在正常组织中是沉默的，但在大多数人实体瘤（包括前列腺癌）中被重新激活和过表达。我们已经进行了广泛的临床前研究，证明用TERTRNA转染的自体DC是体外刺激来自癌症患者PBMC的TERT特异性T细胞应答的显著有效的策略。这些应答主要是CD 8 + T细胞介导的，因为从mRNA转染的DC表达的抗原将优先被引导到内源性I类呈递途径中。鉴于CD 4 + T细胞在诱导和维持抗肿瘤应答中起着至关重要的作用，我们已经表明，含有溶酶体靶向信号LAMP的修饰的TERT转录物（LAMP-TERT）通过将TERT蛋白重定向到II类呈递途径中而导致对TERT特异性CD 4+细胞的刺激增加。在这里，我们建议通过进行I期临床试验将这些临床前发现转化为临床环境，该I期临床试验旨在评估TERTRNA和LAMP-TERTRNA转染的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应答（Aim I）。在本申请的目的2中，我们提出通过测定疫苗接种之前和之后TERT特异性CD 8+和CD 4 + T细胞应答的存在、幅度和持续时间来分析这些T细胞应答，所述方法包括（a）分析通过自动化ELISPOT测定评估的活化T细胞的处理后细胞因子谱的变化;（B）通过四聚体分析测量TERT表位特异性CTL亚群的诱导，（c）测量体内产生的CTL特异性识别和裂解表达TERT的细胞靶标的功能能力，d）通过常规增殖测定或通过使用细胞因子流式细胞术的新的简化全血测定来确定CD 4 + T细胞应答的存在。在本授权的目的3中，我们寻求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驱动的CD 4 + T细胞应答的刺激。拟议的项目将为科学有效的II期研究奠定基础，以评估用TERTRNA转染的DC接种前列腺癌患者的临床疗效，并评估使用TERT作为抗原治疗广泛的癌症的效用，这些癌症过度表达TERT。

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.