Therapeutic use of T cells engineered to produce radiation-inducible cytokines

项目概述：我们建议使用基因工程化的T细胞，在体外条件下递送细胞毒性细胞因子。 放射诱导启动子的控制。当基因工程T细胞定位于肿瘤床时， 通过放射疗法（RT）实现细胞因子的活化，从而使肿瘤暴露于治疗效果 RT和细胞因子治疗的效果。这种“分子开关”策略是由我们的实验室开创的， 重组腺病毒载体在这个概念中，RT激活具有放射诱导启动子的嵌合基因 与细胞因子基因相连此外，RT的直接抗肿瘤作用具有与肿瘤抑制剂协同作用的潜力。 过继转移的T细胞的抗肿瘤作用。 假设：外源性控制肿瘤浸润性T细胞产生细胞因子的能力将（i） 抵消肿瘤的免疫抑制作用，影响关键的抗肿瘤T细胞效应子功能，（ii） 由于细胞因子产生的受控局部活化而使全身毒性最小化，（iii）增强协同作用 RT和过继性转移的T细胞之间的关系，并因此（iv）提高RT的治疗比。 具体目的：1）在逆转录病毒载体中克隆T细胞特异性放射诱导型启动子，其驱动 用于转导T细胞的报告基因，以及使用纵向肿瘤的RT的基因诱导的体内分析 2）在逆转录病毒载体中克隆T细胞特异性放射诱导型启动子以驱动 T细胞中的治疗性细胞因子和用放射诱导的 表达细胞因子的T细胞加上局部RT。 我们的成像技术独特地允许我们检查相同的肿瘤，以及肿瘤的相同区域， 在每次RT给药后数小时或数天，并测量EGFP表达与给药前水平相比的增加。 RT.这将允许在体内比较和验证T细胞特异性启动子的放射诱导，所述T细胞特异性启动子是在大肠杆菌中鉴定的。 目标1.此外，T细胞的抗肿瘤作用将作为癌细胞和血管的破坏而可见，从而 为我们提供了机械数据。我们的提案中的其他尖端技术包括CT扫描图像- 用于以毫米级精度照射小鼠癌组织的引导小动物照射器。 我们提出了RT、基因治疗和过继性T细胞治疗的组合，用于治疗局部晚期乳腺癌。 或转移性癌症是一种全新的治疗方法，因此有可能超越传统的治疗方法。 在癌症治疗中的应用我们的新范例的最初临床应用是治疗难治性固体 肿瘤和寡转移与过继转移的T细胞的联合作用， 细胞因子货物和RT，因为RT已被证明是一种免疫刺激方式;因此，它可能是 翻译我们的方法将导致增加治愈率和显着降低毒性， 癌症患者。值得注意的是，我们的系统可以应用于任何其他需要激活的治疗基因 在附近

Project Summary: We propose to use genetically-engineered T cells to deliver cytotoxic cytokines under the control of a radio-inducible promoter. When the genetically engineered T cells localize to the tumor bed, activation of the cytokine is achieved by radiotherapy (RT), thereby exposing the tumor to the therapeutic effects of both RT and cytokine therapy. This “molecular switch” strategy was pioneered by our laboratory, employing recombinant adenoviral vectors. In this concept, RT activates a chimeric gene with a radio-inducible promoter linked to a cytokine gene. Furthermore, the direct anti-tumor effects of RT have the potential to synergize with the anti-tumor effects of adoptively-transferred T cells. HYPOTHESIS: The ability to exogenously control the production of cytokines by tumor-infiltrating T cells will (i) counteract the immunosuppressive effects of the tumor, affecting a critical anti-tumor T cell effector function, (ii) minimize systemic toxicity due to controlled local activation of cytokine production, (iii) potentiate the synergy between RT and adoptive transferred T cells, and therefore (iv) improve the therapeutic ratio of RT. Specific Aims: 1) Cloning of T cell-specific radio-inducible promoters in retroviral vectors driving expression of a reporter gene for transduction of T cells, and in vivo analysis of gene induction by RT using longitudinal tumor imaging; 2) Cloning of T cell-specific radio-inducible promoters in retroviral vectors to drive expression of therapeutic cytokines in T cells and treatment of murine primary and oligometastatic tumors with radio-inducible cytokines-expressing T cells plus local RT. Our imaging technology uniquely allows us to examine the same tumor, and an identical area of the tumor, hours or days after each RT dose, and measure the increase in EGFP expression compared to levels before RT. This will allow in vivo comparison and validation of radio-induction of T cell-specific promoters identified in Aim 1. Furthermore, anti-tumor effects of T cells will be visible as destruction of cancer cells and vessels, thereby providing us with mechanistic data. Other cutting-edge technologies in our proposal include a CT-scan image- guided small animal irradiator for irradiation of mouse cancer tissues with millimetric precision. Our proposed combination of RT, gene therapy and adoptive T-cell therapy for treatment of locally advanced or metastatic cancer is dramatically new, and therefore has the potential to move beyond traditional approaches in the treatment of cancer. The initial clinical application of our new paradigm is the treatment of refractory solid tumors and oligometastases with the combined effects of adoptively-transferred T cells carrying radio-inducible cytokine cargo and RT, since RT has been shown to be an immunostimulatory modality; therefore, it is likely that translation of our approach would result in increased cures and significantly lower toxicities for cancer patients. Of note, our system could be applied to any other therapeutic gene that needs to be activated locally.