Image-guided irreversible electroporation directed CAR T-cell delivery to solid tumors

项目摘要 肿瘤浸润性淋巴细胞(TILs)可显著提高免疫治疗的成功率。实体瘤 存在阻碍TIL渗透和发挥作用的障碍。增加TILs的策略及其功能面临挑战 这是由于无法直接将T细胞输送到实体瘤中，以及缺乏预测其摄取的技术。 通过这项提议，我们将发展图像引导不可逆电穿孔(Ire)作为一种新颖的应用 在实体瘤中促进TIL的方法。IRE临床上用于治疗实体瘤，通过应用 微秒长的电脉冲，通过细胞膜的快速渗透杀死细胞。独一无二的 IRE的特点是，在杀死细胞的同时，它保留了治疗过程中的细胞外基质和血管 这是刺激局部炎症和促进TIL的理想选择。我们将调查我们提出的 翻译小鼠模型和恶性胸膜间皮瘤(MPM)患者的策略 细胞毒性TIL与免疫抑制细胞的比率较高的实体瘤已被证明 提高患者存活率。在我们的实验中，我们将使用嵌合抗原受体(CAR)T细胞 基因修饰以识别间皮瘤上过度表达的细胞表面抗原--间皮蛋白 癌细胞。基于PI团队之前的工作，我们合理地认为IRE导致的细胞死亡：(I)可以成像 使用程序内FDG-PET；(Ii)可以产生促进TIL的肿瘤内趋化因子梯度；以及 因此，FDG-PET引导的IRE可以作为映射和驱动肿瘤靶向CAR T细胞的替代物 送货。此外，我们的IRE定向CAR T细胞渗透策略可以使用电脉冲重复 参数设计为癌细胞靶向/保留TIL，从而增强免疫治疗效果。 我们工作的长期目标是开发以图像为导向的IRE作为一个平台，以实现可预测、可重现和 可控地将CAR T细胞和其他细胞抗癌疗法输送到实体肿瘤。在目标1中，我们将 创建结合FDG-PET成像和计算机的集成肿瘤免疫微环境图(ITIMM) 模拟，和定量多重免疫荧光组织学，以开发一种工具，将确定T- IRE治疗实体瘤的细胞定位。然后，iTIMM将被用于开发一种临床相关的 与静脉注射T细胞相结合的方案，将用于增加TIL。在目标2中，我们 将使用PET(癌细胞)和生物发光成像(T细胞)的双重成像来验证IRE脉冲 参数，以选择性地丰富和增加MPM内的功能CAR T细胞。在目标3中，我们将执行一项 PET引导下经胸MPM术的初步临床试验验证iTIMM概念 及其对TIL的影响。图像导引IRE对肿瘤微环境的调节 本地化T细胞交付，促进CAR T细胞功能，iTIMM的翻译验证呈现创新 临床使用技术的合理组合，是解决以下关键挑战的雄心勃勃的尝试 实体瘤免疫治疗。

PROJECT ABSTRACT Tumor infiltrating lymphocytes (TILs) can significantly increase the success of immunotherapy. Solid tumors present barriers to TIL penetration and function. Strategies to increase TILs and their function are challenged by the inability to direct the delivery of T cells into solid tumors and a lack of techniques to predict their uptake. Through this proposal, we will develop the use of image-guided irreversible electroporation (IRE) as a novel approach to promote TILs in solid tumors. IRE is clinically used for the treatment of solid tumors by applying microsecond long electric pulses that kill cells through rapid permeabilization of the cell membrane. A unique feature of IRE is that, while killing cells, it spares the extracellular matrix and blood vessels within the treatment region, which is ideal for stimulating local inflammation and promoting TILs. We will investigate our proposed strategy in translational mouse models and in patients with malignant pleural mesothelioma (MPM), which is a solid tumor where a higher ratio of cytotoxic TIL to immunosuppressor cells has already been shown to increase patient survival. In our experiments, we will use chimeric antigen receptor (CAR) T cells that are genetically modified to recognize mesothelin, a cell surface antigen that is overexpressed on mesothelioma cancer cells. Based on prior work by the PIs' team, we rationalize that cell death from IRE: (i) can be imaged using intra-procedural FDG-PET; (ii) can generate an intratumoral chemokine gradient that promotes TILs; and (iii) therefore, FDG-PET guided IRE can serve as a surrogate to both map and drive tumor-targeted CAR T-cell delivery. Furthermore, our strategy of IRE-directed CAR T-cell infiltration can be repeated using electric pulse parameters designed to be cancer cell-targeted/TIL-sparing, thereby augmenting immunotherapeutic efficacy. The long-term goal of our work is to develop image-guided IRE as a platform for predictable, reproducible, and controllable delivery of CAR T cells and other cellular anticancer therapies to solid tumors. In Aim 1, we will create an integrated tumor immune microenvironment map (iTIMM) combining FDG-PET imaging, computer simulations, and quantitative multiplex immunofluorescence histology to develop a tool that will determine T- cell localization in solid tumors treated with IRE. iTIMM will then be used to develop a clinically relevant protocol that, in combination of intravenous administration of T cells, will be used to increase TILs. In Aim 2, we will use dual imaging with PET (cancer cells) and bioluminescence imaging (T cells) to validate IRE pulse parameters to selectively enrich and increase functional CAR T cells within MPM. In Aim 3, we will perform a pilot clinical trial using PET-guided transthoracic IRE of MPM prior to resection to validate the concept of iTIMM and its effect on TILs. The modulation of the tumor microenvironment with image-guided IRE to map and localize T-cell delivery, promoting CAR T cell function, and translational validation of iTIMM presents innovative rational combinations of clinically used techniques and is an ambitious attempt to address a key challenge to solid tumor immunotherapy.