Engineering the ER Stress Response to Promote the Survival and Cytotoxic Specificity of CAR T cells

项目摘要/摘要 嵌合抗原受体(CAR)T细胞已经给癌症治疗带来了革命性的变化，并显示出令人难以置信的 复发或难治性B细胞癌患者的有效率，但对实体瘤无效。为了 在实体瘤中工作，CAR T细胞必须能够在免疫抑制的肿瘤微环境中生存 (TME)并区分存在于肿瘤细胞和健康组织上的抗原。这两个障碍 可以通过使用已知的生物回路对T细胞进行巧妙的合成工程来克服。我们有 确定了一条对多种类型白细胞的生存和功能至关重要的途径，并自然开启 然而，令人惊讶的是，在激活CD8+T细胞的过程中，它并没有在肿瘤微环境中被激活。这里, 我建议综合激活这一途径，以促进CD8+CAR T细胞的细胞毒作用和靶向性。 对肿瘤的特异性。具体地说，我将首先探索这个信号通路的多个方面及其能力 促进CAR T细胞在肿瘤微环境应激下的存活和细胞毒作用。作为一秒钟 为了提高实体瘤的CAR-T细胞特异性，我将设计一种结构，选择性地激活一种 CAR对转录后调控的反应。转录后调控的好处包括 信号的激活和放大。通过使转录后调控因子和靶标 依赖于结合的靶细胞抗原，我将在功能上创建一个需要刺激的“与”逻辑门 从两种不同的抗原中分离出来，以促进CAR T细胞的杀伤。对于这两种策略，我将首先演示它们的 模拟TME条件的原代小鼠和人类细胞的体外疗效，然后在体内使用 同基因移植黑色素瘤小鼠模型。总体而言，我假设这一关键因素的人工激活 T细胞途径将增强CAR-T在固体TME中的行为和控制。

PROJECT SUMMARY/ABSTRACT Chimeric antigen receptor (CAR) T cells have revolutionized cancer therapies and show incredible responses in patients with relapsed or refractory B cell cancers, yet are ineffective in solid tumors. In order to work in solid tumors, CAR T cells must be able to survive the immunosuppressive tumor microenvironment (TME) and distinguish between antigens presented on tumor cells and healthy tissue. Both of these obstacles can be overcome through clever synthetic engineering of the T cell using known biological circuits. We have identified a pathway critical to the survival and function of multiple types of leukocytes and naturally turned on during activation of CD8+ T cells, however surprisingly, it is not activated in the tumor microenvironment. Here, I propose to synthetically activate this pathway in order to promote CD8+ CAR T cell cytotoxicity and on-target, on-tumor specificity. Specifically, I will first explore multiple aspects of this signaling pathway and its ability to promote CAR T cell survival and cytotoxicity under the stresses in the tumor microenvironment. As a second strategy to improve CAR-T cell specificity for solid tumors, I will engineer a construct that selectively activates a CAR in response to post-transcriptional regulation. Benefits of post-transcriptional regulation include speed of activation, and amplification of signal. By making both the post-transcriptional regulator and the target dependent on binding target cell antigens, I will functionally create an “AND” logic gate, requiring stimulation from 2 separate antigens in order to facilitate CAR T cell killing. For both strategies I will first demonstrate their efficacy in vitro with primary mouse and human cells simulating TME conditions, and then in vivo using a syngeneic transplanted melanoma mouse model. Overall, I hypothesize that synthetic activation of this critical T cell pathway will enhance CAR-T behavior and control in the solid TME.