Enhancing Melanoma TIL Efficacy with Multifactor mRNA-Mediated T Cell Reprogramming

项目总结 尽管T细胞介导的免疫反应对免疫治疗的成功至关重要，但这些T细胞 与恶性病变相关的肿瘤通常功能失调，无法控制肿瘤生长。通过以下方式治疗 分离、激活和体外扩增的肿瘤浸润性淋巴细胞(TIL)已被证明非常有效 在一些黑色素瘤患者群体中。然而，相当数量的患者没有反应， 推测是由于多种宿主免疫因素中的一种。TIL作用机制的研究现状 这表明肿瘤特异性效应T细胞的早期强劲扩增和分化程度较低的转移 具有长期存活能力的细胞是成功治疗的关键。前者的证据包括有必要 对于TIL输注时大剂量外源性IL-2，其反应与高频 效应性T细胞，大多数肿瘤杀伤发生在治疗开始后的很早。证据： 后者在许多临床前实验和临床观察中被发现，其中TIL的存在 输液产品中的中央记忆亚集与肿瘤的消退相关。我们的总体目标是 通过生成兼具瞬时能力的TIL产品有效提高TIL的治疗效果 立即杀死肿瘤细胞，以及长期坚持和维持持久的抗肿瘤反应的能力。 为了应对这些挑战，我们开发了强大的方法来用mRNA介导的基因重新编程TIL 心理治疗。该方法具有安全、高效、快速等优点。 生产，严格控制表达水平和同步的多因素重新编程。在预赛中 我们已经开发了一种系统，可以将mRNA的寿命延长一个数量级。我们的单人牢房 对扩张术前后患者TIL的分析发现，扩张术中有两条特定的途径存在缺陷 TIL产品，可能导致其即刻疗效不佳和没有记忆命运。这两个都是 将通过TIL信使核糖核酸重新编程来增强。在目标1中，我们将开发一种增强后扩展的方法 直到存活，而在目标2中，我们将改进中央记忆TIL的生产。我们将对效果进行评估 使用来自多发性黑色素瘤的成对肿瘤/TIL集在TIL产生方面的这些改进 在体外和临床前期人源化小鼠中研究肿瘤介导的TIL激活和肿瘤溶解 使用单细胞分析和先进的空间转录学的模型。此应用程序解决了以下需求 提高黑色素瘤过继细胞免疫疗法的应答率，旨在可翻译为 在不久的将来进行临床试验。

PROJECT SUMMARY Although T cell mediated immune responses are critical for the success of immunotherapy, those T cells associated with malignant lesions are typically dysfunctional and fail to control tumor growth. Treatment with tumor infiltrating lymphocytes (TIL) that are isolated, activated, and expanded ex vivo has proven very effective in some patient populations of melanoma. However, a substantial number of patients do not respond, presumably due to one of a number of host immune factors. Current understanding of TIL mechanism of action suggests that both an early robust expansion of tumor-specific effector T cells and transfer of less differentiated cells with long-term survival capacity are key to a successful therapy. Evidence for the former includes the need for high dose exogenous IL-2 at the time of TIL infusion, the correlation of response with a high frequency of effector T cells, and the majority of tumor killing occurring very early after the initiation of therapy. Evidence for the latter is found in many pre-clinical experiments as well as clinical observations where the presence of TILs from the central memory subset in the infusion product correlates with tumor regression. Our overall goal is to improve TIL therapeutic efficacy through the generation of TIL products with both the transient ability to effectively immediately kill tumor cells as well as the long-term ability to persist and maintain durable anti-tumor responses. To address these challenges we have developed robust methods to reprogram TILs with mRNA-mediated gene therapy. The use of our mRNA approach has the advantages of increased safety, high efficiency, rapid production, tightly controlled expression levels and simultaneous multi-factor reprogramming. In preliminary work we have developed a system that increases mRNA lifespan by an order of magnitude. Our single cell analysis of patient TILs pre- and post-expansion has identified two specific pathways deficient in the expanded TIL product that likely contribute to their poor immediate efficacy and absence of memory fate. Both of these will be augmented by TIL mRNA reprogramming. In Aim 1 we will develop a method to enhance post-expansion TIL survival, while in Aim 2 we will improve the production of central memory TILs. We will evaluate the effect of these improvements in TIL production using paired tumor/TIL sets derived from the melanomas from multiple patients by studying tumor-mediated TIL activation and tumor lysis in vitro and in pre-clinical humanized mouse models using single cell analysis and advanced spatial transcriptomics. This application addresses the need to improve response rates to adoptive cell immunotherapies for melanoma and is designed to be translatable to clinical trials in the near future.