Advancing entirely virus-free CRISPR CAR T cells to clinic- Are they as good as lentiviral CAR?

将完全无病毒的CRISPR CAR T细胞推向临床——它们和慢病毒CAR一样好吗？

• 批准号: MR/Y503496/1
• 负责人: Waseem Qasim
• 金额: $ 31.65万
• 依托单位: University College London
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2024
• 资助国家: 英国
• 起止时间: 2024 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FY503496%2F1
• 关键词: Advancing; entirely; virus; free; CRISPR

We have delivered a number of experimental chimeric antigen receptor (CAR) T cell therapies into clinical trials using specialist facilities at Great Ormond Street Hospital. These have helped treat children & adults with certain types of leukaemia who would otherwise have no further options for their disease. These projects were taken from concept and testing in the lab, through to manufacturing in clean rooms and clinical use in patients. All have relied on using disabled viruses to add genes into T cells to reprogram them. Some viral CAR T cells have undergone extra changes using genome editing - molecular scissors to snip out bits of other genes - and this allowed 'universal' T cells to be used without any matching. Now its becoming possible to use similar technology to both cut genes and insert new ones, and this means we can engineer cells without the need for viruses to add the CAR gene. We think the approach could eliminate supply bottlenecks, especially for making virus stocks for each new CAR, a major hold up issue for clinical trials. We have shown in our lab work so far that the efficiency of CAR insertion using CRISPR is almost as high as when we use viruses, and the cells behave and function just as well in short-term dish experiments. We now need to make sure that the process works at a larger scale, in a way that can be used in clean rooms, and that the cells behave no differently when they activated and responding against human leukemia in animals over a longer period.This application will manufacture and compare CRISPR CAR T cells using existing lentiviral or new virus-free strategies to add the CAR. The tests will be on a scale suitable for future human trials and under conditions that can be transferred to a clean room setting. The cells will be tested in the lab and also in mice carrying human cancers, and analysed in depth to produce maps of which genes are active in the different populations of cells. We will include tests to find out which combinations of edits work best, and include versions where CAR expression is driven by where in the genome it has been inserted.The project will adopt a new CAR against CD38 which is already being tested in our lab. It was derived from the monoclonal antibody Daratumumab, which is being widely investigated against multiple myeloma and autoimmune disorders and so has an extensively mapped safety and toxicity profile. Plans for an 'off-the-shelf' universal CAR38 therapy to explore deep cell-mediated clearance of disease are being formulated, and this project will help determine which strategies to use to manufacture the CAR T cell banks ahead for a trial. If CAR T cell production can be switched to entirely virus-free platforms, it will save time and money, and in the long run allow many more versions to be advanced to a clinical stage.

我们已经使用大奥蒙德街医院的专业设施将许多实验性嵌合抗原受体（CAR）T细胞疗法投入临床试验。这些药物有助于治疗患有某些类型白血病的儿童和成人，否则他们将没有进一步的选择。这些项目从实验室的概念和测试，到洁净室的生产和患者的临床使用。所有这些都依赖于使用失能病毒将基因添加到T细胞中以重新编程它们。一些病毒CAR T细胞使用基因组编辑进行了额外的改变-分子剪刀剪下其他基因的片段-这使得“通用”T细胞可以在没有任何匹配的情况下使用。现在，使用类似的技术来切割基因和插入新的基因已经成为可能，这意味着我们可以在不需要病毒添加CAR基因的情况下设计细胞。我们认为这种方法可以消除供应瓶颈，特别是为每种新的CAR制备病毒储备，这是临床试验的一个主要问题。到目前为止，我们在实验室工作中已经证明，使用CRISPR插入CAR的效率几乎与使用病毒时一样高，并且细胞在短期培养皿实验中的行为和功能也一样好。我们现在需要确保这个过程在更大的规模上工作，以一种可以在洁净室中使用的方式，并且当细胞在动物体内激活并对人类白血病做出反应时，它们的行为在更长的时间内没有不同。这个应用程序将使用现有的慢病毒或新的无病毒策略来制造和比较CRISPR CAR T细胞。这些测试将在适合未来人体试验的规模上进行，并在可以转移到洁净室环境的条件下进行。这些细胞将在实验室和携带人类癌症的小鼠中进行测试，并进行深入分析，以绘制不同细胞群中哪些基因活跃的图谱。我们将进行测试，以找出哪些编辑组合效果最好，并包括CAR表达由其插入基因组中的位置驱动的版本。该项目将采用一种针对CD 38的新CAR，该CAR已经在我们的实验室进行了测试。它来源于单克隆抗体达雷妥尤单抗，目前正在对多发性骨髓瘤和自身免疫性疾病进行广泛研究，因此具有广泛的安全性和毒性特征。目前正在制定一种“现成的”通用CAR 38疗法的计划，以探索深度细胞介导的疾病清除，该项目将有助于确定使用何种策略来制造CAR T细胞库进行试验。如果CAR T细胞的生产可以转换到完全无病毒的平台上，这将节省时间和金钱，从长远来看，可以使更多的版本进入临床阶段。