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.

我们利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，否则他们将没有其他选择来治疗他们的疾病。这些项目从实验室的概念和测试，到洁净室的制造和患者的临床使用。所有这些都依赖于使用失效病毒将基因添加到 T 细胞中以对其进行重新编程。一些病毒 CAR T 细胞通过基因组编辑（分子剪刀剪掉其他基因的片段）进行了额外的改变，这使得“通用”T 细胞无需任何匹配即可使用。现在，使用类似的技术来切割基因和插入新基因已经成为可能，这意味着我们可以改造细胞，而不需要病毒来添加 CAR 基因。我们认为这种方法可以消除供应瓶颈，特别是为每个新 CAR 制造病毒库存，这是临床试验的一个主要阻碍问题。迄今为止，我们的实验室工作已经表明，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。我们现在需要确保该过程以可在洁净室中使用的方式在更大范围内发挥作用，并且细胞在较长时间内激活并针对动物中的人类白血病做出反应时，其行为没有不同。该应用将使用现有的慢病毒或新的无病毒策略来添加 CAR 来制造和比较 CRISPR CAR T 细胞。这些测试的规模将适合未来的人体试验，并且条件可以转移到洁净室环境。这些细胞将在实验室以及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，以确定哪种编辑组合效果最好，并包括 CAR 表达由基因组中插入位置驱动的版本。该项目将采用一种针对 CD38 的新 CAR，该 CAR 已在我们的实验室进行测试。它源自单克隆抗体 Daratumumab，该抗体正在针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。正在制定“现成”通用 CAR38 疗法的计划，以探索深层细胞介导的疾病清除，该项目将有助于确定在试验前使用哪些策略来制造 CAR T 细胞库。如果 CAR T 细胞生产可以切换到完全无病毒的平台，将节省时间和金钱，从长远来看，可以让更多版本进入临床阶段。