There and back again - reprogramming gamma/delta T-cells (gdT) into iPSCs in order to differentiate them back into (gdT) for cancer immunotherapy

人类T细胞产品的生物处理已经开始给癌症治疗带来革命性的变化，特别是在嵌合抗原受体(CAR)用于白血病及其他领域。在这个项目中，我们的目标是在UCL生物工程部门建立我们各自的优势。(例如1，2)和新加坡生物处理技术研究所(BTI)的Steve oh团队(例如3，4)-通过研究一种新的、可能更有用的方法来生产转基因和靶向特异性GDT细胞，用于未来的免疫治疗。BTI已经开发了1)将T细胞重新编程为hiPSC和2)定向分化为造血干细胞(HSC)的专利技术。HSC可以通过组合生长因子或激活转录因子(如Foxp3)进一步分化为T细胞。BTI已经证明了通过激活Pax6转录因子将HiPSC分化为视网膜色素上皮细胞(RPE)的能力。这项技术刚刚作为新的平台专利提交。一种类似的方法可能对促进T细胞分化起作用。学生的目标是通过培养外周血单个核细胞和其他上皮来源的GDT细胞来实现这一点。然后，他将使用已建立的程序(5，6)从GDT中获得HiPSCs，然后这些HiPSCs将被维持并传代为干细胞(SC)，并详细描述其特征。然后，学生将(重新)将GDT衍生的HiPSC重新区分为HSC，基本上如前所述(5，6)。随后，她将尝试从这些HSC中衍生出完全分化的GDT。作为该项目的一部分，我们将能够在SC阶段以一种方便的方式对GDT进行基因工程--我们假设这将导致工程基因及其表达的保留。这可能涉及TCR、共刺激分子、嵌合抗原受体(CARS)或MHC(人类白细胞抗原)1等基因的“敲除”。巴里萨·M等，《科学评论》，2017年，7(1)：28052。Fisher J.等人，Mol Ther.2017年26(2)：3543。[2]Lee等，Biotechnol J.2018，13(4)：e17005674.Sivalingam J.等，血液学，2018,103(7)：e2795。Watanabe D.等，干细胞移植医学。2018年，7(1)：346。曾杰等，《公共科学图书馆·综合》。2019，14(5)：e0216815

Bioprocessing of human T-cell products have begun to revolutionize cancer treatments, in particular in the fields of chimeric antigen receptor (CAR) use in leukaemias and beyond. In this project we aim to build on our respective strengths in the UCL Biochemical Engineering dept. (e.g. 1, 2) and the Steve Oh group at the Bioprocessing Technology Institute (BTI) in Singapore (e.g. 3, 4) - by investigating a novel and potentially more useful way to produce genetically modified and target-specific gdT cells for use in immunotherapies in the future. BTI has developed patented technologies in 1) reprogramming T cells into hiPSC and 2) directed differentiation into haemapoietic stem cells (HSC). HSC can be further differentiated to T cells by a combination of growth factors or activation of transcription factors such as Foxp3. BTI has demonstrated the ability to differentiate hiPSC to retinal pigment epithelial cells (RPE) by activation of Pax6 transcription factor. This technology has just been filed as a new platform patent. A similar approach is likely to work for accelerating T cell differentiation.The student will aim to do this by culturing gdT cells from peripheral blood mononuclear cells and other epithelial sources. He will then derive hiPSCs from the gdT using established procedures (5, 6) These hiPSCs will then be maintained and passaged as stem cells (SC) and characterised in detail. The student will then (re)differentiate the gdT-derived hiPSCs back into HSC, essentially as previously described (5, 6). From these HSC she will subsequently attempt to derive fully differentiated gdT. As part of this project, we will importantly be able to genetically engineer the gdT in a convenient way during the SC stage - which we hypothesise will lead to retention of the engineered gene and its expression. This could involve a TCR, co-stimulatory molecules, chimeric antigen receptors (CARs), or 'knock-out' of genes such as MHC (HLA).1. Barisa M. et al., Sci Rep. 2017, 7(1): 28052. Fisher J. et al., Mol Ther. 2017, 26(2): 3543. Lee et al., Biotechnol J. 2018, 13(4): e17005674. Sivalingam J. et al., Haematologica 2018, 103(7): e2795. Watanabe D. et al., Stem Cells Transl Med. 2018, 7(1): 346. Zeng J. et al., PLoS One. 2019, 14(5): e0216815