Development of a stable, continuous lentiviral (LV) production system for improved gene delivery and cell function

稳定细胞系的可规模化和一致性生产对于改善慢病毒（LV）载体生产是必要的，以确保为基因治疗和基因修饰的细胞治疗提供足够的高质量载体。这需要一种综合方法，同时考虑细胞系的生物工程和上游和下游加工的生物工艺工程。从生物工程的角度来看，该项目将寻求开发能够同时转导和刺激T细胞（LentiStim）的病毒载体，而不需要Dynabeads或抗体，此外还将对各种病毒包膜进行系统的比较研究，以增强T细胞工程。从生物加工的角度来看，该项目将寻求建立一种可扩展的生物工艺，用于生产具有最佳病毒包膜的LentiStim病毒载体，并开发一种端到端的工艺，由此生产的LV将用于使用嵌合抗原受体（CAR）来培养原代T细胞。将在人源化小鼠模型中测试从该过程产生的T细胞的抗肿瘤功效。这将使用具有B细胞淋巴瘤细胞系（Raji）或/和原代淋巴瘤细胞的移植物抗宿主病抗性的NOD/SCID/IL 2 rynull（NSG）小鼠模型来建立。

Scalable and consistent production of stable cell lines for improved lentiviral (LV) vector production is necessary to ensure sufficient supply of high-quality vector for gene therapies and gene-modified cell therapies. This requires an integrated approach with both biological engineering of the cell lines and bioprocess engineering of the upstream and downstream processing being considered in parallel. From a biological engineering perspective, the project will seek to develop viral vectors which enable the simultaneous transduction and stimulation of T-cells (LentiStim) without the need for Dynabeads or antibodies in addition to undertaking a systematic, comparative study of various viral envelopes for enhanced T-cell engineering. From a bioprocessing perspective, the project will seek to establish a scalable bioprocess for the production of LentiStim viral vectors with the optimal viral envelope and develop an end-to-end process whereby the produced LV will be used to transduce primary T-cells with a chimeric antigen receptor (CAR). The T-cells produced from this process will be tested for anti-tumour efficacy in humanized mouse models. This will be established using NOD/SCID/IL2rynull (NSG) mouse model resistant to graft versus host disease with a B cell lymphoma cell line (Raji) or/and primary lymphoma cells.