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

为了确保基因治疗和基因修饰细胞治疗有足够的高质量载体供应，有必要大规模和持续地生产稳定的细胞系，用于改进慢病毒（LV）载体的生产。这需要一种综合的方法，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个可扩展的生物工艺，用于生产具有最佳病毒包膜的LentiStim病毒载体，并开发一个端到端工艺，使所生产的LV用于转导具有嵌合抗原受体（CAR）的原代t细胞。从这一过程中产生的t细胞将在人源化小鼠模型中进行抗肿瘤功效测试。这将使用NOD/SCID/IL2rynull （NSG）小鼠模型，对移植物抗宿主病具有B细胞淋巴瘤细胞系（Raji）或/和原发性淋巴瘤细胞。

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.