EXPANSION AND GENETIC TRANSDUCTION OF EBV-SPECIFIC CTLS

最近的研究表明，EBV特异性CTL对免疫功能低下患者的EBV淋巴瘤的预防和治疗是非常有效的。 然而，这种新兴的治疗策略的广泛应用和扩展到其他EBV相关淋巴增生性疾病（LPD）将需要可靠和有效的手段来产生足够量的肿瘤抗原特异性CTL以用于有效的患者治疗。 提出的研究的主要目的是开发和实施一种独特的方法，用于离体衍生、响应于自体EBV转化的淋巴母细胞样细胞系或LMP2a转导的树突状细胞的抗原特异性扩增，以及在灌注的临床规模生物反应器系统中逆转录病毒介导的EBV特异性CTL的基因标记。 将对以下假设进行检验：在AastromReplicell TM细胞生产系统中，使用流通技术，连续单次培养基交换将能够增强T细胞生产、功能和遗传转导。 第二阶段计划将重点关注符合GMP法规的工艺优化和全自动化。 将在临床试验中测试自动生物反应器系统，以评估基因标记的EBV特异性CTL用于治疗EBV相关淋巴瘤和霍奇金病的安全性和有效性。 自动化临床规模的生物反应器系统将能够有效和可靠地生产用于EBV相关LPD和其他癌症的抗原特异性免疫治疗的高功能基因标记的T细胞。用于治疗量的EBV特异性CTL的离体衍生、扩增和基因标记的封闭的、符合GMP的生物反应器系统对于预防和治疗免疫功能低下患者中的EBV相关LPD具有直接的商业价值。 用于抗原特异性T细胞扩增的自动化方法的开发将减少开放式方法步骤以及设备和劳动力的相关成本，从而使得能够以可靠且成本有效的方式广泛应用基于T细胞的癌症免疫疗法。

Recent studies have demonstrated the EBV-specific CTLs are highly effective for prophylaxis of therapy of EBV-lymphoma in immunocompromised patients. However, widespread application of this emerging treatment strategy and extension to other EBV- associated lymphoproliferative diseases (LPDs) will require a reliable and efficient means to generate sufficient quantities of tumor antigen-specific CTLs for effective patient therapy. The primary objective of the proposed studies is to develop and implement a unique process for ex vivo derivation, antigen- specific expansion in response to autologous EBV-transformed lymphoblastoid cell lines or LMP2a-transduced dendritic cells, and retroviral-mediated gene-marking of EBV-specific CTLs in a perfused clinical-scale bioreactor system. The hypothesis that continuous single-pass medium exchange will enable enhanced T- cell production, function, and genetic transduction using flow- through technology in the AastromReplicell TM Cell Production System will be tested. The Phase II program will focus on optimization and full automation of the processes in compliance with GMP regulations. The automated bioreactor system will be tested in clinical trials to assess the safety and efficacy of gene-marked EBV-specific CTLs for therapy of EBV-associated lymphoma and Hodgkin s disease. An automated clinical scale bioreactor system would enable efficient and reliable production of highly functional gene-marked T-cells for antigen-specific immunotherapy of EBV-associated LPDs and other cancers. PROPOSED COMMERCIAL APPLICATION A closed, GMP-compliant, bioreactor system for ex vivo derivation, expansion and gene-marking of therapeutic quantities of EBV-specific CTLs would have immediate commercial value for prevention and treatment of EBV-associated LPDs in immunocompromised patients. The development of an automated process for antigen-specific T-cell expansion will reduce open- process steps and associated costs in equipment and labor, thus enabling widespread application of T-cell-based cancer immunotherapies in a reliable and cost-effective fashion.