Use of normo-thermic perfused organs to profile PK and infection efficiency of novel viral gene therapy vectors.

有效基因治疗的主要障碍包括血流的快速中和、肝脏中库普弗细胞的高捕获以及靶细胞感染的低效率。有针对性的基因改造可以产生慢病毒载体变体，这可能有可能克服这些障碍。然而，如果没有有效的测试系统来比较新的变种，误导性的数据可能会导致不合适的变种被优先用于临床开发。事实上，目前的临床前模型不能充分概括人类患者的规模和生理状况。啮齿动物是常用的模型，但也是一个特别差的模型，其循环速度比人类快13倍，肝脏内皮细胞隔板间隙比人类宽30-60 nm[Wisse，2008，GT]，血容量和总质量比人类低3000倍，以及非常不同的病毒预暴露和血细胞补体受体图谱[Carlisle，2009，hemed]。我们处于独特的地位，能够提供一个系统，一个常温灌流装置(NTPD)，可以在体外维持人体器官，并允许给它们注射治疗剂。持续灌流允许采集血液和胆汁样本，从而可以确定器官功能/毒性和病毒载体的药代动力学，而通过定期进行穿孔组织活检对病毒基因组和转基因产品进行采样也可以跟踪进入和整合的速率和水平。我们建议，与牛津生物医学公司合作，我们将使用NTPD来描述在牛津生物医学公司开发的新型慢病毒载体变体的PK。

Major barriers to effective gene therapy include rapid neutralisation in the blood stream, high Kupffer cell capture in the liver and low efficiency of target cell infection. Targeted genetic modification can generate lentiviral vector variants which may have the potential to overcome these barriers. However, without valid test systems with which to compare new variants, misleading data may result in unsuitable variants being prioritised for clinical development. Indeed, current pre-clinical models fail to adequately recapitulate the scale and physiology of human patients. Rodents are commonly used but are a particularly poor model, with a circulation rate 13x faster than humans, liver endothelial fenestrae gaps 30-60nm wider than in humans [Wisse,2008,GT], a blood volume and total mass 3000x lower than humans, and a very different history of virus pre-exposure and blood cell complement receptor profile [Carlisle,2009,Blood]. We are uniquely positioned to be able to provide a system, a normothermic perfusion device (NTPD), which can maintain human organs outside the body and permit their dosing with therapeutic agents. Continuous perfusion allows for blood and bile samples to be taken so that organ function/toxicity and viral vector pharmacokinetics can be defined, whilst sampling for virus genomes and transgene products by punch tissue biopsies taken at regular junctures can also track rates and levels of entry and integration. We propose that, in collaboration with Oxford Biomedica, we will use the NTPD to profile the PK of novel Lentiviral vector variants developed in Oxford Biomedica.