Quantitative Analysis of Retroviral Gene Delivery

此项目是主动提交的。该项目的目标是制定策略，以提高逆转录病毒介导的基因转移的效率和实用性。逆转录病毒是目前基因治疗临床试验中广泛使用的载体系统，它促进了逆转录病毒载体技术的快速发展。尽管如此，逆转录病毒的基因转移效率仍然令人失望地低，阻碍了逆转录病毒基因治疗作为一种有效的临床工具的出现。研究发现，带电分子对基因转移过程的效率有很大影响。具体地说，带正电荷的化合物被证明可以促进基因转移，而带负电荷的化合物则抑制这一过程。本项目旨在完成以下工作：(1)详细研究带电化合物促进和抑制逆转录病毒基因转移过程的机制，(2)建立一个数学框架来分析带电化合物对病毒运输和转导的影响，以及(3)使用体外和体内基因治疗方案比较动物模型中的基因转移效率，该基因治疗方案旨在增强逆转录病毒载体和靶细胞之间的静电相互作用。为了实现这些目标，研究人员将利用一套互补的酶和免疫组织化学分析、分子成像技术和细胞荧光分析来量化逆转录病毒基因转移过程的各个步骤。对生物分子运输和动力学反应数据的工程分析将有助于构建逆转录病毒运输的综合模型。

This project is an unsolicited submission. The goal of this project is to develop strategies to enhance the efficiency and utility of retrovirus-mediated gene transfer. Retroviruses are a widely utilized vector system in current gene therapy clinical trials, which has resulted in the rapid advancement of retroviral vector technology. Despite this, retroviral gene transfer efficiency has remained disappointingly low, hampering the emergence of retroviral gene therapy as an effective clinical tool. It has been found that charged molecules greatly influence the efficiency of the gene transfer process. Specifically, positively charged compounds have been shown to enhance gene transfer, whereas negatively charged compounds inhibit the process. This project seeks to accomplish the following: (1) to investigate in detail the mechanism of charged compound enhancement and inhibition of the retroviral gene transfer process, (2) to develop a mathematical framework for analyzing the effect of charged compounds on virus transport and transduction, and (3) to compare gene transfer efficiency in an animal model using ex vivo and in vivo gene therapy protocols optimized to enhance the electrostatic interaction between the retrovirus vector and target cell. To achieve these goals, the investigators will utilize a complementary set of enzymatic and immunohistochemical assays, molecular imaging techniques, and cytofluorometric analyses to quantitate various steps of the retroviral gene transfer process. Engineering analysis of the resultant biomolecular transport and kinetic reaction data will be used to facilitate the construction of an integrative model of retrovirus transport.