Do retroviral vector insertions influence the fate of terminally differentiated cells such as T lymphocytes by insertional mutagenesis?

携带(Y-)逆转录病毒(RV)载体的基因转移到造血干/祖细胞可能严重影响单个细胞的命运，最终导致克隆显性甚至恶性转化。观察到的这些克隆的生存优势归因于随机载体插入原癌基因或其他信号基因。然而，目前尚不清楚逆转录病毒载体插入的如此强烈的副作用是否也会出现在终末分化细胞中。我们在这里的目的是利用外周T细胞来研究这个问题，外周T细胞是各种基因治疗策略的重要靶点。我们将使用Y-逆转录病毒载体和慢病毒载体进行剂量递增的T细胞转导，并在小鼠骨髓移植模型中分析插入位置在体外和体内的长期影响。我们的合作伙伴Baum开发的带有强大启动子/增强子元件的载体将被用来模拟最坏的情况。与SPP1230的其他申请者(Frühauf，von Laer，von Kalle)合作，我们还将分析临床基因治疗样本中T细胞中RV载体的插入情况。总而言之，这些数据应该可以对通过Y-逆转录和慢病毒基因转移导致T淋巴细胞恶性转化的风险获得新的见解。通过一个合作网络，该项目还将对特定战略计划的几个共同平台作出重大贡献，包括。插入基因组学、克隆选择模型、数学建模和矢量学。

Gene transfer with (y-)retroviral (RV) vectors into hematopoietic stem and progenitor cells may heavily influence the fate of single cells eventually leading to clonal dominance or even malignant transformation. The observed survival advantage of those clones has been attributed to random vector insertions into proto-oncogenes or other signaling genes. However, it is not known yet whether such strong side effects of retroviral vector insertions may also be expected with terminally differentiated cells. We here aim at investigating this question using peripheral T cells, an important target of various gene therapy strategies. We will perform dose-escalated T cell transduction with both y-retro and lentiviral vectors and analyze the long-term impact of insertion sites in vitro and in vivo, in murine BMT models. Vectors with strong promotor/enhancer elements developed by our partner Baum will be used to mimic a worst case scenario . In cooperation with other applicants of this SPP1230 (Frühauf, von Laer, von Kalle) we will also analyze RV vector insertions in T cells from clinical gene therapy samples. Together this data should allow to get novel insights into the risk of malignant transformation of T lymphocytes by both y-retro- and lentiviral gene transfer. Through a network of cooperation this project will also significantly contribute to several of the common platforms of the given SPP, incl. insertional genomics, models of clonat selection, mathematical modeling and vectorology.