Engineered long terminal repeats of retroviral vectors enhance transgene expression in hepatocytes in vitro and in vivo.
Engineered long terminal repeats of retroviral vectors enhance transgene expression in hepatocytes in vitro and in vivo.
复制标题
DOI：
10.1016/j.ymthe.2003.08.005
复制
发表时间：
2003-11
期刊：
影响因子：
--
通讯作者：
K. Yamaguchi;K. Itoh;N. Ohnishi;Y. Itoh;C. Baum;T. Tsuji;Toshikazu Nagao;H. Higashitsuji;T. Okanoue;J. Fujita
中科院分区：
文献类型：
--
作者：
K. Yamaguchi;K. Itoh;N. Ohnishi;Y. Itoh;C. Baum;T. Tsuji;Toshikazu Nagao;H. Higashitsuji;T. Okanoue;J. Fujita
为了分析逆转录病毒在肝细胞中表达的重要元件，在肝细胞癌细胞系中分析了脾病灶形成病毒（SFFVp）红细胞增多株长末端重复序列（LTR）U3区的顺式作用元件。双作用元件位于上游区域的直接重复，正调控逆转录病毒的表达，被确定。转录因子NFAT 5和Sp1在多种组织中广泛表达，与这些元件结合。为了增加特异性而不降低逆转录病毒在肝细胞中表达的效力，这些元件被来自B型肝炎病毒增强子II区的序列取代。新型载体SF-Hep 3和SF-Hep 5（用于肝细胞3和5的基于SFFV β的载体）用这些工程化的LTR开发。这些载体的工程化LTR仅在体外增强了肝癌细胞系中逆转录病毒的表达。在鼠肝细胞中，这些载体与基于Moloney鼠白血病病毒的载体或含有野生型SFFV β LTR的载体相比，分别增加转基因表达4- 9倍或3.5- 5倍。
To analyze the important elements for retroviral expression in hepatocytes,cis-acting elements in the U3 region of the long terminal repeat (LTR) of the polycythemic strain of spleen focus-forming virus (SFFVp) were analyzed in a hepatocellular carcinoma cell line. Twocis-acting elements located within the upstream region of the direct repeat, which positively regulated retroviral expression, were identified. Transcription factors NFAT5 and Sp1, which are ubiquitously expressed in a variety of tissues, bound to these elements. To increase specificity without lowering the potency of retroviral expression in hepatocytes, these elements were replaced by a sequence derived from the hepatitis B virus enhancer II region. Novel vectors, SF-Hep3 and SF-Hep5 (SFFVp-based vector for hepatocytes 3 and 5), were developed with these engineered LTRs. The engineered LTRs of these vectors enhanced the retroviral expression only in hepatocellular carcinoma cell linesin vitro. These vectors also increased transgene expression 4- to 9-fold or 3.5- to 5-fold in comparison with a Moloney murine leukemia virus-based vector or a vector containing the wild-type LTR of SFFVp, respectively, in murine hepatocytesin vivo.