Transcriptional targeting to anterior pituitary lactotrophic cells using recombinant adenovirus vectors in vitro and in vivo in normal and estrogen/sulpiride-induced hyperplasic anterior pituitaries
Transcriptional targeting to anterior pituitary lactotrophic cells using recombinant adenovirus vectors in vitro and in vivo in normal and estrogen/sulpiride-induced hyperplasic anterior pituitaries
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DOI：
10.1210/en.141.9.3493
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发表时间：
2000-09-01
期刊：
影响因子：
4.8
通讯作者：
Castro, MG
中科院分区：
文献类型：
--
作者：
Southgate, TD;Windeatt, S;Castro, MG
使用垂体细胞类型特异性启动子是实现病毒载体编码的转基因的垂体细胞类型特异性转录靶向的一种强大的分子工具。最近有人提出，治疗性基因的转录靶向可以作为一种治疗垂体疾病的基因治疗策略。我们成功地利用重组腺病毒载体中编码的人催乳素启动子(HPrl)在体内外靶向单纯疱疹病毒1型胸苷激酶(HSV1-TK)或β-半乳糖苷酶的转基因表达。在功能上，利用hPr1启动子将HSV1-TK的表达限制在乳酸型肿瘤细胞中，导致在更昔洛韦(GCV)存在的情况下，针对特定细胞类型诱导增乳型GH3肿瘤细胞的凋亡。在皮质营养型AtT20细胞系中，我们既没有检测到HSV1-TK的表达，也没有检测到GCV存在时的细胞凋亡。重组腺病毒载体中编码的hPr1启动子也将转基因表达限制在原代培养的垂体前叶(AP)中的哺乳细胞，更重要的是，在体内的垂体前叶内。当将hPr1启动子驱动的HSV1-TK用于雌激素/舒必利诱导的AP内原位增殖的体内模型时，治疗既不能减少腺体的重量，也不能减少体内转导区域内的乳汁细胞数量，也不能降低循环中的PRL水平。这与在相同的实验范式中由人巨细胞病毒启动子(HCMV)驱动HSV1-TK的表达形成了鲜明对比，后者在减轻脑垂体重量和循环PRL水平方面是有效的。我们的研究结果对垂体瘤基因治疗策略的设计具有重要的指导意义。我们证明了体内动物模型的选择，即AP腺体内原位腺瘤的选择，以及特定的基因治疗策略的选择，即使用强大的无处不在的启动子。较弱但细胞类型特异的启动子，决定了实验性的治疗结果。
The use of pituitary cell type-specific promoters is a powerful molecular tool to achieve pituitary cell type-specific transcriptional targeting of transgenes encoded by viral vectors. It has recently been proposed that transcriptional targeting of therapeutic genes could be harnessed as a gene therapy strategy for the treatment of pituitary disease. We describe the successful use of the human PRL promoter (hPrl) encoded within recombinant adenovirus vectors to target transgene expression of Herpes Simplex Virus Type 1-Thymidine Kinase (HSV1-TK) or beta-galactosidase to lactotrophic cells in vitro and in, vivo. Functionally, the restriction of expression of HSV1-TK to lactotrophic tumor cells, using the hPrl promoter, resulted in the cell type-specific induction of apoptosis in the lactotrophic GH3 tumor cell line, in the presence of ganciclovir (GCV). In the corticotrophic AtT20 cell line, we detected neither HSV1-TK expression, nor apoptosis in the presence of GCV. The hPrl promoter encoded within a recombinant adenoviral vector also restricted transgene expression to lactotrophic cells in primary anterior pituitary (AP) cultures, and importantly, within the anterior pituitary gland in vivo. When the HSV1-TK driven by hPrl promoter was used in an in vivo model of estrogen/sulpiride lactotroph induced hyperplasia within the AP in situ, the treatment was not effective in either reducing the weight of the gland, the number of lactotrophic cells within the transduced area in vivo, or the circulating PRL levels. This is in contrast to the human cytomegalovirus promoter (hCMV) driving expression of HSV1-TK in the same experimental paradigm, which was effective in reducing pituitary weight and circulating PRL levels. Our results have important implications in the design of gene therapy strategies for pituitary tumors. We demonstrate that both the choice of the in vivo animal model, i.e. adenoma in the AP gland in situ, and the particular gene therapy strategy chosen, i.e. use of strong ubiquitous promoters us. weaker but cell type-specific promoters, determine the experimental therapeutic outcome.