Herpesvirus saimiri (HVS) has the capacity to incorporate large amounts of heterologous DNA and can infect many different human cell types. To develop its potential as a gene therapy vector, we cloned herpes simplex virus thymidine kinase (TK) gene into the HVS genome in the form of an enhanced green fluorescent protein (EGFP) fusion protein, using a cosmid-based approach. At multiplicity of infection = 100 over 90% of human leukemic K562 and Jurkat cells were transduced with HVS/EGFP-TK. Conditions of no selective pressure expression were maintained at > 92% per cell division. Expression of the EGFP-TK fusion protein rendered transfected leukaemic cells sensitive to cytotoxic treatment with the prodrugs ganciclovir (GCV) and (E)-5-(2-bromovinyl)-2'deoxyuridine (BVDU) at concentrations as low as 10 ng/ml. The viral vector was also screened against a panel of colorectal and pancreatic carcinoma cell lines. All cell lines were transduced but showed a range of sensitivity to infection. Three of the most easily transduced cell lines: Mia PaCa, HCT116 and SW948 transduced with HVS/EGFP-TK were effectively ablated by subsequent treatment with GCV or BVDU. Our results show that in its current form HVS/EGFP-TK could be utilized as an antitumour agent, or it could be developed further by inclusion of a therapeutic gene, with TK presence ensuring a mechanism of controlled removal of modified cells when no longer necessary. These results suggest that HVS/EGFP-TK has a great potential for a number of gene therapy applications.

中文翻译：

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胸腺嘧啶核苷激酶在人类白血病和结肠直肠细胞中的功能性表达，通过基于疱疹病毒赛米里的载体作为EGFP融合蛋白传递。

saimiri疱疹病毒（HVS）具有整合大量异源DNA的能力，并且可以感染许多不同的人类细胞类型。为了开发其作为基因治疗载体的潜力，我们使用基于粘粒的方法，以增强的绿色荧光蛋白（EGFP）融合蛋白的形式将单纯疱疹病毒胸苷激酶（TK）基因克隆到HVS基因组中。感染复数= 100时，用HVS / EGFP-TK转导了90％以上的人类白血病K562和Jurkat细胞。无选择性压力表达的条件保持在每细胞分裂＞ 92％。EGFP-TK融合蛋白的表达使转染的白血病细胞对前药更昔洛韦（GCV）和（E）-5-（2-溴乙烯基）-2'脱氧尿苷（BVDU）的细胞毒性处理敏感，浓度低至10 ng /毫升 还针对一组结肠直肠和胰腺癌细胞系筛选了病毒载体。所有细胞系均被转导，但显示出对感染的敏感性范围。通过随后用GCV或BVDU处理有效消融了三种最容易转导的细胞系：Mia PaCa，HCT116和SW948（通过HVS / EGFP-TK转导）。我们的结果表明，HVS / EGFP-TK以其当前形式可以用作抗肿瘤药，或者可以通过包含治疗基因来进一步开发，而TK的存在确保了不再需要时可控地去除修饰细胞的机制。这些结果表明，HVS / EGFP-TK在许多基因治疗应用中具有巨大潜力。所有细胞系均被转导，但显示出对感染的敏感性范围。通过随后用GCV或BVDU处理有效消融了三个最容易转导的细胞系：Mia PaCa，HCT116和SW948（通过HVS / EGFP-TK转导）。我们的结果表明，HVS / EGFP-TK以其当前形式可以用作抗肿瘤药，或者可以通过包含治疗基因来进一步开发，而TK的存在确保了不再需要时可控地去除修饰细胞的机制。这些结果表明，HVS / EGFP-TK在许多基因治疗应用中具有巨大潜力。所有细胞系均被转导，但显示出对感染的敏感性范围。通过随后用GCV或BVDU处理有效消融了三个最容易转导的细胞系：Mia PaCa，HCT116和SW948（通过HVS / EGFP-TK转导）。我们的结果表明，HVS / EGFP-TK以其当前形式可以用作抗肿瘤药，或者可以通过包含治疗基因来进一步开发，而TK的存在确保了不再需要时可控地去除修饰细胞的机制。这些结果表明，HVS / EGFP-TK在许多基因治疗应用中具有巨大潜力。我们的结果表明，HVS / EGFP-TK以其当前形式可以用作抗肿瘤药，或者可以通过包含治疗基因来进一步开发，而TK的存在确保了不再需要时可控地去除修饰细胞的机制。这些结果表明，HVS / EGFP-TK在许多基因治疗应用中具有巨大潜力。我们的结果表明，HVS / EGFP-TK以其当前形式可以用作抗肿瘤药，或者可以通过包含治疗基因来进一步开发，而TK的存在确保了不再需要时可控地去除修饰细胞的机制。这些结果表明，HVS / EGFP-TK在许多基因治疗应用中具有巨大潜力。