表达可溶性TβRⅡ胞外区腺病毒载体的构建及功能试验研究

Construction of an Adenovirus Vector Expressing the Soluble Extracellular Domain of TGF-βRⅡ and Tests to Determine its Anti-Cancer Effects

目的:探讨表达可溶性TβRⅡ胞外区片段的腺病毒载体构建,及其在肿瘤生物治疗学方面的作用机制和效果。方法:RT-PCR法克隆鼠源性TβRⅡ胞外区编码序列,采用AD-easyXLAdenovirusVectorSystem试剂盒构建表达上述基因的腺病毒载体。通过ELISA试验验证表达可溶性TβRⅡ胞外区序列的腺病毒载体转染细胞后所具有的阻断TGF-β的作用。应用于小鼠体内试验,观察该病毒载体对免疫效应细胞抗肿瘤活性的影响,从而评价其在肿瘤治疗方面的作用。结果:克隆了鼠源性TβRⅡ胞外区编码序列并成功地构建了表达这些基因的腺病毒载体。功能试验表明由腺病毒表达的TβRⅡ胞外区片段具有阻断TGF-β信号通路的作用。体内试验则表明应用于荷瘤小鼠可抑制肿瘤的发展。结论:表达可溶性TβRⅡ胞外区片段的腺病毒载体可显著地提高免疫细胞的杀伤能力,抑制肿瘤生长,为肿瘤的生物学治疗提供了新的研究方向。

Objective： To construct an adenovirus vector expressing the soluble extracellular domain of TGF-βR Ⅱ and to evaluate the function of the vector. Methods： The soluble extracellular domain of the murine TGF-βR Ⅱ gene was cloned by RT-PCR and inserted into an adenovirus vector using the AD-easy XL Adenovirus Vector System. Western blot showed that eukaryotic cells containing the vector expressed the soluble extracellular domain of TGF-βR Ⅱ. The presence of the soluble extracellular domain of TGF-βR Ⅱ was detected using an ELISA, and the anti-cancer effect was evaluated by tumor inhibition tests in vivo. Results： The soluble extracellular domain of the mufine TGF-βR Ⅱ gene was successfully cloned into an adenovirus vector. Functional tests showed that the soluble extracellular domain of TGF-βR Ⅱ could block TGF-β signaling in vitro after infecting eukaryotic cells with the newly constructed adenovirus vector. Tumor inhibition tests showed that the adenovirus vector containing the soluble extracellular domain of TGF-βR Ⅱ could inhibit tumorigenesis in mice. Conclusion： Expression of the soluble extracellular domain of TGF-βR Ⅱ improved the anti-canoer immune response and inhibited cancer progression. Using a construct such as this vector provides a potential new treatment option.