The vascular endothelial growth factor (VEGF) is an essential factor to pathologic angiogenesis. Disruption of VEGF/VEGF receptor interaction in cancer patients inhibits the development of new and pre-existing tumor blood vessels. Consequently, VEGF becomes an important therapeutic target for handling solid tumors. In this work, human VEGF was produced in the culture supernatant of SiHa cells transduced with a replication-defective adenoviral vector (pAdhVEGF₁₂₁) encoding this molecule. The 35 kDa VEGF₁₂₁ homodimer was obtained from clarified culture media as a glycosylated protein. VEGF₁₂₁ expression levels were strictly dependent on the adenoviral viral load used. VEGF₁₂₁ was produced with purity over 98% after a single step chromatography by immobilized metal affinity chromatography. Additionally, VEGF₁₂₁ binds Bevacizumab antibody with a K_D of 7 nM. Biological characterization by mitogenic assay in HUVEC and ECV-304 cells showed that VEGF₁₂₁ stimulates cell proliferation in a dose-dependent manner in both cells. Finally, the neovascularization activity of VEGF₁₂₁ was demonstrated by vascular permeability assays in matrigel plug-bearing mice, showing significantly increased vasculature leakage after treatment with VEGF₁₂₁. Consequently, transduction of SiHa cells with adenovirus is a suitable alternative for manufacture heterologous proteins of therapeutic interest.

中文翻译：

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腺病毒载体转导的SiHa细胞中产生的人血管内皮生长因子的表达和表征。

血管内皮生长因子（VEGF）是病理性血管生成的重要因素。癌症患者中VEGF / VEGF受体相互作用的破坏抑制了新的和先前存在的肿瘤血管的发育。因此，VEGF成为治疗实体瘤的重要治疗靶标。在这项工作中，在用编码该分子的复制缺陷型腺病毒载体（pAdhVEGF ₁₂₁）转导的SiHa细胞的培养上清液中产生了人类VEGF 。35 kDa VEGF ₁₂₁同型二聚体是从澄清的培养基中获得的糖基化蛋白。VEGF ₁₂₁表达水平严格取决于所用的腺病毒载量。血管内皮生长因子₁₂₁通过固定化金属亲和色谱法进行一步色谱后，得到纯度超过98％的产物。另外，VEGF _121以7nM的K _D结合贝伐单抗抗体。在HUVEC和ECV-304细胞中通过有丝分裂测定的生物学特性表明，VEGF ₁₂₁在两种细胞中均以剂量依赖性方式刺激细胞增殖。最后，通过在基质胶带塞小鼠中的血管通透性试验证明了VEGF ₁₂₁的新血管形成活性，显示在用VEGF ₁₂₁处理后血管渗漏显着增加。因此，用腺病毒转导SiHa细胞是制备治疗性异源蛋白质的合适选择。