Key Laboratory of Pediatric Hematology and Oncology Ministry of Health and Pediatric Translational Medicine Institute, Shanghai Children's Medical Center, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
Key Laboratory of Pediatric Hematology and Oncology Ministry of Health and Pediatric Translational Medicine Institute, Shanghai Children's Medical Center, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
Talanta. 2023 Oct 1;263:124722. doi: 10.1016/j.talanta.2023.124722. Epub 2023 May 24.
Screening novel aptamers for recombinant protein detection is of great significance in industrial mass production of antibody drugs. In addition, construction of structurally stable bispecific circular aptamers (bc-apts) may provide a tumor-targeted treatment strategy by simultaneously binding two different cell types. In this study, we obtained a high-affinity hexahistidine tag (His-tag)-binding aptamer 20S and explored its application in recombinant protein detection and T cell-based immunotherapy. We developed a new molecular beacon (MB) 20S-MB to detect His-tagged proteins in vitro and in vivo with high sensitivity and specificity, and the results showed high consistency with the enzyme-linked immunosorbent assay (ELISA). Moreover, we constructed two kinds of bc-apts by cyclizing 20S or another His-tag-binding aptamer, 6H5-MU, with Sgc8, which specifically recognizes protein tyrosine kinase 7 (PTK7) on tumor cells. After forming a complex with His-tagged OKT3, an anti-CD3 antibody for T cell activation, we utilized these aptamer-antibody complexes (ap-ab complex) to enhance cytotoxicity of T cells by linking T cells and target cells together, and 20S-sgc8 exhibited antitumor efficacy superior to that of 6H5-sgc8. In conclusion, we screened a novel His-tag-binding aptamer and used it to construct a new type of MB for rapid detection of recombinant proteins, as well as establish a feasible approach for T cell-based immunotherapy.
筛选新型适体用于重组蛋白检测在抗体药物的工业大规模生产中具有重要意义。此外，构建结构稳定的双特异性环状适体（bc-apts）可能通过同时结合两种不同的细胞类型提供一种肿瘤靶向治疗策略。在本研究中，我们获得了一种高亲和力的六组氨酸标签（His-tag）结合适体 20S，并探索了其在重组蛋白检测和基于 T 细胞的免疫治疗中的应用。我们开发了一种新的分子信标（MB）20S-MB，可高灵敏度和特异性地检测体外和体内的 His 标记蛋白，结果与酶联免疫吸附测定（ELISA）高度一致。此外，我们通过将 20S 或另一种 His 标记结合适体 6H5-MU 环化与 Sgc8 构建了两种 bc-apts，Sgc8 特异性识别肿瘤细胞上的蛋白酪氨酸激酶 7（PTK7）。与激活 T 细胞的抗 CD3 抗体 OKT3 形成复合物后，我们利用这些适体-抗体复合物（ap-ab 复合物）将 T 细胞和靶细胞连接在一起，增强 T 细胞的细胞毒性，20S-sgc8 表现出优于 6H5-sgc8 的抗肿瘤功效。总之，我们筛选了一种新型 His 标记结合适体，并将其用于构建用于快速检测重组蛋白的新型 MB，以及建立基于 T 细胞的免疫治疗的可行方法。
