Hosseinzadeh Farideh, Mohammadi Saeed, Nejatollahi Foroogh
Shiraz HIV/AIDS Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.
Recombinant antibody laboratory, Department of Immunology, Shiraz University of Medical Sciences, Shiraz, Iran.
Rep Biochem Mol Biol. 2017 Oct;6(1):8-14.
Cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) molecules are expressed on T-cells and inhibit their function by inhibiting activation of subsequent T-cell molecular pathways. Blocking of CTLA-4 inhibits the growth of malignant tumor cells. Anti-CTLA-4 monoclonal antibodies activate the immune system against cancer. Due to several advantages of single-chain antibodies (scFvs) compared to monoclonal antibodies in cancer immunotherapy, specific anti-CTLA-4 scFvs (single-chain variable fragment) were selected in this study.
A phage antibody display library of scFvs was analyzed and a panning process was performed against an immunodominant epitope of CTLA-4. PCR and DNA fingerprinting were used to differentiate the specific clones. The specificity of the selected clones was investigated by phage ELISA (Enzyme-linked immunosorbent assay).
Two specific clones with frequencies of 35 and 20% were identified. The clones reacted with the corresponding epitope on ELISA, while no reactivity was observed with an unrelated peptide, M13KO7 helper phage, unrelated scFvs, or no peptide as negative controls.
Targeted therapy against cancer markers is an ideal treatment strategy. Specific human anti-CTLA-4scFvs were selected in this study. These scFvs bound the related epitope. These antibodies have the potential to be used for targeted therapy, where the blocking of CTLA4 receptor is needed. The study suggests further evaluation of the selected scFvs to reveal the effects of the selected antibodies.
细胞毒性T淋巴细胞相关抗原4（CTLA-4）分子在T细胞上表达，并通过抑制后续T细胞分子途径的激活来抑制其功能。阻断CTLA-4可抑制恶性肿瘤细胞的生长。抗CTLA-4单克隆抗体可激活针对癌症的免疫系统。由于在癌症免疫治疗中，单链抗体（scFv）相较于单克隆抗体具有若干优势，本研究选择了特异性抗CTLA-4 scFv（单链可变片段）。
分析scFv的噬菌体抗体展示文库，并针对CTLA-4的免疫显性表位进行淘选过程。采用聚合酶链反应（PCR）和DNA指纹分析来区分特异性克隆。通过噬菌体酶联免疫吸附测定（ELISA）研究所选克隆的特异性。
鉴定出两个频率分别为35%和20%的特异性克隆。这些克隆在ELISA中与相应表位发生反应，而作为阴性对照的无关肽、M13KO7辅助噬菌体、无关scFv或无肽均未观察到反应性。
针对癌症标志物的靶向治疗是一种理想的治疗策略。本研究筛选出了特异性人抗CTLA-4 scFv。这些scFv与相关表位结合。这些抗体有潜力用于需要阻断CTLA-4受体的靶向治疗。该研究建议对所选scFv进行进一步评估，以揭示所选抗体的作用效果。
