Mohammadi Mozafar, Nejatollahi Foroogh, Ghasemi Younes, Faraji Sayyid Nooreddin
Faculty of advanced medical sciences and technologies, Shiraz University of Medical Sciences, Shiraz, Iran.
Recombinant antibody laboratory, Dept. of immunology, Shiraz University of Medical Sciences, Shiraz, Iran.
Appl Biochem Biotechnol. 2017 Jan;181(1):379-390. doi: 10.1007/s12010-016-2218-1. Epub 2016 Aug 27.
Breast cancer is the most common malignancy in women. Altered expression of MUC18, a cell surface receptor, and its interaction with Wnt-5a as its ligand, affects the motility and invasiveness of breast cancer cells. In this study, we explored the Wnt-5a binding site and designed an antigenic epitope on the MUC18 receptor using in silico methods. A specific single-chain variable fragment (scFv) was isolated against the epitope by several panning processes. The binding ability of the scFv to the related epitope was evaluated in ELISA and flow cytometry. The inhibitory effects of the selected scFv on MUC18 positive cell line, MDA-MB231, was assessed by migration and invasion assays. The results demonstrated isolation of specific scFv with frequency of 40 % which showed significant binding with the epitope in both ELISA and fluorescence-activated cell sorting (FACS) analyses. The antibody inhibited the migration (76 %) and invasion (67 %) of MUC18 positive cell line. The results suggest the specific anti-MUC18 scFv as an effective antibody for breast cancer immunotherapy.
乳腺癌是女性中最常见的恶性肿瘤。细胞表面受体MUC18的表达改变及其与作为配体的Wnt-5a的相互作用，会影响乳腺癌细胞的运动性和侵袭性。在本研究中，我们利用计算机模拟方法探索了Wnt-5a的结合位点，并在MUC18受体上设计了一个抗原表位。通过多次淘选过程，分离出了针对该表位的特异性单链可变片段（scFv）。通过酶联免疫吸附测定（ELISA）和流式细胞术评估了scFv与相关表位的结合能力。通过迁移和侵袭试验评估了所选scFv对MUC18阳性细胞系MDA-MB231的抑制作用。结果表明，特异性scFv的分离频率为40%，在ELISA和荧光激活细胞分选（FACS）分析中均显示与表位有显著结合。该抗体抑制了MUC18阳性细胞系的迁移（76%）和侵袭（67%）。结果表明，特异性抗MUC18 scFv是一种用于乳腺癌免疫治疗的有效抗体。
