摘要
目的制备并筛选靶向人表皮生长因子受体3(human epidermal growth factor receptor 3,Her3)的单克隆抗体。方法用CHO系统克隆表达Her3的胞外区,获得重组蛋白h GH-Her3-ECD,用该重组蛋白免疫BALB/c小鼠,并经常规细胞融合杂交瘤技术制备单克隆抗体。对制备的单克隆抗体进行抑制Her3与其配体HRG的结合试验、亲和力测定、亚型鉴定、结合抗原结构域的确定、识别肿瘤细胞表面Her3抗原的FACS分析及对A431细胞增殖抑制分析。结果获得100多株靶向Her3的单克隆抗体,其中927、1002、1050、993、1044、1056株具有较强的抑制Her3与HRG结合活性,均为Ig G1型,亲和力较好。927、1002、1050株与Her3第Ⅰ结构域结合,993、1044、1056株与Her3第Ⅲ结构域结合,均能结合A431和Bx Pc3表面的Her3分子,可明显抑制A431细胞的增殖。结论获得的靶向Her3单克隆抗体基本满足开发治疗性单抗的要求,具有成为治疗性单克隆抗体的潜在条件。
Objective To p repare and screen the monoclonal antibodies(Mc Abs) targeting human epidermal growth factor receptor 3(Her3). Methods The extracellular domain of Her3 protein was expressed by using CHO cell system.BALB / c mice were immunized by the obtained recombinant protein h GH-Her3-ECD, based on which Mc Abs were prepared by using traditional hybridoma technology and screed by blocking Her3-HRG binding assay, affinity assay FACS assay and A431 cell proliferation inhibition assay, identified for subtype, and determined for antigen binding domain.Results Of the more than 100 kinds of Mc Abs obtained, those secreted by strains 927, 1002, 1050, 993, 1044 and 1056 showed strong inhibiting activities to binding of Her3 to HRG, all of which were of subtype Ig G1 and showed good affinities. The Mc Abs secreted by strains 927, 1002 and 1050 showed binding to domain Ⅰof Her3, while those by strains993, 1044 and 1056 to domain Ⅲ, all of which showed binding to the Her3 molecules on surface of A431 and Bx Pc3 cells, and inhibited the proliferation of A431 cells significantly. Conclusion The obtained Mc Abs targeting Her3 basically met the need for development of therapeutic antibodies of cancers.
出处
《中国生物制品学杂志》
CAS
CSCD
2016年第11期1162-1168,共7页
Chinese Journal of Biologicals
基金
上海市2015年度"科技创新行动计划"生物医药领域科技支持项目(15431905100)
关键词
人表皮生长因子受体3
单克隆抗体
肿瘤
Human epidermal growth factor receptor 3(Her3)
Monoclonal antibody(Mc Ab)
Tumor