Hosseini Ghatar Reza, Soltantoyeh Tahereh, Bahadori Motahareh, Khoshnoodi Jalal, Golsaz Shirazi Forough, Jeddi Tehrani Mahmood, Amiri Mohammad Mehdi, Shokri Fazel
Department of Immunology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran.
Iran J Immunol. 2017 Sep;14(3):200-214.
Human epidermal growth factor receptor 2 (HER2) has a crucial role in several malignancies. The extracellular domain of HER2 (HER2-ECD) has been extensively employed as an important target in passive and active immunotherapy. Isolated recombinant prokaryotic HER2-ECD subdomains were previously found to be ineffective in inducing anti-tumor antibody response.
To employ recombinant eukaryotic HER2-ECD subdomains to raise anti-HER2 antibodies and determine their anti-tumor activity in vitro.
Two paired subdomains of HER2-ECD (DI+II and DIII+IV), representing Pertuzumab and Trastuzumab binding domains, respectively, along with the full extracellular domain of HER2 were generated in CHO-K1 cells. Polyclonal antibodies were raised against these subdomains and characterized using ELISA, flow cytometry, and immunoblot and their anti-tumor activity was assessed by XTT assay. The cross-reactivity of these antibodies was specified along with other members of the human HER family.
Similar to Trastuzumab and anti-HER2-ECD antibody, anti-DI+II and DIII+IV polyclonal antibodies reacted with recombinant HER2-ECD and native HER2 expressed on tumor cells. These two polyclonal antibodies were able to inhibit the binding of Pertuzumab and Trastuzumab to HER2, respectively, and did not cross-react with other members of HER family. These antibodies were able to inhibit tumor cell growth in vitro, similar to Trastuzumab.
The high immunogenicity of human HER2 DI+II and DIII+IV subdomains in rabbits and the tumor inhibitory activity of the purified specific antibodies imply that they might be suitable for active immunotherapy in formulation with appropriate adjuvants and in combination with other HER2 specific therapeutics.
人表皮生长因子受体2（HER2）在多种恶性肿瘤中起关键作用。HER2的细胞外结构域（HER2-ECD）已被广泛用作被动和主动免疫治疗的重要靶点。先前发现分离的重组原核HER2-ECD亚结构域在诱导抗肿瘤抗体反应方面无效。
利用重组真核HER2-ECD亚结构域产生抗HER2抗体，并在体外确定其抗肿瘤活性。
在CHO-K1细胞中产生分别代表帕妥珠单抗和曲妥珠单抗结合结构域的两对HER2-ECD亚结构域（DI+II和DIII+IV），以及HER2的完整细胞外结构域。针对这些亚结构域产生多克隆抗体，并通过酶联免疫吸附测定、流式细胞术和免疫印迹进行表征，通过XTT试验评估其抗肿瘤活性。确定这些抗体与人类HER家族其他成员的交叉反应性。
与曲妥珠单抗和抗HER2-ECD抗体类似，抗DI+II和DIII+IV多克隆抗体与重组HER2-ECD和肿瘤细胞上表达的天然HER2反应。这两种多克隆抗体分别能够抑制帕妥珠单抗和曲妥珠单抗与HER2的结合，并且不与HER家族的其他成员发生交叉反应。这些抗体能够在体外抑制肿瘤细胞生长，类似于曲妥珠单抗。
人HER2 DI+II和DIII+IV亚结构域在兔体内具有高免疫原性，纯化的特异性抗体具有肿瘤抑制活性，这表明它们可能适合与适当佐剂配制并与其他HER2特异性治疗药物联合用于主动免疫治疗。
