Department of Biochemistry, University of Zürich, 8057 Zürich, Switzerland.
Structure. 2013 Nov 5;21(11):1979-91. doi: 10.1016/j.str.2013.08.020. Epub 2013 Oct 3.
Human epidermal growth factor receptor-2 (HER2) is a receptor tyrosine kinase directly linked to the growth of malignancies from various origins and a validated target for monoclonal antibodies and kinase inhibitors. Utilizing a new approach with designed ankyrin repeat proteins (DARPins) as alternative binders, we show that binding of two DARPins connected by a short linker, one targeting extracellular subdomain I and the other subdomain IV, causes much stronger cytotoxic effects on the HER2-addicted breast cancer cell line BT474, surpassing the therapeutic antibody trastuzumab. We determined crystal structures of these DARPins in complex with the respective subdomains. Detailed models of the full-length receptor, constrained by its rigid domain structures and its membrane anchoring, explain how the bispecific DARPins connect two membrane-bound HER2 molecules, distorting them such that they cannot form signaling-competent dimers with any EGFR family member, preventing any kinase dimerization, and thus leading to a complete loss of signaling.
人表皮生长因子受体 2（HER2）是一种受体酪氨酸激酶，与多种来源的恶性肿瘤的生长直接相关，也是单克隆抗体和激酶抑制剂的有效靶点。我们利用一种新的方法，设计了锚蛋白重复蛋白（DARPins）作为替代结合物，结果表明，通过短接头连接的两个 DARPins 的结合，一个针对细胞外亚结构域 I，另一个针对亚结构域 IV，对依赖 HER2 的乳腺癌细胞系 BT474 产生更强的细胞毒性作用，超过了治疗性抗体曲妥珠单抗。我们确定了这些 DARPins 与相应亚结构域复合物的晶体结构。受其刚性结构域和膜锚定限制的全长受体的详细模型解释了双特异性 DARPins 如何连接两个膜结合的 HER2 分子，使它们扭曲，以至于它们不能与任何 EGFR 家族成员形成信号功能二聚体，阻止任何激酶二聚化，从而导致信号完全丧失。
