Bispecific antibody (BsAb) has become an important trend in developing next generation biologics therapies. By simultaneously engaging two molecular targets, BsAbs show distinctive mechanism of actions that could lead to clinical benefits unattainable by conventional monoclonal antibodies (mAbs). Successful launch provided clinical validation and encourage more BsAb development in the pipeline of pharmaceutical companies. Fabs-in-tandem immunoglobulin (FIT-Ig™) format was initially described in 2017. This unique design provides a symmetrical and tetravalent IgG-like bispecific molecule with correct association of 2 sets of VH/VL pairs, where two Fabs are fused directly in a crisscross orientation without any mutations or use of peptide linkers. FIT-Ig can be readily made from 2 existing monoclonal antibodies by basic molecular biology techniques with high expression level in mammalian cells, and easily purified to homogeneity using standard approaches without extensive optimization. FIT-Ig molecules exhibit favorable drug-like properties, in vitro and in vivo functions, as well as manufacturing efficiency for commercial development. Here, we provide an example of construction and preliminary characterization of a FIT-Ig molecule with discussions on optimization and general utility.

中文翻译：

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用于双特异性靶向的 Fabs-in-Tandem 免疫球蛋白分子的生成

双特异性抗体（BsAb）已成为开发下一代生物疗法的重要趋势。通过同时结合两个分子靶标，BsAb 显示出独特的作用机制，可以带来传统单克隆抗体 (mAb) 无法获得的临床益处。成功推出提供了临床验证并鼓励制药公司在管道中开发更多 BsAb。Fabs-in-tandem 免疫球蛋白 (FIT-Ig™) 形式最初于 2017 年描述。这种独特的设计提供了对称的四价 IgG 样双特异性分子，正确结合了 2 组 VH/VL 对，其中两个 Fabs 直接融合以纵横交错的方向，没有任何突变或使用肽接头。FIT-Ig 可以很容易地通过基础分子生物学技术从 2 种现有的单克隆抗体中制备，在哺乳动物细胞中具有高表达水平，并且无需大量优化即可使用标准方法轻松纯化至均质。FIT-Ig 分子表现出良好的类药物特性、体外和体内功能，以及商业开发的生产效率。在这里，我们提供了一个 FIT-Ig 分子的构建和初步表征的例子，并讨论了优化和一般效用。