Novel Single Domain Antibodies with Multivalency and Multispecificity

描述(申请人提供)：ErbB/Her受体在传播调节细胞增殖、分化、运动和凋亡的信号方面发挥重要作用。值得注意的是，ErbB/HER受体酪氨酸激酶及其配体家族的异常调节在许多人类肿瘤中是常见的。事实上，EGFR、HER2和HER3已被证实是多种人类恶性肿瘤的诊断和治疗靶点，如肺癌、结直肠癌、乳腺癌、头颈部、胃癌和胰腺癌。因此，全面靶向ErbB/Her家族成员的新型蛋白质生物药物在不同类型癌症的靶向诊断和治疗中具有重要的应用价值。该项目解决了开发具有成本效益的靶向配基的迫切需要，该配基能够识别并以所需的亲和力和多特异性结合她的家庭成员。该项目旨在利用一种名为mRNA显示的强大的体外蛋白质选择方法，从具有异常高多样性的人SDA结构域文库中进化出与EFRG、HER2或HER3胞外结构域紧密和特异结合的人类来源的单域抗体(SDA)。得到的单体SDA将被用作产生异二聚体SDA的基础，该SDA与HER感兴趣的成员在两个非重叠的表位上结合，具有协同亲和力效应。此外，我们将开发一种高效和通用的自组装系统，允许可逆和可控地将HER结合的SDA负载到纳米颗粒上，以获得所需的多重特异性。由此得到的SDA具有许多优点，包括高的靶标结合亲和力，易于获得所需的亲和力，多价和多特异性，由于人类起源而产生的最低免疫原性，较小的尺寸用于更好的组织渗透，此外，由于在细菌中的高表达水平，显著降低了制造成本。这些SDA的简单性和稳定性也极大地促进了它们与各种用于翻译应用的纳米颗粒的结合。由此产生的SDA将通过靶向输送含有针对几个转移特征基因的siRNA的纳米颗粒在胰腺导管腺癌(PDAC)异种移植动物模型中进行体内检测，这些基因对PDAC的肿瘤发生和转移具有重要作用。该项目的成功将导致开发用于癌症诊断和治疗的具有成本效益的、有针对性的生物制药。

DESCRIPTION (provided by applicant): ErbB/HER receptors play essential roles in propagating signals that regulate cell proliferation, differentiation, motility, and apoptosis. Significantly, aberrant regulation of the ErbB/HER family of receptor tyrosine kinases and their ligands is a common occurrence in many human tumors. Indeed, EGFR, HER2, and HER3 have been diagnostically and therapeutically validated as targets in a wide variety of human malignancies such as lung, colorectal, breast, head and neck, gastric and pancreatic cancers. Therefore, novel protein biopharmaceuticals that comprehensively target ErbB/HER family members could have significant applications in targeted diagnosis and therapy of different types of cancer. This project addresses a critical need for developing cost-effective targeting ligands that recognize and bind HER family members with desired avidity and multispecificity. The project is directed at using a powerful in vitro protein selection method called mRNA display to evolve single domain antibodies (SDAs) with human origin that tightly and specifically bind to the extracellular domain of EFRG, HER2 or HER3, respectively, from a human SDA domain library with an unusually high diversity. The resulting monomeric SDAs will be used as the basis for the generation of heterodimeric SDAs that bind to a HER member of interest at two nonoverlapping epitopes with synergistic avidity effect. In addition, we will develop an efficient and universal self-assembly system that allows for reversible and controllable loading of HER-binding SDAs to nanoparticles to acquire desired multispecificity. The resulting SDAs would have a number of advantages, including high target-binding affinity, ease to achieve desired avidity, multivalency and multispecificity, minimal immunogenicity due to human origin, small size for better tissue penetration, in addition to significantly reduced manufacturing costs due to high expression levels in bacteria. The simplicity and stability of these SDAs also greatly facilitate their conjugation with various nanoparticles for translational applications. The resulting SDAs will be examined in vivo in pancreatic ductal adenocarcinoma (PDAC) xenograft animal models by targeted delivery of nanoparticles containing siRNAs against several metastasis signature genes that are important for PDAC tumorigenesis and metastasis. The success of the project will lead to the development of cost-effective, targeted biopharmaceuticals for cancer diagnosis and therapy.