Novel Single Domain Antibodies with Multivalency and Multispecificity

描述（由申请人提供）：ErbB/HER受体在传播调节细胞增殖、分化、运动性和凋亡的信号中发挥重要作用。值得注意的是，ErbB/HER家族受体酪氨酸激酶及其配体的异常调节在许多人类肿瘤中是常见的。事实上，EGFR、HER 2和HER 3已经在诊断和治疗上被验证为多种人类恶性肿瘤如肺癌、结肠直肠癌、乳腺癌、头颈癌、胃癌和胰腺癌的靶标。因此，全面靶向ErbB/HER家族成员的新型蛋白质生物药物可能在不同类型癌症的靶向诊断和治疗中具有重要应用。 该项目解决了开发具有成本效益的靶向配体的关键需求，这些配体以所需的亲合力和多特异性识别并结合HER家族成员。该项目旨在使用一种称为mRNA展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类SDA结构域文库中进化出具有人类起源的单结构域抗体（SDA），这些抗体分别与EFRG，HER 2或HER 3的胞外结构域紧密特异性结合。所得单体SDA将用作产生异二聚体SDA的基础，所述异二聚体SDA在两个非重叠表位处以协同亲合力效应结合感兴趣的HER成员。此外，我们将开发一种有效的和通用的自组装系统，允许可逆和可控的HER-结合SDA的纳米粒子的负载，以获得所需的多特异性。所得SDA将具有许多优点，包括高靶标结合亲和力、易于实现所需亲合力、多价性和多特异性、由于人来源的最小免疫原性、用于更好的组织渗透的小尺寸，以及由于在细菌中的高表达水平而显著降低的制造成本。这些SDA的简单性和稳定性也极大地促进了它们与用于翻译应用的各种纳米颗粒的缀合。将在胰腺导管腺癌（PDAC）异种移植动物模型中通过靶向递送含有针对几种转移标记基因的siRNA的纳米颗粒来体内检查所得SDA，所述转移标记基因对于PDAC肿瘤发生和转移是重要的。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靶向生物药物。

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.