针对孤儿受体 Tie1 的纳米抗体体外抑制癌症血管生成和迁移。

IF 0.1 3区 艺术学 0 MUSIC MUSICAL TIMES Pub Date : 2022-05-23 DOI:10.1007/s00018-022-04336-9
May Meltzer, Noam Eliash, Ziv Azoulay, Uzi Hadad, Niv Papo
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引用次数: 0

摘要

人类信号分子 Tie1 和 Tie2 受体酪氨酸激酶(RTKs)在包括各种癌症在内的许多疾病中发挥着重要的病理生理作用。Tie1 的活性主要通过下游血管生成素-1(Ang1)依赖性激活 Tie2 来介导,因此 Tie 1 和 Tie1/Tie2/Ang1 轴都是有吸引力的治疗干预靶点。然而,开发以 Tie1 为靶点的抑制剂以及了解它们对 Tie2 和 Tie1/Tie2/Ang1 轴的影响仍是一项尚未完成的任务,这主要是因为 Tie1 是一种孤儿受体,很难按免疫抗体库筛选所需的数量生产和使用。在寻找这种孤儿受体的选择性抑制剂时,我们试图利用非免疫纳米抗体的优势(如体积小,可与隐藏的表位结合),同时克服它们的局限性(即表达量低和稳定性差)。因此,我们对酵母表面展示的针对 Tie1 细胞外结构域的原始和预先设计的合成(非免疫)纳米抗体库进行了表达、稳定性和亲和力筛选。筛选出的纳米抗体具有高表达、对 Tie1 具有良好的亲和力和特异性,因此与 Tie1 的结合优于与 Tie2 的结合。通过体外和基于细胞的试验,对这种合成纳米抗体的稳定性、选择性、效力和治疗潜力进行了分析。该纳米抗体可引发 Tie1 依赖性抑制 RTK(Tie2、Akt 和 Fak)磷酸化和血管内皮细胞的血管生成,并抑制人类胶质母细胞瘤细胞的活力和迁移。这项研究为开发纳米抗体作为治疗与 Tie1 激活相关的不同癌症的药物开辟了道路。
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In vitro inhibition of cancer angiogenesis and migration by a nanobody that targets the orphan receptor Tie1.

The human signaling molecules Tie1 and Tie2 receptor tyrosine kinases (RTKs) play important pathophysiological roles in many diseases, including different cancers. The activity of Tie1 is mediated mainly through the downstream angiopoietin-1 (Ang1)-dependent activation of Tie2, rendering both Tie 1 and the Tie1/Tie2/Ang1 axis attractive putative targets for therapeutic intervention. However, the development of inhibitors that target Tie1 and an understanding of their effect on Tie2 and on the Tie1/Tie2/Ang1 axis remain unfulfilled tasks, due, largely, to the facts that Tie1 is an orphan receptor and is difficult to produce and use in the quantities required for immune antibody library screens. In a search for a selective inhibitor of this orphan receptor, we sought to exploit the advantages (e.g., small size that allows binding to hidden epitopes) of non-immune nanobodies and to simultaneously overcome their limitations (i.e., low expression and stability). We thus performed expression, stability, and affinity screens of yeast-surface-displayed naïve and predesigned synthetic (non-immune) nanobody libraries against the Tie1 extracellular domain. The screens yielded a nanobody with high expression and good affinity and specificity for Tie1, thereby yielding preferential binding for Tie1 over Tie2. The stability, selectivity, potency, and therapeutic potential of this synthetic nanobody were profiled using in vitro and cell-based assays. The nanobody triggered Tie1-dependent inhibition of RTK (Tie2, Akt, and Fak) phosphorylation and angiogenesis in endothelial cells, as well as suppression of human glioblastoma cell viability and migration. This study opens the way to developing nanobodies as therapeutics for different cancers associated with Tie1 activation.

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MUSICAL TIMES
MUSICAL TIMES MUSIC-
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