Serum-stable RNA aptamers to urokinase-type plasminogen activator blocking receptor binding
Serum-stable RNA aptamers to urokinase-type plasminogen activator blocking receptor binding
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DOI：
10.1261/rna.2338210
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发表时间：
2010-12-01
期刊：
影响因子：
4.5
通讯作者：
Andreasen, Peter Andre
中科院分区：
文献类型：
--
作者：
Dupont, Daniel Miotto;Madsen, Jeppe Buur;Andreasen, Peter Andre
丝氨酸蛋白酶尿激酶型纤溶酶原激活物(UPA)被广泛认为是抗癌治疗的潜在靶点。它通过uPA受体(UPAR)与细胞表面的结合是其功能的核心，在肿瘤的侵袭和转移中起着重要作用。在目前的研究中，我们使用指数富集法系统进化配体(SELEX)来筛选血清稳定的2‘-氟嘧啶修饰的RNA适配子，该适配子专门针对人uPA并在低纳摩尔浓度下阻断与其受体的相互作用。与适体的抑制功能一致，结合被发现依赖于uPA生长因子结构域的存在，该结构域介导uPAR结合。对最有效的uPA适配子之一uPAAP-12进行了更详细的分析，并且可以在不严重丧失其抑制活性的情况下显著减小其大小。最后，在细胞培养实验中，我们证明了适体的uPA清除作用可以减少uPA-PAI-1复合体对uPA的依赖内吞作用和细胞表面相关的纤溶酶原激活。UPA清除2‘-氟嘧啶修饰的RNA适配子代表了干扰uPA系统病理功能的一种新的有前途的原理。
The serine proteinase urokinase-type plasminogen activator (uPA) is widely recognized as a potential target for anticancer therapy. Its association with cell surfaces through the uPA receptor (uPAR) is central to its function and plays an important role in cancer invasion and metastasis. In the current study, we used systematic evolution of ligands by exponential enrichment (SELEX) to select serum-stable 2'-fluoro-pyrimidine-modified RNA aptamers specifically targeting human uPA and blocking the interaction to its receptor at low nanomolar concentrations. In agreement with the inhibitory function of the aptamers, binding was found to be dependent on the presence of the growth factor domain of uPA, which mediates uPAR binding. One of the most potent uPA aptamers, upanap-12, was analyzed in more detail and could be reduced significantly in size without severe loss of its inhibitory activity. Finally, we show that the uPA-scavenging effect of the aptamers can reduce uPAR-dependent endocytosis of the uPA-PAI-1 complex and cell-surface associated plasminogen activation in cell culture experiments. uPA-scavenging 2'-fluoro-pyrimidine-modified RNA aptamers represent a novel promising principle for interfering with the pathological functions of the uPA system.