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A cross-linked anti-TNF-α aptamer for neutralization of TNF-α-induced cutaneous Shwartzman phenomenon: A simple and novel approach for improving aptamers' affinity and efficiency
Biotechnology Progress ( IF 2.9 ) Pub Date : 2021-07-04 , DOI: 10.1002/btpr.3191
Kazem Mashayekhi 1, 2 , Mojtaba Sankian 3 , Saeed Mohammadian Haftcheshmeh 4 , Ramezan Ali Taheri 5 , Kazem Hassanpour 6 , Gholamreza Farnoosh 7
Affiliation  

To increase the efficiency of aptamers to their targets, a simple and novel method has been developed based on aptamer oligomerization. To this purpose, previously anti-human TNF-α aptamer named T1–T4 was trimerized through a trimethyl aconitate core for neutralization of in vitro and in vivo of TNF-α. At first, 54 mer T1–T4 aptamers with 5′-NH2 groups were covalently coupled to three ester residues in the trimethyl aconitate. In vitro activity of novel anti-TNF-α aptamer and its dissociation constant (Kd) was done using the L929 cell cytotoxicity assay. In vivo anti-TNF-α activity of new oligomerized aptamer was assessed in a mouse model of cutaneous Shwartzman. Anchoring of three T1–T4 aptamers to trimethyl aconitate substituent results in formation of the 162 mer fragment, which was well revealed by gel electrophoresis. In vitro study indicated that the trimerization of T1–T4 aptamer significantly improved its anti-TNF-α activity compared to non-modified aptamers (P < 0.0001) from 40% to 60%. The determination of Kd showed that trimerization could effectively enhance Kd of aptamer from 67 nM to 36 nM. In vivo study showed that trimer aptamer markedly reduced mean scar size from 15.2 ± 1.2 mm to 1.6 ± 0.1 mm (P < 0.0001), which prevent the formation of skin lesions. In vitro and in vivo studies indicate that trimerization of anti-TNF-α aptamer with a novel approach could improve the anti-TNF-α activity and therapeutic efficacy. According to our findings, a new anti-TNF-α aptamer described here could be considered an appropriate therapeutic agent in treating several inflammatory diseases.

中文翻译:

用于中和 TNF-α 诱导的皮肤 Shwartzman 现象的交联抗 TNF-α 适体:一种提高适体亲和力和效率的简单而新颖的方法

为了提高适体对其靶标的效率,已经开发了一种基于适体寡聚化的简单而新颖的方法。为此,以前名为 T1-T4 的抗人 TNF-α 适体通过乌头酸三甲酯核心进行三聚化,以在体外体内中和 TNF-α。起初,具有 5'-NH 2基团的 54 mer T1-T4 适体与乌头酸三甲酯中的三个酯残基共价偶联。使用 L929 细胞毒性试验进行新型抗 TNF-α 适体的体外活性及其解离常数 ( Kd )。体内在皮肤 Shwartzman 的小鼠模型中评估了新的寡聚适体的抗 TNF-α 活性。三个 T1-T4 适体与乌头酸三甲酯取代基的锚定导致形成 162 mer 片段,凝胶电泳很好地揭示了这一点。体外研究表明,与未修饰的适体相比,T1-T4 适体的三聚化显着提高了其抗 TNF-α 活性(P  < 0.0001),从 40% 提高到 60%。K d的测定表明,三聚化可以有效地将适体的K d从67 nM提高到36 nM。体内研究表明,三聚体适体显着减少平均疤痕大小从 15.2 ± 1.2 mm 到 1.6 ± 0.1 mm ( P < 0.0001),防止皮肤损伤的形成。体外体内研究表明,用一种新方法将抗 TNF-α 适体三聚化可以提高抗 TNF-α 活性和治疗效果。根据我们的研究结果,这里描述的一种新的抗 TNF-α 适体可以被认为是治疗几种炎症性疾病的合适治疗剂。
更新日期:2021-07-04
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