Molecular Biology Research Center, School of Life Sciences, Central South University, Changsha, 410078, China; Department of Biochemistry, College of Medicine, Jishou University, Jishou, 416000, China.
Molecular Biology Research Center, School of Life Sciences, Central South University, Changsha, 410078, China.
Biochimie. 2018 Aug;151:150-158. doi: 10.1016/j.biochi.2018.06.003. Epub 2018 Jun 15.
Multiple myeloma (MM) is a malignant plasma cell disease and is considered incurable. Annexin A2 (ANXA2) is closely related to the proliferation and adhesion of MM. Using protein-SELEX, we performed a screen for aptamers that bind GST-ANXA2 from a library, and GST protein was used for negative selection. The enrichment of the ssDNA pool was monitored by filter-binding assay during selection. After nine rounds of screening and high-throughput sequencing, we obtained six candidate aptamers that bind to the ANXA2 protein. The affinities of the candidate aptamers for ANXA2 were determined by ELONA. Binding of aptamer wh6 to the ANXA2 protein and to the MM cell was verified by aptamer pulldown experiment and flow cytometry, respectively. Aptamer wh6 binds the ANXA2 protein with good stability and has a dissociation constant in the nanomolar range. The binding specificity of aptamer wh6 was confirmed in vivo in nude mouse xenografts with MM cells and with MM bone marrow aspirates. Furthermore, aptamer wh6 can block MM cell adhesion to ANXA2 and block the proliferation of MM cells induced by ANXA2. In summary, wh6 can be considered a promising candidate tool for MM diagnosis and treatment.
多发性骨髓瘤（MM）是一种恶性浆细胞疾病，被认为是不可治愈的。膜联蛋白 A2（ANXA2）与 MM 的增殖和黏附密切相关。我们使用蛋白质 SELEX 从文库中筛选与 GST-ANXA2 结合的适体，并用 GST 蛋白进行阴性选择。在筛选过程中，通过滤膜结合实验监测 ssDNA 池的富集情况。经过九轮筛选和高通量测序，我们获得了六个与 ANXA2 蛋白结合的候选适体。通过 ELONA 测定候选适体对 ANXA2 的亲和力。通过适体下拉实验和流式细胞术分别验证了适体 wh6 与 ANXA2 蛋白和 MM 细胞的结合。适体 wh6 与 ANXA2 蛋白具有良好的稳定性结合，解离常数在纳摩尔范围内。在携带 MM 细胞的裸鼠异种移植和 MM 骨髓抽吸物中体内验证了适体 wh6 的结合特异性。此外，适体 wh6 可以阻断 MM 细胞与 ANXA2 的黏附，并阻断 ANXA2 诱导的 MM 细胞增殖。总之，wh6 可以被认为是 MM 诊断和治疗的有前途的候选工具。
