Duan Chengchen, Townley Helen Elizabeth
Nuffield Department of Women's and Reproductive Health, Oxford University John Radcliffe Hospital, Oxford OX3 9DU, UK.
Department of Engineering Science, Oxford University, Oxford OX1 3PJ, UK.
Bioengineering (Basel). 2022 Apr 15;9(4):174. doi: 10.3390/bioengineering9040174.
NELL1 (Neural epidermal growth factor-like (EGFL)-like protein) is an important biomarker associated with tissue and bone development and regeneration. NELL1 upregulation has been linked with metastasis and negative prognosis in rhabdomyosarcoma (RMS). Furthermore, multiple recent studies have also shown the importance of NELL1 in inflammatory bowel disease and membranous nephropathy, amongst other diseases. In this study, several anti-NELL1 DNA aptamers were selected from a randomized ssDNA pool using a fluorescence-guided method and evaluated for their binding affinity and selectivity. Several other methods such as a metabolic assay and confocal microscopy were also applied for the evaluation of the selected aptamers. The top three candidates were evaluated further, and AptNCan3 was shown to have a binding affinity up to 959.2 nM. Selectivity was examined in the RH30 RMS cells that overexpressed NELL1. Both AptNCan2 and AptNCan3 could significantly suppress metabolic activity in RMS cells. AptNCan3 was found to locate on the cell membrane and also on intracellular vesicles, which matched the location of NELL1 shown by antibodies in previous research. These results indicate that the selected anti-NELL1 aptamer showed strong and highly specific binding to NELL1 and therefore has potential to be used for in vitro or in vivo studies and treatments.
NELL1（神经表皮生长因子样（EGFL）样蛋白）是一种与组织和骨骼发育及再生相关的重要生物标志物。在横纹肌肉瘤（RMS）中，NELL1的上调与转移和不良预后有关。此外，最近的多项研究还表明NELL1在炎症性肠病和膜性肾病等多种疾病中具有重要作用。在本研究中，使用荧光引导方法从随机单链DNA库中筛选出几种抗NELL1 DNA适配体，并评估了它们的结合亲和力和选择性。还应用了代谢测定和共聚焦显微镜等其他几种方法来评估所选的适配体。对排名前三的候选者进行了进一步评估，结果显示AptNCan3的结合亲和力高达959.2 nM。在过表达NELL1的RH30 RMS细胞中检测了选择性。AptNCan2和AptNCan3均能显著抑制RMS细胞的代谢活性。发现AptNCan3定位于细胞膜以及细胞内囊泡上，这与先前研究中抗体所示的NELL1定位相匹配。这些结果表明，所选的抗NELL1适配体对NELL1表现出强且高度特异性的结合，因此有潜力用于体外或体内研究及治疗。
