Department of Chemistry, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, Fujian 361005, P. R. China.
Fujian Provincial Key Laboratory of Innovative Drug Target Research, School of Pharmaceutical Sciences, Xiamen University, Xiamen, Fujian 361102, P. R. China.
J Mater Chem B. 2020 Nov 11;8(43):9971-9979. doi: 10.1039/d0tb01091j.
Circulating tumor cell (CTC) detection and enumeration have been considered as a noninvasive biopsy method for the diagnosis, characterization, and monitoring of various types of cancers. However, CTCs are exceptionally rare, which makes CTC detection technologically challenging. In the past few decades, much effort has been focused on highly efficient CTC capture, while the activity of CTCs has often been ignored. Here, we develop an effective method for nondestructive CTC capture, release, and detection. Folic acid (FA), as a targeting molecule, is conjugated on magnetic nanospheres through a cleavable disulfide bond-containing linker (cystamine) and a polyethylene glycol (PEG2k) linker, forming MN@Cys@PEG2k-FA nanoprobes, which can bind with folate receptor (FR) positive CTCs specifically and efficiently, leading to the capture of CTCs with an external magnetic field. When approximately 150 and 10 model CTCs were spiked in 1 mL of lysis blood, 93.1 ± 2.9% and 80.0 ± 9.7% CTCs were recovered, respectively. In total, 81.3 ± 2.6% captured CTCs can be released from MN@Cys@PEG2k-FA magnetic nanospheres by treatment with dithiothreitol. The released CTCs are easily identified from blood cells for specific detection and enumeration combined with immunofluorescence staining with a limit of detection of 10 CTC mL-1 lysed blood. Moreover, the released cells remain healthy with high viability (98.6 ± 0.78%) and can be cultured in vitro without detectable changes in morphology or behavior compared with healthy untreated cells. The high viability of the released CTCs may provide the possibility for downstream proteomics research of CTCs; therefore, cultured CTCs were collected for proteomics. As a result, 3504 proteins were identified. In conclusion, the MN@Cys@PEG2k-FA magnetic nanospheres prepared in this study may be a promising tool for early-stage cancer diagnosis and provide the possibility for downstream analysis of CTCs.
循环肿瘤细胞（CTC）的检测和计数已被认为是一种用于诊断、特征描述和监测各种癌症的非侵入性活检方法。然而，CTC 极为罕见，这使得 CTC 的检测技术极具挑战性。在过去的几十年中，人们致力于高效的 CTC 捕获，而往往忽略了 CTC 的活性。在这里，我们开发了一种用于非破坏性 CTC 捕获、释放和检测的有效方法。叶酸（FA）作为靶向分子，通过含有可裂解二硫键的连接子（半胱胺）和聚乙二醇（PEG2k）连接子连接到磁性纳米球上，形成 MN@Cys@PEG2k-FA 纳米探针，可与叶酸受体（FR）阳性 CTC 特异性和高效结合，从而在外磁场下捕获 CTC。当在 1 毫升裂解血中分别加入约 150 和 10 个模型 CTC 时，分别回收了 93.1±2.9%和 80.0±9.7%的 CTC。总的来说，通过用二硫苏糖醇处理，可以从 MN@Cys@PEG2k-FA 磁性纳米球上释放 81.3±2.6%的捕获 CTC。释放的 CTC 很容易从血细胞中识别出来，用于特定的检测和计数，结合免疫荧光染色，检测限为 10 CTC mL-1 裂解血。此外，与未经处理的健康细胞相比，释放的细胞保持高活力（98.6±0.78%），并且可以在体外培养，形态或行为没有可检测的变化。释放的 CTC 具有高活力，可能为 CTC 的下游蛋白质组学研究提供了可能性；因此，收集培养的 CTC 进行蛋白质组学研究。结果鉴定了 3504 种蛋白质。总之，本研究制备的 MN@Cys@PEG2k-FA 磁性纳米球可能是一种用于早期癌症诊断的有前途的工具，并为 CTC 的下游分析提供了可能性。
