Institute of Functional Nano & Soft Materials (FUNSOM), Jiangsu Key Laboratory for Carbon-Based Functional Materials & Devices, Joint International Research Laboratory of Carbon-Based Functional Materials and Devices, Soochow University, Suzhou, 215123, P. R. China.
Department of thoracic Surgery, The Second Affiliated Hospital of Soochow University, Suzhou, 215123, P. R. China.
Small Methods. 2023 Jul;7(7):e2300096. doi: 10.1002/smtd.202300096. Epub 2023 Apr 22.
The phenotypic changes of circulating tumor cells (CTCs) during the epithelial-mesenchymal transition (EMT) have been a hot topic in tumor biology and cancer therapeutic development. Here, an integrated platform of single-cell fluorescent enzymatic assays with superwetting droplet-array microchips (SDAM) for ultrasensitive functional screening of epithelial-mesenchymal sub-phenotypes of CTCs is reported. The SDAM can generate high-density, volume well-defined droplet (0.66 nL per droplet) arrays isolating single tumor cells via a discontinuous dewetting effect. It enables sensitive detection of MMP9 enzyme activities secreted by single tumor cells, correlating to their epithelial-mesenchymal sub-phenotypes. In the pilot clinical double-blind tests, the authors have demonstrated that SDAM assays allow for rapid identification and functional screening of CTCs with different epithelial-mesenchymal properties. The consistency with the clinical outcomes validates the usefulness of single-cell secreted MMP9 as a biomarker for selective CTC screening and tumor metastasis monitoring. Convenient addressing and recovery of individual CTCs from SDAM have been demonstrated for gene mutation sequencing, immunostaining, and transcriptome analysis, revealing new understandings of the signaling pathways between MMP9 secretion and the EMT regulation of CTCs. The SDAM approach combined with sequencing technologies promises to explore the dynamic EMT plasticity of tumors at the single-cell level.
循环肿瘤细胞 (CTC) 在上皮-间质转化 (EMT) 过程中的表型变化一直是肿瘤生物学和癌症治疗发展的热门话题。在这里，我们报道了一种集成的单细胞荧光酶分析与超润湿液滴阵列微芯片 (SDAM) 的平台，用于上皮-间质亚表型的 CTC 进行超灵敏功能筛选。SDAM 可以通过不连续的去湿效应生成高密度、体积确定的液滴 (每个液滴 0.66 nL) 阵列，从而分离单个肿瘤细胞。它能够敏感地检测单个肿瘤细胞分泌的 MMP9 酶活性，与它们的上皮-间质亚表型相关。在初步的临床双盲测试中，作者证明了 SDAM 检测可以快速识别和筛选具有不同上皮-间质特性的 CTC，并具有功能。单细胞分泌的 MMP9 作为一种用于选择性 CTC 筛选和肿瘤转移监测的生物标志物的实用性，与临床结果一致。已经证明了从 SDAM 方便地寻址和回收单个 CTC 进行基因突变测序、免疫染色和转录组分析，揭示了 MMP9 分泌与 EMT 对 CTC 调控之间信号通路的新认识。SDAM 方法与测序技术相结合，有望在单细胞水平上探索肿瘤 EMT 可塑性的动态变化。
