Zhang Peng, Zhou Xin, Zeng Yong
Department of Chemistry , Ralph N Adams Institute for Bioanalytical Chemistry , University of Kansas , Lawrence , KS 66045 , USA . Email:
University of Kansas Cancer Center , Kansas City , KS 66160 , USA.
Chem Sci. 2019 Apr 22;10(21):5495-5504. doi: 10.1039/c9sc00961b. eCollection 2019 Jun 7.
Circulating exosomes have been studied as a promising biomarker for non-invasive cancer diagnosis, as they are implicated in tumor initiation, progression, and metastasis. However, the clinical significance of circulating exosomes has not been revealed thoroughly, due to the technical limitation in sensitive and multiplexed detection of cargoes on exosomes, such as proteins and nucleic acids. Herein we developed an integrated exosome profiling platform (ExoProfile chip) to afford superior sensitivity and multiplexed capability for quantitative detection of a panel of surface protein markers on exosomes. To achieve this goal, we innovatively constructed 3D porous serpentine nanostructures patterned colloidal self-assembly to provide enormous reaction sites and improve biosensing efficiency of exosomes. Meanwhile, the switchable microfluidic design enabled the simultaneous detection of eight markers on single addition of exosome samples. The ExoProfile chip was validated with purified exosomes from SKOV3 cells, which yielded a limit of detection of 21 exosomes per μL. We applied the ExoProfile chip to clinical analysis of circulating exosomes using only 10 μL ovarian cancer plasma and completing the analysis within 3 h. The diagnostic power of seven markers (EGFR, HER2, CA125, FRα, CD24, EpCAM, and CD9 plus CD63) were evaluated with receiver operator characteristic curve and heatmap clustering. Compared to single biomarker, the combined assessment of a biomarker panel was demonstrated to display improved accuracy in distinguishing early and late stage cancer. The results suggested the ExoProfile chip as a promising platform for molecular fingerprinting of circulating exosomes towards early cancer diagnosis.
循环外泌体已被作为一种有前景的非侵入性癌症诊断生物标志物进行研究，因为它们与肿瘤的发生、发展和转移有关。然而，由于对外泌体上货物（如蛋白质和核酸）进行灵敏和多重检测存在技术限制，循环外泌体的临床意义尚未得到充分揭示。在此，我们开发了一种集成外泌体分析平台（ExoProfile芯片），以提供卓越的灵敏度和多重检测能力，用于定量检测外泌体上的一组表面蛋白标志物。为实现这一目标，我们创新性地构建了图案化胶体自组装的三维多孔蛇形纳米结构，以提供大量反应位点并提高外泌体的生物传感效率。同时，可切换的微流控设计使得在单次添加外泌体样品时能够同时检测八种标志物。ExoProfile芯片用SKOV3细胞纯化的外泌体进行了验证，其检测限为每微升21个外泌体。我们仅使用10微升卵巢癌血浆将ExoProfile芯片应用于循环外泌体的临床分析，并在3小时内完成分析。通过受试者工作特征曲线和热图聚类评估了七种标志物（表皮生长因子受体、人表皮生长因子受体2、糖类抗原125、叶酸受体α、CD24、上皮细胞黏附分子以及CD9加CD63）的诊断能力。与单一生物标志物相比，一组生物标志物的联合评估在区分早期和晚期癌症方面显示出更高的准确性。结果表明ExoProfile芯片是用于循环外泌体分子指纹识别以实现早期癌症诊断的一个有前景的平台。
