Department of Materials Science and Engineering, University of Michigan, Ann Arbor, MI, USA.
Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA.
Neoplasia. 2019 Aug;21(8):822-836. doi: 10.1016/j.neo.2019.06.005. Epub 2019 Jul 9.
Intraperitoneal dissemination of ovarian cancers is preceded by the development of chemoresistant tumors with malignant ascites. Despite the high levels of chemoresistance and relapse observed in ovarian cancers, there are no in vitro models to understand the development of chemoresistance in situ.
We describe a highly integrated approach to establish an in vitro model of chemoresistance and stemness in ovarian cancer, using the 3D hanging drop spheroid platform. The model was established by serially passaging non-adherent spheroids. At each passage, the effectiveness of the model was evaluated via measures of proliferation, response to treatment with cisplatin and a novel ALDH1A inhibitor. Concomitantly, the expression and tumor initiating capacity of cancer stem-like cells (CSCs) was analyzed. RNA-seq was used to establish gene signatures associated with the evolution of tumorigenicity, and chemoresistance. Lastly, a mathematical model was developed to predict the emergence of CSCs during serial passaging of ovarian cancer spheroids.
Our serial passage model demonstrated increased cellular proliferation, enriched CSCs, and emergence of a platinum resistant phenotype. In vivo tumor xenograft assays indicated that later passage spheroids were significantly more tumorigenic with higher CSCs, compared to early passage spheroids. RNA-seq revealed several gene signatures supporting the emergence of CSCs, chemoresistance, and malignant phenotypes, with links to poor clinical prognosis. Our mathematical model predicted the emergence of CSC populations within serially passaged spheroids, concurring with experimentally observed data.
Our integrated approach illustrates the utility of the serial passage spheroid model for examining the emergence and development of chemoresistance in ovarian cancer in a controllable and reproducible format.
卵巢癌的腹腔扩散是在形成具有恶性腹水的耐药肿瘤后发生的。尽管卵巢癌观察到高水平的耐药性和复发，但没有体外模型来了解原位耐药性的发展。
我们描述了一种高度综合的方法，使用 3D 悬滴球体平台来建立卵巢癌的耐药性和干性体外模型。该模型通过连续传代非贴壁球体来建立。在每一轮中，通过增殖、顺铂治疗效果和新型 ALDH1A 抑制剂的反应来评估模型的有效性。同时，分析了癌症干细胞样细胞（CSC）的表达和肿瘤起始能力。RNA-seq 用于建立与肿瘤发生和耐药性演变相关的基因特征。最后，开发了一个数学模型来预测卵巢癌球体连续传代过程中 CSC 的出现。
我们的连续传代模型显示出细胞增殖增加、CSC 富集和出现铂耐药表型。体内肿瘤异种移植实验表明，与早期传代球体相比，晚期传代球体具有更高的肿瘤发生能力和更高的 CSCs。RNA-seq 揭示了几个支持 CSC 出现、耐药性和恶性表型的基因特征，与不良临床预后有关。我们的数学模型预测了连续传代球体中 CSC 群体的出现，与实验观察到的数据一致。
我们的综合方法说明了连续传代球体模型在可控和可重复的格式下检查卵巢癌耐药性的出现和发展的实用性。
