A Novel High-Throughput, High-Content Three-Dimensional Assay for Determination of Tumor Invasion and Dormancy

Metastasis accounts for most cancer deaths, while dormancy of tumor cells leads to unexpected cancer recurrence. These two aspects of cancer remain relatively untreatable in part because current two-dimensional (2D) methods of high-throughput drug screening cannot quantify outcomes related to these phenotypes. Three-dimensional (3D) in-vitro tumor models are a promising alternative because they better recreate the tumor microenvironment and relevant phenotypes. However, outcome measures for high-throughput screening of these systems are often limited to single measures such as metabolic activity using assays that are not standardized or optimized for 3D models. To address this gap, the objective of this work to develop an image-based assay to measure tumor cell health, proliferation, invasion, and dormancy for high-throughput drug screening. Drug dosing experiments were performed using a novel 3D pancreatic tumor metastasis model, followed by application of various dye combinations to assess viability, proliferation, metabolic activity, and invasion. We successfully demonstrated that high throughput imaging and analysis can be performed to quantify proliferation, metabolic activity, and invasion in a single multi-output assay. Proof-of-concept experiments also revealed that while gemcitabine does effectively inhibit cancer cell proliferation it does not kill all cells and may contribute to tumor dormancy. Overall, this work using a novel 3D tumor metastasis model coupled with a multi-output assay serves as a first step toward a drug screening platform that will enable researchers to better correlate in-vitro model results with clinical outcomes related to metastasis and dormancy