Automated Assessment of Cancer Drug Efficacy On Breast Tumor Spheroids in Aggrewell™400 Plates Using Image Cytometry

Tumor spheroid models have proven useful in the study of cancer cell responses to chemotherapeutic compounds by more closely mimicking the 3-dimensional nature of tumors in situ. Their advantages are often offset, however, by protocols that are long, complicated, and expensive. Efforts continue for the development of high-throughput assays that combine the advantages of 3D models with the convenience and simplicity of traditional 2D monolayer methods. Herein, we describe the development of a breast cancer spheroid image cytometry assay using T47D cells in Aggrewell (TM) 400 spheroid plates. Using the Celigo(R) automated imaging system, we developed a method to image and individually track thousands of spheroids within the Aggrewell (TM) 400 microwell plate over time. We demonstrate the use of calcein AM and propidium iodide staining to study the effects of known anti-cancer drugs Doxorubicin, Everolimus, Gemcitabine, Metformin, Paclitaxel and Tamoxifen. We use the image cytometry results to quantify the fluorescence of calcein AM and PI as well as spheroid size in a dose dependent manner for each of the drugs. We observe a dose-dependent reduction in spheroid size and find that it correlates well with the viability obtained from the CellTiter96 (R) endpoint assay. The image cytometry method we demonstrate is a convenient and high-throughput drug-response assay for breast cancer spheroids under 400 mu m in diameter, and may lay a foundation for investigating other three-dimensional spheroids, organoids, and tissue samples.

Automated summary

Tumor spheroid models are important for studying cancer cell responses to chemotherapy. This study developed a breast cancer spheroid image cytometry assay that allows for convenient and high-throughput drug-response testing. The assay uses an automated imaging system to track and analyze thousands of spheroids and employs fluorescent staining to study the effects of anti-cancer drugs. The results show that spheroid size reduction correlates with cell viability, and the method can serve as a foundation for studying other three-dimensional spheroids and tissue samples.