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Quantification of MMP Activity and Inhibition in a 3D Tumor Invasion Model


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September 29, 2015

Authors: Brad Larson, Leonie Rieger, BioTek Instruments, Inc. Winooski, VT; Crystal Falco, Diana Hulboy, Enzium, Inc. Philadelphia, PA


Metastasis, the spread of cancer cells from the primary tumor to secondary locations within the body, is linked to approximately 90% of cancer deaths. Penetration through the basement membrane is a critical step during the metastatic process, and has been linked to the formation of membrane superstructures called invadopodia. While being made of multiple substances, a notable component are matrix metalloproteinases (MMPs), and specifically MMP-14. Mounting preclinical evidence linking MMPs to cancer progression, combined with the issue of overlapping substrate specificity of MMP family members, has made the development of targeted MMP inhibitors an attractive approach to cancer therapy. Therefore, methods to selectively measure MMP-14 activity, specifically within invadopodia, in a sensitive yet easy to perform process, are necessary.

Similarly, appropriate in vitro cell models have been unable to accurately assess the ability of novel therapies to inhibit tumor invasion, including invadopodia formation. Incorporating a three-dimensional (3D) spheroid-type cellular structure that includes co-cultured cell types forming a tumoroid, provides a more predictive model than the use of individual cancer cells cultured on the bottom of a well in traditional two-dimensional (2D) format.

Here we demonstrate the ability to image and quantify MMP-14 activity, in addition to tumor invasion, using a 3D tumoroid cell model. Enzium’s protease activity detection technology was incorporated into the invasion assay procedure to enable simultaneous phenotypic and mechanism of action quantification. The tumoroids comprised primary human dermal fibroblasts and MDA-MB-231 breast adenocarcinoma cells, known to be invasive and metastasize to lung from primary mammary fat pad tumors. The cells were aggregated into 3D structures using Corning Spheroid Microplates containing an Ultra Low Attachment surface. A novel cell imaging multimode reader and cellular analysis algorithms from BioTek Instruments, Inc. were incorporated to provide automated, image-based detection and quantification of invasion and enzyme activity. The combination presents an accurate, easy-to-use method.
Cytation 5