As recent decades have seen extensive resources dedicated to exploring tumor metastasis, discoveries have exposed a need for understanding the intricacies of the tumor microenvironment (TME) in an ex vivo model. Much of the heterogeneity of the disease stems from the complexity of interactions between the cells that comprise the TME, placing its investigation at the heart of developing effective treatment. A number of findings implicate chemokines and their receptors, expressed early in cancer progression, in promoting tumor metastasis. These findings identify several cell types significantly involved in secreting and responding to chemokines, such as tumor cells, fibroblasts, and endothelial cells, among others. Chemokine expression depends not only on the cell types of the TME but also on the conditions in the surrounding matrix, especially interstitial flow. A strong link has been identified between interstitial flow and tumor metastasis. This flow, a result of rapid lymphangiogenesis, correlates with the expression of a number of chemokines responsible for tumor invasion and metastasis. Current limitations for studying the human TME have stalled progress since mouse models cannot fully engage all the relevant human cell types. Thus, the emergence of complex intercellular relationships within the TME generates a need for heterotypic three-dimensional in vitro models to cover the gaps between animal models and human studies. The need for 3D in vitro models is amplified by the exposed importance of interstitial flow. To mimic and examine the breast cancer tumor, we have designed a bioreactor, machined through femtosecond laser-etching technology that allows for co-culture of the numerous cell types that comprise the TME. The design incorporates a semicircular chamber connected to a channel 200 μm in width by a thin porous filter of laser-etched glass. The device is coated with poly-L-lysine and Collagen I to improve cell adhesion. To recreate the TME, the semicircular chamber is loaded with cancer-associated fibroblasts and tumor cells (MCF-7 and MDA-MB-231 cell lines) in a ratio of 3:1 in 3D reconstituted basement membrane with 1.5 mg/mL Collagen I (rat-tail) and 10% Matrigel. This 3D cell culture is grown for 5 days to allow spheroid formation. Human microvascular endothelial cells are cultured to form a confluent monolayer in the channel that is then perfused with medium, of which a higher quantity is aspirated from the outlet to generate interstitial flow through the 3D cell culture. Cancer-associated leukocytes can be infused through the channel to better mimic the TME. This unique and innovative design will allow in vitro visualization of cell migration and expression, which can shed light on the mechanisms of metastasis. Experiments can then be performed to elucidate the specific relationships linking interstitial flow and metastasis.
Citation Format: Kathryn Hockemeyer, Tammy Sobolik, Alexander Terekhov, Melody Swartz, William Hofmeister, John Wikswo, Chris Janetopoulos, Ann Richmond. A three-dimensional in vitro bioreactor for analysis of the tumor microenvironment: investigation of interstitial flow and the mechanism of metastasis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2614. doi:10.1158/1538-7445.AM2013-2614