Understanding the Role of Adipocytes and the Tumor Microenvironment on Doxorubicin Therapy of Breast Cancer in a Tissue Engineered, Patient Specific, High-throughput Biomimetic Platform
Daniel Lara, BS1, Karel-Bart Celie, BA2, Yoshiko Toyoda, BA1, Matthew Wright, BA2, Arash Samadi, BS1, Justin Buro, BA3, Sofya Oshchepkova, BA1, Sophia Hameedi, BA2, Kristy Brown, PhD1, Jason Spector, MD1.
1Weill Cornell Medicine, New York, NY, USA, 2Columbia University Vagelos College of Physicians and Surgeons, New York, NY, USA, 3George Washington University School of Medicine and Health Sciences, Washington, DC, USA.
Introduction: Breast cancer (BC) research encompasses not only the study of tumor behavior, but also requires understanding the innumerable interactions that occur between breast cellular components and the extracellular matrix (ECM) in the development and progression of the disease. Despite extensive research, BC continues to be a leading cause of morbidity and mortality for women across the globe. The lack of three-dimensional models that are able to accurately replicate the tumor microenvironment and tumor-associated stroma has resulted in the unsuccessful translation of most preclinical studies into the clinic. It has been suggested that adipose tissue is a drug-metabolizing organ that protects tumor cells from chemotherapeutic agents. We have developed a tissue engineered, three-dimensional, high-throughput, patient specific, biomimetic model of BC that incorporates patient-derived adipocytes, adipose stromal cells, breast-duct organoids, and BC cells, and studied the effect of treatment with the chemotherapeutic agent doxorubicin on BC cell survival. Methods: Under an approved IRB, breast tissue was acquired from patients and differentially processed to isolate mature adipocytes, stromal cells, and breast organoids which were subsequently co-cultured with cancer cells in a 0.6% type I collagen matrix. Mixtures containing fluorescently tagged MDA-MB-231 or MDA-MB-468 triple negative BC cells at a concentration of 200,000 cells/mL were plated onto 96 well plates. BC cells in plain collagen and biomimetic hydrogels without BC served as controls. Groups composed of three replicates of both Biomimetic and collagen only controls were treated with one time with serially diluted doses of doxorubicin at 0, 0.001, 0.01, 0.1, 1, and 10 uM, in cell culture media and fixed after 3 days in culture, stained, and fluorescently imaged using confocal microscopy. Tissues from three patients were used to test each cell line. Analysis was performed using Imaris™ software. Results: After 3 days of doxorubicin treatment, MDA-MB-231 and MDA-MB-468 cell lines showed increased survival at 10mM doxorubicin (p<0.05) in the biomimetic platform derived from patient tissue, when compared to BC cells cultured in the collagen only controls (Fig 1. A and B). Moreover, fluorescent confocal microscopy demonstrated doxorubicin uptake by adipocytes that was directly proportional to increasing doxorubicin concentrations. No increased doxorubicin fluorescence was detected within the collagen-only controls (Fig 1. C and D). Conclusion: When cultured within a tissue engineered biomimetic platform comprised of patient tissue components, we observed significantly increased survival of two different triple negative breast cancer cell lines (MDA-MB-231 and MDA-MB-468) when compared to cultures in collagen alone. These data clearly demonstrate that the tumor microenvironment and neighboring tissue components modulates the effect of doxorubicin therapy on two different BC cell lines at least partially through adipocyte sequestration of doxorubicin. The ability of our platform to be personalized for individual patients may allow us to not only elucidate how the microenvironment affects BC cells, but also how patient specific tissue impacts the effects of various chemotherapeutic agents.
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