Modeling the oxygen-deprived breast cancer tumor microenvironment within a three-dimensional bioengineered platform that exhibits hypoxia-driven immune evasion

Introduction: Oxygen deprivation within tumors has been associated with enhanced cancer cell survival, altered extracellular matrix (ECM) production, and increased immune evasion in breast cancer (BCa). These outcomes collectively contribute to chemotherapeutic challenges. Current models do not adequately mimic physiologic oxygen levels relevant to breast tissue and tumor-immune interactions. Taking this into consideration, we developed a plasma-derived three-dimensional (3D) model that supports growth of BCa cells, generates physio- and pathophysiologic oxygen levels, and supports prevailing hypoxia-driven tumor outcomes such as immune evasion. Methods: BCa cells (MCF7 and MDA-MB-231) were embedded into 3D scaffolds formed through the cross-linking of patient-derived plasma. The oxygen content within these 3D matrices was engineered and further characterized to recapitulate physiologic blood and breast tissue (3D physiologic, gradient 14-10 kPa) or pathophysiologic (3D tumorous, gradient 9.6-2.4 kPa) levels by the manipulation of height, culture time, oxygen incubation environment, and cellular concentration. BCa cells grown within 3D physiologic and tumorous scaffolds were analyzed by flow cytometry, confocal imaging, or immunohistochemistry for the expression of hypoxia-inducible factor-1 alpha (HIF-1 alpha), proliferation capabilities, and ECM production. Peripheral blood mononuclear cells were incorporated on the top of 3D physiologic and tumorous scaffolds, and the differences in the tumor-infiltrating capabilities of T cells into 3D physiologic and tumorous scaffolds were assessed by flow cytometry. Lastly, hypoxic inhibition was validated as a target to sensitize BCa cells in order to overcome immune surveillance. Results: BCa cells grown in oxygen-deprived 3D tumorous environments showed a significant increase in the expression of HIF-1 alpha and a reduced cell proliferation compared to the BCa cells in 3D physiologic scaffolds. Increased secretion of collagen I, collagen III, laminin, and fibronectin was demonstrated by BCa cells in 3D tumorous compared to 3D physiologic scaffolds. T-cell infiltration (especially CD8 T cells) was significantly impaired within the oxygen-deprived breast cancer tumor microenvironment (TME), and hypoxia inhibition was further confirmed to resensitize BCa cells to cytotoxic CD8 T cells. Conclusions: Modeling of the oxygen-deprived breast cancer TME in our engineered 3D physiologic and tumorous scaffolds supported known intratumoral hypoxia characteristics such as reduced BCa cell proliferation, increased ECM expression, and hindered immune infiltration. Furthermore, hypoxic inhibition was validated as a target to sensitize BCa cells to immune surveillance. These models could serve as a promising platform for the evaluation of immunologic events as well as a drug-screening platform tool to overcome hypoxia-driven immune evasion. Citation Format: Somshuvra Bhattacharya, Kristin Calar, Claire Evans, Pilar de la Puente. Modeling the oxygen-deprived breast cancer tumor microenvironment within a three-dimensional bioengineered platform that exhibits hypoxia-driven immune evasion [abstract]. In: Proceedings of the AACR Special Conference on the Evolving Landscape of Cancer Modeling; 2020 Mar 2-5; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2020;80(11 Suppl):Abstract nr B29.