In Vitro 3D Spheroid Model Preserves Tumor Microenvironment of Hot and Cold Breast Cancer Subtypes.

Abstract

Dynamic interaction of cancer, immune, and stromal cells with extracellular matrix components modulates and resists the response of standard care therapies. To mimic this, an in vitro 3D spheroid model is designed using liquid overlay method to simulate hot (MDA-MB-231) and cold (MCF-7) breast tumor microenvironment (TME). This study shows increased mesenchymal phenotype, stemness, and suppressive microenvironment in MDA-MB-231-spheroids upon exposure to doxorubicin. Intriguingly, the presence of human dermal fibroblasts enhances cancer-associated fibroblast phenotype in MDA-MB-231-spheroids through increased expression of CXCL12 and FSP-1, leading to higher infiltration of immune cells (THP-1 monocytes). However, a suppressive TME is observed in both subtypes, as seen by upregulation of M2-macrophage-specific CD68 and CD206 markers. Specifically, increased PDL-1 expressing tumor-associated macrophages along with FoxP3 expressing T regulatory cells are found in MDA-MB-231-spheroids when cultured with peripheral blood mononuclear cells. Further, it is found that the addition of 1-methyl-tryptophan, a potent indoleamine-2,3-dioxygenase-1 inhibitor, subsides the suppressive phenotype by decreasing the M2 polarization via downregulation of tryptophan metabolism and IL10 expression, particularly in MCF-7 triculture spheroids. Thus, the in vitro 3D spheroid model of TME can be utilized in therapeutics to validate immunomodulatory drugs for various breast cancer subtypes.