3D Model of the Early Melanoma Microenvironment Captures Macrophage Transition into a Tumor-Promoting Phenotype

SIMPLE SUMMARY: We developed a “tumor-in-a-dish” experimental system to study the early events favoring tumor growth and suppression of the immune response in metastatic melanoma. We combined murine melanoma tumor cells with fibroblasts and macrophages in a 3D collagen matrix and characterized how interactions between these three cell types, which are present in the early stages of tumorigenesis, drive immune suppression and the tumor-promoting transition in macrophages that is observed in vivo. Over the course of 7 days in the co-cultures, we quantified the dynamics of cues transmitted by direct cell–cell interactions, through the extracellular matrix and through secretion of immune mediators. We found that macrophages acquired features and a functional profile consistent with those present in in vivo murine melanoma tumors. This system will enable future studies of macrophage–stromal cross-talk in the melanoma microenvironment and provide a platform to test potential therapeutic approaches aimed at stimulating immune activity in macrophages. ABSTRACT: Tumor immune response is shaped by the tumor microenvironment (TME), which often evolves to be immunosuppressive, promoting disease progression and metastasis. An important example is melanoma tumors, which display high numbers of tumor-associated macrophages (TAMs) that are immunosuppressive but also have the potential to restore anti-tumor activity. However, to therapeutically target TAMs, there is a need to understand the early events that shape their tumor-promoting profile. To address this, we built and optimized 3D in vitro co-culture systems, composed of a collagen-I matrix scaffolding murine bone-marrow-derived macrophages (BMDMs), YUMM1.7 melanoma cells, and fibroblasts to recreate the early melanoma TME and study how interactions with fibroblasts and tumor cells modulate macrophage immune activity. We monitored BMDM behavior and interactions through time-lapse imaging and characterized their activation and secretion. We found that stromal cells induced a rapid functional activation, with increased motility and response from BMDMs. Over the course of seven days, BMDMs acquired a phenotype and secretion profile that resembled melanoma TAMs in established tumors. Overall, the direct cell–cell interactions with the stromal components in a 3D environment shape BMDM transition to a TAM-like immunosuppressive state. Our systems will enable future studies of changes in macrophage–stromal cross-talk in the melanoma TME.