Abstract
Tumor-associated macrophages often correlate with tumor progression, and therapies targeting immune cells in tumors have emerged as promising treatments. To select effective therapies, we established an in vitro 3D multicellular spheroid model including cancer cells, fibroblasts, and monocytes. We analyzed monocyte infiltration and differentiation in spheroids generated from fibroblasts and either of the cancer cell lines MCF-7, HT-29, PANC-1, or MIA PaCa-2. Monocytes rapidly infiltrated spheroids and differentiated into mature macrophages with diverse phenotypes in a cancer cell line-dependent manner. MIA PaCa-2 spheroids polarized infiltrating monocytes to M2-like macrophages with high CD206 and CD14 expression, whereas monocytes polarized by MCF-7 spheroids displayed an M1-like phenotype. Monocytes in HT-29 and PANC-1 primarily obtained an M2-like phenotype but also showed upregulation of M1 markers. Analysis of the secretion of 43 soluble factors demonstrated that the cytokine profile between spheroid cultures differed considerably depending on the cancer cell line. Secretion of most of the cytokines increased upon the addition of monocytes resulting in a more inflammatory and pro-tumorigenic environment. These multicellular spheroids can be used to recapitulate the tumor microenvironment and the phenotype of tumor-associated macrophages in vitro and provide more realistic 3D cancer models allowing the in vitro screening of immunotherapeutic compounds.
Highlights
Crosstalk between different cells in the tumor microenvironment (TME) plays an important role in tumor growth and tumor-mediated immune suppression in vivo
After 7 days of co-culture, spheroids generated from MCF-7, HT-29, or PANC-1 obtained a spherical shape and stable appearance, with different cell line-dependent spheroid sizes despite identical seeding conditions (Figure 1A)
Histological examination identified the HT-29 spheroids to be spherical with a central core containing necrotic cells, whereas MIA PaCa-2 and PANC-1 spheroids were looser in the structure, often showing a dense cell core and loose periphery (Figure 1C)
Summary
Crosstalk between different cells in the tumor microenvironment (TME) plays an important role in tumor growth and tumor-mediated immune suppression in vivo. The presence of infiltrating immune cells in tumors often correlates with tumor growth and progression [1,2,3,4]. The TAM compartment is both highly dynamic and tumor-specific. This heterogeneity reflects the ability of macrophages to acquire an entire spectrum of phenotypic, metabolic, and functional profiles, ranging from a pro-inflammatory to an anti-inflammatory state [7]. M1-like TAMs can produce reactive oxygen species, destroy malignant cells, and release pro-inflammatory cytokines, further stimulating anti-cancer immunity [1]. Cancer cells often acquire the ability to polarize infiltrating monocytes toward an M2-like state with immunosuppressive and pro-angiogenic features that promote tumor development and invasion [7,8,9,10]. TAMs can either antagonize, augment, or mediate the antitumor effects of different anti-cancer therapies [6]
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