Related Experiment Video
Updated: Sep 6, 2025
Reprograming Model of Human Monocyte-derived Macrophages for In-vitro Assays
Published on: April 18, 2025
In vitro 2D and 3D cancer models to evaluate compounds that modulate macrophage polarization
Natasha Helleberg Madsen1, Boye Schnack Nielsen1, Jesper Larsen1
1Bioneer A/S, Kogle Allé 2, 2970 Hørsholm, Denmark.
Abstract:
In vitro cancer models that can identify novel immunomodulating compounds are essential. Using a 3D multicellular tumor spheroid (MCTS) model comprising cancer cells, fibroblasts, and macrophages, we tested tumor-associated macrophage (TAM)-inhibiting compounds (CCL2 Ab, CSF1R inhibitor, CSF1R Ab) and TAM-reprograming compounds (poly I:C, CD40 Ab, CD40 ligand) for their effects on monocyte infiltration and polarization in tumor spheroids. For characterization of macrophage polarization, we measured the expression of CD206, CD163, CD86, MHC II, CD40, and CD14 and measured 43 soluble factors in the 3D MCTS cultures. 2D macrophage models were evaluated for comparison. A CSF1R inhibitor prevented infiltration of monocytes into pancreatic cancer spheroids, and macrophages treated with the inhibitor showed decreased expression of M2 markers. Treatment with a CD40 ligand and poly I:C induced M1 macrophage polarization in our models. We propose that these models can be used to improve the drug screening process of anti-cancer immunotherapies targeting macrophages.
Insights
This study developed a 3D multicellular tumor spheroid model to test immunomodulating compounds. A CSF1R inhibitor reduced monocyte infiltration and M2 marker expression in cancer spheroids, while CD40 ligand and poly I:C promoted M1 macrophage polarization.
Area of Science:
- Oncology
- Immunology
- Drug Discovery
Background:
- Developing in vitro cancer models is crucial for identifying novel immunomodulating compounds.
- Tumor-associated macrophages (TAMs) play a significant role in cancer progression and immune evasion.
- Targeting TAMs offers a promising strategy for anti-cancer immunotherapy.
Purpose of the Study:
- To evaluate the efficacy of TAM-inhibiting and TAM-reprograming compounds using a 3D multicellular tumor spheroid (MCTS) model.
- To assess the impact of these compounds on monocyte infiltration and macrophage polarization within tumor spheroids.
- To establish a robust model for screening anti-cancer immunotherapies targeting macrophages.
Main Methods:
- Established a 3D MCTS model incorporating cancer cells, fibroblasts, and macrophages.
- Tested TAM-inhibiting compounds (CCL2 Ab, CSF1R inhibitor, CSF1R Ab) and TAM-reprograming compounds (poly I:C, CD40 Ab, CD40 ligand).
- Characterized macrophage polarization by measuring CD206, CD163, CD86, MHC II, CD40, and CD14 expression, and analyzed 43 soluble factors. Compared results with 2D macrophage models.
Main Results:
- A CSF1R inhibitor effectively prevented monocyte infiltration into pancreatic cancer spheroids.
- Macrophages treated with the CSF1R inhibitor exhibited reduced expression of M2 markers.
- Treatment with CD40 ligand and poly I:C induced M1 macrophage polarization in the 3D MCTS model.
Conclusions:
- The 3D MCTS model is a valuable tool for evaluating immunomodulating compounds targeting macrophages.
- This model can improve the drug screening process for anti-cancer immunotherapies.
- Targeting TAMs with specific inhibitors or reprogramming agents shows potential in modulating the tumor immune microenvironment.
