Abstract
BACKGROUND: The tumor microenvironment (TME) composition is among the critical events leading to the poor prognosis of head and neck cancers. TME includes immune cells, tumor-associated macrophages (TAMs), cancer-associated fibroblasts (CAF), and other non-cancerous cells, which contribute to therapeutic outcome. The heterogeneity of the TME offers a multitude of potential targets for photodynamic therapy (PDT). Advanced 3D models that closely mimic the microenvironment are a promising tool to study tumor-stroma interactions, TAM plasticity and their impact on Foslip(®)-PDT outcome. OBJECTIVE: The aim of this study was to assess the effect of Foslip-PDT on photo-induced macrophage re-education in 3D tri-culture model composed of cancer cells, CAFs, and macrophages. METHODS: A 3D model was established using FaDu cancer cells, MeWo fibroblasts, and PMA-differentiated U937 macrophages. The spheroids were characterized using immunochemistry, immunofluorescence, and qPCR. Foslip-based photoirradiation was applied to spheroids at different fluences to evaluate the photoinduced cell death. Macrophage phenotypes were assessed by flow cytometry. RESULTS: The 3D tri-culture model displayed hallmarks of stromal-tumor interactions, including CAF clustering, macrophage infiltration (~30-40%), and epithelial-mesenchymal transition. Macrophages in the spheroids had prevailing M2 phenotype as deduced from overexpression of immunosuppressive markers (CD163, PDL-1, IL-10). The liposomal photosensitizer Foslip accumulated 2 to 3 times more in macrophage-enriched spheroids, however, PDT induced similar levels of cell death in all tested models. At the same time, Foslip-PDT produced immunomodulatory effect in tri-culture model characterized by the increase of CD80-M1 marker expression and the decrease in the expression of the CD206-M2 marker. CONCLUSION: The 3D tri-culture model integrated essential features of the HNSCC microenvironment. Foslip-PDT was effective in reprograming M2 macrophages to tumor-killing M1 macrophages. This study opens the way to combine direct tumor damage with TME modulation.