Gliomagenesis induces profound changes in the composition of the extracellular matrix (ECM) of the brain. In this study, we identified a cellular population responsible for the increased deposition of collagen I and fibronectin in glioblastoma. Elevated levels of the fibrillar proteins collagen I and fibronectin were associated with the expression of fibroblast activation protein (FAP), which is predominantly found in pericyte-like cells in glioblastoma. FAP+ pericyte-like cells were present in regions rich in collagen I and fibronectin in biopsy material and produced substantially more collagen I and fibronectin in vitro compared to other cell types found in the GBM microenvironment. Using mass spectrometry, we demonstrated that 3D matrices produced by FAP+ pericyte-like cells are rich in collagen I and fibronectin and contain several basement membrane proteins. This expression pattern differed markedly from glioma cells. Finally, we have shown that ECM produced by FAP+ pericyte-like cells enhances the migration of glioma cells including glioma stem-like cells, promotes their adhesion, and activates focal adhesion kinase (FAK) signaling. Taken together, our findings establish FAP+ pericyte-like cells as crucial producers of a complex ECM rich in collagen I and fibronectin, facilitating the dissemination of glioma cells through FAK activation.

• Keywords
• collagen type I, extracellular matrix proteins, fibronectin, glioblastoma, pericytes, proteomics,
• MeSH
• Endopeptidases MeSH
• Extracellular Matrix * metabolism   pathology   MeSH
• Fibronectins * metabolism   MeSH
• Glioblastoma pathology   metabolism   MeSH
• Glioma * pathology   metabolism   MeSH
• Collagen Type I metabolism   MeSH
• Humans MeSH
• Membrane Proteins metabolism   MeSH
• Cell Line, Tumor MeSH
• Tumor Microenvironment physiology   MeSH
• Brain Neoplasms * pathology   metabolism   MeSH
• Pericytes * metabolism   pathology   MeSH
• Cell Movement physiology   MeSH
• Serine Endopeptidases metabolism   MeSH
• Gelatinases metabolism   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Endopeptidases MeSH
• fibroblast activation protein alpha MeSH Browser
• Fibronectins * MeSH
• Collagen Type I MeSH
• Membrane Proteins MeSH
• Serine Endopeptidases MeSH
• Gelatinases MeSH

BACKGROUND & AIMS: Patient-derived organoid cancer models are generated from epithelial tumor cells and reflect tumor characteristics. However, they lack the complexity of the tumor microenvironment, which is a key driver of tumorigenesis and therapy response. Here, we developed a colorectal cancer organoid model that incorporates matched epithelial cells and stromal fibroblasts. METHODS: Primary fibroblasts and tumor cells were isolated from colorectal cancer specimens. Fibroblasts were characterized for their proteome, secretome, and gene expression signatures. Fibroblast/organoid co-cultures were analyzed by immunohistochemistry and compared with their tissue of origin, as well as on gene expression levels compared with standard organoid models. Bioinformatics deconvolution was used to calculate cellular proportions of cell subsets in organoids based on single-cell RNA sequencing data. RESULTS: Normal primary fibroblasts, isolated from tumor adjacent tissue, and cancer associated fibroblasts retained their molecular characteristics in vitro, including higher motility of cancer associated compared with normal fibroblasts. Importantly, both cancer-associated fibroblasts and normal fibroblasts supported cancer cell proliferation in 3D co-cultures, without the addition of classical niche factors. Organoids grown together with fibroblasts displayed a larger cellular heterogeneity of tumor cells compared with mono-cultures and closely resembled the in vivo tumor morphology. Additionally, we observed a mutual crosstalk between tumor cells and fibroblasts in the co-cultures. This was manifested by considerably deregulated pathways such as cell-cell communication and extracellular matrix remodeling in the organoids. Thrombospondin-1 was identified as a critical factor for fibroblast invasiveness. CONCLUSION: We developed a physiological tumor/stroma model, which will be vital as a personalized tumor model to study disease mechanisms and therapy response in colorectal cancer.

• Keywords
• Cancer, Co-cultures, Colorectal Cancer, Fibroblasts, Organoids,
• MeSH
• Cancer-Associated Fibroblasts * metabolism   MeSH
• Fibroblasts metabolism   MeSH
• Coculture Techniques MeSH
• Colorectal Neoplasms * pathology   MeSH
• Humans MeSH
• Tumor Microenvironment MeSH
• Organoids metabolism   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Cancer represents an extremely complicated ecosystem where cancer cells communicate with non-cancer cells present in the tumour niche through intercellular contacts, paracrine production of bioactive factors and extracellular vesicles, such as exosomes [...].

• MeSH
• Ecosystem MeSH
• Exosomes * metabolism   MeSH
• Extracellular Vesicles * metabolism   MeSH
• Humans MeSH
• Cell Communication MeSH
• Tumor Microenvironment MeSH
• Neoplasms * metabolism   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Editorial MeSH

Fibroblast activation protein (FAP) is a membrane-bound protease that is upregulated in a wide range of tumours and viewed as a marker of tumour-promoting stroma. Previously, we demonstrated increased FAP expression in glioblastomas and described its localisation in cancer and stromal cells. In this study, we show that FAP+ stromal cells are mostly localised in the vicinity of activated CD105+ endothelial cells and their quantity positively correlates with glioblastoma vascularisation. FAP+ mesenchymal cells derived from human glioblastomas are non-tumorigenic and mostly lack the cytogenetic aberrations characteristic of glioblastomas. Conditioned media from these cells induce angiogenic sprouting and chemotaxis of endothelial cells and promote migration and growth of glioma cells. In a chorioallantoic membrane assay, co-application of FAP+ mesenchymal cells with glioma cells was associated with enhanced abnormal angiogenesis, as evidenced by an increased number of erythrocytes in vessel-like structures and higher occurrence of haemorrhages. FAP+ mesenchymal cells express proangiogenic factors, but in comparison to normal pericytes exhibit decreased levels of antiangiogenic molecules and an increased Angiopoietin 2/1 ratio. Our results show that FAP+ mesenchymal cells promote angiogenesis and glioma cell migration and growth by paracrine communication and in this manner, they may thus contribute to glioblastoma progression.

• Keywords
• angiogenesis, angiopoietin, fibroblast activation protein, glioblastoma, microenvironment, seprase, vessel destabilisation,
• Publication type
• Journal Article MeSH