Background
Patient-derived colorectal cancer (CRC)organoids (PDO) solely consisting of malignant cells led to major advances inthe understanding of cancer treatments. Yet, a major limitation is the absenceof cells from the tumor microenvironment (TME), thereby prohibiting potentialinvestigation of treatment responses on immune and structural cells. Here weutilize single cell transcriptomics to determine differential responses oftissue derived macrophages and peripheral blood derived monocytic cells toentry into the tumor microenvironment in a simulated ex vivo set up and utilize insights gained from these experimentsto set up a complex primary organotypic assay system consisting of PDO, cancerassociated fibroblasts (CAF) and primary monocytes as surrogate for tumorassociated macrophages (TAM) to probe treatment responses of chemotherapeuticsand oncolytic influenza A viruses.
Materials and Methods
Monocytes and ex vivo derived macrophages were co-cultured with CAF and subjectedto single-cell-RNA-seq. Complex co-cultures were set up, containing CRC PDO,patient matched CAF and monocytic cells and subjected to single cell RNA seq,flow cytometry and cytokine analysis.
Results
We discovered that CAF co-culture resulted in apartial differentiation of monocytes into macrophages and a phenotypic switch,characterized by the expression of major immunosuppressive markers comparableto TAMs in CRC. Macrophages did not appear to gain TAM-like features from CAFco-culture. Oxaliplatin and 5-FU, the standard of care chemotherapy for CRC,induced polarization of TAM-like cells to a pro-inflammatory phenotypecomparable to the immunogenic effects of treatment with an oncolytic virus.
Conclusions
Thus, primary CAF-containing triple culturessuccessfully model TAM-like phenotypes exvivo and allow the assessment of their functional and phenotypic changes inresponse to treatments following a precision medicine approach.
J. Kabiljo: None. A. Theophil: None. J. Homola: None. A. Renner: None. J. Karall: None. N. Hartman: None. S. Stang: None. L. Tran: None. J. Laengle: None. A. Kulu: None. A. Chen: None. M. Fabits: None. V. Atanasova: None. H. Walczak: None. D. Herndler-Brandstetter: None. G. Egger: None. H. Dolznig: None. A. Kusienicka: None. M. Farlik: None. M. Bergmann: None.