Tumor Microenvironment (TME) has been recognized as vital constituent for promoting tumor growth, invasion and metastasis. TME comprises various heterogeneous cells including cancer associated fibroblasts (CAFs) whose tumor-promoting effect has been the main focus of many investigations in which specific key signaling pathways are involved. In a recent study we demonstrated that papillary fibroblasts (PFs) and reticular fibroblasts (RFs) react differently to cutaneous squamous cell carcinoma (cSCC) cell interaction in 3D-human organotypic skin cultures. RFs differentiated towards CAFs and enhanced tumor EMT and invasion. To understand if we can interfere in the differentiation process of PFs, RFs and CAFs, we compared the discrepancy between these fibroblast subtypes, followed by examining multi effects of inhibitors (i) targeting TGFβ, PI3K, mTOR and NOTCH pathways on cSCC cell line MET2, PFs, RFs and CAFs in 2D and 3D cultures. Here we observed that PFs, RFs and CAFs are diverse in the range of morphology, proliferation, and gene expression profile. All inhibitors except TGFβi dampened tumor cell viability while only PI3Ki and mTORi showed such effect on fibroblast subtypes. All inhibitors strongly decreased the expression of CAF markers (e.g. α-SMA, COL11A1) while TGFβi and PI3Ki concurrently increased PF marker (e.g. Decorin, NTN1) expression in CAFs. Co-culturing MET2 with RFs or CAFs led to greater invasion, increased Ki67 expression, enhanced EMT marker (e.g. N-cad, MMP9) expression, elevated cytokine secretion (e.g. IL-6, IL-8) and profound CAF marker (e.g. ASMA, COL11A1) expression compared to PFs, which were further downregulated by the inhibitors with PI3Ki having the strongest effect. In conclusion, we elucidated the discrepancy between PFs, RFs, CAFs and further demonstrated the effect of different pathway inhibitors on both cSCC tumor cell line and fibroblast subtypes. Our study brings insight to the utilize of pathway inhibitors on future TME targeting therapy in cSCC.

Tumor Microenvironment (TME) has been recognized as vital constituent for promoting tumor growth, invasion and metastasis. TME comprises a variety of heterogeneous cells, including cancer associated fibroblasts (CAFs) whose tumor-promoting effect has been the main focus of many investigations in which specific key signaling pathways are involved, including TGFβ, PI3K/Akt/mTOR and NOTCH signaling pathway. Specific inhibitors designed to target key molecules in up-mentioned pathways have been widely studied in cancer research for their functions and anti-tumor mechanisms. However, the regulatory effect of these inhibitors on CAFs, especially on its activation, differentiation and interplay with tumor cells has not yet been elaborately observed. In our previous study, we demonstrated that two fibroblast subtypes isolated from normal human papillary dermis (PFs) and reticular dermis (RFs) respectively showed distinct functions on cutaneous squamous cell carcinoma (cSCC) tumor behavior in 3D organotypic culture. Moreover, RFs differentiated towards CAFs under the stimulation of tumor cell signaling. Here in this study, we included inhibitors (i) targeting TGF-β (SB431542), PI3K (LY294002), mTOR (Everolimus) and NOTCH (PF-03084014) pathways, and observed their effects on tumor cell proliferation, invasion and EMT, as well as the differentiation process of PFs, RFs and CAFs in 3D organotypic culture mimicking cSCC, aiming to bring insight to future utilize of pathway inhibitors on TME targeting therapy.

ESDR461 - De-activation of cancer associated fibroblast by pathway inhibitors in 3D-human organotypic cultures mimicking cutaneous squamous cell carcinoma

The scientific meeting of the European Society for Dermatological Research (ESDR) is held yearly, usually in September. With an international attendance of more than 1000 skin scientists from around the globe, the four-day event includes short oral presentations from peer-reviewed abstracts, keynote lectures from international experts, sponsored symposia, satellite programs, educational events as well as an important focus on posters, networking and building collaborations amongst scientists. For more information about this year’s meeting visit our website.