Identification of small-molecule inhibitors targeting different signaling pathways in cancer-associated fibroblast reprogramming under tumor-stroma interaction

Abstract

Cancer-associated fibroblasts (CAFs) interact reciprocally with tumor cells through various signaling pathways in many cancer types, including cutaneous squamous cell carcinoma. Among normal fibroblast subtypes, papillary fibroblasts (PFs) and reticular fibroblasts (RFs) respond distinctly to tumor cell signaling, leading predominantly to the differentiation of RFs rather than PFs into CAFs. The regulation of subtype differentiation in fibroblasts, however, remains poorly understood

In this study, we assessed the differences between PFs, RFs, and CAFs and examined the effects of small-molecule inhibitors targeting the TGFβ, phosphoinositide 3-kinase/protein kinase B/mTOR, and NOTCH pathways on both the tumor-promoting properties of CAFs and CAF reprogramming in 2D and 3D cultures. Blocking TGFβ and phosphoinositide 3-kinase strongly deactivated CAFs while inducing a PF-like phenotype in both RFs and CAFs.

Three-dimensional coculturing of a cutaneous squamous cell carcinoma cell line (MET2) with RFs or CAFs led to enhanced tumor invasion, RF–CAF transition, and cytokine production. These effects were significantly suppressed by blocking the TGFβ and phosphoinositide 3-kinase/mTOR pathways, but not the NOTCH pathway. In conclusion, this study identified biomarkers for PFs, RFs, and CAFs and demonstrated the differential effects of blocking key signaling pathways in CAFs and tumor cell–CAF interactions. These findings highlight a potential CAF-to-PF reprogramming therapeutic strategy and provide valuable insights into the use of pathway-specific inhibitors in CAF-targeted cancer therapy.