A Wnt-induced phenotypic switch in cancer-associated fibroblasts inhibits EMT in colorectal cancer

Tumor progression is recognized as a result of an evolving crosstalk between tumor cells and their surrounding non-transformed stroma. Although Wnt signaling has been intensively studied in colorectal cancer (CRC), it remains unclear whether activity in the tumor-associated stroma contributes to malignancy. To specifically interfere with stromal signals, we generated Wnt-independent tumor organoids that secrete the Wnt antagonist Sfrp1. Upon subcutaneous transplantation into immuno-competent as well as -deficient mice, we observed strong reduction of tumor growth. Histological and transcriptomic analyses revealed that Sfrp1 induces an EMT (epithelial-mesenchymal transition) phenotype in tumor cells without affecting tumor-intrinsic Wnt signaling, suggesting involvement of non-immune stromal cells. Indeed, blockage of canonical signaling using Sfrp1, Dkk1, or fibroblast-specific genetic ablation of ß-catenin strongly decreased the number of cancer-associated myofibroblasts (myCAFs). Interestingly, we found that the Wnt activity in CAFs is linked with distinct subtypes, where low and high levels induce an inflammatory-like CAF (iCAF) subtype or contractile myCAFs, respectively. Co-culture of tumor organoids with iCAFs resulted in massive upregulation of EMT markers, while myCAFs reverted this phenotype. In summary, we show that tumor growth and malignancy are differentially regulated via district fibroblast subtypes under the influence of juxtacrine Wnt signals. Overall design: Generation of Wnt-independent mouse tumor organoids that overexpress the Wnt antagonist Sfrp1 to block stromal Wnt signals and study the consequences on tumor growth and progression. Generated tumor lines were sequenced in vitro and upon sorting of Epcam+ and Pdgfra+ cells from subcutaneous tumors. Stromal consequences were also studied of human xenografts after Sfrp1 overexpression. Moreover, mouse fibroblasts were cultured in the presence and absence of Wnt stimulation for validation.