LAUSR

Intestinal tumor initiation is a probabilistic process that depends on the competition between mutant and normal epithelial stem cells in crypts. Intestinal stem cells are closely associated with a diverse but poorly characterized network of mesenchymal cell types. It remains unknown whether the physiologic mesenchymal microenvironment of mutant stem cells affects tumor initiation. Here we show that the mesenchymal niche controls tumor initiation in trans. By characterizing the heterogeneity of the intestinal mesenchyme by single cell RNA-seq, we identified a population of rare pericryptal Ptgs2-expressing fibroblasts (RPPFs) that constitutively process arachidonic acid into highly labile prostaglandin E2 (PGE2). Specific ablation of Ptgs2 in fibroblasts was sufficient to prevent tumor initiation in two different models of sporadic, autochthonous tumorigenesis. At the mechanistic level, by performing single-cell RNA-seq analyses of a mesenchymal niche organotypic model, we found that fibroblast-derived PGE2 reprograms intestinal stem cells into Tuft cell precursors. These are characterized by the activation of a strong tumor initiation program driven by the Hippo pathway effector Yap, a molecule indispensable for intestinal tumorigenesis. Organoid experiments established a novel molecular mechanism whereby PGE2 activates Yap dephosphorylation, nuclear translocation, and transcriptional activity by signaling specifically via receptor Ptger4. In vivo, epithelial-specific ablation of Ptger4 misdirected the regenerative reprogramming of stem cells, abrogated Yap target gene activation, and prevented sporadic tumor initiation in mutant mice, thereby demonstrating the robust paracrine control of tumor-initiating stem cells by PGE2/Ptger4. Patient-derived organoid analyses established that PGE2/ PTGER4 also regulate stem cell function in humans. Our study demonstrates for the first time that colorectal cancer initiation is orchestrated by the mesenchymal niche. Moreover, it reveals a mechanism whereby RPPFs exert paracrine control over tumor-initiating stem cells via the druggable PGE2/Ptger4/Yap signaling axis. Citation Format: Manolis Roulis, Emilios Kaklamanos, George Kollias, Richard Flavell. Paracrine orchestration of intestinal tumorigenesis at the mesenchymal-epithelial interface [abstract]. In: Proceedings of the AACR Special Conference on the Hippo Pathway: Signaling, Cancer, and Beyond; 2019 May 8-11; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2020;18(8_Suppl):Abstract nr PR12.