Metastasis of colon cancer requires Dickkopf-2 to generate cancer cells with Paneth cell properties

Metastasis is the leading cause of cancer-related mortality. Paneth cells provide stem cell niche factors in the homeostatic condition, but the underlying mechanisms of cancer stem cell niche development are unclear. Here we report that Dickkopf-2 (DKK2) is essential for Paneth-like cell generation during the metastasis of colon cancer. Splenic injection of DKK2-knockout (KO) cancer organoids into C57BL/6 mice resulted in a significant reduction of liver metastases compared to the control cancer organoids in spite of the presence of carcinogenic mutations in Apc, Kras and Tp53 genes. RNA-seq analysis of DKK2 KO organoids showed reduction of Paneth cell marker genes such as Lysozymes and Defensins. Ingenuity pathway analysis (IPA) of the RNA-seq data and assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) suggested HNF4A as a downstream target of DKK2. Chromatin immunoprecipitation followed by sequencing (Chip-seq) analysis showed that HNF4A binds to the open chromatin regions of Sox9, a well-known transcription factor for Paneth cell differentiation. Reduction of Paneth-like cells in the absence of DKK2 has been confirmed in the murine metastatic model of colon cancer using flow cytometry, immunohistochemistry and single cell RNA-seq. These data suggest the necessity of DKK2 in Paneth-like cell generation. Thus, DKK2 contributes to stem cell niche development during the metastatic progression of colon cancer. Overall design: Dickkopf-2 knockout colon cancer organoids carrying mutations in Apc, Kras and Tp53 genes were applied for ChIP-sequencing using recombinant Anti-HNF4-alpha antibody, ChIP Grade (ab181604) and rabbit IgG isotype antibody.