Porcupine inhibitor suppresses paracrine Wnt-driven growth of Rnf43;Znrf3-mutant neoplasia

Rnf43 (RING finger protein 43) and Znrf3 (zinc/RING finger protein 3) (RZ) are two closely related transmembrane E3 ligases, encoded by Wnt target genes, that remove surface Wnt (wingless-int) receptors. The two genes are mutated in various human cancers. Such tumors are predicted to be hypersensitive to, yet still depend on, secreted Wnts. We previously showed that mutation of RZ in the intestine yields rapidly growing adenomas containing LGR5 ⁺ (leucine-rich repeat-containing G-protein coupled receptor 5) stem cells and Wnt3-producing Paneth cells. We now show that removal of Paneth cells by Math1 mutation inhibits RZ ⁻/⁻ tumor formation. Similarly, deletion of Wnt3 inhibits tumorigenesis. Treatment of mice carrying RZ ⁻/⁻ intestinal neoplasia with a small molecule Wnt secretion inhibitor (porcupine inhibitor C59) strongly inhibited growth, whereas adjacent normal crypts remained intact. These results establish that paracrine Wnt secretion is an essential driver of RZ ⁻/⁻ tumor growth and imply that a therapeutic window exists for the use of porcupine inhibitors for RZ-mutant cancers. Significance Rnf43 (RING finger protein 43) and Znrf3 (zinc/RING finger protein 3), encoded by stem cell-specific Wnt (wingless-int) target genes, constitute a crucial negative feedback loop in the Wnt signaling pathway. Rnf43 is mutated in subsets of human cancers of the colon, pancreas, stomach, ovary, and liver, while Znrf3 is mutated in adrenocortical carcinoma and osteoblastoma. Indeed, when both genes are mutated simultaneously in small intestinal stem cells in mice, tumors arise within a few weeks. Treatment of mice carrying RZ ⁻/⁻ intestinal neoplasia with a small molecule Wnt secretion inhibitor strongly inhibited growth, while adjacent normal crypts remained intact. These results establish that paracrine Wnt secretion is an essential driver of RZ ⁻/⁻ tumor growth and imply that a therapeutic window exists for the use of porcupine inhibitors for RZ-mutant cancers.