PMID- 33067269 OWN - NLM STAT- MEDLINE DCOM- 20210408 LR - 20210408 IS - 1538-7445 (Electronic) IS - 0008-5472 (Linking) VI - 80 IP - 24 DP - 2020 Dec 15 TI - The Functional Landscape of Patient-Derived RNF43 Mutations Predicts Sensitivity to Wnt Inhibition. PG - 5619-5632 LID - 10.1158/0008-5472.CAN-20-0957 [doi] AB - A subset of Wnt-addicted cancers are sensitive to targeted therapies that block Wnt secretion or receptor engagement. RNF43 loss-of-function (LOF) mutations that increase cell surface Wnt receptor abundance cause sensitivity to Wnt inhibitors. However, it is not clear which of the clinically identified RNF43 mutations affect its function in vivo. We assayed 119 missense and 45 truncating RNF43 mutations found in human cancers using a combination of cell-based reporter assays, genome editing, flow cytometry, and immunofluorescence microscopy. Five common germline variants of RNF43 exhibited wild-type activity. Cancer-associated missense mutations in the RING ubiquitin ligase domain and a subset of mutations in the extracellular domain hyperactivate Wnt/beta-catenin signaling through formation of inactive dimers with endogenous RNF43 or ZNRF3. RNF43 C-terminal truncation mutants, including the common G659fs mutant are LOF specifically when endogenous mutations are examined, unlike their behavior in transient transfection assays. Patient-derived xenografts and cell lines with C-terminal truncations showed increased cell surface Frizzled and Wnt/beta-catenin signaling and were responsive to porcupine (PORCN) inhibition in vivo, providing clear evidence of RNF43 impairment. Our study provides potential guidelines for patient assignment, as virtually all RNF43 nonsense and frameshift mutations, including those in the C-terminal domain and a large number of patient-associated missense mutations in the RING domain and N-terminal region compromise its activity, and therefore predict response to upstream Wnt inhibitors in cancers without microsatellite instability. This study expands the landscape of actionable RNF43 mutations, extending the benefit of these therapies to additional patients. SIGNIFICANCE: Systematic examination of patient-derived RNF43 mutations identifies rules to guide patient selection, including that truncation or point mutations in well-defined functional domains sensitize cancers to PORCN inhibitors. CI - (c)2020 American Association for Cancer Research. FAU - Yu, Jia AU - Yu J AD - Program in Cancer and Stem Cell Biology, Duke-NUS Medical School, Singapore, Singapore. FAU - Yusoff, Permeen A Mohamed AU - Yusoff PAM AD - Center for Technology and Development, Duke-NUS Medical School, Singapore, Singapore. FAU - Woutersen, Danielle T J AU - Woutersen DTJ AD - Department of Gastroenterology and Hepatology, Erasmus MC-University Medical Center, Rotterdam, the Netherlands. FAU - Goh, Pamela AU - Goh P AD - Center for Technology and Development, Duke-NUS Medical School, Singapore, Singapore. FAU - Harmston, Nathan AU - Harmston N AUID- ORCID: 0000-0002-8589-2938 AD - Program in Cancer and Stem Cell Biology, Duke-NUS Medical School, Singapore, Singapore. AD - Science Division, Yale-NUS College, Singapore, Singapore. FAU - Smits, Ron AU - Smits R AD - Department of Gastroenterology and Hepatology, Erasmus MC-University Medical Center, Rotterdam, the Netherlands. FAU - Epstein, David M AU - Epstein DM AD - Program in Cancer and Stem Cell Biology, Duke-NUS Medical School, Singapore, Singapore. AD - Center for Technology and Development, Duke-NUS Medical School, Singapore, Singapore. FAU - Virshup, David M AU - Virshup DM AUID- ORCID: 0000-0001-6976-850X AD - Program in Cancer and Stem Cell Biology, Duke-NUS Medical School, Singapore, Singapore. babita.madan@duke-nus.edu.sg david.virshup@duke-nus.edu.sg. AD - Department of Pediatrics, Duke University School of Medicine, Durham, North Carolina. FAU - Madan, Babita AU - Madan B AUID- ORCID: 0000-0003-1065-8589 AD - Program in Cancer and Stem Cell Biology, Duke-NUS Medical School, Singapore, Singapore. babita.madan@duke-nus.edu.sg david.virshup@duke-nus.edu.sg. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20201016 PL - United States TA - Cancer Res JT - Cancer research JID - 2984705R RN - 0 (ETC-159) RN - 0 (Frizzled Receptors) RN - 0 (Heterocyclic Compounds, 4 or More Rings) RN - 0 (Membrane Proteins) RN - EC 2.3.- (Acyltransferases) RN - EC 2.3.1.- (PORCN protein, human) RN - EC 2.3.2.27 (RNF43 protein, human) RN - EC 2.3.2.27 (Ubiquitin-Protein Ligases) SB - IM MH - Acyltransferases/antagonists & inhibitors MH - Animals MH - Cell Line, Tumor MH - Endocytosis/physiology MH - Frizzled Receptors/metabolism MH - Gene Expression Regulation, Neoplastic MH - Heterocyclic Compounds, 4 or More Rings/pharmacology MH - Humans MH - Membrane Proteins/antagonists & inhibitors MH - Mice, Nude MH - *Mutation MH - Neoplasms/drug therapy/*genetics/metabolism MH - Protein Multimerization MH - Ubiquitin-Protein Ligases/*genetics/*metabolism MH - Wnt Signaling Pathway/*drug effects/genetics MH - Xenograft Model Antitumor Assays EDAT- 2020/10/18 06:00 MHDA- 2021/04/10 06:00 CRDT- 2020/10/17 05:24 PHST- 2020/03/25 00:00 [received] PHST- 2020/08/23 00:00 [revised] PHST- 2020/10/12 00:00 [accepted] PHST- 2020/10/18 06:00 [pubmed] PHST- 2021/04/10 06:00 [medline] PHST- 2020/10/17 05:24 [entrez] AID - 0008-5472.CAN-20-0957 [pii] AID - 10.1158/0008-5472.CAN-20-0957 [doi] PST - ppublish SO - Cancer Res. 2020 Dec 15;80(24):5619-5632. doi: 10.1158/0008-5472.CAN-20-0957. Epub 2020 Oct 16.