title: Genome-wide CRISPR screens reveal a Wnt-FZD5 signaling circuit as a druggable vulnerability of RNF43-mutant pancreatic tumors.
abstract: Forward genetic screens with CRISPR-Cas9 genome editing enable high-resolution detection of genetic vulnerabilities in cancer cells. We conducted genome-wide CRISPR-Cas9 screens in RNF43-mutant pancreatic ductal adenocarcinoma (PDAC) cells, which rely on Wnt signaling for proliferation. Through these screens, we discovered a unique requirement for a Wnt signaling circuit: engaging FZD5, one of the ten Frizzled receptors encoded in the human genome. Our results uncover an underappreciated level of context-dependent specificity at the Wnt receptor level. We further derived a panel of recombinant antibodies that reports the expression of nine FZD proteins and confirms that FZD5 functional specificity cannot be explained by protein expression patterns. Additionally, antibodies that specifically bind FZD5 and FZD8 robustly inhibited the growth of RNF43-mutant PDAC cells grown in vitro and as xenografts in vivo, providing orthogonal support for the functional specificity observed genetically. Proliferation of a patient-derived PDAC cell line harboring an RNF43 variant was also selectively inhibited by the FZD5 antibodies, further demonstrating their use as a potential targeted therapy. Tumor organoid cultures from colorectal carcinoma patients that carried RNF43 mutations were also sensitive to the FZD5 antibodies, highlighting the potential generalizability of these findings beyond PDAC. Our results show that CRIPSR-based genetic screens can be leveraged to identify and validate cell surface targets for antibody development and therapy.
authors.affiliation: Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Zachary
authors.initials: Z
authors.lastname: Steinhart
authors.affiliation: Donnelly Centre, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Zvezdan
authors.initials: Z
authors.lastname: Pavlovic
authors.affiliation: Donnelly Centre, University of Toronto, Toronto, Ontario, Canada.|Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Megha
authors.initials: M
authors.lastname: Chandrashekhar
authors.affiliation: Donnelly Centre, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Traver
authors.initials: T
authors.lastname: Hart
authors.affiliation: Donnelly Centre, University of Toronto, Toronto, Ontario, Canada.|The Centre for the Commercialization of Antibodies and Biologics, Toronto, Ontario, Canada.
authors.forename: Xiaowei
authors.initials: X
authors.lastname: Wang
authors.affiliation: Donnelly Centre, University of Toronto, Toronto, Ontario, Canada.|Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Xiaoyu
authors.initials: X
authors.lastname: Zhang
authors.affiliation: Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Mélanie
authors.initials: M
authors.lastname: Robitaille
authors.affiliation: Donnelly Centre, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Kevin R
authors.initials: KR
authors.lastname: Brown
authors.affiliation: Hubrecht Institute for Developmental Biology and Stem Cell Research, University Medical Centre, Utrecht, Foundation, Hubrecht Organoid Technology, Utrecht, the Netherlands.
authors.forename: Sridevi
authors.initials: S
authors.lastname: Jaksani
authors.affiliation: Hubrecht Institute for Developmental Biology and Stem Cell Research, University Medical Centre, Utrecht, Foundation, Hubrecht Organoid Technology, Utrecht, the Netherlands.
authors.forename: René
authors.initials: R
authors.lastname: Overmeer
authors.affiliation: Hubrecht Institute for Developmental Biology and Stem Cell Research, University Medical Centre, Utrecht, Foundation, Hubrecht Organoid Technology, Utrecht, the Netherlands.
authors.forename: Sylvia F
authors.initials: SF
authors.lastname: Boj
authors.affiliation: Donnelly Centre, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Jarrett
authors.initials: J
authors.lastname: Adams
authors.affiliation: Donnelly Centre, University of Toronto, Toronto, Ontario, Canada.|The Centre for the Commercialization of Antibodies and Biologics, Toronto, Ontario, Canada.
authors.forename: James
authors.initials: J
authors.lastname: Pan
authors.affiliation: Hubrecht Institute for Developmental Biology and Stem Cell Research, University Medical Centre, Utrecht, Foundation, Hubrecht Organoid Technology, Utrecht, the Netherlands.
authors.forename: Hans
authors.initials: H
authors.lastname: Clevers
authors.affiliation: Donnelly Centre, University of Toronto, Toronto, Ontario, Canada.|Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Sachdev
authors.initials: S
authors.lastname: Sidhu
authors.affiliation: Donnelly Centre, University of Toronto, Toronto, Ontario, Canada.|Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada.|Canadian Institute for Advanced Research, Toronto, Ontario, Canada.
authors.forename: Jason
authors.initials: J
authors.lastname: Moffat
authors.affiliation: Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Ontario, Canada.|Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Stéphane
authors.identifier: 0000-0001-7241-9044
authors.initials: S
authors.lastname: Angers
mesh_terms: D000230:Adenocarcinoma / Q000235:genetics / Q000378:metabolism; D000818:Animals; D000906:Antibodies / Q000494:pharmacology*; D021441:Carcinoma, Pancreatic Ductal / Q000235:genetics* / Q000378:metabolism; D059447:Cell Cycle Checkpoints / Q000187:drug effects*; D045744:Cell Line, Tumor; D064112:Clustered Regularly Interspaced Short Palindromic Repeats; D015179:Colorectal Neoplasms / Q000235:genetics* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics*; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D051157:Frizzled Receptors / Q000037:antagonists & inhibitors* / Q000378:metabolism; D006801:Humans; D051379:Mice; D016513:Mice, SCID; D058990:Molecular Targeted Therapy; D009368:Neoplasm Transplantation; D015513:Oncogene Proteins / Q000235:genetics*; D009940:Organoids / Q000187:drug effects / Q000378:metabolism; D010190:Pancreatic Neoplasms / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D044767:Ubiquitin-Protein Ligases; D060449:Wnt Signaling Pathway / Q000187:drug effects* / Q000235:genetics
chemical_list: D000906:Antibodies; D004268:DNA-Binding Proteins; C498589:FZD5 protein, human; D051157:Frizzled Receptors; D015513:Oncogene Proteins; D012333:RNA, Messenger; C496945:RNF43 protein, human; D044767:Ubiquitin-Protein Ligases