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Genome-wide CRISPR screens reveal a Wnt–FZD5 signaling circuit as a druggable vulnerability of RNF43-mutant pancreatic tumors

A Corrigendum to this article was published on 01 November 2017

This article has been updated

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.

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Figure 1: Genome-wide CRISPR–Cas9 screens identify genetic vulnerabilities of RNF43-mutant PDAC cells.
Figure 2: FZD5 knockout inhibits proliferation of RNF43-mutant PDAC cell lines and Wnt target genes activation.
Figure 3: Wnt and Frizzled expression patterns are not predictive of essentiality.
Figure 4: Anti-FZD5 antibodies inhibit growth of RNF43-mutant PDAC through cell-cycle arrest.
Figure 5: Anti-FZD5 antibodies inhibit growth of RNF43-mutant PDAC in in vivo xenograft models.
Figure 6: Anti-FZD5 IgG inhibits survival of RNF43-mutant colorectal cancer (CRC) organoids.

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  • 20 September 2017

    In the version of this article initially published, duplicate panels in Figure 5e were incorrectly used in the assembly of the figure, for which all original panels are presented in Supplementary Figure 10. The errors in Figure 5e have been corrected in the HTML and PDF versions of the article.

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Acknowledgements

We wish to thank members of the Angers and Moffat labs for discussions. We are thankful to D. Hedley (Princess Margaret Hospital, Toronto, Canada) and M. Tsao (Ontario Cancer Institute, Toronto, Canada) for providing the patient-derived xenograft cells; P. Mero (Moffat lab) for help with images. This work was supported from grants funded by the Canadian Institutes for Health Research to S.A. (CIHR-273548) and J.M. (CIHR-342551) and by the Ontario Research Fund to S.S. J.M. holds a Canada Research Chair in Functional Genomics of Cancer and S.A. holds a Canada Research Chair in Functional Architecture of Signal Transduction. Some of the equipment used in this study was supported by The 3D (Diet, Digestive Tract and Disease) Centre funded by the Canadian Foundation for Innovation and Ontario Research Fund, project numbers 19442 and 30961.

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Contributions

Conceptualization, S.A., J.M., S.S.; investigation, Z.S., Z.P., M.C., T.H., X.W., X.Z., M.R., S.J., R.O., S.F.B., J.A., K.R.B.; writing, S.A., Z.S., J.M.; funding acquisition, S.A., J.M., S.S.; supervision, S.A., J.M., S.S., H.C., and J.P.

Corresponding authors

Correspondence to Sachdev Sidhu, Jason Moffat or Stéphane Angers.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Figures and Table

Supplementary Figures 1–11 and Supplementary Table 4 (PDF 4227 kb)

Supplementary Table 1

Raw read counts (XLSX 11317 kb)

Supplementary Table 2

Bayes factors (XLSX 1580 kb)

Supplementary Table 3

Differential fitness z-scores (XLSX 1314 kb)

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Steinhart, Z., Pavlovic, Z., Chandrashekhar, M. et al. Genome-wide CRISPR screens reveal a Wnt–FZD5 signaling circuit as a druggable vulnerability of RNF43-mutant pancreatic tumors. Nat Med 23, 60–68 (2017). https://doi.org/10.1038/nm.4219

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