Next-Generation Surrogate Wnts Support Organoid Growth and Deconvolute Frizzled Pleiotropy In Vivo

Yi Miao; Andrew Ha; Wim de Lau; Kanako Yuki; António J M Santos; Changjiang You; Maarten H Geurts; Jens Puschhof; Cayetano Pleguezuelos-Manzano; Weng Chuan Peng; Ramazan Senlice; Carol Piani; Jan W Buikema; Oghenekevwe M Gbenedio; Mario Vallon; Jenny Yuan; Sanne de Haan; Wieger Hemrika; Kathrin Rösch; Luke T Dang; David Baker; Melanie Ott; Philippe Depeille; Sean M Wu; Jarno Drost; Roeland Nusse; Jeroen P Roose; Jacob Piehler; Sylvia F Boj; Claudia Y Janda; Hans Clevers; Calvin J Kuo; K Christopher Garcia

doi:10.1016/j.stem.2020.07.020

Next-Generation Surrogate Wnts Support Organoid Growth and Deconvolute Frizzled Pleiotropy In Vivo

Cell Stem Cell. 2020 Nov 5;27(5):840-851.e6. doi: 10.1016/j.stem.2020.07.020. Epub 2020 Aug 19.

Authors

Yi Miao¹, Andrew Ha², Wim de Lau³, Kanako Yuki⁴, António J M Santos⁴, Changjiang You⁵, Maarten H Geurts³, Jens Puschhof³, Cayetano Pleguezuelos-Manzano³, Weng Chuan Peng⁶, Ramazan Senlice⁷, Carol Piani⁷, Jan W Buikema⁸, Oghenekevwe M Gbenedio⁹, Mario Vallon⁴, Jenny Yuan⁴, Sanne de Haan¹⁰, Wieger Hemrika¹¹, Kathrin Rösch¹², Luke T Dang¹³, David Baker¹³, Melanie Ott¹², Philippe Depeille⁸, Sean M Wu¹⁴, Jarno Drost¹⁰, Roeland Nusse¹⁵, Jeroen P Roose⁹, Jacob Piehler⁵, Sylvia F Boj⁷, Claudia Y Janda¹⁶, Hans Clevers¹⁷, Calvin J Kuo⁴, K Christopher Garcia¹⁸

Affiliations

¹ Department of Molecular and Cellular Physiology, Department of Structural Biology, Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, CA 94305, USA.
² Department of Medicine, Division of Hematology, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Biology, Stanford University School of Medicine, Stanford, CA 94305, USA.
³ Oncode Institute, Hubrecht Institute, University Medical Centre Utrecht, Utrecht, the Netherlands.
⁴ Department of Medicine, Division of Hematology, Stanford University School of Medicine, Stanford, CA 94305, USA.
⁵ Division of Biophysics, Department of Biology, University of Osnabrück, 49076 Osnabrück, Germany.
⁶ Howard Hughes Medical Institute, Department of Developmental Biology, Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Princess Máxima Center for Pediatric Oncology, Utrecht, the Netherlands.
⁷ Foundation Hubrecht Organoid Technology (HUB), Utrecht, the Netherlands.
⁸ Department of Cardiology, University Medical Center Utrecht & Utrecht Regenerative Medicine Center, Utrecht University, 3508 GA Utrecht, the Netherlands.
⁹ Department of Anatomy, University of California, San Francisco, San Francisco, CA, USA.
¹⁰ Oncode Institute, Princess Máxima Center for Pediatric Oncology, Heidelberglaan 25, 3584 CS Utrecht, the Netherlands.
¹¹ U-Protein Express BV, Yalelaan 62, 3584 CM Utrecht, the Netherlands.
¹² Gladstone Institutes and Department of Medicine, University of California, San Francisco, San Francisco, CA, USA.
¹³ Department of Biochemistry, Institute for Protein Design and Howard Hughes Medical Institute, University of Washington, Seattle, WA 98105, USA.
¹⁴ Division of Cardiovascular Medicine, Department of Medicine, Cardiovascular Institute and Institute of Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA.
¹⁵ Howard Hughes Medical Institute, Department of Developmental Biology, Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA.
¹⁶ Princess Máxima Center for Pediatric Oncology, Utrecht, the Netherlands.
¹⁷ Oncode Institute, Hubrecht Institute, University Medical Centre Utrecht, Utrecht, the Netherlands; Oncode Institute, Princess Máxima Center for Pediatric Oncology, Heidelberglaan 25, 3584 CS Utrecht, the Netherlands.
¹⁸ Department of Molecular and Cellular Physiology, Department of Structural Biology, Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, CA 94305, USA. Electronic address: kcgarcia@stanford.edu.

Abstract

Modulation of Wnt signaling has untapped potential in regenerative medicine due to its essential functions in stem cell homeostasis. However, Wnt lipidation and Wnt-Frizzled (Fzd) cross-reactivity have hindered translational Wnt applications. Here, we designed and engineered water-soluble, Fzd subtype-specific "next-generation surrogate" (NGS) Wnts that hetero-dimerize Fzd and Lrp6. NGS Wnt supports long-term expansion of multiple different types of organoids, including kidney, colon, hepatocyte, ovarian, and breast. NGS Wnts are superior to Wnt3a conditioned media in organoid expansion and single-cell organoid outgrowth. Administration of Fzd subtype-specific NGS Wnt in vivo reveals that adult intestinal crypt proliferation can be promoted by agonism of Fzd5 and/or Fzd8 receptors, while a broad spectrum of Fzd receptors can induce liver zonation. Thus, NGS Wnts offer a unified organoid expansion protocol and a laboratory "tool kit" for dissecting the functions of Fzd subtypes in stem cell biology.

Keywords: DARPin; Frizzled; Wnt; canonical Wnt signaling; organoids; protein engineering; regenerative medicine; stem cell; surrogate Wnt.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Frizzled Receptors*
Hepatocytes
Organoids*
Stem Cells
Wnt Signaling Pathway

Substances

Frizzled Receptors

Abstract

Publication types

MeSH terms

Substances

Grants and funding