Norrin mediates tumor-promoting and -suppressive effects in glioblastoma via Notch and Wnt
Ahmed El-Sehemy, … , Peter Dirks, Valerie A. Wallace
Ahmed El-Sehemy, … , Peter Dirks, Valerie A. Wallace
Published March 17, 2020
Citation Information: J Clin Invest. 2020;130(6):3069-3086. https://doi.org/10.1172/JCI128994.
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Research Article Oncology

Norrin mediates tumor-promoting and -suppressive effects in glioblastoma via Notch and Wnt

Abstract

Glioblastoma multiforme (GBM) contains a subpopulation of cells, GBM stem cells (GSCs), that maintain the bulk tumor and represent a key therapeutic target. Norrin is a Wnt ligand that binds Frizzled class receptor 4 (FZD4) to activate canonical Wnt signaling. Although Norrin, encoded by NDP, has a well-described role in vascular development, its function in human tumorigenesis is largely unexplored. Here, we show that NDP expression is enriched in neurological cancers, including GBM, and its levels positively correlated with survival in a GBM subtype defined by low expression of ASCL1, a proneural factor. We investigated the function of Norrin and FZD4 in GSCs and found that it mediated opposing tumor-suppressive and -promoting effects on ASCL1lo and ASCL1hi GSCs. Consistent with a potential tumor-suppressive effect of Norrin suggested by the tumor outcome data, we found that Norrin signaling through FZD4 inhibited growth in ASCL1lo GSCs. In contrast, in ASCL1hi GSCs Norrin promoted Notch signaling, independently of WNT, to promote tumor progression. Forced ASCL1 expression reversed the tumor-suppressive effects of Norrin in ASCL1lo GSCs. Our results identify Norrin as a modulator of human brain cancer progression and reveal an unanticipated Notch-mediated function of Norrin in regulating cancer stem cell biology. This study identifies an unanticipated role of Norrin in human brain cancer progression. In addition, we provide preclinical evidence suggesting Norrin and canonical Wnt signaling as potential therapeutic targets for GBM subtype–restricted cancer stem cells.

Authors

Ahmed El-Sehemy, Hayden Selvadurai, Arturo Ortin-Martinez, Neno Pokrajac, Yasin Mamatjan, Nobuhiko Tachibana, Katherine Rowland, Lilian Lee, Nicole Park, Kenneth Aldape, Peter Dirks, Valerie A. Wallace

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Figure 5

RNA-Seq analysis reveals divergent and overlapping effects of NDP knockdown on gene expression in ASCL1lo and ASCL1hi GSCs.

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RNA-Seq analysis reveals divergent and overlapping effects of NDP knockd...
(A) Venn diagram representing the number of overlapping and common differentially expressed genes in each cell line. Roughly half of G523 hits and one-third of G411 hits are overlapping. Notably, the number of hits in G411 is higher than in G523. (B) Unbiased enrichment maps of the identified hits in both lines (the blue circles represent G411, red circles represent G523). The overlapping sets predominantly center on cell cycle, cell division, and proliferation, while G411 unique sets are related to cytoskeletal rearrangement and movement. Refer to Supplemental Figure 6 for more details. (C) Venn diagram representing the number of unique and common enriched gene sets. (D) Western blot validation of CCNE2 and CCNA2 expression in G411 and G523 cells with NDP knockdown. (E) Log2(fold change) values for selected Wnt targets from the RNA-Seq library results. The selected genes were significantly differentially expressed after NDP knockdown in G411 but not G523 cells. Bars show the average fold change of selected hits for 2 shNDP (shNDP-A, shNDP-C) constructs in each cell line.