Pharmacologic Wnt Inhibition Reduces Proliferation, Survival, and Clonogenicity of Glioblastoma Cells

Abstract

Wingless (WNT) signaling has been shown to be an important pathway in gliomagenesis and in the growth of stem-like glioma cells. Using immunohistochemistry to assess translocation of β-catenin protein, we identified intranuclear staining, which suggest WNT pathway activation, in 8 of 43 (19%) adult and 9 of 30 (30%) pediatric glioblastoma patient surgical samples. WNT activity, evidenced by nuclear β-catenin in our cohort and high expression of its target AXIN2 in published glioma datasets, was associated with shorter patient survival, although this was not statistically significant. We determined the effects of the porcupine inhibitor LGK974 in 3 glioblastoma cell lines with elevated AXIN2 and found that it reduced WNT pathway activity by 50% or more as assessed by T cell factor-luciferase reporters. WNT inhibition led to suppression of growth and proliferation in the cultures and a modest induction of cell death. LGK974 reduced NANOG mRNA levels and the fraction of cells expressing the stem cell marker CD133 in neurosphere cultures, induced glial differentiation, and suppressed clonogenicity. These data indicate that LGK974 represents a promising new agent that can inhibit the canonical WNT pathway in vitro, slow tumor growth and deplete stem-like clonogenic cells, thereby providing further support for targeting WNT in patients with glioblastoma.

Alterations in cellular metabolome after pharmacological inhibition of Notch in glioblastoma cells

“…A number of groups have shown that WNT signaling blockade depletes stem‐like glioma cells . We have recently shown that the porcupine inhibitor LGK974, which targets WNT signaling by modulating ligand processing, can also inhibit the pathway and reduce survival in glioma neurospheres . For these LGK974 studies, LN229 cells were used instead of GBM10, as LN229 is characterized by higher WNT activity similar to that seen in the upper range of snap‐frozen GBM specimens .…”

“…Luminescence was read at an emission wavelength of 490 nm on the Infinite M1000Pro plate reader (Tecan) and normalized to beta‐galactosidase activity. A robust IC 50 was achieved at 5 µM LGK974 at 48 hr as defined by previous experiments …”

“…28 We have recently shown that the porcupine inhibitor LGK974, which targets WNT signaling by modulating ligand processing, can also inhibit the pathway and reduce survival in glioma neurospheres. 26 For these LGK974 studies, LN229 cells were used instead of GBM10, as LN229 is characterized by higher WNT activity similar to that seen in the upper range of snap-frozen GBM specimens. 26 Inhibition of the pathway after LGK974 administration was confirmed by reduction in luciferase WNT reporter signals, with no corresponding effects on glutamate levels (Fig.…”

WNT vampirization by glioblastoma leads to tumor growth and neurodegeneration

“…GB is characterized by the deregulation of many key signaling pathways involving growth, proliferation, survival, and apoptosis, such as p53, pRB, EGFR, PI3K, STAT3 or WNT 32-33 . WNT signaling has long been suggested as a hallmark in gliomagenesis associated with the proliferation of stem-like cells in human GBs 34 . WNT signaling promotes tumoral cell proliferation and dissemination as well as resistance to chemo and radiotherapy (reviewed in 15,35 ).…”

“…It has been identified in 19% of surgical samples from adult GB patients and in 30% of surgical samples from pediatric patients. Moreover, WNT inhibition leads to suppression of tumor growth, proliferation in cultures, and a modest induction of cell death 34 .…”