High-Throughput Drug Screening Identifies a Potent Wnt Inhibitor that Promotes Airway Basal Stem Cell Homeostasis

Abstract

Graphical Abstract Highlights d A dysregulated Wnt/b-catenin axis in human lung premalignancy can be modeled in vitro d High-throughput drug screening identifies the Wnt/b-catenin inhibitor WIC1 d Low-dose treatment with WIC1 promotes differentiation to the ciliated cell fate d WIC1 acts to repress phosphorylation of b-catenin Y489 and Tp63 expression SUMMARYMechanisms underpinning airway epithelial homeostatic maintenance and ways to prevent its dysregulation remain elusive. Herein, we identify that b-catenin phosphorylated at Y489 (p-b-catenin Y489 ) emerges during human squamous lung cancer progression. This led us to develop a model of airway basal stem cell (ABSC) hyperproliferation by driving Wnt/b-catenin signaling, resulting in a morphology that resembles premalignant lesions and loss of ciliated cell differentiation. To identify small molecules that could reverse this process, we performed a high-throughput drug screen for inhibitors of Wnt/ b-catenin signaling. Our studies unveil Wnt inhibitor compound 1 (WIC1), which suppresses T-cell factor/lymphoid enhancer-binding factor (TCF/LEF) activity, reduces ABSC proliferation, induces ciliated cell differentiation, and decreases nuclear p-b-catenin Y489 . Collectively, our work elucidates a dysregulated Wnt/p-b-catenin Y489 axis in lung premalignancy that can be modeled in vitro and identifies a Wnt/ b-catenin inhibitor that promotes airway homeostasis. WIC1 may therefore serve as a tool compound in regenerative medicine studies with implications for restoring normal airway homeostasis after injury.

Wnt signaling in lung development, regeneration, and disease progression

“…Haas et al demonstrated that high Wnt signaling triggers transcription of Tp63 , which then acts as a master regulator of basal cell stemness 66 . A recently identified novel Wnt signaling inhibitor can promote restoration of Wnt-induced dysregulation of proximal airway homeostasis via p-β-catenin Y489 /TP63-related mechanisms 65 (Fig. 3b ), further implicating the importance of the pathway in proximal airway homeostasis.…”

“…While Wnt signaling promotes ciliated cell differentiation, this is not true at all doses of pathway activation 65 . In contrast, high levels of Wnt signaling drive ABSC hyperproliferation concomitant with a phosphorylated form of nuclear β-catenin at Y489 (p-β-catenin Y489 ) and abolishment of differentiation to the ciliated cell fate 65 (Fig. 3b ).…”

“…b In the proximal airway ABSCs employ high levels of Wnt signaling to facilitate proliferation and medium levels of Wnt signaling to promote differentiation to the ciliated cell fate. At the molecular level, this appears to be accomplished by nuclear localization of p-β-catenin Y489 65 . Further, Tp63 is regulated by upstream Wnt signaling to promote basal cell stemness, thereby inhibiting differentiation to ciliated cells 66 .…”

“…Recent work has identified the presence of Wnt-producing (Porcupine+) epithelial cells that are closely apposed to Wnt-responding (LGR5+) epithelial cells in LADC 124 . Human premalignant lesions and squamous lung cancer samples also display increased stromal and epithelial Porcupine as well as nuclear p-β-catenin Y489 65 , further elucidating the Wnt signaling niche networks that are targetable with small molecule inhibitors.…”

The Role of β-Catenin in Th1 Immune Response against Tuberculosis and Profiles of Expression in Patients with Pulmonary Tuberculosis

“…β -Catenin has been thought to be involved in the regulation of cell fate, such as proliferation, differentiation, and apoptosis or T cell development [ 16 , 17 ]. In most published studies, β -catenin has a close association with the pathogenesis of malignant diseases and is a therapeutic target for cancer in view of its function to promote tumor cell proliferation [ 18 ].…”