Optimizations of an ELISA-like high-throughput screening assay for the discovery of β-catenin/TCF4 interaction antagonists.

Zhenghao FU; Gangan Yan; Xiaohong Zhu; Xiaoping Liu; Yunyu Chen

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Optimizations of an ELISA-like high-throughput screening assay for the discovery of β-catenin/TCF4 interaction antagonists.

Author: Zhenghao FU ¹ ; Gangan YAN ¹ ; Xiaohong ZHU ² ; Xiaoping LIU ¹ ; Yunyu CHEN ¹
Author Information

1. Institute for Drug Screening and Evaluation, Wannan Medical College, Wuhu 241002, Anhui, China.
2. Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China.
Publication Type:Journal Article
Keywords: Wnt inhibitor; enzyme-linked immunosorbent assay; high-throughput screening; plumbagin; sanguinarine; β-catenin/TCF4 interaction
MeSH: Carcinoma, Non-Small-Cell Lung; Cell Line, Tumor; Enzyme-Linked Immunosorbent Assay; High-Throughput Screening Assays; Humans; Lung Neoplasms; Transcription Factor 4/genetics*; beta Catenin/genetics*
From: Chinese Journal of Biotechnology 2021;37(8):2878-2889
CountryChina
Language:Chinese
Abstract: In canonical Wnt/β-catenin signaling pathway, β-catenin/TCF4 (T-cell factor 4) interaction plays an important role in the pathogenesis and development of non-small cell lung cancer (NSCLC), and it is tightly associated with the proliferation, chemoresistance, recurrence and metastasis of NSCLC. Therefore, suppressing β-catenin/TCF4 interaction in Wnt/β-catenin signaling pathway would be a new therapeutic avenue against NSCLC metastasis. In this study, considering the principle of enzyme-linked immunosorbent assay (ELISA), an optimized high-throughput screening (HTS) assay was developed for the discovery of β-catenin/TCF4 interaction antagonists. Subsequently, this ELISA-like screening assay was performed using 2 μg/mL GST-TCF4 βBD and 0.5 μg/mL β-catenin, then a high Z' factor of 0.83 was achieved. A pilot screening of a natural product library using this ELISA-like screening assay identified plumbagin as a potential β-catenin/TCF4 interaction antagonist. Plumbagin remarkably inhibited the proliferation of A549, H1299, MCF7 and SW480 cell lines. More importantly, plumbagin significantly suppressed the β-catenin-responsive transcription in TOPFlash assay. In short, this newly developed ELISA-like screening assay will be vital for the rapid screening of novel Wnt inhibitors targeting β-catenin/TCF4 interaction, and this interaction is a potential anticancer target of plumbagin in vitro.