The therapeutic effect of the BRD4-degrading PROTAC A1874 in human colon cancer cells

An-Cheng Qin,Yi-Fan Chen,Xing-Sheng Lu,Shu-Sheng Wang,Yu Song,Yun Gao,Hua Jin,Li-Na Zhou

doi:10.1038/s41419-020-03015-6

Abstract

A1874 is a novel BRD4-degrading proteolysis targeting chimera (PROTAC). In primary colon cancer cells and established HCT116 cells, A1874 potently inhibited cell viability, proliferation, cell cycle progression, as well as cell migration and invasion. The BRD4-degrading PROTAC was able to induce caspase and apoptosis activation in colon cancer cells. Furthermore, A1874-induced degradation of BRD4 protein and downregulated BRD-dependent genes (c-Myc, Bcl-2, and cyclin D1) in colon cancer cells. Significantly, A1874-induced anti-colon cancer cell activity was more potent than the known BRD4 inhibitors (JQ1, CPI203, and I-BET151). In BRD4-knockout colon cancer cells A1874 remained cytotoxic, indicating the existence of BRD4-independent mechanisms. In addition to BRD4 degradation, A1874 cytotoxicity in colon cancer cells was also associated with p53 protein stabilization and reactive oxygen species production. Importantly, the antioxidant N-acetyl-cysteine and the p53 inhibitor pifithrin-α attenuated A1874-induced cell death and apoptosis in colon cancer cells. In vivo, A1874 oral administration potently inhibited colon cancer xenograft growth in severe combined immuno-deficient mice. BRD4 degradation and p53 protein elevation, as well as apoptosis induction and oxidative stress were detected in A1874-treated colon cancer tissues. Together, A1874 inhibits colon cancer cell growth through both BRD4-dependent and -independent mechanisms.

Highlights

Colon cancer has become a common malignancy and a global health issue[1,2], causing significant cancer-related human mortalities each year[3,4]
In pCan[1] cells and pCan[2] cells, A1874-induced viability (CCK-8 optical densities (ODs)) reduction (Fig. 3c), and cell apoptosis were significantly more potent than JQ1, CPI203, and I-BET726. These known inhibitors were utilized at even higher concentrations than A1874. These results suggest that A1874-induced anti-colon cancer cell activity might not be solely dependent on Bromodomain-containing protein 4 (BRD4) protein degradation
CRISPR/Cas9-induced BRD4 KO failed to promote p53 protein upregulation (Fig. 4h) and Reactive oxygen species (ROS) production (CellROX intensity, Fig. 4i) in pCan[1] cells. These results demonstrate that p53 stabilization and oxidative injury act independently of BRD4 protein degradation to participate in A1874induced anti-colon cancer cell activity

Summary

Introduction

Colon cancer has become a common malignancy and a global health issue[1,2], causing significant cancer-related human mortalities each year[3,4]. Current clinical treatments, including chemotherapy, surgery, radiation, and/or molecularly targeted therapies[1,5,6], along with the advanced early screen and diagnosis techniques, have significantly improved the prognosis and five-year overall survival of colon cancer patients[1,5,6]. The prognosis for the advanced, metastatic and recurrent colon cancer patients remains poor[1]. Bromodomain-containing protein 4 (BRD4) is an extensively studied BET (bromodomain and extraterminal domain) family protein, that provides a promising therapeutic target for colon cancer[7]. Recent studies have proposed BRD4 as a novel oncogene as it is overexpressed in colon cancer[12] and many other malignancies[8,11]

Methods

Results

Conclusion