Targeting BET Proteins With a PROTAC Molecule Elicits Potent Anticancer Activity in HCC Cells

Author(s): Huapeng Zhang ¹ ^, ² ^, ³ ^, ⁴ , Gongquan Li ¹ ^, ² ^, ³ ^, ⁴ , Yi Zhang ² ^, ⁵ , Jihua Shi ¹ ^, ² ^, ³ ^, ⁴ , Bing Yan ¹ ^, ² ^, ³ ^, ⁴ , Hongwei Tang ¹ ^, ² ^, ³ ^, ⁴ , Sanyang Chen ¹ ^, ² ^, ³ ^, ⁴ , Jiakai Zhang ¹ ^, ² ^, ³ ^, ⁴ , Peihao Wen ¹ ^, ² ^, ³ ^, ⁴ , Zhihui Wang ¹ ^, ² ^, ³ ^, ⁴ , Chun Pang ¹ ^, ² ^, ³ ^, ⁴ , Jie Li ¹ ^, ² ^, ³ ^, ⁴ , Wenzhi Guo ¹ ^, ² ^, ³ ^, ⁴ , Shuijun Zhang ¹ ^, ² ^, ³ ^, ⁴ ^, ^*

Publication date (Electronic): 14 January 2020

Journal: Frontiers in Oncology

Publisher: Frontiers Media S.A.

Keywords: BET, PROTAC, protein degradation, HCC, apoptosis

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Abstract

Background and Aim: Bromodomain and extraterminal domain (BET) family proteins are epigenetic regulators involved in human malignances. Targeting BET proteins for degradation using proteolysis-targeting chimera (PROTAC) recently has drawn increasing attention in the field of cancer therapeutics. BET proteins have been found to be overexpressed in HCC cells and tumor tissues. However, the biological activity of BET-PROTACs in hepatocellular carcinoma (HCC) remains unclear. In this study, we investigated anti-HCC activity of BETd-260, a BET-PROTAC molecule using in vitro and in vivo models.

Methods: BETd-260-mediated anti-HCC activity was investigated by cell viability, apoptosis assays. Efficacy was examined with a cell lines-derived HCC xenograft model in mice. Anticancer mechanism was investigated by RT-PCR, western blotting and immunohistochemical staining.

Results: BETd-260 potently suppressed cell viability and robustly induced apoptosis in HCC cells. BETd-260 reciprocally modulated the expression of several apoptotic genes in HCC cells, i.e., suppressing the expression of anti-apoptotic Mcl-1, Bcl-2, c-Myc, and X-linked inhibitor of apoptosis (XIAP), whereas increasing the expression of pro-apoptotic Bad. BETd-260 treatment led to disruption of mitochondrial membrane integrity, and triggered apoptosis via intrinsic signaling in HCC cells. BETd-260 triggered apoptosis in HCC xenograft tissue and profoundly inhibited the growth of HCC xenograft tumors in mice.

Conclusion: Our data suggest that pharmacological targeting of BET for degradation may be a novel therapeutic strategy for the treatment of HCC.

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Most cited references 25

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Protacs: chimeric molecules that target proteins to the Skp1-Cullin-F box complex for ubiquitination and degradation.

K M Sakamoto, K. -B. Kim, A Kumagai … (2001)

The intracellular levels of many proteins are regulated by ubiquitin-dependent proteolysis. One of the best-characterized enzymes that catalyzes the attachment of ubiquitin to proteins is a ubiquitin ligase complex, Skp1-Cullin-F box complex containing Hrt1 (SCF). We sought to artificially target a protein to the SCF complex for ubiquitination and degradation. To this end, we tested methionine aminopeptidase-2 (MetAP-2), which covalently binds the angiogenesis inhibitor ovalicin. A chimeric compound, protein-targeting chimeric molecule 1 (Protac-1), was synthesized to recruit MetAP-2 to SCF. One domain of Protac-1 contains the I kappa B alpha phosphopeptide that is recognized by the F-box protein beta-TRCP, whereas the other domain is composed of ovalicin. We show that MetAP-2 can be tethered to SCF(beta-TRCP), ubiquitinated, and degraded in a Protac-1-dependent manner. In the future, this approach may be useful for conditional inactivation of proteins, and for targeting disease-causing proteins for destruction.

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Hijacking the E3 Ubiquitin Ligase Cereblon to Efficiently Target BRD4.

Jing Lu, Yimin Qian, Martha Altieri … (2015)

BRD4, a bromodomain and extraterminal domain (BET) family member, is an attractive target in multiple pathological settings, particularly cancer. While BRD4 inhibitors have shown some promise in MYC-driven malignancies such as Burkitt's lymphoma (BL), we show that BRD4 inhibitors lead to robust BRD4 protein accumulation, which may account for their limited suppression of MYC expression, modest antiproliferative activity, and lack of apoptotic induction. To address these limitations we designed ARV-825, a hetero-bifunctional PROTAC (Proteolysis Targeting Chimera) that recruits BRD4 to the E3 ubiquitin ligase cereblon, leading to fast, efficient, and prolonged degradation of BRD4 in all BL cell lines tested. Consequently, ARV-825 more effectively suppresses c-MYC levels and downstream signaling than small-molecule BRD4 inhibitors, resulting in more effective cell proliferation inhibition and apoptosis induction in BL. Our findings provide strong evidence that cereblon-based PROTACs provide a better and more efficient strategy in targeting BRD4 than traditional small-molecule inhibitors.

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The BH3 mimetic ABT-737 targets selective Bcl-2 proteins and efficiently induces apoptosis via Bak/Bax if Mcl-1 is neutralized.

Mark van Delft, Andrew Wei, Kylie D. Mason … (2006)

Since apoptosis is impaired in malignant cells overexpressing prosurvival Bcl-2 proteins, drugs mimicking their natural antagonists, BH3-only proteins, might overcome chemoresistance. Of seven putative BH3 mimetics tested, only ABT-737 triggered Bax/Bak-mediated apoptosis. Despite its high affinity for Bcl-2, Bcl-x(L), and Bcl-w, many cell types proved refractory to ABT-737. We show that this resistance reflects ABT-737's inability to target another prosurvival relative, Mcl-1. Downregulation of Mcl-1 by several strategies conferred sensitivity to ABT-737. Furthermore, enforced Mcl-1 expression in a mouse lymphoma model conferred resistance. In contrast, cells overexpressing Bcl-2 remained highly sensitive to ABT-737. Hence, ABT-737 should prove efficacious in tumors with low Mcl-1 levels, or when combined with agents that inactivate Mcl-1, even to treat those tumors that overexpress Bcl-2.

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Author and article information

Contributors

Wenzhi Guo: URI : http://loop.frontiersin.org/people/834613/overview

Shuijun Zhang: URI : http://loop.frontiersin.org/people/791513/overview

Journal

Journal ID (nlm-ta): Front Oncol

Journal ID (iso-abbrev): Front Oncol

Journal ID (publisher-id): Front. Oncol.

Title: Frontiers in Oncology

Publisher: Frontiers Media S.A.

ISSN (Electronic): 2234-943X

Publication date (Electronic): 14 January 2020

Publication date Collection: 2019

Volume: 9

Electronic Location Identifier: 1471

Affiliations

[1] ¹Department of Hepatobiliary and Pancreatic Surgery, The First Affiliated Hospital of Zhengzhou University , Zhengzhou, China

[2] ²Open and Key Laboratory of Hepatobiliary & Pancreatic Surgery and Digestive Organ Transplantation at Henan Universities, The First Affiliated Hospital of Zhengzhou University , Zhengzhou, China

[3] ³Zhengzhou Key Laboratory of Hepatobiliary & Pancreatic Diseases and Organ Transplantation , Zhengzhou, China

[4] ⁴Henan Key Laboratory of Digestive Organ Transplantation , Zhengzhou, China

[5] ⁵Department of Orthopaedic Surgery, The First Affiliated Hospital of Zhengzhou University , Zhengzhou, China

Author notes

Edited by: Qingxin Mu, University of Washington, United States

Reviewed by: Sida Liao, TScan Therapeutics, Inc., United States; Ning Wei, University of Pittsburgh, United States

*Correspondence: Shuijun Zhang zhangshuijun@ 123456zzu.edu.cn

This article was submitted to Cancer Molecular Targets and Therapeutics, a section of the journal Frontiers in Oncology

†These authors have contributed equally to this work

Article

DOI: 10.3389/fonc.2019.01471

PMC ID: 6971110

PubMed ID: 31993368

SO-VID: 1e2edc78-f868-4d46-a32f-be5393e65662

License:

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

History

Date received : 22 August 2019

Date accepted : 09 December 2019

Page count

Figures: 6, Tables: 0, Equations: 0, References: 33, Pages: 11, Words: 6322

Funding

Funded by: Natural Science Foundation of Henan Province 10.13039/501100006407

Funded by: Key Scientific Research Project of Colleges and Universities in Henan Province 10.13039/501100013066

Targeting BET Proteins With a PROTAC Molecule Elicits Potent Anticancer Activity in HCC Cells

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Most cited references 25

Protacs: chimeric molecules that target proteins to the Skp1-Cullin-F box complex for ubiquitination and degradation.

Hijacking the E3 Ubiquitin Ligase Cereblon to Efficiently Target BRD4.

The BH3 mimetic ABT-737 targets selective Bcl-2 proteins and efficiently induces apoptosis via Bak/Bax if Mcl-1 is neutralized.

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