Research Paper Volume 16, Issue 6 pp 5526—5544

Cinobufotalin regulates the USP36/c-Myc axis to suppress malignant phenotypes of colon cancer cells in vitro and in vivo

Yongjun Hu1, , Ming Luo1, ,

  • 1 Department of General Surgery, The Second Xiangya Hospital, Central South University, Changsha 410011, China

Received: July 18, 2023       Accepted: January 4, 2024       Published: March 15, 2024
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Copyright: © 2024 Hu and Luo. This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.


Ubiquitin-specific protease 36 (USP36) has been reported to exhibit oncogenic effects in various malignancies, but the function of USP36 in colon cancer progression remains indefinite. Herein, we aimed to determine the role and mechanism of USP36 in malignant phenotypes of colon cancer cells and explore the potential drug targeting USP36. Bioinformatics analyses indicated that USP36 is highly expressed and significantly related to tumor stages in colon cancer. Besides, USP36 was further up-regulated in oxaliplatin (Oxa)-resistant colon cancer cells. Colony formation, Edu staining, Transwell, wound healing, sphere formation, and CCK-8 assays were conducted and showed that the proliferation, Oxa-resistance, migration, stemness, and invasion of HCT116 cells were promoted after overexpressing USP36, while suppressed by USP36 knockdown. Mechanically, USP36 enhances c-Myc protein stabilization in HCT116 cells via deubiquitination. AutoDock tool and ubiquitin-AMC hydrolysis assay identified cinobufotalin (CBF), an anti-tumor drug, maybe a USP36 inhibitor by inhibiting its deubiquitination activity. CBF significantly prohibited proliferation, migration, invasion, and stemness of HCT116 cells and reversed Oxa-resistance, whereas enforced expression of USP36 blocked these effects. Moreover, in vivo analyses confirmed the oncogenic role of USP36 and the therapeutic potential of CBF in the malignancy of colon cancer. In conclusion, CBF may be a promising therapeutic agent for colon cancer due to its regulation of the USP36/c-Myc axis.


USP36: ubiquitin-specific protease 36; CBF: cinobufotalin; Oxa: oxaliplatin; PTMs: post-translational modifications; DUB: deubiquitinating enzyme; USP: ubiquitin-specific proteases; OS: overall survival; DFS: disease-free survival; Co-IP: co-immunoprecipitation; CHX: cycloheximide; Ub: ubiquitin; TIC: tumor-initiating cell; ELDA: Extreme Limiting Dilution Analysis; CSCs: cancer stem cells.