American Association for Cancer Research
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Supplementary Figure S1-S3 from Mcl-1 Degradation Is Required for Targeted Therapeutics to Eradicate Colon Cancer Cells

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journal contribution
posted on 2023-03-31, 01:08 authored by Jingshan Tong, Peng Wang, Shuai Tan, Dongshi Chen, Zaneta Nikolovska-Coleska, Fangdong Zou, Jian Yu, Lin Zhang

Figure S1. Regorafenib and sorafenib did not affect Mcl-1 mRNA expression but induced GSK3β-dependent Mcl-1 degradation. A transfected quadruple mutant in 4 putative phosphorylation sites (S121A/E125A/S159A/T163A) of Mcl-1 was resistant to regorafenib-induced ubiquitination and degradation. Figure S2. Apoptosis induced by regorafenib or sorafenib was suppressed in HCT116 cells with Mcl-1 phosphorylation site mutant knock-in (Mcl-1-KI), inhibition of GSK3β, or knockout of BAX. Figure S3. Mcl-1 degradation and PUMA induction did not affect each other in regorafenib-induced apoptosis. Regorafenib-induced apoptosis in Mcl-1-KI cells could not be restored by the Bcl-2/Bcl-XL inhibitors ABT-737 or ABT-263.



National Natural Science Foundation of China



The Bcl-2 family protein Mcl-1 is often degraded in cancer cells subjected to effective therapeutic treatment, and defective Mcl-1 degradation has been associated with intrinsic and acquired drug resistance. However, a causal relationship between Mcl-1 degradation and anticancer drug responses has not been directly established, especially in solid tumor cells where Mcl-1 inhibition alone is insufficient to trigger cell death. In this study, we present evidence that Mcl-1 participates directly in determining effective therapeutic responses in colon cancer cells. In this setting, Mcl-1 degradation was induced by a variety of multikinase inhibitor drugs, where it relied upon GSK3β phosphorylation and FBW7-dependent ubiquitination. Specific blockade by genetic knock-in (KI) abolished apoptotic responses and conferred resistance to kinase inhibitors. Mcl-1-KI also suppressed the antiangiogenic and anti-hypoxic effects of kinase inhibitors in the tumor microenvironment. Interestingly, these same inhibitors also induced the BH3-only Bcl-2 family protein PUMA, which is required for apoptosis. Degradation-resistant Mcl-1 bound and sequestered PUMA from other prosurvival proteins to maintain cell survival, which was abolished by small-molecule Mcl-1 inhibitors. Our findings establish a pivotal role for Mcl-1 degradation in the response of colon cancer cells to targeted therapeutics, and they provide a useful rational platform to develop Mcl-1–targeting agents that can overcome drug resistance. Cancer Res; 77(9); 2512–21. ©2017 AACR.