An autophagy-dependent cell death of MDA-MB-231 cells triggered by a novel Rhein derivative 4F

Abstract

Triple negative breast cancer (TNBC) has very poor prognosis and no efficacious therapeutic options due to the absence of a validated molecular target. Therefore, novel therapeutic strategies against TNBC are urgently needed. Our team synthesized and screened a series of compounds derived from Rhein, of which 4F was selected for further analysis based on its ability to produce the vacuolated appearance of cells. Using Cell counting kit-8 assay, colony-formation assay, cell apoptosis and cell cycle assay, we compared the antitumor effects of 4F, Rhein and Cisplatin on a TNBC cell line MDA-MB-231 in vitro. The vacuoles in MDA-MB-231 cells were observed and analyzed by hematoxylin–eosin staining and transmission electron microscopy. Autophagy and apoptosis-related proteins including p62, Microtubule Light Chain 3 (LC3), Beclin-1 and Caspase-3 were determined by western blot. The tandem mRFP-GFP-LC3 Lentivirus was used for monitoring the maturation step of autophagosomes. Our data revealed that 4F had lower cytotoxicity to normal breast cell line MCF-10A as compared with positive drug Doxorubicin. Although 4F had better cytotoxicity than Rhein, it had no influence on cells apoptosis in 4F-treated cells. Accumulation of autolysosomes and autophagosomes was observed in 4F-treated MDA-MB-231 cells, accompanied by increased level of Beclin-1 protein. Enhanced autophagic flux was verified by higher ratio of LC3-II/LC3-I, the degradation of p62 protein and alteration in red and green fluorescence puncta. These findings suggested that the process of MDA-MB-231 cell death induced by 4F seemed rely mainly on autophagy rather than apoptosis. 4F may be an alternative drug candidate against TNBC and merits more exploration.

Therapeutic Emergence of Rhein as a Potential Anticancer Drug: A Review of Its Molecular Targets and Anticancer Properties

“…Not only rhein, but also its derivatives showed promising effects in breast cancer. For example, a novel rhein-derived compound, 4F, was found to induce autophagy in MDA-MB-231 cells by upregulating the expression of beclin-1 and causing the degradation of p62 [148]. Furthermore, it was observed that an analog of rhein, rhein lysinate (RHL), inhibited phosphorylation of EGFR, MEK, c-Raf, and ERK and induced apoptosis in MCF-7, SK-Br-3, and MDA-MB-231 cells [46].…”

Autophagy-dependent cell cycle arrest in esophageal cancer cells exposed to dihydroartemisinin

“…Autophagic cell death is one of the major mechanisms that induced programmed cell death. It was found that autophagic cell death played an important role in anticancer drugs [20,21]. DHA could induce autophagy in some human cancer cell lines, including esophageal cancer cells [22][23][24], while the precise mechanisms of DHA on cancer cells were still limited.…”

Rhein Derivative 4F Inhibits the Malignant Phenotype of Breast Cancer by Downregulating Rac1 Protein

“…Furthermore, its inhibition on the proliferation and growth of breast cancer cells was also much stronger than that of Rhein and control, and greatly prolonging the doubling time of cancer cells. Our previous study also proved that derivative 4F is far less cytotoxic to normal breast cells than doxorubicin (Yunfeng et al, 2019b). This indicates that the side chain of Rhein was successfully modified and that derivative 4F is may be a selective anthracycline candidate with low toxicity.…”

“…Moreover, its toxicity to human triple-negative breast cancer MDA-MB-231 cells in this concentration range was time-and dose-dependent ( Figure 2B). The IC 50 values of derivative 4F and Rhein were (12.80 ± 0.83) and (163.96 ± 33.36) mmol/L in MDA-MB-231 cells and (7.54 ± 1.25) mmol/L and (120.19 ± 10.98) mmol/L in MCF-7 cells (Figur2C, G, H) (Yunfeng et al, 2019a;Yunfeng et al, 2019b). The IC 50 of derivative 4F was much lower than that of the lead compound Rhein.…”