Morin overcomes doxorubicin resistance in human breast cancer by inducing DNA damage and modulating the LKB1/AMPK/mTORC1 signaling pathway.

Breast cancer chemoresistance hampers chemotherapy efficacy; researchers investigate the pharmacological activities of natural products for potential solutions. This study aimed to determine the effect of morin, a bioflavonoid isolated from Maclura pomifera, on two Dox-resistant human breast cancer cell lines MDA-MB-231 (MDA-DR) and MCF-7 (MCF-DR). Sulforhodamine B and colony-forming assays demonstrated the cytotoxic effect of morin on both cell lines. Morin induced DNA damage and reduced the DNA repair mechanism, a feature of chemoresistance. In addition, morin reduced the protein expressions of cell cycle regulators, such as cyclin D1, CDK4, cyclin E1, cyclin B1, and p-Rb, thereby halting cell cycle progression. Moreover, morin slightly reduced PARP and Bcl-xL expressions but left LC3-II and RIPK3 expressions unchanged. Annexin-V/7-AAD analysis showed morin increased 7-AAD positive cells and annexin-V positive cells among MDA-DR and MCF-DR cells, respectively. In addition, morin increased p-AMPK and p-LKB1 levels; and, thus, inhibited phosphorylation of the mTOR pathway, but decreased t-AMPK levels by inducing lysosomal degradation, and AICAR, an AMPK activator, reduced Raptor, cyclin D1, CDK4, cyclin E1 and phosphorylated, and total mTOR levels, indicating AMPK is a key player in inducing cell death. Also, morin modulated MAPK phosphorylation and attenuated p-Akt and p-GSK3αß levels; and thus, inhibited cell survival. In addition, morin suppressed tumor growth in our MDA-DR xenografted mouse model. These findings indicate that morin is a potential treatment for Dox-resistant breast cancer and that it does so by inducing DNA damage and modulating the LKB1/AMPK/mTORC1 pathway, along with regulating the MAPK, and Akt/GSK3αß signaling pathways.

Morin Sensitizes MDA-MB-231 Triple-Negative Breast Cancer Cells to Doxorubicin Cytotoxicity by Suppressing FOXM1 and Attenuating EGFR/STAT3 Signaling Pathways.

Considerable emphasis is being placed on combinatorial chemotherapeutic/natural treatments for breast cancer. This study reveals the synergistic anti-tumor activity of morin and Doxorubicin (Dox) co-treatment on MDA-MB-231 triple-negative breast cancer (TNBC) cell proliferation. Morin/Dox treatment promoted Dox uptake and induced DNA damage and formation of nuclear foci of p-H2A.X. Furthermore, DNA repair proteins, RAD51 and survivin, and cell cycle proteins, cyclin B1 and forkhead Box M1 (FOXM1), were induced by Dox alone but attenuated by morin/Dox co-treatment. In addition, Annexin V/7-AAD analysis revealed that necrotic cell death after co-treatment and apoptotic cell death by Dox alone were associated with the induction of cleaved PARP and caspase-7 without Bcl-2 family involvement. FOXM1 inhibition by thiostrepton showed that co-treatment caused FOXM1-mediated cell death. Furthermore, co-treatment downregulated the phosphorylation of EGFR and STAT3. Flow cytometry showed that the accumulation of cells in the G2/M and S phases might be linked to cellular Dox uptake, p21 upregulation, and cyclin D1 downregulation. Taken together, our study shows that the anti-tumor effect of morin/Dox co-treatment is due to the suppression of FOXM1 and attenuation of EGFR/STAT3 signaling pathways in MDA-MB-231 TNBC cells, which suggests that morin offers a means of improving therapeutic efficacy in TNBC patients.

Cell cycle arrest-mediated cell death by morin in MDA-MB-231 triple-negative breast cancer cells.

BACKGROUND: Morin, a flavonoid extracted from Moraceace family and exhibits several pharmacological activities including anti-cancer activity. Although the anticancer activity of morin in breast cancer was estimated in some investigations, the pharmaceutical mechanism has not been fully elucidated. Therefore, we investigated to unveil the detail signaling pathway in morin-treated in MDA-MB-231 triple-negative breast cancer cells. METHODS: The cytotoxicity of morin in MDA-MB-231 cells was confirmed by sulforhodamine B (SRB) assay and colony formation assay. Flow cytometry was performed to examine the cell cycle and cell death patterns and the protein expression and phosphorylation were detected by western blotting. RESULTS: Our results showed that morin inhibited MDA-MB-231 cells proliferation in time and concentration-dependent manner. Morphological changes were observed when treated with various concentration of morin in MDA-MB-231 cells. In regard to protein expression, morin induced the phosphorylation of ERK and p-H2A.X and decreased the level of DNA repair markers, RAD51 and survivin. In addition, flow cytometry showed S and G2/M arrest by morin that was associated with the decrease in the protein expression of cyclin A2 and cyclin B1 and upregulation of p21. Interestingly, annexin V/PI staining result clearly showed that morin induced cell death without apoptosis. Furthermore, attenuated FoxM1 by morin was co-related with cell cycle regulators including p21, cyclin A2 and cyclin B1. CONCLUSION: Taken together, our study indicates that morin-induced cell death of MDA-MB-231 is caused by sustained cell cycle arrest via the induction of p21 expression by activation of ERK and repression of FOXM1 signaling pathways.