The essential role of YAP in ERα36-mediated proliferation and the epithelial-mesenchymal transition in MCF-7 breast cancer cells.

Purpose: Most breast cancers are hormone-receptor-positive, and thus the first-line therapy for them is an anti-estrogen medication such as tamoxifen. If metastasis occurs or resistance to tamoxifen develops, the 5-year survival rates for breast cancer patients significantly decrease. Hence, a better understanding of the molecular mechanisms that contribute to breast cancer aggressiveness is of great importance. ERα36 is an estrogen receptor variant that is known to be upregulated in breast cancer patients receiving tamoxifen treatment or in triple-negative breast cancer cells. However, the specific molecular mechanism underlying ERα36-induced tamoxifen-resistance is not yet fully understood. Methods: ERα36-overexpressing MCF-7 cells were constructed by either plasmid transfection using ERα36 vector or retroviral infection using ERα36-V5-His vector. Target-gene expression was assessed by Western blot analysis and real-time PCR, and YAP activation was evaluated by luciferase assays and immunofluorescence. Cell proliferation and formation of three-dimensional spheroids were evaluated using the IncuCyte S3 Live Cell Analysis System. Results: We found that the expression patterns of Hippo signaling-related genes were significantly changed in ERα36-overexpressing MCF-7 cells compared to MCF-7 cells, which were also similarly observed in tamoxifen-resistant MCF-7 cells. Specifically, the protein expression level and activity of YAP, the core downstream protein of the Hippo pathway, were significantly increased in ERα36-overexpressing MCF-7 cells compared with MCF-7 cells. The aggressive phenotypes acquired by ERα36 overexpression in MCF-7 cells were destroyed by YAP knockout. On this basis, we propose that ERα36 regulates YAP activity by a new mechanism involving Src kinase. Conclusion: Our results suggest that YAP targeting may be a new therapeutic approach to the treatment of advanced breast cancers overexpressing ERα36.

Targeting Cdc20 for cancer therapy.

The Anaphase-Promoting Complex/Cyclosome (APC/C), an E3 ubiquitin ligase, and two co-activators, Cdc20 and Cdh1, enable the ubiquitin-dependent proteasomal degradation of various critical cell cycle regulators and govern cell division in a timely and precise manner. Dysregulated cell cycle events cause uncontrolled cell proliferation, leading to tumorigenesis. Studies have shown that Cdh1 has tumor suppressive activities while Cdc20 has an oncogenic property, suggesting that Cdc20 is an emerging therapeutic target for cancer treatment. Therefore, in this review, we discussed recent findings about the essential roles of APC/C-Cdc20 in cell cycle regulation. Furthermore, we briefly summarized that the regulation of Cdc20 expression levels is strictly controlled to order cell cycle events appropriately. Finally, given the function of Cdc20 as an oncogene, therapeutic interventions targeting Cdc20 activity may be beneficial in cancer treatment.

The Ubiquitin System: An Emerging Therapeutic Target for Lung Cancer.

The ubiquitin system, present in all eukaryotes, contributes to regulating multiple types of cellular protein processes such as cell signaling, cell cycle, and receptor trafficking, and it affects the immune response. In most types of cancer, unusual events in ubiquitin-mediated signaling pathway modulation can lead to a variety of clinical outcomes, including tumor formation and metastasis. Similarly, ubiquitination acts as a core component, which contributes to the alteration of cell signaling activity, dictating biosignal turnover and protein fates. As lung cancer acquires the most commonly mutated proteins, changes in the ubiquitination of the proteins contribute to the development of lung cancer. Various inhibitors targeting the ubiquitin system have been developed for clinical applications in lung cancer treatment. In this review, we summarize the current research advances in therapeutics for lung cancer by targeting the ubiquitin system.

Ubiquitin-Conjugating Enzymes in Cancer.

The ubiquitin-mediated degradation system is responsible for controlling various tumor-promoting processes, including DNA repair, cell cycle arrest, cell proliferation, apoptosis, angiogenesis, migration and invasion, metastasis, and drug resistance. The conjugation of ubiquitin to a target protein is mediated sequentially by the E1 (activating)‒E2 (conjugating)‒E3 (ligating) enzyme cascade. Thus, E2 enzymes act as the central players in the ubiquitination system, modulating various pathophysiological processes in the tumor microenvironment. In this review, we summarize the types and functions of E2s in various types of cancer and discuss the possibility of E2s as targets of anticancer therapeutic strategies.

Essential Role of Polo-like Kinase 1 (Plk1) Oncogene in Tumor Growth and Metastasis of Tamoxifen-Resistant Breast Cancer.

The most common therapy for estrogen receptor-positive breast cancer is antihormone therapy, such as tamoxifen. However, acquisition of resistance to tamoxifen in one third of patients presents a serious clinical problem. Polo-like kinase 1 (Plk1) is a key oncogenic regulator of completion of G2-M phase of the cell cycle. We assessed Plk1 expression in five chemoresistant cancer cell types and found that Plk1 and its downstream phosphatase Cdc25c were selectively overexpressed in tamoxifen-resistant MCF-7 (TAMR-MCF-7) breast cancer cells. Real-time monitoring of cell proliferation also showed that TAMR-MCF-7 cells were more sensitive to inhibition of cell proliferation by the ATP-competitive Plk1 inhibitor BI2536 than were the parent MCF-7 cells. Moreover, BI2536 suppressed expression of epithelial-mesenchymal transition marker proteins and 3D spheroid formation in TAMR-MCF-7 cells. Using TAMR-MCF-7 cell-implanted xenograft and spleen-liver metastasis models, we showed that BI2536 inhibited tumor growth and metastasis in vivo Our results suggest that Plk1 could be a novel target for the treatment of tamoxifen-resistant breast cancer. Mol Cancer Ther; 17(4); 825-37. ©2018 AACR.

Essential role of Notch4/STAT3 signaling in epithelial-mesenchymal transition of tamoxifen-resistant human breast cancer.

We previously demonstrated that tamoxifen (TAM)-resistant human breast cancer (TAMR-MCF-7) cells showed increased expression of mesenchymal marker proteins compared to the parent MCF-7 cells. Notch is functionally important in the promotion of epithelial-mesenchymal transition (EMT) during both development and tumor progression. Notch1 and Notch4 have been reported as prognostic markers in human breast cancer. Here, we indicated that Notch4, but not Notch1, plays a critical role in the regulation of EMT signaling in TAMR-MCF-7 cells. Notch4 suppression by either Notch inhibitors or Notch4 siRNA attenuated EMT signaling. Tyrosine-phosphorylated STAT3 protein is known as a crucial signaling molecule in the regulation of tumorigenesis and metastasis. We found that TAMR-MCF-7 cells exhibited constitutive STAT3 phosphorylation, and Notch inhibition reduced the level of activated STAT3 in TAMR-MCF-7 cells. An intrasplenic injection model of liver metastases was performed using TAMR-MCF-7 cells. Mice injected with N-[N-(3,5-Difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester (DAPT, 10 mg/kg) formed smaller splenic tumors and showed a reduced micrometastatic tumor burden in their livers compared with the control group treated with vehicle. To conclude, Notch4 could be a potential target to prevent metastasis in TAM-resistant breast cancer.