Abstract A46: Hsp90 regulates Twist1 expression through STAT3 to induce epithelial-mesenchymal transition in ovarian cancer
Authors: Chong Kay Yi , Garofalo Francesca , Madarikan Oluwagbemisola , Pitruzzello Nicholas , Tsai Cheng-Hsiu , Bingham Jamie , Yang-Hartwich Yang 
Published: August 2018

Background: Metastatic disease is the leading cause of death from ovarian cancer and its underlying mechanisms are poorly understood. Twist1 is a key driver of epithelial-mesenchymal transition (EMT) and metastasis. Understanding the function and regulation of Twist1 is a vital step in the development of effective treatments for metastatic ovarian cancer. Heat shock protein 90 (Hsp90) is a molecular chaperone that modulates multiple signaling networks, and recent studies have highlighted the roles of extracellular Hsp90 in promoting metastasis in cancer. The roles of Hsp90 in regulating intracellular pathways leading to EMT and metastasis remain largely unknown.

Objective: In our study, we tested the hypothesis that Hsp90 promotes EMT in ovarian cancer through the regulation of Twist1 at the transcriptional level.

Methods: We treated A2780 and SKOV3 with a Hsp90-specific inhibitor, 17-allylamino-17 demethoxygeldanamycin (17-AAG). The effects of Hsp90 inhibition on Twist1 mRNA expression and promoter activity were measured using quantitative PCR and luciferase reporter assays, respectively. Proximity ligation assays were performed to visualize the effects of 17-AAG on the interaction between Hsp90 and transcription factors, followed by chromatin immunoprecipitation to measure the binding of transcription factors to the Twist1 promoter.

Results: Treatment with 17-AAG significantly downregulated Twist1 expression at the mRNA level in A2780, SKOV3, and three ovarian cancer patient-derived cell lines. Hsp90 overexpression substantially induced Twist1 promoter activity while treatment with 17-AAG significantly decreased the activity. Western blotting and immunofluorescent staining revealed the presence of 4 transcription factors known to be clients of Hsp90 and regulators of Twist1, which are β-catenin, signal transducer and activator of transcription 3 (STAT3), hypoxia-inducible factor 1-alpha (HIF-1α), and HIF-1β. We identified that Hsp90 interacts with β-catenin, STAT3, and HIF-1α in our cell lines. We observed that 17-AAG treatment dramatically impaired Hsp90-STAT3 interaction and the binding of STAT3 to the Twist1 promoter. Inhibition of Hsp90 was also shown to block interleukin-6 (IL-6) and transforming growth factor beta (TGF-β)-induced EMT.

Conclusion: Taken together, our findings reveal that STAT3 is dependent on Hsp90 to activate Twist1 expression. Hsp90 plays a critical role in enabling STAT3 to bind to the Twist1 promoter and promote Twist1 transcription leading to EMT. We uncovered a previously unrecognized role of Hsp90, which cooperates with STAT3 in the transcriptional regulation of Twist1 in ovarian cancer cells. Inhibiting Hsp90 using small-molecule inhibitors such as 17-AAG may have potential as a therapeutic strategy to prevent EMT and metastasis in ovarian cancer.

Citation Format: Kay Yi Chong, Francesca Garofalo, Oluwagbemisola Madarikan, Nicholas Pitruzzello, Cheng-Hsiu Tsai, Jamie Bingham, Yang Yang-Hartwich. Hsp90 regulates Twist1 expression through STAT3 to induce epithelial-mesenchymal transition in ovarian cancer. [abstract]. In: Proceedings of the AACR Conference: Addressing Critical Questions in Ovarian Cancer Research and Treatment; Oct 1-4, 2017; Pittsburgh, PA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(15_Suppl):Abstract nr A46.