Abstract 3314: Targeting Hsp90 affects stem cell signaling in triple negative breast cancer

Abstract

Abstract Background: Triple Negative Breast Cancer (TNBC) exhibit dismal survival rates due to their propensity to develop distant metastases. Heat shock protein 90 (Hsp90) is a molecular chaperone that aids in the folding and maturation of various proteins involved in breast cancer progression and resistance to therapy. Many studies have also suggested that breast cancer's ability to proliferate, progress, and spread is due to the presence of a rare subpopulation of cancer stem cells. The aim of this study is to investigate the efficacy of HSP90 inhibitors celastrol and triptolide (from the Chinese herb “Thunder God of Vine” (Tripterygium wilfordii) on stem cells as a novel therapy for TNBC. Methods: We performed in vitro studies using the TNBC cell lines BT20 and MDA-MB-231. We performed mammosphere assays to determine self-renewal capacity. For in vivo, we injected BT20 cells into flanks of athymic nude mice and treated with celastrol and triptolide at 3 mg/Kg bw and 0.25 mg/Kg bw, respectively. Results: Both celastrol and triptolide significantly suppressed mammosphere size and number. Furthermore, expression of breast cancer stem cell markers DCLK1, ALDH1 and CD133 were significantly reduced in the two cell lines following treatment. Flow cytometry also suggested a significant reduction in DCLK1+, ALDH+ and CD133+ cells. Recently, Notch signaling has been shown to be critical for self-renewal of cancer stem cells. In cells treated with either celastrol or triptolide, there was a significant reduction in activated Notch intracellullar domain (NICD), and its downstream target Hes-1. However, in cells where we ectopically overexpressed NICD, neither compound was as potent as control, vector transfected cells in reducing 2D cell proliferation or 3D mammosphere formation, suggesting the direct role for inhibiting Notch activation as a mechanism of action for the two compounds. Furthermore, inhibition of Notch signaling pathway by using Υ-secretase inhibitor DAPT shows further reduction in mammosphere formation and Notch and its downstream target gene. We confirmed these finding in vivo using BT20 tumor xenografts grown in athymic nude mice. There was a reduction in the size and weight of tumors in mice treated with celastrol or triptolide. Western blot data showed that there is a decrease in activated Notch-1 protein and stem cell marker, DCLK1+ and ALDH+ in celastrol and triptolide treated xenograft tissue. Conclusion: Taken together these data suggest that both celastrol and triptolide affect cancer stem cells in TNBC, in part through inhibition of Notch signaling. Citation Format: Prabhu Ramamoorthy, Parthasarathy Rangarajan, Roy Jensen, Shrikant Anant. Targeting Hsp90 affects stem cell signaling in triple negative breast cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3314.