Abstract B18: Estrogen deficiency causes reawakening of dormant disseminated breast cancer cells via angiopoietin-1/Tie2 signaling
Authors: Han Hyun Ho , Cho Nam Hoon , Kang Suki , Lee Joo Hyun , Kim Ji Eun 
Published: December 2015

Introduction: Compared to premenopausal women, postmenopausal women with breast cancer are at increased risks of bone metastasis. When cancer cells migrate into bone marrow (BM), they stay dormant in stable microvasculature but proliferate in unstable microvasculature. Angiopoietin-1(ANGPT1)/Tie2 signaling regulates microvascular stability. ANGPT-1 stabilizes microvasculature, while ANGPT-2 destabilizes it and promotes neovascularization. We hypothesized that postmenopausal estrogen deficiency induces reawakening of the dormant tumor cells in BM through modulation of angiogenesis. We examined the effects of estrogen depletion on the proliferative status of breast cancer cells (BCCs) in a BM microvasculature model, and quantified the changes of ANGPT1 and ANGPT2 expressions in the model before and after estrogen depletion.

Materials and Methods: Bone marrow microvasculature model was generated by using bone marrow mesenchymal stem cells (MSCs) and endothelial cells (ECs). To deplete estrogen in the model, charcoal-stripped serum-containing culture medium with or without 17β-estradiol (E2) were used. ANGPT1 and ANGPT2 protein levels in the model were determined by western blot. Estrogen receptor(ER)-positive MCF-7 and ER-negative MDA-MB-231 breast cancer cell lines were used. Fluorescence-tagged tumor cells were seeded in the BM microvasculature models with or without estrogen and their proliferation were monitored by microscope and microscope fluorescence reader. Recombinant human ANGPT1 (RhANGPT1) and Tie2-specific blocking antibody were used for ANGPT1/Tie2 signaling modulation.

Results: ANGPT2 expression was significantly increased in BM microvasculature model exposed to prolonged estrogen depletion. ANGPT1 expression was not changed. BCCs in the estrogen-depleted BM microvasculature model grew significantly faster than those in the model supplemented with E2. Blocking ANGPT1/Tie2 signaling in the model by Tie2-specific blocking antibody enhanced BCC proliferation in presence of estrogen, whereas RhANGPT1-treatment inhibited BCC proliferation in estrogen-depleted BM microvasculature model. In further, RhANGPT1 protected BCCs from doxorubicin-induced apoptosis. RhANGPT1 inhibited proliferation of Tie2-expressing MDA-MB-231 cells in absence of BM MSCs or ECs, suggesting direct interaction of ANGPT1 and Tie2-expressing tumor cells.

Conclusions: With in vitro disseminated breast cancer cell model, we confirmed that estrogen deficiency modified bone marrow microvasculature to more metastasis-prone environment. We identified the ANGPT1/Tie2 signaling as a key regulator of angiogenic tumor cell dormancy. The results suggest that postmenopausal estrogen deficiency may lead to reawakening of dormant BCCs by altering ANGPT1 and ANGPT2 levels in BM microvasculature. Angiogenic dormancy in association with estrogen implies a new strategy against metastatic recurrence in postmenopausal women with remote breast cancer history.

Citation Format: Hyun Ho Han, Nam Hoon Cho, Suki Kang, Joo Hyun Lee, Ji Eun Kim. Estrogen deficiency causes reawakening of dormant disseminated breast cancer cells via angiopoietin-1/Tie2 signaling. [abstract]. In: Proceedings of the AACR Special Conference: Tumor Angiogenesis and Vascular Normalization: Bench to Bedside to Biomarkers; Mar 5-8, 2015; Orlando, FL. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl):Abstract nr B18.