Abstract C283: Non-malignant cells from tissues targeted by metastatic breast cancer induce tumor cell resistance to antiestrogens and conventional chemotherapeutic agents.

The role of the microenvironment of the metastatic niche on breast cancer (BrCa) de novo or secondary resistance to currently available treatments is poorly understood. To study the tumor-stroma interactions and explore potential therapeutic targets in the context of the local metastatic microenvironment of disseminated BrCa we assembled heterotypic in vitro three-dimensional (3D) tissue cultures comprised of estrogen receptor-positive (ER+) BrCa cells and non-malignant accessory cells from organs frequently targeted by metastatic disease, which model the composition and architecture of metastatic lesions. MCF7, T47D and ZR75-1 cells expressing luciferase were immersed in extracellular matrices (collagen type I and Matrigel) alone or with accessory cells, and exposed to clinically achievable concentrations of FDA-approved antineoplastic agents widely used for the treatment of BrCa. The presence of HS-5 bone marrow stromal cells (BMSCs) induced BrCa cell resistance to vinca alkaloids (vincristine, vinblastin and vinorelbin) and taxanes (docetaxel, paclitaxel, cabazitaxel) both in 2D and 3D co-cultures. BMSCs sensitized the MCF7 to antifolates (methotrexate, pralatrexate) in 2D (but not in 3D) cultures. Importantly, BrCa spheroid response to antiestrogens (hydroxytamoxifen (4-OHT), fulvestrant or raloxifene) in 3D conditions was marked by acinar differentiation of tumor spheroids that resembles normal breast epithelium. Co-culture of BrCa cell and immortalized accessory cells from the bone (BMSCs; HOBIT and hFOB osteoblasts), brain (SVGp12 astrocytes), liver (THLE3 hepatocytes), and lung (MRC9 fibroblasts) in 3D conditions, conferred resistance to 4-OHT, raloxifene, and fulvestrant at clinically relevant doses. MCF7 cells injected s.c. in nude mice together with BMSCs (heterotypic xenografts) had reduced response to tamoxifen compared with monotypic xenografts (MCF7 alone). BMSCs induced in MCF7 cells downregulation of TGFβ2; upregulation of a transcriptional signature enriched for genes associated with high-grade tumors, including members of the JAK/STAT pathway, interferon signal gene signature, signaling through EGFR superfamily members, NFkB and other antiapoptotic pathways; and elevated c-Myc protein levels. The growth of MCF7 cells in co-culture with BMSCs or hepatocytes was inhibited by exogenous TGFβ1/2 or neutralizing antibodies against IL-6, INF-gamma, and TNF-alpha. Inhibition of JAK1/2 using ruxolitinib also inhibited MCF7 cell growth in co-culture conditions. Our results indicate that accessory cells representing the microenvironment of metastatic sites confer BrCa cell resistance to antiestrogens and chemotherapeutic agents in conventional and/or 3D tissue cultures and xenograft models. The mechanisms of stroma-induced drug resistance reveal candidate therapeutic targets for refractory BrCa patients with metastatic disease. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):C283. Citation Format: Eugen Dhimolea, Richard W.J. Groen, Kornelia Polyak, Constantine S. Mitsiades. Non-malignant cells from tissues targeted by metastatic breast cancer induce tumor cell resistance to antiestrogens and conventional chemotherapeutic agents. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr C283.