Abstract 973: Tumor-associated fibroblasts induce long-term changes in breast cancer invasion, cancer stem cell populations, and metastasis through modulation of the Tiam1-ostepontin pathway
Authors: Xu Kun , Tian Xuejun , Naber Stephen P. , Oh Sun , Kuperwasser Charlotte , Buchsbaum Rachel J. 
Published: May 2015

Abstract
We have previously shown that silencing of the Rac exchange factor Tiam1 in tumor-associated fibroblasts induces increased invasion and metastasis in epithelial and cancer cells using 3D-tissue culture models and a mouse model of human breast cancer. Here we investigate the underlying mechanism using a novel method for isolating human breast cancer cells out of 3D spheroid co-cultures containing mammary fibroblasts. Co-culture of human breast cancer cells with Tiam1-deficient fibroblasts induces increased cancer cell invasion into matrix, while co-culture with Tiam1-overexpressing fibroblasts induces decreased invasion into matrix. Strikingly, breast cancer cells isolated from 3D culture with Tiam1-deficient fibroblasts exhibit persistent increases in migration, epithelial-mesenchymal transition (EMT), and cancer stem cell characteristics as assayed by tumorsphere assay, flow cytometry, and limiting dilution mouse xenograft implantation experiments. In contrast, breast cancer cells exposed to Tiam1-overexpressing fibroblasts and then isolated from 3D co-cultures persistently display the opposite phenotypes. The observed changes in the breast cancer cells are dependent on fibroblast-produced osteopontin (OPN), as OPN silencing or antibody-mediated inhibition abrogates the increased invasion, migration, and cancer stem cell populations induced by Tiam1-deficient fibroblasts. Treatment of co-cultures by chemotherapeutic agents does not prevent effects induced by Tiam1-deficient fibroblasts. In contrast, treatment of co-cultures by the OPN inhibitor Agelastatin A reverses all effects induced by Tiam1-deficient fibroblasts, and blocks subsequent development of lung metastases in the mouse xenograft model. Finally, in human breast cancer samples Tiam1 expression is significantly decreased in fibroblasts associated with invasive human breast cancers compared to fibroblasts associated with ductal carcinoma in situ (DCIS). Conversely, OPN expression is significantly increased in fibroblasts associated with invasive breast cancers compared to fibroblasts associated with DCIS. These results suggest that regulated Tiam1 expression in breast cancer-associated fibroblasts modulates breast cancer invasion, EMT, cancer stem cell populations, and metastasis through regulation of fibroblast OPN secretion. At least some breast cancers demonstrate commensurate changes in fibroblast Tiam1 and OPN expression correlating with invasiveness, suggesting a mechanism for therapeutic targeting.
Citation Format: Kun Xu, Xuejun Tian, Stephen P. Naber, Sun Oh, Charlotte Kuperwasser, Rachel J. Buchsbaum. Tumor-associated fibroblasts induce long-term changes in breast cancer invasion, cancer stem cell populations, and metastasis through modulation of the Tiam1-ostepontin pathway. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 973. doi:10.1158/1538-7445.AM2014-973