Carcinomas Deficient Mouse Mammary − β Transforming Growth Factor-Stromally Derived Lysyl Oxidase Promotes Metastasis of Updated

The tumor stromal environment can dictate many aspects of tumor progression. A complete understanding of factors driving stromal activation and their role in tumormetastasis is critical to furthering research with the goal of therapeutic intervention. Polyomamiddle T-inducedmammary carcinomas lacking the type II TGF-b receptor (PyMT) are highly metastatic compared with control PyMT-induced carcinomas (PyMT). We hypothesized that the PyMT-activated stroma interacts with carcinoma cells to promote invasion and metastasis. We show that the extracellular matrix associated with PyMT tumors is stiffer and has more fibrillar collagen and increased expression of the collagen crosslinking enzyme lysyl oxidase (LOX) compared with PyMT mammary carcinomas. Inhibition of LOX activity in PyMT mice had no effect on tumor latency and size, but significantly decreased tumor metastasis through inhibition of tumor cell intravasation. This phenotype was associated with a decrease in keratin 14–positive myoepithelial cells in PyMT tumors following LOX inhibition as well as a decrease in focal adhesion formation. Interestingly, the primary source of LOX was found to be activatedfibroblasts. LOXexpression in thesefibroblasts can be driven bymyeloid cell-derived TGF-b, which is significantly linked to human breast cancer. Overall, stromal expansion in PyMT tumors is likely caused through the modulation of immune cell infiltrates to promote fibroblast activation. This feeds back to the epithelium to promote metastasis by modulating phenotypic characteristics of basal cells. Our data indicate that epithelial induction of microenvironmental changes can play a significant role in tumorigenesis and attenuating these changes can inhibit metastasis. Cancer Res; 73(17); 5336–46. 2013 AACR. Introduction The stromal microenvironment of a tumor is an essential component of tumor progression (1). Composed of various resident and recruited cell types as well as extracellular proteins, the stromal components can determine phenotypic characteristics and ultimately patient outcome. By providing growth factors and other migratory signals as well as depositing scaffolding proteins, the tumor stroma can effectively drive or impede a tumor cell toward intravasation and metastatic colonization (2). Specifically, matrix deposition and remodeling, largely facilitated through fibroblast mediators, promotes tumor growth and migration (3). While stromal influence is acknowledged, a full understanding of the signals driving the formation of a tumor promoting stroma as well as the reciprocal response of the epithelium to these changes has yet to be obtained. Insights into these interactions will provide the backbone for future therapeutic interventions specifically targeting tumor–stromal crosstalk. Extracellular matrix (ECM) proteins, and in particular collagen, are a major component of the tumor microenvironment and exert significant effects on the tumor epithelium (4). Through its integrin mediators, ECM proteins encourage tumor growth and invasiveness. Increased mammographic density, which is significantly associated with collagen levels, independently predicts increased probability of occurrence of breast cancer in patients (5). These results are mimicked in murine models of breast cancer progression in which deposition of collagen that is unable to be proteolytically cleaved results in increased tumor formation as well as increased lung metastasis (6). Recently, it has been appreciated that ECMepithelial crosstalk is not only mediated by the ECM proteins themselves, but by the orientation and crosslinking status of the collagen fibers. Lysyl oxidase (LOX) is a matrix-modifying enzyme that cross-links and stiffens collagen fibers to promote their stability (3). LOX has garnered interest in breast cancer as Authors' Affiliations: Department of Cancer Biology, Vanderbilt University School of Medicine and Vanderbilt-Ingram Cancer Center, Nashville, Tennessee; and Department of Surgery andCenter for Bioengineering and Tissue Regeneration, University of California at San Francisco, San Francisco, California Note: Supplementary data for this article are available at Cancer Research Online (http://cancerres.aacrjournals.org/). Corresponding Author: H.L. Moses, Department of Cancer Biology, Vanderbilt University School of Medicine and Vanderbilt-Ingram Cancer Center, 698 Preston Research Building, 2220 Pierce Ave., Nashville, TN 37212. Phone: 615-936-1782; Fax: 615-936-1790; E-mail: hal.moses@vanderbilt.edu doi: 10.1158/0008-5472.CAN-13-0012 2013 American Association for Cancer Research. Cancer Research Cancer Res; 73(17) September 1, 2013 5336 on August 29, 2014. © 2013 American Association for Cancer Research. cancerres.aacrjournals.org Downloaded from Published OnlineFirst July 15, 2013; DOI: 10.1158/0008-5472.CAN-13-0012