Matrix Metalloproteinase 13 Is Induced in Fibroblasts in Polyomavirus Middle T Antigen-Driven Mammary Carcinoma without Influencing Tumor Progression Boye S. Nielsen1¤a, Mikala Egeblad2, Fritz Rank3, Hanne A. Askautrud2,5, Caroline J. Pennington 4 , Tanja X. Pedersen1¤b, Ib J. Christensen 1 , Dylan R. Edwards4, Zena Werb2, Leif R. Lund 1 * 1 The Finsen Laboratory, Rigshospitalet, Copenhagen, Denmark, 2 Department of Anatomy, University of California San Francisco, San Francisco, California, United States of America, 3 Department of Pathology, Rigshospitalet, Copenhagen, Denmark, 4 School of Biological Sciences, University of East Anglia, Norwich, Norfolk, United Kingdom, 5 Department of Medical Genetics, Ulleva˚l University Hospital and Faculty of Medicine, University of Oslo, Oslo, Norway Abstract Matrix metalloproteinase (MMP) 13 (collagenase 3) is an extracellular matrix remodeling enzyme that is induced in myofibroblasts during the earliest invasive stages of human breast carcinoma, suggesting that it is involved in tumor progression. During progression of mammary carcinomas in the polyoma virus middle T oncogene mouse model (MMTV- PyMT), Mmp13 mRNA was strongly upregulated concurrently with the transition to invasive and metastatic carcinomas. As in human tumors, Mmp13 mRNA was found in myofibroblasts of invasive grade II and III carcinomas, but not in benign grade I and II mammary intraepithelial neoplasias. To determine if MMP13 plays a role in tumor progression, we crossed MMTV-PyMT mice with Mmp13 deficient mice. The absence of MMP13 did not influence tumor growth, vascularization, progression to more advanced tumor stages, or metastasis to the lungs, and the absence of MMP13 was not compensated for by expression of other MMPs or tissue inhibitor of metalloproteinases. However, an increased fraction of thin collagen fibrils was identified in MMTV- PyMT;Mmp132/2 compared to MMTV-PyMT;Mmp13+/+ tumors, showing that collagen metabolism was altered in the absence of MMP13. We conclude that the expression pattern of Mmp13 mRNA in myofibroblasts of invasive carcinomas in the MMTV-PyMT breast cancer model recapitulates the expression pattern observed in human breast cancer. Our results suggest that MMP13 is a marker of carcinoma-associated myofibroblasts of invasive carcinoma, even though it does not make a major contribution to tumor progression in the MMTV-PyMT breast cancer model. Citation: Nielsen BS, Egeblad M, Rank F, Askautrud HA, Pennington CJ, et al. (2008) Matrix Metalloproteinase 13 Is Induced in Fibroblasts in Polyomavirus Middle T Antigen-Driven Mammary Carcinoma without Influencing Tumor Progression. PLoS ONE 3(8): e2959. doi:10.1371/journal.pone.0002959 Editor: Mikhail V. Blagosklonny, Ordway Research Institute, United States of America Received March 26, 2008; Accepted July 2, 2008; Published August 13, 2008 Copyright: 2008 Nielsen et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: The study was financially supported by the Danish Cancer Society, the Meyer Foundation, the Danish Medical Research Council (ME), the European Commission: grant number LSHC-CT-2003-503297 (Cancerdegradome) and FP7 grant (Microenvimet), and by the National Cancer Institute (USA) grants CA057621 and CA072006 (ZW). None of these supporting institutions played any role during the study and manuscript preparation. Competing Interests: The authors have declared that no competing interests exist. * E-mail: lund@inet.uni2.dk €a Current address: Exiqon A/S, Diagnostic Product Development, Vedbæk, Denmark €b Current address: Department of Clinical Biochemistry, Rigshospitalet, Copenhagen, Denmark Introduction The ability of cancer cells to invade or metastasize to distant organs is linked to their ability to traverse the extracellular matrix. The extracellular matrix (ECM) is a heterogeneous protein matrix that consists of a variety of collagens, laminins, fibronectin and proteoglycans [1,2]. Matrix metalloproteinases (MMP) cleave and remodel ECM components [3], and increased activity of some of the MMPs, including MMP2, 9, and 14 can promote cancer cell invasion in vivo [3–7]. MMP13 (collagenase-3) was originally identified in human breast cancer tissue [8]. It is secreted from cells as an inactive zymogen that can be activated by the MMP2/MMP14/tissue inhibitor of metalloproteinase (TIMP2) complex [9] or by plasmin [10]. MMP13 acts in the extracellular environment as a potent collagenase capable of degrading a variety of collagens [11]. Mmp13 mRNA is expressed in a subpopulation of myofibroblasts in invasive ductal breast carcinomas [12], but rarely in normal breast, benign breast lesions and ductal carcinoma in situ (DCIS). Interestingly, the presence of microinvasion in DCIS is associated with focal expression of Mmp13 mRNA in stromal fibroblasts [12,13]. Direct comparison of the Mmp13 mRNA expression pattern with that of the Mmp2, Mmp11 and Mmp14 mRNAs indicates that Mmp13 is unique in this respect since these other MMPs are also present in DCIS in the absence of invasion [12]. These observations raise the question as to whether MMP13 is a rate-limiting proteinase that mediates the initial steps in breast cancer invasion. The role of MMPs and other ECM-degrading proteinases during breast cancer progression has been studied using a variety of murine mammary tumor models [6]. In one model, tumors are induced by expressing the polyoma middle T oncogene under the mouse mammary tumor virus LTR (MMTV-PyMT) [14] leading to early onset hyperplasia in the mammary glands [15]. The PyMT-oncogene induced mammary tumors go through typical pre-invasive, invasive and metastatic phases similarly to human breast cancer [14]. The onset of tumorigenesis in the MMTV- PyMT mouse model is observed at 2–3 weeks after birth, and the non-invasive early stage tumors, mammary intraepithelial neopla- PLoS ONE | www.plosone.org 1 August 2008 | Volume 3 | Issue 8 | e2959