Tissue Inhibitor of Metalloproteinases-1 Promotes Liver Metastasis by Induction of Hepatocyte Growth Factor Signaling Charlotte Kopitz, 1 Michael Gerg, 1 Obul Reddy Bandapalli, 4 Dilek Ister, 1 Caroline J. Pennington, 6 Stephanie Hauser, 1 Christin Flechsig, 1 Hans-Willi Krell, 7 Dalibor Antolovic, 5 Keith Brew, 8 Hideaki Nagase, 9 Manfred Stangl, 2 Claus W. Hann von Weyhern, 3 Bjo¨rn L.D.M. Bru¨cher, 2 Karsten Brand, 4 Lisa M. Coussens, 10 Dylan R. Edwards, 6 and Achim Kru¨ger 1 1 Institut fu¨r Experimentelle Onkologie und Therapieforschung, 2 Chirurgische Klinik, and 3 Institut fu¨r Allgemeine Pathologie und Pathologische Anatomie, Klinikum rechts der Isar der Technischen Universita¨t Mu¨nchen, Munich, Germany; 4 Institute of Pathology and 5 Department of Surgery, University Hospital Heidelberg, Heidelberg, Germany; 6 School of Biological Sciences, University of East Anglia, Norwich, Norfolk, United Kingdom; 7 Roche Diagnostics GmbH, Penzberg, Germany; 8 Department of Biomedical Science, Florida Atlantic University, Boca Raton, Florida; 9 The Kennedy Institute of Rheumatology Division, Imperial College School of Medicine, London, United Kingdom; and 10 Department of Pathology and Comprehensive Cancer Center, University of California, San Francisco, California Abstract Balanced expression of proteases and their inhibitors is one prerequisite of tissue homeostasis. Metastatic spread of tumor cells through the organism depends on proteolytic activity and is the death determinant for cancer patients. Paradoxically, increased expression of tissue inhibitor of metalloproteinases- 1 (TIMP-1), a natural inhibitor of several endometalloprotei- nases, including matrix metalloproteinases and a disintegrin and metalloproteinase-10 (ADAM-10), in cancer patients is negatively correlated with their survival, although TIMP-1 itself inhibits invasion of some tumor cells. Here, we show that elevated stromal expression of TIMP-1 promotes liver metas- tasis in two independent tumor models by inducing the hepatocyte growth factor (HGF) signaling pathway and expression of several metastasis-associated genes, including HGF and HGF-activating proteases, in the liver. We also found in an in vitro assay that suppression of ADAM-10 is in principle able to prevent shedding of cMet, which may be one explanation for the increase of cell-associated HGF receptor cMet in livers with elevated TIMP-1. Similar TIMP-1–associ- ated changes in gene expression were detected in livers of patients with metastatic colorectal cancer. The newly identi- fied role of TIMP-1 to create a prometastatic niche may also explain the TIMP-1 paradoxon. [Cancer Res 2007;67(18):8615–23] Introduction Metastasis and progressive scattering of tumor cells throughout tissues is the main cause of organ failure and subsequent death of cancer patients. Proteolytic remodeling of components of the extracellular matrix (ECM) in tissue interstitium is a prerequisite of the metastatic process (1). Matrix metalloproteinases (MMP) have long been thought to be major regulators of the pericellular protease-dependent steps of tumor progression (2). Altered expression profiles of some MMPs have been correlated with poor clinical prognosis for several human tumor types (3). Based on these facts, the historical view of MMPs and cancer was that they exerted proinvasive and prometastatic activity by mere ECM remodeling (1). Thus, it was hypothesized that therapeutic broad- spectrum inhibition of MMPs would result in antimetastatic activity (1). Indeed, under certain experimental circumstances, overexpression of the endogenous broad-spectrum metalloprotei- nase [including a disintegrin and metalloproteinase-10 (ADAM-10); ref. 4] inhibitor tissue inhibitor of metalloproteinases-1 (TIMP-1; ref. 5) can reduce tumor cell invasion (6, 7). In contrast to these observations are other results, which are actually in agreement with clinical studies showing a correlation between elevated TIMP-1 expression and poor prognosis in many human cancer types (5, 8), where elevated levels of TIMP-1 either had no effect on metastasis (9) or led to increased incidence (10, 11) or growth (12) of primary tumors. These effects of TIMP-1 on cancer development have been attributed to its known proproliferative, antiapoptotic (5), or proangiogenic (13) activities, which are necessary but not sufficient for metastasis. Tumor cell invasion and metastasis are the primary determi- nants of the survival of cancer patients (13). Signaling pathways leading to the expression of proteolytic enzymes have been suggested to be key mediators of invasive growth (14). Hepatocyte growth factor (HGF/scatter factor) signaling regulates a multitude of downstream prometastatic effector molecules, such as urokinase- type plasminogen activator (uPA), uPA receptor (uPAR), plasminogen activator inhibitor-1 (PAI-1), and MMPs (15–17). HGF stimulates metastasis of diverse tumor cells (15) and elevated levels of circulating HGF (18) or aberrant activity of cMet (14) has been detected in patients with many cancer types, including non- Hodgkin’s lymphoma (19) and colorectal cancer (14). Although elevated TIMP-1 expression in these patients correlates with decreased survival, a link between TIMP-1 and metastasis- promoting pathways has previously not been described. Here, we address this issue and report that elevated stromal levels of TIMP-1 induce HGF signaling, leading to enhanced expression of other metastasis-promoting genes. Consequently, susceptibility of the liver for metastasis of tumor cell lines of different origin was shown to be increased, assigning a new role to TIMP-1 as a factor creating a prometastatic niche. Materials and Methods Cell culture and adenoviruses. HEK 293 (20), HT1080lacZ -K15 (21), L-CI.5s (22), and 293T cells (23) were cultured as described previously. Note: Supplementary data for this article are available at Cancer Research Online (http://cancerres.aacrjournals.org/). Requests for reprints: Achim Kru¨ger, Institut fu¨r Experimentelle Onkologie und Therapieforschung, Klinikum rechts der Isar der Technischen Universita¨t Mu¨nchen, Ismaninger Str. 22, D-81675 Munich, Germany. Phone: 49-89-4140-4463; Fax: 49-89- 4140-6182; E-mail: achim.krueger@lrz.tu-muenchen.de. I2007 American Association for Cancer Research. doi:10.1158/0008-5472.CAN-07-0232 www.aacrjournals.org 8615 Cancer Res 2007; 67: (18). 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