The initial invasive processes during cancer development remain largely unknown. Stromelysin-3/matrix metalloproteinase 11 (ST3/MMP11) is associated with tumor invasion and poor prognosis. We present novel evidence that adipocytes present at human breast tumor invasive front are induced by cancer cells to express ST3. Using mouse syngeneic model, light and electron microscopy showed that in ST3-deficient mice but not in wild-type mice, forced cancer cell-adipocyte interaction/crosstalk results in adipocyte membrane alteration, allowing cancer cell fat infiltration and death. Thus, adipocytes are involved in initial cancer cell survival into connective tissue, and this effect is ST3 mediated. This suggested that ST3 might play a role in adipocyte metabolism. Accordingly, ST3-deficient mice exhibited fat excess and increased mRNA levels of peroxisome proliferator-activated receptor ; (PPAR;) and adipocyte protein 2 (aP2) adipogenic markers, indicating that, in vivo, ST3 negatively regulates fat homeostasis. Moreover, ST3-deficient mouse embryonic fibroblasts exhibited a dramatic enhanced potential to differentiate into adipocytes associated with increased PPAR; and aP2 expression, and recombinant ST3 treatment reverted their differentiation. Thus, in vitro, ST3 reduces adipocyte differentiation in an autocrine manner. High fibroblasts/adipocytes ratio is a stroma feature, and peritumoral fibroblast origin remains debated. Our results support the concept that invading cancer cells aberrantly restore the negative ST3 function on adipogenesis into proximal adipocytes/preadipocytes, leading to the accumulation/maintenance of a particular peritumoral fibroblast subpopulation. Accordingly, in human breast tumors, we observed that ST3-expressing peritumoral fibroblasts are distinct from A-smooth muscle actinexpressing myofibroblasts. This constitutes the first report of implication of a MMP in cancer cell-adipocyte interaction/ crosstalk during early steps of connective tissue invasion. (Cancer Res 2005; 65(23): 10862-71)

Transforming growth factor B1 (TGFB) is a tumor suppressor during the initial stage of tumorigenesis, but it can switch to a tumor promoter during neoplastic progression. Ionizing radiation (IR), both a carcinogen and a therapeutic agent, induces TGFB activation in vivo. We now show that IR sensitizes human mammary epithelial cells (HMEC) to undergo TGFB-mediated epithelial to mesenchymal transition (EMT). Nonmalignant HMEC (MCF10A, HMT3522 S1, and 184v) were irradiated with 2 Gy shortly after attachment in monolayer culture or treated with a low concentration of TGFB (0.4 ng/mL) or double treated. All double-treated (IR + TGFB) HMEC underwent a morphologic shift from cuboidal to spindle shaped. This phenotype was accompanied by a decreased expression of epithelial markers E-cadherin, Bcatenin, and ZO-1, remodeling of the actin cytoskeleton, and increased expression of mesenchymal markers N-cadherin, fibronectin, and vimentin. Furthermore, double treatment increased cell motility, promoted invasion, and disrupted acinar morphogenesis of cells subsequently plated in Matrigel. Neither radiation nor TGFB alone elicited EMT, although IR increased chronic TGFB signaling and activity. Gene expression profiling revealed that double-treated cells exhibit a specific 10-gene signature associated with Erk/mitogenactivated protein kinase (MAPK) signaling. We hypothesized that IR-induced MAPK activation primes nonmalignant HMEC to undergo TGFB-mediated EMT. Consistent with this, Erk phosphorylation was transiently induced by irradiation and persisted in irradiated cells treated with TGFB, and treatment with U0126, a MAP/Erk kinase (MEK) inhibitor, blocked the EMT phenotype. Together, these data show that the interactions between radiation-induced signaling pathways elicit heritable phenotypes that could contribute to neoplastic progression. [Cancer Res 2007;67(18):8662-70]

Purpose: Metastatic melanomas are generally resistant to chemotherapy and radiation, even when wild-type for p53. These tumors often grow in small nests where many of the cells have little contact with extracellular matrix (ECM). Previous work showed that M21melanomas undergo apoptosis in response to chemotherapy when cells are adherent to ECM but not in suspension. Thus, reduced integrin-dependent adhesion to ECM could mediate therapy resistance. The goal of this study was to test whether stimulation of integrin signaling could increase chemotherapeutic efficacy. Experimental Design: Colony forming assays and survival assays were used to test the responses of melanoma lines in vitro. Severe combined immunodeficient mice with subcutaneous human melanomas received chemotherapy with or without reagents that stimulate integrin signaling; tumor volume was then monitored over time. Results: Clonal growth assays confirmed that M21 cells showed reduced sensitivity to the chemotherapeutic drug 1-h-D-arabinofuranosylcytosine (araC). When five additional primary melanoma lines were screened, 80% showed higher sensitivity when adherent compared with suspended. Subcutaneous M21 tumors in vivo showed minimal ECM between tumor cells. To evaluate the importance of integrin signaling in chemoresistance in this model, mice were treated with araC, with or without the multivalent snake venom disintegrin contortrostatin or the activating anti-h1 integrin antibody TS2/16. Although araC, TS2/16, or contortrostatin alone had little effect on M21 tumor growth, combining araC with either integrin signaling reagents strongly reduced growth (P = 0001). Conclusions: Loss of integrin-mediated adhesion is rate limiting for therapeutic response in this model. Combining chemotherapy with reagents that stimulate integrin signaling may therefore provide a new approach to therapy.Melanomas are usually resistant to chemotherapy and radiation even at early stages (1). Surgery is therefore the major mode of treatment, which, if unsuccessful, leaves few options. As a consequence, 5-year survival rates for patients with unresectable or metastatic disease are <10% (2). Because melanomas often have wild-type p53 genes and lack known defects in other DNA damage and apoptosis pathways, the reasons for chemotherapy resistance are poorly understood.Integrin-mediated adhesion promotes the transmission of many signaling pathways initiated by growth factor receptors, including Erk mitogen-activated protein kinase, phosphatidylinositol-3-OH-kinase, and Rho family GTPases (3). As a result of these synergies, many cell types require integrin-mediated adhesion to extracellular matrix (ECM) for survival (4). This mechanism, however, is generally lost in metastatic cancers. Among cells that survive well in suspension, a subset exhibits a synergy between adhesion and DNA damage pathways (5, 6). These include mouse embryo fibroblasts, human fibrosarcoma, and human melanoma. In these systems, including melanomas, loss of adhesion results in decreased p53 level...

We hypothesize that identification of the circumstances and tumors in which TGFbeta manipulation enhances tumor cell radiosensitivity, while protecting normal tissues, could significantly increase therapeutic index.