TRAF4 is a novel phosphoinositide-binding protein modulating tight junctions and favoring cell migration.

Tumor necrosis factor (TNF) receptor-associated factor 4 (TRAF4) is frequently overexpressed in carcinomas, suggesting a specific role in cancer. Although TRAF4 protein is predominantly found at tight junctions (TJs) in normal mammary epithelial cells (MECs), it accumulates in the cytoplasm of malignant MECs. How TRAF4 is recruited and functions at TJs is unclear. Here we show that TRAF4 possesses a novel phosphoinositide (PIP)-binding domain crucial for its recruitment to TJs. Of interest, this property is shared by the other members of the TRAF protein family. Indeed, the TRAF domain of all TRAF proteins (TRAF1 to TRAF6) is a bona fide PIP-binding domain. Molecular and structural analyses revealed that the TRAF domain of TRAF4 exists as a trimer that binds up to three lipids using basic residues exposed at its surface. Cellular studies indicated that TRAF4 acts as a negative regulator of TJ and increases cell migration. These functions are dependent from its ability to interact with PIPs. Our results suggest that TRAF4 overexpression might contribute to breast cancer progression by destabilizing TJs and favoring cell migration.

STARD3 or STARD3NL and VAP form a novel molecular tether between late endosomes and the ER.

Inter-organelle membrane contacts sites (MCSs) are specific subcellular regions favoring the exchange of metabolites and information. We investigated the potential role of the late-endosomal membrane-anchored proteins StAR related lipid transfer domain-3 (STARD3) and STARD3 N-terminal like (STARD3NL) in the formation of MCSs involving late-endosomes (LEs). We demonstrate that both STARD3 and STARD3NL create MCSs between LEs and the endoplasmic reticulum (ER). STARD3 and STARD3NL use a conserved two phenylalanines in an acidic tract (FFAT)-motif to interact with ER-anchored VAP proteins. Together, they form an LE-ER tethering complex allowing heterologous membrane apposition. This LE-ER tethering complex affects organelle dynamics by altering the formation of endosomal tubules. An in situ proximity ligation assay between STARD3, STARD3NL and VAP proteins identified endogenous LE-ER MCS. Thus, we report here the identification of proteins involved in inter-organellar interaction.

[The tumoral stroma, a breeding ground for cancer cells]. / Le stroma tumoral - un terreau fertile pour la cellule cancéreuse.

Carcinomas are constituted by malignant epithelial cells and the tumor microenvironment also called tumoral stroma. The present non-exhaustive review will focus on cellular and molecular key events implicating cancer-associated fibroblasts (CAF) and the extracellular matrix (ECM) in the stroma remodeling processes occuring during tumor invasive steps. We will also discuss the place of the stroma in the cancerology today, the recent progresses done and its usefulness to design new therapies.

Inhibition of cyclooxygenase-2 causes regression of gastric adenomas in trefoil factor 1 deficient mice.

Cyclooxygenase-2 (Cox-2) expression is a marker of reduced survival in gastric cancer patients, and inhibition of Cox-2 suppresses gastrointestinal carcinogenesis in experimental animal models. To investigate the role of Cox-2 in gastric carcinogenesis in vivo, we utilized trefoil factor 1 (Tff1) deficient mice, which model the neoplastic process of the stomach by developing gastric adenomas with full penetrance. These tumors express Cox-2 protein and mRNA, and we have now investigated the effects of genetic deletion of the mouse Cox-2 gene [also known as prostaglandin-endoperoxide synthase 2 (Ptgs2)] and a Cox-2 selective drug celecoxib. Our results show that genetic deletion of Cox-2 in the Tff1 deleted background resulted in reduced adenoma size and ulceration with a chronic inflammatory reaction at the site of the adenoma. To characterize the effect of Cox-2 inhibition in more detail, mice that had already developed an adenoma were fed with celecoxib for 8-14 weeks, which resulted in disruption of the adenoma that ranged from superficial erosion to deep ulcerated destruction accompanied with chronic inflammation. Importantly, mice fed with celecoxib for 16 weeks, followed by control food for 9 weeks, redeveloped a complete adenoma with no detectable inflammatory process. Finally, we determined the identity of the Cox-2 expressing cells and found them to be fibroblasts. Our results show that inhibition of Cox-2 is sufficient to reversibly disrupt gastric adenomas in mice.

Inhibition of gastric carcinogenesis by the hormone gastrin is mediated by suppression of TFF1 epigenetic silencing.

BACKGROUND & AIMS: Epigenetic alterations have been correlated with field cancerization in human patients, but evidence from experimental models that specific epigenetic changes can initiate cancer has been lacking. Although hormones have been associated with cancer risk, the mechanisms have not been determined. The peptide hormone gastrin exerts a suppressive effect on antral gastric carcinogenesis. METHODS: N-methyl-N-nitrosourea (MNU)-dependent gastric cancer was investigated in hypergastrinemic (INS-GAS), gastrin-deficient (GAS(-/-)), Tff1-deficient (Tff1(+/-)), and wild-type (WT) mice. Epigenetic alterations of the trefoil factor 1 (TFF1) tumor suppressor gene were evaluated in vitro and in vivo. RESULTS: Human intestinal-type gastric cancers in the antrum exhibited progressive TFF1 repression and promoter hypermethylation. Mice treated with MNU exhibited a field defect characterized by widespread Tff1 repression associated with histone H3 lysine 9 methylation and H3 deacetylation at the Tff1 promoter in epithelial cells. In MNU-induced advanced cancers, DNA methylation at the Tff1 promoter was observed. Tumor induction and Tff1 repression were increased in MNU-treated mice by Helicobacter infection. Hypergastrinemia suppressed MNU-dependent tumor initiation and progression in a manner that correlated with gene silencing and epigenetic alterations of Tff1. In contrast, homozygous gastrin-deficient and heterozygous Tff1-deficient mice showed enhanced MNU-dependent field defects and cancer initiation compared with WT mice. In gastric cancer cells, gastrin stimulation partially reversed the epigenetic silencing in the TFF1 promoter. CONCLUSIONS: Initiation of antral gastric cancer is associated with progressive epigenetic silencing of TFF1, which can be suppressed by the hormone gastrin.

Methyl donor deficiency affects fetal programming of gastric ghrelin cell organization and function in the rat.

Methyl donor deficiency (MDD) during pregnancy influences intrauterine development. Ghrelin is expressed in the stomach of fetuses and influences fetal growth, but MDD influence on gastric ghrelin is unknown. We examined the gastric ghrelin system in MDD-induced intrauterine growth retardation. By using specific markers and approaches (such as periodic acid-Schiff, bromodeoxyuridine, homocysteine, terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling, immunostaining, reverse transcription-polymerase chain reaction), we studied the gastric oxyntic mucosa cellular organization and ghrelin gene expression in the mucosa in 20-day-old fetuses and weanling pups, and plasma ghrelin concentration in weanling rat pups of dams either normally fed or deprived of choline, folate, vitamin B6, and vitamin B12 during gestation and suckling periods. MDD fetuses weighed less than controls; the weight deficit reached 57% at weaning (P < 0.001). Both at the end of gestation and at weaning, they presented with an aberrant gastric oxyntic mucosa formation with loss of cell polarity, anarchic cell migration, abnormal progenitor differentiation, apoptosis, and signs of surface layer erosion. Ghrelin cells were abnormally located in the pit region of oxyntic glands. At weaning, plasma ghrelin levels were decreased (-28%; P < 0.001) despite unchanged mRNA expression in the stomach. This decrease was associated with lower body weight. Taken together, these data indicate that one mechanism through which MDD influences fetal programming is the remodeling of gastric cellular organization, leading to dysfunction of the ghrelin system and dramatic effects on growth.

Intake of grape-derived polyphenols reduces C26 tumor growth by inhibiting angiogenesis and inducing apoptosis.

This study evaluated the in vivo antitumor activity of grape-derived polyphenols. BALB/c mice were subcutaneously implanted with C26 colon carcinoma cells, and 2 d later they received either solvent or red wine polyphenols (RWPs) (100 mg/kg/d, human equivalent dose approximately 500 mg/d) in the drinking water for 25 d. Wistar rats received either solvent or RWPs (100 mg/kg/d, human equivalent dose approximately 1000 mg/d) in the drinking water 1 wk before injection of azoxymethane and were studied 10 wk later. In mice, RWPs inhibited tumor growth by 31%, reduced tumor vascularization and the number of lung metastases, decreased proliferation as indicated by down-regulation of Ki67, cyclin D1, and UHRF1, and increased apoptosis as indicated by TUNEL staining and active caspase-3 levels in tumor cells. RWPs reduced expression of VEGF, matrix metalloproteinase (MMP)-2, MMP-9, and cyclooxygenase-2 and increased expression of tumor suppressor genes p16(INK4A), p53, and p73 in tumor cells. In rats, RWPs reduced by 49% the number of azoxymethane-induced aberrant crypt foci (preneoplastic lesions) in colon. Thus, RWPs effectively reduced the development of colon carcinoma tumors in vivo by blunting tumor vascularization and by inhibiting proliferation and promoting apoptosis of tumor cells subsequent to an up-regulation of tumor suppressor genes.

Copper binds the carboxy-terminus of trefoil protein 1 (TFF1), favoring its homodimerization and motogenic activity.

Trefoil protein 1 (TFF1) is a small secreted protein belonging to the trefoil factor family of proteins, that are present mainly in the gastrointestinal (GI) tract and play pivotal roles as motogenic factors in epithelial restitution, cell motility, and other incompletely characterized biological processes. We previously reported the up-regulation of TFF1 gene in copper deficient rats and the unexpected property of the peptide to selectively bind copper. Following the previous evidence, here we report the characterization of the copper binding site by fluorescence quenching spectroscopy and mass spectrometric analyses. We demonstrate that Cys58 and at least three Glu surrounding residues surrounding it, are essential to efficiently bind copper. Moreover, copper binding promotes the TFF1 homodimerization, thus increasing its motogenic activity in in vitro wound healing assays. Copper levels could then modulate the TFF1 functions in the GI tract, as well as its postulated role in cancer progression and invasion.

Matrix metalloproteinase 11/stromelysin-3 exerts both activator and repressor functions during the hematogenous metastatic process in mice.

MMP11 expression is a poor prognosis factor in human carcinomas. Although it has been shown to favor primary tumor development, its role in metastatic processes remains unclear. We studied the hematogenous metastatic activity of C26 mouse colon cancer cells injected into the tail vain of wild-type or MMP11-deficient mice during 2 months. Using X-ray computed tomography to image metastasis development in recipient living mice, lung metastases were found to occur earlier and to grow faster in wild-type mice. Histological analyses of the lung, liver, kidney, adrenal gland, mammary gland, ovary and salivary gland, performed at the end of experiment, also showed lower numbers of metastases in wild-type mice, regardless of organ. Lung metastases showed similar Factor VIII-positive vascular networks regardless of the mouse MMP11 status. However, those found in MMP11-deficient mice also exhibited vessel-like structures that did not express Factor VIII, Lyve-1 and vimentin, and were not stained with PAS. Consequently, they did not correspond to vascular or lymphatic vessels or to vascular mimicry channels. Collectively, these results revealed significant spatio-temporal variability that is dependent on host MMP11 status. Furthermore, they point-out the paradoxical role of MMP11 in favoring the onset and growth of lung metastases but limiting lung foci number, and inhibiting the cancer cell dissemination to other organs. These data highlight the complexity of the metastatic process in which the same factor can play activator or repressor functions depending on the metastatic step.

Caveolin-1 (P132L), a common breast cancer mutation, confers mammary cell invasiveness and defines a novel stem cell/metastasis-associated gene signature.

Here we used the Met-1 cell line in an orthotopic transplantation model in FVB/N mice to dissect the role of the Cav-1(P132L) mutation in human breast cancer. Identical experiments were performed in parallel with wild-type Cav-1. Cav-1(P132L) up-regulated the expression of estrogen receptor-alpha as predicted, because only estrogen receptor-alpha-positive patients have been shown to harbor Cav-1(P132L) mutations. In the context of primary tumor formation, Cav-1(P132L) behaved as a loss-of-function mutation, lacking any tumor suppressor activity. In contrast, Cav-1(P132L) caused significant increases in cell migration, invasion, and experimental metastasis, consistent with a gain-of-function mutation. To identify possible molecular mechanism(s) underlying this invasive gain-of-function activity, we performed unbiased gene expression profiling. From this analysis, we show that the Cav-1(P132L) expression signature contains numerous genes that have been previously associated with cell migration, invasion, and metastasis. These include i) secreted growth factors and extracellular matrix proteins (Cyr61, Plf, Pthlh, Serpinb5, Tnc, and Wnt10a), ii) proteases that generate EGF and HGF (Adamts1 and St14), and iii) tyrosine kinase substrates and integrin signaling/adapter proteins (Akap13, Cdcp1, Ddef1, Eps15, Foxf1a, Gab2, Hs2st1, and Itgb4). Several of the P132L-specific genes are also highly expressed in stem/progenitor cells or are associated with myoepithelial cells, suggestive of an epithelial-mesenchymal transition. These results directly support clinical data showing that patients harboring Cav-1 mutations are more likely to undergo recurrence and metastasis.

Regulation of cell migration and survival by focal adhesion targeting of Lasp-1.

Large-scale proteomic and functional analysis of isolated pseudopodia revealed the Lim, actin, and SH3 domain protein (Lasp-1) as a novel protein necessary for cell migration, but not adhesion to, the extracellular matrix (ECM). Lasp-1 is a ubiquitously expressed actin-binding protein with a unique domain configuration containing SH3 and LIM domains, and is overexpressed in 8-12% of human breast cancers. We find that stimulation of nonmotile and quiescent cells with growth factors or ECM proteins facilitates Lasp-1 relocalization from the cell periphery to the leading edge of the pseudopodium, where it associates with nascent focal complexes and areas of actin polymerization. Interestingly, although Lasp-1 dynamics in migratory cells occur independently of c-Abl kinase activity and tyrosine phosphorylation, c-Abl activation by apoptotic agents specifically promotes phosphorylation of Lasp-1 at tyrosine 171, which is associated with the loss of Lasp-1 localization to focal adhesions and induction of cell death. Thus, Lasp-1 is a dynamic focal adhesion protein necessary for cell migration and survival in response to growth factors and ECM proteins.

The trefoil factor 1 participates in gastrointestinal cell differentiation by delaying G1-S phase transition and reducing apoptosis.

Trefoil factor (TFF)1 is synthesized and secreted by the normal stomach mucosa and by the gastrointestinal cells of injured tissues. The link between mouse TFF1 inactivation and the fully penetrant antropyloric tumor phenotype prompted the classification of TFF1 as a gastric tumor suppressor gene. Accordingly, altered expression, deletion, and/or mutations of the TFF1 gene are frequently observed in human gastric carcinomas. The present study was undertaken to address the nature of the cellular and molecular mechanisms targeted by TFF1 signalling. TFF1 effects were investigated in IEC18, HCT116, and AGS gastrointestinal cells treated with recombinant human TFF1, and in stably transfected HCT116 cells synthesizing constitutive or doxycycline-induced human TFF1. We observed that TFF1 triggers two types of cellular responses. On one hand, TFF1 lowers cell proliferation by delaying G1-S cell phase transition. This results from a TFF1-mediated increase in the levels of cyclin-dependent kinase inhibitors of both the INK4 and CIP subfamilies, leading to lower E2F transcriptional activity. On the other hand, TFF1 protects cells from chemical-, anchorage-free-, or Bad-induced apoptosis. In this process, TFF1 signalling targets the active form of caspase-9. Together, these results provide the first evidence of a dual antiproliferative and antiapoptotic role for TFF1. Similar paradoxical functions have been reported for tumor suppressor genes involved in cell differentiation, a function consistent with TFF1.

Pregnancy and post-partum breast cancer: a prospective study.

BACKGROUND: The concomitant occurrence of breast cancer and pregnancy remains a challenging clinical situation combining ethical and medical problems. There are few prospective data on pregnancy-associated breast cancer (PABC) whose incidence continuously increases. PATIENTS AND METHODS: Forty patients with PABC were compared with 61 non-pregnant, age-matched patients with infiltrative breast carcinomas (BC) diagnosed and followed since 1982. RESULTS: Although PABC and BC tumor size, grade and type, and lymphovascular and lymphnode invasion were similar, the BC cases showed better overall--(p=0.0001) and disease-free (p-0.015) survival. Moreover, the outcome of pregnant patients was worse than post-partum patients (p=0.017). Importantly, the number of PABC patients receiving hormonotherapy was lower than the BC patients (p<0.0004), due to lower estrogen receptor (ER) (p=0.038) and progesterone receptor (PR) (p=0.008) immunohisto-chemical (IH) levels. Retrospective estrogen-regulated pS2/trefoil factor 1 (pS2/TFF1) immunohistochemitry showed no difference between PABC and BC. All the children delivered were healthy. CONCLUSION: Pregnancy and the post-partum period increase breast cancer aggressiveness, pregnancy being the most detrimental. PABC hormone-dependence is under-estimated using ER and PR, and pS2/TFF1 might help in its determination. Appropriate treatment does not impair child outcome.

Copper-binding activity of Trefoil factor 1 (TFF1): a new perspective in the study of the multifunctional roles of TFFs.

Trefoil factors (TFFs) are gastrointestinal peptides playing an essential role in the epithelial restitution. Among the three known TFF peptides, TFF1 is characterized by three disulfide bonds producing a compact globular structure and an extended and disordered tail formed by amino- and carboxy-termini. The presence of a cysteine surrounded by several negatively charged residues in this region of the protein, highly conserved in different species, suggests the possible formation of a metal-binding site. Affinity chromatography and mass spectrometric analyses allowed us to demonstrate a selective binding affinity of TFF1 for copper. The binding induces conformational changes in the tertiary structure as demonstrated by circular dichroism experiments, while limited proteolysis revealed an altered access to the cleavage sites in the amino- and carboxy-termini. The results of this study reveal a new property of TFF1 and suggest that copper could influence its biological activities by interfering with the dimerization of the peptide and/or the interaction with mucins or putative TFF receptors.

Overexpression of the cholesterol-binding protein MLN64 induces liver damage in the mouse.

AIM: To examine the in vivo phenotype associated with hepatic metastatic lymph node 64 (MLN64) over-expression. METHODS: Recombinant-adenovirus-mediated MLN64 gene transfer was used to overexpress MLN64 in the livers of C57BL/6 mice. We measured the effects of MLN64 overexpression on hepatic cholesterol content, bile flow, biliary lipid secretion and apoptosis markers. For in vitro studies cultured CHO cells with transient MLN64 overexpression were utilized and apoptosis by TUNEL assay was measured. RESULTS: Livers from Ad.MLN64-infected mice exhibited early onset of liver damage and apoptosis. This response correlated with increases in liver cholesterol content and biliary bile acid concentration, and impaired bile flow. We investigated whether liver MLN64 expression could be modulated in a murine model of hepatic injury. We found increased hepatic MLN64 mRNA and protein levels in mice with chenodeoxycholic acid-induced liver damage. In addition, cultured CHO cells with transient MLN64 overexpression showed increased apoptosis. CONCLUSION: In summary, hepatic MLN64 over-expression induced damage and apoptosis in murine livers and altered cholesterol metabolism. Further studies are required to elucidate the relevance of these findings under physiologic and disease conditions.

TRAF4, the unique family member.

The fourth member of the TRAF protein family (TRAF4) presents several characteristics that distinguish it from the other members of the family. These characteristics concern the primary sequence of the protein, a strong evolutionary conservation, and a tightly regulated physiological expression during development. The subcellular localization of TRAF4 is controversial as it has been detected at the cell membrane, in the cytoplasm and in the nucleus. Using mouse and fly models, it has been established that TRAF4 is a key molecule in diverse ontogenic processes, particularly in the nervous system. However, the molecular mechanisms of action of TRAF4 remain evasive as it was found to interact with diverse types of proteins, leading either to pro-apoptotic or anti-apoptotic functions. Finally, few studies implicated TRAF4 in human diseases.

Stromelysin-3 is a potent negative regulator of adipogenesis participating to cancer cell-adipocyte interaction/crosstalk at the tumor invasive front.

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 gamma (PPARgamma) 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 PPARgamma 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 alpha-smooth muscle actin-expressing myofibroblasts. This constitutes the first report of implication of a MMP in cancer cell-adipocyte interaction/crosstalk during early steps of connective tissue invasion.

Cytotoxicity of a natural anthraquinone (Aloin) against human breast cancer cell lines with and without ErbB-2: topoisomerase IIalpha coamplification.

In the present study the cytotoxic activity of aloin, a natural anthracycline from Aloe plant, is reported against two human breast cancer cell lines; without (MCF-7) and with (SKBR-3) erbB-2-topoIIalpha coamplification. MCF-7cell line was shown to be more sensitive to aloin than SKBR-3 demonstrated by MTT and clonogenic assays, from which IC50 and 50% ICF values are reported to be 60 microg/ml, respectively, in the former cell line and as high as 150 and 80 microg/ml, respectively, in the latter, which are still far below the maximum tolerated dose of the compound. The effect of aloin is suggested to be brought about by more than one mechanism depending on the dose level and tumor phenotype. This was demonstrated by flow cytometric analysis, fluorescence microscopy and western blot analysis, which revealed that aloin at higher concentrations caused a reduction in the proportion of cells undergoing mitosis by induction of apoptosis, inhibition of topo II alpha protein expression and downregulation of cyclin B1 protein expression in MCF-7 cell line, whereas erbB-2 protein expression was not affected. Topo IIalpha protein expression was mildly downregulated in SKBR-3 cell line at higher concentrations only.

Cyclooxygenase-2 expression and effect of celecoxib in gastric adenomas of trefoil factor 1-deficient mice.

Expression of cyclooxygenase-2 (Cox-2) is elevated in gastric adenocarcinomas and precursor lesions leading to this disease. Mice deficient for trefoil factor 1 (TFF1) develop a pyloric adenoma with full penetrance. Because inhibition of Cox-2 suppresses tumor growth in several animal models, we studied expression of Cox-2 and effect of a selective Cox-2 inhibitor celecoxib in gastrointestinal tissues of the TFF1-deficient mice. Cox-2 mRNA and protein were strongly expressed in the pyloric adenomas of the TFF1(-/-) mice as detected by in situ hybridization and immunohistochemistry. Nonneoplastic gastrointestinal tissues of wild-type or TFF1(-/-) mice expressed low or nondetectable levels of Cox-2. Celecoxib (1600 ppm p.o. for 3 months) caused ulceration and inflammation of the adenoma in all treated TFF1(-/-) mice (n = 7). This effect of the drug was adenoma specific, because no histological alterations were observed in the non-neoplastic gastric or intestinal tissues in the TFF1(-/-) or wild-type mice receiving the drug treatment. All untreated TFF1(-/-) mice had an adenoma (n = 7), but none demonstrated the combination of ulceration and inflammation. Our data show that Cox-2 is expressed in gastric adenomas of the TFF1(-/-) mice and suggest that inhibition of Cox-2 disturbs the integrity of the adenoma by promoting ulceration and inflammation. These findings support the effort to initiate clinical studies to investigate the effect of Cox-2 inhibitors as a chemotherapeutic modality for dysplasias of the stomach.

Dual stromelysin-3 function during natural mouse mammary tumor virus-ras tumor progression.

In human carcinomas, stromelysin-3/matrix metalloproteinase 11 (ST3, MMP11) expression by nonmalignant fibroblastic cells located in the immediate vicinity of cancer cells is a bad prognostic factor. Using mouse models of primary tumors, it has been demonstrated that ST3 is a key player during local invasion, favoring cancer cell survival in connective tissue through an antiapoptotic function. To investigate the ST3 impact on additional phases of cancer cell invasion, we developed mammary gland cancer prone MMTV-ras transgenic mice in wild-type (ras+/+;ST3+/+) or ST3-deficient (ras+/+;ST3-/-) genotype and studied their whole natural cancer history. The tumor-free survival and delay between the first ras oncogenic hit and primary tumor appearance increased in ras+/+;ST3-/- mice (P < 0.000001 and <0.0000007, respectively). A systematic search for occult primary tumors and metastases revealed, in addition to a lower total number and size of primary tumors (P < 0.02), an unexpected higher number of metastases (P < 0.01) in ras+/+;ST3-/- mice. Moreover, for a similar number and size of primary invasive tumors, ras+/+;ST3-/- mice developed more metastases, indicating that the cancer cells evolving in ST3-deficient stroma have an increased potential to hematogenous dissemination. We conclude that the ST3 microenvironment is a consistently active partner of invading cancer cells but that its function differs throughout cancer progression, being tumor enhancer or repressor in processes leading to local or distal invasion. Such a dual effect for an MMP might shed light, at least partially, for the absence of survival benefit for patients included in anti-MMP clinical trials.