Extracellular Matrix Molecule tenascin-C Research Articles

Targeting the MAtrix REgulating MOtif abolishes several hallmarks of cancer, triggering antitumor immunity

Tumor-targeted therapies have often been inefficient due to the lack of concomitant control over the tumor microenvironment. Using an immunocompetent autologous breast cancer model, we investigated a MAtrix REgulating MOtif (MAREMO)-mimicking peptide, which inhibits the protumorigenic extracellular matrix (ECM) molecule tenascin-C that activates several cancer hallmarks. In cultured cells, targeting the MAREMO blocks tenascin-C signaling involved in cell adhesion and immune-suppression by inhibiting tenascin-C interactions with fibronectin, TGFβ, CXCL12, and others, thereby blocking downstream events. Using RNASequencing and various genetic, molecular, in situ, and in vivo assays, we demonstrate that the MAREMO peptide similarly blocks multiple tenascin-C functions in vivo. This includes releasing tumor-infiltrating leukocytes, including CD8+ T cells, from the stroma. The MAREMO peptide also triggers interferon signaling, restoring antitumor immunity, contributing to tumor growth inhibition and reduced dissemination. The MAREMO peptide targets tumor cells directly by promoting growth suppression and inhibiting phenotypic plasticity, subsequently enhancing responsiveness to the endogenous death inducer tumor necrosis factor-related apoptosis-inducing ligand, as shown by a loss-of-function approach. Moreover, the MAREMO peptide largely subdues the tumor bed by depleting fibroblasts, repressing tenascin-C and other ECM molecules, and restoring the function of the few remaining blood vessels. In conclusion, targeting tenascin-C with a MAREMO peptide represents a powerful anticancer strategy with a broad inhibition of several cancer hallmarks.

Advances on the roles of tenascin-C in cancer.

ABSTRACTThe roles of the extracellular matrix molecule tenascin-C (TNC) in health and disease have been extensively reviewed since its discovery over 40 years ago. Here, we will describe recent insights into the roles of TNC in tumorigenesis, angiogenesis, immunity and metastasis. In addition to high levels of expression in tumors, and during chronic inflammation, and bacterial and viral infection, TNC is also expressed in lymphoid organs. This supports potential roles for TNC in immunity control. Advances using murine models with engineered TNC levels were instrumental in the discovery of important functions of TNC as a danger-associated molecular pattern (DAMP) molecule in tissue repair and revealed multiple TNC actions in tumor progression. TNC acts through distinct mechanisms on many different cell types with immune cells coming into focus as important targets of TNC in cancer. We will describe how this knowledge could be exploited for cancer disease management, in particular for immune (checkpoint) therapies.

The extracellular matrix molecule tenascin-C modulates cell cycle progression and motility of adult neural stem/progenitor cells from the subependymal zone

Adult neurogenesis has been described in two canonical regions of the adult central nervous system (CNS) of rodents, the subgranular zone (SGZ) of the hippocampus and the subependymal zone (SEZ) of the lateral ventricles. The stem cell niche of the SEZ provides a privileged environment composed of a specialized extracellular matrix (ECM) that comprises the glycoproteins tenascin-C (Tnc) and laminin-1 (LN1). In the present study, we investigated the function of these ECM glycoproteins in the adult stem cell niche. Adult neural stem/progenitor cells (aNSPCs) of the SEZ were prepared from wild type (Tnc+/+) and Tnc knockout (Tnc−/−) mice and analyzed using molecular and cell biological approaches. A delayed maturation of aNSPCs in Tnc−/− tissue was reflected by a reduced capacity to form neurospheres in response to epidermal growth factor (EGF). To examine a potential influence of the ECM on cell proliferation, aNSPCs of both genotypes were studied by cell tracking using digital video microscopy. aNSPCs were cultivated on three different substrates, namely, poly-d-lysine (PDL) and PDL replenished with either LN1 or Tnc for up to 6 days in vitro. On each of the three substrates aNSPCs displayed lineage trees that could be investigated with regard to cell cycle length. The latter appeared reduced in Tnc−/− aNSPCs on PDL and LN1 substrates, less so on Tnc that seemed to compensate the absence of the ECM compound to some extent. Close inspection of the lineage trees revealed a subpopulation of late dividing aNSPCslate that engaged into cycling after a notable delay. aNSPCslate exhibited a clearly different morphology, with a larger cell body and conspicuous processes. aNSPCslate reiterated the reduction in cell cycle length on all substrates tested, which was not rescued on Tnc substrates. When the migratory activity of aNSPC-derived progeny was determined, Tnc−/− neuroblasts displayed significantly longer migration tracks. This was traced to an increased rate of migration episodes compared to the wild-type cells that rested for longer time periods. We conclude that Tnc intervenes in the proliferation of aNSPCs and modulates the motility of neuroblasts in the niche of the SEZ.

Modulating tenascin-C functions by targeting the MAtrix REgulating MOtif, “MAREMO”

The extracellular matrix molecule Tenascin-C (TNC) promotes cancer and chronic inflammation by multiple mechanisms. Recently, TNC was shown to promote an immune suppressive tumor microenvironment (TME) through binding soluble chemoattracting factors, thus retaining leukocytes in the stroma. TNC also binds to fibronectin (FN) and other molecules, raising the question of a potential common TNC binding mechanism. By sequence comparison of two TNC-interacting domains in FN, the fifth (FN5) and thirteenth (FN13) fibronectin type III domains we identified a MAtrix REgulating MOtif “MAREMO” or M-motif that is highly conserved amongst vertebrates. By sequence analysis, structural modeling and functional analysis we found also putative M-motifs in TNC itself. We showed by negative staining electron microscopic imaging that the M-motif in FN mediates interactions with FN as well as with TNC. We generated two M-motif mimetic peptides P5 and P13 resembling the M-motif in FN5 and FN13, respectively. By using structural information we modelled binding of these M-motif mimetics revealing a putative MAREMO binding site MBS in FN5 and TN3, respectively overlapping with the M-motif. We further demonstrated that the M-motif mimetic peptides blocked several functions of TNC, such as binding of TNC to FN, cell rounding on a mixed FN/TNC substratum, FN matrix expression and subsequent assembly, TNC-induced signaling and gene expression, TNC chemokine binding and dendritic cell retention, thus providing novel opportunities to inhibit TNC actions. Our results suggest that targeting the MAREMO/MBS interaction could be exploited for reducing inflammation and matrix functions in cancer and fibrosis.

The expression of tenascin-C in neural stem/progenitor cells is stimulated by the growth factors EGF and FGF-2, but not by TGF\u03b21

Neural stem/progenitor cells (NSPCs) rely on internal and external cues determining their lineage decisions during brain development. The progenitor cells of the embryonic mammalian forebrain reside in the ventricular and subventricular zones of the lateral ventricles, where they proliferate, generate neurons and glial cells, and respond to external cues like growth factors. The extracellular matrix (ECM) surrounds NSPCs and influences the cell fate by providing mechanical scaffold, trophic support, and instructive signals. The ECM molecule tenascin-C (Tnc) is expressed in the proliferative zones of the developing forebrain and involved in the proliferation and maturation of NSPCs. Here, we analyzed the regulation of the Tnc gene expression by NSPCs cultivated under the influence of different growth factors. We observed that the epidermal growth factor (EGF) and the fibroblast growth factor (FGF)-2 strongly increased the expression of Tnc, whereas the transforming growth factor (TGF)β 1 had no effect on Tnc gene expression, in contrast to previous findings in cell cultures of neural and non-neural origin. The stimulation of the Tnc gene expression induced by EGF or FGF-2 was reversible and seen in constantly treated as well as short term stimulated NSPC cultures. The activation depended on the presence of the respective receptors, which was slightly different in cortical and striatal NSPC cultures. Our results confirm the influence of extracellular stimuli regulating the expression of factors that form a niche for NSPCs during embryonic forebrain development.

Impact of Tenascin-C on Radiotherapy in a Novel Syngeneic Oral Squamous Cell Carcinoma Model With Spontaneous Dissemination to the Lymph Nodes.

Radiotherapy, the most frequent treatment of oral squamous cell carcinomas (OSCC) besides surgery is employed to kill tumor cells but, radiotherapy may also promote tumor relapse where the immune-suppressive tumor microenvironment (TME) could be instrumental. We established a novel syngeneic grafting model from a carcinogen-induced tongue tumor, OSCC13, to address the impact of radiotherapy on OSCC. This model revealed similarities with human OSCC, recapitulating carcinogen-induced mutations found in smoking associated human tongue tumors, abundant tumor infiltrating leukocytes (TIL) and, spontaneous tumor cell dissemination to the local lymph nodes. Cultured OSCC13 cells and OSCC13-derived tongue tumors were sensitive to irradiation. At the chosen dose of 2 Gy mimicking treatment of human OSCC patients not all tumor cells were killed allowing to investigate effects on the TME. By investigating expression of the extracellular matrix molecule tenascin-C (TNC), an indicator of an immune suppressive TME, we observed high local TNC expression and TIL infiltration in the irradiated tumors. In a TNC knockout host the TME appeared less immune suppressive with a tendency towards more tumor regression than in WT conditions. Altogether, our novel syngeneic tongue OSCC grafting model, sharing important features with the human OSCC disease could be relevant for future anti-cancer targeting of OSCC by radiotherapy and other therapeutic approaches.

Novel Human Tenascin-C Function-Blocking Camel Single Domain Nanobodies.

The extracellular matrix (ECM) molecule Tenascin-C (TNC) is well-known to promote tumor progression by multiple mechanisms. However, reliable TNC detection in tissues of tumor banks remains limited. Therefore, we generated dromedary single-domain nanobodies Nb3 and Nb4 highly specific for human TNC (hTNC) and characterized the interaction with TNC by several approaches including ELISA, western blot, isothermal fluorescence titration and negative electron microscopic imaging. Our results revealed binding of both nanobodies to distinct sequences within fibronectin type III repeats of hTNC. By immunofluroescence and immunohistochemical imaging we observed that both nanobodies detected TNC expression in PFA and paraffin embedded human tissue from ulcerative colitis, solid tumors and liver metastasis. As TNC impairs cell adhesion to fibronectin we determined whether the nanobodies abolished this TNC function. Indeed, Nb3 and Nb4 restored adhesion of tumor and mesangial cells on a fibronectin/TNC substratum. We recently showed that TNC orchestrates the immune-suppressive tumor microenvironment involving chemoretention, causing tethering of CD11c+ myeloid/dendritic cells in the stroma. Here, we document that immobilization of DC2.4 dendritic cells by a CCL21 adsorbed TNC substratum was blocked by both nanobodies. Altogether, our novel TNC specific nanobodies could offer valuable tools for detection of TNC in the clinical practice and may be useful to inhibit the immune-suppressive and other functions of TNC in cancer and other diseases.

Loss of the Extracellular Matrix Molecule Tenascin-C Leads to Absence of Reactive Gliosis and Promotes Anti-inflammatory Cytokine Expression in an Autoimmune Glaucoma Mouse Model.

Previous studies demonstrated that retinal damage correlates with a massive remodeling of extracellular matrix (ECM) molecules and reactive gliosis. However, the functional significance of the ECM in retinal neurodegeneration is still unknown. In the present study, we used an intraocular pressure (IOP) independent experimental autoimmune glaucoma (EAG) mouse model to examine the role of the ECM glycoprotein tenascin-C (Tnc). Wild type (WT ONA) and Tnc knockout (KO ONA) mice were immunized with an optic nerve antigen (ONA) homogenate and control groups (CO) obtained sodium chloride (WT CO, KO CO). IOP was measured weekly and electroretinographies were recorded at the end of the study. Ten weeks after immunization, we analyzed retinal ganglion cells (RGCs), glial cells, and the expression of different cytokines in retina and optic nerve tissue in all four groups. IOP and retinal function were comparable in all groups. Although RGC loss was less severe in KO ONA, WT as well as KO mice displayed a significant cell loss after immunization. Compared to KO ONA, less βIII-tubulin+ axons, and downregulated oligodendrocyte markers were noted in WT ONA optic nerves. In retina and optic nerve, we found an enhanced GFAP+ staining area of astrocytes in immunized WT. A significantly higher number of retinal Iba1+ microglia was found in WT ONA, while a lower number of Iba1+ cells was observed in KO ONA. Furthermore, an increased expression of the glial markers Gfap, Iba1, Nos2, and Cd68 was detected in retinal and optic nerve tissue of WT ONA, whereas comparable levels were observed in KO ONA. In addition, pro-inflammatory Tnfa expression was upregulated in WT ONA, but downregulated in KO ONA. Vice versa, a significantly increased anti-inflammatory Tgfb1 expression was measured in KO ONA animals. We conclude that Tnc plays an important role in glial and inflammatory response during retinal neurodegeneration. Our results provide evidence that Tnc is involved in glaucomatous damage by regulating retinal glial activation and cytokine release. Thus, this transgenic EAG mouse model for the first time offers the possibility to investigate IOP-independent glaucomatous damage in direct relation to ECM remodeling.

Tumor Penetrating Peptide-Functionalized Tenascin-C Antibody for Glioblastoma Targeting.

Conjugation to clinical-grade tumor penetrating iRGD peptide is a widely used strategy to improve tumor homing, extravasation, and penetration of cancer drugs and tumor imaging agents. The C domain of the extracellular matrix molecule Tenascin-C (TNC-C) is upregulated in solid tumors and represents an attractive target for clinical-grade single-chain antibody- based vehicles for tumor delivery drugs and imaging agents. To study the effect of C-terminal genetic fusion of the iRGD peptide to recombinant anti- TNC-C single-chain antibody clone G11 on systemic tumor homing and extravasation. Enzyme-linked immunosorbent assay was used to study the interaction of parental and iRGD-fused anti-TNC-C single-chain antibodies with C domain of tenascin-C and αVβ3 integrins. For systemic homing studies, fluorescein-labeled ScFV G11-iRGD and ScFV G11 antibodies were administered in U87-MG glioblastoma xenograft mice, and their biodistribution was studied by confocal imaging of tissue sections stained with markers of blood vessels and Tenascin C immunoreactivity. In a cell-free system, iRGD fusion to ScFV G11 conferred the antibody has a robust ability to bind αVβ3 integrins. The fluorescein labeling of ScFV G11-iRGD did not affect its target binding activity. In U87-MG mice, iRGD fusion to ScFV G11 antibodies improved their homing to tumor blood vessels, extravasation, and penetration of tumor parenchyma. The genetic fusion of iRGD tumor penetrating peptide to non-internalizing affinity targeting ligands may improve their tumor tropism and parenchymal penetration for more efficient delivery of imaging and therapeutic agents into solid tumor lesions.

Tenascin-C Orchestrates an Immune-Suppressive Tumor Microenvironment in Oral Squamous Cell Carcinoma.

Inherent immune suppression represents a major challenge in the treatment of human cancer. The extracellular matrix molecule tenascin-C promotes cancer by multiple mechanisms, yet the roles of tenascin-C in tumor immunity are incompletely understood. Using a 4NQO-induced oral squamous cell carcinoma (OSCC) model with abundant and absent tenascin-C, we demonstrated that tenascin-C enforced an immune-suppressive lymphoid stroma via CCL21/CCR7 signaling, leading to increased metastatic tumors. Through TLR4, tenascin-C increased expression of CCR7 in CD11c+ myeloid cells. By inducing CCL21 in lymphatic endothelial cells via integrin α9β1 and binding to CCL21, tenascin-C immobilized CD11c+ cells in the stroma. Inversion of the lymph node-to-tumor CCL21 gradient, recruitment of T regulatory cells, high expression of anti-inflammatory cytokines, and matrisomal components were hallmarks of the tenascin-C-instructed lymphoid stroma. Ablation of tenascin-C or CCR7 blockade inhibited the lymphoid immune-suppressive stromal properties, reducing tumor growth, progression, and metastasis. Thus, targeting CCR7 could be relevant in human head and neck tumors, as high tenascin-C expression and an immune-suppressive stroma correlate to poor patient survival.

Elimination of the four extracellular matrix molecules tenascin-C, tenascin-R, brevican and neurocan alters the ratio of excitatory and inhibitory synapses

The synaptic transmission in the mammalian brain is not limited to the interplay between the pre- and the postsynapse of neurons, but involves also astrocytes as well as extracellular matrix (ECM) molecules. Glycoproteins, proteoglycans and hyaluronic acid of the ECM pervade the pericellular environment and condense to special superstructures termed perineuronal nets (PNN) that surround a subpopulation of CNS neurons. The present study focuses on the analysis of PNNs in a quadruple knockout mouse deficient for the ECM molecules tenascin-C (TnC), tenascin-R (TnR), neurocan and brevican. Here, we analysed the proportion of excitatory and inhibitory synapses and performed electrophysiological recordings of the spontaneous neuronal network activity of hippocampal neurons in vitro. While we found an increase in the number of excitatory synaptic molecules in the quadruple knockout cultures, the number of inhibitory synaptic molecules was significantly reduced. This observation was complemented with an enhancement of the neuronal network activity level. The in vivo analysis of PNNs in the hippocampus of the quadruple knockout mouse revealed a reduction of PNN size and complexity in the CA2 region. In addition, a microarray analysis of the postnatal day (P) 21 hippocampus was performed unravelling an altered gene expression in the quadruple knockout hippocampus.

Tenascin-C increases lung metastasis by impacting blood vessel invasions

Metastasis is a major cause of death in cancer patients. The extracellular matrix molecule tenascin-C is a known promoter of metastasis, however the underlying mechanisms are not well understood. To further analyze the impact of tenascin-C on cancer progression we generated MMTV-NeuNT mice that develop spontaneous mammary tumors, on a tenascin-C knockout background. We also developed a syngeneic orthotopic model in which tumor cells derived from a MMTV-NeuNT tumor. Tumor cells were transfected with control shRNA or with shRNA to knockdown tenascin-C expression and, were grafted into the mammary gland of immune competent, wildtype or tenascin-C knockout mice. We show that stromal-derived tenascin-C increases metastasis by reducing apoptosis and inducing the cellular plasticity of cancer cells located in pulmonary blood vessels invasions (BVI), before extravasation. We characterized BVI as organized structures of tightly packed aggregates of proliferating tumor cells with epithelial characteristics, surrounded by Fsp1+ cells, internally located platelets and, a luminal monolayer of endothelial cells. We found extracellular matrix, in particular, tenascin-C, between the stromal cells and the tumor cell cluster. In mice lacking stromal-derived tenascin-C, the organization of pulmonary BVI was significantly affected, revealing novel functions of host-derived tenascin-C in supporting the integrity of the endothelial cell coat, increasing platelet abundance, tumor cell survival, epithelial plasticity, thereby promoting overall lung metastasis. Many effects of tenascin-C observed in BVI including enhancement of cellular plasticity, survival and migration, could be explained by activation of TGF-β signaling. Finally, in several human cancers, we also observed BVI to be surrounded by an endothelial monolayer and to express tenascin-C. Expression of tenascin-C is specific to BVI and is not observed in lymphatic vascular invasions frequent in breast cancer, which lack an endothelial lining. Given that BVI have prognostic significance for many tumor types, such as shorter cancer patient survival, increased metastasis, vessel occlusion, and organ failure, our data revealing a novel mechanism by which stromal tenascin-C promotes metastasis in human cancer, may have potential for diagnosis and therapy.

Serum Levels and Urinary Excretion of Tenascin-C and TIMP-1 in Acute Kidney Injury.

Both the extracellular matrix molecule tenascin-C (Tn-C) and tissue inhibitors of metalloproteinases (TIMPs) have a role in tissue injury, inflammation, and remodeling. In this pilot study, we tried to evaluate the role of these markers in acute kidney injury (AKI). A total of 52 subjects were enrolled in this study. Group 1 consisted of 27 patients with AKI (stage 1, 2, and 3), and Group 2 consisted of 25 age- and gender-matched healthy subjects. Serum and urine samples (to determine Tn-C and TIMP-1) were obtained from the participants at the beginning of the study. Second samples were obtained from Group 1 patients when renal function improved (at discharge). Serum TIMP-1 concentrations (admission and discharge) were higher in Group 1 than Group 2 (p = 0.0001 for both comparisons). Tn-C excretion in spot urine was significantly higher in healthy controls than at the admission levels of the patient group (p = 0.036). However, TIMP-1 excretion in spot urine was lower in healthy controls than in admission and discharge levels of the patient group (p = 0.0001 for both comparisons). Our results show that these biomarkers (especially TIMP-1) may have a role in the pathophysiology of AKI. Further studies are needed in this field.

Investigating cell-type specific functions of tenascin-C.

The extracellular matrix molecule tenascin-C (TNC) has received a lot of attention since its discovery 30 years ago because of its multiple roles in tissue repair, and in pathologies such as chronic inflammation, fibrosis, and cancer. Mouse models with high or no TNC expression have enabled the validation of key roles for TNC in immunity and angiogenesis. In parallel, many approaches including primary cell or organ cultures have shed light on the cellular and molecular mechanisms by which TNC exerts its multiple actions in vivo. Here, we will describe assays that investigate its antiadhesive properties and that measure the effect of TNC on the actin cytoskeleton, cell survival, proliferation, and migration. We will also describe assays to assess the impact of TNC on endothelial and immune cells in cell and organ culture, and to compare the responses of fibroblasts from normal and diseased tissues.

How to detect and purify tenascin-C.

The extracellular matrix molecule tenascin-C (TNC) was discovered over 30 years ago, and its tightly regulated pattern of expression since sparked keen interest in the scientific community. In adult tissues, TNC expression is restricted to specific niches and areas of active remodeling or high mechanical strain. However, while most healthy tissues contain little TNC, its transient expression upon cellular stress or tissue injury helps to mediate repair and restore homeostasis. Persistent expression of TNC is associated with chronic inflammation, fibrosis, and cancer, where methods for its detection are emerging as a reliable means to predict disease onset, prognosis, and response to treatment. Because studying the expression of this large matrix molecule is not always straightforward, here we describe basic techniques to examine tissue levels of TNC mRNA and protein. We also describe methods for purifying recombinant TNC, knocking down its expression, and creating cell-derived matrices with or without TNC within.

Spatial organization of the tenascin-C microenvironment in experimental and human cancer

The extracellular matrix (ECM) molecule tenascin-C (TNC) promotes tumor progression. This has recently been demonstrated in the stochastic murine RIP1-Tag2 insulinoma model, engineered to either express TNC abundantly or to be devoid of TNC. However, our knowledge about organization of the TNC microenvironment is scant. Here we determined the spatial distribution of TNC together with other ECM molecules in murine RIP1-Tag2 insulinoma and human cancer tissue (insulinoma and colorectal carcinoma). We found that TNC is organized in matrix tracks together with other ECM molecules of the AngioMatrix signature, a previously described gene expression profile that characterizes the angiogenic switch. Moreover, stromal cells including endothelial cells, fibroblasts and leukocytes were enriched in the TNC tracks. Thus, TNC tracks may provide niches for stromal cells and regulate their behavior. Given similarities of TNC rich niches for stromal cells in human insulinoma and colon cancer, we propose that the RIP1-Tag2 model may be useful for providing insights into the contribution of the tumor stroma specific ECM as promoter of cancer progression.

Tenascin-C Downregulates Wnt Inhibitor Dickkopf-1, Promoting Tumorigenesis in a Neuroendocrine Tumor Model

The extracellular matrix molecule tenascin-C (TNC) is a major component of the cancer-specific matrix, and high TNC expression is linked to poor prognosis in several cancers. To provide a comprehensive understanding of TNC's functions in cancer, we established an immune-competent transgenic mouse model of pancreatic β-cell carcinogenesis with varying levels of TNC expression and compared stochastic neuroendocrine tumor formation in abundance or absence of TNC. We show that TNC promotes tumor cell survival, the angiogenic switch, more and leaky vessels, carcinoma progression, and lung micrometastasis. TNC downregulates Dickkopf-1 (DKK1) promoter activity through the blocking of actin stress fiber formation, activates Wnt signaling, and induces Wnt target genes in tumor and endothelial cells. Our results implicate DKK1 downregulation as an important mechanism underlying TNC-enhanced tumor progression through the provision of a proangiogenic tumor microenvironment.

The role of Tenascin C in the lymphoid progenitor cell niche

Hemopoietic stem cells (HSCs) are extrinsically controlled by the bone marrow (BM) microenvironment. Mice devoid of the extracellular matrix molecule Tenascin-C (TNC) were reported to develop normally. The current study explores the relationship between TNC and hemopoiesis, from HSCs within their niche to maturing progenitors in alternate niches. Although the absence of TNC did not alter the size of the BM stem cell pool, we report decreased thymic T cell progenitors with redistribution to other lymphoid organs, suggesting an anchoring role for TNC. TNC did not play an essential role in stem and progenitor cell homing to BM, but significantly altered lymphoid primed progenitor cell homing. These cells express the TNC receptor, integrin α9β1, with the same reduced homing evident in the absence of this integrin. The absence of TNC also resulted in an increased proportion and number of mature circulating T cells. In addition, the absence of TNC significantly impaired hemopoietic reconstitution after transplant and increased stem and progenitor cell mobilization. In summary, our analysis revealed unidentified roles for TNC in hemopoiesis: in lineage commitment of thymic T cell progenitors, peripheral T cell migration, and hemopoietic reconstitution.

Abstract 429: The value of Tenascin-C expression in neoadjuvant breast cancer therapy

Abstract Introduction: The extra cellular matrix molecule Tenascin-C (TNC) in its many isoforms is highly expressed during tissue repair and pathological conditions such as cancer and stress. Utilising the selective pressure of neoadjuvant chemotherapy on carcinoma cells we screened for a subgroup of breast cancer patients with elevated TNC expression and a bad therapy response. TNC expression was correlated with biomarkers, histopathological (stage) and clinical data. Breast cancer cell lines were tested on TNC levels and their expression behaviour under competitive conditions. Methods: Paraffin sections from 68 preoperatively treated breast cancer patients were used for automated immunohistochemistry (IHC) with antibodies targeting TNC, AE1/AE3, Ki67, CD3 and p53. In addition F-18 FDG PET-CT was performed to evaluate therapy response. Protein- and gene expression experiments performed onto six breast cancer cell lines (MCF-7, MDA-MB231, MDA-MB453, SK-BR3, SUM-159, T47-D) showed TNC-expression in MDA-MB453 breast cancer cell line. Gene expression of TNC in hedgehog positive cells (MCF-7) was evaluated after co-cultivation with TNC positive cells (MDA-MB453) by qRT-PCR. The qRT-PCR results confirmed the protein expression results on in vitro stained cell-slides after cell-cell interactions and different growth times. Growth and migration behaviour was studied in 3D-matrigels. Results: We demonstrate that TNC expression in breast cancer is immensely reduced after neoadjuvant chemotherapy (43% of patients lost TNC expression) and that expression profiles after treatment correlate with tumor grade, lymph node status and estrogen receptor status, indicating prognostic value, probably for endocrine treatment. Post-therapeutical correlation and co-localisation of the proliferation marker Ki67 in cancer cells and TNC in the surrounding tumor-stroma of patients having a less beneficial response to chemotherapy. The evaluation of expression levels revealed a linear correlation of TNC and Ki67. Submitting cell lines to selective environmental pressure on cell chamber slides, we observed differentiated staining patterns and cellular localization for TNC. After co-cultivation with MDA-MB453, the hedgehog positive cell line MCF-7 expressed highly elevated levels of TNC. Conclusions: These findings concur with the role of TNC in tissue reorganisation, wound healing and it's growth promoting properties in cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 429. doi:10.1158/1538-7445.AM2011-429

The role of tenascin-C in tissue injury and tumorigenesis

The extracellular matrix molecule tenascin-C is highly expressed during embryonic development, tissue repair and in pathological situations such as chronic inflammation and cancer. Tenascin-C interacts with several other extracellular matrix molecules and cell-surface receptors, thus affecting tissue architecture, tissue resilience and cell responses. Tenascin-C modulates cell migration, proliferation and cellular signaling through induction of pro-inflammatory cytokines and oncogenic signaling molecules amongst other mechanisms. Given the causal role of inflammation in cancer progression, common mechanisms might be controlled by tenascin-C during both events. Drugs targeting the expression or function of tenascin-C or the tenascin-C protein itself are currently being developed and some drugs have already reached advanced clinical trials. This generates hope that increased knowledge about tenascin-C will further improve management of diseases with high tenascin-C expression such as chronic inflammation, heart failure, artheriosclerosis and cancer.