The urokinase receptor-derived cyclic peptide [SRSRY] suppresses neovascularization and intravasation of osteosarcoma and chondrosarcoma cells.
ABSTRACT: The receptor for the urokinase-type plasminogen activator (uPAR) is a widely recognized master regulator of cell migration and uPAR88-92 is the minimal sequence required to induce cell motility and angiogenesis by interacting with the formyl peptide receptor type 1 (FPR1). In this study, we present evidence that the cyclization of the uPAR88-92 sequence generates a new potent inhibitor of migration, and extracellular matrix invasion of human osteosarcoma and chondrosarcoma cells expressing comparable levels of FPR1 on cell surface. In vitro, the cyclized peptide [SRSRY] prevents formation of capillary-like tubes by endothelial cells co-cultured with chondrosarcoma cells and trans-endothelial migration of osteosarcoma and chondrosarcoma cells. When chondrosarcoma cells were subcutaneously injected in nude mice, tumor size, intra-tumoral microvessel density and circulating tumor cells in blood samples collected before the sacrifice, were significantly reduced in animals treated daily with i.p-administration of 6 mg/Kg [SRSRY] as compared to animals treated with vehicle only. Our findings indicate that [SRSRY] prevents three key events occurring during the metastatic process of osteosarcoma and chondrosarcoma cells: the extracellular matrix invasion, the formation of a capillary network and the entry into bloodstream.
Project description:The receptor for the urokinase-type plasminogen activator (uPAR) is a widely recognized master regulator of cell migration and uPAR88-92 is the minimal sequence required to induce cell motility. We and others have previously documented that the uPAR88-92 sequence, even in the form of synthetic linear peptide (SRSRY), interacts with the formyl peptide receptor type 1 (FPR1), henceforth inducing cell migration of several cell lines, including monocytes. FPR1 is mainly expressed by mammalian phagocytic leukocytes and plays a crucial role in chemotaxis. In this study, we present evidence that the cyclization of the SRSRY sequence generates a new potent and stable inhibitor of monocyte trafficking. In rat basophilic leukaemia RBL-2H3/ETFR cells expressing high levels of constitutively activated FPR1, the cyclic SRSRY peptide ([SRSRY]) blocks FPR1 mediated cell migration by interfering with both internalization and ligand-uptake of FPR1. Similarly to RBL-2H3/ETFR cells, [SRSRY] competes with fMLF for binding to FPR1 and prevents agonist-induced FPR1 internalization in human monocyte THP-1 cells. Unlike scramble [RSSYR], [SRSRY] inhibits fMLF-directed migration of monocytes in a dose-dependent manner, with IC50 value of 0.01 nM. PMA-differentiated THP-1 cell exposure to fMLF gradient causes a marked cytoskeletal re-organization with the formation of F-actin rich pseudopodia that are prevented by the addition of [SRSRY]. Furthermore, [SRSRY] prevents migration of human primary monocytes and trans-endothelial migration of monocytes. Our findings indicate that [SRSRY] is a new FPR1 inhibitor which may suggest the development of new drugs for treating pathological conditions sustained by increased motility of monocytes, such as chronic inflammatory diseases.
Project description:Chondrosarcoma is the second most prevalent general primary tumor of bone following osteosarcoma. Chondrosarcoma development may be linked to angiogenesis, which is principally elicited by vascular endothelial growth factor-A (VEGF-A). VEGF-A level has been recognized as a prognostic marker in angiogenesis. WNT1-inducible signaling pathway protein-3 (WISP)-3/CCN6 belongs to the CCN family and is involved in regulating several cellular functions, including cell proliferation, differentiation, and migration. Nevertheless, the effect of WISP-3 on VEGF-A production and angiogenesis in human chondrosarcoma remains largely unknown. This current study shows that WISP-3 promoted VEGF-A production and induced angiogenesis of human endothelial progenitor cells. Moreover, WISP-3-enhanced VEGF-A expression and angiogenesis involved the c-Src and p38 signaling pathways, while miR-452 expression was negatively affected by WISP-3 via the c-Src and p38 pathways. Our results illustrate the clinical significance of WISP-3, VEGF-A and miR-452 in human chondrosarcoma patients. WISP-3 may illustrate a novel therapeutic target in the metastasis and angiogenesis of chondrosarcoma.
Project description:Chondrosarcoma is a type of highly malignant tumor with a potent capacity for local invasion and causing distant metastasis. Chondrosarcoma shows a predilection for metastasis to the lungs. Hepatocyte growth factor (HGF) has been demonstrated to stimulate cancer proliferation, migration, and metastasis. However, the effect of HGF on migration activity of human chondrosarcoma cells is not well known. Here, we found that human chondrosarcoma tissues demonstrated significant expression of HGF, which was higher than that in normal cartilage. We also found that HGF increased the migration and expression of matrix metalloproteinase (MMP)-2 in human chondrosarcoma cells. c-Met inhibitor and siRNA reduced HGF-increased cell migration and MMP-2 expression. HGF treatment resulted in activation of the phosphatidylinositol 3'-kinase (PI3K)/Akt/PKC?/NF-?B pathway, and HGF-induced expression of MMP-2 and cell migration was inhibited by specific inhibitors or siRNA-knockdown of PI3K, Akt, PKC?, and NF-?B cascades. Taken together, our results indicated that HGF enhances migration of chondrosarcoma cells by increasing MMP-2 expression through the c-Met receptor/PI3K/Akt/PKC?/NF-?B signal transduction pathway.
Project description:Chondrosarcoma is a primary malignant bone cancer, with a potent capacity to invade locally and cause distant metastasis; it has a poor prognosis and shows a predilection for metastasis to the lungs. Brain derived neurotrophic factor (BDNF) is a small-molecule protein from the neurotrophin family of growth factors that is associated with the disease status and outcomes of cancers. However, the effect of BDNF on migration activity in human chondrosarcoma cells is mostly unknown. Here, we found that human chondrosarcoma tissues showed significant expression of BDNF, which was higher than that in normal cartilage and primary chondrocytes. We also found that BDNF increased the migration and expression of ?5 integrin in human chondrosarcoma cells. In addition, knockdown of BDNF expression markedly inhibited migratory activity. BDNF-mediated migration and ?5 integrin up-regulation were attenuated by antibody, inhibitor, or siRNA against the TrkB receptor. Pretreatment of chondrosarcoma cells with PI3K, Akt, and NF-?B inhibitors or mutants also abolished BDNF-promoted migration and integrin expression. The PI3K, Akt, and NF-?B signaling pathway was activated after BDNF treatment. Taken together, our results indicate that BDNF enhances the migration of chondrosarcoma by increasing ?5 integrin expression through a signal transduction pathway that involves the TrkB receptor, PI3K, Akt, and NF-?B. BDNF thus represents a promising new target for treating chondrosarcoma metastasis.
Project description:Chondrosarcomas are primary malignant bone tumors that have a poor prognosis. WNT1-inducible signaling pathway protein-3 (WISP-3, also termed CCN6) belongs to the CCN family of proteins and is implicated in the regulation of various cellular functions, such as cell proliferation, differentiation, and migration. It is unknown as to whether CCN6 affects human chondrosarcoma metastasis. We show how CCN6 promotes chondrosarcoma cell migration and invasion via matrix metallopeptidase-9 (MMP)-9 expression. These effects were abolished by pretreatment of chondrosarcoma cells with PI3K, Akt, mTOR, and NF-?B inhibitors or short interfering (si)RNAs. Our investigations indicate that CCN6 facilitates metastasis through the PI3K/Akt/mTOR/NF-?B signaling pathway. CCN6 and MMP-9 expression was markedly increased in the highly migratory JJ012(S10) cell line compared with the primordial cell line (JJ012) in both in vitro and in vivo experiments. CCN6 knockdown suppressed MMP-9 production in JJ012(S10) cells and attenuated cell migration and invasion ability. Importantly, CCN6 knockdown profoundly inhibited chondrosarcoma cell metastasis to lung. Our findings reveal an important mechanism underlying CCN6-induced metastasis and they highlight the clinical significance between CCN6 and MMP-9 in regard to human chondrosarcoma. CCN6 appears to be a promising therapeutic target in chondrosarcoma metastasis.
Project description:BACKGROUND: Cyclooxygenase (COX)-2, the inducible isoform of prostaglandin (PG) synthase, has been implicated in tumor metastasis. Interaction of COX-2 with its specific EP receptors on the surface of cancer cells has been reported to induce cancer invasion. However, the effects of COX-2 on migration activity in human chondrosarcoma cells are mostly unknown. In this study, we examined whether COX-2 and EP interaction are involved in metastasis of human chondrosarcoma. RESULTS: We found that over-expression of COX-2 or exogenous PGE2 increased the migration of human chondrosarcoma cells. We also found that human chondrosarcoma tissues and chondrosarcoma cell lines had significant expression of the COX-2 which was higher than that in normal cartilage. By using pharmacological inhibitors or activators or genetic inhibition by the EP receptors, we discovered that the EP1 receptor but not other PGE receptors is involved in PGE2-mediated cell migration and alpha2beta1 integrin expression. Furthermore, we found that human chondrosarcoma tissues expressed a higher level of EP1 receptor than normal cartilage. PGE2-mediated migration and integrin up-regulation were attenuated by phospholipase C (PLC), protein kinase C (PKC) and c-Src inhibitor. Activation of the PLCbeta, PKCalpha, c-Src and NF-kappaB signaling pathway after PGE2 treatment was demonstrated, and PGE2-induced expression of integrin and migration activity were inhibited by the specific inhibitor, siRNA and mutants of PLC, PKC, c-Src and NF-kappaB cascades. CONCLUSIONS: Our results indicated that PGE2 enhances the migration of chondrosarcoma cells by increasing alpha2beta1 integrin expression through the EP1/PLC/PKCalpha/c-Src/NF-kappaB signal transduction pathway.
Project description:Accumulating evidence indicates that dysregulation of miRNAs could contribute to tumor growth and metastasis of chondrosarcoma by infuencing cell proliferation and invasion. In the current study, we are interested to examine the role of miRNAs in the carcinogenesis and progression of chondrosarcoma. Here, using comparative miRNA profiling of tissues and cells of chondrosarcoma and cartilage, we identified miR-494 as a commonly downregulated miRNA in the tissues of patients with chondrosarcoma and chondrosarcoma cancer cell line, and upregulation of miR-494 could inhibit proliferation and invasion of chondrosarcoma cancer cells in vivo and in vitro. Moreover, our data demonstrated that SOX9, the essential regulator of the process of cartilage differentiation, was the direct target and functional mediator of miR-494 in chondrosarcoma cells. And downregulation of SOX9 could also inhibit migration and invasion of chondrosarcoma cells. In the last, we identified low expression of miR-494 was significantly correlated with poor overall survival and prognosis of chondrosarcoma patients. Thus, miR-494 may be a new common therapeutic target and prognosis biomarker for chondrosarcoma.
Project description:Resistin is a recently discovered adipocyte-secreting adipokine, which may play a critical role in modulating cancer pathogenesis. Chondrosarcoma is a highly malignant tumor known to frequently metastasize; however, the role of resistin in the metastasis of human chondrosarcoma is largely unknown. Here, we found that the expression of resistin was higher in chondrosarcoma biopsy tissues than in normal cartilage. Moreover, treatment with resistin increased matrix metalloproteinase (MMP)-2 expression and promoted cell migration in human chondrosarcoma cells. Co-transfection with microRNA (miR)-519d mimic resulted in reversed resistin-mediated cell migration and MMP-2 expression. Additionally, AMP-activated protein kinase (AMPK) and p38 inhibitors or siRNAs reduced the resistin-increased cell migration and miR-519d suppression, and inhibition of resistin expression resulted in suppression of MMP-2 expression and lung metastasis in vivo. Taken together, our results indicate that resistin promotes chondrosarcoma metastasis and MMP-2 expression through activation of the AMPK/p38 signaling pathway and down-regulation of miR-519d expression. Therefore, resistin may represent a potential novel molecular therapeutic target in chondrosarcoma metastasis.
Project description:Chondrosarcoma is a heterogeneous collection of malignant bone tumors and is the second most common primary malignancy of bone after osteosarcoma. Recent work has identified frequent, recurrent mutations in IDH1 or IDH2 in nearly half of central chondrosarcomas. However, there has been little systematic genomic analysis of this tumor type, and, thus, the contribution of other genes is unclear. Here we report comprehensive genomic analyses of 49 individuals with chondrosarcoma (cases). We identified hypermutability of the major cartilage collagen gene COL2A1, with insertions, deletions and rearrangements identified in 37% of cases. The patterns of mutation were consistent with selection for variants likely to impair normal collagen biosynthesis. In addition, we identified mutations in IDH1 or IDH2 (59%), TP53 (20%), the RB1 pathway (33%) and Hedgehog signaling (18%).
Project description:Chemokine (C-C motif) ligand 3 (CCL3), also known as macrophage inflammatory protein-1?, is a cytokine involved in inflammation and activation of polymorphonuclear leukocytes. CCL3 has been detected in infiltrating cells and tumor cells. Chondrosarcoma is a highly malignant tumor that causes distant metastasis. However, the effect of CCL3 on human chondrosarcoma metastasis is still unknown. Here, we found that CCL3 increased cellular migration and expression of matrix metalloproteinase (MMP)-2 in human chondrosarcoma cells. Pre-treatment of cells with the MMP-2 inhibitor or transfection with MMP-2 specific siRNA abolished CCL3-induced cell migration. CCL3 has been reported to exert its effects through activation of its specific receptor, CC chemokine receptor 5 (CCR5). The CCR5 and AMP-activated protein kinase (AMPK) inhibitor or siRNA also attenuated CCL3-upregulated cell motility and MMP-2 expression. CCL3-induced expression of MMP-2 and migration were also inhibited by specific inhibitors, and inactive mutants of AMPK, p38 mitogen activated protein kinase (p38 or p38-MAPK), and nuclear factor ?B (NF-?B) cascades. On the other hand, CCL3 treatment demonstrably activated AMPK, p38, and NF-?B signaling pathways. Furthermore, the expression levels of CCL3, CCR5, and MMP-2 were correlated in human chondrosarcoma specimens. Taken together, our results indicate that CCL3 enhances the migratory ability of human chondrosarcoma cells by increasing MMP-2 expression via the CCR5, AMPK, p38, and NF-?B pathways.