sq-23377 and Neoplasm-Metastasis

Upon activation by its ligand hepatocyte growth factor/scatter factor, the receptor tyrosine kinase Met promotes survival, proliferation, and migration of epithelial cells during embryogenesis. Deregulated Met signaling can also promote cancer progression and metastasis. Met belongs to the functional family of dependence receptors whose activity switches from pro-survival to pro-apoptotic during apoptosis upon caspase cleavage. Although apoptosis resistance is a hallmark of cancer cells, some remain sensitive to other cell death processes, including necrosis induced by calcium stress. The role and fate of Met during necrotic cell death are unknown. Following treatment with calcium ionophores, cell lines and primary cells undergo necrosis, and the full-length Met receptor is efficiently degraded. This degradation is achieved by double cleavage of Met in its extracellular domain by a metalloprotease of the A disintegrin and metalloproteinase (ADAM) family and in its intracellular domain by calpains (calcium-dependent proteases). These cleavages separate the Met extracellular region from its kinase domain, thus preventing Met activity and its potential pro-survival activity. Although the intracellular fragment is very similar to the fragment generated by caspases, it displays no pro-apoptotic property, likely because of the presence of the last few amino acids of Met, known to inhibit this pro-apoptotic function. The fragments identified here are observed in lung tumors overexpressing the Met receptor, along with fragments previously identified, suggesting that proteolytic cleavages of Met are involved in its degradation in tumor tissues. Thus, Met is a modulator of necrosis, able to protect cells when activated by its ligand but efficiently degraded by proteolysis when this process is engaged.

Topics: ADAM Proteins; Animals; Apoptosis; Calcium; Calpain; Caspases; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cell Survival; Enzyme Activation; Epithelial Cells; HEK293 Cells; Hepatocyte Growth Factor; Humans; Ionomycin; Lung Neoplasms; Mice; Necrosis; Neoplasm Metastasis; Proto-Oncogene Proteins c-met; RNA Interference; RNA, Small Interfering; Signal Transduction

Despite lots of research efforts, the pathology of head and neck cancer remains elusive. Accumulating evidence suggests that the innate and adaptive immunity plays an important role in HNSCC (Head and Neck Squamous Cell Carcinoma) development. Recently, a new T helper cell subset additional to the classical Th1 and Th2 cells was identified called Th17 cells, due to their secretion of IL-17. However, Th17 cells also produce additional proinflammatory cytokines and many other cytokines are involved in their differentiation and expansion. It was shown that Th17 cells play a prominent role in host defense but are also associated with the development of autoimmune diseases. The role of Th17 cells in cancer pathogenesis remains nebulous.. Th17 cells of peripheral blood, primary tumors and metastatic lymph nodes were FACS analyzed for their CD161 expression. Supernatants of the permanent HNSCC cell line BHY were used to induce Th17 cells by HNSCC tumor mileu.. Here we show that Th17 cells from patients with HNSCC downregulate the Th17 cell surface receptor CD161 in peripheral blood as well as in primary tumors and especially in metastatic lymph nodes.. We have showed for the first time alterations of Th17 cell phenotype in HNSCC patients.

Topics: Aged; Brefeldin A; Carcinoma, Squamous Cell; Culture Media, Conditioned; Head and Neck Neoplasms; Humans; Ionomycin; Leukocytes, Mononuclear; Lymph Nodes; Lymphocyte Activation; Middle Aged; Neoplasm Metastasis; NK Cell Lectin-Like Receptor Subfamily B; Tetradecanoylphorbol Acetate; Th17 Cells

The NFAT (nuclear factor of activated T cells) family of transcription factors plays a fundamental role in the transcriptional regulation of the immune response. However, NFATs are ubiquitously expressed, and recent evidence points to their important functions in human epithelial cells and carcinomas. Specifically, NFAT has been shown to be active in human breast and colon carcinoma cells and to promote their invasion through Matrigel. The mechanisms by which NFAT promotes invasion have not been defined. To identify NFAT target genes that induce carcinoma invasion, we have established stable breast cancer cell lines that inducibly express transcriptionally active NFAT. Gene expression profiling by cDNA microarray of cells induced to express NFAT revealed up-regulation of cyclooxygenase-2 (COX-2). Increased NFAT expression and activity induced COX-2 expression as well as prostaglandin E2 synthesis. This induction was more prominent when NFAT was activated by phorbol 12-myristate 13-acetate and calcium ionophore ionomycin and was blocked by the NFAT antagonist cyclosporin A. Breast cancer cells with elevated COX-2 expression showed increased invasion through Matrigel, and this was reduced in cells treated with COX-2 inhibitors. Conversely, loss of NFAT1 protein expression using small interfering RNA led to a reduction in COX-2 transcription and reduced invasion. Similarly, Matrigel invasion was reduced in cells in which COX-2 expression was reduced using specific siRNA. These findings demonstrate that NFAT promotes breast cancer cell invasion through the induction of COX-2 and the synthesis of prostaglandins.

Topics: Breast Neoplasms; Cell Line, Tumor; Collagen; Cyclooxygenase 2; Cyclosporine; Dinoprostone; Drug Combinations; Humans; Ionomycin; Ionophores; Laminin; Neoplasm Invasiveness; Neoplasm Metastasis; NFATC Transcription Factors; Proteoglycans; RNA, Small Interfering; Tetradecanoylphorbol Acetate

We have isolated a novel soluble factor(s), neutrophil activator of matrix metalloproteinases (NAM), secreted by unstimulated normal human peripheral blood neutrophils that causes the activation of cell secreted promatrix metalloproteinase-2 (proMMP-2). Partially purified preparations of NAM have been isolated from the conditioned media of neutrophils employing gelatin-Sepharose chromatography and differential membrane filter centrifugation. NAM activity, as assessed by exposing primary human umbilical vein endothelial cells (HUVEC) or HT1080 cells to NAM followed by gelatin zymography, was seen within one hour. Tissue inhibitor of metalloproteinase-2 (TIMP-2) and hydroxamic acid derived inhibitors of MMPs (CT1746 and BB94) abrogated the activation of proMMP-2 by NAM, while inhibitors of serine and cysteine proteases showed no effect. NAM also produced an increase in TIMP-2 binding to HUVEC and HT1080 cell surfaces that was inhibited by TIMP-2, CT1746, and BB94. Time-dependent increases in MT1-MMP protein and mRNA were seen following the addition of NAM to cells. These data support a role for NAM in cancer dissemination.

Topics: Adult; Amides; Calcimycin; Cathepsin G; Cathepsins; Cells, Cultured; Culture Media, Conditioned; Endothelium, Vascular; Enzyme Activation; Enzyme Precursors; Humans; Inflammation; Ionomycin; Matrix Metalloproteinase 14; Matrix Metalloproteinase 2; Neoplasm Invasiveness; Neoplasm Metastasis; Neutrophils; Oligopeptides; Pancreatic Elastase; Phenylalanine; Protease Inhibitors; Proteins; RNA, Messenger; Serine Endopeptidases; Substrate Specificity; Tetradecanoylphorbol Acetate; Thiophenes; Tissue Inhibitor of Metalloproteinase-2; Umbilical Veins

Lymphocytes obtained from tumor-draining lymph nodes (DLN) can have potent in vivo antitumor activity after in vitro activation with bryostatin 1 and ionomycin. However, the presence of visceral metastases in the donor can inhibit the effectiveness of such lymphocytes. In the present study, we tested the ability of low-dose cyclophosphamide to overcome metastasis-induced immunosuppression in a murine model.. Mice were injected with MCA-105 sarcoma cells in the footpad alone or in the footpad and the tail vein to establish lung metastases. Cyclophosphamide was given i.p. 1 day before harvesting the draining popliteal lymph nodes. For all donor groups, DLN cells were activated with 5 nM bryostatin 1 and 1 microM ionomycin and cultured for 7 days in 20 U/ml IL-2. Activated DLN cells were then adoptively transferred to syngeneic mice with 3-day lung metastases.. The adoptive transfer of DLN cells from mice with footpad tumors only significantly reduced the number of lung metastases compared to untreated mice. However, activated DLN cells obtained from mice with both footpad and lung tumors were significantly less effective. Treatment of similar donor mice with 10 mg/kg cyclophosphamide significantly improved the anti-tumor activity of adoptively transferred cells. This dose of cyclophosphamide did not reduce the number of cells obtained from each lymph node or the expansion of cell numbers in vitro.. These results suggest that the administration of low-dose cyclophosphamide prior to harvesting DLN cells may improve the success of adoptive immunotherapy in cancer patients.

Topics: Animals; Bryostatins; Cyclophosphamide; Female; Immune Tolerance; Immunotherapy, Adoptive; Ionomycin; Lactones; Lung Neoplasms; Lymphocyte Activation; Lymphocytes, Tumor-Infiltrating; Macrolides; Mice; Mice, Inbred C57BL; Mitogens; Neoplasm Metastasis; Neoplasm Transplantation; Sarcoma, Experimental

Treatment of human cancer with tumour-specific T lymphocytes is limited by the frequent unavailability of autologous tumour to stimulate T-cell growth and by the toxicity associated with high-dose interleukin-2 (IL-2) treatment. In the present study we demonstrate that Bryostatin 1 (B) plus ionomycin (I) can substitute for tumour antigen and activate tumour-bearing hosts' T-cells which provide long-term protection against tumour challenge after adoptive transfer. Lymphocytes obtained from the popliteal lymph nodes (DLN) draining an MCA-105 footpad sarcoma were stimulated with B/I, and then cultured for 7 days with 20 U ml-1 IL-2. This in vitro stimulation protocol consistently expanded cell numbers greater than 20-fold during 7 days. Mice given B/I-stimulated draining lymph node (DLN) cells were protected from specific i.v. tumour challenge for at least 15 weeks after adoptive transfer, even in the absence of IL-2 treatment. Tumour immunity conferred by B/I-activated DLN cells was systemic and independent of host T-cells. However, resistance to tumour challenge was lost when either CD4+ or CD8+ T-cells were depleted in vivo. These studies indicate that DLN cells activated with bryostatin 1 and ionomycin persist long-term in vivo as functional memory cells after adoptive transfer.

Topics: Adjuvants, Immunologic; Animals; Antineoplastic Agents; Bryostatins; Evaluation Studies as Topic; Female; Immunotherapy, Adoptive; Ionomycin; Lactones; Lymphocyte Activation; Macrolides; Mice; Mice, Inbred C57BL; Neoplasm Metastasis; Neoplasm Transplantation; Sarcoma, Experimental; T-Lymphocytes; Time Factors