Direct correlation between nitric oxide synthase II inducibility and metastatic ability of UV-2237 murine fibrosarcoma cells carrying mutant p53.
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The relationship between nitric oxide synthase II (NOS II) inducibility and the metastatic ability of UV-2237 murine fibrosarcoma cells was determined. Highly metastatic cells survived to produce numerous lung metastases after i.v. injection in syngeneic C3H/HeN mice, whereas poorly metastatic cells did not. Highly metastatic clones exhibited higher levels of NOS II than did poorly metastatic clones in response to interleukin 1alpha and IFN-gamma stimulation. Furthermore, both poorly and highly metastatic clones contained an identical p53 mutation. Overexpression of NOS II in a highly metastatic clone by transfection with NOS II gene retarded tumor growth and completely suppressed metastasis. Our data indicate that a low to moderate level of NOS II expression directly correlates with metastatic ability of UV-2237 fibrosarcoma cells carrying mutant p53 and that a high level of nitric oxide production suppresses tumor growth and metastasis. (+info)
The insulin-like growth factor axis and prostate cancer: lessons from the transgenic adenocarcinoma of mouse prostate (TRAMP) model.
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We have characterized the temporal expression of the insulin-like growth factor (IGF) axis in the transgenic adenocarcinoma of mouse prostate (TRAMP) model as prostate cancer progression in this model closely mimics that observed in the human disease, and the model provides samples representing the earliest stages of prostate cancer that are clinically the most difficult to obtain. We report that prostate-specific IGF-I mRNA expression increased during prostate cancer progression in TRAMP mice and was elevated in the accompanying metastatic lesions, whereas prostatic IGF-I mRNA remained at nontransgenic levels in androgen-independent disease. Expression of IGF-II mRNA, however, was reduced in primary prostate cancer, metastatic lesions, and androgen-independent disease. Expression of type-1 IGF receptor (IGF1R) mRNA, encoding the cognate receptor for both IGF-I and IGF-II, as well as type-2 IGF receptor (IGF2R) mRNA was not found to be altered during primary prostate cancer progression in intact TRAMP mice but was dramatically reduced in metastatic lesions and in androgen-independent disease. Similar to reports from clinical disease, serum IGF-I levels were observed to increase precociously in TRAMP mice early in disease progression but remained at nontransgenic levels after castration. Elevated serum levels of IGF-binding protein 2 were observed to correlate with advanced prostate cancer in the TRAMP model. Together these observations implicate IGF-I as an important factor during the initiation and progression of primary prostate cancer and provide evidence that there is a strong selection against expression of IGF1R and IGF2R in metastatic and androgen-independent disease. (+info)
Overexpression of sialomucin complex, a rat homologue of MUC4, inhibits tumor killing by lymphokine-activated killer cells.
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Sialomucin complex (SMC) is a large heterodimeric glycoprotein complex composed of a mucin subunit ascites sialoglycoprotein-1 and a transmembrane subunit ascites sialoglycoprotein-2. It is a rat homologue of human mucin gene MUC4 and is abundantly expressed on the cell surface of highly metastatic ascites 13762 rat mammary adenocarcinoma cells. Because of their extended and rigid structures, mucin-type glycoproteins are suggested to have suppressing effects on cell-cell and cell-matrix interactions. During the metastatic process, these effects presumably cause tumor cell detachment from the primary tumor mass and facilitate escape of the tumor cells from immunosurveillance. Analyses of human breast cancer cells in solid tumors and tumor effusions showed that the more aggressive cells in effusions are stained with polyclonal antibodies against SMC more frequently than cells in solid tumors, suggesting a role for MUC4/SMC in tumor progression and metastasis. Previously, we generated recombinant cDNAs for SMC that vary in the number of mucin repeats to study the putative functions of SMC in tumor metastasis. These cDNAs were transfected into human cancer cell lines and tested for the effect of the expression of this gene. Here, using a tetracycline-responsive inducible expression system, we demonstrate that overexpression of SMC masks the surface antigens on target tumor cells and effectively suppresses tumor cell killing by cytotoxic lymphocytes. This effect results from the ability of SMC to block killer cell binding to the tumor cells and is dependent on both overexpression of the mucin and the number of mucin repeats in the expressed SMC. These results provide an explanation for the proposed role of SMC/MUC4 in tumor progression. (+info)
Overexpression of alpha1-6 fucosyltransferase in hepatoma cells suppresses intrahepatic metastasis after splenic injection in athymic mice.
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Changes in oligosaccharide structures alter the biological functions of cancer cells. Alpha1-6 fucosyltransferase (alpha1-6FucT) catalyzes the transfer of fucose to the innermost GlcNAc in N-glycans. Although alpha1-6FucT is barely detected in normal liver, it is enhanced during rat hepatocarcinogenesis and in human hepatoma. To understand the biological meaning of the alpha1-6FucT in hepatoma, especially in terms of metastasis, we established human hepatoma cell lines, which express high levels of alpha1-6FucT by transfection of the alpha1-6FucT gene and investigated intrahepatic metastasis after splenic injection to athymic mice. Tumor formation in the liver was dramatically suppressed in the alpha1-6FucT transfectants (1 of 9 and 1 of 10 in alpha1-6FucT transfectants versus 6 of 9 and 6 of 9 in controls). Although there were no differences in terms of cell invasiveness to a Matrigel or in terms of cytotoxicity to interleukin 2-treated lymphocytes between alpha1-6FucT transfectants and control cells, cell adhesion to mice hepatocytes and nonparenchymal liver cells in culture was significantly inhibited in alpha1-6FucT transfectants, compared to the controls. Attachment of alpha1-6FucT transfectants to a fibronectin-coated dish was decreased compared to controls because alpha5beta1 integrin was more strongly alpha1-6 fucosylated in the alpha1-6FucT transfectants. Two-dimensional electrophoresis followed by lectin blot showed that certain glycoproteins (Mr 50,000-150,000, pI 4.8-5.5) were alpha1-6 fucosylated and might be linked to suppression of intrahepatic metastasis. This is the first demonstration of the biological significance of alpha1-6 fucosylation on N-glycans in hepatoma cells under in vivo conditions. (+info)
Deficient activity of von Willebrand's factor-cleaving protease in patients with disseminated malignancies.
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An aberrant platelet immunorelated glycoprotein Ib (GPIb) receptor expressed by human tumor cells appears to participate in primary adhesive interactions required for the metastatic process. Hence, we questioned whether plasma von Willebrand's factor (vWf), its adhesive ligand, manifested comparable anomalies in patients with disseminated tumors. Plasma specimens from patients with disseminated metastases showed 68% (P < 0.013), 91% (P < 0.0009), and 207% (P < 0.0009) enhancements in FVIII:C activity, vWf-related antigen levels, and ristocetin co-factor activity, respectively, whereas their SDS-agarose electrophoretic analysis demonstrated a 165% (P < 0.001) increase in the highly polymeric forms of vWf compared to control preparations from patients with corresponding, localized solid tumors. Substantially reduced levels of vWf-cleaving protease activity were observed in study patient specimens, with no plasma inhibitors detectable. The clinical presence and absence of tumor metastases correlated significantly with vWf-cleaving enzyme activities of < or = 15% and > or = 88%, respectively (n = 20; P < 0.0001). Finally, with an in vitro model system, tumor-induced platelet aggregation was enhanced by 127% (P < 0.001) in study patient platelet-rich plasma (PRP) compared to control PRP and could be completely inhibited (P < 0.0009) when both tumor cells and their PRP substrates were incubated with monoclonal antibodies directed against the vWf binding epitope of GPIb alpha and against the GPIb binding epitope of plasma vWf, respectively. Unusually large vWf multimers observed in patients with disseminated tumors probably result from deficient vWf-cleaving protease activity and may represent a novel mechanism regulating primary platelet-tumor adhesive interactions involved in the metastatic process. (+info)
Metastasis-association of the rat ortholog of the human epithelial glycoprotein antigen EGP314.
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Screening for surface molecules expressed by metastasizing rat tumors had revealed evidence for metastasis-association of a molecule also expressed on epithelial cells. The similarity to the expression profile of the panepithelial glycoprotein EGP314 prompted us to isolate and sequence the gene and to explore functional features of the molecule in transfected tumor lines. The molecule D5.7A, named according to the antibody, D5.7, used for selection, indeed, is the ortholog of EGP314 with 92% and 80% identity to the murine and the human molecules. Like EGP314, D5.7A has a particular cleavage site, a small cleavage product being resolved under reducing conditions from the membrane anchored part of the molecule. Transfection of a low metastasizing fibrosarcoma, pheochromoblastoma and adenocarcinoma revealed that expression of D5.7A facilitates tumor progression. Depending on the origin of the tumor, D5.7A transfectants either metastasized via the lymphatic system (pheochromoblastoma, adenocarcinoma) or hematogeneously (fibrosarcoma). Particularly after proteolytic cleavage, D5.7A facilitated cell - cell adhesion and provided a proliferative signal upon crosslinking. Thus, the rat ortholog of EGP314 is involved in metastasis formation. Importantly, its functional activities apparently rely on proteolytic cleavage. These findings provide a first evidence on how a panepithelial marker can be involved in tumor progression. (+info)
A novel immunological model for the study of prostate cancer.
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The Dunning R-3327 rat prostatic adenocarcinoma is a widely accepted model for in vivo experimental studies of prostate cancer. We have previously derived phenotypically distinct cell lines from a s.c. tumor resulting from the inoculation of the R-3327-5 subclone into Copenhagen rats. In this study, we report studies using a gelatin sponge model for the delivery of tumor cells and the retrieval of tumor-specific leukocytes responsive to different prostatic cell lines. S.c. preimplanted sponges were inoculated with tumor cells previously selected for differential properties of tumor formation and metastasis and examined for leukocyte content at time points of 1, 3, and 5 weeks after tumor cell inoculation. Cytospin and flow cytometric analyses revealed fewer tumor-associated leukocytes present in sponges inoculated with tumorigenic R-3327-5' and R-3327-5'B lines, with lesser sponge degradation, than in experiments with the nontumorigenic R-3327-5'A line, suggestive of a tumor cell-induced immunomodulatory mechanism. Morphological studies indicate an intermittent tumor growth pattern that gradually disappears in sponges inoculated with the nontumorigenic R-3327-5'A cells but a robust growth pattern in sponges inoculated with the tumorigenic cell lines. Cytokine analyses show the secretion of higher levels of active transforming growth factor-beta by the more invasive and metastatic lines. Total transforming growth factor-beta levels are higher in the epithelial, tumorigenic R-3327-5'B line. Additionally, the more tumorigenic lines secrete interleukin 10, a potent immunosuppressive molecule. In this report, we demonstrate the ability to retrieve viable leukocyte populations from a prostate tumor line bearing sponges, which offers an important model for further in vitro and in vivo manipulations and holds promise for testing adoptive immunotherapeutic strategies. (+info)
Methylation of the CD44 metastasis suppressor gene in human prostate cancer.
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Previous studies demonstrated that CD44 is a metastasis suppressor gene for prostate cancer and that the expression of CD44 both at mRNA and protein levels is down-regulated during prostate cancer progression, with down-regulation being correlated with higher tumor grade, aneuploidy, and distant metastasis. In this study, we evaluated DNA hypermethylation as a potential mechanism accompanying this decreased CD44 expression in human prostate cancer. Nucleotide sequence analysis revealed a CpG island in the CD44 transcriptional regulatory region. We found that cytosine methylation of CD44 promoter occurs in CD44-negative prostate cancer cell line (i.e., LNCaP) but not in prostate cancer cell lines (i.e., TSU, PC3, and DU145) expressing this gene. In addition, we examined methylation status of CD44 in 84 matched normal and cancer prostate specimens. Hypermethylation of the 5' CpG island of CD44 gene was observed in 31 of 40 primary prostate cancer specimens, 3 of 4 distant organ site metastases obtained at autopsy from men who died of prostate cancer, and 4 of the 40 matched normal tissues. These results demonstrated that methylation of the 5' CpG island of CD44 gene is closely associated with transcriptional inactivation, resulting in a decreased expression of CD44 in human prostate cancer. (+info)