Physiopathology of liver metastasis.
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Humans↗
Biomedical subjects
M M Mareel
Publications and source records attributed to M M Mareel.
Bile acids stimulate invasion and haptotaxis in human colorectal cancer cells through activation of multiple oncogenic signaling pathways.
Bile acids are implicated in colorectal carcinogenesis as evidenced by epidemiological and experimental studies. We examined whether bile acids stimulate cellular invasion of human colorectal and dog kidney epithelial cells at different stages of tumor progression. Colon PC/AA/C1, PCmsrc, and HCT-8/E11 cells and kidney MDCKT23 cells were seeded on top of collagen type I gels and invasive cells were counted after 24 h incubation. Activation of the Rac1 and RhoA small GTPases was investigated by pull-down assays. Haptotaxis was analysed with modified Boyden chambers. Lithocholic acid, chenodeoxycholic acid, cholic acid and deoxycholic acid stimulated cellular invasion of SRC- and RhoA-transformed PCmsrc and MDCKT23-RhoAV14 cells, and of HCT-8/E11 cells originating from a sporadic tumor, but were ineffective in premalignant PC/AA/C1 and MDCKT23 cells. Bile acid-stimulated invasion occurred through stimulation of haptotaxis and was dependent on the RhoA/Rho-kinase pathway and signaling cascades using protein kinase C, mitogen-activated protein kinase, and cyclooxygenase-2. Accordingly, BA-induced invasion was associated with activation of the Rac1 and RhoA GTPases and expression of the farnesoid X receptor. We conclude that bile acids stimulate invasion and haptotaxis in colorectal cancer cells via several cancer invasion signaling pathways.
Bile Acids and Salts↗
The role of bile acids in carcinogenesis.
Bile acids play a role in colorectal carcinogenesis as evidenced by epidemiological and experimental studies. Some bile acids stimulate growth of normal colonic and adenoma cells, but not of colorectal cancer cells. Moreover, bile acids stimulate invasion of colorectal cancer cells, at least in vitro. One possible mechanism of action is bile acid-induced DNA binding and transactivation of the activator protein-1 (AP-1) by co-operate activation of extracellular signal-regulated kinases (ERKs) and PKC signaling. In the present paper, we review the mechanisms by which bile acids influence carcinogenesis.
Animals↗
Alkyl-lysophospholipid 1-O-octadecyl-2-O-methyl- glycerophosphocholine induces invasion through episialin-mediated neutralization of E-cadherin in human mammary MCF-7 cells in vitro.
1-O-octadecyl-2-O-methyl-glycerophosphocholine (ET-18-OMe) is an analogue of the naturally occurring 2-lysophosphatidylcholine belonging to the class of antitumor lipids. Previously, we demonstrated that ET-18-OMe modulates cell-cell adhesion of human breast cancer MCF-7 cells. In the present study, we tested the effect of ET-18-OMe on adhesion, invasion and localisation of episialin and E-cadherin in MCF-7/AZ cells expressing a functional E-cadherin/catenin complex. The MCF-7/6 human breast cancer cells were used as negative control since their E-cadherin/catenin complex is functional in cells grown on solid substrate but not in suspension. The function of E-cadherin, a calcium-dependent transmembrane cell-cell adhesion and signal-transducing molecule, is disturbed in invasive cancers by mutation, loss of mRNA stability, proteolytic degradation, tyrosine phosphorylation of associated proteins and large cell-associated proteoglycans or mucin-like molecules such as episialin. Episialin, also called MUC1, is an anti-adhesion molecule that by its large number of glycosylated tandem repeats can sterically hinder the adhesive properties of other glycoproteins. ET-18-OMe inhibited the E-cadherin functions of MCF-7/AZ cells as measured by inhibition of fast and slow aggregation and by the induction of collagen invasion. These effects were enhanced by MB2, an antibody against E-cadherin and blocked by monoclonal antibodies (MAbs) 214D4 or M8 against episialin. ET-18-OMe had no influence on tyrosine phosphorylation of beta-catenin and the E-cadherin/catenin complex remained intact. Transcription, translation, protein turnover and cell surface localisation of episialin were not altered. ET-18-OMe induced finger-like extensions with clustering of episialin together with E-cadherin and carcinoembryonic antigen but not with occludin. In cells in suspension, ET-18-OMe caused a shift in the flow-cytometric profile of episialin toward a lower intensity for MCF-7/AZ cells. In contrast with MCF-7/AZ cells, the adhesion-deficient and noninvasive MCF-7/6 cells showed neither morphotypic changes nor induction of aggregation nor invasion in collagen I upon treatment with ET-18-OMe. Co-localisation of episialin with E-cadherin was rarely observed. We conclude that in the human breast cancer cells MCF-7/AZ, E-cadherin and episialin are key molecular players in the regulation of promotion and suppression of cell-cell adhesion and invasion.
Antibodies, Monoclonal↗
Inhibition of tamoxifen's therapeutic benefit by tangeretin in mammary cancer.
Tangeretin, a molecule present in citrus fruits and in certain 'natural' menopausal medications, is an effective tumour growth and invasion inhibitor in vitro of human MCF 7/6 breast cancer cells. However, when added to the drinking water of MCF 7/6 tumour-bearing mice it neutralises the beneficial tumour-suppressing effect of tamoxifen. Tangeretin reduces the number of natural killer cells. This may explain why the beneficial suppressive effect of tangeretin on MCF 7/6 cell proliferation in vitro is completely counteracted in vivo.
Animals↗
Internalization of the E-cadherin/catenin complex and scattering of human mammary carcinoma cells MCF-7/AZ after treatment with conditioned medium from human skin squamous carcinoma cells COLO 16.
Cytokines and other paracrine or autocrine factors functionally modulate the invasion-suppressor and signal-transducing E-cadherin/catenin complex. We have used conditioned medium from human squamous carcinoma COLO 16 cells (CM COLO 16) as a source of such factors to modulate the E-cadherin/catenin complex in human breast carcinoma MCF-7 cells. CM COLO 16 induces scattering of MCF-7/AZ, but not of MCF-7/6 cells on tissue culture plastic substratum, and reduces aggregation of MCF-7/AZ cells in suspension. Insulin-like growth factor I counteracts this reduction of aggregation. Confocal laser scanning microscopy of immunocytochemical stainings shows loss of the honeycomb pattern of E-cadherin, alpha-catenin and beta-catenin, and internalization of those elements. Cell surface biotinylation shows a decrease in membrane-bound E-cadherin. Immunoprecipitation and cell fractionation show that the composition of the complex is maintained. Interleukin-1, interleukin-6, granulocyte-monocyte colony stimulating factor, stem cell factor, scatter factor/hepatocyte growth factor and transforming growth factor-beta, added separately to MCF-7/AZ cells, could not mimic the effects of CM COLO 16. Neither could we find evidence that the 80 kDa extracellular fragment of E-cadherin is implicated in scattering of MCF-7/AZ cells. This fragment is present in CM COLO 16, but it is also produced by the MCF-7/AZ cells themselves, even at higher levels. Our data point toward cytoplasmic internalization induced by paracrine factors as one of the downregulating mechanisms for the E-cadherin/catenin complex.
Biotinylation↗
Influence of tangeretin on tamoxifen's therapeutic benefit in mammary cancer.
BACKGROUND: Tamoxifen and the citrus flavonoid tangeretin exhibit similar inhibitory effects on the growth and invasive properties of human mammary cancer cells in vitro; furthermore, the two agents have displayed additive effects in vitro. In this study, we examined whether tangeretin would enhance tamoxifen's therapeutic benefit in vivo. METHODS: Female nude mice (n = 80) were inoculated subcutaneously with human MCF-7/6 mammary adenocarcinoma cells. Groups of 20 mice were treated orally by adding the following substances to their drinking water: tamoxifen (3 x 10(-5) M), tangeretin (1 x 10(-4) M), tamoxifen plus tangeretin (3 x 10(-5) M plus 1 x 10(-4) M), or solvent. RESULTS AND CONCLUSIONS: Oral treatment of mice with tamoxifen resulted in a statistically significant inhibition of tumor growth compared with solvent treatment (two-sided P = .001). Treatment with tangeretin did not inhibit tumor growth, and addition of this compound to drinking water with tamoxifen completely neutralized tamoxifen's inhibitory effect. The median survival time of tumor-bearing mice treated with tamoxifen plus tangeretin was reduced in comparison with that of mice treated with tamoxifen alone (14 versus 56 weeks; two-sided P = .002). Tangeretin (1 x 10(-6) M or higher) inhibited the cytolytic effect of murine natural killer cells on MCF-7/6 cells in vitro, which may explain why tamoxifen-induced inhibition of tumor growth in mice is abolished when tangeretin is present in drinking water. IMPLICATIONS: We describe an in vivo model to study potential interference of dietary compounds, such as flavonoids, with tamoxifen, which could lead to reduced efficacy of adjuvant therapy. In our study, the tumor growth-inhibiting effect of oral tamoxifen was reversed upon addition of tangeretin to the diet. Our data argue against excessive consumption of tangeretin-added products and supplements by patients with mammary cancer during tamoxifen treatment.
ATP Binding Cassette Transporter, Subfamily B, Mem↗
The alphaE-catenin gene (CTNNA1) acts as an invasion-suppressor gene in human colon cancer cells.
The acquisition of invasiveness is a crucial step in the malignant progression of cancer. In cancers of the colon and of other organs the E-cadherin/catenin complex, which is implicated in homotypic cell-cell adhesion as well as in signal transduction, serves as a powerful inhibitor of invasion. We show here that one allele of the alphaE-catenin (CTNNA1) gene is mutated in the human colon cancer cell family HCT-8, which is identical to HCT-15, DLD-1 and HRT-18. Genetic instability, due to mutations in the HMSH6 (also called GTBP) mismatch repair gene, results in the spontaneous occurrence of invasive variants, all carrying either a mutation or exon skipping in the second alphaE-catenin allele. The alphaE-catenin gene is therefore, an invasion-suppressor gene in accordance with the two-hit model of Knudsen for tumour-suppressor genes.
Alleles↗
Sialylation of E-cadherin does not change the spontaneous or ET-18-OMe-mediated aggregation of MCF-7 human breast cancer cells.
We have investigated the role of sialylation on cell-cell adhesion mediated by E-cadherin. Two MCF-7 human breast cancer cell variants were studied: MCF-7/AZ cells showed a spontaneous cell-cell adhesion in the fast and slow aggregation assay. whereas the adhesion deficient MCF-7/6 cell variant failed to form larger aggregates, suggesting that E-cadherin was not functional under the conditions of both assays. We measured the sialyltransferase activities using Galbeta1-3GalNAcalpha-O-benzyl and Galbeta1-4GlcNAcalpha-O-benzyl as acceptor substrates as well as mRNA levels of four sialyltransferases, ST3Gal I, ST3Gal III, ST3Gal IV, ST6Gal I, using multiplex RT-PCR in MCF-7 cell variants. The alpha2-6 and alpha2-3 sialylation of E-cadherin was investigated by immuno-blot using Sambucus nigra agglutinin and Maackia amurensis agglutinin. Compared to the adhesion-proficient MCF-7/AZ cells, the adhesion-deficient MCF-7/6 cell line apparently lacks ST6Gal I mRNA, has a lower ST3Gal I mRNA, a lower ST3Gal I sialyltransferase activity, and no alpha2-3 linked sialic acid moieties on E-cadherin. The potential anti-cancer drug 1-O-octadecyl-2-O-methylglycero-3-phosphocholine (ET-18-OMe, 48 h, 25 microg/ml) belonging to the class of alkyllysophospholipids restored the E-cadherin function in the adhesion-deficient MCF-7/6 cells as evidenced by an increased aggregation. ET-18-OMe caused loss of ST6Gal I mRNA in MCF-7/AZ cells but no changes of sialyltransferase activities or sialic acid moieties on E-cadherin could be observed. We conclude that Ca2+-dependent, E-cadherin-specific homotypic adhesion of MCF-7/AZ or MCF-7/6 cells treated with ET-18-OMe was not affected by sialylation of E-cadherin.
Antineoplastic Agents↗
hMSH6 deficiency and inactivation of the alphaE-catenin invasion-suppressor gene in HCT-8 colon cancer cells.
Transition from an epithelioid (E) to a round (R) morphotype, in the human colon cancer cell line HCT-8, is associated with loss or truncation of alphaE-catenin and acquisition of invasiveness in organ culture. In E clones, like in parental HCT-8 cells, one allele of the alphaE-catenin gene (CTNNA1) is mutated. HCT-8 cells have also a 'Microsatelite Instability-High' (MSI-H) phenotype presumably due to a mutated hMSH6 gene. Fusion of E type cells doubles the wild type CTNNA1 alleles and prevents the loss of alphaE-catenin. Introduction of an extra chromosome 2, carrying a wild type hMSH6 gene, restores post-replicative mismatch repair and also prevents the frequent inactivation of the remaining wild type CTNNA1 allele.
Alleles↗
Did the four human cancer cell lines DLD-1, HCT-15, HCT-8, and HRT-18 originate from one and the same patient?
Four human colon cancer cell lines, HCT-8, HRT-18, DLD-1, and HCT-15, with an epithelioid morphotype reproducibly formed alpha-catenin-deficient round cells. Using DNA fingerprinting, we found that these four cell lines have an identical genetic background. Our finding strongly suggests a genetic background for the reproducible loss of alpha-catenin and the ensuing acquisition of invasiveness in all four cell lines.
Colonic Neoplasms↗
Mutation of alpha-catenin results in invasiveness of human HCT-8 colon cancer cells.
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Cadherins↗
Patterns of alpha- and beta-catenin and E-cadherin expression in colorectal adenomas and carcinomas.
Previous in vitro and in vivo model studies have shown that when E-cadherin expression in carcinoma cells is reduced, invasive behaviour ensues. The situation in human cancer in vivo, however, appears to be more complex, as immunohistochemically determined E-cadherin expression in various carcinomas, including colorectal cancer, does not always correlate with invasive growth. Loss of cell adhesion during invasion in spite of E-cadherin expression might be associated with a defective cadherin-catenin complex. The expression of alpha- and beta-catenin in comparison with E-cadherin was therefore examined in colorectal adenomas and carcinomas and in lymph node and liver metastases. In normal colonic mucosa, alpha- and beta-catenin immunoreactivity occurred along the lateral plasma membranes of the epithelial cells, in a pattern identical to E-cadherin staining. A similar pattern was found in colorectal adenomas and in most malignancies. In general, in neoplastic epithelia, the majority of the cancer cells displayed a normal (matching) pattern of E-cadherin and catenin expression. It is concluded that the patterns of expression of E-cadherin and alpha- and beta-catenin are very similar in colorectal neoplasms. This observation indicates that invasion in colorectal cancer is not paralleled by consistent loss of expression of the components of the cadherin-catenin complex.
Adenocarcinoma↗
Anti-invasive activity of alkaloids and polyphenolics in vitro.
Invasiveness, the ability of certain tumour cells to migrate beyond their natural tissue boundaries, often leads to metastasis, and usually determines the fatal outcome of cancer. The need for anti-invasive agents has led us to search for possibly active compounds among alkaloids and polyphenolics. One hundred compounds were screened in an assay based on the confrontation of invasive human MCF-7/6 mammary carcinoma cells with fragments of normal embryonic chick heart in vitro. Anti-invasive activity was frequently found among chalcones having a prenyl group. Six compounds were found to inhibit invasion when added to the culture medium at concentrations as low as 1 microM. For at least three of them the anti-invasive effect could be associated with a cytotoxic effect on the MCF-7/6 cells, but not on the heart tissue. This selective cytotoxicity was substantiated by different methods, such as histology and growth assays (volume measurements, cell counts, MTT and sulforhodamine B assays). The anti-invasive effects of the compounds could neither be ascribed to induction of apoptosis nor to the promotion of cell-cell adhesion. Our data indicate that among the alkaloids and polyphenolics a number of molecules can inhibit growth and invasion of human mammary cancer cells via selective cytotoxicity.
Alkaloids↗
Functional downregulation of the E-cadherin/catenin complex leads to loss of contact inhibition of motility and of mitochondrial activity, but not of growth in confluent epithelial cell cultures.
When epithelial cells reach confluency in vitro, a number of energy-requiring activities such as growth and motility are contact-inhibited. We investigated the possible role of the E-cadherin/catenin complex, which acts as an invasion suppressor, in contact inhibition. Three strategies for modulation of the complex were used. Firstly, the cell-cell adhesion and signal transduction functions of E-cadherin were neutralized immunologically in human MCF-7/6 mammary carcinoma cells possessing a complete complex. Secondly, the effect of E-cadherin transfection in E-cadherin negative cell lines was investigated. Thirdly, alpha-catenin deficient variants of the human HCT-8/S11 colon carcinoma cell line were compared with their parent cells. In confluent cultures functional downregulation of the E-cadherin/catenin complex did not alter cell growth nor saturation density. This was shown by cell number counts, protein staining assays, cell cycle analysis, proliferation markers (Ki67 and Proliferating Cell Nuclear Antigen) and apoptosis assays. However, confluent cells with a functionally deficient complex showed positional instability and enhanced succinate dehydrogenase-mediated mitochondrial 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyl) tetrazolium bromide (MTT) conversion, as compared to cells with an active complex. Our data indicate that contact inhibition of motility and of mitochondrial enzyme activity, but not of growth is regulated by the E-cadherin/catenin complex in epithelial cells.
Cadherins↗
The E-cadherin/catenin complex in invasion and metastasis.
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Adenocarcinoma↗
Transition from the noninvasive to the invasive phenotype and loss of alpha-catenin in human colon cancer cells.
Loss of epithelioid organization in carcinoma cell lines has been related to invasiveness and poor differentiation of tumors. We investigated the invasion in vitro of various human colon cancer cell lines. Most cell lines were noninvasive into chick heart fragments, and this correlated with an epithelioid morphotype. Only cell lines COLO320DM, SW620, and variants of HCT-8 and DLD-1 were invasive and nonepithelioid. We examined in these cell lines whether invasiveness was related to changes in the structure and function of the E-cadherin/catenin complex. E-cadherin functions as an invasion suppressor and as a cell-cell adhesion molecule when linked to the cytoskeleton via alpha-catenin plus beta- or gamma-catenin. All noninvasive cell lines showed E-cadherin linked to these catenins. The E-cadherin-dependent cell-cell adhesion function in these cell lines was demonstrated by two assays in vitro. It was interesting that all invasive cell lines showed a dysfunctional E-cadherin/catenin complex. COLO320DM, SW480, and SW620 cells were defective in E-cadherin expression, whereas the invasive variants of HCT-8 and DLD-1 lacked the alpha-catenin protein. From clonal epithelioid HCT-8 cultures with functional E-cadherin/catenin complexes, we subcloned, repeatedly, round cell variants that were again invasive and expressed no alpha-catenin protein. Our data suggest that reproducible transformations toward a more invasive phenotype in HCT-8 cells are associated with down-regulation of alpha-catenin. The mechanisms of this transformation and the level of alpha-catenin down-regulation are currently investigated.
Amino Acid Sequence↗
Capillary gas chromatography of hexadecylphosphocholine in Caco-2T cells and cell culture media.
The gas-chromatographic determination of hexadecylphosphocholine (HePC), an experimental antitumor agent of the alkyllysophospholipid group, in Caco-2T cell culture and cell culture media is described. The Caco-2T cells were treated with HePC at a concentration of 40 micrograms/ml (9.8 microM) and the uptake of the drug into the cells (calculated per milligram protein) was measured after 48 h culture (37 degrees C, 10% CO2). Also, a reversibility test for another 48 h was carried out in which the retention of the drug was measured. The toxicity of HePC on Caco-2T cells in viability assays was determined. Before the capillary gas-chromatographic determination, sample cleanup was performed by solid-phase extraction (SPE) on a weak cation-exchange column of the CBA (carboxylic acid) type. For quantitation, racemic 1-O-octadecyl-2-O-methylglycero-3-phosphorylcholine (ET-18-OMe) was added as internal standard, followed by derivatization with trimethylsilylbromide. The results showed that HePC taken up by the cells during 48 h of treatment was still detectable 48 h after removal of the drug from the medium.
Antineoplastic Agents↗