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F Vignon

Publications and source records attributed to F Vignon.

At least 37 records · Page 2Linked to original sources
PubMed1990

Interactions of cathepsin-D and insulin-like growth factor-II (IGF-II) on the IGF-II/mannose-6-phosphate receptor in human breast cancer cells and possible consequences on mitogenic activity of IGF-II.

The lysosomal enzyme cathepsin-D (cath-D) and insulin-like growth factor-II (IGF-II), which share a common IGF-II/mannose-6-phosphate (M6P) transmembrane receptor, are both synthesized and secreted by breast cancer cells, upon which they might exert an intracrine/autocrine control on proliferation. We have evaluated the binding of 125I-immunopurified human cath-D in different breast cell membrane preparations. The concentration of high affinity M6P reversible binding sites (mean Kd, 0.85 nM) varied among the different breast cancer cells (0-0.82 pmol/mg membrane protein), but there was no correlation between the presence of steroid receptor and M6P-dependent binding. Cross-linking experiments with [125I]cath-D and [125I]IGF-II showed the formation of complexes with the 270,000 mol wt IGF-II/M6P receptor molecule which migrated, respectively, at 330,000 and 270,000 mol wt in 3-10% gradient sodium dodecyl sulfate-polyacrylamide gels. [125I]IGF-II cross-linking was increased by M6P (20% above control), whereas cath-D strongly inhibited IGF-II interaction by 80%. Conversely, IGF-II reduced [125I]cath-D cross-linking by 55%. Direct ligand binding on receptors transferred onto nitrocellulose sheets by Western blotting confirmed the interaction of both ligands on the same receptor molecule. By studying IGF-II's growth-promoting activity in these cells in a wide range of concentrations, we show that IGF-II triggers its mitogenic response via IGF-II/M6P receptor at low concentrations, whereas it is mainly acting via IGF-I receptor at high concentrations. Three lines of evidences lead us to that conclusion.(ABSTRACT TRUNCATED AT 250 WORDS)

Binding, Competitive↗
PubMed1990

Regulation, clinical and biological significance of cathepsin D in breast cancer.

The lysosomal protease, pro-cathepsin D, is overexpressed and secreted by human breast cancers. In estrogen-responsive breast cancer cell lines, estrogens and growth factors stimulate cathepsin D expression through distinct mechanisms. Clinical studies indicate that high cathepsin D concentration in primary breast cancers is correlated with an increased risk of metastasis and particularly useful to orientate node-negative tumors towards an adjuvant therapy.

Breast Neoplasms↗
PubMed1989

Lipid composition of human seminal plasma.

This study aims at determining the amounts of cholesterol, triglycerides, phospholipids, and nonesterified fatty acids in man's seminal liquid and determining their possible variations linked with the ways of taking and congealing samples. It concludes the determinations of lipids in human seminal liquid are reproducible; the way of taking samples has no real influence; however, it seems best to centrifuge sperm immediately after liquefaction to avoid use of triglycerides and NEFA by the spermatozoa.

Cholesterol↗
PubMed1989

A two-site immunoenzymometric assay of 52-kDa pro-cathepsin D, and its use in human breast diseases.

After isolating monoclonal antibodies specific for the 52-kDa precursor of cathepsin D (cath-D), which is secreted in excess in both hormone-dependent and hormone-independent breast cancer, we developed a two-step double-determinant immunoenzymometric assay that is specific for this pro-enzyme. The assay combines the use of a monoclonal antibody specific for the precursor and bound to microtiter plates, and a second antibody directed against a smaller processed form of the mature enzyme, coupled to alkaline phosphatase. The specificity of the assay relies on separate and sequential additions of the antigen and the conjugated second antibody. It allows rapid measurement of the analyte in plasma and cytosols of normal and neoplastic mammary tissues, with a detection limit of 5 fmol and a maximal interassay coefficient of variation of 9%. This assay is particularly useful for tissue cytosol samples where the pro-enzyme form co-exists with large quantities of the mature processed forms of the enzyme. Comparative assays of 52-kDa pro-cath-D and total cath-D in cytosols of breast cancers and benign mastopathies indicate that the present assay better discriminates between benign and cancerous mammary tumors.

Antibodies, Monoclonal↗
PubMed1989

[Update on the mechanism of action of synthetic antiestrogens].

Synthetic antiestrogens are drugs commonly used in the endocrine therapy of breast cancer. Their pharmacology is complex, most of them presenting a partial estrogenic effect. Their action via the estrogen receptor (ER) has been better understood through the use of cellular models. The antihormone-ER complex binds to estrogen responsive elements on DNA, but is unable to mimic exactly the transcriptional activation induced by the estradiol-ER complex. The antiproliferative effect of these antihormones, mainly cystostatic but also cytotoxic, is the result of the competitive inhibition of estrogens, and might also involve interaction with other cellular mediators (stimulation of inhibitory factors, and growth factors inhibition). The term antiestrogen seems therefore to be too restrictive and it may be more accurate to name them estrogen-receptor mediated drugs inhibiting cell proliferation.

Breast↗
PubMed1989

[Comparative study of the lipid composition of seminal fluid and ovulatory peritoneal fluid in humans].

Sperm capacitation is accompanied by changes in the lipid composition of the sperm membrane. The purpose of this study is to compare the lipid composition of the semen with that of peritoneal fluid. These two media have opposite effects on sperm capacitation. Cholesterol, triglyceride and phospholipids were measured in seminal fluid (15 specimens) obtained by sperm centrifugation immediately after liquefaction and in the peritoneal fluid at ovulation (15 specimens) obtained through transvaginal puncture. The lipid composition of these two media is different from that of serum. Seminal fluid is characterized by a high level of phospholipids, 14.79 +/- 3.67 g/l as against 0.53 +/- 0.12 g/l in peritoneal fluid. The level of cholesterol is identical in the two media. The cholesterol/phospholipid ratio is therefore lower in seminal fluid than in peritoneal fluid (0.022/0.79). Lecithins are raised in peritoneal fluid as are sphingomyelins and the phosphatidylethanolamines, which are the most important phospholipids in seminal fluid. It therefore can be postulated that the cholesterol/phospholipid ratio, which is low in seminal fluid, and its richness in sphingomyelin brings about stabilization of the membrane cover which corresponds to a decapacitated state, whereas the cholesterol/phospholipid ratio, which is higher in peritoneal fluid, makes for greater fluidity, and this corresponds to the state of capacitation. These preliminary results obtained from human material do not in any way contradict those obtained from animal studies.

Ascitic Fluid↗
PubMed1988

Characterization and properties of two monoclonal antibodies specific for the Mr 52,000 precursor of cathepsin D in human breast cancer cells.

The Mr 52,000 estrogen-induced protein secreted by MCF7 cells has been identified as a procathepsin D (procath D), which increases cell growth in vitro, may stimulate invasiveness by digesting extracellular matrix and appears to be a tissue marker for predicting relapses in breast cancer patients. The protease is also present within mammary cells as Mr 48,000, 34,000, 14,000 mature forms that were also recognized by the previously described antibodies to the Mr 52,000 protein (M. Garcia, F. Capony, D. Derocq, D. Simon, B. Pau, and H. Rochefort, Cancer Res., 45: 709-716, 1985). Using selective screening with a [35S]methionine-labeled MCF7 cell lysate, we have now isolated two new monoclonal antibodies interacting exclusively with the Mr 52,000 procath D. The two monoclonal antibodies, M2E8* and D9H8*, purified from ascitic fluids are IgG1. Their respective Kds for Mr 52,000 procath D are 0.96 and 0.18 nM. They are directed against two separate domains of the proenzyme that differ from the three domains of previously described antibodies. They both interact with the deglycosylated protein and recognize the autoactivated secreted proenzyme (Mr 51,000), which is devoid of the first part of the NH2-terminal end. By immunodetection of cathepsin D proteolytic activity in plasma, these two antibodies were found to recognize selectively human cathepsin D but not the cathepsin D of other species (rat, mouse, rabbit, goat, and horse) whereas antibodies to mature cathepsin D were less species specific. Using sequential passages on concanavalin A-Sepharose and Sepharose matrices coupled to antibodies to the precursor and antibodies to the mature cathepsin D, we separately purified to homogeneity the Mr 52,000 procath D form and its processed cellular forms, whose biological activities can now be assessed independently. The two monoclonal antibodies were also shown to inhibit the uptake and processing of the Mr 52,000 cathepsin D in MCF7 cells (mostly D9H8*) and to decrease its proteolytic activity, mostly on extracellular matrix (M2E8*). These two monoclonal antibodies are therefore new tools for studying the function, regulation, cellular processing, and localization of procath D as well as the clinical significance of its concentration in normal and tumoral mammary epithelial cells.

Antibodies, Monoclonal↗
PubMed1988

Structure, function, regulation and clinical significance of the 52K pro-cathepsin D secreted by breast cancer cells.

In estrogen-receptor-positive human breast cancer cell lines (MCF7, ZR75-1), estrogens specifically increase the secretion into the culture medium of a 52,000 Da (52K) glycoprotein and stimulate cell proliferation. The 52K protein has been purified to homogeneity using monoclonal antibodies and identified as the secreted precursor of a cathepsin D bearing mannose-6-phosphate signals. The secreted precursor 52K protein is mitogenic in vitro in estrogen-deprived MCF7 cells, can be taken up by these cells via mannose-6-phosphate receptors, and can degrade extracellular matrix and proteoglycans following its auto-activation. The protease is also produced constitutively by ER-negative cell lines, and is inducible by tamoxifen in some antiestrogen-resistant variants. The corresponding cDNA has been cloned using N-terminal sequencing of the protein and monoclonal antibodies. Its complete sequencing indicates a strong homology with pro-cathepsin D of normal tissues. Using a cDNA probe, the regulation of 52K cathepsin D mRNA by estrogens and antiestrogens has been studied and chromosome localization determined by in situ hybridization. Clinical studies using both immunohistochemistry and immunoenzymatic assay of breast cancer cytosol have shown that the concentration of total cellular cathepsin D (52K + 48K + 34K) is related to the proliferation of mammary ducts and to the prognosis of breast cancer. Its cytosolic concentration in primary tumors of postmenopausal patients is correlated slightly with lymph node invasion and significantly with shorter disease-free intervals in a 6-year retrospective study with the Danish Breast Cancer Groups and Finsen Institute (S. Thorpe et al.).(ABSTRACT TRUNCATED AT 250 WORDS)

Biomarkers, Tumor↗
PubMed1987

Antiestrogens inhibit the mitogenic effect of growth factors on breast cancer cells in the total absence of estrogens.

The antiproliferative effect of antiestrogens in breast cancer is believed to be entirely due to the inhibition of estrogen induced growth. We show here that non-steroidal antiestrogens inhibit the growth of the human breast cancer MCF7 cells in the complete absence of estrogens (phenol-red-free medium) when cell proliferation is stimulated by insulin or epidermal growth factor. This non-antiestrogenic effect of antiestrogens is, however, mediated by accessible estrogen receptor sites, as it is not observed in receptor negative hormone-independent breast cancers, and is rescued by estradiol but not by insulin. We conclude that antiestrogens inhibit cell proliferation by inhibiting growth factor action as well as estrogen action and that in both cases, accessible estrogen receptors are required.

Breast Neoplasms↗
PubMed1987

Steroid receptor-mediated cytotoxicity of an antiestrogen and an antiprogestin in breast cancer cells.

The antiproliferative and cytotoxic effects of 4-hydroxytamoxifen, an antiestrogen with a high affinity for the estrogen receptor, and of 17 beta-hydroxy-11 beta-(4-methylaminophenyl)-17-(1-propynyl)estra-4,9-dien-3- one-6-7 (RU486), an antiprogestin with a high affinity for the progestin receptor, have been studied on human breast cancer cell lines in culture. The number of dead cells was evaluated by several techniques (trypan blue stain exclusion, DNA cleavage, lactic dehydrogenase activity, morphological changes, and cloning efficiency in soft agar) and found to be increased both by the antiestrogen and the antiprogestin at concentrations correlating with the affinities for their respective receptors. This cytotoxic effect was prevented by the occupation of the respective receptors with estrogen and progestin and was not found in the estrogen receptor- and progestin receptor-negative MDA MB 231 and BT20 cell lines. The contrast between the ultrastructural modifications of chromatin and the integrity of mitochondria suggested that the antihormone-induced cell death was by apoptosis. We conclude that in addition to the receptor-mediated cytostatic activity and the nonspecific cytotoxic activity, antiestrogens trigger a third type of effect that we designate as "receptor-mediated cytotoxic." Similar conclusions can be drawn for the antiprogestin RU486, indicating moreover that the antihormone and antiproliferative activities of this drug are clearly dissociated. The mechanism of these receptor-mediated cytotoxic activities of antiestrogen and antiprogesterone is not known but does not seem to be explained entirely by the antihormone activity of these drugs.

Breast Neoplasms↗
PubMed1987

Estrogen-induced lysosomal proteases secreted by breast cancer cells: a role in carcinogenesis?

In an attempt to understand the mechanism by which estrogens stimulate cell proliferation and mammary carcinogenesis, metastatic human breast cancer cell lines (MCF7, ZR75-1) were found to secrete a 52,000 dalton (52K) protein under estrogen stimulation. Following its purification to homogeneity, the 52K protein was identified as a secreted procathepsin-D-like aspartyl protease bearing mannose-6-phosphate signals. This precursor displays an in vitro autocrine mitogenic activity on estrogen-deprived MCF7 cells and is able to degrade basement membrane and proteoglycans following its autoactivation. The total protease (52K + 48K and 34K) was detected and assayed by monoclonal antibodies and was found to be highly concentrated in proliferative and cystic mastopathies. In breast cancer, its cytosolic concentration appears to be correlated more to tumor invasiveness than to hormone responsiveness. The mRNA of the 52K protease accumulates rapidly following estradiol treatment, as was shown by Northern blot analysis with cloned cDNA. The 52K cathepsin-D-like protease is the first example of a lysosomal protease induced by estrogens in cancer cells. Results obtained using different approaches suggest that two cysteinyl cathepsins are also related to cell transformation and invasiveness. It has been proposed that cathepsin-B is involved in breast cancer and metastatic melanoma, and its regulation by estrogen has been shown in the rat uterus. Cathepsin-L corresponds to the major excreted protein (MEP) whose synthesis and secretion are markedly increased by transformation of NIH 3T3 cells with Ki ras and are regulated by several growth factors. In addition to secreted autocrine growth factors and to other proteases (plasminogen activator, collagenase), lysosomal cathepsins may therefore play an important role in the process of tumor growth and invasion as long as their precursor is secreted abundantly.

Breast Neoplasms↗
PubMed1987

Difference between R5020 and the antiprogestin RU486 in antiproliferative effects on human breast cancer cells.

We have compared the effects of the progestin R5020 and the antiprogestin RU486 on the growth of the MCF-7 and T47D breast cancer cell lines. Differences between the two compounds were demonstrated in several parameters. 1. Estradiol was required for the efficient inhibition of cell growth of both lines by R5020 but not by RU486. Therefore in the total absence of estrogen (phenol-red free medium), the effects of the two drugs on cell growth were dissociated, RU486 remaining inhibitory while R5020 was inactive. 2. The proteins secreted by cells were differently affected, since R5020 induced a 48K protein and decreased the production of the estrogen-regulated 52K protein, while RU486 had no effect on these two parameters. 3. The morphology of cells treated by R5020 was more altered in the presence of estradiol than in its absence, while that of cells treated by RU486 was not affected whether or not estradiol was present. 4. There was a greater reduction of estrogen receptor sites in MCF-7 cells produced by R5020 than by RU486. Even though the two drugs appear to act through the same progesterone receptor and to inhibit total protein secretion, it is likely that they exert their antiproliferative effects on cultured breast cancer cells by different mechanisms. R5020 antagonizes the stimulation produced by estradiol. RU486 by contrast exerts a more direct progesterone receptor mediated inhibitory effect requiring no synergism by estradiol and therefore does not act through a partial progestin activity.

Breast Neoplasms↗
PubMed1987

The estrogen-regulated 52K-cathepsin-D in breast cancer: from biology to clinical applications.

We have studied estrogen-regulated proteins in an attempt to understand the mechanism by which estrogens stimulate cell proliferation and mammary carcinogenesis. In estrogen receptor positive human breast cancer cell lines (MCF7, ZR75-1) estrogens specifically increase the production into the culture medium of a 52,000 daltons (52K) glycoprotein. Several high affinity monoclonal antibodies to the partially purified secretory 52K protein have allowed to purify to homogeneity this protein and its cellular processed products. The 52K protein has been identified as the secreted precursor of a cathepsin-D like protease bearing mannose-6-phosphate signals and routed to lysosomes via mannose-6-phosphate receptor. The protease is mitogenic in vitro on estrogen deprived MCF7 cells and is able to degrade basement membrane and proteoglycans following its activation. The cellular related proteins, as detected by immunohistochemistry and immunoassay are more concentrated in proliferative mammary ducts than in resting ducts and their concentration in breast cancer cytosol appears to be more correlated with lymph nodes invasion and disease free survival (with S. Thorpe, Copenhagen) than with the estrogen receptor (RE) level. The protein is also produced constitutively by RE-negative cell lines, while in some antiestrogen resistant variants, it becomes inducible by tamoxifen, contrary to the wild type MCF7 cells. Cloning of its cDNA in lambda gt11 has allowed to show that the mRNA is rapidly induced by estrogens and to sequence the protein and compare it to that of the normal human kidney cathepsin-D.(ABSTRACT TRUNCATED AT 250 WORDS)

Breast Neoplasms↗
PubMed1987

Phosphorylation, glycosylation, and proteolytic activity of the 52-kD estrogen-induced protein secreted by MCF7 cells.

We have studied the posttranslational modifications of the 52-kD protein, an estrogen-regulated autocrine mitogen secreted by several human breast cancer cells in culture (Westley, B., and H. Rochefort, 1980, Cell, 20:353-362). The secreted 52-kD protein was found to be phosphorylated mostly (94%) on high-mannose N-linked oligosaccharide chains, and mannose-6-phosphate signals were identified. The phosphate signal was totally removed by alkaline phosphatase hydrolysis. The secreted 52-kD protein was partly taken up by MCF7 cells via mannose-6-phosphate receptors and processed into 48- and 34-kD protein moieties as with lysosomal hydrolases. By electron microscopy, immunoperoxidase staining revealed most of the reactive proteins in lysosomes. After complete purification by immunoaffinity chromatography, we identified both the secreted 52-kD protein and its processed cellular forms as aspartic and acidic proteinases specifically inhibited by pepstatin. The 52-kD protease is secreted in breast cancer cells under its inactive proenzyme form, which can be autoactivated at acidic pH with a slight decrease of molecular mass. The enzyme of breast cancer cells, when compared with cathepsin D(s) of normal tissue, was found to be similar in molecular weight, enzymatic activities (inhibitors, substrates, specific activities), and immunoreactivity. However, the 52-kD protein and its cellular processed forms of breast cancer cells were totally sensitive to endo-beta-N-acetylglucosaminidase H (Endo H), whereas several cellular cathepsin D(s) of normal tissue were partially Endo H-resistant. This difference, in addition to others concerning tissue distribution, mitogenic activity and hormonal regulation, strongly suggests that the 52-kD cathepsin D-like enzyme of breast cancer cells is different from previously described cathepsin D(s). The 52-kD estrogen-induced lysosomal proteinase may have important functions in facilitating the mammary cancer cells to proliferate, migrate, and metastasize.

Breast Neoplasms↗
PubMed1987

[Study of salivary cortisol using radioimmunological assay. Diagnostic value].

Salivary cortisol levels reflect the biologically active "free" fraction of blood cortisol. The authors describe the results obtained with the aim of a radio-immunoassay commercial serum cortisol kit, without prealable extraction in different physiological and pathological situations. Salivary cortisol determination appears performant both in nycthemeral studies and in stimulation or freination tests.

Adult↗