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H Rochefort

Publications and source records attributed to H Rochefort.

At least 19 recordsLinked to original sources
PubMed1992

Activation of estrogen receptor transfected into a receptor-negative breast cancer cell line decreases the metastatic and invasive potential of the cells.

Breast cancers containing estrogen receptors are responsive to antiestrogen treatment and have a better prognosis than estrogen receptor-negative tumors. The loss of estrogen and progesterone receptors appears to be associated with a progression to less-differentiated tumors. We transfected the human estrogen receptor into the estrogen receptor-negative metastatic breast cancer cell line MDA-MB-231 in an attempt to restore their sensitivity to antiestrogens. Two stable sublines of MDA-MB-231 cells (HC1 and HE5) expressing functional estrogen receptors were studied for their ability to grow and invade in vitro and to metastasize in athymic nude mice. The number and size of lung metastases developed by these two sublines in ovariectomized nude mice was not markedly altered by tamoxifen but was inhibited 3-fold by estradiol. Estradiol also significantly inhibited in vitro cell proliferation of these sublines and their invasiveness in Matrigel, a reconstituted basement membrane, whereas the antiestrogens 4-hydroxytamoxifen and ICI 164,384 reversed these effects. These results show that estradiol inhibits the metastatic ability of estrogen receptor-negative breast cancer cells following transfection with the estrogen receptor, whereas estrogen receptor-positive breast cancers are stimulated by estrogen, indicating that factors other than the estrogen receptor are involved in progression toward hormone independence. Reactivation or transfer of the estrogen receptor gene can therefore be considered as therapeutic approaches to hormone-independent cancers.

Animals↗
PubMed1992

Inhibition of breast cancer growth by suramin.

In this study, the polyanionic compound suramin was shown to be a potent in vitro growth inhibitor of both hormone-insensitive, estrogen receptor-negative human breast cancer cells (MDA MB231 and SK-BR-3) and hormone-responsive, estrogen receptor-positive human breast cancer cells (ZR 75-1, T47D, and MCF7). The inhibitory effect of suramin was dose dependent, with a median effective dose varying from 7 microM for MDA MB231 cells to 50 microM for MCF7 cells. This result indicated that estrogen receptor-negative cells were more sensitive to the drug. In MCF7 cells, not only did suramin block the mitogenic action of growth factors such as epidermal growth factor (EGF) and insulin-like growth factors I and II (IGF-I and IGF-II, respectively), but it also totally abolished the increase in cell proliferation induced by the steroid hormone 17 beta-estradiol (E2). Maximal inhibition was obtained after 5 days of suramin treatment, and inhibition either was partially reversed by E2, IGF-I, and IGF-II or was not reversible by EGF following removal of drug. In addition, suramin significantly decreased synthesis and secretion of the lysosomal enzyme cathepsin D, which was shown to be associated with a high risk of breast tumor metastasis. These results therefore suggest that, because of its effects on growth and cathepsin D secretion, suramin might be a helpful additional therapeutic tool for breast cancer patients, especially for patients with estrogen receptor-negative tumors which are insensitive to antihormonal strategies.

Breast Neoplasms↗
PubMed1992

Interactions of pro-cathepsin D and IGF-II on the mannose-6-phosphate/IGF-II receptor.

The mannose-6-phosphate (Man-6P)/IGF-II receptor is a multifunctional receptor which binds with a high affinity on distinct sites two strikingly different classes of ligands: IGF-II, and Man-6P bearing molecules such as lysosomal enzymes or other biologically relevant ligands (TGF beta precursor, EGF receptor, proliferin...). Binding of each ligand on its cognate site is severely decreased in the presence of the other type of ligand, thus revealing that the two distinct sites are strongly interacting (steric hindrance, conformational change). Any imbalance in ligands and receptor concentration in various pathological situations (transformation, tumor, altered hormonal levels...) is thus likely to perturb their associated biological functions in the targeting and routing of lysosomal enzymes or Man-6P ligands or in the autocrine/paracrine IGF-II--induced cellular responses.

Binding Sites↗
PubMed1992

Immunostaining of cathepsin D in breast cancer: quantification by computerised image analysis and correlation with cytosolic assay.

Cathepsin-D (cath-D) was quantified in 34 breast cancer specimens by immunohistochemical staining of frozen sections with a computer image analysis and the results were compared with the corresponding cytosolic assay. Cath-D concentrations varied from 0 to 420 arbitrary units (AU). Tumour cells were more intensely stained than peritumoral tissue with the D7E3 mouse monoclonal antibody than with rabbit polyclonal antibodies. There was a good correlation (r = 0.80) between cath-D values obtained either by immunohistochemistry with D7E3 antibody or by cytosolic immunoenzymatic assay. However, with a cut-off of 50 AU, 3 out of 25 patients had higher immunohistochemical values and 2 had higher cytosolic values. Therefore, quantification of cath-D concentration in tissue section by immunostaining and a computerised image analyser, which is the only technique available for small tumours, should provide similar prognostic information to that obtained by assaying cath-D in the cytosol.

Animals↗
PubMed1992

Estradiol stimulates cell growth and secretion of procathepsin D and a 120-kilodalton protein in the human ovarian cancer cell line BG-1.

Ovarian cancers are highly invasive. In a first attempt to define the hormones and factors involved in the control of tumor invasion and metastasis, we have used the human ovarian cancer cell line BG-1 which contains both estrogen and progesterone receptors. Protein synthesis and secretion was assayed by [35S]methionine incorporation and polyacrylamide gel electrophoresis followed by fluorography. Three responses to estradiol were found: 1) procathepsin D secretion was increased, whereas the corresponding intracellular proteins were not significantly affected; 2) an abundant but nonidentified 120-kilodalton (kDa) estrogen-induced secreted glycoprotein, different from CA125, was detected for the first time; and 3) the number of cells as determined by DNA assay was markedly stimulated, reaching a higher level of confluency. The antiestrogen OH-tamoxifen was weakly agonist at low concentrations to stimulate cell growth but was a pure antagonist on the 120-kDa protein. The steroid specificity of these responses strongly suggests that they are mediated by the estrogen receptor. We conclude that cathepsin D secretion is specifically stimulated by estrogen in this ovarian cancer cell line as it is in estrogen receptor-positive breast cancer cells. Both cathepsin D and a newly described 120-kDa secreted glycoprotein are potential markers of hormone responsiveness and/or aggressiveness which deserve to be further studied in clinical ovarian cancers.

Cathepsin D↗
PubMed1992

Biological and clinical significance of cathepsin D in breast cancer.

Cathepsin D, an aspartyl protease of lysosomes, is overproduced and hypersecreted by breast cancer cells. The prognostic value of its immunoassay in breast cancer cytosol is reviewed from the first retrospective clinical studies available, which show a strong correlation between high concentrations of cathepsin D in the cytosol of primary tumor and further occurrence of metastasis. This new prognostic factor is induced by estrogen in hormone dependent breast cancer but expressed at a high level in hormone independent breast cancer and appears to be independent of other more classical factors. Its value in node negative patients varies according to the studies. In nude mice, transfection of cathepsin D cDNA into tumor cells increases their metastatic potential, suggesting that overexpression of this protease may be one of the factors responsible for metastasis in human breast cancer. The mechanism by which this protease might facilitate metastasis in vivo is still unknown, even though cathepsin D has the potential to initiate a proteolytic cascade, to degrade extracellular matrix and to liberate FGFs like growth factors from the matrix. These studies should stimulate the search for new therapeutical agents in order to inhibit cathepsin D action.

Animals↗
PubMed1992

Cathepsin D concentration in breast cancer cytosols: correlation with disease-free interval and overall survival.

Cathepsin D (CD) is an aspartyl protease implicated in cancer metastasis. In this study of 331 patients, we show that patients with primary breast carcinomas containing high concentrations of CD have a significantly shorter disease-free interval (chi-square = 4.28, P < 0.05) and overall survival (chi-square = 7.7, P < 0.01) than patients with low concentrations. CD as a prognostic marker for overall survival was equally valuable for women younger (chi-square = 4.39, P < 0.05) and older (chi-square = 3.97, P < 0.05) than 50 years. CD was also a significant prognostic marker for overall survival within the estradiol receptor (ER)-positive subgroup of patients (chi-square = 5.79, P < 0.025), but not in the ER-negative subgroup. Patients with tumors containing high concentrations of CD and low concentrations of ER had shorter disease-free intervals (chi-square = 15.1, P < 0.001) and lower overall survival (chi-square = 20.9, P < 0.001) than patients with high concentrations of ER but low concentrations of CD.

Biomarkers, Tumor↗
PubMed1991

The anti-progestin RU486 stabilizes the progestin-induced fatty acid synthetase mRNA but does not stimulate its transcription.

Progestins induce fatty acid synthetase (FAS) in breast cancer cell lines, both increasing its gene transcription and mRNA stabilization (Joyeux, C., Rochefort, H., and Chalbos, D. (1989) Mol. Endocrinol. 4, 681-686). In vitro run-on transcription assays show that RU486, in contrast to progestin, inhibits FAS transcription by 40-50%. Moreover and surprisingly, anti-progestin RU486 also stabilizes FAS mRNA 3- to 4-fold in MCF7 cells as measured by chase experiments in the presence of actinomycin D or cordycepin or after short cell labeling with [3H]uridine. Dexamethasone is inefficient in increasing the half-life of FAS mRNA in MCF7 cells. RU486 had no effect on MDA-MB 231 cells which contain glucocorticoid but no progesterone receptors, indicating that the progesterone receptor is implicated in this regulation. RU486-induced mRNA stabilization allows delayed accumulation of FAS mRNA. These results indicate that the progesterone receptor can be activated separately to stimulate gene transcription or stabilize mRNA.

Cell Line↗
PubMed1991

Cathepsin D gene of human MCF7 cells contains estrogen-responsive sequences in its 5' proximal flanking region.

Cathepsin D is a lysosomal protease produced and secreted in excess by most human breast cancer cells. In MCF7 cells, estrogens stimulate cathepsin D expression at the mRNA level via a mechanism independent of de novo protein synthesis. We have isolated the human cathepsin D gene and its 5' flanking sequences from a MCF7 genomic library. To demonstrate its transcriptional estrogen regulation, we constructed chimeric recombinants bearing different fragments of the cathepsin D gene 5' proximal region inserted in front of the bacterial chloramphenicol acetyl transferase reporter gene. By transient cotransfection with the estrogen receptor expression vector into MCF7 cells, we showed that a 240 bp fragment located in the 5' proximal region of the gene was able to mediate transcriptional estrogen activation. This induction was concentration-dependent and suppressed by the antiestrogen ICI 164, 384.

Breast Neoplasms↗
PubMed1991

Progestin treatment depresses estrogen receptor but not cathepsin D levels in needle aspirates of benign breast disease.

We studied the effect of a progestin (lynestrenol) on estrogen receptors (ER) and cathepsin D (cath-D) levels immunochemically in successive fine needle aspirates of benign breast disease. Fibrocystic disease was the main pathology (43 out of 47 patients). Thirty-one patients were treated with 10 mg of lynestrenol daily from the fifth to the twenty-fifth day of the menstrual cycle for 1 to 3 months. Sixteen untreated patients were used as controls. Lynestrenol significantly decreased the percentage of ER stained cells. This is in agreement with the antiestrogenic effect of progestin and, for the first time, indicates that in vivo progestin may decrease the stimulatory effect of estrogens on mammary cells by decreasing their estrogen receptor content. No effect of progestin on cath-D level was found throughout the whole population. However, this level varied more between aspirates of each patient in the treated group than in the control group, suggesting heterogeneity in patient responses to progestin. Since cath-D may have a role in carcinogenesis, clinical follow-up of these patients and more detailed studies are required to determine whether this progestin-challenge test has any value for detecting high risk mastopathies and for predicting effectiveness of treatment.

Adult↗
PubMed1991

MCF7 mammary cancer cells respond to bFGF and internalize it following its release from extracellular matrix: a permissive role of cathepsin D.

High and low affinity receptors for basic fibroblast growth factor (bFGF) were detected by binding experiments on MCF7 breast cancer cells. These cells were stimulated for growth by physiological concentrations of bFGF. However, in contrast to endothelial cells, these MCF7 cells did not produce detectable amounts of biologically active bFGF or related heparin-binding growth factor(s) of the FGF family. In vitro, the cathepsin D (cath-D) secreted by MCF7 cells was able to digest extracellular matrix (ECM) and to release ECM-bound 125I-bFGF. When MCF7 cells were cultured on ECM containing bound bFGF, they internalized bFGF, which was slowly and partially proteolyzed in the cells. Processing occurred in acidic compartments and was inhibited by leupeptin. Pepstatin A, an inhibitor of aspartyl proteases, had no effect on the processing but reduced internalization of matrix-bound bFGF by MCF7 cells. Taken together, these results suggest a cooperation between cath-D and bFGF, by which the protease could facilitate the release of bFGF from ECM and its subsequent use by breast cancer cells and/or adjacent cells involved in angiogenesis.

Breast Neoplasms↗
PubMed1991

Hormonal regulation of cathepsin D following transfection of the estrogen or progesterone receptor into three sex steroid hormone resistant cancer cell lines.

The cathepsin D gene is differentially regulated by estrogens in hormone responsive breast cancer cells, by progestins in normal human endometrium and is highly expressed but not regulated by these steroids in estrogen (RE)- and progesterone receptor (RP)-negative breast cancer cells. We have stably transfected the RE-negative breast cancer cell line MDA-MB 231 and the Hela cell line with an expression vector for the human RE. The endogenous cathepsin D which is constitutively expressed was further stimulated by estradiol. However, the growth of both cell lines was not stimulated by estradiol and could not be inhibited by the antiestrogen ICI 164,384. By contrast, the cathepsin D gene in the estrogen responsive Ishikawa endometrial cancer cell line was unresponsive to estrogen or to progesterone even following stable transfection of expression vectors for the RP (both A and B isoforms). We conclude that the cathepsin D gene is potentially responsive to estrogens in MDA-MB 231 and Hela cells, which therefore express all of the transcriptional machinery (except the RE) necessary for this regulation. By contrast, cathepsin D remains unresponsive to estrogen and progesterone in Ishikawa cells. The cathepsin D gene is one of the first examples of an endogenous steroid responsive gene which can be controlled by steroids following stable transfection of a steroid receptor.

Breast Neoplasms↗
PubMed1991

Increased plasma cathepsin D concentration in hepatic carcinoma and cirrhosis but not in breast cancer.

Using a sandwich enzyme-linked immunoassay, plasma total cathepsin D concentration was assayed in 40 breast cancer patients and 84 patients with various liver diseases and compared to that of 52 normal subjects. There were no significant variations found in breast cancer patients related to tumor size, node invasiveness or metastases. In normal women, cathepsin D levels were slightly but not significantly increased in the luteal phase and in pregnancy. By contrast, plasma cathepsin D concentration was significantly increased in 70-75% of patients with liver disease (cirrhosis, hepatocarcinoma, hepatitis), but not in those with liver steatosis. Cathepsin D was independent of most of the plasma hepatic function tests and was correlated with alpha-fetoprotein in cirrhosis and with alpha-fucosidase in primary hepatocellular carcinoma. We conclude that plasma cathepsin D is not a useful marker in breast cancer. However, since the cellular level of this protease is associated with risk of metastasis in breast cancer, clinical follow-up will be required to test whether high cathepsin D plasma concentration has any prognostic value in liver cirrhosis and primary hepatocarcinoma.

Breast Neoplasms↗
PubMed1991

Mechanism of action of high-affinity antiestrogens. An overview.

This is a short review on the mechanism of action of high-affinity antiestrogens at the cellular and molecular level. Past and recent data from this and other laboratories are discussed concerning the mode of action of antiestrogens in controlling gene expression and cell proliferation, including their mode of action as inhibitors of growth factor action and oncogene expression. It has been proposed that antiestrogens may not simply act to inhibit estrogen action but may also directly trigger cell growth inhibition via the estrogen receptor in the absence of estrogen. "Estrogen receptor mediated growth inhibitors" may therefore be a more appropriate and general term for defining these estrogen analogues.

Animals↗
PubMed1991

Estradiol down-regulates the mannose-6-phosphate/insulin-like growth factor-II receptor gene and induces cathepsin-D in breast cancer cells: a receptor saturation mechanism to increase the secretion of lysosomal proenzymes.

We have studied the regulation by estradiol of the mannose-6-phosphate (Man-6-P)/insulin-like growth factor-II (IGF-II) receptor concentration in different breast cancer cell lines. The mRNA level was assayed by Northern blot using the H5.1 cDNA probe. The protein level was assayed by Western ligand blot, by binding saturation with [125I]procathepsin-D on total membrane preparations, and by immunoprecipitation of 35S-labeled proteins. In three estrogen receptor-positive cell lines (MCF7, T47D, and ZR75-1), estradiol specifically decreased the steady state level of the Man-6-P/IGF-II receptor protein and mRNA. Moreover, in different cell lines and in primary culture of normal mammary cells, the secretion of procathepsin-D was inversely correlated with the level of Man-6-P/IGF-II receptor protein and mRNA. We conclude that estradiol down-regulates the Man-6-P/IGF-II receptor in breast cancer cells. Since two of its ligands, procathepsin-D and IGF-II, are induced by estrogen, we propose that the Man-6-P/IGF-II receptor becomes saturated after estrogen treatment. This model might explain the previously described estrogen-induced secretion of procathepsin-D and other lysosomal proenzymes routed by the same transport system.

Breast Neoplasms↗
PubMed1991

Decrease of c-erbB-2 and c-myc RNA levels in tamoxifen-treated breast cancer.

The c-myc, c-erbB-2, hst and int-2 oncogenes are frequently amplified and/or overexpressed in human breast carcinomas. We studied the effect of tamoxifen on RNA levels of these oncogenes in 19 breast cancer patients treated for 3 weeks prior to surgery as compared with 22 control patients. RNA levels were measured by in situ hybridization coupled with computer-aided quantification. c-myc and c-erbB-2 expression was high in the control population (mean values: 23.4 and 29.1 grains/cell respectively) and significantly decreased in the tamoxifen-treated population (mean values: 14.6 and 7.4 grains/cell respectively) (P = 0.018, P = 0.003 respectively); hst and int-2 RNA levels were low (2-6 grains/cell) and not significantly altered by the treatment. There was a correlation between gene amplification and expression for c-erbB-2 (P = 0.0005) and hst (P = 0.02) in the control population. Elevated c-erbB-2 RNA level was correlated with the absence of estrogen (P = 0.02) or progesterone (P = 0.05) receptors. In the ER+ population, the tamoxifen-treated group had significantly lower c-myc expression levels than the control group (P = 0.04) which is in agreement with the estrogen induction of c-myc in ER+ T47D cell line and its inhibition by antiestrogens. Surprisingly, c-erbB-2 expression in the tamoxifen-treated group was significantly diminished in the ER- (P = 0.02) and PR- (P = 0.01) populations. This effect was not observed in the ER- BT474 cell line. These results suggest that in vivo tamoxifen decreases c-myc and c-erbB-2 RNA levels in breast cancer cells via two different mechanisms. To our knowledge this is the first evidence of in vivo down regulation of a gene by tamoxifen in ER- breast cancer cells.

Adult↗