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Biomedical subjects

G Leclercq

Publications and source records attributed to G Leclercq.

At least 163 records · Page 9Linked to original sources
PubMed1985

In vitro synergism between estrogens and cytotoxic agents.

Two lines of research intending to achieve synergism between cytotoxic agents and estrogens for breast cancer treatment are pursued in our laboratory. According to a screening procedure we select cytotoxic-linked estrogens which bind to estrogen receptors and thereby would be specifically concentrated into the tumor cells. A mesylate derivative of estrone has emerged from our investigations. This compound displays a very strong binding affinity for the receptors and inhibits the growth of the receptor-positive MCF-7 breast cancer cell line. The lack of growth inhibition in the receptor-negative line Evsa-T indicates that it is devoid of major non-specific cytotoxicity. We attempt to enhance the vulnerability of the tumor cells by producing an estrogen-induced modification of their chromatin. The in vitro exposure of isolated uterine nuclei to uterine cytosol preincubated with estradiol increases their ability to bind [3H]actinomycin D. Identical results are obtained with MXT mouse mammary tumors. Experiments are in progress to settle whether these changes are associated with enhanced cell killing.

Animals↗
PubMed1985

Assay for estrogen and progesterone receptors of breast cancer cell lines in monolayer culture.

A whole-cell assay for measuring estrogen (ER) and progesterone (PgR) receptors in monolayer culture of human breast cancer cell lines is described. It is based on the measurement of incorporated tritiated ligands during 50 min of incubation (i.e. [3H]estradiol for ER, [3H]ORG-2058 for PgR). The assay fulfills all criteria of specificity as shown by competitive studies and measurements of the dissociation constants of the binding reactions. Moreover, a subcellular fractionation of MCF-7 labeled cells revealed that the majority of incorporated steroids was associated with the nuclear fraction. This finding is consistent with the concept of nuclear location of steroid-receptor complexes. Cultures in the presence of 10(-8) M estradiol indicated that the methodology is adequate for detecting the well-known estrogenic induction of PgR synthesis. The assay proved suitable for the quantitative assessment of the receptor content of various neoplastic (MCF-7; ZR-75-1, Cama-1, Evsa-T) and non-neoplastic (HBL-100) cell lines. The methodology has the other advantages of being simple and rapid, of requiring small amounts of cells and of allowing histological examination of the latter before, during and after biochemical analysis.

Binding, Competitive↗
PubMed1985

Chemiluminescence assay of phagocyte activity in mice bearing neuroblastoma: effect of in vitro culture of the tumor.

Phagocyte activity in mice bearing neuroblastoma (NB) has been assayed using a chemiluminescence (CL) technique and proves to be dependent on the pretreatment of the NB cells used to induce the tumor. Indeed, original NB C 1300 tumor, which is maintained by serial in vivo passages induces highly triggered peritoneal phagocytes, producing large amounts of reactive oxygen intermediates (ROI). On the contrary, C 1300 cells cultured in vitro for a few passages and established lines as well as clones derived from the original tumor fail to induce this triggering. However, this decreased phagocyte stimulation can be regained at full extent after a few in vivo passages of the tumor cells.

Animals↗
PubMed1985

Hydroxy derivatives of tamoxifen.

In the exploration of the structural features that affect the RBA (binding affinity for the estrogen receptor of rat uterus relative to that of estradiol) in the tamoxifen [trans-(Z)-1-[4-[2-(dimethylamino)ethoxy]phenyl ]-1,2-diphenyl-1-butene] series, several derivatives variously substituted in the 1-phenyl group have been synthesized. [In the tamoxifen series, the descriptors E and Z, which define the configuration of the geometrical isomers and depend on the location and nature of substituents in the aromatic moieties and the ethyl group, may vary, although the relative configuration (cis or trans) does not. In order to avoid confusion the terms cis and trans will be used in this paper to refer to the relative positions of the 4-[2-(dimethylamino)ethoxy]phenyl and ethyl (or hydroxyethyl, hydroxypropyl, or bromo) substituents attached to the ethene moiety.] The final stage of each synthesis involved acid-catalyzed dehydration of a tertiary alcohol, and, in contrast to the known 3- and 4-hydroxy derivatives which were obtained as near-equimolar cis,trans mixtures, only the trans forms of the 2-hydroxy, 2-methyl, 2,4-dihydroxy, and 4-hydroxy-2-methyl derivatives were obtained. Also, in contrast to the trans forms of the 3- and 4-hydroxy derivatives, which are readily equilibrated to cis,trans mixtures, the trans 2-hydroxy derivative could not be isomerized. Tamoxifen and 2-methyltamoxifen had similar RBA's (approximately 1% of that of E2), but that of 2-hydroxytamoxifen was much lower (0.1%). Introduction of a second hydroxyl group (2,4-dihydroxy derivative) enhanced the RBA, and for the 4-hydroxy-2-methyl derivative, the RBA and growth inhibitory activity against the MCF-7 mammary tumor cell line in vitro were high and comparable to those of 4-hydroxytamoxifen, a metabolite of the parent drug. Tamoxifen derivatives hydroxylated at positions 3 or 4 of the 1-butene moiety and the 5-hydroxy-1-pentene analogue were also synthesized, but they had very low RBA values.

Binding, Competitive↗
PubMed1985

Histological staining by tritiated actinomycin D: a useful tool to study neoplastic cell heterogeneity and estrogen sensitivity of mammary tumors.

The MXT mouse transplantable mammary cancer is characterized by significant amounts of steroid hormone receptors. Moreover this model, as other solid tumors, contains diverse subpopulations of neoplastic cells. In the present study we have examined this pattern as well as the influence of exogenous physiologic doses of estradiol on tumor cell subpopulations. Their identification was undertaken at the morphological level employing incubation of tritiated actinomycin D with fixed material followed by autoradiography. This semiquantitative cytochemical method has proved itself to be a useful tool in order to assess the cellular heterogeneity of carcinoma cells.

Animals↗
PubMed1985

Tritiated actinomycin-D staining method: a valuable tool to study oestrogen receptor-induced modifications of transcriptional activity in normal and neoplastic cells.

Exposure at 4 degrees C of purified cell nuclei from uterus and hormone-sensitive mammary cancer to their own cytosol preincubated with oestradiol produced a significant increase in their ability to bind [3H] actinomycin D (3H-AMD). This increase did not occur under conditions preventing the transfer of the oestrogen-receptor into the cell nucleus, nor in cancer cell nuclei devoid of oestrogen-receptors. Uterine cytosol preincubated with the strong antioestrogen 4-hydroxytamoxifen did not modify the 3H-AMD binding capacity of the uterine nuclei but significantly suppressed the increase induced by oestradiol. Moreover, ultrastructural study of tumour nuclei revealed that the oestradiol receptor-induced increase was associated with a marked chromatin dispersion. These results strongly suggest that the 3H-AMD staining method is a valuable tool for the assessment of oestrogen receptor-induced modifications of transcriptional activity in normal and pathological tissue as well.

Animals↗
PubMed1985

Effect of castration and 17 beta-estradiol pulse on cell proliferation in the uterus and the MXT mouse mammary tumor.

The effects of a single 17 beta-estradiol (E2) injection on cell proliferation were studied in 3 groups of 30 mice transplanted with the MXT ovary-dependent mammary tumor. In group A, all animals were castrated prior to tumor implantation; groups B and C had intact ovaries at the time of transplantation, but group B was left intact throughout the experiment, while group C underwent castration 4 weeks later. On day 40 in groups B and C and on day 80 in group A, in which tumor development was significantly delayed, the same procedure for testing the effects of E2 was applied: Ten controls received 0.1 ml saline ip and were killed on the next day; 4 lots of 5 mice received 0.25 micrograms E2 ip and were killed one by one at 12-hour intervals. Exactly 1 hour prior to sacrifice, each animal received 25 microCi [methyl-3H]thymidine ip. Histologic sections of tumors and uteri were processed for autoradiography, and nuclear thymidine (dThd) labeling indices (LI) were determined. All tumors of group A grafted under unfavorable hormonal conditions were poorly differentiated, and E2 injection induced no appreciable changes in their dThd LI. Tumors B and C were well-differentiated adenocarcinomas, in which E2 induced significant modifications of cell proliferation. In group B, complex changes in dThd LI occurred in tumors as well as in uteri, probably due to interferences with the ovarian hormonal production. In group C, E2 produced a marked rise in dThd LI in tumors, lasting from the 12th to the 36th hour after its injection. Stimulation was maximum at the 24th hour, representing a 2.8-fold increase over mean basal dThd LI. It is concluded that the presence of an intact ovarian function at the time of transplantation is critical for maintaining the properties of hormone dependence in MXT tumors. In mice castrated after tumor implantation, a single E2 injection induces a marked and partially synchronous proliferation of neoplastic cells. The hypothesis that such hormonal manipulation might amplify the killing effect of cell cycle- or phase-specific cytotoxic drugs could be adequately tested with this model.

Adenocarcinoma↗
PubMed1985

[New method for the determination of estrogen and progesterone receptors in human breast cell lines].

A method for the determination of estrogen and progesterone receptor levels in human mammary cell lines (MCF-7, Cama-1, ZR-75-1, Evsa-T and HBL-100) is described. Cells cultured as monolayers were incubated with the tritiated steroids, [3H]-17 beta-Estradiol or [3H] ORG-2058. Binding of steroids to receptors was a function of cellular uptake. Incubation periods of 50 min were sufficient to attain maximum intracellular incorporation. The binding of 17 beta-E2 and ORG-2058 to MCF-7 cells, a phenomenon which is saturable at low concentrations for the radioactive ligand, is a linear function of the number of cells assayed (Interval: 2.5 X 10(4) to 1.5 X 10(6) cells per well). Binding data and their Scatchard plot allowed for the calculation of affinity and capacity values. Thus, for ER, Kd = 2.0 +/- 0.5 X 10(-10) M and n = 3.76 +/- 0.91 Fmol/microgram DNA, and for PgR Kd = 2.0 +/- 0.2 X 10(-10) M and n = 14.02 +/- 2.30 Fmol/microgram DNA (Mean +/- SD). Binding specificity of 17 beta-Estradiol and ORG-2058 to MCF-7 cells was analysed by means of study on the inhibitory effect of increasing concentrations of unlabelled competitors: 17 beta-Estradiol, ORG-2058, Estrone, DES, R-5020, Cortisol, Androsterone and Testosterone. Only pharmacological doses of some of the mentioned molecules produce displacement of the hormonereceptor binding. This phenomenon appears to be related to the affinity of these chemical compounds for the receptor macromolecules to which estrogens and progesterone bind.

Binding, Competitive↗
PubMed1984

Synthesis and evaluation of novel N-substituted N'-(3-hydroxy-17-oxoestra-1,3,5(10)-trien-2- and -4-yl)thiourea derivatives for binding to the estrogen receptor and cytotoxic activity on MCF-7 cells.

A novel series of estrone derivatives having a free 3-phenolic group with the 2- or 4-position substituted with a thiourea function was synthesized. None of the products showed significant binding to the estrogen receptor, and the cytotoxic activity on MCF-7 cells for VII and X was weak.

Animals↗