[Report by the German Society of Senology].
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Breast Neoplasms↗
Biomedical subjects
K D Schulz
Publications and source records attributed to K D Schulz.
European guidelines for quality assurance in the surgical management of mammographically detected lesions. European Breast Cancer Working Group.
The European Guidelines developed for mammography screening have contributed to the general discussion on quality assurance and the important tasks of the health professionals dealing with breast cancer screening. The cooperation of each medical discipline is of utmost importance in order to achieve optimal results and eventually a mortality reduction. The following guidelines are based on the British NHS quality assurance guidelines for surgeons in breast cancer screening and modified to meet the different needs in the European Countries. The term "surgeon" denotes a medical doctor trained and involved in the surgical treatment of breast diseases. The members of the working group who participated in order to adapt these guidelines are listed above. We hope that this document will contribute towards a more comprehensive approach of breast cancer screening-detected lesions throughout Europe.
Breast Neoplasms↗
Establishment of new epithelial carcinoma cell lines by blocking monolayer formation.
Human endometrial carcinoma cell lines were established by using a new cell culture technique. The malignant endometrial tumors were grossly disaggregated by mechanical means and cultivated in suspension culture. Adhesion to the bottom of the culture flasks was prevented by first coating the flasks with a thin agarose layer. Four cell lines were derived from 17 samples by this new technique. The cell lines obtained in this way were fully characterized, including karyotyping, intermediate filament staining and transplantation to nude mice. This new technique of initial suspension culture may also be applicable to other human tumors that are equally difficult to cultivate in vitro.
Animals↗
[After care of breast carcinoma patients--recommendations from the Berlin consensus conference].
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Aftercare↗
Androgen-like and anti-androgen-like effects of antiprogestins in human mammary cancer cells.
In addition to their antiprogestational activity, the antiprogestins RU486, ZK98.299 and ZK98.734 possess varying antiglucocorticoid as well as androgen-like or antiandrogen-like properties in human mammary cancer cells. The human mammary cancer cell line MFM-223, which contains only androgen receptors, was used as a model to investigate androgen receptor mediated effects of these antiprogestins. Proliferation of MFM-223 cells is inhibited by androgens and does not respond to oestrogens, progestins and glucocorticoids. As shown in proliferation assays, ZK98.734 was a strong inhibitor of cell proliferation. This effect was antagonised by the antiandrogen hydroxyflutamide. ZK98.734 was found to displace [3H]R1881 from the androgen receptor in MFM-223 cells, substantiating the involvement of the androgen receptor. The antiprogestin ZK98.299 failed to influence the proliferation of MFM-223 cells. ZK98.299 did not bind to the androgen receptor and was devoid of androgenic or antiandrogenic activity. RU486 bound to the androgen receptor. It was a weak inhibitor of MFM-223 cell proliferation, but the inhibition of proliferation by RU486 was not antagonised by hydroxyflutamide. This effect was probably not mediated by the androgen receptor. RU486 had antiandrogenic activity in this cell line, as it antagonised the inhibitory effect of dihydrotestosterone at a 100-molar excess. These results were confirmed by transfection experiments with an MMTV-CAT construct in the same cell line, demonstrating the biological function of the ZK98.734-androgen receptor complex. ZK98.299 and RU486 were not able to induce CAT activity. The different androgenic or antiandrogenic properties of the antiprogestins investigated should be considered when selecting antiprogestational properties of the antiprogestins investigated should be considered when selecting antiprogestational compounds for clinical applications, as a partial androgenic activity may be of benefit in breast cancer but can have undesired side-effects in other diseases.
Androgen Antagonists↗
Androgen receptor mediated growth control of breast cancer and endometrial cancer modulated by antiandrogen- and androgen-like steroids.
Androgens are involved in many regulatory processes in mammary and endometrial epithelium, but their role in the development and progression of breast and endometrial carcinoma is poorly understood. Androgen receptors (AR) are found in normal epithelium as well as in more than 50% of specimen from both tumor types. The occurrence of AR is correlated with estrogen and progesterone receptors. Androgen receptor positive cell lines were established during the last few years in our laboratory from malignant mammary (MFM-223) and endometrial (MFE-296) tumors supplementing the small number of androgen-responsive cell lines published so far. In this paper some aspects of the role of androgens in these two types of hormone responsive female cancers are presented. The proliferation of ZR-75-1, MFM-223 and MFE-296 cells is inhibited by androgens. The progestin medroxyprogesterone acetate inhibits the proliferation of estrogen- and progesterone receptor negative MFM-223 cells via the androgen receptor. Some steroid metabolites with distinct estrogenic properties like androst-5-ene-3 beta,17 beta-diol possess androgenic properties in this model system. Androgens stimulate the in vitro secretion of gross cystic disease fluid proteins by human mammary cancer cells. These proteins are normally found in benign breast cysts in vivo. The occurrence of gross cystic disease is correlated with an increased risk of breast cancer. The AR is autoregulated in MFM-223 mammary cancer cells on the protein and mRNA level. In MFE-296 cells with endometrial origin AR protein was increased after incubation with androgens.
Androgen Antagonists↗
Modulatory actions of progesterone on gonadotropin-releasing hormone-induced arachidonic acid liberation from perifused rat pituitary cells.
The stimulatory action of gonadotropin-releasing hormone (GnRH) on gonadotropin secretion from cultured rat pituitary cells is modulated by estradiol and progesterone. Recent studies provided evidence that both steroids exert effects on different pathways of GnRH signal transduction, which might be responsible for their actions on luteinizing hormone (LH) release. Here we investigated whether the steroids are able to modulate GnRH-induced liberation of arachidonic acid, which is thought to be involved in GnRH signal transduction. Pituitary cells obtained from female rats were treated for 48 h with vehicle, 1 nmol/l estradiol or 1 nmol/l estradiol + 100 nmol/l progesterone, 48 h with 1 nmol/l estradiol and 2 h with 100 nmol/l progesterone. In addition, these cells were prelabeled with [3H]arachidonic acid. Then the cells were transferred to a perifusion system and challenged with a 6-min pulse of 100 nmol/l GnRH. Estradiol treatment enhanced the LH secretory response while GnRH-induced [3H]arachidonic acid liberation remained unaffected. However, progesterone modulated both LH secretion and [3H]arachidonic acid release in response to the GnRH stimulus. The short-term progesterone treatment paradigm enhanced the LH and arachidonic acid responses by up to 160 +/- 13 and 204 +/- 18%, respectively, while long-term treatment was inhibitory (59 +/- 9 and 63 +/- 4% vs control). Because arachidonic acid has been shown to be involved in GnRH signal transduction, it seems reasonable to speculate that the actions of progesterone described in the present study are related to its modulatory effect on GnRH-induced LH secretion.
Animals↗
Expression of the messenger RNAs for luteinizing hormone-releasing hormone (LHRH) and its receptor in human ovarian epithelial carcinoma.
Recently we reported the presence of specific high affinity binding sites for luteinizing hormone-releasing hormone (LHRH) and its analogues (Kd = 1.5 or 1.7 nM) in the human epithelial ovarian cancer cell lines EFO-21 and EFO-27. The proliferation of these cell lines was inhibited by nM concentrations of a LHRH agonist. This study was performed to ascertain whether these ovarian cancer cell lines produce LHRH and whether the high affinity LHRH binding site found previously was identical to the pituitary LHRH receptor. Significant amounts of immunoreactive LHRH were found in the extracts of both the EFO-21 cell line (449 +/- 56 fmol/10(6) cells) and the EFO-27 line (409 +/- 76 fmol/10(6) cells). LHRH bioactivity of these extracts, assessed in terms of release of luteinizing hormone by rat pituitary cells, was comparable to that of authentic LHRH. EFO-21 and EFO-27 cells expressed the mRNAs for both human LHRH and human LHRH receptor as assessed by reverse transcriptase-PCR using oligonucleotide primers according to published sequences. In addition, in eight of eight biopsy samples of human epithelial ovarian cancers we detected mRNA for LHRH, six of these specimens expressed the mRNA representing the LHRH receptor. These data support the concept that human epithelial ovarian cancers might have a local system based on LHRH to regulate cell proliferation. It is still obscure at present whether LHRH produced locally has a stimulatory, inhibitory, or no impact on the proliferation of ovarian cancer cells. However, exogenous LHRH agonists seem to have clear antiproliferative activity, probably mediated through LHRH receptors. This finding might provide the base for novel approaches in the therapy of epithelial ovarian cancer.
Animals↗
Ovarian steroids modulate gonadotropin-releasing hormone-induced biphasic luteinizing hormone secretory responses and inositol phosphate accumulation in rat anterior pituitary cells and alpha T3-1 gonadotrophs.
The ovarian steroids estradiol and progesterone act as important modulators of GnRH-induced luteinizing hormone (LH) secretion from anterior pituitary cells. Recently, we demonstrated that the steroids are able to influence GnRH-stimulated Ca2+ mobilization from extra- and intracellular sources. Here we investigated the actions of estradiol and progesterone on GnRH-induced biphasic LH secretory responses in the model of perifused female rat pituitary cells. A 20 min GnRH stimulus elicited biphasic LH responses composed of an initial peak followed by a prolonged plateau phase. Both phases were equally enhanced by long-term (48 h) estradiol treatment. This action was facilitated by subsequent short-term progesterone treatment. In contrast, combined treatment with estradiol and progesterone for 48 h led to inhibited LH secretory profiles. To determine the steroid actions on the extracellular Ca2+ independent component of LH secretion we performed experiments using cells that were perifused with Ca2+ deficient medium. Under these conditions the cells responded exclusively with a single peak phase of LH secretion, which was augmented or inhibited by estradiol and progesterone treatment as described above. To test the hypothesis that an effect of estradiol and progesterone on GnRH-induced polyphophoinositide hydrolysis is responsible for their modulatory actions on Ca2+ signals and LH secretion we measured inositol phosphate (IP) accumulation after different steroid treatment paradigms in rat pituitary cells and alpha T3-1 immortalized gonadotrophs. GnRH-induced IP production was enhanced by long-term estradiol treatment. Short-term exposure of estradiol-primed cells to progesterone did not lead to significant changes of IP production. The long-term progesterone treatment paradigm enhanced GnRH-induced IP formation, while it decreased Ca2+ signals and LH secretion. Alpha T3-1 cells were used to perform more detailed analysis of IP formation. The actions of estradiol and progesterone on the production of inositol mono-, bis-, and trisphosphates were similar to those observed in the mixed cell population. It is concluded that estradiol and progesterone modulate both peak and plateau phases of GnRH-stimulated LH secretory responses, effects which are associated with their impact on Ca2+ signals. Our findings argue against a role of IP modulation in the mechanism of progesterone actions on Ca2+ signaling and LH secretion in gonadotrophs. Such a mechanism might be involved in the positive effects of estradiol in these cells.
Animals↗
[Metastases in breast carcinoma].
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Breast Neoplasms↗
Androgen responsiveness of the new human endometrial cancer cell line MFE-296.
MFE-296 endometrial cancer cells express androgen receptors in vitro. These cells, which are tumorigenic in nude mice, are derived from a moderately differentiated human endometrial adenocarcinoma. They express vimentin and the cytokeratins 7, 8, 18, and 19. Karyotyping revealed near-tetraploidy for most of the cells. No marker chromosomes were observed. DNA analyses confirmed the genetic identity of the cell line and the patient from whom the cell line was derived. Proliferation of MFE-296 cells was inhibited by the progestin R5020 and the androgen dihydrotestosterone (DHT). The inhibition of proliferation by DHT was antagonized by the antiandrogen Casodex, demonstrating the involvement of the androgen receptor. Androgen binding was determined at 22,000 binding sites per cell using a whole-cell assay (KD = 0.05 nM) and 30 fmol/mg protein with the dextran charcoal method; 7 fmol/mg protein of progesterone receptors were found, whereas estrogen receptors were below 5 fmol/mg protein. The androgen receptor was functionally intact, as demonstrated by transfection experiments with a reporter-gene construct, containing an androgen-responsive element. In MFE-296 cells the content of the androgen receptor was up-regulated by its own ligand.
Androgens↗
Production of insulin-like growth factor binding proteins by human ovarian carcinoma cells.
Cells of the human ovarian carcinoma lines EFO-21, EFO-27, MFO-35 and MFO-36 secrete binding proteins for insulin-like growth factors (IGFBPs) into their culture media. By sodium dodecyl sulfate/polyacrylamide gel electrophoresis (SDS-PAGE) and ligand blotting, seven groups of IGFBPs with molecular masses of 25, 30 (doublet), 34, 37, 40, 45, and 50 kDa were observed, depending on the cell line under investigation. By Northern blot analyses using cDNAs or oligonucleotides specific for the six types of IGFBP (IGFBP-1 to IGFBP-6), mRNA for all IGFBPs tested except for IGFBP-1 could be detected in the ovarian carcinoma cell extracts. In detail, analysis of EFO-21 protein products by SDS-PAGE yielded IGFBPs of 25, 34, and 50 kDa; extracts of EFO-21 cells contained mRNAs for IGFBP-2, -3, -4, and -6. EFO-27 cells produced IGFBPs of 40 kDa and 45 kDa as determined by SDS-PAGE, and mRNAs for IGFBP-3, -4, and -6 were detected. In the conditioned medium of MFO-35 cells, IGFBPs of 25, 30 (doublet), 34, 37, 40, and 45 kDa were observed by SDS-PAGE, while mRNAs for the five proteins IGFBP-2 to IGFBP-6 were found. MFO-36 cells produced IGFBPs of 34 kDa and 50 kDa as determined by SDS-PAGE, and the cells expressed mRNAs for IGFBP-2, -3, -4, and -6. In relation to published molecular mass data of the known IGFBPs, the size of the secreted proteins could be correlated to the mRNA patterns expressed by the ovarian carcinoma cells. It is concluded that ovarian carcinoma cells frequently express IGFBP-3, -4, and -6 and, to a lesser extent, IGFBP-2; the expression of IGFBP-5 appears as a rather rare event, while IGFBP-1 was not found to be expressed in ovarian carcinoma cells.
Carrier Proteins↗
Urinary excretion of growth factors in patients with ovarian cancer.
The levels of epidermal growth factor (EGF) and transforming growth factor-alpha (TGF-alpha) were analysed in 24-h urine samples from patients with ovarian malignancies, benign ovarian tumours, and healthy controls by specific radioimmunoassays. No significant difference in total urinary immunoreactive EGF excretion between the groups was detected. However, 79% (23/29) of the patients with ovarian carcinomas excreted TGF-alpha (median 12.6 pmol/24 h), whereas only 17% (2/12) of the patients with benign ovarian tumours and 23% (3/13) of the controls did so. The difference between cancer patients and controls was highly significant (P < 0.001). Analyses of the urine samples separated by gel filtration revealed a greater molecular heterogeneity of EGF and TGF-alpha in cancer patients than in controls. High and low molecular weight forms of EGF were able to bind to the EGF receptor and to induce anchorage-independent growth. After surgical reduction of the tumour, a distinct decrease of urinary high molecular weight forms was observed. Thus, some macromolecular growth factors seem to be associated with epithelial ovarian carcinomas.
Biomarkers, Tumor↗
Modulation of calcium signaling and LH secretion by progesterone in pituitary gonadotrophs and clonal pituitary cells.
In estradiol-treated pituitary cells, progesterone enhances gonadotropin-releasing hormone (GnRH)-induced LH secretion from cultured rat pituitary cells during short-term treatment but attenuates this response during prolonged treatment. In the present study, the effects of gonadal steroids on GnRH-induced cytoplasmic calcium ([Ca2+]i) responses in gonadotrophs were analyzed in rat pituitary cells and immortalized (alpha T3-1) murine gonadotrophs. Ca2+ responses were measured in cell suspensions and single gonadotrophs, loaded with Fura-2 or Indo-1, respectively, and pretreated for 48 h with 1 nM estradiol with or without 100 nM progesterone, or for 48 h with 1 nM estradiol and then for 3 h with 100 nM progesterone. In cells of the alpha T3-1 gonadotroph lineage, GnRH elicited biphasic Ca2+ signals composed of an initial peak response followed by a prolonged plateau phase. The amplitudes of both the extracellular Ca(2+)-independent spike phase and the extracellular Ca(2+)-dependent plateau phase were enhanced or inhibited by short- or long-term progesterone treatment, respectively. In single pituitary gonadotrophs, GnRH (0.5 nM) elicited oscillatory responses due to intermittent release and uptake of Ca2+ from intracellular stores. Treatment with progesterone shifted the oscillatory signal toward biphasic (3 h) or subthreshold (48 h) response profiles, revealing a steroid-induced change in the pattern of Ca2+ mobilization. In addition to these agonist-induced responses, the transient [Ca2+]i responses of pituitary cells and individual gonadotrophs to high K+ were enhanced or inhibited after short- or long-term progesterone treatment, respectively. These actions were correlated with the effects of progesterone on K(+)-induced LH secretion. The [Ca2+]i and LH secretory responses to phorbol ester treatment were also enhanced by short-term exposure of the cells to progesterone. The results demonstrate that the stimulatory and inhibitory effects of progesterone on agonist-induced Ca2+ signaling result from changes in Ca2+ mobilization and entry, and contribute to the modulatory actions of the steroid on GnRH-induced LH secretion.
Animals↗
Modulatory action of progesterone and progesterone antagonists on hypothalamic-pituitary function.
The ability of ovarian steroids to sensitize and desensitize the pituitary gonadotroph to hypothalamic gonadotrophin-releasing hormone (GnRH) is essential for their modulatory actions on gonadotrophin secretion. The time-dependent actions of progesterone on GnRH-stimulated gonadotrophin secretion from cultured pituitary cells obtained from female rats were examined. Progesterone induced an acute stimulatory effect on luteinizing hormone (LH) and follicle-stimulating hormone (FSH) secretion in cell perifusion studies, from as early as 50 min after the onset of progesterone treatment. Long-term incubation (52 h) of pituitary cells in static culture reduced the responsiveness of the gonadotroph to GnRH. The antiprogestins RU486, ZK 98.299, and ZK 98.734 blocked both the acute facilitatory and the long-term inhibitory action of progesterone. In the absence of progesterone, the antiprogestins per se induced marked inhibitory and stimulatory effects on GnRH-stimulated LH secretion. In brief, short-term treatment of non-oestrogen-primed cells with antiprogestins was ineffective (ZK compounds) or reduced LH secretion (RU486), while long-term treatment was stimulatory. Oestrogen-primed cells exerted exclusively inhibitory effects on GnRH-induced LH secretion. In conclusion, antiprogestins are effective antagonists of progesterone actions in the gonadotroph. However, they exert diverse actions on gonadotrophin secretion in the absence of progesterone, which might interfere with their antagonistic properties.
Animals↗
Expression of luteinizing hormone releasing hormone and its mRNA in human endometrial cancer cell lines.
Recent results have shown that specific binding sites for luteinizing hormone-releasing hormone (LHRH) are present in biopsy samples of human endometrial cancer and in the human endometrial cancer cell lines Ishikawa and HEC-1A. The proliferation of these cell lines was retarded by LHRH analogs. The present study was undertaken to determine whether these endometrial tumor cells also produce LHRH or an LHRH-like peptide which could serve as natural ligand for the LHRH binding sites. Using a specific antibody, LHRH-immunoreactivity was detected in extracts of Ishikawa (426 +/- 84 fmol/10(6) cells) and HEC-1A (368 +/- 41 fmol/10(6) cells) cells. LHRH-like bioactivity of these samples was assessed in a rat pituitary cell culture system. The release of luteinizing hormone induced by endometrial cancer cell extracts corresponded to that obtained with comparable amounts of authentic LHRH. The expression of the mRNA for LHRH could be demonstrated by reverse transcriptase - polymerase chain reaction using specific primers according to the published sequence and by subsequent Southern blot analysis. The presence of immuno- and bioactive LHRH-like factors and the demonstration of expression of the mRNA for LHRH in two human endometrial cancer cell lines supports the concept of an autocrine regulatory system based on LHRH in endometrial cancer.
Base Sequence↗
High affinity binding and direct antiproliferative effects of LHRH analogues in human ovarian cancer cell lines.
Recently, specific binding sites for luteinizing hormone releasing hormone (LHRH) and its analogues have been demonstrated in biopsy samples of human epithelial ovarian cancer. Their biological significance remained obscure. In this study we ascertained whether such LHRH-binding sites are also present in the human epithelial ovarian cancer cell lines EFO-21 and EFO-27 and if they could mediate antiproliferative effects of LHRH analogues. Using [125I, D-Trp6]LHRH, a high affinity/low capacity binding site was detected in both lines: EFO-21 (Kd1 = 1.5 x 10(-9) M; binding capacity (Bmax1) = 4.9 fmol/10(6) cells) and EFO-27 (Kd1 = 1.7 x 10(-9) M; Bmax1 = 3 fmol/10(6) cells). In addition, a second class of low affinity/high capacity binding sites (EFO-21: Kd2 = 7.5 x 10(-6) M; Bmax2 = 24 pmol/10(6) cells; EFO-27: Kd2 = 4.3 x 10(-6) M; Bmax2 = 14.5 pmol/10(6) cells) was demonstrated. Specific binding of [125I, D-Trp6]LHRH was displaced with nearly equal efficiency by unlabeled [D-Trp6]LHRH, the LHRH-antagonists SB-75 and Hoe-013, and by native LHRH but not by unrelated peptides such as oxytocin and somatostatin. In the presence of 10(-5) M agonist [D-Trp6]LHRH, the proliferation of both cell lines was significantly reduced to 77% of controls after 24 h and to approx. 60% after 6 days. Lower concentrations (10(-9) M) of the agonist, significantly decreased the proliferation to 87.5% for EFO-21 and 86% for EFO-27 after 6 days. These antiproliferative effects were enhanced by increasing doses of [D-Trp6]LHRH and were maximal at 10(-5) M (EFO-21: 65.5% of control, EFO-27: 68% of control). Similar dose-dependent antiproliferative effects were obtained in EFO-21 line with the LHRH-antagonists SB-75 and Hoe-013, while these analogues had no effects on the proliferation of EFO-27 cells. SB-75 partly antagonized the antiproliferative effect of [D-Trp6]LHRH in a dose dependent way in the EFO-27 line. These data suggest that LHRH analogues can directly inhibit the in vitro proliferation of human ovarian cancer cells. This effect might be mediated through the high affinity LHRH binding sites.
Cell Division↗
Medroxyprogesterone acetate inhibits the proliferation of estrogen- and progesterone-receptor negative MFM-223 human mammary cancer cells via the androgen receptor.
This study demonstrates for the first time, that medroxyprogesterone acetate (MPA) inhibits the proliferation of the estrogen and progesterone receptor negative mammary cancer cell line MFM-223 via the androgen receptor. MPA is a progestin, which is used in the hormonal treatment of disseminated breast cancer. It binds to the progesterone, androgen, and glucocorticoid receptor and may exert its antiproliferative effects via different receptors. MFM-223 human mammary cancer cells contain a very high level of androgen receptors (160 fmol/mg protein) and low levels of estrogen, progesterone, and glucocorticoid receptors (< 20 fmol/mg protein). This cell line provides therefore a good model system to analyze the possible role of the androgen receptor in the action of MPA avoiding interference with other steroid hormone receptors. Effective inhibition of proliferation is achieved by 10 nM MPA or 1 nM of the androgen dihydrotestosterone, corresponding well to the binding affinities of both compounds (3.6 and 0.18 nM, respectively). The involvement of the androgen receptor was confirmed by competition experiments with antiandrogens. Furthermore, MFM-DHT cells, which are an androgen resistant subline of MFM-223 cells, are also resistant to MPA. This data supports the involvement of the androgen receptor in the action of MPA and additionally rules out direct hormone-independent cytotoxic effects of MPA.
Androgen Receptor Antagonists↗