Insulin-like growth factor I expression by tumors of neuroectodermal origin with the t(11;22) chromosomal translocation. A potential autocrine growth factor.

Expression of insulin-like growth factor I (IGF-I) mRNA by some tumor cell lines of neuroectodermal origin has been described. To further explore the significance of IGF-I mRNA expression in these tumors, a more extensive analysis was performed. Most (9 of 10) neuroectodermal tumor cell lines with a t(11;22) translocation (primitive neuroectodermal tumor [PNET], Ewing's sarcoma, esthesioneuroblastoma) expressed IGF-I mRNA, whereas 0 of 15 cell lines without the translocation (PNET, neuroblastoma) expressed IGF-I. Furthermore, inasmuch as all neuroblastoma (12 of 12) cell lines examined expressed IGF-II RNA, the pattern of IGF expression could distinguish between these closely related tumors. CHP-100, a PNET cell line with the t(11;22) translocation, was shown to secrete both IGF-I protein and an IGF binding protein, IGFBP-2. This cell line also expressed the type I IGF receptor mRNA, and blockade of this receptor by a monoclonal antibody (alpha IR3) inhibited serum-free growth. These data demonstrate that IGF-I expression is a property of neuroectodermal tumors with a t(11;22) translocation and that interruption of an IGF-I autocrine loop inhibits the growth of these tumor cells.

Oestrogen/growth factor cross-talk in breast carcinoma: a specific target for novel antioestrogens.

Breast cancer cells express receptors for and are sensitive to a variety of steroids, polypeptide hormones and growth factors; the blocking of and/or the interference with their biochemical pathways could represent a new approach to breast tumour therapy. Antioestrogens achieve such a goal by competing with oestradiol for binding to the oestrogen receptors through which intracellular effects of the hormone are mediated. Tamoxifen has undergone the most extensive clinical evaluations and represents the treatment of choice for the endocrine management of breast cancer. Nevertheless, it is well known that tamoxifen retains agonist activity both in vitro and in vivo. To circumvent this disadvantage, new molecules with steroid-like structure, represented by ICI 164,384 and ICI 182,780, have been synthesized. In this review, Alessandra de Cupis and Roberto Favoni review data about the cross-talk between the two major families of breast cancer growth regulator: oestrogens and growth factors, focusing on the use of nonsteroidal antioestrogens and the new generation of steroidal antioestrogens as possible specifically targeted inhibitors of breast tumour proliferation.

Steroidal and nonsteroidal oestrogen antagonists in breast cancer: basic and clinical appraisal.

Endogenous sex hormones, such as oestrogens, control the development and growth of breast carcinoma. Blocking the binding between the steroid hormone and its receptor using anti-oestrogenic compounds is one of the possible ways of impeding the action of oestrogens at the level of their target tissues. The nonsteroidal, triphenylethylenic, anti-oestrogen tamoxifen has so far been the standard compound for endocrine treatment of breast cancer. Nevertheless, it forms a complex with the oestrogen receptor which gives rise to a range of biopharmacological events, ranging from full oestrogen antagonism to partial agonism. Moreover, its use is limited by the possible onset of drug resistance in many patients. In spite of these restrictions, tamoxifen has proved to be very helpful as a starting point for the development of new, more efficacious compounds. In this article, Roberto Favoni and Alessandra de Cupis review and discuss tamoxifen, several of its derivatives and the steroidal-like anti-oestrogens in terms of laboratory development, pharmacology and clinical evaluation.

Analysis of insulin-like growth factor I gene expression in malignancy: evidence for a paracrine role in human breast cancer.

Insulin-like growth factor I (IGF-I) activity has been reported to be produced by several human cancers. Identification of RNAs transcribed from the IGF-I gene has been complicated by the detection of multiple hybridizing bands on Northern analysis. To determine if any of these RNAs are transcribed from the IGF-I gene, we have used a sensitive and specific ribonuclease (RNAse) protection assay for IGF-I. We have also studied the breast cancer tissue expression of IGF-I using in situ hybridization histochemistry. We have found no IGF-I mRNA in breast (zero of 11) or colon cancer (zero of 9) cell lines; both of these tumors have been previously reported to express IGF-I mRNA. However, three of three neuroepithelioma and one of two Ewing's sarcoma cell lines express IGF-I mRNA; therefore, in these tumors IGF-I may be an autocrine growth factor. In contrast to breast cancer cell lines, RNA extracted from breast tissues has easily detectable IGF-I mRNA. In situ hybridizations show that IGF-I mRNA is expressed in the stromal cells, and not by normal or malignant epithelial cells. These findings suggest that although IGF-I is not produced by breast epithelial cells it may function as either a paracrine stimulator of epithelial cells or an autocrine stimulator of stromal cells.

Suramin-induced growth inhibition and insulin-like growth factor-I binding blockade in human breast carcinoma cell lines: potentially related events.

Suramin, a polyanionic drug used in the treatment of trypanosomiasis and onchocerciasis, inhibits growth factor-induced mitogenesis in several human tumours. We have investigated the effect of suramin on human breast cancer cell lines (HBCCL). By cell counts and thymidine incorporation we found that 50 to 400 micrograms/ml suramin inhibits the proliferation of HBCCL in a dose-dependent and reversible fashion (ID50 approximately 200 micrograms/ml for MCF-7 and MDA-MB 231). Radioreceptor and affinity cross-linking assays showed that suramin was also able to reduce the binding of insulin-like growth factor I (IGF-I) to its receptor (40-50% inhibition at 100 micrograms/ml). Our results indicate that the drug does not affect the IGF-I receptor (IGF-I-R), but binds directly to the IGF-I peptide. In conclusion, the strict correlation observed between suramin inhibition of proliferation and IGF-I binding on HBCCL suggests a possible therapeutic role for this molecule as an antineoplastic drug in human breast tumours.

Anti-insulin-like growth factor-I activity of a novel polysulphonated distamycin A derivative in human lung cancer cell lines.

1. The purpose of this study was to investigate the antiproliferative effect and the modulation of the mitogenic insulin-like growth factor-I (IGF-I) system by FCE 26644 and FCE 27784, two polyanionic sulphonated distamycin A derivative compounds, on two human non-small cell lung cancer (N-SCLC) cell lines. 2. For cell growth studies the colorimetric MTT and the thymidine incorporation assays were performed; the presence of IGF-I and IGF-binding proteins in conditioned media was revealed by radioimmunoassay and Western ligand blot, respectively. Variations at the IGF-I-receptor level were tested by binding studies on cell monolayers. 3. A significant concentration- and time-dependent cytostatic activity of FCE 26644 (IC50 approximately 200 micrograms ml-1 at 72 h) compared to its analogue FCE 27784 (IC50 > 800 micrograms ml-1) was observed in both cell lines studied. The IGF-I-stimulated proliferation of the IGF-I-responsive A549 cell line was abolished by 24 h of FCE 26644 treatment whereas FCE 27784 was inactive. FCE 26644 increased (4 to 6 fold) the secretion of IGF-I-like material and reduced the IGF-I binding (IC50 > 100 micrograms ml-1) in both A549 and Ca-Lu-1 cell lines. FCE 26644 (100 micrograms ml-1) did not affect the KD (approximately 0.5 nM) but reduced the Bmax and the number of receptor sites (50%). 4. Our findings demonstrate that the ability to down-regulate the cell proliferation of N-SCLC cell lines, shown by FCE 26644, depends at least partially, on interference with the "IGF-I mitogenic system'.

Comparison between novel steroid-like and conventional nonsteroidal antioestrogens in inhibiting oestradiol- and IGF-I-induced proliferation of human breast cancer-derived cells.

1. This study has two specific aims: (a) to compare the antioestrogenic activity of two steroidal analogues of 17 beta-oestradiol, the 7 alpha-alkylamide, ICI 164,384 and the 7 alpha-alkylsulphinylamide, ICI 182,780, with that of the triphenylethylene-derived compound 4OH-tamoxifen on a pool of human breast cancer cell lines (HBCCL) with a range of hormonal responsiveness and acquired anti-oestrogen resistance and (b) to investigate the ability of such antioestrogens to modulate the potent breast carcinoma growth-stimulatory activity of the 'IGF-I system'. 2. For the chemosensitivity investigations we used a long-term colorimetric and the short-term thymidine incorporation assay; we analysed IGF-I in conditioned media by a radioimmunoassay, IGF-I mRNA in the cells by RT-PCR and molecular species of IGF-I-binding proteins, secreted in conditioned media, by Western ligand blot. IGF-I receptors were assayed on cell monolayers by binding studies and by Scatchard analysis, we calculated KD, Bmax and sites/cell. 3. Our results indicate that ICI 182,780 and ICI 164,384 are 1.5-5.5 fold more potent than 4OH-tamoxifen in inhibiting the basal proliferation of oestrogen-receptor positive (ER+) breast cancer cell lines. Moreover we demonstrate the capacity of ICI 182,780 and ICI 164,384 to reduce, in a time-dependent fashion, oestrogen- and/or IGF-I-stimulated growth of ER+cell lines, possibly by negatively interfering with an IGF-I-like material secretion and IGF-I-receptor number. 4. Our data provide the first evidence that, on ER+human breast carcinoma cell lines, steroidal antioestrogens inhibit cell growth and modulate the IGF-I mitogenic system. The mechanism of this latter effect has yet to be identified.

Pre-clinical evaluation of new antineoplastic agents in NSCLC cell lines: evidence of histological subtype-dependent cytotoxicity.

The antiproliferative effect of paclitaxel, docetaxel, gemcitabine, topotecan, SN-38 and cis-platin was studied on 5 non-small cell lung cancer (NSCLC) cell lines, 3 of which were adenocarcinoma (ADK) and 2 squamous cell carcinoma (SCC). Cellular chemosensitivity was determined using the MTT in vitro assay after 48, 72 and 96 h of exposure to drug in concentration ranging from 0.001 to 100 microM. A concentration-dependent cell growth inhibition was observed for paclitaxel, gemcitabine, topotecan, SN-38 and cis-platin in all cell lines tested. Docetaxel showed a concentration-independent cytotoxicity and was 104 times more potent than cis-platin (IC50 = 0. 001 vs. 10 microM). Paclitaxel, gemcitabine, topotecan and SN-38 were 102 times more potent than cis-platin, with median IC50 = 0.1 microM at 72 h. The level of drug-induced cell growth inhibition appeared to be correlated, for some of the six drugs tested, with the tumor histological subtype. In particular, topotecan and cis-platin were more active in squamous cell carcinoma than in adenocarcinoma cell lines (p=0.006 and 0.001 respectively at 0.1 microM concentration), while paclitaxel was more active in ADK than in SCC cell lines (p=0.004 at 0.01 microM concentration). Ca-Lu-6, a cell line that, contrary to most other lung cancer cell lines, is wild-type for most oncogenes/tumor suppressor genes, was by far the most sensitive cell line used (p=0.002, 0.003, 0.01 for paclitaxel, topotecan and cis-platin respectively, at 1 microM concentration), showing a >50% growth inhibition to new drugs at a concentration of 0.01 microM. In conclusion, all these new compounds tested were found to be more potent than cis-platin in affecting cellular proliferation of six NSCLC cell lines studied. We suggest that the specific histological subtype and molecular pattern of the cell line being treated could affect the antiproliferative effect of these drugs.

Interaction between novel anticancer agents and radiation in non-small cell lung cancer cell lines.

Integration of chemotherapy and radiation is the standard practice in the management of locally advanced inoperable NSCLC. To assess the biological interaction between third generation chemotherapeutic agents and radiation in non-small cell lung cancer (NSCLC) in vitro, we tested a number of different drugs (paclitaxel, docetaxel, gemcitabine, topotecan, SN-38 and cisplatin) combined with radiation, in lung cancer cell lines. Cellular chemosensitivity was determined, using the semi-automated colorimetric MTT assay, after 48, 72 and 96 h of exposure to increasing drug concentrations, (0.001-100 microM) and radiation doses (100-400 cGy). Cell lines used were the adenocarcinoma (ADK), A-549, and the squamous-cell carcinoma (SCC), LX-1. Cells were pre-treated with anticancer agents at 24, 12 and 0 h before irradiation. Cytofluorimetric cell cycle analysis was performed. A significant S-phase block or a G(2)/M block was seen with gemcitabine and topotecan or paclitaxel pre-treatment, respectively. Apoptosis was seen only after paclitaxel exposure in the A-549 cell line. Despite a similar pattern of cell-kinetic changes induced by chemotherapy pre-treatment in all cell lines, the adenocarcinoma A-549 cell line was not radiosensitized by any of the anticancer agents tested, whereas synergism was observed in the LX-1 squamous carcinoma cell line, when exposed to gemcitabine, SN-38, topotecan and cisplatin. Paclitaxel, despite a favourable cell cycle effect, was not found to be synergistic with radiotherapy in our experimental model. In conclusion, the observed synergism appears to be dose- and timing-independent and seems to be related to the histological subtype being present in SCC only. Favourable perturbation of the cell cycle is evident with all the new agents tested in both cell types, but was not sufficient to produce synergism with radiation.

Insulin-like growth factor-I (IGF-I) and IGF-binding proteins blood serum levels in women with early- and late-stage breast cancer: mutual relationship and possible correlations with patients' hormonal status.

The pathogenesis and progression of breast cancer involve complex interactions between hormones and polypeptide growth factors such as insulin-like growth factor-I (IGF-I). IGF-I has been found in stromal fibroblasts derived from malignant and benign breast tissue and it is a mitogen for several breast cancer cell lines. It circulates bound to specific high-affinity binding proteins, which could act as either positive or negative modulators of tumorigenesis. This study has been addressed to characterize IGF-I and its binding proteins in the serum of 85 unselected patients with early breast cancer. The IGF-I concentration was assessed by radioimmunoassay of 69 out of 85 samples before and after dissociation of the IGF-I and IGF-binding protein (IGF-BP) complex whereas IGF-BP of all 85 sera were analyzed by Western ligand blotting; estradiol and progesterone were measured by radioimmunoassay in native serum samples. In our study no differences in IGF-I serum levels between pre- and post-menopausal patients were observed. Patients with higher estradiol and progesterone serum levels did not present different IGF-I concentrations compared to patients with lower serum levels. Furthermore, IGF-I median values were not found to depend on estrogen receptor (ER) status. A heterogeneous quali-quantitative molecular pattern of binding proteins was detected: IGF-BP3 and IGF-BP1 were the most and the least expressed respectively. No correlations between ER status, or parameters related to the hormonal status, and IGF-I or binding proteins expression were observed. No significant differences in IGF-I concentration and IGF-BP expression were observed between cancer patients and a control group matched for age and menopausal status. Finally, preliminary collection of 20 sera derived from patients with late breast cancer was analyzed for IGF-I and its binding proteins content.

Suramin interferes with auto/paracrine insulin-like growth factor I-controlled proliferative loop on human lung cancer cell lines.

Human non-small cell lung cancer (N-SCLC), a common malignancy generally unmanageable by conventional cytotoxic chemotherapy, represents a major world health burden. Suramin, a polyanionic drug which appears to interfere with growth-factor/receptor interaction, has recently been shown to be cytostatic for small cell lung cancer cells; it may also be effective for N-SCLC. As insulin-like growth factor I (IGF-I) is a known progression agent for N-SCLC, we have examined the effects of suramin on the 'IGF-I system' in a panel of human N-SCLC cell lines. Colorimetric and thymidine incorporation assays were used to assess cell chemosensitivity whereas a radio-receptor assay was employed to evaluate IGF-I/receptor binding. Suramin reversibly reduced, in a concentration- and time-dependent manner, the growth of each N-SCLC cell line examined either cultured in serum-containing or serum-free medium. Furthermore, suramin caused a concentration-related inhibition of labeled IGF-I peptide specific binding on all cell lines studied. Suramin caused a significant reduction in the Bmax values with only weak variations in the affinity constants (Kd). We hypothesize that suramin interference with IGF-I mitogenic activity is a pathway by which this drug produces its effect in vitro. These data indicate further studies on the mechanism of action and pharmacology of suramin in vivo are warranted.

The insulin-like growth factors, their receptors, and their binding proteins in human breast cancer.

Various investigators have shown that the IGFs are mitogens for breast cancer cells. The expression of the IGF receptors is seen in most breast cancer cell lines and tissues, suggesting that most breast cancers have the ability to respond to the IGFs. Although authentic IGF-I is not expressed by breast cancer cell lines, it is possible that an IGF-related peptide that can be detected immunologically is expressed. Furthermore, in estrogen responsive xenotransplants, changes in the level of IGF-II mRNA correlate directly with estrogen-mediated changes in tumor growth. These observations suggest that IGF-II may be important in tumorigenesis and may serve as an autocrine growth stimulator of breast cancer cells. When human breast cancer tissues are studied, IGF-I and IGF-II mRNA expression are commonly seen. However, in situ hybridization studies suggest that IGF-I mRNA is expressed mainly by the stromal elements, while IGF-II mRNA can be found both in stroma and malignant epithelial cells. These observations support the studies done with breast cancer cell lines; IGF-I may stimulate cells via a paracrine pathway, while IGF-II may act as both an autocrine and paracrine growth factor. In addition, IGF-BPs are commonly expressed by breast cancer cells in culture, and it is possible that expression of the IGF-BPs act to modulate the effects of either IGF-I or IGF-II. We propose that the IGFs are important stimulators of breast cancer cells and that their growth promoting effects may be mediated by autocrine, paracrine, or endocrine mechanisms. Furthermore, interactions between the stroma and malignant epithelial cells may be important in regulating the growth of breast cancer. The biological importance of a fibroblast-epithelial cell interaction has been demonstrated in a normal mouse mammary cell line; morphological and functional changes in epithelial cells were induced when the cells were in direct contact with fibroblasts. Similar mechanisms may be important in malignant breast epithelial cells. For example, many breast cancer cells produce platelet-derived growth factor (PDGF) yet have no PDGF receptor. PDGF has been demonstrated to increase IGF-I production by fibroblasts, and a dual paracrine pathway involving PDGF and IGF-I expression by epithelial cells and stromal cells could be envisioned. The pathways through which the IGF system may function in human breast cancer are schematically represented in figure 1. Further work in our laboratory is directed at clarifying the role for the IGFs in breast cancer growth.

Manipulation of the growth rate of human breast cancer cells by antiandrogen followed by chemotherapy.

The effects induced by the antiandrogen Cyproterone Acetate (CPA) on the proliferation of EVSA-T human breast cancer cells endowed with androgen receptors were studied. Kinetic analyses were carried out by two autoradiographic techniques measuring the percentage of cells in S-phase and the growth fraction (GF). The exposure of the cultures to CPA for 24 h caused a marked inhibition on S-phase cells without significantly affecting the CF. The accumulation of cells in G1-phase, confirmed by cytometric analysis, was rescued to the S compartment by replacing the culture medium 24 h after CPA administration. Exposure of EVSA-T to Doxorubicin or Methotrexate after CPA and medium change at the time of maximal proliferative recruitment, led to an enhancement of cytotoxicity as demonstrated by colony survival assay.

Doxorubicin cytotoxicity to P388 lymphocytic leukemia as determined by alkaline elution and established assays.

Very large variations exist in the response of individual tumors to antineoplastic agents, even when the tumors are apparently very similar from the point of view of stage and histological classification. It has been recognized for a long time that methods capable of revealing the specific chemosensitivity of individual tumors could be useful for an individual optimization of a chemotherapeutic protocol. The Tumor Colony Forming Assay (TCFA) and the Biochemical Antimetabolic Assay (BAA) have been proposed for this purpose. Their main limitation is a consequence of the fact that the capability of in vitro growth is required from cells of a tumor grown in vivo. This is often lacking or very poor in the first in vitro passages. In this work we have investigated the possibility of using a sensitive method for evaluating DNA damage, the Alkaline Elution technique (AE). Cells treated in vivo can be easily tested directly for DNA damage. No cell proliferation in vitro is required. It is not required that the measured effect is the specific cause of cell death. A P388 Doxorubicin sensitive line and a resistant subline were tested. Correct correlations between DNA damage and chemosensitivity were obtained working both in vivo and in vitro. This test could be useful for assessing the chemosensitivity in vivo of alkylating and intercalating agents.

Responsiveness to hormone, growth factor and drug treatment of a human breast cancer cell line: comparison between early and late cultures.

Growth rate, morphology, and responsiveness to mitogenic stimuli and pharmacological treatments were evaluated in early and late cell passages derived from the same clone of the widely used MCF-7 human breast adenocarcinoma cell line. Our results indicate dissimilarities between early (E) and late (L) passages for some of the parameters analyzed. The cells that underwent many subcultivations grew faster than the others; both appeared homogeneous in size and shape. The E cells, subcultured for almost 1 yr, displayed higher sensitivity to the mitogenic action of both estradiol, according to the level of estrogen receptor, and insulin-like growth factor-I than did the L cells, kept in culture for more than 10 yr. Cell responsiveness to two drugs, a novel steroid antiestrogen and a polysulfonated distamycin A derivative, was more pronounced in the early cultures only at the longer time of exposure to the higher concentration of the estrogen antagonist. In addition, a drug-induced inhibition of insulin-like growth factor-I binding to its receptor was shown in both E and L cells, the latter being less sensitive than the former when exposed to the antiestrogen. Finally, MCF-7 E and L cells showed similar behavior when drug-induced apoptosis was tested.

Establishment and preliminary characterization of human malignant mesothelioma cell lines.

Malignant mesothelioma is an aggressive and rare tumor of the mesothelium which arises in serosal cavities and is strongly related to asbestos exposure. The incidence of this tumor is still increasing because of the widespread use of asbestos over the past years and the long latency of this malignancy. The paucity of well-characterized in vitro human models and the consequent shortage of experimental pharmacological studies on mesothelioma reduce the success of its clinical management. It is well known that established human cell lines represent a rapid and advantageous system for in vitro studies of several diseases, such as mesothelioma. Thus, the aim of our study was to establish and characterize a panel of selected human malignant mesothelioma cell lines in order to highlight the biology of this tumor and to test the effectiveness of new antiproliferative compounds. During the last year, we collected 10 pleural effusion samples from patients with diagnosed and pharmacologically untreated mesothelioma. Among these, we were able to isolate three continuously growing cell lines, identified as IST-Mes1, IST-Mes2 and IST-Mes3. Another previously established malignant mesothelioma cell line (MPP89) has also been included in this study. The cells in culture appeared morphologically heterogeneous: three of them (IST-Mes1, IST-Mes2, and MPP89) were spindle-shaped, and upon reaching confluence, they assumed the characteristic cobblestone-like pattern, whereas IST-Mes3 showed mixed sizes and shapes. Cell growth studies revealed that all cell lines reach exponential growth phase within 4-7 days after plating with a doubling time ranging from 37-87 hours. Finally, insulin-like growth factor-I did not stimulate cellular proliferation of both the IST-Mes1 and IST-Mes2 cell lines.

The role of polypeptide growth factors in human carcinomas: new targets for a novel pharmacological approach.

The processes of cellular proliferation and progressive acquisition of a specialized phenotype show a high degree of coordination. In particular, these complex signaling networks mediating cell growth, differentiation, migration, and apoptosis are regulated in part by polypeptide growth factors that can act, by autocrine and/or paracrine mechanisms of action, as positive or negative modulators. Because these growth factors are unable to cross the hydrophobic cell membrane, they exert their effects via binding to cell surface receptors, most of which possess intrinsic tyrosine kinase activity. Owing to the interaction of polypeptide growth factors with their specific transmembrane receptors, a cascade of intracellular biochemical signals, resulting in the activation and repression of various subsets of genes, is triggered. One of the major incentives for studying factors that regulate processes of proliferation and differentiation is the recognition of their involvement in tumorigenesis. Genetic aberrations in growth factors signaling pathways are, in fact, inextricably linked to cancer. Malignant cells arise as a result of a stepwise progression of genetic events characterized by the unregulated expression of growth factors or components of their signaling networks. The main aim of this review is to examine the current understanding of the crucial contribution that several growth factors may have on transformation, tumorigenesis, and progression in several human tumors among the most widespread in western countries. For this purpose, we will analyze the chemistry and the molecular organization of the most important growth factors and their specific receptors. In addition, we will focus on the mechanisms of signal transduction, the complex cascade of biochemical events ensued from the growth factor/receptor binding. The present knowledge of the role of growth factor biochemical signaling networks in cancer leads to improvements not only in diagnosis and prognosis for this disease, but also for new and more targeted therapeutic intervention. The second part of this review will focus on the novel pharmacological approaches for cancer therapy that have been developed already or are being developed with the aim to specifically interfere at various steps of the growth factors signaling pathways.

New vs old fashioned oestradiol antagonists in mammary carcinoma: 'in vitro' and 'in vivo' pharmacological approaches.

The rationale underlying therapeutic strategies designed to inhibit the action of endogenous sex hormones in malignant breast cells is provided by the demonstration of their involvement in supporting the development and growth of breast carcinoma. The surgical removal of steroid-secreting glands, in order to reduce the level of oestrogens reaching their target tissues, has for years been substituted by the so-called endocrinotherapeutic approach, which is based on the counteraction of the steroid hormone activity by the hormonal receptor blockade with suitable antioestrogenic compounds. Over the past 25 years, the non-steroidal oestrogen antagonist tamoxifen has become the standard endocrine treatment for breast cancer. The triphenylethylene-derivative compound competes efficiently for binding to the oestrogen receptor, but the complex retains some transcriptional activity. Consequently, tamoxifen exhibits, both ' in vitro and in vivo ', a range of biological activity from full oestrogen antagonism to partial agonism. There is also evidence suggesting that the agonist activity of this compound may ultimately stimulate breast tumour growth, thus causing some treatment failures. Moreover, the use of tamoxifen is limited by the possible onset of drug-resistance in many patients. Nevertheless, widely tested tamoxifen has proved to be very helpful for the development of new compounds to be used as long-term adjuvant therapy or as preventive agents. These novel oestrogen antagonists belong to two major classes: tamoxifen analogs and new pure steroidal-like antioestrogens. The search for and development of compounds devoid of tamoxifen cross-resistance, with a safer toxicity profile as well as the lack of oestrogenic effects, provide the bases to improve the current therapeutic applications of antioestrogens.

Identification of insulin-like growth factor binding proteins in breast cancer cells.

The insulin-like growth factors (IGFs) are potent mitogens for some breast cancer cell lines. Recent evidence suggests that IGF-induced mitogenesis may be influenced by specific IGF binding proteins (IGFBPs). In this study, breast cancer cell lines were examined for IGFBP protein and mRNA expression. Western ligand blot examination of conditioned media from breast cancer cell lines suggested that the IGFBP protein expression was heterogeneous. Although all breast cancer cell lines expressed a 24 kDa binding protein, MCF-7, an estrogen receptor positive (ER+) cell line, expressed a IGFBP compatible with reported sizes for IGFBP-2. Estrogen receptor negative (ER-) cells (MDA-MD-231, Hs578T) secreted IGFBPs consistent with sizes reported for IGFBP-1 and -3. Examination of mRNA expression supported these findings; IGFBP-2 was seen in all (4/4) ER+ cell lines while high levels of IGFBP-3 were found in ER- cell lines (3/5), although lower levels of IGFBP-3 mRNA could be found in some ER+ cell lines. In MCF-7 cells, steady state levels of IGFBP-3 mRNA were decreased by estradiol, while IGFBP-2 mRNA levels were slightly increased. These data suggest that IGFBP expression by breast cancer cells is heterogeneous, that the pattern of IGFBP expression is different between ER+ and ER- cell lines, and that in ER+ cells IGFBP mRNA may be regulated by estrogens. Thus, the IGFBPs may play an important role in mediating the mitogenic response of breast cancer cells to the IGFs.

Proliferative activity of human solid tumors evaluated by thymidine labeling index and primer-dependent alpha-DNA polymerase.

Proliferating cells, independent of their position in the cell cycle, contain a primer-dependent alpha-DNA polymerase (PDP); we have, therefore, tested the feasibility of the PDP assay in breast, ovarian, and head and neck tumors. Experimental results demonstrate that the PDP labeling index (PDP-LI) is constantly superior to the thymidine labeling index (TLI), which indicates the percentage of S phase cells. The PDP-LI/TLI ratios observed were 5.0 in breast cancer, 3.6 in ovarian cancer, and 2.5 in head and neck tumors. Higher PDP-LI scores have been obtained in metastatic breast tumor samples (PDP-LI = 8.8) compared to nonmetastatic tumors (PDP-LI = 4.9).