Marc Lacroix

Relevance of breast cancer cell lines as models for breast tumours: an update

The number of available breast cancer cell (BCC) lines is small, and only a very few of them have been extensively studied. Whether they are representative of the tumours from which they originated remains a matter of debate. Whether their diversity mirrors the well-known inter-tumoural heterogeneity is another essential question. While numerous similarities have long been found between cell lines and tumours, recent technical advances, including the use of micro-arrays and comparative genetic analysis, have brought new data to the discussion. This paper presents most of the BCC lines that have been described in some detail to date. It evaluates the accuracy of the few of them widely used (MCF-7, T-47D, BT-474, SK-BR-3, MDA-MB-231, Hs578T) as tumour models. It is concluded that BCC lines are likely to reflect, to a large extent, the features of cancer cells in vivo. The importance of oestrogen receptor-alpha (gene ESR1) and Her-2/neu (ERBB2) as classifiers for cell lines and tumours is underlined. The recourse to a larger set of cell lines is suggested since the exact origin of some of the widely used lines remains ambiguous. Investigations on additional specific lines are expected to improve our knowledge of BCC and of the dialogue that these maintain with their surrounding normal cells in vivo.

Significance, detection and markers of disseminated breast cancer cells

The development of distant metastases is the major cause of death from breast cancer. In order to predict and prevent tumour spreading, many attempts are being made to detect small numbers of tumour cells that have shed from the primary lesions and have moved to lymph nodes, blood or bone marrow. This article presents the advantages and the limitations of techniques used for disseminated tumour cells (DTC) detection. DTC markers are listed and the most currently used of them (KRT19, CEACAM5, TACSTD1, MUC1, EGFR, ERBB2, SCGB2A2, SCGB2A1, SCGB1D2, PIP, SBEM, TFF1, TFF3, ANKRD30A, SPDEF, ESR1, SERPINB5 and GABRP) are discussed, notably on the basis of recent data on breast tumour portraits (luminal epithelial-like, basal/myoepithelial-like and ERBB2). The significance of DTC for the prognosis and prediction of response to therapy is examined. DTC viability, the notion of cell dormancy and the concept of breast cancer stem cells are also discussed.

Persistent use of false cell lines

From HeLa and its multiple identities, to MDA‐MB‐435, erroneously and widely used as breast cancer cells, the history of cancer cell lines is rich in misidentification and cross‐contamination events. Despite the fact that these problems were regularly signaled during the last decades, many actors of research still seem to ignore them. A never‐ending story? Solutions exist, notably based on recent technical advances in cell line authentication (short tandem repeat analysis). However, a collaborative action involving users of cell lines, cell banks, journals and funding agencies is needed to achieve success.

About GATA3, HNF3A, and XBP1, three genes co-expressed with the oestrogen receptor-α gene (ESR1) in breast cancer

In breast tumours and breast cancer cell (BCC) lines, microarray analyses have revealed that a series of genes are expressed in close association with the oestrogen receptor-alpha (ER-alpha) gene, ESR1. Three of them, GATA3, HNF3A (also known as FOXA1), and XBP1 encode transcription factors. Here, we present these factors and we discuss their potential involvement in the ER-alpha-mediated actions in BCC. We notably show the relations that exist, or that might exist, between these factors and the oestrogen-inducible trefoil factor TFF1.

Estrogenic effect of a series of bisphenol analogues on gene and protein expression in MCF-7 breast cancer cells.

Bisphenols constitute a family of compounds, which includes many substances that have as a common chemical structure two phenolic rings joined together through a bridging carbon. In the present study, we aimed to determine whether several events triggered by 17 beta-estradiol (E(2)) in MCF-7 breast cancer cells were also observed in response to various bisphenol-A (BPA) analogues. We studied the expression of estrogen controlled genes by measuring the induction of pS2 (mRNA and protein) and progesterone receptor (PgR) as well as the expression of a luciferase reporter gene transfected into MVLN cells. These data were compared to the cell proliferation potency and effectiveness as the latest expression of estrogen controlled functions. Bisphenols showed an agonistic effect in all our assays, suggesting that these compounds may act through all the response pathways triggered by the natural hormone. We found differences between the assays in the potency of bisphenols, defined as the minimum concentration required to produce a maximal effect. In the cell proliferation assay, all tested compounds needed a lower concentration than in the other assays to give maximal response. Our results suggest that the polarity and nature of the substituent in the central carbon determines the estrogenic potency. Presence of two propyl chains at the central carbon appears to confer the greatest potency in both gene and protein expression assays.

Estrogen Receptor Alpha: Impact of Ligands on Intracellular Shuttling and Turnover Rate in Breast Cancer Cells

Estrogen receptors (alpha and beta) are members of the steroid/thyroid nuclear receptors superfamily of ligand-dependent transcription factors. Impact of the alpha isoform of estrogen receptor (ER) on breast cancer etiology and progression is now well established. Current therapeutic strategy to treat ER-positive breast cancer relies on the blockade of ER trancriptional activity by antiestrogens. Data accumulated during the last five years on the mechanism of action of ER enable one to foresee new strategies. These data indeed reveal that ER is not statically bound to DNA at promoter sites of genes regulating cell proliferation and/or differentiation, but rather behaves as a very mobile protein continuously shuttling between targets located within various cellular compartments (i.e. membrane, microsomes, nucleus...). This allows the receptor to generate both non-genomic and genomic responses. Ligands, growth factors and second messengers produced downstream of activated membrane receptors modulate ER-mediated responses by interfering with the traffic patterns of the receptor, as well as by locally blocking its transient anchorage. Changes in ER turnover rate associated with these regulatory processes seem also to strongly influence the ability of the receptor to mediate gene transactivation. The present paper surveys these biological data and analyzes how they may be integrated into new drug design programs aimed at expanding our therapeutic armamentarium against breast cancer.

Production and regulation of interleukin-11 by breast cancer cells

We have studied the production of interleukin-11 (Il-11) in 13 breast cancer cell (BCC) lines. Two of these cell lines (MDA-MB-231 and Hs578T) expressed the cytokine at both the protein and mRNA levels. Il-11 did not modulate the growth of five BCC lines examined, including the two cytokine-producing BCC lines. The production of Il-11 was increased by transforming growth factor-beta1 in a dose-dependent manner with a rapid (2 h) and transient (24 h) mRNA induction, but not by epidermal growth factor, insulin-like growth factor-I and -II, basic fibroblast growth factor, platelet-derived growth factor or parathyroid hormone. The cyclic AMP inducer, forskolin, and the activator of protein kinase C, phorbol 12-myristate 13-acetate, also stimulated the production of Il-11. Besides Il-11, MDA-MB-231 and Hs578T were the only BCC lines to produce interleukin-6 (Il-6) protein and mRNA. Since Il-11 and Il-6 are potent stimulators of osteoclast development and bone is a major source of TGF-beta1, our data suggest that Il-11, together with Il-6, contributes to the high bone destructive capacity of MDA-MB-231 cells and could play a role in breast cancer-induced osteolysis.

The "portrait" of hereditary breast cancer.

SummaryFive to ten per cent of all breast carcinomas are of hereditary origin. Many of them have been associated to mutations in the BRCA1 and BRCA2 susceptibility genes. No “BRCA3” gene has been found to account for the non-BRCA1/BRCA2 breast cancer (BRCAx) families, and BRCAx tumors are increasingly believed to originate from multiple distinct genetic events. Phenotype studies have questioned the existence of specific “portraits” among hereditary breast carcinomas (HBC). They have shown that most BRCA1 tumors have a “basal (epithelial)-like” aspect, while BRCA2 and BRCAx HBC are more heterogeneous. HBC have also been submitted to genetic analyses, notably with the objective of resolving the heterogeneity of BRCAx lesions. The present review aims to summarize recent data on BRCA1, BRCA2, and BRCAx HBC, including hypotheses on the origin of BRCA1 tumors and their paradoxical relations to estrogen-sensitivity.

Antiestrogenic activity of two 11β-estradiol derivatives on MCF-7 breast cancer cells

Two 11 beta-derivatives of estradiol (E2) were tested for their potential antiestrogenic activity in the MCF-7 breast cancer model: one contained a phenoxydimethylaminoethyl side-chain (RU 39,411), the other a pentafluoropentylsulfinyl side-chain (RU 58,668). The former compound displayed mixed estrogenic/antiestrogenic properties, while the latter indicated only an antiestrogenic activity. Both the compounds produced a growth inhibition of MCF-7 cells at doses related to their binding affinity for the estrogen receptor (ER); E2 suppressed this inhibition. The compounds also down-regulated the estrogen binding capacity of the cells but failed to reduce ER mRNA levels, indicating that the grafting of their side-chains prevented this antagonistic effect usually observed with steroidal estrogens. Assessment of ER levels by enzyme immunoassay revealed a marked increase with RU 39,411 and a decrease with RU 58,668; different mechanisms of action should, therefore, be considered. Finally, the estrogenic activity of RU 39,411 was demonstrated by its strong ability to induce synthesis of the progesterone receptor; RU 58,668 failed to display this agonistic activity.

Secretory Products of Breast Cancer Cells Specifically Affect Human Osteoblastic Cells: Partial Characterization of Active Factors

The pathogenesis of tumor‐induced osteolysis (TIO) following breast cancer metastases in bone remains unclear. We postulated that osteoblasts could be target cells for the secretory products of breast cancer cells. We previously showed that serum‐free conditioned medium (CM) of the breast cancer cell line MCF‐7 inhibits DNA synthesis by 75% of control values in osteoblast‐like cells SaOS‐2 and that this effect is only in a minor part due to transforming growth factor β secretion. To establish the specificity of our observations and to look for other biologically active factors, we have tested the effects of medium conditioned by several cancer and noncancer cell lines (breast, colon, placenta, or fibrosarcoma) on the proliferation of osteoblast‐like cells (SaOS‐2, MG‐63), normal human osteoblasts, human fibrosarcoma cells, and normal human fibroblasts. Culture medium (1:2) of the breast cancer cell lines MCF‐7, T‐47D, MDA‐MB‐231, and SK‐BR‐3 inhibited by 25–50% the proliferation of osteoblast‐like cells SaOS‐2, MG‐63, and normal osteoblasts as evaluated by the MTT survival test or [3H]thymidine incorporation. MCF‐7 cells completely inhibited the proliferation of normal human osteoblasts in coculture. This inhibitory effect was reversible and not due to cytotoxicity. Moreover, the cyclic adenosine monophosphate (cAMP) response to parathyroid hormone (PTH) of osteoblast‐like cells SaOS‐2 was also increased by 100–240% by the same CM. Such activities were, however, not detected in medium from the breast noncancer cell line HBL‐100 or in the medium conditioned by non–breast cancer cell lines (COLO 320DM, HT‐29, JAR, or HT‐1080). Medium from the breast cancer cells had no effect on normal human fibroblasts or fibrosarcoma cells (HT‐1080), suggesting the specificity of their action on human osteoblasts. After partial purification by ultrafiltration and size‐exclusion chromatography, we found that medium of T‐47D cells contained at least three nonprostanoid factors of low molecular weights (apparent MW of 700, 1500, and 4000 D) which affected human osteoblast‐like cells. These factors were heat stable and could be peptides without disulfide bonds. In summary, our data show that human breast cancer cells release soluble factors that inhibit osteoblast proliferation and increase their cAMP response to PTH, indicating that osteoblasts could be important target cells for breast cancer cells and could be involved in the process of TIO.