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Biobanks in general, and specifically tumour banks, are considered as essential tools for the development of translational and clinical research in biology and oncology. Biobank tasks include the collection and preservation of biological samples, and their association with information that will be essential for further scientific use ("annotations" that allow for the "qualification" of biological samples in biological resource). A collection is made of a series of biological resource that are representative of a homogeneous group of individuals or patients that are defined on the basis of clinical or biological information. Collections are used by scientists that are aware of their existence. In the absence of a published catalogue, this awareness is most often limited to research teams that are geographically close, or to investigators who already established collaborative projects with medical teams within the hospital that operates the tumour bank. Publications of catalogues, especially digitalized and online catalogues, should foster the development of high-level, large-scale and multicentric scientific projects. In addition, tumour banks will formalize rules that allow publication of collections, and upstream, rules that are used to qualify biological samples in biological resource: this should translate in an improved overall quality of samples and annotations. Tumour bank catalogues remain relatively few; however, some recent achievements established the "proof of concept" and already raise questions regarding rules for publication. It will be important to demonstrate that these high expectations translate into measurable benefits.

Quantitative microscopy by image analysis allows not only to measure various parameters on each cell but also to consider the global population as a whole. In the hypothesis that cell position is reflecting the relational and dynamical structure of the system, spatial arrangement analysis may help to show up intercellular communication (interactions and control systems via contact or diffusible factors). We describe a topographical analysis method used to study these neighbour relationships, and thus the sociological behaviour of the cells. It is applied to the study of the effect of estrogenic and antiestrogenic treatments on a breast cancer cell line (MCF-7). It shows up that estrogens increase proliferation and induce an unusual topographical behaviour, notably in cell cycle phases: cells in S phases are very randomly distributed. It points out the role of estrogens on the cells neighbour relationships inducing the way to a permissive proliferation context. This effect is reversed by antiestrogenic treatment after a few days. Antiestrogenic treatment alone increases the proliferation constraint.

It is now clearly established that proteolytic enzymes, and in particular plasminogen activator (uPA), play an important role in breaking down the extracellular matrix, which is considered to be a step in metastasis formation. Plasminogen activators are controlled at various levels. Two inhibitors, PAI-1 and PAI-2, have been identified, the latter being more specific for uPA. In attempts to determine their prognostic value, it is essential to investigate the relative importance of these parameters and their interactions. We used an immunoenzymatic method to assay uPA, PAI-1 and PAI-2 antigens in cytosols prepared from 314 primary breast tumors. The patients were followed up for a minimum of six years and all relevant clinical and laboratory findings had been recorded. Univariate analysis confirmed the poor outcome of patients whose tumors contained large amounts of uPA and PAI-1. In addition, low levels of PAI-2 correlated with shorter disease-free survival in the overall population (P = 0.02), post-menopausal women (P = 0.02) and women without lymph node involvement (P = 0.02). Multivariate analysis using the "Main Effects" Cox model identified node involvement, macroscopic tumor size and PAI-2 as significant variables. The "interactive" Cox model, taking into account interactions between uPA and its two inhibitors, identified a first subgroup with a very poor prognosis associating either high levels of PAI-1 with low levels of PAI-2 in the overall population as well as following stratification for axillary node negative disease, or high levels of uPA with low levels of PAI-2 in the group of menopausal women. We conclude that PAI-1 provides the same prognostic informations as uPA, and does not appear to play its role as an inhibitor. In contrast, PAI-2 increased the prognostic value of both uPA, particularly in post-menopausal women, as well as PAI-1 in a subgroup of axillary node negative patients.

The appearance of mucositis is a frequent and painful secondary effect of anticancer chemotherapy. Patients who develop oral toxicity during the first course of treatment will almost assuredly show identical side effects during each subsequent course unless the drugs are changed or the doses are lowered. In the absence of an efficacious antidote or preventive prophylaxis for such lesions to date, this report presents the results of a preliminary retrospective non-randomized study of the effect of soft-laser treatments on mucositis in cancer patients receiving combination chemotherapy, including 5-fluorouracil. Iatrogenic mucositis was observed during 43% of 53 chemotherapy cycles in the case control population. Curative laser therapy reduced the time to repair lesions and the rate of therapeutic modifications. For patients who received soft-laser therapy as a preventive measure, the incidence of oral complications was reduced to 6% during 101 cycles of chemotherapy. All of these patients, even those who have encountered mucositis before receiving preventive laser therapy, terminated their cancer therapy as originally scheduled. Well designed and carefully controlled trials will be necessary to define the place of helium-neon laser therapy in the repair and prevention of oral complications due to cancer chemotherapy.

Preservation of the shape and the integrity of multicellular eukaryotes needs rigorous cell proliferation monitoring. During the prereplicative G1 phase, a finely adjusted and specific control supervises the proliferant/non proliferant states of the cells. Some molecular mechanisms of growth regulation have been identified in recent years. Changes in normal cell attachment on extracellular matrix and intercellular chemical signalling (secretion of informative molecules) activate intracellular signals for division. The transduction mechanisms of the extracellular signalling to the nucleus have been partially elucidated for steroid hormones and growth factors. Molecular biology research and proto-oncogene discoveries have led to considerable progress in understanding the role of these normal genes in the control of cellular proliferation. The initiation of the response to extracellular factors requires: i), direct transducers (specific binding of the steroid hormone on its cytoplasmic or nuclear receptor and high affinity binding of this activated complex to specific DNA sequences); and ii) indirect transducers (binding of growth factors on extracellular domains of specific receptor proteins which convert this extracellular event into several intracellular signals, secondary messengers, protein kinases and specific nuclear regulatory factors). Whatever the transduction system, nuclear events control transcription of growth regulatory genes. The series of enzymatic reactions set in motion by indirect transduction systems require strict regulation systems, the diversity and the complexity of which has been perceived in studies on jun and fos gene families. Each proliferation step is governed by growth stimulators and growth inhibitors, the transformation of normal cells to cancer cells resulting from alterations of these regulatory process. Independent of extracellular stimuli and of their transfer to the nucleus, intracellular controls coordinate cell cycle phases (G1, S, G2 and M) to produce daughter cells identical to the original cell. Two control points are particularly critical: one in G1 (the "start" point) and the other in G2 just before mitosis. Although intermediate steps between extracellular and intracellular controls are still unknown, yeast gene analyses have allowed determination of molecular regulatory mechanisms implicated in the passage of these critical points. A considerable advance was made by the discovery that some of the involved components presented strong sequence and function homologies in organisms from yeast to man, suggesting a phyllogenetically conserved mechanism. It seems likely that the phosphorylation state of protein p34, its association with a G1-phase specific cyclin or a M-phase specific cyclin, and its protein kinase activity regulate the proliferation state of higher eukaryotic cells. In spite of significant advances, much research is still necessary to elucidate all the mechanisms involved in cell cycle control.

This study concerns the correlation between ER and PR status, menopausal status and the effect of adjuvant hormonotherapy in high risk primary breast cancer patients. We have compared the results obtained in a randomized trial (Institut Paoli-Calmettes, Marseille) with those of a historic series (Centre René Huguenin, Saint-Cloud). The patients presented the same clinical and histological criteria and received identical therapeutic protocols (chemotherapy and/or hormonotherapy). Compared with patients receiving no adjuvant treatment, it appeared that: 1) in post-menopausal patients, a significant detrimental effect of tamoxifen was found in ER-negative patients, while in ER-positive patients hormonal treatment was well correlated with both the presence and level of steroid receptors; 2) in pre-menopausal patients, hormonal therapy (oophorectomy + tamoxifen) appeared to be mediated by a complex mechanism involving more than an ER-positive cell population. In the light of the published results, the present findings underline the importance of reevaluating the indications of hormonotherapy in terms of hormone receptors and menopausal status. They also indicate the importance of biological factors in the evaluation of response to therapy. They can identify, subsets of patients in whom a given therapeutic protocol is detrimental, even though it may be beneficial for the overall population. heterogeneity in response to therapy among patient subsets is one of the most important problems which confronts medical statisticians and clinical investigators.

In order to test the qualities of the 2 assays, in the same laboratory and on the same tumors, a single-point dextran-coated charcoal radioligand binding assay (RLA-DCC) and Abbott enzyme immunoassay (EIA) were used for more than two years to perform estrogen receptor determinations on cytosols from 2,134 breast cancers. Statistical analysis of the data was performed according to the method of Passing-Bablock. The final regression curve between EIA (y) and RLA-DCC (x) was excellent y = 1.187 X fmol/mg of protein. However, from 1986 to 1988, a great variability was observed for this correlation. We report the study of this variability, which could be explained by several factors, especially calibration problems for the immunoassay kits and changes in our technical team. The binding assay appears to be more sensitive to the technicians' experience than the immunoassay. Technical points are discussed, particularly cytosol preparation and KCl presence or absence in the homogeneisation buffer. The conditions allowing for optimal correlation and routine determination fiability can therefore be defined.

The total amount of cathepsin-D (mature forms and pro-enzyme especially) in the cytosol of 88 breast cancers including 85 primary cancers and 11 axillary lymph node metastasis was measured by way of immuno-radiometric assay. Maximum follow-up is 58 months. Cathepsin-D was found to be independent of clinical and biological parameters including axillary lymph node involvement and oestradiol receptors (RE). Univariate analysis has pointed out a significant linkage between overall survival and cathepsin-D, using a cut-off level of 30 pmol/mg protein which is the most discriminating value. Cathepsin-D appeared to be particularly useful in lymph-node-positive and RE-negative patients but it was not significant in the node-negative population. Multivariate analysis of the overall survival (Cox model) revealed that it was strongly related to oestradiol receptor, lymph node involvement and cathepsin-D.

Estrogen receptors (ER) were investigated using immunohistochemical techniques in 54 cases and biochemical techniques in 38 cases on a series of biopsy specimens of normal, hyperplasic and malignant mammary tissue (intraductal carcinoma). Immunohistochemical data were submitted to quantitative and semi-quantitative analysis (SAMBA 200, TITN). On normal tissue, immunostaining is bright and evenly distributed in galactophores and ductulo-lobular junctions; it is unevenly distributed but consistently present in lobules and increasing after menopause. On hyperplasic tissue, immunostaining of papillary cystadenomas and epitheliosis and fibroadenosis zones was similar to normal tissue with respect to the intensity and heterogeneity. Conversely, their immunohistochemical features are close to blunt duct adenosis and atypical lobular hyperplasia of ductulo-lobular junctions. On malignant tissue, immunostaining is unpredictable, it can be evenly distributed, unevenly distributed in patches or gradients, or totally absent. Immunohistochemical techniques can only be used to assess the intensity and distribution of ER in function of cell type in normal tissue and during a cancerous process. Thus strong receptivity of ductulo-lobular junctions may reinforce the claim that these structures are the targets for estrogen co-carcinogenic substances. The heterogeneity of ER at the onset of carcinoma suggests the possibility of several pathway of development. The fact that myoepithelial cell never seem to display estrogen type receptivity makes it logical to seek a special sensitivity for them.

Estrogen receptor determinations were performed on 241 cytosols from 160 breast cancer tumors, using both dextran coated charcoal method (D.C.C.) and monoclonal antibodies (Abbott's ER-EIA kit) in order to compare the two methods, and to evaluate the clinical usefulness of this new immunological simplified assay. Intra- and inter-assay reproducibility were studied during a six month period. Intra-assay of both methods were lower than 5%. Inter-assay variation coefficients of ER-EIA studied on 35 standard curves varied from 12.5% (standard: 0) to 6.7% (standard: 250 fmoles/ml). ER determination of 80 human breast cancers were performed both by EIA and Scatchard analysis. The regression curve obtained was (EIA) = 1.04 (Scatchard) + 21 fmoles/mg of proteins (r = 0.963). With 153 breast cancer cytosols whose volume was too small for multipoint Scatchard analysis, EIA results were compared to results obtained by a "near saturating" concentration of tritiated ligand (5 nM). The regression curve obtained was (EIA) = 1.34 (5nM) + 5 fmoles/mg proteins (r = 0.978). These results can be compared with those obtained between Scatchard and 5 nM: (Scatchard) = 1.29 (5nM) - 9 fmoles/mg proteins (r = 0.985). Reproducibility was studied on clinical specimens assayed at two different periods during the clinical evaluation. Regression curves obtained were (2nd assay) = 1.05 (1st assay) - 5.5 fmoles/mg proteins for EIA, and (2nd assay) = 0.96 (1st assay) + 9 fmoles/mg proteins for DCC method. EIA assay presented an high stability for protein concentrations very low, up to 0.2 mg/ml. Finally, a very good correlation was obtained between ER-EIA and DCC method, and ER-EIA seemed specially well fitted to small tumors.

An estrogen receptor immunocytochemical assay (ER-ICA) was applied to 130 malignant breast carcinomas and the results were compared to those of steroid binding assays performed on cytosol extracts of the same tumors. Also laminin (lam) distribution was studied in the same tumors. A semi quantitative analysis and a computerized image analysis system (SAMBA 200 TITN) were used to evaluate the positive ER and lam immunostaining. Positive ER immunostaining was always located in the nucleus of tumor cells and of normal cells in adjacent breast tissue. When immunohistochemical staining was correlated to biochemical assay there was a 88% correlation staining intensity and percentage of positive cells significantly increased (p less than 0.01) with cytosolic ER levels. Lam was observed within vascular and epithelial basement membranes (BMs). Lam staining displayed a continuous linear pattern in intraductal carcinomas or was heterogeneously distributed with a discontinuous linear pattern in invasive carcinomas. No intracellular lam staining was detected. In tumors, laminin immunostaining revealed often multilayered BMs and abnormal multilayered BMs in blood vessels in the tumor stroma. These results indicated that (ER-ICA) is to date the most reliable histochemical method for ER detection and correlated in 88% of the cases with ER biochemical assay ER-ICA provides additional information for heterogeneous ER distribution within tumors ER-ICA as a qualitative method is unable to replace the quantitative ER determination obtained with biochemical assay ER-ICA based on ER antigenic site detection is complementary to biochemical assay based on ER functional site determination laminin immunostaing constitutes a new approach to the heterogeneous BM changes occurring in carcinomas, and permits a better understanding of cell diffusion processes and of stroma-tumor cells interactions: the consistent extracellular lam distribution in contact with the stroma, indicates that the latter plays an important role in the assembly of BM components the SAMBA 200 permits a reliable accurate evaluation of the percentage of the immunostained cells and surfaces.

Laminin distribution in tissues from surgically removed thyroids consisting in normal gland (5), Basedow's disease (5), thyroiditis (5), follicular adenomas (8), papillary carcinomas (8), follicular carcinomas (6) was studied using rabbit laminin antibody raised against murine laminin. Immunoperoxidase technique (Avidin-Biotin-Peroxidase Complex) was performed on (1) paraffin sections of fixed tissue (2) frozen sections for light microscopy examination. Vibratome thick sections (100 micron) and pre-embedding technique were used for electron microscopy study. Positive staining, was obtained only on frozen and vibratome sections and was found within basement membranes but never in epithelial cell cytoplasm. Laminin had a similar distribution in follicular adenomas, Basedow's disease and normal tissue. Nests of damaged cells in Hashimoto's thyroiditis lacked positive laminin immunostaining. In papillary carcinomas positive staining was found beneath the epithelial cells along the cores. In well differentiated follicular carcinomas the perifollicular laminin staining was preserved, whereas in poorly differentiated follicular carcinomas laminin staining was barely visible or absent.

Breast cancer specimens from 115 patients were assayed for the presence of estrogen receptors (ER) using a monoclonal antibody (H222 SP gamma) and an immuno-histochemical method ER-ICA. The specific ER immuno-staining was observed in cell nuclei. The intensity of the immuno-staining level (ISL) and the percentage of positive cells (PC) variable within tumors tissue, were graded (from 0 to 3): intensity level (IL) and percentage of positive cells (PC). (IL + PC) were correlated to biochemical ER assay using radiolabeled ligands ER - RLA. Both ER - ICA and ER - RLA were correlated in 87.9% of the cases. Discrepancies between both assays appeared mainly due to heterogeneity of the tissue samples. From this study it is concluded that ER - ICA is a reliable histochemical method since well correlated with biochemical assay, easily and rapidly performable, and constitutes a new approach method for ER detection in tissue, particularly valuable in minimal breast cancer, and precancerous lesions.

In 318 cases of human primary breast carcinoma, the presence and content of estrogen receptor (ER) have been correlated to histopathological features of these carcinomas. We have evidenced that: a) there is a relationship between the ER presence, the histoprognostic grade and the histological type; b) there is no correlation between the frequency of ER presence, the neoplasic cellularity and the stromal reaction. The presence of ER appears to be related to a differentiated state of the carcinoma and to a moderate neoplasic activity. These data show that ER, correlated with histopathological features, could be used as an estimation's factor of the cellular activity of a human mammary carcinoma, thus as a factor of better prognostic value for the evolution of this tumor.

Since early antiquity malignant tumors have been recognized and characterized by: 1) their particular local-regional invasive properties. Developing irregularly from the primary tumor mass, the invasion of adjacent tissues often displays classical crab-like images which distinguish, macroscopically and microscopically, malignant tumors from benign tumors. 2) their metastatic capacities. Even though they have been taught since the time of Hippocrates, the specific clinical characteristics of malignant tumors have only recently been apprehended and studied on cellular and biochemical levels. The development of the metastatic phenomenon is a complex chain of events limited to only a few tumor subpopulations. Like every biological study knowledge has progressed over the past years only by concentration of efforts on some of these stages: Invasion associated with disorganization and destruction of basement membranes; Migration of metastatic tumor cells in the stroma and bridging of vascular basal membranes; Colonization of target tissues by recognition and penetration of specific vascular membranes, establishment and concentric development within these tissues. These three stages result from successive interactions between multiple cellular and biochemical phenomena. This review is a schema of our present knowledge of tumor invasion, and more precisely of that concerning the relations between tumor cells and extracellular matrices, particularly basal membranes, leading to a definition of the cellular phenotype with a high metastatic potential.

Progesterone receptors (PR) from human breast tumors were assayed by a new method using monoclonal antibodies immobilized on beads (PR-EIA, Abbott Laboratories). EIA results were compared to those obtained with the dextran-coated charcoal method using a tritiated ligand (ORG 2058). The precision and reproducibility of the EIA method were studied over a 3-month period: intra-assay coefficients of variation were less than 6% for the range of the assay (between 5-250 fmol/ml), and inter-assay coefficients of variation calculated on 13 consecutive standard curves were less than 10%, except for standard 0 (33%). PR assays performed on 78 tumors both by EIA and radioligand (RLA) were compared. The linear regression obtained was: PR-EIA = 0.81 RLA+1 fmol/mg protein (r = 0.88). For reproducibility studies, cytosols were assayed twice during a period ranging from 1 week-3 months, both by EIA and RLA. The linear regression obtained between the second assay (B) and the first assay (A) was: B = 0.98 A + 11 fmol/mg protein for RLA (r = 0.98), and B = 0.99 A-7 fmol/mg protein for EIA (r = 0.98). To study the effect of KCl on PR-EIA, 26 tumors were homogenized in 0.4 M KCl Tris buffer and assayed both by EIA and RLA. A good correlation was obtained between the 2 methods, but higher values were obtained with PR-EIA (P = 1.6) in comparison with RLA. The addition of KCl to the cytosol showed that KCl had no effect on EIA results, but significantly lowered RLA results. To study the effect of KCl on progesterone receptor isoforms, cytosols were analyzed by chromatography on TSK G3000 SW columns, and the presence of PR was detected in each fraction by both EIA and RLA. In the absence of KCl, only the oligomeric form of PR was observed; however with both techniques, detection of this form different from tumor to tumor, emphasizing the inter-tumoral molecular heterogeneity of PR. After PR isoform dissociation by KCl (0.4 M) and chromatographic analysis of the forms obtained, monoclonal antibodies detected PR molecular forms different from those observed by radioligand; furthermore, chromatographic patterns obtained were different from one tumor to another and confirmed the inter-tumoral molecular heterogeneity of the progesterone receptor.

Like other solid tumors ovarian tumors have been intensely investigated these past years to isolate specific tumoral antigens. These antigens are now used by clinical teams for post therapeutic follow-up. However, out of the 25 major antigens actually described in ovarian carcinomas, the only one that can be used today is the carcinoembryonic antigen (CEA). Moreover, an international collaborative work has demonstrated cross reacting antigenicity between these major tumoral antigens. Cellular enzymes (galactosyl transferase and sialyl-transferase) and hormonal receptors for proteinic hormones such as steroid hormones) are being studied but cannot be used as screening markers at this time.