New mechanistic insights of integrin ß1 in breast cancer bone colonization.

Bone metastasis is a frequent and life-threatening complication of breast cancer. The molecular mechanisms supporting the establishment of breast cancer cells in the skeleton are still not fully understood, which may be attributed to the lack of suitable models that interrogate interactions between human breast cancer cells and the bone microenvironment. Although it is well-known that integrins mediate adhesion of malignant cells to bone extracellular matrix, their role during bone colonization remains unclear. Here, the role of ß1 integrins in bone colonization was investigated using tissue-engineered humanized in vitro and in vivo bone models. In vitro, bone-metastatic breast cancer cells with suppressed integrin ß1 expression showed reduced attachment, spreading, and migration within human bone matrix compared to control cells. Cell proliferation in vitro was not affected by ß1 integrin knockdown, yet tumor growth in vivo within humanized bone microenvironments was significantly inhibited upon ß1 integrin suppression, as revealed by quantitative in/ex vivo fluorescence imaging and histological analysis. Tumor cells invaded bone marrow spaces in the humanized bone and formed osteolytic lesions; osteoclastic bone resorption was, however, not reduced by ß1 integrin knockdown. Taken together, we demonstrate that ß1 integrins have a pivotal role in bone colonization using unique tissue-engineered humanized bone models.

Radiological pitfall: Siliconoma in internal mammary lymph node mimics breast cancer recurrence.

Siliconomas are rarely found in internal mammary lymph nodes in the context of ruptured, ipsilateral, silicone breast implants. However, they can sometimes cause a diagnostic dilemma, as in the presented case. We discuss the diagnostic pitfalls that can arise from misinterpreting a siliconoma for a metastatic lymph node, review the literature, and suggest appropriate diagnostic approaches.

Epstein-Barr virus-positive diffuse large B-cell lymphoma of the elderly expresses EBNA3A with conserved CD8 T-cell epitopes.

Post-transplantation lymphoproliferative disorders (PTLD) arise in the immunosuppressed and are frequently Epstein-Barr virus (EBV) associated. The most common PTLD histological sub-type is diffuse large B-cell lymphoma (EBV+DLBCL-PTLD). Restoration of EBV-specific T-cell immunity can induce EBV+DLBCL-PTLD regression. The most frequent B-cell lymphoma in the immunocompetent is also DLBCL. 'EBV-positive DLBCL of the elderly' (EBV+DLBCL) is a rare but well-recognized DLBCL entity that occurs in the overtly immunocompetent, that has an adverse outcome relative to EBV-negative DLBCL. Unlike PTLD (which is classified as viral latency III), literature suggests EBV+DLBCL is typically latency II, i.e. expression is limited to the immuno-subdominant EBNA1, LMP1 and LMP2 EBV-proteins. If correct, this would be a major impediment for T-cell immunotherapeutic strategies. Unexpectedly we observed EBV+DLBCL-PTLD and EBV+DLBCL both shared features consistent with type III EBV-latency, including expression of the immuno-dominant EBNA3A protein. Extensive analysis showed frequent polymorphisms in EB-NA1 and LMP1 functionally defined CD8+ T-cell epitope encoding regions, whereas EBNA3A polymorphisms were very rare making this an attractive immunotherapy target. As with EBV+DLBCL-PTLD, the antigen presenting machinery within lymphomatous nodes was intact. EBV+DLBCL express EBNA3A suggesting it is amenable to immunotherapeutic strategies.

Galectin-1 mediated suppression of Epstein-Barr virus specific T-cell immunity in classic Hodgkin lymphoma.

In Hodgkin lymphoma (HL), the malignant Hodgkin Reed-Sternberg cells interact with the host microenvironment to create an immunosuppressive network that protects the lymphoma from immune attack. These mechanisms are not fully understood. We examined the role of the immunomodulatory protein galectin-1 (Gal-1) on Epstein-Barr virus-specific CD8(+) T cell responses in HL. Initial studies indicated Gal-1 expression in all in vitro established Hodgkin Reed-Sternberg cell lines. In situ analysis revealed Gal-1 expression in 26 of 42 classic HL, whereas Gal-1 was uniformly negative in nodular lymphocyte predominant HL. Gal-1(hi) expression was associated with male gender, older patients, reduced CD8(+) T cell infiltration at the tumor site, and most importantly, an impaired latent membrane protein 1 and 2-specific CD8(+) T-cell responses. In vitro exposure to recombinant Gal-1 inhibited proliferation and interferon-gamma expression by Epstein-Barr virus-specific T cells. These observations provide an important link between the Gal-1-mediated immunomodulatory networks and loss of antigen-specific T-cell function in classic HL.

FGFR1 and WT1 are markers of human prostate cancer progression.

BACKGROUND: Androgen-independent prostate adenocarcinomas are responsible for about 6% of overall cancer deaths in men. METHODS: We used DNA microarrays to identify genes related to the transition between androgen-dependent and androgen-independent stages in the LuCaP 23.1 xenograft model of prostate adenocarcinoma. The expression of the proteins encoded by these genes was then assessed by immunohistochemistry on tissue microarrays (TMA) including human prostate carcinoma samples issued from 85 patients who had undergone radical prostatectomy. RESULTS: FGFR1, TACC1 and WT1 gene expression levels were associated with the androgen-independent stage in xenografts and human prostate carcinoma samples. MART1 protein expression was correlated with pT2 tumor stages. CONCLUSION: Our results suggest that each of these four genes may play a role, or at least reflect a stage of prostate carcinoma growth/development/progression.

Preoperative systemic chemotherapy does not modify strategy of liver resection.

BACKGROUND/AIMS: Most patients who undergo surgical resection of colorectal liver metastases have been previously treated with chemotherapy as adjuvant therapy or as a primary treatment. The aim of this retrospective study was to compare morbidity, mortality, liver function and histology of the liver specimens in patients undergoing colorectal liver metastases (CRLM) resection with and without a history of previous chemotherapy. METHODOLOGY: Records of 210 patients who underwent CRLM resection in our institution between January 1996 and March 2003 were retrospectively reviewed. We selected for further analysis medical charts of 40 patients who didn't receive a combined treatment concurrently to liver resection. Group I included 25 patients who did not receive adjuvant chemotherapy. Group II included 15 patients who received chemotherapy in the 3 months before liver resection. RESULTS: Postoperative mortality was 0% and 0.7% in group I and II respectively. Specific liver complications and pulmonary complications were similar in the two groups: 20% and 32% us. 33% respectively. The mean length of stay in the hospital was similar in the two groups (19 +/- 14 vs. 14 +/- 8). Changes of liver function test were similar in the two groups. Pathologic examination of liver specimens was not different in the two groups. CONCLUSIONS: Preoperative systemic chemotherapy didn't increase the risk of liver resection.

"Sugar" tumor of the pancreas: a rare entity that is diagnosable on preoperative fine-needle biopsies.

This study is the second to report a pancreatic "sugar" tumor (ST) case. This ST was incidentally discovered in a 31-year-old woman using computed tomography scan (CT scan) for work-up of a hepatic focal nodular hyperplasia. Both CT scan and endoluminal ultrasonography (EUS) features evoked a 15-mm large benign endocrine tumor. Pathological examination of EUS-guided fine-needle aspiration biopsies could not confirm this diagnosis. Laparoscopic corporeo-caudal pancreatectomy was performed. The tumor was intrapancreatic, well circumscribed, and organized in sheets of epithelioid cells. The tumor cells expressed HMB-45 but did not express epithelial or endocrine immunohistochemical markers. These histophenotypic features are those of an extra pulmonary ST, which belong to the PEComa family of tumors. Retrospective examination of preoperative biopsies evidenced the same histophenotypic features. This observation highlights that STs should be considered in preoperative differential diagnosis of pancreas tumors, since they may be treated by limited surgical resection.

Identification of TCL1A as an immunohistochemical marker of adverse outcome in diffuse large B-cell lymphomas.

We used a combination of DNA-microarray and tissue-microarray (TMA) analyses to identify markers that could be routinely used to predict the outcome of diffuse large-B-cell lymphoma (DLCL) patients. Gene expression profiling was performed using DNA-microarrays on 52 tumour biopsy samples [31 DLCL and 21 follicular lymphomas (FL)] from 48 patients (28 DLCL and 20 FL). T-cell leukemia/lymphoma-1A (TCL1A) mRNA overexpression was correlated with relapse in DLCL patients. TMA analysis was applied on a distinct series of 36 formalin-fixed, paraffin-embedded DLCL samples and showed that TCL1A immunoexpression was correlated with either higher relapse (p=0.02) or lower 5-year overall survival (p=0.009) rates. Moreover, the prognostic value of TCL1A was independent from IPI in our series. Our data suggest that TCL1A immunodetection is an independent marker of adverse outcome that could be used in routine settings for the management of DLCL patients.

Prognostic significance of angiogenesis evaluated by CD105 expression compared to CD31 in 905 breast carcinomas: correlation with long-term patient outcome.

Our purpose was to determine the respective prognostic significance of CD105 and CD31 immunoexpression in node negative patients with breast carcinoma, since angiogenesis induces blood borne metastases and death in carcinomas. CD105 (endoglin) has been reported as expressed by activated endothelial cells and consequently should better reflect neoangiogenesis in malignant tumors. Comparison of CD31 and CD105 immunocytochemical expression was undertaken in a series of 905 breast carcinomas. Results were compared to patients' long-term (median = 11.3 years) outcome. Univariate (Kaplan-Meier) analysis showed that the number of CD105+ microvessels (cut-off 15 vessels) correlated significantly with poor overall survival (p=0.001). This correlation was less significant in node negative patients (p=0.035). The number of CD31+ microvessels (cut-off 25 vessels) similarly correlated with poor survival (p=0.032) but not in the subgroup of node negative patients. Marked CD105 expression also correlated with a high risk for metastasis in all patients (p=0.0002) and in the subset of node negative patients (p=0.001). Similarly metastasis risk in node negative patients correlated with marked CD31 immunocytochemical expression (p=0.02). Multivariate analysis (Cox model) identified CD105, but not CD31 immunoexpression, as an independent prognostic indicator. Our results suggest that: i) in breast carcinomas, immunoselection of microvessels containing activated CD105 labelled endothelial cells is endowed with a stronger prognostic significance, as compared to CD31 vessels labelling; ii) the CD105 immunoexpression may be considered as a potential tool for selecting node negative patients with a poorer outcome and higher metastasis risk; iii) in these patients specific antiangiogenic therapy targeted by anti-CD105 conjugates can be further developed.

[Prognostic significance of VEGF receptors, VEGFR-1 (Flt-1) and VEGFR-2 (KDR/Flk-1) in breast carcinoma]. / Valeur pronostique de l'expression des recepteurs du VEGF, VEGFR-1 (FLT-1) et VEGFR-2 (KDR/FLK-1) dans le cancer du sein.

OBJECTIVES: The aim of this study was to investigate the prognostic value of vascular endothelial growth factor (VEGF) receptors, VEGFR-1 (Flt-1) and VEGFR-2 (KDR/Flk-1) in breast carcinoma. METHODS: VEGF receptor expression was investigated using immunohistochemical assays with monoclonal antibodies on frozen sections in a series of 918 patients and was correlated with prognostic parameters and with long-term follow-up (median, 11.3 years). VEGFR-1 and VEGFR-2 immunostained surface was evaluated in percentage of the total tumor specimen surface by light microscopy (x100). RESULTS: VEGFR-1 and VEGFR-2 were strongly expressed in endothelial cells within blood microvessels, and weakly in tumor cells. Univariate (Kaplan Meier) analysis showed that VEGFR-1 positive tumor surface (cut off=5%) was not correlated with survival, but was significantly correlated with high metastasis risk (p=0.03) and relapse (p=0.01) in all patients, and in those with node negative tumors (p=0.001 and p=0.01 respectively). In multivariate analysis (Cox model), VEGFR-1 expression was identified as an independent prognostic indicator. Univariate analysis showed that VEGFR-2 positive tumor surface (cut off=10%) was not correlated with survival or with metastasis risk and relapse. CONCLUSION: Our results show that VEGFR-1 immunohistochemical expression permits the identification of patients with poor outcome, particularly those with node negative tumors, with high risk of metastasis and relapse. VEGFR-1 immunodetection may further be considered as a potential tool for evaluating tumor agressiveness and therapeutic strategies in breast cancer.

Characterization of Hodgkin's lymphoma-like undifferentiated carcinoma of the nasopharyngeal type as a particular UCNT subtype mimicking Hodgkin's lymphoma.

Undifferentiated carcinoma of the nasopharyngeal type (UCNT) that histologically mimics Hodgkin's lymphoma (HL) ("Hodgkin's lymphoma-like UCNT"--HL-like UCNT) is known as a diagnostic pitfall. Using immunohistochemistry, Western blot and cDNA array technology, we wanted to document its phenotypical and molecular characteristics. We report herein 5 cases of UCNT that morphologically mimic HL and 3 classical UCNT cases. We compared the expression profiles of a thousand selected genes in HL-like UCNT and in classical UCNT cases. No difference in the profile of EBV infection was noted between the HL-like UCNT and control cases. Significant differences were detected in the expression of genes involved in the matrix modelling, angiogenesis, apoptosis and regulation of the Th-2 interleukins. The eosinophil chemoattractant eotaxin was expressed in the stroma of HL-like UCNT, but not in the control cases. The eotaxin receptor CCR3 was expressed in both stromal and carcinoma cell populations of HL-like UCNT, this pattern being similar to the one observed in HL. These results show that UCNT morphologically resembling HL share also some specific phenotypical and molecular features with HL, and might deserve to be isolated as a particular UCNT subtype.

Constitutive nuclear localization and initial cytoplasmic apoptotic activation of endogenous caspase-3 evidenced by confocal microscopy.

The localization of caspases and their substrates in different cellular compartments may be one way to regulate apoptosis. Caspase-3-dependent proteolysis of inhibitor caspase-activated deoxyribonuclease (ICAD) activates caspase-activated deoxyribonuclease (CAD), which induces apoptotic internucleosomal DNA degradation. The nuclear localization of ICAD, pro- and active-caspase-3 molecules remains a controversial issue. Using a combination of immunodetection of endogenous molecules and confocal microscopy, we analysed the kinetics of the procaspase-3 and CAD activation induced by FAS triggering in Jurkat cells. Through a semi-quantitative image analysis, we showed a constitutive nuclear localization of pro-caspase 3 and ICAD in non-apoptotic cells. FAS stimulation induced 7A6 apoptotic antigen expression, which could be related to three different sequential patterns of nuclear chromatin organization. Active-caspase-3 first appeared in the cytoplasm and was next observed in the nucleus. Simultaneously, the amount of ICAD located in the nucleus decreased, whereas the amount of ICAD located in the cytoplasm remained unchanged. Thus, our experiments using in situ immunodetection of endogenous molecules show that the ICAD cleavage by the active-caspase-3 probably takes place in the nucleus. These results provide new perspectives about the subcellular compartmentation and traffic of caspases during the apoptotic process.

[Accuracy of intraoperative frozen section diagnosis in non palpable breast lesions: a series of 791 cases]. / Efficacité de l'examen peropératoire des lésions infracliniques mammaires: études sur une série de 791 cas.

The aim of this study is to estimate the accuracy and reliability of intraoperative frozen section of nonpalpable breast lesions. In fact, frozen section of palpable breast lesions has proven to be valid, but its use in breast infraclinic lesions has been discussed recently, with the publication of European recommendations. Diagnosis on frozen section was routinely performed in a serie of 791 patients with nonpalpable mammographically detected abnormalities breast lesions from January 1990 through July 2000. The initial frozen section diagnoses with known mammographic pattern were compared with the diagnoses obtained on permanent paraffin sections to estimate the accuracy of frozen sections. Frozen section diagnosis was delayed until final diagnosis assessed on permanent paraffin sections in only 8 cases (1%). Frozen section diagnoses were accurate in 744 of 783 cases (95%). The diagnosis was modified on the basis of permanent sections in 39 cases; consisting of 39 false negative. No false positive diagnosis was noted. Sensitivity and specificity of frozen section diagnoses were 87,69 and 100, respectively. When the comparison between frozen and permanent section was analyzed according to the mammographic pattern, the sensitivity among patients with microcalcifications was lower (75,23) than among patients with opacities (93,86). When frozen section and permanent section diagnoses were related according to the mammographic size ( 10 mm) the sensitivity among patients with opacities measuring less than 10 mm was lower (91,75) than among patients with opacities larger than 10 mm (95,65) and the sensitivity among patients with microcalcifications larger than 10 mm was greater (77,14) than among patients with microcalcifications size less than 10 mm (74,28). This results are similar to those obtained on palpable breast lesions, and show that frozen section is a feasible and reliable procedure in nonpalpable breast lesions, particularly more relevant in mammographic opacities than in microcalcifications, whatever the lesion size.

CD105 expression is a marker of high metastatic risk and poor outcome in breast carcinomas. Correlations between immunohistochemical analysis and long-term follow-up in a series of 929 patients.

CD105 (endoglin) is expressed significantly in activated endothelial cells in culture and in tumor microvessels. Quantification of CD105 immunocytochemical expression that may be clinically relevant has not been accurately evaluated. We studied CD105 expression on frozen tissue sections by using immunohistochemical assays in a series of 929 patients and correlated the findings with long-term follow-up (median, 11.3 years). Univariate (Kaplan-Meier) analysis showed that the number of CD105+ microvessels (cutoff, 15 vessels) correlated significantly with poor overall survival among all patients (P = .001). This correlation was less significant in node-negative patients (P = .035). Marked CD105 expression also correlated with a high risk for metastasis among all patients (P = .006) and among node-negative patients (P = .001). Multivariate analysis (Cox model) identified CD105 immunodetection as an independent prognostic indicator. Our results suggest that immunohistochemical expression of CD105 has practical clinical relevance for identifying node-negative patients with a poor prognosis. Moreover, immunodetection of CD105 also may be considered a potential tool for selecting patients who could benefit from specific antiangiogenic therapy, using anti-CD105 conjugates.

[The immunohistochemical expression of CD105 is a marker for high metastatic risk and worse prognosis in breast cancers]. / L'expression immunohistochimique du CD105 est un marqueur de risque métastatique élevé et de mauvais pronostic dans les cancers du sein.

The quantification of angiogenesis in human solid tumors has been shown to be an indicator of prognosis and tumor microvasculature is a candidate target for antiangiogenic therapy. CD105 (endoglin) is significantly expressed in activated endothelial cells in culture and in tumor microvessels. Quantification of CD105 immunocytochemical expression that may be of significant clinical relevance, has not been accurately evaluated as yet. In the present report, CD105 expression on frozen sections was investigated using immunohistochemical assays in a series of 929 patients and correlated with long-term (median = 11.3 years) follow-up. The CD105 immunostaining was observed on endothelial cells mostly in small cells. The number of vessels and the immunostained surface were evaluated in so called "hot spots" within tumor stroma. Both the number of vessels and immunostained surface were correlated to the patients' outcome (overall survival, disease free survival, metastases) in the whole group of patients and also specifically in node negative subgroup. Univariate (Kaplan Meier) analysis showed that the number of CD105 positive microvessels (cut-off n = 15) was significantly correlated with poor overall survival, among all patients (p = 0.001). This correlation was less significant in the group of node negative patients (p = 0.035). Marked CD105 expression was also correlated with high metastasis risk among all patients (p = 0.006) and among node negative patients as well (p = 0.001). In multivariate analysis (Cox model) CD105 immunodetection was identified as an independent prognostic indicator. Our results suggest that CD105 immunohistochemical expression has a practical clinical relevance for identifying node negative patients with poor prognosis. Moreover, the CD105 immunodetection may also be considered as a potential tool for selecting patients that could benefit from specific antiangiogenic therapy, using anti CD105 conjugates.