Lack Of Bcl-2 Expression Research Articles

Tumor immune microenvironment and apoptotic markers in breast cancer patients carrying <I>BRCA1</I> gene mutations

Background. It is suggested that defects in BRCA1 / 2 genes contribute to a high mutational load and high immunogenicity, which modulates immune microenvironment. At the same time, it was shown that BRCA1 / 2-associated breast cancer tumors do not belong to the category of immunoactive ones. These tumors have low expression of immune response genes and exhibit an immunosuppressive type of microenvironment. This indicates the need of antitumor immune response modulation and maintaining of the optimal balance of tumor CD4/CD8 T-lymphocytes ratio. In addition, there is evidence of the additional evaluation of TP53 mutation in these tumors and disruption of the cell death process, which can also be a factor of resistance to therapy, including PARP inhibitors, and serve as a therapeutic target.Materials and methods. The prospective study included 20 patients with BRCA1-associated breast cancer. BRCA1 / 2 mutations (BRCA1 185delAG, 4153delA, 5382insC, 3819delGTAAA, 3875delGTCT, 300T>G, 2080delA, BRCA2 6174delT) were detected in by real-time polymerase chain reaction. Immunohistochemical study was performed on paraffin embedded tissue blocks by an automated method on a ThermoScentific immunohistotainer using monoclonal antibodies. The expression of markers of tumor-infiltrating CD4+ and CD8+ T-lymphocytes, markers of macrophages (CD68, CD163), apoptosis (Bcl-2, p53), cell adhesion markers (E-cadherin, β-catenin) in breast cancer in carriers of BRCA1 mutations was assessed.Results. High CD4/CD8 ratio, which characterizes immunosuppressive microenvironment, occurred in 75 % of cases. BRCA1 5382insC mutation is associated with high level of CD4+ TILs (p˂0.05), G2 is associated with a low CD4/CD8 ratio (p = 0.039) and a high level of CD163 (p = 0.02, AUC = 0.739); T1 correlates with high levels of CD8+ TILs (p = 0.038) and high levels of CD163 (p = 0.033). High Ki-67 is associated with a lack of Bcl-2 expression (p = 0.04) and a low level of E-cadherin (p = 0.02). Negative expression of Bcl-2 occurred in 75 % of cases. High level of p53 expression has been described as the main type of expression in these tumors, suggesting a combination of TB53 and BRCA1 mutations and a violation of cell death mechanism of in these tumors.Conclusion. Breast cancer tumors of patients with hereditary mutations in BRCA1 gene demonstrate immunosuppressive type of microenvironment and a violation of the cell death mechanism. The main directions of future therapy of these tumors may include tumor immune microenvironment modification and activation of cell death mechanisms.

47P Immune tumor microenvironment and markers of apoptosis in breast cancer in carriers of hereditary mutations in the BRCA1 gene

Tumor immune microenvironment is currently recognized as an important participant in carcinogenesis, influencing the behavior of tumor cells and treatment efficacy. It is suggested that defects in BRCA1/2 genes contribute to a high mutation load and high immunogenicity, which modulates immune microenvironment. At the same time, it was shown that BRCA1/2-associated breast cancer tumors do not belong to the category of immunoactive ones. In addition, there is evidence of the additional evaluation of TP53 mutation in these tumors and disruption of the cell death process, which can also be a factor of resistance to therapy. 40 patients with breast cancer with BRCA1 mutations 185delAG, 4153delA, 5382insC, 3819DELGTAA, 3875delGTCT, 300T>G, 2080delA and BRCA2 (6174delT)) determined by real-time polymerase chain reaction (PCR) were included. Immunohistochemical study was performed using monoclonal antibodies to Ki67, p53, CD8, CD4, CD68, CD163, BCL2. In the presented study patients with BRCA1-associated breast cancer, had high low tumor CD4/CD8 ratio in 50% of cases, which characterizes an immunosuppressive microenvironment. Mutation BRCA1 5382insC is associated with high level of CD4+ T-lymphocytes (p˂0.05), G3 is associated with a low CD4/CD8 ratio (p=0.056) and a low levels of CD68+M1, CD163+M2-macrophages (p=0,022, p=0.002); T1 correlates with high levels of CD8+ TILs (p=0.038) and high levels of CD163 (p=0.033). High Ki-67 is associated with a lack of BCL2 expression (p=0.04). Negative expression of BCL2 occurred in 52.5% of cases. A high level of p53 expression is described as the main type of expression in these tumors, suggesting a combination of TB53 and BRCA1 mutations and a violation of the mechanism of cell death in these patients. The immunosuppressive type of microenvironment prevails in the tumors of breast cancer patients with hereditary mutations in the BRCA1 gene, where a violation of the mechanism of cell death in BRCA1-deficient breast cancer was revealed. The main directions of future therapy of these tumors may be modification of the immune microenvironment and activation of the mechanisms of cell death.

The influence of erythropoietin on apoptosis and fibrosis in the early phase of chronic pancreatitis in rats.

IntroductionChronic pancreatitis (CP) is a continuing, inflammatory process of the pancreas, characterised by irreversible morphological changes. The identification of pancreatic stellate cells resulted in the development of research on the pathogenesis of CP. Erythropoietin (Epo) regulates the interaction between apoptosis and inflammation of the brain, kidney, and heart muscle. Erythropoietin receptors were also found in the pancreas, in particular on the islet cells. Our objective was to evaluate the influence of Epo on fibrosis and apoptosis in experimental CP.Material and methodsThe experiments were performed on 48 male Wistar rats (250–350 g). The animals were divided into six equal groups (I – control, II – chronic cerulein – induced pancreatitis, III – 1 ml of Epo sc, IV – 0.5 ml of Epo sc, V – CP treated with 1 ml Epo, VI – CP treated with 0.5 ml Epo). The blood for gelatinases and pancreata for the morphological examinations and immunohistochemistry were collected.ResultsA slight reduction of interstitial oedema and less severe fibrosis were noticed in the groups treated with Epo. Reduced expression of caspase-3 and α-actin, and a lack of Bcl-2 expression were observed in areas with inflammation. There was no expression of caspase-9 observed in all groups. There were no statistically significant differences between the groups in the activity of gelatinases.ConclusionsErythropoietin seems to have the effect of reducing fibrosis and apoptosis in an experimental model of CP.

Lack of Bcl-2 expression in feline follicular lymphomas.

Bcl-2, an anti-apoptotic protein, is commonly overexpressed in follicular lymphomas in humans. This is usually the result of a chromosomal translocation that transposes the Bcl-2 gene into the immunoglobulin gene locus. The immunohistochemical assessment of this overexpression can be used as a tool for the differentiation of follicular lymphoma and follicular hyperplasia. In cats, little information about the expression of Bcl-2 in follicular lymphoma exists. We investigated 18 follicular lymphomas histologically and immunohistochemically for the expression of Bcl-2, CD3, CD45R, and feline leukemia virus. Clonality was assessed by PCR for antigen receptor gene rearrangements. Although the histology resembled that of their human counterparts, diffuse expression of Bcl-2 within the follicles of the feline lymphomas, as seen in human cases, was not present. Only single cells within the follicles, comparable to the reactive controls, were positive for Bcl-2 expression. The mean survival time of 4.6 y confirmed the indolent character of the tumor. None of the clinical parameters assessed were statistically significant predictors of survival. Furthermore, a statistically significant difference in survival of animals with or without anti-neoplastic therapy was also not demonstrable.

Apoptotic Blocks in Primary Non-Hodgkin B-Cell Lymphomas Identified By BH3 Profiling

Background: The anti-apoptotic protein BCL2 is expressed in most non-Hodgkin lymphomas (NHL) including chronic lymphocytic leukemias (CLL), mantle cell lymphomas (MCL), follicular lymphomas (FL), diffuse large B cell lymphomas (DLBCL) and marginal zone lymphomas (MZL). BCL2 expression is associated with an inferior survival when co-expressed with MYC, an oncogene that induces cellular proliferation, especially in high-grade lymphomas with translocations in MYC and BCL2(HGBL-DH). Venetoclax, a selective BCL2 inhibitor, is effective in CLL, but has modest clinical activity in other NHLs. CLL has been shown to be “primed” and BCL2-dependent, predicting for a favorable response to venetoclax using BH3 profiling. This technique measures mitochondrial outer membrane depolarization (MOMP) after exposure to synthetic pro-apoptotic BH3 peptides that have different affinities to anti-apoptotic proteins BCL2, BCLXL, MCL1 and BCLW. Cells with functional BAX/BAK undergo MOMP after exposure to pro-apoptotic proteins BIM/BID. “Primed” cells undergo MOMP after exposure to BIM, BID and PUMA, where the latter inhibits all anti-apoptotic proteins. Thus, inhibiting the anti-apoptotic proteins would initiate apoptosis. We hypothesized that BH3 profiling of NHLs could help us understand the mechanisms of venetoclax-resistance in BCL2+ NHL. This has not yet been performed due to the rarity of samples archived as a viable cell suspension.Methods: We performed BH3 profiling on 138 B-cell NHLs (36 CLL/SLL, 42 FL, 38 DLBCL, 10 HGBL-DH, 13 MCL, and 3 MZLs) and 34 controls (14 tonsils and 20 peripheral blood mononuclear cells). NHLs were taken prior to (n=107) or after therapy (n= 31). A T-test and ANOVA were performed to determine the differences in MOMP between NHLs subtypes and normal B cells. We also profiled HGBL-DH cell lines to determine if chemotherapy (cyclophosphamide, doxorubicin, vincristine, dexamethasone and bendamustine) could potentiate venetoclax-responses. We measured the levels of anti-apoptotic proteins and MYC by immunoblot, speculating that proteins with short half-lives (MYC and MCL1) may fall upon cell-cycle arrest.Results: Normal B and T lymphocytes were primed and depended on BCL2, MCL1 and BCL-XL. NHLs were primed in 85% (121/138) of cases, 5% (6/138) were unprimed (no PUMA response) and 10% (14/138) were incompetent to undergo MOMP (no BIM/BID response), indicating dysfunctional BAX/BAK. The latter apoptotic defect was mainly seen in DLBCL (10/14). CLL and HGBL-DH had significantly higher venetoclax responses than DLBCL, MCL and FL (all p <0.05). Responses to venetoclax were only observed in samples that expressed BCL2 protein. In BCL2+ NHLs, increased MOMP was achieved by combining venetoclax with the MCL1-specific inhibitor MS1. Inhibiting BCLXL or BCLW did not increase MOMP, suggesting that MCL1 inhibits apoptosis in the absence of BCL2. Profiling HGBL-DH cell lines OCI-Ly8 and Toledo after chemotherapy, only doxorubicin and vincristine increased priming (PUMA response), decreased the levels of MYC and MCL1 proteins and potentiated venetoclax-induced apoptosis (all p <0.05).Conclusion: CLL and HGBL-DH were the most BCL2-dependent NHLs and had the highest responses to venetoclax. Venetoclax-resistance in our NHLs was due to the presence of MCL1, the lack of BCL2 expression and dysfunctional BAX/BAK. In the former scenario, resistance may be overcome by combining venetoclax with vincristine and doxorubicin, two drugs used in standard RCHOP. DisclosuresJohnson:Seattle Genetics: Honoraria; Merck: Consultancy, Honoraria; AbbVie Inc.: Consultancy, Honoraria, Research Funding; Lundbeck: Consultancy, Honoraria, Other: travel, Research Funding; Bristol-Myers Squibb: Consultancy, Honoraria; Roche: Consultancy, Honoraria, Other: travel, Research Funding. Steidl:Juno Therapeutics: Consultancy; Roche: Consultancy; Bristol-Myers Squibb: Research Funding; Tioma: Research Funding; Seattle Genetics: Consultancy; Nanostring: Patents & Royalties: patent holding. Scott:Janssen: Research Funding; NanoString: Patents & Royalties: Named Inventor on a patent licensed to NanoString Technologies, Research Funding; Celgene: Consultancy, Honoraria; Roche: Research Funding. Letai:Flash Therapeutics: Equity Ownership; Vivid Biosciences: Equity Ownership; AbbVie: Consultancy, Other: Lab research report; Novartis: Consultancy, Other: Lab research report; AstraZeneca: Consultancy, Other: Lab research report.

Granular Cell Basal Cell Carcinoma

Granular cell basal cell carcinoma (BCC), characterized by large cells with abundant eosinophilic cytoplasm and conspicuous granularity, is an extremely rare variant of BCC with only 14 cases reported in the literature to date. A case of an 82-year-old man with clinically suspected BCC of the face is presented. Microscopic examination demonstrated characteristic morphologic and immunophenotypic features of granular cell BCC, as well as novel expression of p16 and lack of bcl-2 expression, both of which are previously undescribed in granular BCCs in the literature to date. Although very rare, this entity is important to include in the differential of any nodular cutaneous neoplasm with granular features. The rarity of this lesion makes immunohistochemistry especially helpful. As in other BCCs, granular cell BCCs typically strongly express Ber-EP4 and cytokeratins. The granular BCCs are characteristically periodic acid-Schiff positive, but show no expression of S100 protein. The rarity of the granular cell BCC and the aggressive biological behavior of the entities that may share similar histologic features make arriving at the correct diagnosis paramount to appropriate clinical management. The fifteenth case of granular cell BCC with subsequent review of the literature is reported, with particular focus on the immunohistochemical characteristics.

Utility of BCL2, PD1, and CD25 Immunohistochemical Expression in the Diagnosis of T-cell Lymphomas

T-cell lymphomas (TCLs) are a heterogenous group of diseases that show histologic and immunophenotypic features overlapping with reactive lymphoid proliferations and often require the use of ancillary testing for accurate diagnosis. The oncoprotein, bcl-2, is expressed in various types of lymphoma. At present, expression of this protein is useful for distinguishing several B-cell lymphomas. Although there are some anecdotal reports that the lack of bcl-2 expression by T cells might also be a useful marker for the diagnosis of TCL, there are no focused studies to address this hypothesis. Another antigen with value in TCL diagnosis is programmed death-1 (PD-1), a marker of follicular helper T cells, which has been reported to be sensitive in the detection of angioimmunoblastic TCL and peripheral T-cell lymphoma, unclassified. However, several reports have also shown that PD-1-positive cells may be increased in a number of settings other than TCL, including reactive and atypical lymphadenopathies. Finally, lymphoma cells express a variety of cytokine receptors and signaling molecules that are current or potential targets for immunomodulatory therapy. One such target is the interleukin (IL)-2 receptor (CD25), which is acted on by denileukin diftitox/ONTAK, a recombinant diphtheria toxin-IL-2 fusion protein. Selection of suitable patients for therapy often includes pretreatment assessment of CD25 expression in tumor cells. In order to further assess the diagnostic and therapeutic utility of these antigens, we compared the expression of the CD25, PD-1, and bcl-2 in 119 cases of T-cell non-Hodgkin lymphoma using immunohistochemical techniques applied to routinely processed and paraffin-embedded tissues. We show that lack of expression of bcl-2 was observed in 52% cases of TCL and may aid in identification of neoplastic T-cell populations. In combination, bcl-2, CD25, and PD-1 provide diagnostic utility and may aid in selecting appropriate patients for immunomodulatory therapy.

Osteoblastoma of the mandible: a case report with immunohistochemical evaluation

Background: Osteoblastoma, a rare osteoblastic tumor, constitutes approximately 1% of all primary bone tu- mors. Aim: The purpose of this report is to present an osteoblastoma of the mandible, with particular emphasis on the differential diagnosis of this rare tu- mor. Methods-Results: the lesion showed osteoblast- like cells, stromal cells, osteoclast-like cells and inter- lacing trabeculae of osteoid. Because of the high cel- lularity of the tumor, immunohistochemical analysis was performed using the cell proliferation marker Ki-67, the p53 and the anti-apoptotic protein Bcl-2. The lesion demonstrated low to moderate prolifera- tive activity and no immunoreactivity was detected for p53. An interesting finding was the Bcl-2 expres- sion by the multinucleated osteoclast-like giant cells, in contrast to the lack of Bcl-2 expression from osteo- blast-like and stromal tumor cells. Conclusion: To our knowledge the Bcl-2 expression has not yet been in- vestigated in benign or malignant osteoblastic tumors. This finding may be related to the molecular mecha- nisms regulating the apoptosis of osteoclast-like tu- mor cells or their function.

Down-regulation of the miRNA master regulators Drosha and Dicer is associated with specific subgroups of breast cancer

Down-regulation of Drosha and Dicer has been suggested to be of prognostic value in some cancers. The aims of our study were to investigate the down-regulation of Drosha and Dicer in breast cancers and its associations with clinicopathological features, molecular subtypes and outcome. Drosha and Dicer expression was assessed with real-time RT-PCR in 245 patients with breast cancer receiving adjuvant anthracycline-based chemotherapy and compared to expression levels of normal breast tissue. Drosha down-regulation was observed in 18% of cases and was associated with high grade, high Ki-67, lack of Bcl2 expression, HER2 over-expression and gene amplification and TOPO2A gene amplification. Dicer down-regulation was found in 46% of cases and was associated with lack of expression of ER, PR and Bcl2 and with high grade, high Ki-67, triple-negative and basal-like phenotypes. Drosha and Dicer were concurrently down-regulated in 15% of cases and significantly associated with high grade and high Ki-67 index. No significant associations between down-regulation of Drosha and/or Dicer and outcome were observed. Our results suggest that down-regulation of Drosha and/or Dicer is not robustly associated with the outcome of breast cancer patients treated with adjuvant anthracycline-based chemotherapy but preferentially observed in distinct subgroups of breast cancer.

Lack of Bcl-2 Expression in Follicular Lymphoma May Be Caused by Mutations in the BCL2 Gene or by Absence of the t(14;18) Translocation

Lack of Bcl-2 Expression in Follicular Lymphoma May Be Caused by Mutations in the BCL2 Gene or by Absence of the t(14;18) Translocation

Recurrence and survival predictive value of phenotypic expression of Bcl-2 varies with tumor stage of colorectal adenocarcinoma

Although decreased or lack of expression of Bcl-2 has been correlated with advanced tumor stage and shortened patient survival in colorectal cancer (CRC), its value in predicting the recurrence has not been well explored. Therefore, we assessed the usefulness of phenotypic expression of Bcl-2 in non-Hispanic Caucasian patients with CRCs in identifying risk of recurrence. Archival tissues of 92 Stage II and 66 Stage III primary CRCs were evaluated for immunohistochemical expression of Bcl-2. None of these patients received either pre- or post-surgical adjuvant therapies. Kaplan-Meier and Cox proportional hazards methods were used to estimate the rates of recurrence and survival according to Bcl-2 expression. Decreased expression of Bcl-2 was associated with an increased rate of recurrence in patients with Stage II CRCs (5-year log-rank test P=0.0015; Hazard Ratio (HR)=3.90, 95%C.I.:1.55-9.77) but not with Stage III CRCs (5-year log-rank test P=0.6058; HR=1.07, 95%C.I.:0.47-2.45) after adjusting for other demographic and clinicopathological features. Furthermore, decreased expression of Bcl-2 was an indicator of short survival in patients with Stage II CRCs but not with Stage III CRCs. Thus, decreased or lack of Bcl-2 expression in primary CRCs may serve as a molecular biomarker of high risk of recurrence for Caucasian patients with Stage II CRCs. These findings might be useful in identifying biologically aggressive phenotypes of Stage II CRCs, and may aid the oncologist in designing maximally appropriate therapeutic regimens.

Bcl-2 as Prognostic Factor in Head and Neck Squamous Cell Carcinoma

A series of 66 cases of oral squamous cell carcinoma (OSCC) was retrospectively analyzed by immunohisto-chemistry for bcl-2 expression to verify its predictive value for clinical outcome in patients with OSCC. After grouping for bcl-2 expression, OSCCs were statistically analyzed for the variables age, gender, histological grading (G), TNM, staging, recurrence, and overall survival rate. Univariate and multivariate (Cox regression) analyses were performed. Thirty-six OSCC (54.5%) showed expression for bcl-2, whereas 30 (44.5%) were negative. No statistical association was found between bcl-2 expression and any variables considered at baseline. Overall disease-specific survival rate at 72 months was 51%, independently from the extent of the tumor. In terms of prognostic significance, the bcl-2-positive group showed more than 60% survival at 72 months whereas the bcl-2-negative group showed none. An independent association of bcl-2 expression was found with an improved overall survival rate (p = 0.048), although grading and staging were established to be the best baseline markers of prognosis. On the basis of these results, it is possible to suggest bcl-2 as an early marker of prognosis: lack of bcl-2 expression could constitute a hallmark of aggressive biological behavior in OSCC.

Expression of BAG-1 and BcL-2 Proteins Before and After Neoadjuvant Chemotherapy of Locally Advanced Breast Cancer

It has been suggested that expression of anti-apoptotic proteins such as Bcl-2 or BAG-1 may confer cellular resistance to chemotherapy. A corollary of this hypothesis is that expression of these proteins may predict clinical response to treatment and that Bcl-2– or BAG-1–positive cells may selectively be enriched in postchemotherapy tissue specimens. The goal of this exploratory pilot study was to assess these two predictions by using immunohistochemistry in 29 paired pre- and postchemotherapy breast tissue specimens obtained from patients who underwent preoperative doxorubicin-based chemotherapy. All breast cancers expressed BAG-1 protein, and, in individual tumors, 40–100% of neoplastic cells stained positive for this protein. Homogenous cytoplasmic staining was typically observed, though neoplastic cells also showed nuclear staining in many specimens. We found no correlation between prechemotherapy expression of BAG-1 and subsequent pathological response to cytotoxic therapy. Paired pre- and posttreatment specimens showed similar levels of BAG-1 expression when residual tumor could be assessed. Bcl-2 was expressed in 55% of cancers and was localized to the cytoplasm. Absence of Bcl-2 expression in prechemotherapy specimens was associated with more frequent complete pathological response (58% vs. 20%; p = 0.04). However, similar to BAG-1, no difference between pre- and posttherapy expression of Bcl-2 was observed in neoplastic cells in paired tissue specimens. These observations suggest that BAG-1 contributes an important cellular function to breast epithelial cells, which is reflected by its ubiquitous expression in these tissues. However, it does not appear to determine response to doxorubicin-based chemotherapy. In contrast, lack of Bcl-2 expression was associated with a higher probability of complete pathological response to doxorubicin-based chemotherapy.

Bcl-2 expression is a prognostic factor in the subgroups of patients with colorectal cancer

The prognostic significance of Bcl-2 expression in colorectal cancer has been intensively studied, however, the results were controversial in the whole group of colorectal cancer patients. We proposed that one of the main reasons for such controversial results may be that Bcl-2 played variable roles in the subgroup of patients. We, therefore, investigated the prognostic importance of Bcl-2 expression by using immunohistochemistry in the various subgroups of 147 patients with colorectal cancer. Among these tumours, 85 (58%) expressed Bcl-2 protein and 62 (42%) were negative. Bcl-2 expression was positively related to DCC expression (p=0.0002). Survival analyses in the subgroups of the patients showed that lack of Bcl-2 expression was related to a worse prognosis in the male patients (p=0.02) but not in female patients (p=0.53), in the patients with DNA diploid tumours (p=0.005) not in the patients with non-diploid tumours (p=0.46), and in the patients with ras negative tumours (p=0.01) not in the patients with ras positive tumours (p=0.25). Bcl-2 expression was not related to prognosis in the total group of the patients (p=0.20). In conclusion, Bcl-2 protein may play variable prognostic roles in the subgroups of the patients with colorectal cancer. Analysis of Bcl-2 expression in the tumour may be of value in predicting prognosis and therapeutic response.

Immunohistochemically Detectable Bcl-2 Expression in Metastatic Melanoma: Association with Survival and Treatment Response

Objective: The antiapoptotic protein Bcl-2 is supposed to influence the treatment responsiveness of different malignancies. In the present study the prognostic and predictive significance of Bcl-2 expression for survival and response to an administered therapy was explored in patients with metastatic melanoma. Also, the correlation between Bcl-2 expression and proliferation activity of tumor cells was defined to examine the regulatory role of Bcl-2 in proliferation. Materials and Methods: Sixty metastatic melanomas obtained from patients treated with chemoimmunotherapy were examined by immunohistochemistry with anti-Bcl-2 and anti-Ki-67 (MIB-1) antibodies. Proliferation activity was expressed in percentages as MIB-1 index. Results: The presence of Bcl-2 immunoreactivity was associated with a significantly lower MIB-1 index (p = 0.016), and a longer disease-free survival (p = 0.004). The lack of Bcl-2 expression was related to a higher response rate to therapy in comparison to a diffuse and focal pattern of Bcl-2 expression (p = 0.017). Although the presence of Bcl-2 immunoreactivity as such did not correlate with survival after the initiation of chemoimmunotherapy, the focal Bcl-2 expression pattern was strongly associated with a worse prognosis compared to a diffuse expression or a lack of Bcl-2 staining (p < 0.0001). Conclusions: Our results support the role of Bcl-2 in the regulation of cell proliferation and suggest that an increase of metastatic potential and progression of malignant melanoma is associated with a loss of Bcl-2 expression. The lack of Bcl-2 expression could be a predictor of the response to chemoimmunotherapy, whereas the Bcl-2 expression pattern, possibly indicating the heterogeneity of the tumors, might be a potential prognostic factor for survival after the initiation of therapy.

Expression of CD40, CD44, bcl-2 antigens and rate of cell proliferation on fine needle aspirates from metastatic melanoma.

The clinical behaviour of melanoma is often unpredictable using clinical and histological criteria. Tumour cell markers related to cell cycle regulation, apoptosis, cell-cell interactions and cell proliferation might improve the possibility of predicting the clinical course of melanoma. The aim of the present study was to refine prognostic criteria by an immunocytochemical investigation of CD44, CD40, bcl-2 antigens and cell proliferation in tumour cells aspirated from metastases of malignant melanoma. CD40 is a cell surface receptor shown to be expressed by lymphomas as well as carcinomas, and is thought to play a central role in the process of tumour progression. CD44 is a transmembrane glycoprotein, which is involved in growth signal transmission of importance in the binding of tumour cells to endothelium, cell migration and enhancement of cell motility, which makes it of interest to study in relation to the metastasizing capacity of tumours. The bcl-2 protein is active in the process of programmed cell death (apoptosis) as an antiapoptotic agent and its expression may reflect tumour progression. Mean/median percentages of tumour cell positivity were 8.5/3.0 for CD40, 76.1/86.3 for CD44 and 7.4/3.3 for bcl-2. A significant correlation was observed between expression of apoptosis-associated bcl-2 antigen and overall survival (r = 0.33). The CD44 positive cell fraction was higher in patients with short overall survival than those with long survival but this difference was not statistically significant. The expression of CD40 did not correlate with overall survival. The mean/median proliferation fraction assessed by MIB-1 monoclonal antibody was 25.8/23.9 and showed a significant correlation with survival after diagnosis of melanoma metastasis (r = 0.32). Lack of bcl-2 expression and a high proportion of tumour cells expressing Ki-67 antigen are predictors of poor prognosis that are independent of the traditionally accepted Breslow's thickness of the primary melanomas.

Prognostic significance of immunohistochemically localized biomarkers in stage II and stage III breast cancer: a multivariate analysis.

The aim was to investigate the expression of a panel of biomarkers such as prolactin (PRL), p53, Bcl-2, c-erb B2, Ki-67, CD44, and factor VIII-related antigen (FVIII-RA) in primary tumors of stage II and stage III breast cancer and its correlation with disease prognostication. The streptavidin-biotin peroxidase complex technique was used for the detection of these antigens. Cytoplasmic staining pattern was observed for PRL, Bcl-2, and Ki-67. Staining pattern for p53 was nuclear. Membranous and/or cytoplasmic staining was noted for c-erb B2 and CD44. Microvessel staining was noted for FVIII-RA. Of the 93 primary breast tumors analyzed, positivity for PRL was noted in 82%, for p53 in 56%, for Bcl-2 in 73%, for c-erb B2 in 68%, and for Ki-67 and CD44 in 78% each. The microvessel count (MVC) for FVIII-RA ranged from 0.0 to 29.0, with a median of 6.0, which was used as a cutoff. MVC > or = 6.0 was noted in 51% of breast tumors. With increasing tumor size, the higher frequency of positivity of MVC > or = 6.0 (P = .0001), CD44 (P = .001), PRL (P = .002), and c-erb B2 (P = .008), and higher frequency of Bcl-2 negativity (P = .033), was noted. In stage III patients, a higher positivity of the following biomarkers was noted, compared with stage II patients: MVC > or = 6.0 (P = .0004), PRL (P = .0002), c-erb B2 (P = .001), and CD44 (P = .005). Further, Bcl-2 positivity was significantly lower in patients with stage III disease compared with those with stage II disease (P = .024). In patients with nodal involvement, the frequency of c-erb B2 (P = .006), MVC > or = 6.0 (P = .011), and PRL (P = .032) was higher than in those without nodal involvement. Moreover, in these patients, with the increase in the number of involved lymph nodes, there was a significant increase in frequency of CD44+ (P = .0004) and PRL+ (P = .013) tumors. Abnormal expression of one biomarker was seen in 7% of tumors, of two biomarkers in 4%, of three in 15%, of four in 19%, of five in 28%, of six in 20%, and of all seven biomarkers in 7% of tumors. The frequency of an increasing number of biomarkers coexpressed was higher in stage III patients compared with stage II patients (P = .00003). In the total number of patients (n = 93), tumors with Bcl-2 negativity (P = .00001), MVC > or = 6.0 (P = .001), PRL positivity (P = .02), and CD44 positivity (P = .034) had a significantly poorer overall survival (OS) compared with their respective counterparts. In stage II patients (n = 40), only p53 expression was significantly associated with reduced relapse-free survival (P = .009) and OS (P = .040). In multivariate analysis, p53 expression was an independent prognostic factor that influenced relapse-free survival (P = .034) of stage II breast cancer patients. However, it failed to attain statistical significance for OS. In stage III patients (n = 53), tumors with Bcl-2 negativity (P = .0005) and MVC > or = 6.0 (P = .039) had a significantly poorer OS compared with their respective counterparts. In multivariate analysis of stage III patients, Bcl-2 was the only independent prognostic factor (P = .001) for predicting OS. There was a significant association between coexpression of the biomarkers and OS (P = .001). The OS rates decreased with the increase in number of abnormally expressed biomarkers. p53 expression in primary tumors was an independent prognostic factor that influenced relapse-free survival in patients with stage II disease. In stage III patients, lack of Bcl-2 expression was independently associated with a poor prognosis and, thus, may be an indicator of aggressive phenotype.

Clinical Significance of the t(14;18) And BCL2 Overexpression in Follicular Large Cell Lymphoma

Follicular large cell lymphoma (FLCL) is an aggressive disease that responds to anthracy-cline-containing chemotherapy much like diffuse large B-cell lymphoma (DLBCL). Since the t(14; 18) and/or bcl2 protein expression are less common in FLCL than in its low-grade counterparts, we sought to determine whether these features were predictive of survival as in DLBCL. We studied 50 patients with FLCL who were treated with curative intent. The t(14; 18) was found by cytogenetic analysis in 56% of the patients and bcl2 protein was expressed by the tumor cells in 73%, but neither was predictive of survival. However, abnormalities of chromosome 17p and the presence of trisomy 21 were adverse predictors of survival, as were a number of clinical features. We conclude that neither the absence of the t(14; 18) nor the lack of bcl2 expression explain the good response of a subset of patients with FLCL to curative therapy.

Bcl-2 EXPRESSION IN PLEURAL AND EXTRAPLEURAL SOLITARY FIBROUS TUMOURS

This study evaluated the immunoreactivity for bcl-2, a molecule involved in the control of programmed cell death, in cases of pleural (14) and extrapleural (2) solitary fibrous tumour (SFT), malignant mesotheliomas of different histological types, and a variety of extrapleural CD34-positive and CD34-negative spindle-cell tumours. In all SFTs, strong and diffuse immunostaining was demonstrated with anti-bel-2 antibody, sharply contrasting with the complete lack of staining observed in all mesotheliomas. The specificity of immunodetection of bcl-2 in SFT was confirmed by immunoblot analysis, showing a band consistent with the bcl-2 protein. At extrapleural locations, strong bcl-2 immunoreactivity was observed in Schwannoma (2/3 cases), synovial sarcoma (4/4 cases), and all cases of CD34-positive gastrointestinal stromal tumour (GIST; 10/10 cases). Most sarcomas were bcl-2-negative. Lack of bcl-2 expression was demonstrated in tumours which can pose problems in the differential diagnosis of SFT and can exhibit haemangiopericytoma-like features, including haemangiopericytoma (3 cases), dermatofibrosarcoma protuberans (16 cases), and deep-seated fibrous histiocytoma (3 cases). The constitutive expression of bcl-2 in SFT widens the spectrum of available markers for these tumours, providing a useful adjunct to their differential diagnosis in difficult cases at pleural and extrapleural sites, and contributing to the understanding of their histogenesis and molecular pathogenesis.

BCL-2 But Not Its Epstein-Barr Virus-Encoded Homologue, BHRF1, Is Commonly Expressed in Posttransplantation Lymphoproliferative Disorders

Posttransplantation lymphoproliferative disease (PTLD) is virtually always associated with Epstein-Barr virus (EBV) infection. BCL-2 and other proteins that confer resistance to apoptosis have been implicated in the pathogenesis of a variety of malignancies including lymphomas. One EBV protein, BHRF1, is a homologue of BCL-2, whereas another, the latency membrane protein 1 (LMP-1), upregulates BCL-2 expression in vitro. In the present study, we used immunohistochemistry to study the expression of these viral and cellular proteins as well as a variety of other EBV-encoded proteins in PTLD. BHRF1 was not detected in any PTLD specimen, whereas BCL-2 was shown in 12 of 17 lesions examined. With one exception, all LMP1-positive cases also expressed BCL-2 and the absence of LMP1 was always associated with a lack of BCL-2 expression. The results do not support a role for the EBV homologue of BCL-2 in PTLD, but they do support a role for viral induction of BCL-2 expression.