Dishevelled family proteins in serous ovarian carcinomas: a clinicopathologic and molecular study

Dishevelled family proteins (DVL1, DVL2, and DVL3) are cytoplasmic mediators involved in canonical and non-canonical Wnt signaling that are important for embryonic development. Since Wnt signaling promotes cell proliferation and invasion, its increased activation is associated with cancer development as well. To get deeper insight into the behavior of Dishevelled proteins in cancer, we studied their expression in serous ovarian carcinomas [both low- (LGSC) and high-grade (HGSC)], and HGSC cell lines OVCAR5, OVCAR8, and OVSAHO. DVL protein expression in serous ovarian carcinomas tissues was analyzed using immunohistochemistry, while DVL protein and mRNA expressions in HGSC cell lines were analyzed using Western blot and quantitative real-time PCR. DVL1 protein expression was significantly higher in LGSC compared with normal ovarian tissue, while DVL3 was overexpressed in both LGSC and HGSC. DVL2 and DVL3 protein expression was higher in HGSC cell lines when compared with normal control cell line FNE1, while DVL1, DVL2, and DVL3 mRNA expression was significantly increased only in OVSAHO cell line. Survival analysis revealed no significant impact of DVL proteins on patients’ outcome. Our data show an active involvement of Dishevelled family proteins in serous ovarian carcinomas. Further studies should confirm the clinical relevance of these observations.     

Elevated expression of E-cadherin and alpha-, beta-, and gamma-catenins in metastatic lesions compared with primary epithelial ovarian carcinomas

E-cadherin and catenins play key roles in cell adhesion and motility. Little is known about the changes in expression of these molecules in the progression of ovarian carcinomas. In the present study, the immunohistochemical expression of E-cadherin and alpha-, beta-, and gamma-catenins was examined in 77 cases of ovarian carcinoma. In addition, the expression of these molecules was evaluated in 26 matched pairs of primary and metastatic lesions of advanced ovarian carcinomas. Of the 77 primary lesions, positive staining for E-cadherin and alpha-, beta-, and gamma-catenin was observed in 75 (97%), 63 (82%), 71 (92%) and 57 (74%) cases, respectively. Positivity for E-cadherin and alpha-, beta-, and gamma-catenin was significantly decreased in stage III and IV tumors compared with stage I and II tumors, suggesting that expression of the cadherin-catenin complex is reduced with the advancing stages of a tumor. Interestingly, expression of E-cadherin and alpha-, beta-, and gamma-catenin in the lesions of peritoneal dissemination was significantly increased compared with the primary lesions. These findings suggest that expression of the cadherin-catenin complex changes markedly and that reexpression may occur during the peritoneal dissemination of ovarian carcinoma cells.     

Thymidine phosphorylase expression results in a decrease in apoptosis and increase in intratumoral microvessel density in human gastric carcinomas

Thymidine phosphorylase (dThdPase) / platelet- derived endothelial cell growth factor (PD-ECGF) is expressed at higher levels in a variety of human carcinomas than in adjacent normal tissue. The higher expression is associated with an increase in intratumoral microvessel density (IMVD) and an unfavorable patient prognosis. This study examined the role of dThdPase in apoptosis, IMVD and p53 expression in human gastric carcinomas. dThdPase expression was noted in 12 (35.3%) of 34 early carcinomas, and in 20 (55.6%) of 36 advanced carcinomas. At least 10 areas consisting of carcinoma cells with diffuse dThdPase expression from the 32 dThdPase-positive tumors (category I), and 10 areas without dThdPase expression from the 38 negative tumors (category II) were selected from each case. For early gastric carcinoma, the mean IMVD was 88.8±19.4 in category I and 61.4±17.3 in category II carcinomas, while for advanced gastric carcinoma, the mean IMVD was 98.8±21.0 in category I and 76.0±27.1 in category II carcinomas. The mean IMVD was significantly higher in category I than in category II tumors (P<0.05). The mean apoptotic index (AI: percentage of apoptotic cells) was 1.95±1.30 in category I, and 3.76±1.49 in category II carcinomas for early gastric carcinoma, and 1.51±0.98 in category I and 2.14±0.66 in category II carcinomas for advanced gastric carcinoma, the value of the mean AI being significantly (P<0.05) higher in dThdPase- negative tumors (category II) than in the positive tu-mors (category I), regardless of tumor stage or histological type. There was a significant inverse correlation (P<0.001) between AI and IMVD. These results indicate that dThdPase expression is associated with both an increase in intratumoral microvessels and a decrease in apoptosis in human gastric carcinomas. Received: 8 November 1999 / Accepted: 20 January 2000     

Microvessel density,endothelial-cell proliferation and carbonic anhydrase IX expression in haematological malignancies,bone-marrow metastases and monoclonal gammopathy of undetermined significance

The aim of the study was to compare the angiogenic status, potential qualitative differences in microvessels and carbonic anhydrase IX expression in bone-marrow (BM) metastases and different haematological tumours at time of diagnosis. The microvessel density (MVD), endothelial-cell proliferation (ECP) and carbonic anhydrase IX (CA IX) immunoreactivity were determined on 210 trephine biopsies from 57 patients with multiple myeloma (MM), 13 with acute myeloid leukaemia (AML), 48 with chronic myeloproliferative syndrome (CMPS), 26 with chronic lymphocytic leukaemia (CLL), 25 with epithelial BM metastases, 18 with monoclonal gammopathy of undetermined significance (MGUS) and from a control group composed of 23 patients without haematological neoplasm. There was an increased MVD and ECP in epithelial BM metastases, MM, AML, CMPS and in a part of CLL. While an ECP greater than 0 was detected in 72% of MM, 75% of CMPS and 92% of AML, it was invariably observed (100%) in the BM metastases. The absence of ECP together with a MVD comparable with the control group in our MGUS cases supports the view that MGUS is a pre-angiogenic condition. Qualitative differences in microvessels were associated with growth patterns in MM and CLL and were observed between the different entities of CMPS. In one-third of the epithelial BM metastases, there was a focal CA IX immunoreactivity, which was never observed in the haematological diseases.     

Intracytoplasmic lumina and mucinous inclusions in ovarian carcinomas

Intracytoplasmic mucinous inclusions and lumina have been previously described in non-glandular neoplasms such as urothelial carcinoma. We describe their presence in 93% of non-mucinous ovarian carcinomas. They were found in abundance in all 25 cases (100%) of clear cell carcinoma, in 48 of 50 cases (96%) of serous carcinoma and 20 of 25 cases (80%) of endometrioid carcinoma. The degree of the differentiation of the tumour did not influence the number of inclusions or lumina observed. These results suggest that the presence of intracytoplasmic lumina and mucinous inclusions is more widespread than hitherto appreciated. Their presence in an otherwise poorly differentiated metastatic carcinoma might, at the very least, prompt one to consider the ovary as a possible primary site. In addition, an abundance of intracytoplasmic mucinous inclusions and lumina with microcyst formation, in an otherwise poorly differentiated malignant primary ovarian epithelial tumour, might suggest the possibility of a clear cell carcinoma.     

Elevated expression of E-cadherin and α-, β-, and γ-catenins in metastatic lesions compared with primary epithelial ovarian carcinomas

E-cadherin and catenins play key roles in cell adhesion and motility. Little is known about the changes in expression of these molecules in the progression of ovarian carcinomas. In the present study, the immunohistochemical expression of E-cadherin and α-, β-, and γ-catenins was examined in 77 cases of ovarian carcinoma. In addition, the expression of these molecules was evaluated in 26 matched pairs of primary and metastatic lesions of advanced ovarian carcinomas. Of the 77 primary lesions, positive staining for E-cadherin and α-, β-, and γ-catenin was observed in 75 (97%), 63 (82%), 71 (92%) and 57 (74%) cases, respectively. Positivity for E-cadherin and α-, β-, and γ-catenin was significantly decreased in stage III and IV tumors compared with stage I and II tumors, suggesting that expression of the cadherin-catenin complex is reduced with the advancing stages of a tumor. Interestingly, expression of E-cadherin and α-, β-, and γ-catenin in the lesions of peritoneal dissemination was significantly increased compared with the primary lesions. These findings suggest that expression of the cadherin-catenin complex changes markedly and that reexpression may occur during the peritoneal dissemination of ovarian carcinoma cells.     

Prostaglandins E in primary tumor, metastases, and ascitic fluid of patients with ovarian cancer

The content of prostaglandins E (PGE) is compared in primary tumors, metastases, and ascitic fluid of patients with ovarian cancer, in benign tumors, and in neoplastic ovarian disorders. The PGE content in malignant tumors, neoplastic disorders, and benign tumors is higher than in normal tissues. A relationship between the PGE content, on the one hand, and the degree of differentiation and histological variant of malignant tumor, on the other, is revealed. The PGE content in the ascitic fluid of patients increases with the degree of tumor malignancy. The variants of ovarian cancer therapy with preparations modifying the arachidonic acid metabolism are considered. Translated fromByulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 123, No. 1, pp. 83–86, January, 1997     

Microvessel density in human normal and neoplastic parathyroids

Objective Information on angiogenesis in parathyroid pathology is scanty and in particular no data are available in parathyroid carcinomas. The aim of this study was to analyze angiogenesis as microvascular density (MVD) in parathyroid neoplastic progression from normal gland to adenoma and carcinoma. Methods Sections from formalin-fixed, paraffin-embedded specimens of 33 normal parathyroids, 43 sporadic parathyroid adenomas, and 6 parathyroid carcinomas were cut for immunohistochemistry using anti-endothelial marker CD34. MVD was evaluated in each specimen as number microvessels per mm². MVD data were compared with some anatomoclinical parameters as tumor size, serum calcium, and parathyroid hormone (PTH) level. Results All normal parathyroid glands, all carcinomas, and 8 adenomas out of 43 (18%) showed MVD less than 100 microvessels/mm² (median 70.8; 95% Cl 66.9–88.5); in the majority of parathyroid adenomas (n=35; 82%) the number of microvessels/mm² was higher than 100 (median 188.3; 95% Cl 174.9–210.1). In adenomas both preoperative serum intact PTH concentration and the diameters were significantly and inversely related to the microvessel density (r=0.320, p<0.05 and r=0.334, p<0.05, respectively). Conclusions This study shows that in parathyroid adenomas MVD is heterogeneous and negatively related to the endocrine activity (secretory status and tumor size). Therefore, angiogenesis in parathyroid adenomas and carcinomas appears to be an early event, which does not follow a parallel increase in size.     

Microvessel densities and microvascular architecture in colorectal carcinomas and their liver metastases: significant correlation of high microvessel densities with better survival

AIMS: Microvessel densities in cancers have been shown to be a prognostic factor for some types of cancer. For colorectal cancer, however, the situation is far from clear. METHODS: A consecutive series of 173 colorectal carcinomas was investigated, and to these were added 55 liver metastases originating from colorectal cancer. Microvessels were counted in hotspots (factor VIII immunostaining, 0.74 mm2). The capillary architecture was scored according to the degree of order and envelopment of the neoplastic glands. Endothelial proliferation was determined by factor VIII/Ki67 double labelling. RESULTS: Mean microvessel densities were 51.8 for colorectal carcinomas (range 8-140) and 31.9 for liver metastases (range 3-101). Stratification according to stage, depth of infiltration and nodal involvement showed a significant inverse relation with increase. Mean microvessel densities in primaries were significantly higher than in metastases. Kaplan-Meier analysis showed a significantly higher cancer-specific survival for high microvessel densities (median as cut-off) and for a more ordered microvascular architecture. Endothelial proliferation in carcinomas was significantly higher than in normal mucosa. CONCLUSIONS: Contrary to other types of cancer, for colorectal cancer high microvessel densities confer good rather than poor prognosis. We hypothesize that neoangiogenesis, though extant in colorectal cancer, is not rate-limiting in the metastatic cascade.     

Prognostic significance of microvessel density and vascular endothelial growth factor expression in sinonasal carcinomas

The prognostic significance of microvessel density and proliferative activity of the neoplastic cells, evaluated respectively by CD31 and Ki-67 positivity, and immunohistochemical expression of vascular endothelial growth factor (VEGF) was retrospectively investigated in 105 cases of sinonasal carcinoma (80 surgical specimens and 25 biopsies). The most represented histologic types were intestinal-type adenocarcinoma found in 36 patients (34.3%), squamous cell carcinoma (SCC) in 34 (32.4%), mucinous adenocarcinoma (mainly made up of signet-ring cell patterns) in 15 (14.3%), and adenoid cystic carcinoma in 7 (6.7%). Microvessel density values (in vessels per square millimeter), VEGF, and Ki-67 were not dependent on histologic type but were rather correlated to the histologic grading in SCC. Clinical data were available for 92 (87.6%) of 105 patients, with minimum follow-up of 48 months. Most of the patients (81.5%) were at an advanced stage (T3-T4) at diagnosis. The values of all markers were correlated to tumor stage (P = .03). Multivariate analysis showed that both microvessel density and proliferative activity of the neoplastic cells were independent prognostic parameters (mortality hazard ratio, 1.33 and 1.60, respectively). Although VEGF expression was not correlated to prognosis on the whole series (P = .06), it was a powerful prognostic marker when the analysis was restricted to the group of SCCs (hazard ratio, 3.02; 90% confidence interval, 1.58-5.80). These results show that tumor neoangiogenesis, expressed by microvessel density, together with proliferative activity, is a pathologic marker with a strong prognostic impact in sinonasal carcinomas. Therefore, it may be a useful tool in this field so as to carry out therapeutic protocol planning, which may be further enhanced by the adoption of the more recent antiangiogenic molecules.     

p53 and erbB-2 are not associated in matched cases of primary and metastatic ovarian carcinomas

Ovarian cancer has a high mortality rate largely due to the limited number of ovarian carcinomas detected at an early stage. Understanding the molecular changes occurring during the progression of ovarian carcinoma would aid in the development of therapies that may inhibit or target metastasis. Primary and metastatic lesions from 54 and 40 patients with advanced ovarian carcinoma, respectively (including matched primary and metastatic lesions from 30 patients) were evaluated for nuclear accumulation of p53 (clone BP53-12-1) and cytoplasmic and membranous immunostaining of p185 erbB-2 (clone 3B5) by immunohistochemistry. No differences in the immunostaining of p53 and p185erbB-2 (cytoplasm or membrane) were observed between primary and metastatic lesions of the matched cases. Similarly, no differences in the proportion of positive cases of p53 between primary and metastatic lesions of the matched cases was observed. Thus, novel therapies that target p53 or p185erbB-2 can utilize specimens from either primary or metastatic lesions to characterize these targets prior to therapy. Spearman correlations between p53 and p185erbB-2 (cytoplasm or membrane) immunohistochemistry scores were insignificant for the matched cases, all primary lesions, and all metastatic lesions. Also, no significant associations occurred between nuclear accumulation of p53 (positive versus negative) and phenotypic expression of p185erbB-2 (cytoplasm or membrane) immunostaining scores for the matched cases, all primary lesions, and all metastatic lesions. Thus, the nuclear accumulation of p53 and immunostaining of p185erbB-2 in the cytoplasm or on the cellular membranes are independent.     

Increased expression of fascin, motility associated protein, in cell cultures derived from ovarian cancer and in borderline and carcinomatous ovarian tumors

Fascin bundles actin microfilaments within dynamic cellular structures such as microspikes, stress fibers and membrane ruffles. Fascin overexpression induces membrane protrusions and increased cell motility, and is highly expressed in various transformed cells, and in specialized normal cells including neuronal, endothelial and dendritic cells. In breast cancer, fascin expression correlates with high-grade tumors. To investigate whether fascin might be a predictor factor for ovarian cancer progression, eighteen cell cultures derived from ovarian cancer, and thirty four archival paraffin-embedded material of normal versus borderline and carcinomatous ovaries were stained by immunocytochemistry and immunohistochemistry with fascin Mab 55K-2. Overall expression of the fascin protein was found in 50% (9/18) of cell cultures derived from original samples of ovarian tumors. Expression of fascin protein was found in 67% (6/9) of cell cultures derived from patients diagnosed with stage IV disease, and 33% (3/9) of cell cultures from patients diagnosed with stage II/III. There was no clear relationship between fascin expression and histologic types, tumor grade, or DNA ploidy. However, 75% of cell cultures, which developed into a xenograft after intraperitoneal inoculation, showed fascin expression, while 86% of non-tumorigenic cell cultures did not show fascin expression. Expression of fascin in these established ovarian tumor cell cultures was significantly associated with the ability for these cells to grow intraperitoneally (P<0.05). Furthermore, fascin was never expressed in normal epithelial ovarian tissues, but was present in all pathologic ovaries. Both diffuse and focal patterns were observed in borderline ovarian tumors (67% and 33%), advanced primary ovarian cancer (67% and 33%) and metastatic ovarian cancer (89% and 11%). Therefore, our data suggest that fascin could serve as a prognostic factor for abnormal ovarian epithelial pathology and could be a novel target for the treatment of ovarian cancer.     

Angiogenesis in salivary carcinomas with and without myoepithelial differentiation

To investigate whether salivary carcinomas with and without myoepithelial differentiation could present differences regarding degree of angiogenesis, we compared tumor vascularization between adenoid cystic (31 cases) and epithelial-myoepithelial carcinomas (14) versus mucoepidermoid (37) carcinoma. The expression of peroxiredoxin I was also studied to verify the potential relationship between cellular metabolism and microvascular density. Microvascular density for CD34 and CD105 were significantly lower in carcinomas with myoepithelial differentiation. However, no correlation was found between degree of angiogenesis and amounts of myoepithelial cells. High-grade peroxiredoxin I expression was found in 73.7% of mucoepidermoid carcinomas, whereas 85.1% of carcinomas with myoepithelial differentiation presented low-grade expression. In conclusion, carcinomas with myoepithelial differentiation, regardless of the amounts of myoepithelial cells, are associated to a significantly lower vascular density. The reasons for this lower angiogenic activity remain to be determined but could be related to metabolic characteristics of the cancer cells.     

WAVEs亚家族在卵巢癌中的表达及临床意义

目的 探讨WAVEs亚家族在卵巢癌(oc)中的表达及其与临床病理特征的关系.方法 用免疫组化法检测60例OC、25例卵巢交界性肿瘤、30例卵巢良性肿瘤和21例正常卵巢组织中WAVEs蛋白的表达,结合临床病理资料进行分析;反转录聚合酶链反应法(RT-PCR)和Western blot分别检测WAVEs mRNA和蛋白表达.结果 1)WAVE1在OC中的表达显著高于正常卵巢组织、卵巢良性组织和卵巢交界性组织(P＜0.01).2)WAVE1在OC中的表达与临床分期、病理分化程度及Ca-125水平密切相关(.P＜0.05),而与年龄、肿瘤大小及腹水量无关.3)WAVE1在OC中mRNA、蛋白水平显著高于正常卵巢组织、卵巢良性肿瘤和交界性肿瘤(P＜0.01).而WAVE2、WAVE3在各组表达均低.结论 WAVE1在OC中高表达,可能与其发生发展及侵袭转移有关,可望成为卵巢癌治疗的靶标.     

Comparison of vascularity and angiogenesis in primary invasive mammary carcinomas and in their respective axillary lymph node metastases

It is well established that the ability of a neoplasm to induce a blood supply from a pre-existing circulation (angiogenesis) is a major factor in tumour growth, invasion and metastasis. However, the angiogenic potential of metastases and their subsequent growth have not been extensively studied. The question arises: can metastatic clones induce the same level of angiogenesis as in the primary neoplasm they emanated from? In this study it is hypothesised that in the same patient the level of vascularity and angiogenesis is the same in both the primary invasive ductal carcinoma and in the axillary lymph node metastasis at the time of surgery, according to Kerbels theory of clonal-dominance. To directly address the hypothesis, morphological measures of the established blood/lymphatic circulation (vascularity) as well as estimates of angiogenesis (endothelial cell proliferation) were measured in primary tumours and directly compared to the same parameters in the corresponding lymph node metastasis in a case by case basis (n=17). The results demonstrate varying associations between the level of vascularity and angiogenesis between matched individual tumours and their metastatic lymph nodal deposits. It is possible that either variations in the angiogenic characteristics of the metastasising clone or local or systemic promoters or inhibitors of angiogenesis influence tumour angiogenesis at the different sites.     

DNA profiling of primary serous ovarian and fallopian tube carcinomas with array comparative genomic hybridization and multiplex ligation-dependent probe amplification

Primary serous ovarian carcinoma (OVCA) and serous Fallopian tube carcinoma (FTC), both belonging to the BRCA-linked tumour spectrum, share many properties and are treated similarly. However, a detailed molecular comparison has been lacking. We hypothesized that comparative genomic studies of serous OVCAs and FTCs should point to gene regions critically involved in their tumorigenesis. Array comparative genomic hybridization (array CGH) analysis indicated that serous OVCAs and serous FTCs displayed common but also more distinctive patterns of recurrent changes. Targeted gene identification using a dedicated multiplex ligation-dependent probe amplification (MLPA) probe set directly identified EIF2C2 on 8q as a potentially important driver gene. Other previously unappreciated gained/amplified genes included PSMB4 on 1q, MTSS1 on 8q, TEAD4 and TSPAN9 on 12p, and BCAS4 on 20q. SPINT2 and ACTN4 on 19q were predominantly found in FTCs. Gains/amplifications of CCNE1 and MYC, often in conjunction with changes in genes of the AKT pathway, EVI1 and PTK2, seemed to be involved at earlier stages, whereas changes of ERBB2 were associated with advanced stages. The only BRCA1-mutated FTC shared common denominators with the sporadic tumours. In conclusion, the data suggest that serous OVCAs and FTCs, although related, exhibit differences in genomic profiles. In addition to known pathways, new genes/pathways are likely to be involved, with changes in an miRNA-associated gene, EIF2C2, as one important new feature. Dedicated MLPA sets constitute potentially important tools for differential diagnosis and may provide footholds for tailored therapy.