Activity-based protein profiling reveals active serine proteases that drive malignancy of human ovarian clear cell carcinoma.

Ovarian clear cell carcinoma (OCCC) is an understudied poor prognosis subtype of ovarian cancer lacking in effective targeted therapies. Efforts to define molecular drivers of OCCC malignancy may lead to new therapeutic targets and approaches. Among potential targets are secreted proteases, enzymes which in many cancers serve as key drivers of malignant progression. Here, we found that inhibitors of trypsin-like serine proteases suppressed malignant phenotypes of OCCC cell lines. To identify the proteases responsible for malignancy in OCCC, we employed activity-based protein profiling to directly analyze enzyme activity. We developed an activity-based probe featuring an arginine diphenylphosphonate warhead to detect active serine proteases of trypsin-like specificity and a biotin handle to facilitate affinity purification of labeled proteases. Using this probe, we identified active trypsin-like serine proteases within the complex proteomes secreted by OCCC cell lines, including two proteases in common, tissue plasminogen activator and urokinase-type plasminogen activator. Further interrogation of these proteases showed that both were involved in cancer cell invasion and proliferation of OCCC cells and were also detected in in vivo models of OCCC. We conclude the detection of tissue plasminogen activator and urokinase-type plasminogen activator as catalytically active proteases and significant drivers of the malignant phenotype may point to these enzymes as targets for new therapeutic strategies in OCCC. Our activity-based probe and profiling methodology will also serve as a valuable tool for detection of active trypsin-like serine proteases in models of other cancers and other diseases.

Identification of novel ALK rearrangements in gynecologic clear cell carcinoma.

Clear cell carcinomas (CCCs) of the gynecologic tract are aggressive tumors with high resistance rate to conventional platinum-based chemotherapies. Currently, the molecular features of these tumors remain largely unknown and there is no targeted therapy available. The aim of our study was to identify anaplastic lymphoma kinase (ALK) translocations, a potential molecular target for therapy. Ninety-seven patients with gynecologic CCC (62 ovarian, 27 uterine corpus and 8 uterine cervical) were screened for ALK rearrangement and ALK copy number gain using an ALK break-apart fluorescence in situ hybridization probe. The genomic landscape of all cases with ALK rearrangements and 10 random cases with ALK copy number gain was queried using a hybrid capture-based DNA next-generation sequencing assay and an Illumina Fusion RNA assay. Findings were then correlated with ALK immunohistochemistry (clone D5F3) expression. ALK rearrangement was detected in 5% (5/97) and ALK copy number gain in 79% (77/97) of gynecologic CCCs. Next-generation sequencing in ALK-rearranged CCCs identified a novel BABAM2-ALK fusion in one case. ALK translocation partners were not identified in the remaining cases. Our findings show that ALK fusion, which is targetable in other cancers, may be a pathogenetic mechanism in a small number of gynecologic CCCs.

Clinical and pathological associations of PTEN expression in ovarian cancer: a multicentre study from the Ovarian Tumour Tissue Analysis Consortium.

BACKGROUND: PTEN loss is a putative driver in histotypes of ovarian cancer (high-grade serous (HGSOC), endometrioid (ENOC), clear cell (CCOC), mucinous (MOC), low-grade serous (LGSOC)). We aimed to characterise PTEN expression as a biomarker in epithelial ovarian cancer in a large population-based study. METHODS: Tumours from 5400 patients from a multicentre observational, prospective cohort study of the Ovarian Tumour Tissue Analysis Consortium were used to evaluate associations between immunohistochemical PTEN patterns and overall survival time, age, stage, grade, residual tumour, CD8+ tumour-infiltrating lymphocytes (TIL) counts, expression of oestrogen receptor (ER), progesterone receptor (PR) and androgen receptor (AR) by means of Cox proportional hazard models and generalised Cochran-Mantel-Haenszel tests. RESULTS: Downregulation of cytoplasmic PTEN expression was most frequent in ENOC (most frequently in younger patients; p value = 0.0001) and CCOC and was associated with longer overall survival in HGSOC (hazard ratio: 0.78, 95% CI: 0.65-0.94, p value = 0.022). PTEN expression was associated with ER, PR and AR expression (p values: 0.0008, 0.062 and 0.0002, respectively) in HGSOC and with lower CD8 counts in CCOC (p value < 0.0001). Heterogeneous expression of PTEN was more prevalent in advanced HGSOC (p value = 0.019) and associated with higher CD8 counts (p value = 0.0016). CONCLUSIONS: PTEN loss is a frequent driver in ovarian carcinoma associating distinctly with expression of hormonal receptors and CD8+ TIL counts in HGSOC and CCOC histotypes.

High expression of glutamate-ammonia ligase is associated with unfavorable prognosis in patients with ovarian cancer.

Glutamate-ammonia ligase (GLUL), which is also called GS (glutamine synthetase), is the enzyme that catalyzes the synthesis of glutamine from glutamate and ammonia in an ATP-dependent reaction. Here, we found higher expression of GLUL in the ovarian cancer patients was associated with worse disease-free survival (DFS) and overall survival (OS). In addition, GLUL was heterogeneously expressed in various ovarian cancer cells. The mRNA and protein expression levels of GLUL in NIH:OVCAR-3 and ES-2 cells were obviously higher than that in the other types of ovarian cancer cells. Knockdown of GLUL in NIH:OVCAR-3 or ES-2 cells could significantly decrease the proliferation ability. Furthermore, GLUL knockdown markedly inhibited the p38 MAPK signaling pathway in NIH:OVCAR-3 or ES-2 cells. Our findings suggest that decreasing expression of GLUL may be a new approach that can be used for ovarian cancer treatment.

Frequent Mismatch Repair Protein Deficiency in Mixed Endometrioid and Clear Cell Carcinoma of the Endometrium.

Mixed endometrioid and clear cell carcinoma of the endometrium refers to a scenario in which the tumor exhibits histologic features of both endometrioid and clear cell carcinoma. We observed a tendency for these tumors to occur in a mismatch repair (MMR) protein-deficient molecular background in a prior study that examined a small cohort of mixed-type endometrial carcinomas. The aim of this study was to determine the rate of MMR protein deficiency in a larger series of endometrial mixed endometrioid and clear cell carcinomas, through a retrospective survey of MLH1, PMS2, MSH2, and MSH6 expression in such tumors at 5 tertiary centers. A total of 41 cases were identified and 27 (66%) tumors demonstrated MMR protein deficiency with a comparable frequency across the contributing centers (ranging from 56% to 83%). Among the MMR protein-deficient cases, 59% showed concurrent MLH1 and PMS2 loss, 33% showed concurrent MSH2 and MSH6 loss, and 4% showed isolated PMS2 or MSH6 loss. Compared with a previously published series of 15 pure endometrial clear cell carcinomas, mixed endometrioid and clear cell carcinomas are associated with significantly better disease-specific survival (P=0.02). In summary, endometrial carcinomas with mixed endometrioid and clear cell histology are frequently MMR protein deficient. This finding has implications both for our understanding of its tumor biology and for the identification of patients with potential Lynch syndrome.

Inhibition of Aurora Kinase A Synergistically Enhances Cytotoxicity in Ovarian Clear Cell Carcinoma Cell Lines Induced by Cisplatin: A Potential Treatment Strategy.

OBJECTIVE: This study aims to clarify the incidence of Aurora kinase A (Aurora-A) protein expression and its correlation with clinical parameters in ovarian clear cell carcinoma (OCCC) tumor tissues. In addition, we assessed the efficacy of ENMD-2076, a novel selective Aurora-A inhibitor, in combination with chemotherapeutic agents for the treatment of OCCC. METHODS/MATERIALS: Aurora-A protein expression was determined by immunohistochemical staining of OCCC specimens from 56 patients to evaluate its correlation with clinical outcomes in OCCC. In the in vitro study, 6 OCCC cell lines were exposed to ENMD-2076 in combination with cisplatin, SN38, doxorubicin, or paclitaxel, and cell proliferation, cell cycle distribution, and apoptosis were assessed. RESULTS: The 5-year survival rates of International Federation of Gynecology and Obstetrics stages IC3 to IV patients with intermediate or strong Aurora-A expression were significantly lower than those of patients with negative or weak Aurora-A expression. Increased Aurora-A expression was associated with significantly worse overall survival of International Federation of Gynecology and Obstetrics stages IC3 to IV patients (21% vs 77%). Multivariate analysis revealed that Aurora-A expression was an independent prognostic factor for stages IC3 to IV OCCC patients. Furthermore, synergistic effects were observed with ENMD-2076 in combination with cisplatin or SN-38 in 4 of the 6 tested cell lines. ENMD-2076 dramatically enhanced apoptosis and cell cycle arrest at the G2/M phase induced by cisplatin. CONCLUSIONS: Aurora-A is a promising biomarker that is predictive of patient outcomes and a potential target for OCCC. The results suggested that chemotherapy, including ENMD-2076 in combination with cisplatin, is a potential treatment modality for patients with OCCC.

Frequent somatic mutations of the telomerase reverse transcriptase promoter in ovarian clear cell carcinoma but not in other major types of gynaecological malignancy.

Up-regulated expression of telomerase reverse transcriptase (TERT) and subsequent maintenance of telomere length are essential in tumour development. Recent studies have implicated somatic gain-of-function mutations at the TERT promoter as one of the mechanisms that promote transcriptional activation of TERT; however, it remains unclear whether this genetic abnormality is prevalent in gynaecological neoplasms. We performed mutational analysis in a total of 525 gynaecological cancers, and correlated TERT promoter mutations with clinicopathological features. With the exception of ovarian clear cell carcinomas, in which mutations were found in 37 (15.9%) of 233 cases, the majority of gynaecological malignancies were wild-type. TERT promoter mutation does not appear to be an early event during oncogenesis, as it was not detected in the contiguous endometriosis associated with ovarian clear cell carcinoma. Ovarian clear cell carcinoma cell lines with TERT promoter mutations exhibited higher TERT mRNA expression than those with wild-type sequences (p = 0.0238). TERT promoter mutation tended to be mutually exclusive with loss of ARID1A protein expression (p = 4.4 × 10(-9) ) and PIK3CA mutation (p = 0.0019) in ovarian clear cell carcinomas. No associations with disease-specific survival were observed for ovarian clear cell carcinoma. The above results, in conjunction with our previous report showing longer telomeres in ovarian clear cell carcinomas relative to other types of ovarian cancer, suggests that aberrations in telomere biology may play an important role in the pathogenesis of ovarian clear cell carcinoma.

The Effect of Cyclooxygenase-2 Expression in Uterine Carcinosarcoma on Survival: A Reassessment Based on Mature Data.

OBJECTIVE: To reassess the effect cyclooxygenase-2 (COX-2) expression in carcinosarcoma on survival based on mature 5-year survival data. METHOD: A comparison of 5-year survival of 27 patients with carcinosarcoma according to the presence of COX-2 immunohistochemical staining and staining score was performed. RESULTS: The 5-year survival of those with positive and negative COX-2 staining was statistically not different. However, there was a clear trend for more favorable 5-year survival in patients with a high staining score than in those with a low score, and the difference was of borderline significance (38.5% vs 7.1%; P = 0.06). CONCLUSION: In view of the role of COX-2 in carcinogenesis, our finding that COX-2 expression may confer a better survival in patients with carcinosarcoma is intriguing. Larger studies are indicated to elucidate the effect of COX-2 expression on survival in patients with carcinosarcoma because this may have therapeutic implications.

Fatty acid synthase expression associated with NAC1 is a potential therapeutic target in ovarian clear cell carcinomas.

BACKGROUND: This study examined the clinical significance of NAC1 and the expression level of its potential downstream target fatty acid synthase (FASN) in ovarian clear cell carcinomas (OCCCs), and evaluated the NAC1/FASN pathway as a potential therapeutic target. METHODS: NAC1 and FASN expression and NACC1 gene amplification were assessed in ovarian cancers by immunohistochemistry, fluorescence in situ hybridisation, and clinical data collected by a retrospective chart review. C75, a FASN inhibitor, was used to assess whether this pathway represented a therapeutic target in OCCC. RESULTS: High NAC1 expression was most frequent in clear cell tumours (40.0%:24/60). NACC1 gene amplification was identified in none of the 58 OCCCs. The frequency of NACC1 gene amplification was significantly higher in the high-grade serous histology than in the clear cell histology (P<0.01). NAC1 expression was significantly correlated with FASN expression in both OCCC samples and OCCC cell lines. Either high NAC1 expression or high FASN expression significantly correlated with shorter progression-free and overall survival (P=0.002 and 0.0048). NAC1 overexpression stimulated FASN expression, and NAC1 silencing using siRNA decreased FASN expression in OCCC cell lines. Profound growth inhibition was observed in C75-treated carcinoma cells with FASN overexpression when compared with the response in carcinoma cells without FASN expression. CONCLUSION: These findings indicate that NAC1/FASN overexpression is critical to the growth and survival of a subset of OCCC. The FASN silencing by the C75-induced phenotypes depends on the expression status of the targeted cell line. Therefore, NAC1/FASN pathway-targeted therapy may benefit selected OCCC patients.

PPM1D is a potential therapeutic target in ovarian clear cell carcinomas.

PURPOSE: To identify therapeutic targets in ovarian clear cell carcinomas, a chemoresistant and aggressive type of ovarian cancer. EXPERIMENTAL DESIGN: Twelve ovarian clear cell carcinoma cell lines were subjected to tiling path microarray comparative genomic hybridization and genome-wide expression profiling analysis. Regions of high-level amplification were defined and genes whose expression levels were determined by copy number and correlated with gene amplification were identified. The effects of inhibition of PPM1D were assessed using short hairpin RNA constructs and a small-molecule inhibitor (CCT007093). The prevalence of PPM1D amplification and mRNA expression was determined using chromogenic in situ hybridization and quantitative real-time reverse transcription-PCR in a cohort of pure ovarian clear cell carcinomas and on an independent series of unselected epithelial ovarian cancers. RESULTS: Array-based comparative genomic hybridization analysis revealed regions of high-level amplification on 1q32, 1q42, 2q11, 3q24-q26, 5p15, 7p21-p22, 11q13.2-q13.4, 11q22, 17q21-q22, 17q23.2, 19q12-q13, and 20q13.2. Thirty-four genes mapping to these regions displayed expression levels that correlated with copy number gains/amplification. PPM1D had significantly higher levels of mRNA expression in ovarian clear cell carcinoma cell lines harboring gains/amplifications of 17q23.2. PPM1D inhibition revealed that PPM1D expression and phosphatase activity are selectively required for the survival of ovarian clear cell carcinoma cell lines with 17q23.2 amplification. PPM1D amplification was significantly associated with ovarian clear cell carcinoma histology (P = 0.0003) and found in 10% of primary ovarian clear cell carcinomas. PPM1D expression levels were significantly correlated with PPM1D gene amplification in primary ovarian clear cell carcinomas. CONCLUSION: Our data provide strong circumstantial evidence that PPM1D is a potential therapeutic target for a subgroup of ovarian clear cell carcinomas.

Expression of alpha-methylacyl-CoA racemase in vaginal gastric-type adenocarcinoma and uterine clear cell carcinoma.

BACKGROUND: Alpha-methylacyl-coenzyme A racemase (AMACR, P504S) is a commonly used marker in immunohistochemical diagnosis of prostate cancer. Recent studies identified P504S markers of the clear cell histotype in the ovary and/or endometrium. Gastric-type adenocarcinoma (GAS) is difficult to diagnose histologically, particularly when there is crossover with clear cell carcinoma (CCC). However, the significance of P504S for differentially diagnosing GAS and CCC is unclear. AIM: To evaluate P504S as a potential diagnostic marker of GAS and CCC. SETTINGS AND DESIGN: We analyzed P504S expression in 48 cervical carcinomas (32 GAS and 16 CCC), as well as the expression of other markers including hepatocyte nuclear factor-1 beta (HNF-1ß) and NapsinA. MATERIAL AND METHODS: The expression differences of HNF-1ß, NapsinA, and P504S in GAS and CCC were detected by immunohistochemistry. Immunohistochemical histoscores based on the intensity and extent of staining were calculated. RESULTS: The positive rates of HNF-1ß in GAS and CCC were 90.32% and 75%, respectively. (χ2 = 2.251, P = 0.663). The positive rates of NapsinA in GAS and CCC were 19.36% and 81.25%, respectively. (χ2 = 47.332, P < 0.01). The positive rates of P504S in GAS and CCC were 16.13% and 81.25%, respectively. (χ2 = 41.420, P < 0.01). HNF-1ß was frequently expressed in GAS and CCC, while NapsinA and P504S were frequently expressed in CCC, and reduced or lost in GAS. CONCLUSION: NapsinA and P504S can be used to differentiate between GAS and CCC.

Expression of membrane type 1 matrix metalloproteinase (MMP-14) in epithelial ovarian cancer: high level expression in clear cell carcinoma.

OBJECTIVE: Clear cell carcinomas of the ovary constitute approximately 5% of all ovarian neoplasms and have a distinct gene expression profile relative to other ovarian carcinoma histotypes. Tumors often present as an early stage large pelvic mass with a high degree of recurrence and frequent early metastasis. Matrix metalloproteinases (MMPs) play a role in intraperitoneal metastasis through breakdown of cell-cell and cell-matrix barriers, enabling anchoring of secondary lesions and promoting proliferation in a geometrically constrained matrix environment. The objective of this study was to evaluate MMP expression in ovarian clear cell carcinoma. METHODS: Immunohistochemistry was used to evaluate expression of membrane type 1 MMP (MMP-14), MMP-2 and MMP-9 in a panel of ovarian tumors. Western blotting and gelatin zymography were used to examine MMP-14 expression and activity in the clear cell carcinoma cell line ES2. The ability of ES2 cells to invade and proliferate within three-dimensional collagen gels was evaluated. RESULTS: High level expression of MMP-14 and MMP-2 were observed in ovarian clear cell carcinoma relative to other histotypes (94-95% strong positive). MMP-14 was expressed and active in cultured ES2 cells. ES2 cells also exhibited MMP-dependent invasion of and proliferation within three-dimensional collagen gels. CONCLUSIONS: The high level expression of MMP-14 together with in vitro functional analyses suggest that MMP-14 may contribute to both the proliferative capacity and the enhanced parenchymal metastasis of ovarian clear cell carcinoma.

PTEN promoter methylation and LOH of 10q22-23 locus in PTEN expression of ovarian clear cell adenocarcinomas.

OBJECTIVES: Loss of phosphatase and tensin homolog (PTEN) expression is common in ovarian clear cell adenocarcinomas (OCCA), but PTEN mutations are not frequently observed in OCCA. The mechanism of PTEN gene silencing in OCCA is still not clear. MATERIALS AND METHODS: Immunohistochemical analysis of PTEN expression was performed in 40 OCCA paraffin-embedded tissues. PTEN promoter methylation in 24 OCCA tissues and 5 OCCA cell lines was examined by methylation-specific PCR. Eighteen OCCA patients and 13 serous adenocarcinomas were analyzed for loss of heterozygosity (LOH) at 10q23 with five polymorphic markers. RESULTS: Of the 40 OCCAs, 37.5% (15/40) had reduced PTEN immunoreactivity, LOH was found in 33% (6/18) of OCCAs, and 31% (4/13) of serous adenocarcinomas. In the 33% of OCCAs with LOH, only 33% (2/6) lost PTEN expression. PTEN promoter was unmethylated in 5 OCCA cell lines and 24 OCCA tissues detected by MSP-PCR. No significant correlation between PTEN expression and advanced stage disease or overall survival was found. CONCLUSION: Our results indicate that reduced PTEN expression was detected in more than one third of OCCA cases. Neither PTEN promoter methylation nor LOH at 10q23 locus is significantly related to PTEN inactivation and is not an adverse prognostic factor in OCCA.

High expression of N-acetylglucosaminyltransferase V in mucinous tumors of the ovary.

N-acetylglucosaminyltransferase V (GnT-V) is an enzyme that catalyzes beta1-6 branching of N-acetylglucosamine (GlcNAc) on asparagine-linked oligosaccharides of cell proteins. The present study aimed to investigate GnT-V expression and its prognostic significance in epithelial ovarian cancer. GnT-V expression was studied by immunohistochemistry in 83 surgically resected ovarian cancers, and the staining intensity was evaluated. High GnT-V expression in cancer cells was found in 17 (20.5%) of the 83 cases, and was positively correlated with early FIGO staging. In the histological type, mucinous adenocarcinoma showed significantly strong immunostaining compared to the non-mucinous type (P<0.001). In 36 mucinous tumors, the GnT-V immunostaining score was significantly higher in cancer than in benign and borderline tumors (P<0.001). NOM-1, a human ovarian mucinous adenocarcinoma cell line, expressed strong GnT-V protein and swainsonine treatment suppressed beta1-6GlcNAc branching and reduced migration ability significantly (P<0.001). These results suggested that GnT-V might be involved in the malignant potential of mucinous ovarian cancer.

Glutathione peroxidase 3 is a candidate mechanism of anticancer drug resistance of ovarian clear cell adenocarcinoma.

Ovarian clear cell adenocarcinoma has low sensitivity to platinum drugs. The molecular-biological mechanism of the low sensitivity has not been clarified. The objective of this study was to identify candidate genes associated with low sensitivity of clear cell adenocarcinoma to platinum drugs. Exhaustive gene profiling of 4 ovarian clear cell adenocarcinoma cell lines, KK, OVMANA, OVSAYO, and RMG-1 and 4 ovarian serous adenocarcinoma cell lines, KF, HRA, SHIN-3 and KOC-2S was performed by DNA microarray. Obtained candidate genes were suppressed by RNA interference and changes in the cisplatin sensitivity of clear cell adenocarcinoma cells were observed. Six genes including the glutathione peroxidase 3 (GPX3) gene were identified to be highly expressed in clear cell adenocarcinoma by DNA microarray analysis. GPX3 suppression by RNA interference increased cisplatin sensitivity 3.3-4.2-fold in 3 of the 4 clear cell adenocarcinoma cell lines. GPX3 was identified to be a gene highly expressed in clear cell adenocarcinoma. Since GPX3 suppression increased the cisplatin sensitivity of clear cell adenocarcinoma cells, GPX3 may be a candidate gene associated with the low cisplatin sensitivity of clear cell adenocarcinoma.

Napsin-A, a Possible Diagnostic Marker for Differentiating Clear Cell Ovarian Carcinoma From Other High-grade Ovarian Carcinomas.

Ovarian clear cell carcinoma (CCC) is divergent from other types of epithelial ovarian carcinoma in terms of clinicopathologic and molecular features. It should be separated from other high-grade carcinomas of the ovary for appropriate treatment. Napsin A is a reliable marker for adenocarcinoma of the lungs, but its role in ovarian epithelial carcinomas is vague. We investigated the expression of a panel of TTF-1, paired box 8, estrogen receptor, Wilms tumor 1, and Napsin A in 100 cases of high-grade ovarian carcinomas. All the examined cases were TTF-1 negative and paired box 8 positive. The 2 biomarkers estrogen receptor together with Wilms tumor 1 can separate CCC from endometriod carcinoma, yet this cannot be carried out in the case of serous and mucinous carcinomas of high grade. Napsin A can differentiate CCC with high sensitivity and specificity. It can be concluded that Napsin A is a sensitive and specific marker for CCC of the ovary. However, an entire marker panel may be useful for distinguishing ovarian CCC from other mimics.

Renal papillary adenoma--a putative precursor of papillary renal cell carcinoma.

The precursor lesions of renal cell carcinoma (RCC) are unknown. The purpose of this study is to determine the incidence, histomorphological features, and immunohistochemical features of papillary adenoma and elucidate its potential relationship to RCC. We reviewed 542 consecutive nephrectomy specimens over an 8-year period. Immunohistochemistry was carried out with antibodies specific for alpha-methyl-coenzyme A racemase (AMACR) and glutathione S-transferase alpha (clear-cell RCC marker). Thirty-eight (7%) nephrectomy specimens showed histologic evidence of papillary adenoma. Of these 38 cases, 18 (47%) arose in the setting of papillary RCC (PRCC). Seven papillary adenomas (18%) occurred in the setting of acquired polycystic kidney disease (APKD), 6 in clear-cell RCCs, 3 in chromophobe RCCs, 2 in end-stage kidney disease, 1 in oncocytoma, 1 in angiomyolipoma, and 1 in renal schwannoma. Furthermore, papillary adenomas were more commonly found in kidneys removed for PRCC (25%, 18/71) than in kidneys harboring clear-cell RCC (1.9%, 6/318). Histomorphologically, papillary adenomas were characterized by varying proportions of papillae and tubules formed by cuboidal cells with scant basophilic cytoplasm similar to those in type 1 PRCC. Adenomas associated with PRCC tend to be multiple in number (61% [11/18] of cases had >2 adenomas; mean, 5). In contrast, 100% of papillary adenomas arising in other conditions had less than 2 adenomas. Most of the adenomas (82%, 31/38) stained strongly for AMACR in a fashion similar to that of PRCC. The 7 AMACR-negative cases all arose in the setting of APKD. In this study of surgical specimens, the high coincidence, multifocality, and histologic and immunohistochemical similarities between papillary adenoma and PRCC suggest that the 2 are strongly associated and may represent a continuum of 1 biologic process. In contrast, adenomas associated with APKD exhibit distinct morphological and immunohistochemical features and, therefore, may have an entirely different pathogenesis.

EGFR and VEGFR2 protein expressions in bone metastases of clear cell renal cancer.

BACKGROUND: EGFR and VEGFR2 protein expressions are hallmarks of clear cell renal cancer (RCC) with questionable prognostic impact. The skeletal system is one of the most common metastatic sites of RCC. Unfortunately, there are no data for EGFR and VEGFR2 protein expression in such lesions. MATERIALS AND METHODS: Twenty cases of bone metastatic clear cell RCC were analyzed. EGFR and VEGFR2 proteins were detected by immunohistochemistry and analyzed by morphometry scoring both % positivity and the intensity. RESULTS: EGFR protein scores were significantly reduced in bone metastases of RCC due to the reduction of EGFR protein expression in about one third of the cases (7/20). The VEGFR2 protein-positive phenotype of clear cell RCC was relatively frequent (7/20, 35%), but was lost in bone metastases (2/20, 10%). CONCLUSION: These data suggest a phenotypic/genotypic change of clear cell RCC during the progression to bones and warrant further investigation.

Expression of NEU3 (plasma membrane-associated sialidase) in clear cell adenocarcinoma of the ovary: its relationship with T factor of pTNM classification.

Cell surface carbohydrate expression strongly influences the biological characteristics of cancer cells. Especially, it is known that the change of sialic acid expression could be related to the invasive and metastatic potentials of tumors. This study aimed to investigate sialidase expression of ovarian cancer cells and to evaluate the relationship between plasma membrane-associated sialidase (NEU3) expression and various clinicopathological factors in ovarian clear cell adenocarcinoma patients. In 18 cell lines derived from human ovarian cancers (including clear cell, mucinous, and serous adenocarcinoma), sialidase mRNA expression was evaluated by RT-PCR. NEU1 and NEU3 expression levels were found to be elevated in most cell lines while NEU2 and NEU4 expression was rarely elevated. Interestingly, NEU3 expression was detected in all clear cell adenocarcinoma cell lines. In 71 patients with ovarian clear cell adenocarcinoma, treated at Keio University Hospital from February 1983 to February 2002, NEU3 expression was examined by immunohistochemical staining of surgical specimens and clinicopathological factors were reviewed. NEU3 expression was found to be positive in 77.5% of all cases. Furthermore, a high level of NEU3 expression was significantly correlated with T3 factor of pTNM classification on univariate and multivariate analysis. This is the first report to show that NEU3 is expressed in most of ovarian clear cell adenocarcinoma. And our results show that NEU3 expression is correlated with T factor (pTNM classification) in ovarian clear cell adenocarcinoma.

Alterations of the DNA repair gene OGG1 in human clear cell carcinomas of the kidney.

The OGG1 gene, which codes for a DNA repair protein with antimutator activity, is located on chromosome 3p25, a frequent site of allelic deletions in many types of human tumors, including renal clear cell cancers. We present the analysis of 99 renal tumors for alterations in the OGG1 gene to determine its association with tumorigenesis. Loss of heterozygosity in the 3p25 region was found for 85% of the informative cases. We detected somatic missense mutations of the OGG1 gene in 4 of the 99 tumor samples. Biochemical analysis of the mutant proteins revealed that a substitution at codon 46 impairs the enzymatic activity. We also describe the occurrence of several polymorphisms as well as aberrantly spliced OGG1 transcripts.