BACKGROUND: This study examined the clinical significance of CCNE1 (Cyclin E1) amplification and assessed whether CCNE1 is a potential therapeutic target in ovarian cancer. METHODS: CCNE1 expression and amplification in ovarian cancer was assessed by immunohistochemistry, fluorescence in situ hybridization and clinical data collected by retrospective chart review. CCNE1 gene knockdown using silencing RNA and a CCNE1 gene transfection system were used to asses CCNE1 function in tissue samples of ovarian cancer. RESULTS: Gene amplification was identified in 18 (20.4%) of 88 ovarian carcinomas. CCNE1 copy number significantly correlated with CCNE1 protein expression (r ¼ 0.522, P < .0001). CCNE1 amplification significantly correlated with shorter disease-free survival and overall survival (P < .001). There were nonsignificant trends between high protein expression and poor disease-free survival (P ¼ .2865) and overall survival (P ¼ .1248). Multivariate analysis showed gene amplification was an independent prognostic factor for disease-free survival and overall survival after standard platinum-taxane chemotherapy (P ¼ .0274, P ¼ .0023). Profound growth inhibition and apoptosis were observed in silencing RNA-treated cancer cells with gene amplification compared with results in cancer cells with CCNE1 moderate expression without gene amplification or with low CCNE1 expression. CCNE1 overexpression stimulated proliferation in ovarian cancer cell lines ES2 and TOV-21G, which have lower endogenous CCNE1 expression. CONCLUSIONS: These findings indicate that CCNE1 overexpression is critical to growth and survival of ovarian cancer tumors with CCNE1 gene amplification. Furthermore, they suggest that CCNE1 silencing RNA-induced phenotypes depend on amplification status of ovarian cancers. Therefore, CCNE1-targeted therapy may benefit ovarian cancer patients with CCNE1 amplification.

Recently, the ARID1A gene has been identified as a novel tumor suppressor in ovarian clear cell carcinoma. The prognostic significance of the loss of ARID1A expression is not known. The current study was designed to evaluate whether ARID1A was a prognostic factor for progression, survival, and chemoresistance in ovarian clear cell carcinoma. A total of 60 patients, who were surgically treated for primary ovarian clear cell adenocarcinoma, were enrolled. Surgical specimens were examined for ARID1A protein expression by immunohistochemistry. The correlations between the loss of ARID1A expression and clinicopathological characteristics, prognosis, and chemosensitivity were investigated. Loss of ARID1A expression was identified in 9 (15.0%) of 60 ovarian clear cell carcinoma samples. Loss of ARID1A staining intensity (0 þ ) was more frequently found in cells of clear cell carcinomas than in high-grade serous carcinomas (Po0.01). Loss of ARID1A expression was significantly correlated with advanced FIGO stage and high CA125 levels (P ¼ 0.02, 0.01). There were no significant correlations between loss of ARID1A expression and patient age, status of residual tumor, Ki-67 labeling index, or the status of endometriosis. Loss of ARID1A correlated with shorter progression-free survival of patients with clear cell carcinomas treated with platinum-based chemotherapy (Po0.01). Loss of ARID1A expression tended to correlate with shorter overall survival in patients with ovarian clear cell carcinomas treated with platinum-based chemotherapy. When data were stratified for the multivariate analysis, only the loss of ARID1A expression remained a significant (P ¼ 0.03) predictor of reduced progressionfree survival. Of the 60 patients with ovarian clear cell carcinomas, 14 patients had measurable residual tumor after primary cytoreductive surgery. Tumors with loss of ARID1A expression were more likely to be chemoresistant than tumors with positive ARID1A expression (100.0 vs 40.0%, P ¼ 0.04). This study demonstrates that loss of ARID1A in ovarian clear cell carcinoma is a negative prognostic factor in patients treated with platinum-based chemotherapy. Measurement of ARID1A expression may be a method to predict resistance to platinum-based chemotherapy in patients with ovarian clear cell carcinoma.

Sumary Decreased expression of E-cadherin (E-CD). a homotypic intercellular adhesion molecule, is considered to elicit detachment of tumour cells from primary lesions, which is the first stage of metastasis. Since renal cell cancer (RCC) shows a relatively high frequency of metastasis, we focused our interest on E-CD expression in RCC and its clinicopathological implications. We examined E-CD expression in normnal kidney and RCC by immunohistochemical staining. In normal kidney, E-CD expression was localised in distal tubules and collecting ducts. In RCC, 20 of 106 primary lesions (18.9%) expressed E-CD. whereas none showed positive staining for eight metastatic lesions. There was a statistically significant correlation between loss of E-CD expression and advanced stages of RCC. Kaplan-Meier analysis showed better prognosis in the group with preserved E-CD expression than without E-CD expression (Cox -Mantel test. P = 0.022, the average follow-up was 32 months or until death). This study suggests that the patients with decreased E-CD expression may be associated with metastasis, resulting in poor prognosis. However, frequency of E-CD expression in RCC is lower than in other cancers, which may be derived from the localised distribution of E-CD expression in normal kidney.Keywords E-cadherin; renal cell cancer: metastasis It is well known that cadherins, a family of Ca2+-dependent intercellular adhesion molecules, play essential roles in organogenesis and in the maintenance of normal structure and function (Behrens et al., 1985;Eidelman et al., 1989;Takeichi, 1991). Recently, E-cadherin (E-CD), a subclass of cadherins, has come to be considered to be important as an inhibitory factor in metastasis (Behrens et al., 1985; Frixen et al., 1991;Takeichi, 1991 (1991) showed that loss of E-CD can generate an invasive phenotype, which can be prevented by transfection with E-CD cDNA. Clinically, the correlation between decreased E-CD expression and advanced stages or dedifferentiation was also reported in a variety of tumours (Frixen et al., 1991;Shiozaki et al., 1991; Oka et al., 1992;Umbas et al., 1992;Bringuier et al., 1993; Oka et al., 1993;Terpe et al., 1993). However, reports in relation to the prognosis of malignant tumours have been rare until now (Bringuier et al., 1993 Immunohistochemical staining Anti-E-CD monoclonal antibody (HECD-1, Takara Biomedicals, Tokyo, Japan) was used in this study, and immunoperoxidase staining was performed by modifying the streptavidin-biotin bridge technique described previously (Katagiri et al., 1993

This study examined the status of KRAS and BRAF mutations, in relation to extracellular signal-regulated protein kinase (ERK) activation in 58 ovarian carcinomas to clarify the clinicopathological and prognostic significance of KRAS/BRAF mutations. Somatic mutations of either KRAS or BRAF were identified in 12 (20.6%) out of 58 ovarian carcinomas. The frequency of KRAS/BRAF mutations in conventional serous high-grade carcinomas (4.0% : 1/25) was significantly lower than that in the other histological type (32.3% : 10/31). Phosphorylated ERK1/2 (p-ERK1/2) expression was identified in 18 (38.2%) out of 45 ovarian carcinomas. KRAS/BRAF mutation was significantly correlated with International Federation of Gynecology and Obstetrics (FIGO) stage I, II (Po0.001), and p-ERK1/2 (Po0.001). No significant correlations between KRAS/BRAF mutations or p-ERK1/2 expression and overall survival were found in patients with ovarian carcinoma treated with platinum and taxane chemotherapy (P ¼ 0.2460, P ¼ 0.9339, respectively). Next, to clarify the roles of ERK1/2 activation in ovarian cancers harbouring KRAS or BRAF mutations, we inactivated ERK1/2 in ovarian cancer cells using CI-1040. Cl-1040 is a compound that selectively inhibits MAP kinase kinase (MEK), an upstream regulator of ERK1/2, and thus prevents ERK1/2 activation. Profound growth inhibition and apoptosis were observed in CI-1040-treated cancer cells with mutations in either KRAS or BRAF in comparison with the ovarian cancer cells containing wild-type sequences. This was evident in both in vitro and in vivo studies. The findings in this study indicate that an activated ERK1/2 pathway is critical to tumour growth and survival of ovarian cancers with KRAS or BRAF mutations. Furthermore, they suggest that the CI-1040-induced phenotypes depend on the mutational status of KRAS and BRAF in ovarian cancers. Therefore, ovarian cancer patients with KRAS or BRAF mutations may benefit from CI-1040 treatment.

Background:The aim of this study was to investigate the patterns of epidermal growth factor receptor (EGFR) overexpression, EGFR gene amplification, and the presence of activating mutations in the tyrosine kinase domain of this gene in squamous cell carcinomas and adenocarcinomas/adenosquamous carcinomas of the uterine cervix.Methods:The EGFR expression, amplification, and mutation in cervical carcinomas were assessed by immunohistochemistry, fluorescence in situ hybridisation, and PCR–SSCP, respectively, and correlated with clinical data collected by a retrospective chart review. A functional assessment was performed by inactivating EGFR in cervical cancer cells with the potent inhibitor AG1478.Results:Immunohistochemical analysis revealed that 6 out of 59 (10.2%) cervical squamous cell carcinomas showed significant amplification of the EGFR locus, whereas none of the 52 adeno/adenosquamous cell carcinomas had detectable EGFR amplification (P<0.05). The EGFR amplification significantly correlated with shorter overall survival (P=0.001) in cervical squamous cell carcinomas. Multivariate analysis showed that EGFR gene amplification was an independent prognostic factor for overall survival (P=0.011). None of the squamous cell carcinomas (0%: 0 out of 32) had detectable oncogenic mutations in EGFR exons 18 through 21. The frequencies of KRAS and BRAF mutations were very low in both squamous and adeno/adenosquamous cell carcinomas. Sensitivity of cervical cancer cells to AG1478 depended on the presence of EGFR overexpression. AG1478-induced EGFR inactivation in cell lines with EGFR overexpression significantly suppressed tumour development and progression in a mouse xenograft model.Conclusion:Our data suggest that EGFR signalling is important in a subset of cervical squamous cell carcinomas and that anti-EGFR therapy may benefit patients who carry the 7p11.2 amplicon in their tumours.

BACKGROUND:The goal of this study was to examine the clinical significance of ZNF217 amplification and assess whether ZNF217 could be a potential therapeutic target in ovarian clear cell carcinoma (OCCC). METHODS: ZNF217 expression and amplification in OCCC was assessed by immunohistochemistry, fluorescence in situ hybridization, and clinical data collected via a retrospective chart review. ZNF217 gene knockdown using silencing RNA (siRNA) was used to assess ZNF217 functions in OCCC cell lines. RESULTS: Gene amplification was identified in 12 of 60 (20.0%) OCCCs. ZNF217 copy number correlated significantly with ZNF217 protein expression (r ¼ 0.341; P<.01). ZNF217 amplification correlated significantly with shorter progression-free (P ¼.0042) and overall (P ¼.0199) survival. There were nonsignificant trends between high ZNF217 protein expression and poor progression-free (P ¼.2594) and overall (P ¼.2199) survival. Multivariate analysis revealed ZNF217 gene amplification to be an independent prognostic factor for progression-free and overall survival after standard platinum agent-based chemotherapy (P ¼.0339 and P ¼.031, respectively). Profound growth inhibition and apoptosis were observed in ZNF217 siRNA-treated cancer cells with gene amplification compared with cancer cells with ZNF217 moderate expression without ZNF217 gene amplification or with low ZNF217 expression. CONCLUSION: These findings indicate that ZNF217 overexpression is critical to growth and survival of OCCCs with ZNF217 gene amplification. Furthermore, they suggest that ZNF217 siRNAinduced phenotypes depend on amplification status of OCCCs. Therefore, ZNF217-targeted therapy may benefit OCCC patients with ZNF217 amplification.

BACKGROUND Evidence of systemic inflammation, i.e., elevation of serum C–reactive protein, interleukin‐6, and/or the erythrocyte sedimentation rate, is correlated to poorer prognosis of patients with renal cell carcinoma (RCC). Serum amyloid A (SAA) has been recognized mainly as acute‐phase reactant. METHODS Serum SAA from 72 patients with RCC were examined. Thirty‐eight of 72 patients with RCC had elevated SAA compared with 17 healthy donors. RESULTS The disease specific survival rate was significantly lower in the elevated SAA group, and SAA level was shown to be an independent prognostic factor by univariate and multivariate analysis. CONCLUSIONS Evaluation of serum SM level in RCC patients may be a useful prognostic indicator. Cancer 2001;92:2072–5. © 2001 American Cancer Society.

This study examined the clinical significance of Notch3 expression and assessed its usefulness as a potential therapeutic target in chemoresistant ovarian cancer. Notch3 expression was assessed with immunohistochemical examination, and clinical variables were collected with a retrospective chart review. Notch3 siRNA or γ-secretase inhibitor was used to assess Notch3 function in ovarian cancer cell lines. Notch3 overexpression correlated with shorter progression-free/overall survival in patients with advanced stage (stage III, IV) ovarian carcinoma treated with platinum and taxane. Three of 5 patients showed increased Notch3 immunostaining in recurrent tumors compared with corresponding primary tumors. Notch3 overexpression was observed in both the cisplatin-resistant KFr13 and cisplatin/paclitaxel-resistant KFr13Tx cells. Inactivation of Notch3 by γ-secretase inhibitor or siRNA decreased cell proliferation and induced apoptosis in the KFr13 and KFr13Tx cells. Our findings suggest that Notch3 expression may be related to chemoresistance and that the Notch3 pathway may represent a novel therapeutic target for advanced stage chemoresistant ovarian cancer.