Focal Adhesion Kinase Silencing Augments Docetaxel-Mediated Apoptosis in Ovarian Cancer Cells

Docetaxel causes cell death through induction of apoptosis; however, cell death characteristics for docetaxel have not yet been fully elucidated. We examined the role of focal adhesion kinase (FAK) cleavage in docetaxel-mediated apoptosis

Therapeutic Targeting of ATP7B in Ovarian Carcinoma.

PURPOSE: Resistance to platinum chemotherapy remains a significant problem in ovarian carcinoma. Here, we examined the biological mechanisms and therapeutic potential of targeting a critical platinum resistance gene, ATP7B, using both in vitro and in vivo models. EXPERIMENTAL DESIGN: Expression of ATP7A and ATP7B was examined in ovarian cancer cell lines by real-time reverse transcription-PCR and Western blot analysis. ATP7A and ATP7B gene silencing was achieved with targeted small interfering RNA (siRNA) and its effects on cell viability and DNA adduct formation were examined. For in vivo therapy experiments, siRNA was incorporated into the neutral nanoliposome 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine (DOPC). RESULTS: ATP7A and ATP7B genes were expressed at higher levels in platinum-resistant cells compared with sensitive cells; however, only differences in ATP7B reached statistical significance. ATP7A gene silencing had no significant effect on the sensitivity of resistant cells to cisplatin, but ATP7B silencing resulted in 2.5-fold reduction of cisplatin IC(50) levels and increased DNA adduct formation in cisplatin-resistant cells (A2780-CP20 and RMG2). Cisplatin was found to bind to the NH(2)-terminal copper-binding domain of ATP7B, which might be a contributing factor to cisplatin resistance. For in vivo therapy experiments, ATP7B siRNA was incorporated into DOPC and was highly effective in reducing tumor growth in combination with cisplatin (70-88% reduction in both models compared with controls). This reduction in tumor growth was accompanied by reduced proliferation, increased tumor cell apoptosis, and reduced angiogenesis. CONCLUSION: These data provide a new understanding of cisplatin resistance in cancer cells and may have implications for therapeutic reversal of drug resistance

Decreased expression of RNA interference machinery, Dicer and Drosha, is associated with poor outcome in ovarian cancer patients

The clinical and functional significance of RNA interference (RNAi) machinery, Dicer and Drosha, in ovarian cancer is not known and was examined. Dicer and Drosha expression was measured in ovarian cancer cell lines (n=8) and invasive epithelial ovarian cancer specimens (n=111) and correlated with clinical outcome. Validation was performed with previously published cohorts of ovarian, breast, and lung cancer patients. Anti-Galectin-3 siRNA and shRNA transfections were used for in vitro functional studies. Dicer and Drosha mRNA and protein levels were decreased in 37% to 63% of ovarian cancer cell lines and in 60% and 51% of human ovarian cancer specimens, respectively. Low Dicer was significantly associated with advanced tumor stage (p=0.007), and low Drosha with suboptimal surgical cytoreduction (p=0.02). Tumors with both high Dicer and Drosha were associated with increased median patient survival (>11 years vs. 2.66 years for other groups; p<0.001). In multivariate analysis, high Dicer (HR=0.48; p=0.02), high-grade histology (HR=2.46; p=0.03), and poor chemoresponse (HR=3.95; p<0.001) were identified as independent predictors of disease-specific survival. Findings of poor clinical outcome with low Dicer expression were validated in separate cohorts of cancer patients. Galectin-3 silencing with siRNA transfection was superior to shRNA in cell lines with low Dicer (78-95% vs. 4-8% compared to non-targeting sequences), and similar in cell lines with high Dicer. Our findings demonstrate the clinical and functional impact of RNAi machinery alterations in ovarian carcinoma and support the use of siRNA constructs that do not require endogenous Dicer and Drosha for therapeutic applications

Expression of imatinib mesylate-targeted kinases in endometrial carcinoma

Imatinib mesylate is a tyrosine kinase inhibitor that specifically targets c-Kit, Abl, and platelet-derived growth factor receptor (PDGFR). It has been shown to be an effective treatment for patients with chronic myelogenous leukemia (CML) and gastrointestinal stromal tumors (GIST). These cancers are characterized by activating mutations of the Abl and c-Kit tyrosine kinases, respectively. To determine whether imatinib mesylate could be a potentially useful agent in the treatment of endometrial cancer, we assessed the expressions of Abl, c-Kit, and PDGFR in both primary and recurrent endometrial carcinoma. We performed immunohistochemical analysis on formalin-fixed, paraffin-embedded sections from 63 patients: 33 with endometrioid endometrial carcinoma (EEC), 11 with uterine papillary serous carcinoma (UPSC), 12 with recurrent EEC, and seven with recurrent UPSC. The sections were stained with commercially available antibodies for Abl, PDGFR, and c-Kit. The sections were also stained for phosphorylated Abl and phosphorylated PDGFR. Among the primary EEC, 28/33 (85%) stained positively for Abl and 30/33 (91%) were positive for PDGFR. Of the primary UPSC, 8/11 (73%) were positive for Abl. In addition, 8/11 (73%) of the primary UPSC tumors were positive for PDGFR. Neither the primary EEC (0/33) nor the primary UPSC (0/11) expressed c-Kit. Of the recurrent EEC tumors, 11/12 (92%) were positive for Abl expression, 12/12 (100%) were positive for PDGFR, and 2/8 (25%) were positive for c-Kit. Of the recurrent UPSC, 6/7 (86%) were positive for Abl, 7/7 (100%) were positive for PDGFR, and 2/4 (50%) for c-Kit. In addition, the majority of primary and recurrent tumors were positive for phosphorylated Abl (primary EEC, 91%; primary UPSC, 64%; recurrent EEC, 83%; recurrent UPSC, 86%), and phosphorylated PDGFR (primary EEC, 46%; primary UPSC, 40%; recurrent EEC, 58%; recurrent UPSC, 100%). Within the EEC primary tumors, the differences in kinase expression by grade of tumor were not significant except for PDGFR kinase; the lower grade tumors (1 and 2) had more PDGFR expression than the grade 3 tumors (P < 0.05). The majority of primary and recurrent EEC, as well as primary and recurrent UPSC express Abl and PDGFR. This preclinical data suggest that imatinib mesylate may be useful in the treatment of patients with endometrial carcinoma

Molecular determinants of tumor differentiation in papillary serous ovarian carcinoma.

In epithelial ovarian cancer, tumor grade is an independent prognosticator whose molecular determinants remain unknown. We investigated patterns of gene expression in well- and poorly differentiated serous papillary ovarian and peritoneal carcinomas with cDNA microarrays. A 6500-feature cDNA microarray was used for comparison of the molecular profiles of eight grade III and four grade I stage III serous papillary adenocarcinomas. With a modified F-test in conjunction with random permutations, 99 genes whose expression was significantly different between grade I and grade III tumors were identified (P < 0.01). A disproportionate number of these differentially expressed genes were located on the chromosomal regions 20q13 and all exhibited higher expression in grade III tumors. Interphase fluorescent in situ hybridization demonstrated 20q13 amplification in two of the four grade III and none of the three grade I tumors available for evaluation. Several centrosome-related genes also showed higher expression in grade III tumors. We propose a model in which tumor differentiation is inversely correlated with the overexpression of several oncogenes located on 20q13, a common amplicon in ovarian and numerous other cancers. Dysregulation of centrosome function is one potential mechanistic link between genetic/epigenetic changes and the poorly differentiated phenotype in ovarian cancer.Journal Articleinfo:eu-repo/semantics/publishe

Ubiquitin Carboxyl-Terminal Hydrolase L1 (UCHL1) Promotes Uterine Serous Cancer Cell Proliferation and Cell Cycle Progression

Uterine serous carcinoma (USC) is the most aggressive form of endometrial cancer, with poor survival rates and high recurrence risk. Therefore, the purpose of this study was to identify therapeutic targets that could aid in the management of USC. By analyzing endometrial cancer samples from The Cancer Genome Atlas (TCGA), we found Ubiquitin Carboxyl-Terminal Hydrolase L1 (UCHL1) to be highly expressed in USC and to correlate with poorer overall survival. UCHL1 silencing reduced cell proliferation in vitro and in vivo, cyclin B1 protein levels and cell cycle progression. Further studies showed that UCHL1 interacts with cyclin B1 and increases cyclin B1 protein stability by deubiquitination. Treatment of USC-bearing mice with the UCHL1-specific inhibitor reduced tumor growth and improved overall survival. Our findings suggest that cyclin B1 is a novel target of UCHL1 and targeting UCHL1 is a potential therapeutic strategy for USC