Lingya Pan

Increased Expression of Annexin A3 Is a Mechanism of Platinum Resistance in Ovarian Cancer

Resistance to platinum drugs has emerged as a major obstacle in the treatment of ovarian cancers. Through proteomic analysis, we have found that the expression of annexin A3, a member of the Ca(2+) and phospholipid-binding annexin family, is significantly increased in platinum-resistant ovarian cell lines. Anti-annexin A3 immunostaining indicated that cancers from platinum-resistant patients also possess higher levels of annexin A3 than those from platinum-sensitive patients. Although expression of annexin A3 made susceptible ovarian cancer cells more resistant to platinum, expression of antisense annexin A3 downregulated its expression and rendered the resistant cells more sensitive to platinum. In athymic mice, the growth of tumors from inoculated SKOV3 cells was inhibited by the administration of platinum, whereas tumors from annexin A3-expressing SKOV3/Ann were resistant to platinum treatment. Interestingly, the intracellular platinum concentration and platinum-DNA binding are significantly lower in annexin A3-overexpressing cells than those in parental cells. The lower cisplatin concentration was also accompanied by reduced induction of p53, which could be restored by downregulation of annexin A3. These results indicate that increased expression of annexin A3 is a mechanism of platinum resistance in ovarian cancer. It seems to act by preventing uptake or accumulation of platinum in cells. Therefore, it is conceivable that annexin A3 could be a target for therapeutic intervention and may also serve as a biomarker for drug resistance in ovarian cancer patients.

Secretion of annexin A3 from ovarian cancer cells and its association with platinum resistance in ovarian cancer patients.

Early detection of resistance to platinum‐based therapy is critical for improving the treatment of ovarian cancers. We have previously found that increased expression of annexin A3 is a mechanism for platinum resistance in ovarian cancer cells. Here we demonstrate that annexin A3 can be detected in the culture medium of ovarian cancer cells, particularly these cells that express high levels of annexin A3. Levels of annexin A3 were then determined in sera from ovarian cancer patients using an enzyme‐linked immunosorbent assay. Compared with those from normal donors, sera from ovarian cancer patients contain significantly higher levels of annexin A3. Furthermore, serum levels of annexin A3 were significantly higher in platinum‐resistant patients than in platinum‐sensitive patients. To gain insight into the mechanism of secretion, the ovarian cancer cell lines were examined using both transmission electron microscopy and immunoelectron microscopy. Compared with parent cells, there are significantly more vesicles in the cytoplasm of ovarian cancer cells that express high levels of annexin A3, and at least some vesicles are annexin A3‐positive. Moreover, some vesicles appear to be fused with the cell membrane, suggesting that annexin A3 secretion may be associated with exocytosis and the release of exosomes. This is supported by our observation that ovarian cancer cells expressing higher levels of annexin A3 released increased numbers of exosomes. Furthermore, annexin A3 can be detected in exosomes released from cisplatin‐resistant cells (SKOV3/Cis) by immunoblotting and immunoelectron microscopy.

Mitochondrial comparative proteomics of human ovarian cancer cells and their platinum-resistant sublines.

Resistance to platinum‐based chemotherapy is the major obstacle to successful treatment of ovarian cancer. It is evident that mitochondrial defects and the dysfunctions of oxidative phosphorylation and energy production in ovarian cancer cells were directly related to their resistance to platinum drugs. Using 2‐D DIGE, we compared mitochondrial proteins from two platinum‐sensitive human ovarian cancer cell lines (SKOV3 and A2780) with that of four platinum‐resistant sublines (SKOV3/CDDP, SKOV3/CBP, A2780/CDDP, and A2780/CBP). Among the 236 differentially expressed spots, five mitochondrial proteins (ATP‐α, PRDX3, PHB, ETF, and ALDH) that participate in the electron transport respiratory chain were identified through mass spectrometry. All of them are downregulated in one or two of the platinum‐resistant cell lines. Three proteins (ATP‐α, PRDX3, and PHB) were validated by using western blot and immunohistochemistry. There is a significant decrease of PHB in tumor tissues from ovarian cancer patients who were resistant to platinum‐based chemotherapies. This is the first direct mitochondrial proteomic comparison between platinum‐sensitive and resistant ovarian cancer cells. These studies demonstrated that 2‐D DIGE‐based proteomic analysis could be a powerful tool to investigate limited mitochondrial proteins, and the association of PHB expression with platinum resistance indicates that mitochondria defects may contribute to platinum resistance in ovarian cancer cells.

Cell-cycle synchronization reverses Taxol resistance of human ovarian cancer cell lines

BackgroundTaxol is a powerful chemotherapy agent leading to mitotic arrest and cell death; however, its clinical efficacy has been hampered due to the development of drug resistance. Taxol specifically targets the cell cycle. Progress through mitosis (M stage) is an absolute requirement for drug-induced death because cell death is markedly reduced in cells blocked at the G1-S transition. The measured doubling time for ovarian cancer cells is about 27 h. As such, during treatment with Taxol most of the cells are not in the M stage of the cell cycle. Thus, the effect of cell-cycle synchronization was investigated in regard to reversing Taxol resistance in ovarian cancer cells.MethodsGiemsa-Wright staining was used for assessing the morphology of the cells. The doubling time of the cells was calculated using formula as follows: Td = In2/slope. The resistant index and cell cycle were measured via MTT assays and flow cytometry. Thymidine was used to induce cell-cycle synchronization, and cell apoptosis rates following exposure to Taxol were measured using a flow cytometer.ResultsThe growth doubling time of two Taxol-resistant cell lines were longer than that of Taxol-sensitive cells. Apoptotic rates in Taxol-sensitive and -resistant cell lines after synchronization and exposure to Taxol were all higher compared to unsynchronized controls (p <0.05).ConclusionsSynchronization of the cell-cycle resulted in an increased effectiveness of Taxol toward ovarian cancer cell lines. We speculated that formation of drug resistance toward Taxol in ovarian cancer could be partly attributed to the longer doubling time of these cells.

The target therapy of ovarian clear cell carcinoma

Clear cell adenocarcinoma (CCC) of the ovary accounts for 10% of epithelial ovarian cancer and is a distinct entity from other epithelial ovarian carcinomas. It arises from the endometriosis. CCC has specific biological and clinical behavior. Compared with other histological types, CCC shows a chemoresistant phenotype, which leads to poorer prognosis. Thus, development of new target-based therapies remains an unmet need for these patients. Mutations in the gene ARID1A have been found to occur in high frequency in CCC. The majority of these mutations lead to a loss of expression of the ARID1A protein, which is a subunit of the SWItch/Sucrose NonFermentable (SWI/SNF) chromatin remodeling complex and considered as a bona fide tumor suppressor. Upregulation of the PIK3/AKT/mTOR pathway, particularly through mutations of PIK3CA and inactivation of PTEN, is involved in tumorigenesis of CCC. Targeting angiogenesis, the Met protooncogene pathway, and HER2 are also discussed in this review.

Oncologic and reproductive outcomes after fertility‐sparing management with oral progestin for women with complex endometrial hyperplasia and endometrial cancer

To investigate the oncologic and reproductive outcomes after progestin treatment of complex endometrial hyperplasia (CEH) and grade 1 endometrial carcinoma (EC).

A survival analysis comparing women with ovarian low-grade serous carcinoma to those with high-grade histology

Ovarian low-grade serous carcinoma (LGSC) and high-grade serous carcinoma have distinct molecular profiles, clinical behaviors, and treatment responses. The survival advantage for patients with low-grade carcinoma compared with patients with high-grade histology remains controversial. We retrospectively reviewed the medical charts of 381 patients with ovarian serous carcinoma at Peking Union Medical College Hospital from 2007 to 2010. Patients were classified into two groups according to MD Anderson two-tier system: 35 (9.2%) cases with LGSC and 346 with high-grade serous carcinoma. Patients with low-grade serous ovarian cancer had a significantly younger age at diagnosis (46 versus 56 years, P=0.046), and their median progression-free survival (PFS) and overall survival values were 35.0 and 54.0 months, respectively. A multivariate analysis showed that, for serous ovarian cancer, the histological grade was a significant prognostic factor for PFS but not for overall survival (P=0.022 and P=0.0566, respectively). When stratified by the existence of a residual disease, patients with low-grade disease who underwent cytoreductive surgery without macroscopic residual disease (>1 cm) had a significantly improved median PFS time (36.0 months) compared with that of patients with high-grade carcinoma who received optimal cytoreductive surgery (16.0 months, P=0.017). Conversely, patients with low-grade and high-grade carcinoma who were left with macroscopic residue (>1 cm) experienced a similarly shorter median PFS (10.0 and 13.0 months, respectively, P=0.871). The International Federation of Gynecology and Obstetrics stage and residual disease were significant prognostic factors of low-grade carcinoma, while positive ascites was associated with a worse PFS value. Our data showed that LGSC is a different entity from high-grade carcinoma and that LGSC was associated with improved PFS after optimal cytoreductive surgery but not suboptimal operation.

Quantitative proteomic analysis of mitochondria from human ovarian cancer cells and their paclitaxel-resistant sublines.

Paclitaxel resistance is a major obstacle for the treatment of ovarian cancer. The chemoresistance mechanisms are partly related to the mitochondria. Identification of the relevant proteins in mitochondria will help in clarifying the possible mechanisms and in selecting effective chemotherapy for patients with paclitaxel resistance. In the present study, mitochondria from two paclitaxel‐sensitive human ovarian cancer cell lines (SKOV3 and A2780) and their corresponding resistant cell lines (SKOV3‐TR and A2780‐TR) were isolated. Guanidine‐modified acetyl‐stable isotope labeling and liquid chromatography‐hybrid linear ion trap Fourier‐transform ion cyclotron resonance mass spectrometry (LC‐FTICR MS) were performed to find the expressed differential proteins. Comparative proteomic analysis revealed eight differentially expressed proteins in the ovarian cancer cells and their paclitaxel‐resistant sublines. Among them, mimitin and 14‐3‐3 ζ/δ were selected for further research. The effects of mimitin and 14‐3‐3 ζ/δ were explored using specific siRNA interference in ovarian cancer cell lines and immunohistochemistry in human tissue specimens. The downregulation of mimitin and 14‐3‐3 ζ/δ using specific siRNA in paclitaxel‐resistant ovarian cancer cells led to an increase in the resistance index to paclitaxel. Multivariate analyses demonstrated that lower expression levels of the mimitin and 14‐3‐3 ζ/δ proteins were positively associated with shorter progression‐free survival (PFS) and overall survival (OS) in patients with primary ovarian cancer (mimitin: PFS: P = 0.041, OS: P = 0.003; 14‐3‐3 ζ/δ: PFS: P = 0.031, OS: P = 0.011). Mimitin and 14‐3‐3 protein ζ/δ are potential markers of paclitaxel resistance and prognostic factors in ovarian cancer.

The safety of postoperative hormone replacement therapy in epithelial ovarian cancer patients in China

Abstract Objective This study sought to determine whether postoperative hormone replacement therapy (HRT) has a negative influence on the progression-free and overall survival of epithelial ovarian carcinoma patients. Methods A retrospective chart review identified 77 patients with invasive epithelial ovarian cancer who had received HRT after primary surgical treatment from January 1995 to December 2010 at Peking Union Medical College Hospital. A 1 : 1 cohort of patients with the same diagnosis who did not receive HRT were matched by age and stage. An analysis of both progression-free survival and overall survival was performed using Cox proportional hazards models. Results According to the univariate analysis, HRT did not significantly influence progression-free or overall survival. Similarly, different types of HRT (estrogen alone, tibolone alone or an estrogen–tibolone combination) had no significant effect on the prognosis of epithelial ovarian cancer patients. The FIGO stage, differentiation, histological type and resection status were significantly correlated with progression-free survival and, except for histological type, these factors also significantly influenced overall survival. Finally, the multivariate analysis demonstrated that the strongest independent variable in predicting both progression-free survival and overall survival was the FIGO stage of the disease. Conclusion This study supports the hypothesis that postoperative HRT does not have a negative effect on the progression-free and overall survival of epithelial ovarian cancer patients. However, a multicenter study is needed to support and extend our findings.

Annexin A3 Is a Potential Predictor of Platinum Resistance in Epithelial Ovarian Cancer Patients in a Prospective Cohort.

Epithelial ovarian cancer (EOC) is the leading cause of death among gynecological malignancies and is rarely cured in the recurrent setting, mainly because of progressive chemoresistance, especially platinum resistance. In our previous studies, the platinum-resistance-related protein, annexin A3, was selected by comparative proteomics. In this study, we detected serum annexin A3 levels using a self-developed chemiluminescence immunoassay kit in a prospective EOC patient cohort. We also evaluated the capacity of serum annexin A3 levels to predict platinum resistance. Serum annexin A3 levels in healthy women exhibited a similar normal distribution (Z=0.723, P=0.673), allowing determination of a normal cutoff level of 0.11-1.45 ng/mL. Of the 89 EOC patients, 21 were platinum resistant and 68 were platinum sensitive. Residual disease after primary surgery (p=0.004) and serum annexin A3 levels (p=0.036) were both independent factors associated with platinum resistance. The AUC was 0.733 (95% confidence interval (CI), 0.627-0.823). The optimal cutoff value for serum annexin A3 levels was 2.05 ng/mL. Multivariate logistic analysis showed that expression of annexin A3 as assessed by immunohistochemistry (P=0.005) and residual tumor size (P=0.000) had a significant influence on platinum resistance. The AUC of ROC curve of annexin A3 expression by immunohistochemistry was 0.664 (95% CI, 0.554-0.763) and the cut off value was “>=moderate scores”. In conclusion, we demonstrate that annexin A3 is a secreted protein that may be measured in the peripheral blood using a self-developed, chemiluminescence immunoassay kit. Serum annexin A3 levels may be a potential predictor of platinum resistance in epithelial ovarian cancer patients.