Aldehyde dehydrogenase 1 family member A2 (ALDH1A2) is a rate-limiting enzyme involved in cellular retinoic acid synthesis. However, its functional role in ovarian cancer remains elusive. Here, we found that ALDH1A2 was the most prominently downregulated gene among ALDH family members in ovarian cancer cells, according to complementary DNA microarray data. Low ALDH1A2 expression was associated with unfavorable prognosis and shorter disease-free and overall survival for ovarian cancer patients. Notably, hypermethylation of ALDH1A2 was significantly higher in ovarian cancer cell lines when compared to that in immortalized human ovarian surface epithelial cell lines. ALDH1A2 expression was restored in various ovarian cancer cell lines after treatment with the DNA methylation inhibitor 5-aza-2′-deoxycytidine. Furthermore, silencing DNA methyltransferase 1 (DNMT1) or 3B (DNMT3B) restored ALDH1A2 expression in ovarian cancer cell lines. Functional studies revealed that forced ALDH1A2 expression significantly impaired the proliferation of ovarian cancer cells and their invasive activity. To the best of our knowledge, this is the first study to show that ALDH1A2 expression is regulated by the epigenetic regulation of DNMTs, and subsequently that it might act as a tumor suppressor in ovarian cancer, further suggesting that enhancing ALDH1A2-linked signaling might provide new opportunities for therapeutic intervention in ovarian cancer.

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Hormone receptor expression patterns often correlate with infiltration of specific lymphocytes in tumors. Specifically, the presence of specific tumor-infiltrating lymphocytes (TILs) with particular hormone receptor expression is reportedly associated with breast cancer, however, this has not been revealed in epithelial ovarian cancer (EOC). Therefore, we investigated the association between hormone receptor expression and TILs in EOC. Here we found that ERα, AR, and GR expression increased in EOC, while PR was significantly reduced and ERβ expression showed a reduced trend compared to normal epithelium. Cluster analysis indicated poor disease-free survival (DFS) in AR+/GR+/PR+ subgroup (triple dominant group); while the Cox proportional-hazards model highlighted the triple dominant group as an independent prognostic factor for DFS. In addition, significant upregulation of FoxP3+ TILs, PD-1, and PD-L1 was observed in the triple dominant group compared to other groups. NanoString analyses further suggested that tumor necrosis factor (TNF) and/or NF-κB signaling pathways were activated with significant upregulation of RELA, MAP3K5, TNFAIP3, BCL2L1, RIPK1, TRAF2, PARP1, and AKT1 in the triple dominant EOC group. The triple dominant subgroup correlates with poor prognosis in EOC. Moreover, the TNF and/or NF-κB signaling pathways may be responsible for hormone-mediated inhibition of the immune microenvironment.

The effect of hyperbaric oxygen is known to increase survival of ischemic tissue but its mechanism is not fully understood. The purpose of this study was to evaluate the effect of hyperbaric oxygen on a rat musculocutaneous flap versus ischemia-reperfusion injury, focusing on the mechanism involving the expression of adhesion molecules such as intercellular adhesion molecule-1 (ICAM-1) of endothelial cells and CD18 of neutrophils. A transverse rectus abdominis musculocutaneous (TRAM) flap (6 x 5 cm) supplied by a single superior epigastric vascular pedicle was elevated in 100 Sprague-Dawley rats. The rats were divided into 4 groups: group 0, sham (n = 10); group I, 4 hours of ischemia followed by reperfusion (n = 30); group II, 4 hours of ischemia and hyperbaric oxygen (100% oxygen, 2.5 atm absolute, during the last 90 minutes of ischemia before reperfusion) followed by reperfusion (n = 30); and group III, 4 hours of ischemia followed by reperfusion and hyperbaric oxygen (100% oxygen, 2.5 atm absolute, after reperfusion for 90 minutes; n = 30). The study consisted of gross examination for flap survival, histology, immunohistochemical staining, myeloperoxidase assay, flow cytometric study of CD18, and Northern blot analysis on ICAM-1 messenger ribonucleic acid expression. Gross measurement of the flap showed increased survival in groups II and III compared with group I (P < 0.05). The leukocytes adherent to the endothelium were counted at 24 hours and on day 5. Group I leukocytes were significantly increased compared with groups II and III (P < 0.05). The myeloperoxidase assay of TRAM flaps at 24 hours revealed a significant increase in group I compared with groups II and III (P < 0.05). The expression of CD18 was similar between groups I, II, and III. Immunohistochemical staining for ICAM-1 and Northern blot analysis on ICAM-1 messenger ribonucleic acid showed decreased expression in groups II and III compared with group I. Throughout the analysis, groups II and III did not show any significant differences. These results suggests that hyperbaric oxygen reduces the accumulation of leukocytes in the TRAM flap, but not enough to prevent adhesion of neutrophils on endothelial cells; ischemia-reperfusion injury increases the expression of CD18 and ICAM-1 and causes increased adhesion of leukocytes on the endothelium; hyperbaric oxygen does not alter the expression of CD18 but decreases the expression of ICAM-1; and the point of application for hyperbaric oxygen, whether applied before or after reperfusion, did not show any differences in outcome. In conclusion, the application of hyperbaric oxygen against ischemia-reperfusion injury increases flap survival and the beneficial effect may be explained by a protective mechanism involving downregulation of ICAM-1 on endothelial cells.

Background: Mitochondria are essential for energy conversation and cellular metabolism in eukaryotic cell. Mitochondria contain mitochondrial proteins that are encoded in the nucleus and synthesized in cytosol. Synthesized mitochondrial proteins are specially sorted by sub-mitochondrial compartment and then transported to their functional location. If mitochondrial proteins are not transported to their specific regions, it is reported that Mitochondria has a tendency to be implicated in various diseases including cancer. Mitochondria consist of mitochondrial outer membrane and inner membrane, especially, TOM complex are integrated in mitochondrial outer membrane. TOM40 is a central component of TOM complex has General import pore (GIP) to pass mitochondrial pre-protein. The goal of this study is to investigate the function of TOM40 in ovarian cancer. Methods: At first, we performed microarray to identify new target gene in ovarian cancer for nine human ovarian cancer cell line and seven human ovarian surface epithelial cell lines (HOSEs). We investigated target gene expression level in ovarian cancer cell line by RT-PCR and Western blot. To evaluate the association of target gene expression level and clinicopatholigical parameter in ovarian cancer, we carried out immunohistochemistry by using normal and ovarian cancer tissue. Second, we constructed stable DOV-13 and RMUG-S cell lines for overexpression and knock down of TOM40 protein using lentiviral system. We carried out proliferation, invasion and migration assay of each individual stable cell line. Results: We identified TOM40 as a new target gene for ovarian cancer by microarray data that was overexpressed in all nine human ovarian cancer cell lines than HOSEs. TOM40 protein level in borderline and cancer tissue was higher than normal tissue and benign tissue. TOM40 immuno-staining score of normal ovarian tissues is 1.23 (95% CI, 0.44-2.01) that compare with TOM40 immuno-staining score of benign, borderline, and epithelial ovarian tumors is 1.23 (95% CI, 0.44-2.01), 1.41 (95% CI, 0.45-2.37), 3.42 (95% CI, 2.65-4.20), and 5.04 (95% CI, 4.82-5.27), respectively. We understood TOM40 expression level of DOV-13 cells and RMUG-S cells is lower and higher than other ovarian cancer cell line. TOM40 overexpression in DOV-13 cell line inhibited the cell proliferation, invasion and migration than that of control cell line. TOM40 knock down in RMUG-S cell line induced cell proliferation, invasion and migration abilities than in control RMUG-S cell line. Conclusion: It is important to transport pathways of mitochondrial proteins, actually, the paper discussing directly association of TOM complex involving mitochondrial proteins transport and various diseases including cancer is not found. Our results suggest that TOM40 can be a role as putative tumor suppressor for ovarian cancer. Citation Format: Sol Kim, Hanbyoul Cho, Wookyeom Yang, Hyunja Kwon, Ha yeon Shin, Eunju Lee, Eun-Suk Kang, Jae-hoon Kim. Overexpression of TOM40 (translocase in the outer mitochondrial membrane 40) inhibits the cell proliferation, invasion and migration abilities in ovarian cancer cell lines. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2456. doi:10.1158/1538-7445.AM2014-2456

Background: S100A14 is an EF-hand containing calcium protein of the S100 protein family that exerts its biological effects on different types of cancers. The roles of S100A14 in tumorigenesis and its underlying mechanism have not been fully understood. This study aims to investigate the clinical significance and possible mechanisms of action of S100A14 in the growth and metastasis of epithelial ovarian cancer (EOC). Method: S100A14 expression in ovarian cancer was studied using a total of 104 ovarian tissue specimens (13 normal ovarian tissues, 10 benign ovarian tumors, 10 borderline ovarian tumors, and 71 EOC tissues) via immunohistochemistry. We investigated the possible functions of S100A14 in EOC stable cell by lentivirus-mediated overexpression and short hairpin RNA (shRNA). Results: S100A14 protein was significantly over-expressed in EOC cells and tissues. S100A14 expression was significantly associated with advanced stage (P&lt;0.001), serous type (P=0.004), and poor tumor grade (P&lt;0.001). Patients with S100A14 overexpression had shorter overall (Hazard Ratio = 4.53 [1.16-17.69], P=0.029) and disease-free survival (Hazard Ratio = 3.10 [1.22-7.89], P=0.017) in multivariate analysis. In vitro analysis of cell function revealed that cell proliferation, migration and invasion were regulated by S100A14. We investigated the underlying mechanism and found that the phosphorylation levels of AKT have increased after S100A14 overexpression. Inhibition of AKT by a specific AKT inhibitor (MK-2206) and PI3K inhibitor (wortmannin) at least partly reversed the AKT-phosphorylation events in S100A14-overexpressed cells. Conclusions: The data revealed the aberrant expression of S100A14 in ovarian cancer, suggesting a potentially important role of S100A14 in ovarian carcinogenesis; S100A14 regulated AKT via phosphorylation of its ser473 site also implies that S100A14 could be used as a novel therapeutic target for ovarian cancer treatment. Citation Format: Hanbyoul Cho, Maria Lee, Ha-Yeon Shin, Wookyeom Yang, Eun-ju Lee, Hyunja Kwon, Sol Kim, Jae-Hoon Kim. S100A14 plays an important role in cell growth and metastasis of epithelial ovarian cancer through PI3K-AKT pathway. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2857. doi:10.1158/1538-7445.AM2014-2857

Background Gene expression pattern analysis comparing normal cells with cancer cells are performed by many competitors in the worldwide lab. Cancer has many different species in itself due to its individual differences, variations and heterogeneity. Therefore, it is necessary to spend more time in gene expression pattern analysis using microarray or whole genome sequencing. The purpose of this study was to identify genes and proteins that are highly differentially expressed in newly established epithelial ovarian cancer (EOC) cell lines, and to use this knowledge for the development of novel diagnostic and prognostic markers for EOC. We further performed the validation studies of several candidate genes. Results Comparison of gene expression patterns using microarray analysis enabled us to identify 84 genes that were commonly up-regulated and 132 genes that were down-regulated in cancer cell lines (up; &gt; 2-fold, down; &lt; 3-fold, P &lt; 0.05). In 2-DE and MALDI-TOF/PMF, 31 up-regulated spots that had at least two-fold differences between the 3 EOC cell lines and HOSE cells used as controls were observed. 15 up-regulated genes in cell cycle related genes and 9 down-regulated genes in apoptosis related confirmed gene expression using quantitative RT-PCR. Pathway analysis was validated in cell cycle related genes in six EOC cell lines. Conclusions The data shows the aberrant expression of several cell cycle related genes in EOC and that Cyclin-dependent kinase 1 (CDK1) pathway may play a significant role in ovarian carcinogenesis, implying that CDK1 pathway could be used as a novel therapeutic target for ovarian cancer treatment. Citation Format: wookyeom yang, hanbyoul Cho, hayeon Shin, eunju Lee, hyunja Kwon, sol Kim, jaehoon Kim. Molecular biological characterization of newly established epithelial ovarian cancer (EOC) cell lines : The cyclin-dependent kinase 1 pathway. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2369. doi:10.1158/1538-7445.AM2014-2369

Background: Reptin is also known as RuvBl2, Tip48, TIP49b, ECP51 and CGI-46, is a member of AAA+ (ATPases associated with diverse cellular activities) superfamily. Reptin is associated with cell viability and development of S. cerevisiae, D. melanogaster, X. laevis, or D. rerio has a lot of functions including transcriptional regulation, DNA damage repair and telomerase activity. Self-reactive IgG autoantibodies are present on human sera.Each self-reactive IgG autoantibody derived from pre- and post-treatment of ovarian cancer patients is competitively combined with cancer tissue gotten during the operation, self-reactive IgG autoantibodies bound to tumor antigens. As a result, self-reactive IgG autoantibodies may be decreased after treatment. In this study, we have identified and investigated the function of Reptin as an available therapeutic biomarker for ovarian cancer using difference of self-reactive IgG autoantibody amounts. Method: We carried out the two-dimensional differential gel electrophoresis analysis of immunoprecipitated tumor antigens (2D-DITA) to compare with expression level of autoantibodies in pre- and post-treatment sera of 14 ovarian cancer patients. Autoantibody differentiated between pre- and post-treatment sera was identified by MALDI-TOF. We performed qRT-PCR with 8 human ovarian surface epitheliums (HOSEs) and 14 ovarian cancer cell lines for validation studies. Reptin was also detected in other ovarian cancer tissues by immunohistochemistry (IHC). We constructed SKOV-3 cell line for overexpression Reptin using lentiviral system. For investigating function of Reptin, we performed proliferation, wound healing, and invasion assay. Results: We identified Reptin as a tumor-associated antigen of epithelial ovarian cancer. Relative mRNA level of Reptin showed higher levels in ovarian cancer cell lines than in HOSEs. Reptin protein levels were significantly upregulated in ovarian cancer tissues than normal, benign, and borderline ovarian tumors. In SKOV-3 cells, overexpression of Reptin induced inhibition of cell proliferation (P&lt;0.0001), migration and invasion (P&lt;0.0001). Conclusion: Our results demonstrate that Reptin is a tumor -associated antigen of ovarian cancer, i.e. tumor suppressor. Citation Format: Eun-ju Lee, Hanbyoul Cho, Ha-Yeon Shin, Wookyeom Yang, Hyunja Kwon, Sol Kim, Jae-Hoon Kim. Reptin as a tumor-associated antigen of epithelial ovarian cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1864. doi:10.1158/1538-7445.AM2014-1864