Human Ovarian Adenocarcinoma Research Articles

The effect of endostatin mediated by human mesenchymal stem cells on ovarian cancer cells in vitro

Endostatin is the most potent inhibitor of tumor angiogenesis. However, endostatin protein has a short half-time and virus-mediated endostatin gene therapy has serious toxicity, which limits the application of endostatin in clinical therapy. Mesenchymal stem cells (MSCs) are considered to be able to accumulate at the site of cancers with high specificity and may be used as a new delivery of endostatin. The MSCs from the human bone marrow were transfected with recombinant adenovirus encoding endostatin and EGFP (MSC-EN cells). The tropism capacity of MSCs was quantitatively assayed in vitro using the Millicell system. To investigate the impact of secreted endostatin on cancer cells, SKOV3 cells were co-cultured with MSC-EN cells in Millicell for 48 h, then apoptosis and cell cycle were analyzed on a flow cytometer. In contrast with 293 cells and saline, SKOV3 cells significantly stimulated migration of MSCs, the number reached 919.67 +/- 19.96 (P < 0.05). The endostatin produced by MSC-EN cells made 13.08 +/- 0.21% SKOV3 cells undergo early stage apoptosis (control 3.23 +/- 0.73%, P < 0.05) and 82.05 +/- 2.65% SKOV3 cells accumulate in the G0/G1 phase (control 66.51 +/- 2.91%, P < 0.05). We found that MSCs possessed great migratory capacity in vitro and the human ovarian adenocarcinoma cell line SKOV3 could significantly induce the migration of MSCs. Our results provided evidence that MSCs could be utilized as a powerful delivery system of endostatin. The endostatin produced by MSC-EN cells could inhibit the proliferation of SKOV3 cells.

HOXA10 is Overexpressed in Human Ovarian Clear Cell Adenocarcinoma and Correlates With Poor Survival

Human homeobox gene (HOX) A10 is a homeobox allotype gene of the HOXA family in the HOX family. Human homeobox gene A10 may play an important role in cancer development. However, the role of HOXA10 in the carcinogenesis of ovarian clear cell adenocarcinoma (OCCA) has not been established. We have evaluated the prognostic significance of HOXA10 expression for human OCCA and the effects of HOXA10 on proliferation, motility, and invasion of OCCA cells. We found that HOXA10 was not expressed in normal ovarian epithelium, ovarian endometrial cysts, and ovarian serous carcinomas, but 20 (68.9%) of the 29 OCCAs were positive for the expression of HOXA10. Human homeobox gene A10 expression was negatively correlated to the 5-year survival of OCCA patients (R = -0.442, P = 0.043). When a HOXA10 expression vector was stably transfected into a human OCCA cell line, ES-2, the proliferation rate of ES-2-HOXA10 was much higher than the vector control, the motility of ES-2-HOXA10 cells was significantly increased compared with the control (P < 0.05), and the invasion of ES-2-HOXA10 cells was also much higher than the vector control (P < 0.01 at 5 hours and 12 hours after scratching). In conclusion, HOXA10 was overexpressed in OCCA and was correlated with poor survival. HOXA10 promotes proliferation, migration, and invasion of OCCA cells. Human homeobox gene A10 could be a promising prognostic marker for OCCA.

IRF9 is a Key Factor for Eliciting the Antiproliferative Activity of IFN-α

A number of tumors are still resistant to the antiproliferative activity of human interferon (IFN)-alpha. The Janus kinases/Signal Transducers and Activators of Transcription (JAK-STAT) pathway plays an important role in initial IFN signaling. To enhance the antiproliferative activity of IFN-alpha, it is important to elucidate which factors in the JAK-STAT pathway play a key role in eliciting this activity. In human ovarian adenocarcinoma OVCAR3 cells sensitive to both IFN-alpha and IFN-gamma, only IFN regulatory factor 9 (IRF9)-RNA interference (RNAi) completely inhibited the antiproliferative activity of IFN-alpha among the intracellular JAK-STAT pathway factors. Conversely, Stat1-RNAi did not inhibit the antiproliferative activity of IFN-alpha, whereas it partially inhibited that of IFN-gamma. As a cell death pathway, it is reported that tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptosis through TRAIL-receptor (R) 1 and TRAIL-R2. In IFN-alpha-treated OVCAR3 cells, IRF9-RNAi inhibited transcription of TRAIL whereas Stat1-RNAi did not, suggesting that the transcription of TRAIL induced by IFN-alpha predominantly required IRF9. Furthermore, IFN-stimulated response element-like motifs of TRAIL bound to IFN-stimulated gene factor 3 (ISGF3) complex after IFN-alpha treatment. Subsequently, TRAIL-R2-RNAi inhibited both antiproliferative activities of IFN-alpha and TRAIL, suggesting that TRAIL-R2 mediated both IFN-alpha and TRAIL signals to elicit their antiproliferative activities. Finally, IRF9 overexpression facilitated IFN-alpha-induced apoptosis in T98G (human glioblastoma multiforme) cells, which were resistant to IFN-alpha. Thus, this study suggests that IRF9 is the key factor for eliciting the antiproliferative activity of IFN-alpha and TRAIL may be one of the potential mediators.

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.

Endogenous Human CaMKII Inhibitory Protein Suppresses Tumor Growth by Inducing Cell Cycle Arrest and Apoptosis through Down-regulation of the Phosphatidylinositide 3-Kinase/Akt/HDM2 Pathway

Inhibition of calcium/calmodulin-dependent protein kinase II (CaMKII) results in hypophosphorylation of CaMKII substrates and in some cases suppresses cell growth. We previously presented the first report of the human CaMKII inhibitory protein, hCaMKIINbeta. Here we report the functional characterization of hCaMKIINbeta in ovarian cancer cells. We showed that hCaMKIINbeta was highly expressed in normal ovarian tissues but was not detected in human ovarian adenocarcinoma, indicating that decreased expression of hCaMKIINbeta may be involved in the pathogenesis of human ovarian adenocarcinoma. As an endogenous CaMKII inhibitor, hCaMKIINbeta could significantly inhibit the growth of human ovarian cancer cells in vitro. In vivo, hCaMKIINbeta decreased the tumorigenicity and growth of HO-8910PM human ovarian cancer cells and prolonged the survival of tumor-bearing mice. hCaMKIINbeta blocked cell cycle progression and induced apoptosis of HO-8910PM cells, which was correlated with the up-regulation of p21, p53, and Bax and the down-regulation of cyclin A, cyclin D1, cyclin E, CDK2, phosphorylated retinoblastoma, and Bcl-2. We further demonstrated that hCaMKIINbeta-mediated CaMKII inhibition suppressed Akt activation, leading to the down-regulation of HDM2, which was responsible for the up-regulation of p53 and p21 in human ovarian cancer cells. The tumor-suppressive effect and the negative expression in human ovarian cancer tissues suggest that hCaMKIINbeta may play an important role in the regulation of tumor cell growth, possibly contributing to the development of new therapeutic strategies for ovarian cancer.

1042 Human homeobox gene (HOX) A10 is overexpressed in human ovarian clear cell adenocarcinoma and correlates with poor survival

Heat shock protein 60 (HSP60) participates in protein assembly, folding and transport. Increased HSP60 mRNA expression is associated with cisplatin resistance in some in vitro models and with shorter survival among ovarian cancer patients. HSP60 mRNA expression was quantitated in three independent model systems by reverse-transcription PCR (A2780 human ovarian carcinoma cell line and sublines selected for cisplatin or oxaliplatin resistance in vitro and also in the UCRU-BL13 human bladder carcinoma cell line and a cisplatin-resistant subline). Increased HSP60 mRNA expression was observed in all resistant sublines (range 2.5–15 fold), correlated with relative resistance to cisplatin and oxaliplatin. No differences in HSP60 gene copy number were apparent in resistant sublines. These data provide further evidence of a strong association between in vitro resistance to platinum compounds and increased HSP60 mRNA expression.

Polyethylene glycol-complexed cationic liposome for enhanced cellular uptake and anticancer activity

Liposomes as one of the efficient drug carriers have some shortcomings such as their relatively short blood circulation time, fast clearance from human body by reticuloendothelial system (RES) and limited intracellular uptake to target cells. In this study, polyethylene glycol (PEG)-complexed cationic liposomes (PCL) were prepared by ionic complex of cationically charged liposomes with carboxylated polyethylene glycol (mPEG-COOH). The cationic liposomes had approximately 98.6 ± 1.0 nm of mean particle diameter and 45.5 ± 1.1 mV of zeta potential value. While, the PCL had 110.1 ± 1.2 nm of mean particle diameter and 18.4 ± 0.8 mV of zeta potential value as a result of the ionic complex of mPEG-COOH with cationic liposomes. Loading efficiency of model drug, doxorubicin, into cationic liposomes or PCL was about 96.0 ± 0.7%. Results of intracellular uptake evaluated by flow cytometry and fluorescence microscopy studies showed higher intracellular uptake of PCL than that of Doxil ®. In addition, in vitro cytotoxicity of PCL was comparable to cationic liposomes. In pharmacokinetic study in rats, PCL showed slightly lower plasma level of DOX than that of Doxil ®. In vivo antitumor activity of DOX-loaded PCL was comparable to that of Doxil ® against human SKOV-3 ovarian adenocarcinoma xenograft rat model. Consequently, the PCL, of which surface was complexed with PEG by ionic complex may be applicable as drug delivery carriers for increasing therapeutic efficacy of anticancer drugs.

Radiolabeled 5-Iodo-3′-O-(17β-succinyl-5α-androstan-3-one)-2′-deoxyuridine and Its 5′-Monophosphate for Imaging and Therapy of Androgen Receptor-Positive Cancers: Synthesis and Biological Evaluation

High levels of androgen receptor (AR) are often indicative of recurrent, advanced, or metastatic cancers. These conditions are also characterized by a high proliferative fraction. 5-Radioiodo-3'-O-(17beta-succinyl-5alpha-androstan-3-one)-2'-deoxyuridine 8 and 5-radioiodo-3'-O-(17beta-succinyl-5alpha-androstan-3-one)-2'-deoxyuridin-5'-yl monophosphate 13 target AR. They are also degraded intracellularly to 5-radioiodo-2'-deoxyuridine 1 and its monophosphate 20, respectively, which can participate in the DNA synthesis. Both drugs were prepared at the no-carrier-added level. Precursors and methods are readily adaptable to radiolabeling with various radiohalides suitable for SPECT and PET imaging, as well as endoradiotherapy. In vitro and in vivo studies confirm the AR-dependent interactions. Both drugs bind to sex hormone binding globulin. This binding significantly improves their stability in serum. Biodistribution and imaging studies show preferential uptake and retention of 8 and 13 in ip xenografts of human ovarian adenocarcinoma cells NIH:OVCAR-3, which overexpress AR. When these drugs are administered at therapeutic dose levels, a significant tumor growth arrest is observed.

Chitosan–phospholipid blend for sustained and localized delivery of docetaxel to the peritoneal cavity

Localized and sustained delivery of chemotherapeutics presents a “magic bullet” effect by providing high drug concentrations at the target site, extended drug exposure and reduced systemic toxicity. In the present study, an injectable chitosan–phospholipid (PoLi gel) blend is put forth as a strategy to achieve sustained and localized delivery of docetaxel (DTX) following intraperitoneal (IP) administration. The stability of the blend was confirmed in vitro, by turbidity measurements and attributed to specific molecular interactions and the organization of the materials within the blend, as evidenced by FTIR analysis and confocal laser scanning microscopy, respectively. The chitosan and phospholipid were found to colocalize in regions surrounding a mean object area of 11.2 μm 2 with colocalization coefficients of 43% and 46% for the chitosan and phospholipid, respectively. The PoLi gel blend afforded sustained drug release as seen both in vitro (2.4 ± 0.7% DTX per day) and in vivo (4.4 ± 0.7% DTX per day). Constant concentrations of DTX were observed over a 2-week period in plasma and relevant peritoneal tissues, with no signs of toxicity or inflammation, following IP administration of the blend in healthy CD-1 mice. At DTX doses of 28.8 and 19.2 mg/kg, the blend showed significant tumor inhibition of 87.3 ± 9.3% and 74.1 ± 25.9%, respectively, in a murine xenograft model of human ovarian adenocarcinoma. This localized delivery system has shown excellent potential for sustained IP treatment of cancers, such as ovarian, that reside in the peritoneal cavity.

Coadministration of Paclitaxel and Curcumin in Nanoemulsion Formulations To Overcome Multidrug Resistance in Tumor Cells

Development of multidrug resistance (MDR) against a variety of conventional and novel chemotherapeutic agents is a significant challenge in effective cancer therapy. Over the last several years, we have focused on a multimodal therapeutic strategy to overcome tumor MDR by enhancing the delivery efficiency to the tumor mass and lowering the apoptotic threshold by modulation of the intracellular signaling mechanisms. In this study, we have examined augmentation of therapeutic efficacy upon coadministration of paclitaxel (PTX) and curcumin (CUR), an inhibitor of nuclear factor kappa B (NFkappaB) as well as a potent down-regulator of ABC transporters, in wild-type SKOV3 and drug resistant SKOV3(TR) human ovarian adenocarcinoma cells. PTX and CUR were encapsulated in flaxseed oil containing nanoemulsion formulations. The results showed that the encapsulated drugs were effectively delivered intracellular in both SKOV3 and SKOV3(TR) cells. CUR administration was shown to inhibit NFkappaB activity and down regulate P-glycoprotein expression in resistant cells. Combination PTX and CUR therapy, especially when administered in the nanoemulsion formulations, was very effective in enhancing the cytotoxicity in wild-type and resistant cells by promoting the apoptotic response. Overall, this cotherapy strategy has significant promise in the clinical management of refractory diseases, especially in ovarian cancer.

Combination chemotherapy of oxaliplatin and 5‐fluorouracil may be an effective regimen for mucinous adenocarcinoma of the ovary: A potential treatment strategy

Resistance of ovarian mucinous adenocarcinoma to standard chemotherapy with paclitaxel and carboplatin is associated with poor prognosis, and an effective treatment is needed. The present study aimed to identify an effective chemotherapy for ovarian mucinous adenocarcinoma. Five human ovarian mucinous adenocarcinoma cell lines (MN-1, OMC-1, RMUG-L, RMUG-S, TU-OM-1) were used in this study. Sensitivity of the cells to the anticancer agents was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, and we assessed drug sensitivity by calculating the assay area under the curve for each agent. Protein expression was confirmed by Western blot analysis. We also examined the efficacy of combination chemotherapy on survival in a xenograft model of nude mice. The IC(50) to anticancer agents ranged widely. The assay area under the curve indicated that two of five cell lines (MN-1, TU-OM-1) were sensitive to oxaliplatin, 5-fluorouracil and etoposide, and only one (TU-OM-1) was sensitive to 7-ethyl-10-hydroxycamptothecin, which is an active metabolite of camptothecin. All cell lines were resistant to cisplatin and paclitaxel. The combination of oxaliplatin and 5-fluorouracil resulted in additive or synergistic effects on all cell lines. The combination of oxaliplatin and 5-fluorouracil significantly prolonged survival in a ovarian mucinous adenocarcinoma xenograft model of nude mice. Protein expression levels of the excision repair cross-complementation group 1 were lower in oxaliplatin sensitive cell lines. Exposure to 5-fluorouracil down-regulated cross-complementation group 1 expression in ovarian mucinous adenocarcinoma cells. We conclude that combination chemotherapy consisting of oxaliplatin and 5-fluorouracil was an effective treatment for ovarian mucinous adenocarcinoma and may be a pivotal candidate for a novel treatment strategy.

One- and two-photon activated phototoxicity of conjugated porphyrin dimers with high two-photon absorption cross sections

Two-photon excited photodynamic therapy (PDT) has the potential to provide a highly targeted treatment for neoplastic diseases, as excitation can be pin-pointed to small volumes at the laser focus. In addition, two-photon PDT offers deeper penetration into mammalian tissue due to the longer wavelength of irradiation. Here we report the one-photon and two-photon excited PDT results for a collection of conjugated porphyrin dimers with high two-photon absorption cross sections. These dimers demonstrate high one-photon PDT efficacy against a human ovarian adenocarcinoma cell line (SK-OV-3) and exhibit no significant dark-toxicity at concentrations of up to 20 microM. Their one-photon excited PDT efficiencies, following irradiation at 657 nm, approach that of Visudyne, a drug used clinically for PDT. We investigated and optimised the effect of the photosensitizer concentration, incubation time and the light dose on the PDT efficacy of these dimers. These studies led to the selection of P2C2-NMeI as the most effective porphyrin dimer. We have demonstrated that P2C2-NMeI undergoes a two-photon activated process following excitation at 920 nm (3.6-6.8 mW, 300 fs, 90 MHz) and compared it to Visudyne. We conclude that the in vitro two-photon PDT efficacy of P2C2-NMeI is about twice that of Visudyne. This result highlights the potential of this series of porphyrin dimers for two-photon PDT.

A 2-substituted prodiginine, 2-(p-hydroxybenzyl)prodigiosin, from Pseudoalteromonas rubra.

In the course of work aimed at the discovery of new pharmaceutical lead compounds from marine bacteria, a lipophilic extract of the bacterium Pseudoalteromonas rubra displayed significant cytotoxicity against SKOV-3, a human ovarian adenocarcinoma cell line. Bioassay-directed fractionation of this extract resulted in the isolation of a series of known and new prodiginine-type azafulvenes. The structure of the major metabolite was elucidated by interpretation of spectroscopic data as a 2-substituted prodigiosin, which we named 2-(p-hydroxybenzyl)prodigiosin (HBPG).

Establishment and Characterization of the RMG-V Cell Line from Human Ovarian Clear Cell Adenocarcinoma

A cell line, designated as RMG-V, was established from a patient with clear cell adenocarcinoma of the ovary. The cell line has grown without interruption and has been propagated continuously by serial passaging (more than 36 times) over 5 years. The cells are spindle-shaped, display neoplastic and pleomorphic features, and grow in a jigsaw puzzle-like arrangement while forming monolayers without contact inhibition. These cells proliferate rapidly, and the population doubling time is about 15.5 hours. The number of chromosomes ranges between 77 and 85, with a modal number of 83.

3 Roles of JAK-STAT pathway factors in antiproliferative activities of human IFN-α and IFN-γ: A comprehensive assessment in human ovarian adenocarcinoma OVCAR3 cells

3 Roles of JAK-STAT pathway factors in antiproliferative activities of human IFN-α and IFN-γ: A comprehensive assessment in human ovarian adenocarcinoma OVCAR3 cells

Evaluations of combination MDR-1 gene silencing and paclitaxel administration in biodegradable polymeric nanoparticle formulations to overcome multidrug resistance in cancer cells

In this study, the effect of MDR-1 gene silencing, using small interfering RNA (siRNA), and paclitaxel (PTX) co-therapy in overcoming tumor multidrug resistance was examined. Poly(ethylene oxide)-modified poly(beta-amino ester) (PEO-PbAE) and PEO-modified poly(epsilon-caprolactone) (PEO-PCL) nanoparticles were formulated to efficiently encapsulate MDR-1 silencing siRNA and PTX, respectively. Upon administration in multidrug resistant SKOV3(TR) human ovarian adenocarcinoma cells, siRNA-mediated MDR-1 gene silencing was evident at 100 nM dose. Combination of MDR-1 gene silencing and nanoparticle-mediated delivery significantly influenced the cytotoxic activity of PTX in SKOV3(TR) cells similar to what was observed in drug sensitive SKOV3 cells. We speculate that the enhancement in cytotoxicity was due to an increase in intracellular drug accumulation upon MDR-1 gene silencing leading to an apoptotic cell-kill effect. Taken together, these preliminary results are highly encouraging for the development of combination nano-therapeutic strategies that combine gene silencing and drug delivery to provide more potent therapeutic effect, especially in refractory tumors.

Adaptor protein Crk is implicated in mucus formation in mucinous epithelial ovarian cancer (mEOC) cells MCAS

The mucus production is an indicator for the histological grade of mucinous epithelial ovarian cancer (mEOC). In our previous study, Crk expression was targeted in the human ovarian mucinous adenocarcinoma cell line MCAS through RNA interference, resulting in the establishment of Crk knock down cells. These cells exhibited decreased tumorigenic potential both in vitro and in vivo. The purpose of this study was to investigate if there is any change in the capability of forming mucus in these Crk knock down cells. Cytoplasmic periodic acid Schiff (PAS) staining and particle excluding assay were conducted to assess the mucus formation within and around cells, respectively. Additionally, the amount of mucus formed in tumor lumps from nude mice model was measured following HE and PAS staining. The increased mucus production in Crk knockdown mEOC cells (MCAS) was manifested by increased number of enlarged cells filled with vacuoles-like mucus observed by phase-contrast microscope and cytoplasmic PAS staining; and enhanced mucus secretion was represented by the assembly of pericellular matrix in particle excluding assay and increased mucus area in tumor lumps from nude mice models. The course of carcinogenesis in mEOC is associated with the altered pattern of mucus production and secretion. The adaptor protein Crk is implicated in both pathways.

Modulation of Drug Resistance in Ovarian Adenocarcinoma by Enhancing Intracellular Ceramide Using Tamoxifen-Loaded Biodegradable Polymeric Nanoparticles

To modulate intracellular ceramide levels and lower the apoptotic threshold in multidrug-resistant ovarian adenocarcinoma, we have examined the efficacy and preliminary safety of tamoxifen coadministration with paclitaxel in biodegradable poly(ethylene oxide)-modified poly(epsilon-caprolactone) (PEO-PCL) nanoparticles. In vitro cytotoxicity and proapoptotic activity of paclitaxel and tamoxifen, either as single agent or in combination, was examined in wild-type (SKOV3) and MDR-1-positive (SKOV3TR) human ovarian adenocarcinoma cells. Subcutaneous SKOV3 and SKOV3TR xenografts were established in female nu/nu mice, and this model was used to evaluate the antitumor efficacy and preliminary safety. Paclitaxel (20 mg/kg) and tamoxifen (70 mg/kg) were administered i.v. either as a single agent or in combination in aqueous solution and in PEO-PCL nanoparticles. In vitro cytotoxicity results showed that administration of paclitaxel and tamoxifen in combination lowered the IC50 of paclitaxel by 10-fold in SKOV3 cells and by >3-fold in SKOV3TR cells. The combination paclitaxel/tamoxifen co-therapy showed even more pronounced effect when administered in nanoparticle formulations. Upon i.v. administration of paclitaxel/tamoxifen combination in PEO-PCL nanoparticle formulations, significant enhancement in antitumor efficacy was observed. Furthermore, the combination paclitaxel/tamoxifen therapy did not induce any acute toxicity as measured by body weight changes, blood cell counts, and hepatotoxicity. The results of this study show that combination of paclitaxel and tamoxifen in biodegradable PEO-PCL nanoparticles can serve as an effective clinically translatable strategy to overcome multidrug resistance in ovarian cancer.

CAMP response element–binding protein is expressed at high levels in human ovarian adenocarcinoma and regulates ovarian tumor cell proliferation

Approximately 90% of human ovarian tumors result from transformation of ovarian surface epithelial cells. It has been hypothesized that repeated destruction of the epithelial cells during ovulation, followed by proliferation and migration of epithelial cells to restore the ovarian surface, renders these cells susceptible to mutagenic events. One of the proteins found to promote ovarian surface epithelial cell survival and proliferation was the transcription factor, cAMP response element-binding protein (CREB). Thus, the objective of this study was to determine whether CREB was also highly expressed in tumor cells originating from the ovarian epithelium. Using an ovarian cancer tissue array, it was observed that approximately 54% of the epithelial-derived human ovarian tumors displayed moderate or high levels of CREB immunostaining, while none of the normal ovarian samples did. Comparison of CREB levels in a human ovarian tumor cell line to those of a normal ovarian epithelial cell line revealed elevated levels of CREB and phosphorylated CREB in the ovarian tumor cells. To determine whether CREB regulated proliferation and/or apoptosis in the ovarian tumor cell line, CREB expression was suppressed using RNA interference. Decreased CREB expression significantly reduced ovarian tumor cell proliferation, while there was no effect on apoptosis in these cells. Finally, we showed that CREB is highly expressed in an in vivo murine model of ovarian tumorigenesis. Therefore, CREB is frequently overexpressed in ovarian cancer where it appears to promote cell proliferation.

Human monoclonal antibody for ovarian clear cell carcinoma-2, a human monoclonal antibody with antitumor activity against ovarian cancer cells that recognizes CA125-like antigen

In recent years, antibody therapy employing monoclonal antibodies has become a new approach for treating cancer. This study was performed to establish a human monoclonal antibody recognizing an epitope related to CA125 using KM mice and to assess its reactivity with ovarian cancer cells. A human ovarian clear cell adenocarcinoma cell line (RMG-I) was used to immunize KM mice, and hybridoma supernatant was obtained by a standard method employing enzyme-linked immunosorbent assay screening. Next, selection of hybridomas was performed with two antibodies (MA602-1 and MA602-6) and a sandwich immunoassay for CA125-like antigen, and then the limiting dilution was used to obtain a human monoclonal antibody. Immunohistochemical reactivity of this antibody (human monoclonal antibody for ovarian clear cell carcinoma-2 [HMOCC-2]) with ovarian cancer was assessed, while its specificity was analyzed by Western blotting. Various antibodies were used to identify the epitope targeted by HMOCC-2. Finally, the antitumor effect of HMOCC-2 was assessed by intraperitoneal administration to SCID (severe combined immunodeficiency) mice with heterografts of RMG-I tumors. HMOCC-2 showed a positive reaction with 60% (63/105) of ovarian cancer specimens. Western blotting of the membrane fraction of RMG-I revealed several bands at 120-250 kd. HMOCC-2 recognized the CA125-like antigens identified by several antibodies. HMOCC-2 also exhibited significant antitumor activity (P < 0.01) against ovarian cancer heterografts. HMOCC-2 reacts specifically with ovarian cancer cells via a target epitope analogous to that of CA125 and also exhibits activity against ovarian tumors. These findings suggest that it may have the potential to be employed clinically for molecular-targeting therapy.