Human Ovarian Cancer Lines Research Articles

NK-92MI Cells Engineered with Anti-claudin-6 Chimeric Antigen Receptors in Immunotherapy for Ovarian Cancer.

Background: The application of chimeric antigen receptor (CAR) NK cells in solid tumors is hindered by lack of tumor-specific targets and inefficient CAR-NK cell efficacy. Claudin-6 (CLDN6) has been reported to be overexpressed in ovarian cancer and may be an attractive target for CAR-NK cells immunotherapy. However, the feasibility of using anti-CLDN6 CAR-NK cells to treat ovarian cancer remains to be explored. Methods: CLDN6 expression in primary human ovarian cancer, normal tissues and cell lines were detected by immunohistochemistry and western blot. Two types of third-generation CAR NK-92MI cells targeting CLDN6, CLDN6-CAR1 NK-92MI cells with domains containing self-activated elements (NKG2D, 2B4) and CLDN6-CAR2 NK-92MI cells with classical domains (CD28, 4-1BB) were constructed by lentivirus transfection, sorted by flow cytometry and verified by western blot and qPCR. OVCAR-3, SK-OV-3, A2780, Hey and PC-3 cells expressing the GFP and luciferase genes were transduced. Subcutaneous and intraperitoneal tumor models were established via NSG mice. The ability of CLDN6-CAR NK cells to kill CLDN6-positive ovarian cancer cells were evaluated in vitro and in vivo by live cell imaging and bioluminescence imaging. Results: Both CLDN6-CAR1 and CLDN6-CAR2 NK-92MI cells could specifically killed CLDN6-positive ovarian cancer cells (OVCAR-3, SK-OV-3, A2780 and Hey), rather than CLDN6 negative cell (PC-3), in vitro. CLDN6-CAR1 NK-92MI cells with domains containing self-activated elements (NKG2D, 2B4) exhibited stronger cytotoxicity than CLDN6-CAR2 NK-92MI cells with classical domains (CD28, 4-1BB). Furthermore, CLDN6-CAR1 NK cells could effectively eliminate ovarian cancer cells in subcutaneous and intraperitoneal tumor models. More importantly, CAR-NK cells combined with immune checkpoint inhibitors, anti-PD-L1, could synergistically enhance the antitumor efficacy of CLDN6-targeted CAR-NK cells. Conclusions: These results indicate that CLDN6-CAR NK cells possess strong antitumor activity and represent a promising immunotherapeutic modality for ovarian cancer.

Pharmacological tumor PDL1 depletion with chlorambucil treats ovarian cancer and melanoma: improves antitumor immunity and renders anti-PDL1-resistant tumors anti-PDL1-sensitive through NK cell effects

BackgroundTumor intracellular programmed cell death ligand-1 (PDL1) mediates pathologic signals that regulate clinical treatment responses distinctly from surface-expressed PDL1 targeted by αPDL1 immune checkpoint blockade antibodies.MethodsWe performed a drug screen...

Liposomal paclitaxel induces apoptosis, cell death, inhibition of migration capacity and antitumoral activity in ovarian cancer

The main goal of this study is to evaluate the efficacy of the paclitaxel (PTX) drug formulated with a liposomal nanosystem (L-PTX) in a peritoneal carcinomatosis derived from ovarian cancer. In vitro cell viability studies with the human ovarian cancer line A2780 showed a 50% decrease in the inhibitory concentration for L-PTX compared to free PTX. A2780 cells treated with the L-PTX formulation demonstrated a reduced capacity to form colonies in comparison to those treated with PTX. Cell death following L-PTX administration hinted at apoptosis, with most cells undergoing initial apoptosis. A2780 cells exhibited an inhibitory migration profile when analyzed by Wound Healing and real-time cell analysis (xCELLigence) methods after L-PTX administration. This inhibition was related to decreased expression of the zinc finger E-box-binding homeobox 1 (ZEB1) and transforming growth factor 2 (TGF-β2) genes. In vivoL-PTX administration strongly inhibited tumor cell proliferation in ovarian peritoneal carcinomatosis derived from ovarian cancer, indicating higher antitumor activity than PTX. L-PTX formulation did not show toxicity in the mice model. This study demonstrated that liposomal paclitaxel formulations are less toxic to normal tissues than free paclitaxel and are more effective in inhibiting tumor cell proliferation/migration and inducing ZEB1/TGF-β2 gene expression.

Antiparasitic mebendazole (MBZ) effectively overcomes cisplatin resistance in human ovarian cancer cells by inhibiting multiple cancer-associated signaling pathways.

Ovarian cancer is the third most common cancer and the second most common cause of gynecologic cancer death in women. Its routine clinical management includes surgical resection and systemic therapy with chemotherapeutics. While the first-line systemic therapy requires the combined use of platinum-based agents and paclitaxel, many ovarian cancer patients have recurrence and eventually succumb to chemoresistance. Thus, it is imperative to develop new strategies to overcome recurrence and chemoresistance of ovarian cancer. Repurposing previously-approved drugs is a cost-effective strategy for cancer drug discovery. The antiparasitic drug mebendazole (MBZ) is one of the most promising drugs with repurposing potential. Here, we investigate whether MBZ can overcome cisplatin resistance and sensitize chemoresistant ovarian cancer cells to cisplatin. We first established and characterized two stable and robust cisplatin-resistant (CR) human ovarian cancer lines and demonstrated that MBZ markedly inhibited cell proliferation, suppressed cell wounding healing/migration, and induced apoptosis in both parental and CR cells at low micromole range. Mechanistically, MBZ was revealed to inhibit multiple cancer-related signal pathways including ELK/SRF, NFKB, MYC/MAX, and E2F/DP1 in cisplatin-resistant ovarian cancer cells. We further showed that MBZ synergized with cisplatin to suppress cell proliferation, induce cell apoptosis, and blunt tumor growth in xenograft tumor model of human cisplatin-resistant ovarian cancer cells. Collectively, our findings suggest that MBZ may be repurposed as a synergistic sensitizer of cisplatin in treating chemoresistant human ovarian cancer, which warrants further clinical studies.

IPSC-Derived NK Cells Synergize with T Cells and Anti-PD-1 Antibody to Mediate Durable Anti-Tumor Responses In Vivo

iPSC-Derived NK Cells Synergize with T Cells and Anti-PD-1 Antibody to Mediate Durable Anti-Tumor Responses In Vivo

Palladacyclopentadienyl complexes bearing purine‐based N‐heterocyclic carbenes: A new class of promising antiproliferative agents against human ovarian cancer

A complete protocol for the synthesis of new palladacyclopentadienyl complexes with purine‐based carbenes as supporting ligands is described. The new organometallic compounds were exhaustively characterized using NMR and infrared spectroscopies and elemental analysis. The single‐crystal X‐ray structure of complex 2b coordinating also a triphenylphosphine was resolved. Some of these complexes showed an antiproliferative activity comparable to or better than that of cisplatin on two human ovarian cancer lines: A2780 (cisplatin‐sensitive) and A2780cis (cisplatin‐resistant). Moreover, for complexes 2 and 3 (coordinating one purine‐based N‐heterocyclic carbene ligand and one phosphine) the cytotoxicity is associated with an evident induction of apoptosis. Finally, complexes 3, bearing one purine‐based N‐heterocyclic carbene ligand and one 1,3,5‐triaza‐7‐phosphaadamantane, proved practically inactive on non‐tumour fibroblast cells (MRC‐5).

Abstract B56: PTEN loss in the fallopian tube induces hyperplasia and ovarian tumor formation

Abstract Introduction: Many ovarian cancers likely originate in the fallopian tube and then colonize the ovary. An FTE origin for some HGSC is well established and there is increasing evidence that some endometrioid cancers may also originate in the FT. The loss of PTEN was found in 33% of STIC lesions. According to the TCGA, the PI3K/AKT pathway is altered in 45% of HGSC, and this could amplify downstream signaling similarly to the loss of PTEN, such as pAKT activation. PTEN mutations have also been found in about 20% of endometrioid ovarian carcinoma. Purpose of the Study: The signaling events involved in the onset of ovarian cancer from the fallopian tube epithelium (FTE) are crucial for early detection and treatment of the disease, but they remain poorly defined. Therefore, we aim to identify drivers of transformation and tumorigenesis from FTE. Experimental Procedures: We employed cells engineered with PTEN knocked down using shRNA and performed RNA-seq to identify the pathways involved in tumorigenesis. To uncover the mechanism underlying the invasion of cancerous fallopian tube cells to the ovary, PTEN-deficient cells were tagged with green fluorescent protein to study colonization of the ovary ex vivo. Finally, we generated a novel transgenic model of PAX8-driven loss of PTEN in the FTE to study early lesions in FT and ovarian cancer progression. Results: Conditional homozygous knockout of PTEN mediated by PAX8-cre recombinase was sufficient to drive endometrioid and serous borderline ovarian carcinoma, providing the first model of FTE-derived borderline tumors. In addition, heterozygous PTEN deletion in the FTE resulted in hyperplasia, providing a model to study early events of ovarian cancer pathogenesis. Loss of PTEN increased cell migration, invasion, and upregulated WNT4, a key regulator of Müllerian duct development during embryogenesis. An inhibitor of the WNT canonical and noncanonical pathways, LKG-974, as well as knockdown of WNT4 by siRNA, inhibited loss of PTEN-induced migration, whereas inhibitors of the canonical pathway alone (PRI724) did not. Further investigation revealed that WNT4 was required for increased migration and colonization of the ovary by PTEN-deficient oviductal cells in a β-catenin-independent manner. WNT4 was expressed in human ovarian cancer lines, especially in OVCAR8 and OVCAR4, and in several cancer histotypes, as revealed by the TMA analysis using IHC. LKG-974 also inhibited migration in the human ovarian cancer lines OVCAR4 and OVCAR8. Conclusions: Together, these findings provide a novel model to study the mechanism of fallopian tube tumor initiation and invasion to the ovary mediated by loss of PTEN, which may help to define early events of human ovarian carcinogenesis. Citation Format: Angela Russo, Austin Czarnecki, Matthew Dean, Wei Jian-Jun. PTEN loss in the fallopian tube induces hyperplasia and ovarian tumor formation. [abstract]. In: Proceedings of the AACR Conference: Addressing Critical Questions in Ovarian Cancer Research and Treatment; Oct 1-4, 2017; Pittsburgh, PA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(15_Suppl):Abstract nr B56.

Development and Characterization of Solid Lipid Nanoparticles Loaded with a Highly Active Doxorubicin Derivative.

Solid lipid nanoparticles (SLNs) comprise a versatile drug delivery system that has been developed for the treatment of a variety of diseases. The present study will investigate the feasibility of entrapping an active doxorubicin prodrug (a squalenoyl-derivative) in SLNs. The doxorubicin derivative-loaded SLNs are spherically shaped, have a mean diameter of 300–400 nm and show 85% w/w drug entrapment efficiency. The effects on cell growth of loaded SLNs, free doxorubicin and the prodrug have been examined using cytotoxicity and colony-forming assays in both human ovarian cancer line A2780 wild-type and doxorubicin-resistant cells. Further assessments as to the treatment’s ability to induce cell death by apoptosis have been carried out by analyzing annexin-V staining and the activation of caspase-3. The in vitro data demonstrate that the delivery of the squalenoyl-doxorubicin derivative by SLNs increases its cytotoxic activity, as well as its apoptosis effect. This effect was particularly evident in doxorubicin-resistant cells.

Strong antitumor synergy between DNA crosslinking and HSP90 inhibition causes massive premitotic DNA fragmentation in ovarian cancer cells.

All current regimens for treating ovarian cancer center around carboplatin as standard first line. The HSP90 inhibitor ganetespib is currently being assessed in advanced clinical oncology trials. Thus, we tested the combined effects of ganetespib and carboplatin on a panel of 15 human ovarian cancer lines. Strikingly, the two drugs strongly synergized in cytotoxicity in tumor cells lacking wild-type p53. Mechanistically, ganetespib and carboplatin in combination, but not individually, induced persistent DNA damage causing massive global chromosome fragmentation. Live-cell microscopy revealed chromosome fragmentation occurring to a dramatic degree when cells condensed their chromatin in preparation for mitosis, followed by cell death in mitosis or upon aberrant exit from mitosis. HSP90 inhibition caused the rapid decay of key components of the Fanconi anemia pathway required for repair of carboplatin-induced interstrand crosslinks (ICLs), as well as of cell cycle checkpoint mediators. Overexpressing FancA rescued the DNA damage induced by the drug combination, indicating that FancA is indeed a key client of Hsp90 that enables cancer cell survival in the presence of ICLs. Conversely, depletion of nuclease DNA2 prevented chromosomal fragmentation, pointing to an imbalance of defective repair in the face of uncontrolled nuclease activity as mechanistic basis for the observed premitotic DNA fragmentation. Importantly, the drug combination induced robust antitumor activity in xenograft models, again accompanied with depletion of FancA. In sum, our findings indicate that ganetespib strongly potentiates the antitumor efficacy of carboplatin by causing combined inhibition of DNA repair and cell cycle control mechanisms, thus triggering global chromosome disruption, aberrant mitosis and cell death.

Enhanced targeting of mitochondrial peroxide defense by the combined use of thiosemicarbazones and inhibitors of thioredoxin reductase

Peroxiredoxin-3 (Prx3) accounts for about 90% of mitochondrial peroxidase activity, and its marked upregulation in many cancers is important for cell survival. Prx3 oxidation can critically alter peroxide signaling and defense and can be a seminal event in promoting cell death. Here it is shown that this mechanism can be exploited pharmacologically by combinations of clinically available drugs that compromise Prx3 function in different ways. Clinically relevant levels of the thiosemicarbazone iron chelators triapine (Tp) and 2,2′-Dipyridyl-N,N-dimethylsemicarbazone (Dp44mT) promote selective oxidation of mitochondrial Prx3, but not cytosolic Prx1, in multiple human lung and ovarian cancer lines. Decreased cell survival closely correlates with Prx3 oxidation. However, Prx3 oxidation is not merely an indicator of cell death as cytotoxic concentrations of cisplatin do not cause Prx3 oxidation. The siRNA-mediated suppression of either Prx3 or thioredoxin-2, which supports Prx3, enhances Tp’s cytotoxicity. Tp-mediated Prx3 oxidation is driven by enhanced peroxide generation, but not by nitric oxide. Many tumors overexpress thioredoxin reductase (TrxR) which supports Prx activity. Direct inhibitors of TrxR (e.g. auranofin, cisplatin) markedly enhanced Tp’s cytotoxicity, and auranofin enhanced Prx3 oxidation by low dose Tp. Together, these results support an important role for Prx3 oxidation in the cytotoxicity of Tp, and demonstrate that TrxR inhibitors can significantly enhance Tp’s cytotoxicity. Thiosemicarbazone-based regimens could prove effective for targeting Prx3 in a variety of cancers.

OR36 Ligands distinct to ovarian cancer Hla-A*02:01

Aim T cells recognize cancer cells via HLA/peptide complexes, and class I HLA-restricted tumor infiltrating T cells correspond to favorable prognosis in ovarian cancer, the most deadly gynecologic disease. The identification of tumor-marking HLA/peptide complexes empowers bourgeoning cancer immunotherapy strategies such as therapeutic vaccination or transgenic T cell therapy. While predictive algorithms are highly useful, a large proteomic study can both inform predictive strategies and directly identify novel cancer-associated peptides. Here, ligand elution and peptide sequencing was used to characterize ligands defining the HLA-A*02:01 of cancerous ovarian cells. Methods A secreted form of HLA-A*02:01 was stably transfected into the human ovarian cancer lines SKOV-3, A2780, OV-90 as well as the immortalized normal ovarian line FHIOSE. HLA was harvested from bioreactors of each cell line and purified by affinity chromatography. Purified peptides were fractionated by two dimensions of high-pressure liquid chromatography followed by tandem mass spectrometry and PEAKS software for peptide sequencing. Ingenuity Pathway Analysis and DAVID Bioinformatics Databases were used for source protein analysis. Results Over 30,000 peptides on average were identified at a 1% false discovery rate for each line, with peptide lengths matching expected frequencies. A clear overarching HLA-A*02:01 motif was observed, and novel length-specific motif nuances were detectable. Source protein enrichments were identified, and multiple novel as well as previously predicted peptides were found from characterized ovarian cancer antigens such as p53, human epidermal growth factor receptor 2 (HER2), mesothelin, vascular endothelial growth factor alpha (VEGFA), and epidermal growth factor receptor (EGFR). Finally, a comparative analysis between cell lines revealed consistent and defining ligands among the ovarian cancer lines. Conclusions Class I HLA gathered from tumor cells in this large-scale study provided for the direct characterization of the cancerous ligandome with unprecedented breadth. This brought to light individual peptides consistent and distinct to the HLA-A*02:01 of cancerous cells, and these ligands represent candidate peptides for use in ovarian cancer immunotherapy.

Bioactive Oleanane Glycosides from Polyscias duplicata from the Madagascar Dry Forest [1

As part of the International Cooperative Biodiversity Group (ICBG) program, in a search for antiproliferative compounds, an ethanol extract of Polyscias duplicata was investigated due to its antiproliferative activity against the A2780 human ovarian cell cancer line (IC50 6 µg/mL). Seven known oleanane glycosides, 3β-[(α-L-arabinopyranosyl)oxy]-16α-hydroxyolean-12-en-28-oic acid (1, IC50 8 µM), 3β-[(α-L-arabinopyranosyl)oxy]-16α,23-dihydroxyolean-12-en-18-oic acid (2, IC50 13 µM), 3β-[(O-β-D-glucopyranosyl-(t-->3)-α-L-arabinopyranosyl)oxy]-16α-hydroxyolean-12-en-28-oic acid (3, IC50 7 µM), 3β-[(O-α-L-rhamnopyranosyl-(1-2)-α-L-arabinopyranosyl)oxy]-16α-hydroxyolean-12-en-28-oic acid (4, IC50 2.8 µM), 3β-[(O-β-D-glucopyranosyl-(l-->3)-α-L- arabinopyranosyl)oxy]-23-hydroxyolean-12-en-28-oic acid (5, IC50 10 µM), β-[(O-α-L-rhamnopyranosyl-(-1.2)-α-L-arabinopyranosyl)oxy]-23-hydroxyolean-12-en-28-oic acid (6, IC50 3.4 µM), and 3β-[(α-L-arabinopyranosyl)oxy]-23-hydroxyolean-12-en-28-oic acid (7, IC50 3.4 µM) were isolated, and their structures determined using spectroscopic methods.

Silymarin induces cell cycle arrest and apoptosis in ovarian cancer cells

The polyphenolic flavonoid silymarin that is the milk thistle extract has been found to possess an anti-cancer effect against various human epithelial cancers. In this study, to explore the regulative effect of silymarin on human ovarian cancer line A2780s and PA-1 cells, 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide assay and flow cytometry were respectively used to determine the inhibitory effect of silymarin on the both cell lines, and to measure their cell cycle progression. Apoptosis induction and mitochondrial membrane potential damage were separately detected by terminal deoxynucleotidyl transferase-mediated 2′-deoxyuridine 5′-triphosphate nick end labeling assay and 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazolylcarbocyanine iodide staining. Additionally, western blotting was applied to determine cytochrome C release and expression levels of p53, p21, p27, p16, CDK2, Bax, Bcl-2, procaspase-9, procaspase-3, cleaved caspase-9 and caspase-3 proteins. The activity of caspase-9 and caspase-3 was measured using Caspase-Glo-9 and Caspase-Glo-3 assay. The results indicated that silymarin effectively suppressed cell growth in a dose- and time-dependent manner, and arrested cell cycle progression at G1/S phase in A2780s and PA-1 cells via up-regulation of p53, p21, and p27 protein expression, and down-regulation of CDK2 protein expression. Additionally, silymarin treatment for 24h at 50 and 100µg/ml resulted in a reduction of mitochondrial membrane potential and cytochrome C release, and significantly induced apoptosis in A2780s and PA-1 cells by increasing Bax and decreasing Bcl-2 protein expression, and activation of caspase-9 and caspase-3. Therefore, silymarin is a possible potential candidate for the prevention and treatment of ovarian cancer.

The effect of curcumol on protein expression of JAK2/STAT3 signaling pathway in human ovarian cancer line SKOV3

Objective: To study the effects of curcumol on the protein expression of JAK2 and STAT3 in SKOV3 and to investigate its treatment on molecular mechanism of ovarian cancer. Methods: Choose curcumol of different concentrations to act on human ovarian cancer cell line SKOV3, and extract the corresponding cell protein, and detect the protein expression of JAK2 and STAT3 by western blotting. Results: The protein expression of JAK2 and STAT3 in SKOV3 are significantly inhabited by curcumol, and its strength will enhance with the increase in drug concentration, and it shows in a dose-dependent manner. Conclusion: Curcumol can significantly inhabit the proliferation of SKOV3 cells, and induce apoptosis, and achieve its mechanism by regulating the protein expression of JAK2 and STAT3.

Abstract 3620: A multiplex quantitative immunofluorescence assay for DNA damage repair in response to cytotoxic treatment

Abstract The use of DNA damaging agents remains one of the most common approaches to cancer therapy. The type of damage caused by the agent will determine which of the five main DNA damage repair (DDR) processes will respond. Initially we are developing multiplex quantitative immunoflourescence assays (qIFAs) for biomarkers of active Homologous Recombination (HR) and Nucleotide Excision Repair (NER) pathway and plan to develop mutiplex assays for the other repair pathways in the near future. The first multiplex panel to quantify DDR response has been developed to include Nbs1 phosphorylated at Serine343 and γH2AX, both early markers of DNA double strand breaks as well as ERCC1, which plays a key role in NER. Quantitation of repair pathway activation after drug treatment builds upon our validated qIFA for γH2AX, which scores individual nuclei as positive or negative for the biomarker and quantifies the γH2AX response as the fraction of nuclear area that is occupied by γH2AX foci. In contrast, the multiplex qIFA was designed to quantify the global DNA repair protein response to a variety of cytotoxic, genotoxic and targeted agents administered singly and in combination. Initial evaluations were performed in vitro on a panel of cell lines, characterized for repair pathway defects: the BRCA1-null human ovarian cancer line UWB1.289 (ATCC# CRL-2945), its isogenic line with restored BRCA1 named UWB1.289+BRCA1 (ATCC# CRL-2946), and SKOV3 (BRCA1 WT). Both gemcitabine and cisplatin increased pNbs1 both in UWB1.289 and UWB1.289+BRCA1, but not in SKOV3. Co-expression of pNbs1 with γH2AX was observed but was not 100% concordant. Combination treatment with a topoisomerase 1 inhibitor and PARP1/2 inhibitor (the indenoisoquinoline NSC 724998 with ABT-888) increased nuclear ERCC1 only in UWB1.289, but not in the isogenic UWB1.289+BRCA1 line, or in the ovarian lines SKOV3 and IGROV1, or the human fibroblast line GM00637. Assessment of drug-induced DDR response in a panel of cell lines with known repair defects is planned with the expectation that this will provide a scientific foundation for interpreting qIFA results when the assay is used to analyze clinical biopsies, and will assist in the evaluation of combination therapy strategies, such as synthetic lethality, designed to take advantage of DNA repair defects. Funded by NCI Contract No. HHSN261200800001E. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3620. doi:1538-7445.AM2012-3620

Protein profiles in human ovarian cancer cell lines correspond to their metabolic activity and to metabolic profiles of respective tumor xenografts

Many solid tumors show a large variability in glycolytic activity and lactate accumulation, which has been correlated with different metastatic spread, radioresistance and patient survival. To investigate potential differences in protein profiles underlying these metabolic variances, the highly glycolytic human ovarian cancer cell line OC316 was investigated and compared with the less glycolytic line IGROV-1. Extracellular acidification and oxygen consumption were analyzed with an extracellular flux analyzer. Glycolysis-associated proteins, including specific membrane transporters, were quantified through in-cell western analyses. Metabolic properties of corresponding tumor xenografts were assessed via induced metabolic bioluminescence imaging. Extracellular flux analyses revealed elevated bioenergetics of OC316 cells. Hexokinase II, pyruvate kinase, pyruvate dehydrogenase E1 beta subunit and pyruvate dehydrogenase kinase 1, as well as the glucose transporter 1 and the monocarboxylate transporter 4, were overexpressed in these cells compared with IGROV-1. When generating tumor xenografts in SCID mice, cells maintained their glycolytic behavior, i.e. OC316 showed higher lactate concentrations than IGROV-1 tumors. In summary, a congruency between protein profiles and metabolic properties has been demonstrated in the human ovarian cancer lines investigated. Also, a perpetuation of glycolytic characteristics during the transition from in vitro to the in vivo situation has been documented. This model system could be useful for systematic studies on therapeutic intervention by manipulation of tumor glycolysis and associated pathways.

Hyperactive EGF receptor, Jaks and Stat3 signaling promote enhanced colony-forming ability, motility and migration of cisplatin-resistant ovarian cancer cells

We present evidence that the cisplatin-resistant human ovarian cancer lines, A2780S/CP1 (S/CP1), A2780S/CP3 (S/CP3), and A2780S/CP5 (S/CP5), derived by subjecting the sensitive A2780S ovarian cancer line to multiple rounds of cisplatin treatments followed by recovery and are resistant to 1, 3, and 5 μM cisplatin, respectively, have increased colony-forming ability and altered morphology that is consistent with enhanced motility, migration, and invasiveness in vitro. The malignant phenotype progresses with increasing resistance and is associated with hyperactive epidermal growth factor receptor (EGFR)/extracellular signal-regulated kinase (Erk)1/2 and Janus kinases (Jaks), aberrant Signal Transducer and Activator of Transcription (Stat) 3 activation promoted by EGFR and Jaks, and epithelial-mesenchymal transition (EMT) in vitro. Survivin and FLIP anti-apoptotic factors, vascular endothelial growth factor (VEGF), and matrix metalloproteinase activities are also elevated in the resistant cells. Accordingly, the ectopic expression of constitutively-active Stat3C in the sensitive A2780S cells diminished cisplatin sensitivity. The inhibition of EGFR or Stat3 activity repressed Survivin, VEGF and Vimentin expression and the colony-forming potential, viability, motility, and migration of the resistant cells, and sensitized them to cisplatin. Analysis of human ovarian cancer patients’ tumor tissues shows aberrantly-active EGFR and Stat3 that in certain cases correlate with Vimentin over-expression. Intra-peritoneal mouse xenograft studies revealed, compared to the sensitive A2780S line that had low tumor incidence restricted to the ovary, a high tumor incidence for the resistant S/CP3 and S/CP5 lines that formed tumor nodules at several locations on the small-intestine and colon, and which responded poorly to cisplatin, but were sensitive to concurrent treatment with cisplatin and EGFR or Stat3 inhibitor. Hyperactive EGFR signaling through Stat3 and the Jak-Stat3 activity together promote ovarian cancer progression to cisplatin resistance and therefore represent targets for preventing the development of cisplatin resistance and the recurrent disease during cisplatin therapy in ovarian cancer.

A randomized phase II study of paclitaxel/carboplatin with or without sorafenib in the first-line treatment of patients with stage III/IV epithelial ovarian cancer.

TPS257 Background: The majority of patients diagnosed with ovarian cancer have advanced disease at the time of diagnosis, and are incurable with surgical therapy. Systemic chemotherapy, following initial cytoreductive surgery, has markedly improved the treatment of patients with advanced ovarian cancer. Standard first-line combination chemotherapy includes a taxane and a platinum agent. Further improvement in the first-line treatment of ovarian cancer will likely depend on the successful incorporation of novel, targeted agents into therapy. Several angiogenesis inhibitors have shown activity against refractory ovarian cancer. Sorafenib efficiently blocks the VEGF receptor and demonstrates antitumor activity in murine xenografts using a human ovarian cancer line (SKOV3). This randomized phase II study compares the efficacy and toxicity of a standard paclitaxel/carboplatin regimen, either alone or with the addition of sorafenib, in the first-line treatment of patients with stage III/IV ovarian cancer following cytoreductive surgery. Randomization of 120 patients will allow detection of an improvement in 2 yr PFS from 45% to 67% (α=.10). Methods: Eligibility: All patients must have previously untreated, histologically confirmed, stage III or IV epithelial ovarian carcinoma, with no residual tumor nodule > 3 cm, no involvement of the bowel (i.e. invasion into bowel wall) and no intestinal obstruction after surgical cytoreduction. Trial Design: Following recovery from cytoreductive surgery patients are randomized (1:1) to receive treatment with paclitaxel/carboplatin ± sorafenib. Regimen A: paclitaxel 175mg/m2, 1-3 hr infusion, day 1; carboplatin AUC=6.0 IV day 1; sorafenib 400 mg bid orally, days 1-21. Regimen B: paclitaxel/carboplatin (same doses as Regimen A). Cycles are repeated every 21 days. Patients are reevaluated every 6 weeks; responding/stable patients continue treatment for 6 cycles. After 6 cycles, responding patients on Regimen B continue single agent sorafenib to complete a 12 month course. Current Enrollment: 55 of 120 patients are currently enrolled. No significant financial relationships to disclose.

Enhancement of cisplatin sensitivity by NSC109268 in budding yeast and human cancer cells is associated with inhibition of S-phase progression

NSC109268 has been described previously as inhibitor of proteasomal degradation and of phosphatase 2Calpha. In a yeast screen, we isolated NSC109268 as an agent altering sensitivity to DNA-damaging agents. We found that NSC109268 and the related compound NSC109272 enhance cellular sensitivity to cis- and transplatin but reduce sensitivity to nitrogen mustard. We explored if similar effects could be found in human cancer cells and if cell cycle analysis could hint at the underlying molecular mechanism. Haploid yeast cells were treated in suspension with platinum agents and nitrogen mustard alone or in combination with NSC compounds, and survival was measured by colony-formation assays. Sensitivity of ovarian and prostate cancer cells toward these treatments was evaluated using the MTS assay. Cell cycle progression was determined by flow cytometry. The enhancement of cisplatin sensitivity by NSC109268 found in yeast was confirmed in cisplatin-sensitive and cisplatin-resistant human ovarian cancer lines and in prostate cancer cells. In yeast and in human carcinoma cells, a correlation of enhanced sensitivity with delaying S-phase progression was revealed. The known activities of NSC109268 as proteasome or phosphatase inhibitor could explain the phenotype of S-phase delay by assuming a higher initial DNA damage load, inhibition of DNA translesion synthesis or extended checkpoint arrest.

Inhibitory effect of antisense oligonucleotide of integrin-linked kinase on cell proliferation of human epithelial ovarian cancer

To explore the inhibitory effect of integrin-linked kinase antisense oligonucleotide (ILK-ASODN) on cell proliferation in human ovarian cancer cell line (HO8910). We transfected ILK-ASODN into HO8910 to block ILK gene expression, measured the expression levels of integrin-linked kinase (ILK) mRNA by RT-PCR and ILK protein by western-blotting; the inhibiting effects of the transfection on HO8910 proliferation, the cell cycles, and cell apoptosis were assessed by water soluble tetrazolium-1 (WST-1) and flow cytometry (FCM). After transfection of ILK-ASODN, the expression levels of ILK mRNA decreased significantly in groups D, E, F being 0.307 +/- 0.011, 0.198 +/- 0.008, 0, respectively, when compared with those of the two control groups of A and B (P < 0.05). The expression levels of ILK protein of the groups D, E and F decreased significantly also, being 26.3 +/- 0.8, 20.6 +/- 0.4 and 0, respectively. HO8910 cell proliferation was inhibited significantly, and the rates of apoptosis of the groups D, E and F increased significantly, being 7.31%, 8.84% and 11.27% respectively. The cell population increased in G0/G1 phase of the groups D, E and F, being 49.25%, 56.28% and 67.61% respectively, significantly different in comparison with those of groups of A and B (P < 0.01). Transfection of ILK-ASODN into human ovarian cancer line inhibited cancer cell proliferation significantly.