Sphingosine kinase 1 as a potential therapeutic target in epithelial ovarian cancer

Sphingosine kinase 1 (SK1) is over‐expressed in multiple types of human cancer. SK1 has growth‐promoting effects and has been proposed as a potential therapeutic target. We investigated the therapeutic effects of SK1 inhibition in epithelial ovarian carcinoma (EOC). SK1 siRNA or inhibitors were tested in EOC cell lines, including A2780, SKOV3ip1, A2780‐CP20, SKOV3‐TR, ES2 and RMG2. Cells were treated with SK inhibitor or FTY720, and cell proliferation, apoptosis, angiogenesis and invasion were examined by MTT, FACS, ELISA and wound‐healing assays, respectively. In vivo experiments were performed to test the effects of FTY720 on tumor growth in orthotopic mouse xenografts of EOC cell lines A2780 or SKOV3ip1 and a patient‐derived xenograft (PDX) model of clear cell ovarian carcinoma (CCC). Blocking SK1 with siRNA or inhibitors significantly reduced proliferation, angiogenesis and invasion, and increased apoptosis in chemosensitive (A2780 and SKOV3ip1) and chemoresistant (A2780‐CP20, SKOV3‐TR, ES2 and RMG2) EOC cells. SK1 inhibitors also decreased the intracellular enzymatic activity of SK1. Furthermore, FTY720 treatment significantly decreased the in vivo tumor weight in xenograft models of established cell lines (A2780 and SKOV3ip1) and a PDX model for CCC compared to control (p < 0.05). These results support therapeutic targeting of SK1 as a potential new strategy for EOC.     

Proton pump inhibitors enhance the effects of cytotoxic agents in chemoresistant epithelial ovarian carcinoma

This study was designed to investigate whether proton pump inhibitors (PPI, V-ATPase blocker) could increase the effect of cytotoxic agents in chemoresistant epithelial ovarian cancer (EOC). Expression of V-ATPase protein was evaluated in patients with EOC using immunohistochemistry, and patient survival was compared based on expression of V-ATPase mRNA from a TCGA data set. In vitro, EOC cell lines were treated with chemotherapeutic agents with or without V-ATPase siRNA or PPI (omeprazole) pretreatment. Cell survival and apoptosis was assessed using MTT assay and ELISA, respectively. In vivo experiments were performed to confirm the synergistic effect with omeprazole and paclitaxel on tumor growth in orthotopic and patient-derived xenograft (PDX) mouse models. Expression of V-ATPase protein in ovarian cancer tissues was observed in 44 patients (44/59, 74.6%). Higher expression of V-ATPase mRNA was associated with poorer overall survival in TCGA data. Inhibition of V-ATPase by siRNA or omeprazole significantly increased cytotoxicity or apoptosis to paclitaxel in chemoresistant (HeyA8-MDR, SKOV3-TR) and clear cell carcinoma cells (ES-2, RMG-1), but not in chemosensitive cells (HeyA8, SKOV3ip1). Moreover, the combination of omeprazole and paclitaxel significantly decreased the total tumor weight compared with paclitaxel alone in a chemoresistant EOC animal model and a PDX model of clear cell carcinoma. However, this finding was not observed in chemosensitive EOC animal models. These results show that omeprazole pretreatment can increase the effect of chemotherapeutic agents in chemoresistant EOC and clear cell carcinoma via reduction of the acidic tumor microenvironment.     

Hospicells (ascites‐derived stromal cells) promote tumorigenicity and angiogenesis

The microenvironment is known to play a dominant role in cancer progression. Cells closely associated with tumoral cells, named hospicells, have been recently isolated from the ascites of ovarian cancer patients. Whilst these cells present no specific markers from known cell lineages, they do share some homology with bone marrow‐derived or adipose tissue‐derived human mesenchymal stem cells (CD9, CD10, CD29, CD146, CD166, HLA‐1). We studied the role of hospicells in ovarian carcinoma progression. In vitro, these cells had no effect on the growth of human ovarian carcinoma cell lines OVCAR‐3, SKOV‐1 and IGROV‐1. In vivo, their co‐injection with adenocarcinoma cells enhanced tumor growth whatever the tumor model used (subcutaneous and intraperitoneally established xenografts in athymic mice). In addition, their injection increased the development of ascites in tumor‐bearing mice. Fluorescent macroscopy revealed an association between hospicells and ovarian adenocarcinoma cells within the tumor mass. Tumors obtained by coinjection of hospicells and human ovarian adenocarcinoma cells presented an increased microvascularization indicating that the hospicells could promote tumorigenicity of ovarian tumor cells in vivovia their action on angiogenesis. This effect on angiogenesis could be attributed to the increased HIF1α and VEGF expression associated with the presence of the hospicells. Collectively, these data indicate a role for these ascite‐derived stromal cells in promoting tumor growth by increasing angiogenesis.     

Collagen type VI regulates the CDK4/6-p-Rb signaling pathway and promotes ovarian cancer invasiveness,stemness, and metastasis

The expression of collagen VI in primary ovarian tumors may correlate with tumor grade and response to chemotherapy. We have sought to elucidate the role of collagen VI in promoting ovarian cancer tumor growth and metastasis. Here we examined the effects of collagen VI on ovarian carcinoma stromal progenitor cells (OCSPCs). Epithelial-like OCSPCs (epi-OCSPCs) and mesenchymal-like OCSPCs (msc-OCSPCs) were analyzed by liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS). Differentially expressed genes were integrated with survival-related genes using The Cancer Genome Atlas (TCGA) data and confirmed in our samples. The roles of candidate genes and signaling pathways were further explored. We found that SKOV3/msc-OCSPCs possessed greater migration, invasion, and spheroid formation than SKOV3/epi-OCSPCs (P < 0.001). Expression of collagen alpha-3 (VI; COL6A3), which encodes collagen VI, was 90-fold higher in msc-OCSPCs than in epi-OCSPCs. Analysis of TCGA data and our samples indicated that high expression of COL6A3 was correlated with advanced-stage carcinoma (P < 0.01) and shorter overall survival (P < 0.01). In vitro, adding collagen VI, msc-OCSPCs, or knockdown collagen VI in msc-OCSPCs to epithelial ovarian carcinoma (EOC) cells augmented or decreased invasion and spheroid formation. Tumor dissemination to the peritoneal cavity and lung in mice following intraperitoneal coinjection with msc-OCSPCs and SKOV3-Luc cells and intravenous injection with COL6A3 and ES2 cells derived spheroids was significantly greater compare to coinjection with SKOV3-Luc cells alone or in combination with msc-OCSPCs/shCOL6A3 cells and msc-OCSPCs and ES2 derived spheroids. Knockdown of COL6A3 abolished the expression of DNMT1, CDK4, CDK6, and p-Rb in msc-OCSPCs and EOC spheroids. In contrast, overexpression of COL6A3 enhanced the expression of CDK4, CDK6, and p-Rb in SKOV3 cells. EOC spheroid formation, invasion, tumor growth, and metastasis were inhibited when COL6A3 downstream signaling pathway was blocked using CDK4/6 inhibitor LEE011. Our results suggested that collagen VI regulates the CDK4/6-p-Rb signaling pathway and promotes EOC invasiveness, stemness, and metastasis.     

The effects of selenium on tumor growth in epithelial ovarian carcinoma

Objective

Epidemiological studies suggest that selenium protects against the development of several cancers. Selenium (sodium selenite) has been reported to interfere with cell growth and proliferation, and to induce cell death. In this study, we tested whether selenium could have growth-inhibiting effect in ovarian cancer cells and an orthotopic animal model.

Methods

Cell growth in selenium-treated cells was determined in human ovarian cancer cells, A2780, HeyA8, and SKOV3ip1 using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide (MTT) assay. Animal experiment of selenium with paclitaxel was performed using SKOV3ip1 cells in nude mice to evaluate their inhibiting effect for tumor growth. In addition, another animal experiment of paclitaxel with or without selenium was performed to assess the effect of survival and food intake in mice.

Results

The in vitro growth of selenium-treated cells was significantly decreased dose-dependently in A2780, HeyA8, and SKOV3ip1 cells. Therapy experiment in mice was started 1 week after injection of the SKOV3ip1 cells. Treatment with selenium (1.5 mg/kg, 3 times/week) and paclitaxel injection showed no addictive effect of the inhibition of tumor growth. However, combination of selenium and paclitaxel showed the slightly increased food intake compared with paclitaxel alone.

Conclusion

Although selenium has growth-inhibiting effect in ovarian carcinoma cells in vitro, there is no additive effect on tumor growth in mice treated with combination of paclitaxel and selenium. However, food intake is slightly higher in selenium-treated mice during chemotherapy.     

Functional significance of VEGFR‐2 on ovarian cancer cells

Vascular endothelial growth factor receptor (VEGFR) has recently been discovered on ovarian cancer cells, but its functional significance is unknown and is the focus of this study. By protein analysis, A2780‐par and HeyA8 ovarian cancer cell lines expressed VEGFR‐1 and HeyA8 A2774, and SKOV3ip1 expressed VEGFR‐2. By in situ hybridization (ISH), 85% of human ovarian cancer specimens showed moderate to high VEGFR‐2 expression, whereas only 15% showed moderate to high VEGFR‐1 expression. By immunofluorescence, little or no VEGFR‐2 was detected in normal ovarian surface epithelial cells, whereas expression was detected in 75% of invasive ovarian cancer specimens. To differentiate between the effects of tumor versus host expression of VEGFR, nude mice were injected with SKOV3ip1 cells and treated with either human VEGFR‐2 specific antibody (1121B), murine VEGFR‐2 specific antibody (DC101) or the combination. Treatment with 1121B reduced SKOV3ip1 cell migration by 68% (p < 0.01) and invasion by 72% (p < 0.01), but exposure to VEGFR‐1 antibody had no effect. Treatment with 1121B effectively blocked VEGF‐induced phosphorylation of p130Cas. In vivo treatment with either DC101 or 1121B significantly reduced tumor growth alone and in combination in the SKOV3ip1 and A2774 models. Decreased tumor burden after treatment with DC101 or 1121B correlated with increased tumor cell apoptosis, decreased proliferative index, and decreased microvessel density. These effects were significantly greater in the combination group (p < 0.001). We show functionally active VEGFR‐2 is present on most ovarian cancer cells. The observed anti‐tumor activity of VEGF‐targeted therapies may be mediated by both anti‐angiogenic and direct anti‐tumor effects. © 2008 Wiley‐Liss, Inc.     

Phenethyl isothiocyanate suppresses the metastasis of ovarian cancer associated with the inhibition of CRM1-mediated nuclear export and mTOR-STAT3 pathway

Epithelial ovarian cancer is prone to metastasizing at an early stage, but their mechanisms remain unclear. CRM1 is an important nuclear exportin and inhibitors targeting CRM1 has been explored as an anti-cancer strategy. In previous study, we observed that PEITC could combine with the hydrophobic pocket of CRM1. In this study, we focused on the effects of PEITC on EOC and its mechanisms. Results showed that IC₅₀ values of PEITC on SKOV3 and HO8910 cell line were 42.14 μM and 37.29 μM, respectively. PEITC inhibits the migration and invasion of SKOV3 and HO8910 cells in vitro. Oral administration of 10 μmol PEITC suppressed the metastasis of EOC in a xenograft mouse model in vivo. PEITC treatment decreased the expressions of CRM1 and mTOR (cargo protein of CRM1) in EOC cell lines and in xenograft mouse tissues. Moreover, CRM1-mediated nuclear export was attenuated by PEITC, mTOR accumulated in nucleus, expressions of mTOR^S2448 and downstream effectors STAT3^S727, MMP2 and MMP9 were decreased in a dose- and time-dependent manner. Furthermore, immunohistochemical analysis showed that CRM1 and mTOR were increased in EOC tissues compared with benign ovarian tumors, and related with advanced stage, type II EOC, positive peritoneal cytology and decreased overall survival. In addition, CRM1 was positively correlated with mTOR levels. In conclusion, our data demonstrated that PEITC suppresses the metastasis of EOC through inhibiting CRM1-mediated nuclear export, subsequently suppressing the mTOR-STAT3 pathway. Both CRM1 and mTOR were increased in EOC patients, providing a rationale for further clinical investigation of PEITC in EOC treatment.     

The growth and metastasis of human, HER-2/neu-overexpressing tumor cell lines in male SCID mice

HER-2/neu is overexpressed on a variety of human adenocarcinomas and overexpression has been associated with a poor prognosis. For this reason, HER-2 has become an attractive target for immunotherapy. To facilitate testing of anti-HER-2-monoclonal antibodies (MAbs) and immunotoxins (ITs), we have evaluated the in vivo growth and metastatic spread of three HER-2-overexpressing human breast cancer cell lines (BT474, MDA-MB-453 and HCC1954) and one ovarian cancer cell line (SKOV3.ip1) in pre-irradiated male SCID mice using subcutaneous (s.c.), intravenous (i.v.) and intraperitoneal (i.p.) routes of injection. All the cell lines tested grew as s.c. tumors and the growth of BT474 and MDA-MB-453 cells after s.c. injection was improved by co-inoculation with Matrigel. Metastases to the lungs were detectable by PCR or histopathology after s.c. injection of BT474 and to a much lesser extent after s.c. injection of HCC1954, MD-MB-453 and SKOV3.ip1cells. I.P. injection of HCC1954 and SKOV3.ip1 cells produced fatal ascites while i.v. injection of SKOV3.ip1, but not BT474 or MDA-MB-453 cells, resulted in infiltration of lungs and death within 9–11 weeks.     

Tumor-Selective Response to Antibody-Mediated Targeting of αvβ3 Integrin in Ovarian Cancer

The α_vβ₃ integrin is expressed on proliferating endothelial cells and some cancer cells, but its expression on ovarian cancer cells and its potential as a therapeutic target are unknown. In this study, expression of the α_vβ₃ integrin on ovarian cancer cell lines and murine endothelial cells was tested, and the effect of a fully humanized monoclonal antibody against α_vβ₃, Abegrin (etaracizumab), on cell invasion, viability, tumor growth, and the Akt pathway were examined in vitro and in vivo. We found that etaracizumab recognizes α_vβ₃ on the ovarian cancer cell lines SKOV3ip1, HeyA8, and A2780ip2 (at low levels) but not on murine endothelial cells. Etaracizumab treatment decreased ovarian cancer proliferation and invasion. In vivo, tumor-bearing mice treated with etaracizumab alone gave variable results. There was no effect on A2780ip2 growth, but a 36% to 49% tumor weight reduction in the SKOV3ip1 and HeyA8 models was found (P < .05). However, combined etaracizumab and paclitaxel was superior to paclitaxel in the SKOV3ip1 and A2780ip2 models (by 51–73%, P < .001) but not in the HeyA8 model. Treatment with etaracizumab was then noted to decrease p-Akt and p-mTOR in SKOV3ip1, but not in HeyA8, which is Akt-independent. Tumors resected after therapy showed that etaracizumab treatment reduced the proliferating cell nuclear antigen index but not microvessel density. This study identifies tumor cell α_vβ₃ integrin as an attractive target and defines the Akt pathway as a predictor of response to function-blocking antibody.     

Long noncoding RNA TC0101441 induces epithelial–mesenchymal transition in epithelial ovarian cancer metastasis by downregulating KiSS1

Peritoneal metastasis is a critical feature and clinical challenge in epithelial ovarian cancer (EOC). We previously identified a novel long noncoding RNA (lncRNA, TC0101441) in epithelial ovarian cancer (EOC) using microarrays. However, the impact of TC0101441 on EOC metastasis and prognosis remains unclear. TC0101441 expression in EOC tissues and its correlation with clinicopathological factors and prognosis were examined. A series of in vitro and in vivo assays were performed to elucidate the roles and mechanism of TC0101441 in EOC metastasis. We found that TC0101441 levels were elevated in EOC tissues compared with those in normal controls and significantly correlated with an advanced clinical stage and lymph node metastasis. TC0101441 was determined to be an independent prognostic predictor of overall survival (OS) and disease-free survival (DFS). Furthermore, loss-of-function assays showed that TC0101441 promoted the invasive and metastatic capacities of EOC cells both in vitro and in vivo. Mechanistically, the prometastatic effects of TC0101441 were linked to the induction of epithelial–mesenchymal transition (EMT). Importantly, KiSS1 was identified as a downstream target gene of TC0101441 and was downregulated by TC0101441 in EOC cells. After TC0101441 was silenced, the corresponding phenotypes of EOC cell invasion and EMT were reversed by the overexpression of KiSS1. Taken together, our data suggest that TC0101441 functions as a potential promigratory/invasive oncogene by promoting EMT and metastasis in EOC through downregulation of KiSS1, which may represent a novel prognostic marker and therapeutic target in EOC.     

Monitoring response to cytostatic cisplatin in a HER2(+) ovary cancer model by MRI and in vitro and in vivo MR spectroscopy

Background:

Limited knowledge is available on alterations induced by cytostatic drugs on magnetic resonance spectroscopy (MRS) and imaging (MRI) parameters of human cancers, in absence of apoptosis or cytotoxicity. We here investigated the effects of a cytostatic cisplatin (CDDP) treatment on ¹H MRS and MRI of HER2-overexpressing epithelial ovarian cancer (EOC) cells and in vivo xenografts.

Methods:

High-resolution MRS analyses were performed on in vivo passaged SKOV3.ip cells and cell/tissue extracts (16.4 or 9.4 T). In vivo MRI/MRS quantitative analyses (4.7 T) were conducted on xenografts obtained by subcutaneous implantation of SKOV3.ip cells in SCID mice. The apparent diffusion coefficient (ADC) and metabolite levels were measured.

Results:

CDDP-induced cytostatic effects were associated with a metabolic shift of cancer cells towards accumulation of MRS-detected neutral lipids, whereas the total choline profile failed to be perturbed in both cultured cells and xenografts. In vivo MRI examinations showed delayed tumour growth in the CDDP-treated group, associated with early reduction of the ADC mean value.

Conclusion:

This study provides an integrated set of information on cancer metabolism and physiology for monitoring the response of an EOC model to a cytostatic chemotherapy, as a basis for improving the interpretation of non-invasive MR examinations of EOC patients.     

Chemotherapy sensitivity testing on ovarian cancer cells isolated from malignant ascites

Background: In epithelial ovarian cancer (EOC), 15–20% of the tumors do not respond to first-line chemotherapy (paclitaxel with platinum-based therapy), and in recurrences this number increases. Our aim is to determine the feasibility of cell proliferation assays of tumor cells isolated from malignant ascites to predict in vitro chemotherapy sensitivity, and to correlate these results with clinical outcome.Materials and Methods: Ascites was collected from twenty women with advanced EOC. Cell samples were enriched for tumor cells and EOC origin was confirmed by intracellular staining of CK7, surface staining of CA125 and EpCAM, and HE4 gene expression. In vitro sensitivity to chemotherapy was determined in cell proliferation assays using intracellular ATP content as an indirect measure of cell number. In vitro drug response was quantified by calculation of the drug concentration at which cell growth was inhibited with 50%. Clinical outcome was determined using post-treatment CA125 level.Results: Cell samples of twenty patients were collected, of which three samples that failed to proliferate were excluded in the analysis (15%). Three other samples were excluded, because clinical outcome could not be determined correctly. In twelve of the fourteen remaining cases (86%) in vitro drug sensitivity and clinical outcome corresponded, while in two samples (14%) there was no correspondence.Conclusions: Our study demonstrates the feasibility of drug sensitivity tests using tumor cells isolated from ascites of advanced EOC patients. Larger observational studies are required to confirm the correlation between the in vitro sensitivity and clinical outcome.     

An Orally Available Small-Molecule Inhibitor of c-Met,PF-2341066, Reduces Tumor Burden and Metastasis in a Preclinical Model of Ovarian Cancer Metastasis

Deregulated expression of the hepatocyte growth factor (HGF) receptor, c-Met, in cancer contributes to tumor progression and metastasis. The objective of this study was to determine whether blocking c-Met with an orally available c-Met inhibitor, PF-2341066, reduces tumor burden and increases survival in a xenograft model of ovarian cancer metastasis. Treatment of mice injected interperitoneally with SKOV3ip1 cells showed reduced overall tumor burden. Tumor weight and the number of metastases were reduced by 55% (P < .0005) and 62% (P < .0001), respectively. Treatment also increased median survival from 45 to 62 days (P = .0003). In vitro, PF-2341066 reduced HGF-stimulated phosphorylation of c-Met in the tyrosine kinase domain as well as phosphorylation of the downstream signaling effectors, Akt and Erk. It was apparent that inhibition of the pathways was functionally important because HGF-induced branching morphogenesis was also inhibited. In addition, proliferation and adhesion to various extracellular matrices were inhibited by treatment with PF-2341066, and the activity of matrix metalloproteinases was decreased in tumor tissue from treated mice compared with those receiving vehicle. Overall, these data indicate that PF-2341066 effectively reduces tumor burden in an in vivo model of ovarian cancer metastasis and may be a good therapeutic candidate in the treatment of patients with ovarian cancer.     

SMYD3 promotes implant metastasis of ovarian cancer via H3K4 trimethylation of integrin promoters

Detachment of cancer cells from the primary tumor and formation of spheroids in ascites is required for implantation metastasis in epithelial ovarian cancer (EOC), but the underlying mechanism of this process has not been thoroughly elucidated. To mimic this process, ovarian cancer cells were grown in 3D and 2D culture. Hey and OVCA433 spheroids exhibited decreased cell proliferation and enhanced adhesion and invasion. SMYD3 expression was elevated in ovarian carcinoma spheroids in association with increased H3K4 methylation. Depletion of SMYD3 by transient siRNA, stable shRNA knockdown and the SMYD3 inhibitor BCI-121 all decreased spheroid invasion and adhesion. Gene expression arrays revealed downregulation of integrin family members. Inhibition assays confirmed that invasion and adhesion of spheroids are mediated by ITGB6 and ITGAM. SMYD3-deficient cells regained the ability to invade and adhere after forced overexpression of SMYD3, ITGB6 and ITGAM. However, this biological ability was not restored by forced overexpression of SMYD3 in ITGB6- and/or ITGAM-deficient cancer cells. SMYD3 and H3K4me3 binding at the ITGB6 and ITGAM promoters was increased in spheroids compared to that in monolayer cells, and the binding was decreased when SMYD3 expression was inhibited, consistent with the expression changes in integrins. SMYD3 expression and integrin-mediated adhesion were also activated in an intraperitoneal xenograft model and in EOC patient spheroids. In vivo, SMYD3 knockdown inhibited tumor metastasis and reduced ascites volume in both the intraperitoneal xenograft model and a PDX model. Overall, our results suggest that the SMYD3-H3K4me3-integrin pathway plays a crucial role in ovarian cancer metastasis to the peritoneal surface.     

NPPB is a novel candidate biomarker expressed by cancer‐associated fibroblasts in epithelial ovarian cancer

Most solid tumors contain cancer‐associated fibroblasts (CAFs) that support tumorigenesis and malignant progression. However, the cellular origins of CAFs in epithelial ovarian cancers (EOCs) remain poorly understood, and their utility as a source of clinical biomarkers for cancer diagnosis has not been explored in great depth. Here, we report establishing in vitro and in vivo models of CAFs in ovarian cancer development. Normal ovarian fibroblasts and mesenchymal stem cells cultured in the presence of EOC cells acquired a CAF‐like phenotype, and promoted EOC cell migration in vitro. CAFs also promoted ovarian cancer growth in vivo in both subcutaneous and intraperitoneal murine xenograft assays. Molecular profiling of CAFs identified gene expression signatures that were highly enriched for extracellular and secreted proteins. We identified novel candidate CAF‐specific biomarkers for ovarian cancer including NPPB, which was expressed in the stroma of 60% primary ovarian cancer tissues (n = 145) but not in the stroma of normal ovaries (n = 4). NPPB is a secreted protein that was also elevated in the blood of 50% of women with ovarian cancer (n = 8). Taken together, these data suggest that the tumor stroma is a novel source of biomarkers, including NPPB, that may be of clinical utility for detection of EOC.     

两种荷人卵巢癌腹腔移植瘤的免疫重建SCID小鼠模型的建立及比较

背景与目的:卵巢癌的死亡率在妇科恶性肿瘤居首位。目前,常规的手术、化疗和放疗难以再提高患者的生存率,因此,生物治疗成为卵巢癌的第四大治疗模式,而生物治疗的评价需要合适的动物模型。本实验目的是建立两种荷人卵巢癌腹腔移植瘤的免疫重建的严重联合免疫缺陷(severecombinedimmunodeficiency,SCID)小鼠模型,通过比较选出较适宜的模型。方法:分别用人卵巢癌细胞株SKOV3细胞和SKOV3.ip1细胞腹腔注射6只和10只C.B17/SCID小鼠建立腹腔瘤模型,比较其生物学、组织学和免疫学特性;并将SKOV3.ip1组小鼠的腹水转种6只小鼠的腹腔,观察成瘤情况。以上22只SCID小鼠均用人外周血淋巴细胞腹腔注射进行人免疫功能重建。结果:用SKOV3细胞和SKOV3.ip1细胞分别建立的两种模型小鼠成瘤率均为100%;成瘤潜伏期分别为20～41天和22～30天(P>0.05);生存期分别为50～78天和32～43天(P<0.0001)。SKOV3.ip1组小鼠的腹水进行转种后有83.3%(5/6)能形成腹腔肿瘤和腹水。两种模型小鼠尸检发现肿瘤分布相似,均形成广泛的盆腔播散;但SKOV3.ip1组伴有0.35～5.60ml血性腹水,而SKOV3组仅有1只有0.20ml腹水。组织学显示两种模型都保持了人卵巢浆液性乳头状腺癌的特点,免疫组化结果表明两种模型都表达卵巢癌相关抗原OC166-9。72.7%(16/22)     

High galectin-1 expression correlates with poor prognosis and is involved in epithelial ovarian cancer proliferation and invasion

PurposeGalectin-1 (Gal-1) is a 14-kDa laminin-binding galectin involved in several biological events including regulation of tumour proliferation and metastasis. In this study, we investigated the clinical significance of Gal-1 expression and its functional role in cell proliferation and invasion in epithelial ovarian cancer (EOC).Experimental designWe evaluated the expression of Gal-1 in 52 serous, 11 endometrioid, and 3 mucinous type EOC tumour samples from 66 patients by immunohistochemistry. In vitro experiments were performed to determine the function of Gal-1 in cell survival, proliferation, and invasion in EOC cells using siRNA and anginex, a Gal-1 inhibitor, as well as recombinant Gal-1 protein.ResultsPatients with strong Gal-1 peritumoural staining had poorer progression-free survival (PFS) than patients with weak peritumoural staining (p = 0.03). Inhibition of Gal-1 by siRNA or anginex resulted in the inhibition of cell growth and proliferation of HeyA8 and SKOV3ip1 cells. Moreover, the ability of cells to migrate was significantly reduced by treatment of cells with Gal-1 siRNA but was increased by treatment of cells with recombinant Gal-1. When we evaluated the interaction between fibroblasts (T HESCs) and cancer cells (A2780-CP20), we found that MMP-2 expression in cancer cells was affected by Gal-1 secreted by fibroblast cells, which suggests that Gal-1 in human fibroblasts might affect the invasive abilities of tumour cells.ConclusionOur results suggest that Gal-1 expression is a potential prognostic factor for PFS and that Gal-1 could be a novel treatment target in EOC patients.     

Therapeutic role of EF24 targeting glucose transporter 1‐mediated metabolism and metastasis in ovarian cancer cells

Cancer cells require glucose to support their rapid growth through a process known as aerobic glycolysis, or the Warburg effect. As in ovarian cancer cells, increased metabolic activity and glucose concentration has been linked to aggressiveness of cancer. However, it is unclear as to whether targeting the glycolytic pathway may kill the malignant cells and likely have broad therapeutic implications against ovarian cancer metastasis. In the present research, we found that EF24, a HIF‐1α inhibitor, could significantly block glucose uptake, the rate of glycolysis, and lactate production compared with vehicle treatment in SKOV‐3, A2780 and OVCAR‐3 cells. These results might possibly contribute to the further observation that EF24 could inhibit ovarian cancer cell migration and invasion from wound healing and Transwell assays. Furthermore, as an important mediator of glucose metabolism, glucose transporter 1 (Glut1) was found to contribute to the function of EF24 in both energy metabolism and metastasis. To examine the effect of EF24 and the mediated role of Glut1 in vivo in a xenograph subcutaneous tumor model, intraperitoneal metastasis and lung metastasis model were introduced. Our results indicated that EF24 treatment could inhibit tumor growth, intraperitoneal metastasis and lung metastasis of SKOV‐3 cells, and Glut1 is a possible mediator for the role of EF24. In conclusion, our results highlight that an anti‐cancer reagent with an inhibiting effect on energy metabolism could inhibit metastasis, and EF24 is a possible candidate for anti‐metastasis therapeutic applications for ovarian cancer.     

Efficient inhibition of intra-peritoneal tumor growth and dissemination of human ovarian carcinoma cells in nude mice by anti-L1-cell adhesion molecule monoclonal antibody treatment

The L1 cell adhesion molecule is implicated in the control of proliferation, migration, and invasion of several tumor cell types in vitro. Recently, L1 overexpression was found to correlate with tumor progression of ovarian carcinoma, one of the most common causes of cancer-related deaths in gynecologic malignant diseases. To evaluate L1 as a potential target for ovarian cancer therapy, we investigated the effects of anti-L1 monoclonal antibodies (chCE7 and L1-11A) on proliferation and migration of L1-positive human SKOV3ip ovarian carcinoma cells in vitro and the therapeutic efficacy of L1-11A against i.p. SKOV3ip tumor growth in nude mice. In vitro, both anti-L1 antibodies efficiently inhibited the proliferation of SKOV3ip cells as well as other L1-expressing tumor cell lines (renal carcinoma, neuroblastoma, and colon carcinoma). On two cell lines, hyper-cross-linking of L1-11A with a secondary antibody was necessary for significant inhibition of proliferation, indicating that cross-linking of L1 is required for the antiproliferative effect. L1-negative prostate carcinoma cells were not influenced by antibody treatment. Biweekly treatment of ovarian carcinoma-bearing mice with L1-11A led to a dose-dependent and significant reduction of tumor burden (up to -63.5%) and ascites formation (up to -75%). This effect was associated with reduced proliferation within the tumors. L1-directed antibody-based inhibition of peritoneal growth and dissemination of human ovarian carcinoma cells represents important proof-of-principle for the development of a new therapy against one of the leading gynecologic malignant diseases.