Ascites Production Research Articles

Proteomic identification of fucosylated haptoglobin alpha isoforms in ascitic fluids and its localization in ovarian carcinoma tissues from Mexican patients

BackgroundOvarian cancer is the most lethal gynecologic disease due to delayed diagnosis, and ascites production is a characteristic of patients in advanced stages. The aim of this study was to perform the proteomic analysis of ascitic fluids of Mexican patients with ovarian carcinoma, in order to detect proteins with a differential expression pattern in the continuing search to identify biomarkers for this disease.MethodsSamples were collected from 50 patients from the Instituto Nacional de Cancerología of México under informed consent and with approval of the bioethics and scientific committees. After elimination of abundant proteins (Albumin/IgGs) samples were processed for 2D electrophoresis and further protein identification by Mass Spectrometry (MALDI-TOF). Molecules of interest were followed by western blot and lectin binding assays, and their tissue location by histo-immunofluorescence and confocal analysis.Results and discussionAn area with a differential expression pattern among samples was located in the 2D gels. Identified proteins were 6 alpha 1 isoforms and 1 alpha 2 isoform of Haptoglobin, and 2 isoforms of Transthyretin. While Transthyretin isoforms were constitutively expressed in all samples, clear differences in the expression pattern of Haptoglobin alpha isoforms were found. Moreover, increased levels of fucosylation of Haptoglobin alpha isoforms analyzed in 40 samples by Aleuria aurantia lectin binding by 1D overlay assay showed a positive correlation with advanced stages of the disease. Tissue detection of Haptoglobin and its fucosylated form, by histo-immunofluorescence in biopsies of ovarian cancer, also showed a correlation with ovarian cancer progression. Moreover, results show that fucosylated Haptoglobin is produced by tumor cells.ConclusionsIncreased numbers of highly fucosylated Haptoglobin alpha isoforms in ascitic fluids and the presence of fucosylated Haptoglobin in tumor tissues of ovarian cancer Mexican patients associated with advanced stages of the disease, reinforce the potential of fucosylated Haptoglobin alpha isoforms to be characterized as biomarkers for disease progression.

The efficacy and safety outcomes of bevacizumab and systemic therapy in metastatic colorectal cancer patients with peritoneal disease in the AGITG MAX clinical trial and in nontrial patients in two cancer centers.

595 Background: Metastatic colorectal cancer with peritoneal disease (PmCRC) represents a phenotype that may derive a greater clinical benefit from bevacizumab (BEV) due to the effect of VEGF on peritoneal spread and ascites production. However, there is concern of greater adverse events, in particular GI perforations. We investigated the clinical benefit and safety of adding BEV to chemotherapy in PmCRC in clinical trial and nontrial populations. Methods: We compared PFS and OS in PmCRC patients receiving first-line chemotherapy with and without BEV in: (1) the AGITG MAX trial (capecitabine [CAP] vs CAP/BEV +/- mitomycin C) and (2) two cancer centers in New South Wales between Jan 2005 and Dec 2012 (any regimen +/- BEV). Secondary endpoints included chemotherapy duration and GI adverse events. Time to event outcomes were estimated using the method of Kaplan-Meier and comparisons made using the logrank test. Proportional hazards models were used to obtain hazard ratios for regression analysis. Results: See Table. 84 MAX and 69 nontrial pts had PmCRC and there was a 37% reduction in the risk of progression in those receiving BEV, which is similar when compared to the nonperitoneal disease pts (HR= 0.63 95% CI 0.52-0.79 p value < 0.001). OS in the PmCRC group was significantly worse than the nonperitoneal group (14.3 months vs 18.8 months p value = 0.02). Chemotherapy duration was similar across the groups. The rate of notable GI adverse events, including GI perforation was not increased in the PmCRC group receiving BEV in either cohort. One pt had a GI perforation after receiving CAP+BEV. Conclusions: Pts with mCRC and peritoneal disease receiving first-line therapy in trial and nontrial populations appear to derive a similar proportional benefit when BEV is added to systemic therapy without an increase in GI adverse events. However, this subgroup of patients continues to have a worse prognosis. [Table: see text]

Inhibitory effects of aromatase inhibitor on estrogen receptor-alpha positive ovarian cancer in mice

BackgroundEstrogen causes proliferation of ovarian cancer cells. Although hormone therapy with an anti-estrogen agent is an optional therapy for recurrent epithelial ovarian cancers, both basic and clinical researches are insufficient. We here examine the efficacy of an aromatase inhibitor (AI) for peritonitis carcinomatosa, the late stage of ovarian cancer.MethodsEstrogen receptor (ER)α was assayed in four ovarian cancer cell lines by the RT-PCR method. Using ovariectomized nude mice, peritonitis carcinomatosa consisting of OVCAR-3 cells with the strongest ERα expression or DISS cells with weaker ERα expression was prepared. The survival period was compared between the letrozole group (5 mg/kg/day orally; n = 10) and the control group (n = 10). In addition, the degree of angiogenesis and occurrence of apoptosis were compared using tumor tissue from the abdominal cavity. The expression of aromatase and the protein involving in ERα signaling were examined in tumors immunohistochemically.ResultsSurvival period in OVCAR-3 tumors was significantly prolonged in the letrozole group, compared with the control group (P < 0.05), whereas that in DISS tumors was not different between the both groups. The microvessel density in tumors and expression of VEGF decreased significantly in the letrozole group compared to the control group. The incidence of apoptosis did not differ significantly between these groups. No adverse event was observed accompanying the administration of letrozole. The expressions of aromatase, ERα and FOXP1 that is associated with ERα signaling were reduced in tumors by letrozole administration.ConclusionsLetrozole was effective for ovarian cancers with abundant expression of ERα. Inhibition of angiogenesis and of ascites production appeared to contribute to prolongation of the survival period.

Is the Indication of Liver Resection for Hepatocellular Carcinoma Expanding with the Application of Laparoscopic Approach?

Liver resection (LR) for the patients of hepatocellular carcinoma, often with chronic liver disease, have high risks of developing significant postoperative complications and multicentric metachronous repeat lesions with the need of repeat treatments. Reduction of surgery-induced parenchymal injury and destruction of the collateral blood/lymphatic flow, which leads to less post-operative ascites production, and facilitation of repeat LR with less adhesion and improved vision/manipulation between adhesions are among the advantages of laparoscopic LR. These characteristics of laparoscopic LR may lead to expanding indication of LR.

Folate-linked lipoplexes for short hairpin RNA targeting claudin-3 delivery in ovarian cancer xenografts

Ovarian cancers highly overexpress folate receptor α (FRα) and claudin3 (CLDN3), both of which are associated with tumor progression and poor prognosis of patients. Downregulation of FRα and CLDN3 in ovarian cancer may suppress tumor growth and promote benign differentiation of tumor. In this study, F-P-LP/CLDN3, a FRα targeted liposome loading with short hairpin RNA (shRNA) targeting CLDN3 was prepared and the pharmaceutical properties were characterized. Then, the antitumor effect of F-P-LP/CLDN3 was studied in an in vivo model of advanced ovarian cancer. Compared with Control, F-P-LP/CLDN3 promoted benign differentiation of tumor and achieved about 90% tumor growth inhibition. In the meantime, malignant ascites production was completely inhibited, and tumor nodule number and tumor weight were significantly reduced (p<0.001). FRα and CLDN3 were downregulated together in tumor tissues treated by F-P-LP/CLDN3. The antitumor mechanisms were achieved by promoting tumor cell apoptosis, inhibiting tumor cell proliferation and reducing microvessel density. Finally, safety evaluation indicated that F-P-LP/CLDN3 was a safe formulation in intraperitoneally administered cancer therapy. We come to a conclusion that F-P-LP/CLDN3 is a potential targeting formulation for ovarian cancer gene therapy.

Should studies of maintenance therapy be maintained in women with ovarian cancer?

Should studies of maintenance therapy be maintained in women with ovarian cancer?

Pure laparoscopic hepatectomy for hepatocellular carcinoma with chronic liver disease

Pure laparoscopic hepatectomy is a less invasive procedure than conventional open hepatectomy for the resection of hepatic lesions. Increases in experiences with the technique, in combination with advances in technology, have promoted the popularity of pure laparoscopic hepatectomy. However, indications for usage and potential contraindications of the procedure remain unresolved. The characteristics and specific advantages of the procedure, especially for hepatocellular carcinoma (HCC) patients with chronic liver diseases, are reviewed and discussed in this paper. For cirrhotic patients with liver tumors, pure laparoscopic hepatectomy minimizes destruction of the collateral blood and lymphatic flow from laparotomy and mobilization, and mesenchymal injury from compression. Therefore, pure laparoscopic hepatectomy has the specific advantage of minimal postoperative ascites production that leads to lowering the risk of disturbance in water or electrolyte balance and hypoproteinemia. It minimizes complications that routinely trigger postoperative serious liver failure. Under adequate patient positioning and port arrangement, the partial resection of the liver in the area of subphrenic space, peri-inferior vena cava area or next to the attachment of retro-peritoneum is facilitated in pure laparoscopic surgery by providing good vision and manipulation in the small operative field. Furthermore, the features of reduced post-operative adhesion, good vision, and manipulation within the small area between the adhesions make this procedure safer in the context of repeat hepatectomy procedures. These improved features are especially advantageous for patients with liver cirrhosis and multicentric and/or metachronous HCCs.

Pseudomyxoma peritonei et pleurae – egy ritka betegségről kezelt esetünk kapcsán

Pseudomyxoma Peritonei (PMP) is a rare condition, which arises from the abdominal cavity and it may also involve the pleural cavity occasionally. Concomitant involvement of both cavities is rare. The disease is characterized by production of mucinous ascites, and a low-grade carcinoma deriving mostly from the appendix. Pleural seeding may be due to diaphragmatic perforation during surgical intervention, tumour progression, or persistent congenital malformation between the abdominal and pleural cavities. A 44 year-old female patient was treated with pleuropulmonary involvement of pseudomyxoma. Since both the lungs and pleura were involved, radical resection could not have been carried out, and cytoreduction with intraoperative chemotherapy was applied only. Pleural progression of pseudomyxoma is uncommon and carries a poor prognosis. Management is similar to abdominal PMP, which involves aggressive cytoreduction and local and/or systemic chemotherapy, if possible. PMP is a very uncommon disease with poor prognosis. Extra-abdominal manifestation is rare, too. However, early recognition of the condition may enhance treatment options.

Portal hypertension-related inflammatory phenotypes: From a vitelline and amniotic point of view

Prehepatic portal hypertension induces a splanchnic low-grade inflammatory response that could switch to high-grade inflammation with the development of severe and life-threatening complications when associated with chronic liver disease. The extraembryonic origin of the portal system maybe determines the regression to an extraembryonic phenotype, i.e., vitellogenic and amniotic, during the evolution of both types of portal hypertension. Thus, prehepatic portal hypertension, or compensated hypertension by portal vein ligation in the rat, is associated with molecular mechanisms related to vitellogenesis, where hepatic steatosis and splanchnic angiogenesis stand out. In turn, extrahepatic cholestasis in the rat induces intrahepatic portal hypertension, or decompensated hypertension, with ascites and hepatorenal syndrome. The splanchnic interstitium, the mesenteric lymphatic system, and the peritoneal mesothelium seem to create an inflammatory pathway that could have a key pathophysiological relevance in the production of ascites. The hypothetical comparison between the ascitic and the amniotic fluid also allows for translational investigation. The induced regression of the splanchnic system to extraembryonic functions by portal hypertension highlights the great relevance of the extraem-bryonic structures even during postnatal life.

Significance of vascular endothelial growth factor in growth and peritoneal dissemination of ovarian cancer.

Vascular endothelial growth factor (VEGF) is a key regulator of angiogenesis which drives endothelial cell survival, proliferation, and migration while increasing vascular permeability. Playing an important role in the physiology of normal ovaries, VEGF has also been implicated in the pathogenesis of ovarian cancer. Essentially by promoting tumor angiogenesis and enhancing vascular permeability, VEGF contributes to the development of peritoneal carcinomatosis associated with malignant ascites formation, the characteristic feature of advanced ovarian cancer at diagnosis. In both experimental and clinical studies, VEGF levels have been inversely correlated with survival. Moreover, VEGF inhibition has been shown to inhibit tumor growth and ascites production and to suppress tumor invasion and metastasis. These findings have laid the basis for the clinical evaluation of agents targeting VEGF signaling pathway in patients with ovarian cancer. In this review, we will focus on VEGF involvement in the pathophysiology of ovarian cancer and its contribution to the disease progression and dissemination.

Anti-MUC1 Monoclonal Antibody (C595) and Docetaxel Markedly Reduce Tumor Burden and Ascites, and Prolong Survival in an in vivo Ovarian Cancer Model

MUC1 is associated with cellular transformation and tumorigenicity and is considered as an important tumor-associated antigen (TAA) for cancer therapy. We previously reported that anti-MUC1 monoclonal antibody C595 (MAb C595) plus docetaxel (DTX) increased efficacy of DTX alone and caused cultured human epithelial ovarian cancer (EOC) cells to undergo apoptosis. To further study the mechanisms of this combination-mediated apoptosis, we investigated the effectiveness of this combination therapy in vivo in an intraperitoneal (i.p.) EOC mouse model. OVCAR-3 cells were implanted intraperitoneally in female athymic nude mice and allowed to grow tumor and ascites. Mice were then treated with single MAb C595, DTX, combination test (MAb C595 and DTX), combination control (negative MAb IgG3 and DTX) or vehicle control i.p for 3 weeks. Treated mice were killed 4 weeks post-treatment. Ascites volume, tumor weight, CA125 levels from ascites and survival of animals were assessed. The expression of MUC1, CD31, Ki-67, TUNEL and apoptotic proteins in tumor xenografts was evaluated by immunohistochemistry. MAb C595 alone inhibited i.p. tumor growth and ascites production in a dose-dependent manner but did not obviously prevent tumor development. However, combination test significantly reduced ascites volume, tumor growth and metastases, CA125 levels in ascites and improved survival of treated mice compared with single agent-treated mice, combination control or vehicle control-treated mice (P<0.05). The data was in a good agreement with that from cultured cells in vitro. The mechanisms behind the observed effects could be through targeting MUC1 antigens, inhibition of tumor angiogenesis, and induction of apoptosis. Our results suggest that this combination approach can effectively reduce tumor burden and ascites, prolong survival of animals through induction of tumor apoptosis and necrosis, and may provide a potential therapy for advanced metastatic EOC.

The mechanism of endostar on ascites formation in vivo and in vitro.

e21013 Background: Angiogenesis and vessel hyperpermeability are the two important factors leading to the formation of ascites. Many studies have demonstrated that Endostar strongly inhibits the growth of a variety of murine and xenotransplanted human tumors by suppressing the neovascularization. The study was designed to explore the mechanism of action correlated with tumor ascites in vivo and in vitro. Methods: In vivo, the mice models of tumor ascites with S180 and H22 cell lines were established. Model animals were randomly distributed into 4 groups with 12 mice respectively. 3 groups received different doses of endostar treatment.The control group received 0.9% normal saline. Four mice in each group were undertaken peritoneal capillar permeability experiments. The mice were slowly injected with 0.25 ml of 5 % Evan blue dye from vena caudalis. After 2 h, the mice were killed to collect ascites. The optical density of Evan blue was measured at 540 nm wavelength to indirectly reflect the concentration of Evan blue dye. In vitro, MTT assay was performed to evaluate the proliferation of S180 and H22 cells in the absence or presence of endostar. Tumor cells (1×104cells/well) were incubated with various concentrations of endostar. Invasiveness of Endostar on tumor cells was determinated by Transwell invasion chamber model. Results: In vivo. Results showed that Endostar (dose at 8,16mg/kg) could markedly suppress ascites production and peritoneum permeability, and down-regulated the levels of VEGF, MMP-2 and CD44v6 in blood serum and ascites fluid (P<0.05, respectively). Endostar showed mild apoptosis and cell cycle on S180 and H22 cells (P>0.05). In vitro. Results from incubation of S180 cells and H22 cells with various concentrations of endostar suggested significant inhibition of VEGF, MMP-2 and CD44v6 by ELISA (P<0.05, respectively),and down-regulated the expressions of VEGFmRNA and OPNmRNA by Real-time PCR, and reduced the expressions of OPN, αVβ3 integrin and HIF-1α by Western blot analysis. Conclusions: Those results revealed for the first time that Endostar could profoundly suppress ascites formation through the expressions of VEGF, MMP2, CD44v6, OPN and αVβ3 integrin, rather than induced cells apoptosis and proliferation.

Endostar inhibits malignant ascites production in mice model bearing tumor ascites.

e21014 Background: Endostar is a modified rh-endostatin expressed and purified in E.coli, an additional nine-amino acid sequence is added at the N-terminal of the protein, which improves the stability and activity of the protein. A series of studies have demonstrated that endostar strongly inhibits the growth of a variety of murine and xenotransplanted human tumors by suppressing the neovascularization. The present study was designed to investigate inhibitory effects of endostar in mice model. Methods: The mice models of tumor ascites with S180 and H22 cells by intraperitoneal inoculation were established. Model animals were randomly distributed into 4 groups with 12 mice respectively. 3 groups received different doses of endostar treatment: 4, 8, and 16 mg/kg, respectively. The control group received 0.9% normal saline.The mice were killed to collect the ascites. Then we analyzed the membrane permeability according to it.Apoptosis and cell cycle of tumor cells in the ascites fluid were detected by flow cytometry. The level of VEGF was determined by ELISA in blood serum and ascites fluid. Kaplan-Meier analysis was performed to determine animal survival time. Results: Establishment of mice model bearing tumor ascites was successful with S180 cells and H22 cells by intraperitoneal inoculation. Doses of 8, 16 mg/kg endostar could suppress volume of ascites, and the number of tumor cells and red cells in the ascites fluid (P<0.05, respectively).The different doses of endostar (dose at 4, 8 and 16mg/kg) could profoundly depress peritoneum permeability (P<0.05, respectively). The 8, 16mg/kg endostar could reduce tumor burden of abdominal cavity, and down-regulate the level of VEGF in blood serum and ascites fluid (P<0.05, respectively). Endostar induced mild apoptosis and cell cycle on S180 and H22 cells (P>0.05, respectively). And survival time of 8, 16mg/kg endostar-treated mice was longer than that of control group mice. Conclusions: Results revealed for the first time that endostar could obviously suppress ascites formation, as well as prolonging survival time of test mice bearing S180 and H22 tumor ascites. Endostar is expected to be a novel therapeutic strategy and drug for treating advanced stage cancer with malignant ascites.

Impact of portal venous hemodynamics on indices of liver function and graft regeneration after right lobe living donor liver transplantation

The aim of this study was to evaluate the effects of portal hemodynamics on indices of liver function and graft regeneration in patients after adult right lobe living donor liver transplantation (R-LDLT). Sixty-four patients who underwent R-LDLT and had an uneventful postoperative course were enrolled in this study. The contribution of portal flow was greater to the recipient grafts versus the donor livers (90.74% versus 69.12%, P < 0.0001). Portal flow variations decreased significantly during the first 10 days after R-LDLT (P < 0.0001); variations in the hepatic arterial flow were more constant during this period (P = 0.812). The mean portal venous pressure (PVP) before recipient hepatectomy (the initial PVP) was 23.1 ± 4.0 mm Hg; the mean PVP after reperfusion (the final PVP) was 15.0 ± 4.3 mm Hg (P < 0.0001). Furthermore, the mean hepatic portal venous gradient (ie, PVP - central venous pressure) before recipient hepatectomy was 17.1 ± 4.3 mm Hg; it decreased to 10.6 ± 4.5 mm Hg after reperfusion (P < 0.0001). These findings suggest that after graft reperfusion, the vascular resistance of the hepatic parenchyma decreased, and there was an associated mild decrease in the portal hypertension. Multiple regression analysis indicated that PVPs correlated significantly with indices of liver function after living donor liver transplantation (P < 0.05). Patients were separated into 4 groups according to their PVP values: group A (initial PVP ≥ 23 mm Hg, final PVP ≥ 15 mm Hg), group B (initial PVP < 23 mm Hg, final PVP ≥ 15 mm Hg), group C (initial PVP ≥ 23 mm Hg, final PVP < 15 mm Hg), and group D (initial PVP < 23 mm Hg, final PVP < 15 mm Hg). Immediately after R-LDLT, the peak values for aspartate aminotransferase, alanine aminotransferase, the international normalized ratio and the average ascites production varied appreciably in these groups. The regeneration rate of the liver graft 3 months after R-LDLT was significantly greater in group A versus the other groups. In conclusion, PVP is a significant hemodynamic factor that influences the functional status of the liver and graft regeneration after R-LDLT.

Topotecan as a molecular targeting agent which blocks the Akt and VEGF cascade in platinum-resistant ovarian cancers

(Objective) Topotecan, a novel topoisomerase-1 inhibitor, is a drug that appears to be effective against platinum-resistant ovarian cancers. However, the molecular mechanisms by which Topotecan treatment inhibits cancer cell proliferation are unclear. We investigated whether Topotecan increases the efficacy of Cisplatin in platinum-resistant ovarian cancer models in vitro and in vivo. (Methods) We used Cisplatin-resistant Caov-3 cells and Cisplatin-sensitive A2780 cells. We examined the effect of Cisplatin and Topotecan on the cell viability of Caov-3 and A2780 cells by MTS assay. We examined the Akt kinase activity, VEGF and HIF-1α expression after Cisplatin and Topotecan by a Western blot analysis. Moreover, we also evaluated the effects of Cisplatin and Topotecan on the intraabdominal dissemination of ovarian cancer in vivo. (Results) Topotecan significantly inhibited Cisplatin-induced Akt activation in Caov-3 cells, but not in A2780 cells. In the presence of Topotecan, Cisplatin-induced growth inhibition and apoptosis were significantly enhanced in Caov-3 cells. Topotecan inhibited not only Cisplatin-induced Akt activation but also VEGF and HIF-1α expression. Moreover, treatment with Topotecan increased the efficacy of Cisplatin-induced growth inhibition in the intraabdominal dissemination and production of ascites in athymic nude mice inoculated with Caov-3 cells. (Conclusion) We herein demonstrated that Topotecan inhibits Akt kinase activity and VEGF transcriptional activation after Cisplatin treatment in platinum-resistant ovarian cancers. We clarified how Topotecan enhanced the clinical activity in the platinum-resistant ovarian cancer. These results provide a rationale for using Topotecan in clinical regimens aimed at molecular targeting agents in platinum-resistant ovarian cancers.

Development of a syngeneic mouse model of epithelial ovarian cancer.

BackgroundMost cases of ovarian cancer are epithelial in origin and diagnosed at advanced stage when the cancer is widely disseminated in the peritoneal cavity. The objective of this study was to establish an immunocompetent syngeneic mouse model of disseminated epithelial ovarian cancer (EOC) to facilitate laboratory-based studies of ovarian tumor biology and preclinical therapeutic strategies.MethodsIndividual lines of TgMISIIR-TAg transgenic mice were phenotypically characterized and backcrossed to inbred C57BL/6 mice. In addition to a previously described line of EOC-prone mice, two lines (TgMISIIR-TAg-Low) were isolated that express the oncogenic transgene, but have little or no susceptibility to tumor development. Independent murine ovarian carcinoma (MOVCAR) cell lines were established from the ascites of tumor-bearing C57BL/6 TgMISIIR-TAg transgenic mice, characterized and tested for engraftment in the following recipient mice: 1) severe immunocompromised immunodeficient (SCID), 2) wild type C57BL/6, 3) oophorectomized tumor-prone C57BL/6 TgMISIIR-TAg transgenic and 4) non-tumor prone C57BL/6 TgMISIIR-TAg-Low transgenic. Lastly, MOVCAR cells transduced with a luciferase reporter were implanted in TgMISIIR-TAg-Low mice and in vivo tumor growth monitored by non-invasive optical imaging.ResultsEngraftment of MOVCAR cells by i.p. injection resulted in the development of disseminated peritoneal carcinomatosis in SCID, but not wild type C57BL/6 mice. Oophorectomized tumor-prone TgMISIIR-TAg mice developed peritoneal carcinomas with high frequency, rendering them unsuitable as allograft recipients. Orthotopic or pseudo-orthotopic implantation of MOVCAR cells in TgMISIIR-TAg-Low mice resulted in the development of disseminated peritoneal tumors, frequently accompanied by the production of malignant ascites. Tumors arising in the engrafted mice bore histopathological resemblance to human high-grade serous EOC and exhibited a similar pattern of peritoneal disease spread.ConclusionsA syngeneic mouse model of human EOC was created by pseudo-orthotopic and orthotopic implantation of MOVCAR cells in a susceptible inbred transgenic host. This immunocompetent syngeneic mouse model presents a flexible system that can be used to study the consequences of altered gene expression (e.g., by ectopic expression or RNA interference strategies) in an established MOVCAR tumor cell line within the ovarian tumor microenvironment and for the development and analysis of preclinical therapeutic agents including EOC vaccines and immunotherapeutic agents.

Effect of the Secretory Small GTPase Rab27B on Breast Cancer Growth, Invasion, and Metastasis

BACKGROUND Secretory GTPases like Rab27B control vesicle exocytosis and deliver critical proinvasive growth regulators into the tumor microenvironment. The expression and role of Rab27B in breast cancer were unknown. METHODS Expression of green fluorescent protein (GFP) fused with wild-type Rab3D, Rab27A, or Rab27B, or Rab27B point mutants defective in GTP/GDP binding or geranylgeranylation, or transient silencing RNA to the same proteins was used to study Rab27B in estrogen receptor (ER)-positive human breast cancer cell lines (MCF-7, T47D, and ZR75.1). Cell cycle progression was evaluated by flow cytometry, western blotting, and measurement of cell proliferation rates, and invasion was assessed using Matrigel and native type I collagen substrates. Orthotopic tumor growth, local invasion, and metastasis were analyzed in mouse xenograft models. Mass spectrometry identified proinvasive growth regulators that were secreted in the presence of Rab27B. Rab27B protein levels were evaluated by immunohistochemistry in 59 clinical breast cancer specimens, and Rab3D, Rab27A, and Rab27B mRNA levels were analyzed by quantitative real-time polymerase chain reaction in 20 specimens. Statistical tests were two-sided. RESULTS Increased expression of Rab27B promoted G(1) to S phase cell cycle transition, proliferation and invasiveness of cells in culture, and invasive tumor growth and hemorrhagic ascites production in a xenograft mouse model (n = 10; at 10 weeks, survival of MCF-7 GFP- vs GFP-Rab27B-injected mice was 100% vs 62.5%, hazard ratio = 0.26, 95% confidence interval = 0.08 to 0.88, P = .03). Mass spectrometric analysis of purified Rab27B-secretory vesicles identified heat-shock protein 90alpha as key proinvasive growth regulator. Heat-shock protein 90alpha secretion was Rab27B-dependent and was required for matrix metalloproteinase-2 activation. All Rab27B-mediated functional responses were GTP- and geranylgeranyl-dependent. Presence of endogenous Rab27B mRNA and protein, but not of Rab3D or Rab27A mRNA, was associated with lymph node metastasis (P < .001) and differentiation grade (P = .001) in ER-positive human breast tumors. CONCLUSIONS Rab27B regulates invasive growth and metastasis in ER-positive breast cancer cell lines, and increased expression is associated with poor prognosis in humans.

Effect of a topoisomerase-1 inhibitor (topotecan) on the efficacy of cisplatin in in vitro and in vivo platinum-resistant ovarian cancer models.

e13160 Background: Topotecan, a novel topoisomerase-1 inhibitor, is one of the drugs that might be effective against platinum-resistant epithelial ovarian cancer. However, the molecular mechanisms by which topotecan treatment inhibits cancer cell proliferation are unclear. The PI3K/Akt cascade is known to be an important survival factor in the signal transduction cascades involved in the cell growth. We investigated whether topotecan increases the efficacy of cisplatin in in vitro and in vivo ovarian cancer models. Methods: Mice: BALB/c Slc-nu/nu. Caov-3 cells: The cells are cisplatin-resistant human-derived ovarian cancer cell (mucinous adenocarcinoma). 5×106 Caov-3 cells were injected i.p. into 6-week-old female athymic nude mice (n=20). Athymic nude mice were injected i.p.with Caov-3 cells. Two weeks after inoculation, athymic nude mice inoculated i.p. with Caov-3 cells were randomized into four groups treated with the following for 6 weeks: (a) vehicle (PBS) alone; (b) CDDP (cisplatin 5 mg/kg) once a week; (c) TPT (topotecan 12.5 mg/kg) once a week; (d) CDDP + TPT (cisplatin 5mg/kg + topotecan 12.5 mg/kg) once a week. Results: Topotecan significantly inhibited the cisplatin-induced Akt activation in Caov-3 cells, but not in A2780 cells. In the presence of topotecan, cisplatin-induced inhibition and apoptosis was significantly enhanced in Caov-3 cells. Moreover, treatment with topotecan increased the efficacy of the cisplatin-induced inhibition in the intraabdominal dissemination and production of ascites in athymic nude mice inoculated i.p. with Caov-3 cells. Topotecan inhibited not only the cisplatin-induced Akt activity but also VEGF transcriptional activation and HIF-1a expression which was induced by cisplatin. Conclusions: These results suggest that combination therapy by cisplatin and topotecan would increase the therapeutic efficacy of the platinum-resistant epithelial ovarian cancer. No significant financial relationships to disclose.

Fifteen millimeters of mercury

Fifteen millimeters of mercury

Inhibitory effect of Ad-hTERTp-HSV-TK/GCV system on mice ascites production planted by liver cancer cell line

Objective To observe the effect of Ad-bTERTp-HSV-TK/GCV system on malignant ascites of mice and probe into its mechanism of action.Methods The SX1 inbred strain mice were injected with H22 cell line of liver cancer and were divided into 4 groups at random.The mice in each group were given corresponding treatment after 48 hours.The production of ascites and survival period were evaluated. The apoptosis rates of tumor cells were detected by FCM.Morphological changes of tumor cells were studied by electromicroscope.Results Compared with other groups.Ad-hTERTp-HSV-TK/GCV Can obviously inhibit the production of ascites(P<0.01),prolong the survival period (P<0.01),and apoptosis rate in this group (27.12±2.12)% was significantly higher than that in other groups.No obvious side effect Was found during the treatment.Conclusion Ad-hTERTp-HSV-TK/GCV system Can inhibit production of ascites and prolong the survical period of mice by inducing apoptosis of hepatoma cells,which is a safe and feasible treatment for hepatoma therapy. Key words: Liver neoplasms,experimental; Gene therapy; Kinase adenosine; Adenoviruses.human; Ascites