Mouse Model Of Epithelial Ovarian Cancer Research Articles

Anticancer Effects of BAF312 (Siponimod) in Epithelial Ovarian Cancer.

Epithelial ovarian cancer (EOC) is a lethal disease that is the fifth leading cause of cancer-related death in women. BAF312 (siponimod) is a potent and selective sphingosine-1-phosphate (S1P) receptor modulator that has been approved as a treatment for multiple sclerosis. In addition to its immunomodulatory action, BAF312 shows preclinical antitumor effects in several cancer types. This study sought to determine whether BAF312 had anticancer properties against EOC using in vitro and in vivo models. EOC cell lines A2780, SKOV3ip1, A2780-CP20, and SKOV3-TR were treated with BAF312 and tested for cell proliferation, apoptosis, and migration using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, fluorescence-activated cell sorting, and migration assays. We investigated the expression of sphingosine-1-phosphate receptor 1 (S1PR1) in most EOC cell lines through western blot analysis. To investigate potential mechanisms, western blot analysis was used to assess the expression of AKT serine/threonine kinase 1 (AKT) and extracellular-regulated kinase (ERK) after BAF312 treatment. We also created poly(D,L-lactide-co-glycolide) nanoparticles encapsulating BAF312 (PLGA-NP-BAF312) for in vivo therapy. The average size and zeta potential of PLGA-NP-BAF312 were determined using dynamic light scattering. The therapeutic efficacy of PLGA-NP-BAF312 was tested in an A2780 tumor-bearing orthotopic mouse model of EOC. S1PR1 was overexpressed in most EOC cell lines. BAF312 significantly reduced cell proliferation and migration while inducing significant apoptosis in all EOC cell lines. PLGA-NP-BAF312 treatment significantly reduced tumor weights in A2780 tumor-bearing mice. Furthermore, the anticancer effects of BAF312 were associated with reduced phosphorylation of ERK and AKT. Our findings show that BAF312 has significant anticancer effects in EOC cells by inhibiting the ERK and AKT pathways, and might potentially be used to treat EOCs.

Abstract 354: The progression of changes in cardiac structure and function in mice with epithelial ovarian cancer

Abstract Introduction: Cancer cachexia, characterized by the progressive loss of muscle mass (i.e., atrophy), has a significant impact on patient prognosis and quality of life. While skeletal muscle atrophy and weakness are well characterized in cancer patients, the progression of cancer-induced cardiac atrophy has received little attention. Preliminary work from our lab has shown that cardiac dysfunction, measured by invasive hemodynamic assessment, precedes the development of metastatic disease in a mouse model of epithelial ovarian cancer (EOC). Yet, the time course of changes in cardiac structure and function throughout the progression of ovarian cancer remain unknown. Thus, we non-invasively assessed changes in cardiac structure and function through the progression of EOC. Methods: To investigate the impact of advanced ovarian cancer on cardiac structure and function, we used an orthotopic, syngeneic mouse model of EOC. This model was generated by injecting transformed murine ovarian surface epithelial cells (ID8; 1.0 × 106) or saline (surgical sham) directly under the ovarian bursa of syngeneic mice. Echocardiography was used to assess changes in cardiac structure and function throughout the progression of EOC at days 45, 60, 75, and 90 post-tumor induction (PTI). Morphometric assessments at each timepoint were used to determine the presence of cardiac atrophy. Results: Echocardiography showed that EOC mice had preserved left ventricular (LV) function (i.e., ejection fraction) with no change in structural dimensions at days 45, 60 and 75 PTI. By day 90 PTI, LV wall thickness and end-systolic dimension decreased, suggesting evidence of cardiac atrophy. Consistent with echocardiographic data, morphometric assessment showed profound atrophy at 90 days PTI, but preserved cardiac mass at all other timepoints compared to shams. Conclusion: Together, this work demonstrates that when non-invasive techniques are used to measure changes in cardiac structure, atrophy is not evident until advanced stage ovarian cancer (i.e., 90 days PTI). Despite this, preliminary work from our lab has shown that cardiac dysfunction is present in EOC mice by 45 days PTI when assessed by invasive hemodynamics. Although echocardiography is a widely used technique for measuring indices of cardiac structure and function, our work demonstrates it lacks sensitivity to detect cardiac deficits until the overt stages of disease that accompany cancer cachexia. Thus, it is important to consider that subclinical cardiac dysfunction may exist in the early stages of cancer progression, however, these changes may go undetected unless more sensitive hemodynamic assessments are employed. Citation Format: Bridget Coyle-Asbil, Leslie M. Ogilvie, Madison Pereira, Kathy Matuszewska, Luca J. Delfinis, Shivam Gandhi, Keith R. Brunt, Christopher G. Perry, Jim Petrik, Jeremy A. Simpson. The progression of changes in cardiac structure and function in mice with epithelial ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 354.

Anti-cancer effect of fenbendazole-incorporated PLGA nanoparticles in ovarian cancer.

Fenbendazole (FZ) has potential anti-cancer effects, but its poor water solubility limits its use for cancer therapy. In this study, we investigated the anti-cancer effect of FZ with different drug delivery methods on epithelial ovarian cancer (EOC) in both in vitro and in vivo models. EOC cell lines were treated with FZ and cell proliferation was assessed. The effect of FZ on tumor growth in cell line xenograft mouse model of EOC was examined according to the delivery route, including oral and intraperitoneal administration. To improve the systemic delivery of FZ by converting fat-soluble drugs to hydrophilic, we prepared FZ-encapsulated poly(D,L-lactide-co-glycolide) acid (PLGA) nanoparticles (FZ-PLGA-NPs). We investigated the preclinical efficacy of FZ-PLGA-NPs by analyzing cell proliferation, apoptosis, and in vivo models including cell lines and patient-derived xenograft (PDX) of EOC. FZ significantly decreased cell proliferation of both chemosensitive and chemoresistant EOC cells. However, in cell line xenograft mouse models, there was no effect of oral FZ treatment on tumor reduction. When administered intraperitoneally, FZ was not absorbed but aggregated in the intraperitoneal space. We synthesized FZ-PLGA-NPs to obtain water solubility and enhance drug absorption. FZ-PLGA-NPs significantly decreased cell proliferation in EOC cell lines. Intravenous injection of FZ-PLGA-NP in xenograft mouse models with HeyA8 and HeyA8-MDR significantly reduced tumor weight compared to the control group. FZ-PLGA-NPs showed anti-cancer effects in PDX model as well. FZ-incorporated PLGA nanoparticles exerted significant anti-cancer effects in EOC cells and xenograft models including PDX. These results warrant further investigation in clinical trials.

DPP4 Inhibitor Sitagliptin Enhances Lymphocyte Recruitment and Prolongs Survival in a Syngeneic Ovarian Cancer Mouse Model.

Simple SummaryThe role of immunity in the development and progression of epithelial ovarian cancer (EOC) is well established. Poor T-cell infiltration is associated with mortality in EOC patients, and recent evidence has suggested that the enzyme DPP4 plays a role in this process. The aim of our study was to evaluate the potential of the clinically-approved DPP4-inhibitor sitagliptin to improve immune responses in mice with EOC. We showed that sitagliptin improved CD8+ T-cell responses in an EOC mouse model, consequently reducing metastatic burden and prolonging survival. These data provide a rationale for the use of DPP4-inhibitors as a second-line treatment for EOC.Immunity plays a key role in epithelial ovarian cancer (EOC) progression with a well-documented correlation between patient survival and high intratumoral CD8+ to T regulatory cell (Treg) ratios. We previously identified dysregulated DPP4 activity in EOCs as a potentially immune-disruptive influence contributing to a reduction in CXCR3-mediated T-cell infiltration in solid tumours. We therefore hypothesized that inhibition of DPP4 activity by sitagliptin, an FDA-approved inhibitor, would improve T-cell infiltration and function in a syngeneic ID8 mouse model of EOC. Daily oral sitagliptin at 50 mg/kg was provided to mice with established primary EOCs. Sitagliptin treatment decreased metastatic tumour burden and significantly increased overall survival and was associated with significant changes to the immune landscape. Sitagliptin increased overall CXCR3-mediated CD8+ T-cell trafficking to the tumour and enhanced the activation and proliferation of CD8+ T-cells in tumour tissue and the peritoneal cavity. Substantial reductions in suppressive cytokines, including CCL2, CCL17, CCL22 and IL-10, were also noted and were associated with reduced CD4+ CD25+ Foxp3+ Treg recruitment in the tumour. Combination therapy with paclitaxel, however, typical of standard-of-care for patients in palliative care, abolished CXCR3-specific T-cell recruitment stimulated by sitagliptin. Our data suggest that sitagliptin may be suitable as an adjunct therapy for patients between chemotherapy cycles as a novel approach to enhance immunity, optimise T-cell-mediated function and improve overall survival.

AAV-mediated expression of 3TSR inhibits tumor and metastatic lesion development and extends survival in a murine model of epithelial ovarian carcinoma.

An integral step in the development of solid tumors is the recruitment of blood vessels to fuel tumor growth. Antiangiogenic therapies can inhibit this process and control solid tumor growth. Thrombospondin-1 is an antiangiogenic protein possessing three type I repeats (3TSR) near the center of the protein and a CD47-binding peptide (CD47) in its C-terminus. Previously, we showed that treatment with recombinant 3TSR induces tumor regression, normalizes tumor vasculature, and improves uptake of chemotherapy drugs in an orthotopic, syngeneic mouse model of advanced stage epithelial ovarian cancer (EOC). While effective, this intervention required daily intraperitoneal injections. To circumvent this, here we employ adeno-associated virus (AAV) gene therapy vectors to express 3TSR alone or in combination with the CD47-binding peptide of TSP-1 and evaluate the impact on tumor development and survival in a mouse model of EOC. A single intraperitoneal injection of 1 × 1011 vg of AAV expressing 3TSR, CD47-binding peptide, or 3TSR + CD47 effectively suppressed primary tumor growth; however, only AAV-3TSR was able to inhibit development of secondary lesions at 90-days post-tumor implantation and significantly improve survival. Taken together, AAV-mediated expression of 3TSR appears safe and effective at inhibiting tumor development and represents a novel, less invasive approach for treating ovarian carcinoma.

A Syngeneic Mouse Model of Epithelial Ovarian Cancer Port Site Metastases

Epithelial ovarian cancer (EOC) is a deadly gynecologic malignancy, but animal models for the study of EOC pathophysiology and drug efficacy are limited. Based on the finding that women with EOC are at risk for metastasis at a trocar site after laparoscopy, we developed a syngeneic murine model of port-site metastasis of EOC. We leveraged the ID8 murine EOC cell line to induce intra-peritoneal tumors in mice. Once durable intraperitoneal tumor was confirmed with bioluminescence imaging, intra-abdominal wall tumors were induced by abdominal wall puncture with a hollow bore needle. This resulted in a robust system in which C57BL/6 mice developed metastatic deposits at a rate of 66.7% ± 10.77; no intra-abdominal wall metastases were seen in control samples (P = .0003, CI 41.16–90.84). Immunodeficient NOD SCID gamma mice developed puncture site metastases in 70% ± 10.0 of mice and also had no metastases documented in control sites (P = .002, CI 42.24–97.76). In addition we were able to demonstrate the presence of immune infiltrates within the metastatic deposits of C57BL/6 mice via IHC. Therefore, in this study we demonstrate the predictable development of invasive abdominal wall metastases in a syngeneic mouse model of EOC. This model enables studies of the metastatic process and provides a novel system in which to test the effect of therapies on a clinically-relevant model in an immune competent mouse.

Epigenetic therapy activates type I interferon signaling in murine ovarian cancer to reduce immunosuppression and tumor burden

Ovarian cancer is the most lethal of all gynecological cancers, and there is an urgent unmet need to develop new therapies. Epithelial ovarian cancer (EOC) is characterized by an immune suppressive microenvironment, and response of ovarian cancers to immune therapies has thus far been disappointing. We now find, in a mouse model of EOC, that clinically relevant doses of DNA methyltransferase and histone deacetylase inhibitors (DNMTi and HDACi, respectively) reduce the immune suppressive microenvironment through type I IFN signaling and improve response to immune checkpoint therapy. These data indicate that the type I IFN response is required for effective in vivo antitumorigenic actions of the DNMTi 5-azacytidine (AZA). Through type I IFN signaling, AZA increases the numbers of CD45+ immune cells and the percentage of active CD8+ T and natural killer (NK) cells in the tumor microenvironment, while reducing tumor burden and extending survival. AZA also increases viral defense gene expression in both tumor and immune cells, and reduces the percentage of macrophages and myeloid-derived suppressor cells in the tumor microenvironment. The addition of an HDACi to AZA enhances the modulation of the immune microenvironment, specifically increasing T and NK cell activation and reducing macrophages over AZA treatment alone, while further increasing the survival of the mice. Finally, a triple combination of DNMTi/HDACi plus the immune checkpoint inhibitor α-PD-1 provides the best antitumor effect and longest overall survival, and may be an attractive candidate for future clinical trials in ovarian cancer.

Targeting of free fatty acid receptor 1 in EOC: A novel strategy to restrict the adipocyte-EOC dependence

ObjectivesAdipocyte derived free fatty acids (FFA) promote epithelial ovarian cancer (EOC) by acting as a fuel source to support the energy requirement of the cancer cells. FFA may also exert biological effects through signaling pathways. Recently, a family of FFA activated G-protein coupled receptors (FFAR/GPCRs) was identified. Our objective was to investigate the role of FFAR/GPCRs in EOC and assess their potential as therapeutic targets. MethodsThe mRNA (RT-PCR) expression of FFAR/GPCR family members (FFAR1/GPR40; FFAR2/GPR43, FFAR3/GPR41, FFAR4/GPR120 and GPR84) was examined in: (1) a syngeneic mouse model of EOC fed high energy diet (60% fat) or regular diet (30% fat), (2) EOC cell lines exposed to free fatty acids and (3) specimens from 13 histologically normal ovaries and 28 high grade ovarian serous carcinomas. The GPR 40 antagonist, GW1100, was used to inhibit FFAR1/GPR40 and cell survival was assayed by MTT in various cell lines. ResultsHigh Grade Serous carcinoma specimens expressed significantly increased GPR40 compared to normal ovaries (p=0.0020). Higher expression was noted in advanced stage disease. ID8 ovarian tumors from mice fed with high fat diet also showed higher GPR40 expression. Exposing EOC cells to FFAs, increased GPR40 expression. Treatment of EOC cell lines with GW100 resulted in growth inhibition and was associated with an alteration in their energy metabolism. ConclusionFFA-induced cancer cell growth may be partly mediated through FFAR1/GPR40. Targeting of FFAR1/GPR40 may be an attractive treatment strategy in EOC, and possibly offers a targeted treatment for a subset of EOC patients.

Abstract A42: TWIST1 as a driver of accelerated tumor growth and drug resistance: Potential novel pathways and therapeutics.

Abstract Introduction: Epithelial ovarian cancer (EOC) is particularly deadly due to its fast rate of growth, dissemination, and chemoresistant recurrences. Novel therapies are urgently needed to limit metastasis and restore platinum sensitivity. We have investigated the role of TWIST1, a basic helix-loop-helix transcription factor, in these processes. TWIST1 is a well-documented regulator of metastasis from solid tumors, which has been linked to cancer stem cell phenotypes, drug resistance, and poor prognosis in several tumor types. Here we present in vitro and in vivo data supporting TWIST1's role in EOC growth and drug resistance, and propose a therapeutic modality to target TWIST1. Methodology: In vitro and in vivo studies: We created two cell lines, derived from the EOC line OVCAR8, that differ only in their TWIST1 expression. The cells were stably transfected with a G418-selectible vector containing either TWIST1 or an shRNA targeting TWIST1, under the control of the CMV viral promoter. Cells from each line were injected IP (intraperitoneally) into NSG mice, allowed to grow for 7 weeks, and tumor burden was evaluated at necropsy. In addition, RNA from each of these OVCAR8 cell lines (TWIST1 overexpressing versus knockdown) was isolated and used for RNA-seq analysis. Raw data was processed using the Galaxy online platform, and differentially expressed genes were further analyzed using Ingenuity Pathway Analysis. To determine the effect of TWIST1 on drug resistance, OVCAR8 cells with and without TWIST1 were treated with varying doses of cisplatin, and survival was quantified using colorimetric cytotoxicity assays. Therapeutic Application: Since we and others have shown that TWIST1's transactivation/protein binding (WR) domain is required for TWIST1 function, we produced a fusion protein comprised of GFP and the WR domain, and used CoIP to evaluate its ability to competitively inhibit TWIST1 binding in HEK 293 cells. Results: TWIST1 knockdown resulted in reduced tumor size and overall tumor burden in an NSG mouse model of EOC. Specifically, all mice receiving TWIST1-overexpressing cells developed primary ovarian tumors which were scored +4 on H&E by a veterinary pathologist, whereas only 50% of animals with TWIST1 knockdown developed ovarian tumors, and only 1 of 4 mice had a tumor that scored +4. In addition, fewer metastases were present in mice receiving TWIST knockdown cells, and no mice with TWIST1 knockdown tumors produced ascites. RNA-seq analysis of the TWIST overexpressing versus knockdown cell lines revealed marked differences in genes and pathways responsible for cell migration and cell survival, including upregulation of genes implicated in metastasis, DNA repair, and resistance to apoptosis in TWIST1 overexpressing cells. TWIST1 overexpressing cells also showed less cell death following cisplatin treatment. Finally, in our in vitro inhibitor tests, a GFP-WR domain fusion protein successfully reduced TWIST1 binding to its partner protein NFkB-p65 in HEK 293 cells. Conclusions: The importance of TWIST1 for tumor growth has been demonstrated for multiple tumor types – including ovarian cancer, as presented here. Our RNA-seq analysis reveals a complex network of TWIST1-mediated factors implicated in cell movement, survival, and resistance to DNA damaging agents, and we are working to determine the most important of these pathways in EOC. Most importantly, we have shown that TWIST1 expression in EOC cells leads to greater tumor burden in a mouse model, and by knocking down TWIST1, we were able to prevent peritoneal metastases and reduce tumor size. Therefore, we believe that TWIST1 is an attractive therapeutic target in EOC, owing to its roles at the crossroads of metastasis, drug resistance, and tumor growth. Citation Format: Cai M. Roberts, Michelle A. Tran, James Finlay, Sophia Allaf Shahin, Joana Loeza, Leslie Cisneros, Emily Ye, Thanh Dellinger, Carlotta A. Glackin. TWIST1 as a driver of accelerated tumor growth and drug resistance: Potential novel pathways and therapeutics. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: Exploiting Vulnerabilities; Oct 17-20, 2015; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(2 Suppl):Abstract nr A42.

Targeting myeloid cells in the tumor microenvironment enhances vaccine efficacy in murine epithelial ovarian cancer.

Epithelial ovarian cancer (EOC) is typically diagnosed at advanced stages, and is associated with a high relapse rate. Patients in remission are ideal candidates for immunotherapy aimed at cure or prolonging disease-free periods. However, immunosuppressive pathways in the tumor microenvironment are obstacles to durable anti-tumor immunity. In a metastatic syngeneic mouse model of EOC, immunosuppressive macrophages and myeloid-derived suppressor cells (MDSCs) accumulate in the local tumor environment. In addition, resident peritoneal macrophages from non-tumor-bearing mice were highly immunosuppressive, abrogating stimulated T cell proliferation in a cell contact-dependent manner. Immunization with microparticles containing TLR9 and NOD-2 ligands (MIS416) significantly prolonged survival in tumor-bearing mice. The strategy of MIS416 immunization followed by anti-CD11b administration further delayed tumor progression, thereby establishing the proof of principle that myeloid depletion can enhance vaccine efficacy. In patients with advanced EOC, ascites analysis showed substantial heterogeneity in the relative proportions of myeloid subsets and their immunosuppressive properties. Together, these findings point to immunosuppressive myeloid cells in the EOC microenvironment as targets to enhance vaccination. Further studies of myeloid cell accumulation and functional phenotypes in the EOC microenvironment may identify patients who are likely to benefit from vaccination combined with approaches that deplete tumor-associated myeloid cells.

Combined In Vivo Molecular and Anatomic Imaging for Detection of Ovarian Carcinoma-Associated Protease Activity and Integrin Expression in Mice

Most patients with epithelial ovarian cancer (EOC) experience drug-resistant disease recurrence. Identification of new treatments is a high priority, and preclinical studies in mouse models of EOC may expedite this goal. We previously developed methods for magnetic resonance imaging (MRI) for tumor detection and quantification in a transgenic mouse model of EOC. The goal of this study was to determine whether three-dimensional (3D) fluorescence molecular tomography (FMT) and fluorescent molecular imaging probes could be effectively used for in vivo detection of ovarian tumors and response to therapy. Ovarian tumor-bearing TgMISIIR-TAg mice injected with fluorescent probes were subjected to MRI and FMT. Tumor-specific probe retention was identified in vivo by alignment of the 3D data sets, confirmed by ex vivo fluorescent imaging and correlated with histopathologic findings. Mice were treated with standard chemotherapy, and changes in fluorescent probe binding were detected by MRI and FMT. Ovarian tumors were detected using probes specific for cathepsin proteases, matrix metalloproteinases (MMPs), and integrin αvβ3. Cathepsin and integrin αvβ3 probe activation and retention correlated strongly with tumor volume. MMP probe activation was readily detected in tumors but correlated less strongly with tumor volume. Tumor regression associated with response to therapy was detected and quantified by serial MRI and FMT. These results demonstrate the feasibility and sensitivity of FMT for detection and quantification of tumor-associated biologic targets in ovarian tumors and support the translational utility of molecular imaging to assess functional response to therapy in mouse models of EOC.

Abstract 3513: The anti-tumor effect of activated natural killer cells in xenograft mouse models with epithelial ovarian cancer

Abstract Objectives: Ovarian cancer is the most common cause of death from gynecologic cancer in women. Because of the lack of effective screening and warning sign, 60-70% of patients were diagnosed at advanced stage. At present the treatment of advanced ovarian cancer is cytoreductive surgery followed by chemotherapy, and the overall response rate is 70-80%. But 50-75% of responders will relapse within about 18 -24 months. Therefore, new strategies of treatment, for examples, immunotherapy or targeted therapy, are necessary. This study is to evaluate the anti-tumor effect of activated natural killer (NK) cells in xenograft mouse model with epithelial ovarian cancer(EOC). Methods: We have made the OVCAR3-Luc stable cell line by transient transfection of pGL3-Luciferase (G418) gene. The xenograft mouse model of EOC was constructed by the intra-peritoneal injection of OVCAR3-Luc ovarian cancer cell line to the NOD/scid mice. After we confirm the tumor formation with IVIS spectrum, activated NK cells were injected with intra-peritoneal route. There was no treatment to the negative control and paclitaxel (10mg/kg) was used to the positive control. The tumor growth was evaluated by serial IVIS spectrum and biopsy at 8 weeks after any treatment. Results: Activated NK cells were injected at concentration 1 × 107 cells per mouse with intra-peritoneal route. This procedure was repeated 5 times. The NK cells significantly inhibited the tumor growth of OVCAR3-Luc-induced NOD/scid mice, up to the 57.1% comparing the negative control. Although paclitaxel decrease 69.3% of the tumor growth, it is not different to that of NK cell treating mice. Furthermore, side effects involving hair loss, abnormal behavior and weight low were not shown in NK cell-injected mouse models. Conclusion: We confirmed that activated NK cells are able to inhibit ovarian tumor growth without side effect in xenograft models. Therefore, this research demonstrates that NK cell-based immunotherapy might be desirable to the patients with ovarian cancer as an adoptive immunotherapy. 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 3513. doi:1538-7445.AM2012-3513

The Thrombospondin-1 Mimetic ABT-510 Increases the Uptake and Effectiveness of Cisplatin and Paclitaxel in a Mouse Model of Epithelial Ovarian Cancer

Epithelial ovarian cancer (EOC) comprises approximately 90% of ovarian cancers and arises from the surface epithelium. Typical treatment of EOC involves cytoreductive surgery combined with chemotherapy. More recent therapies have targeted the tumor vasculature using antiangiogenic compounds such as thrombospondin-1 (TSP-1). TSP-1 mimetic peptides such as ABT-510 have been created and have been in various clinical trials. We have previously shown that ABT-510 reduces abnormal vasculature associated with tumor tissue and increases the presence of mature blood vessels. It has been hypothesized that treatment with antiangiogenic compounds would allow increased delivery of cytotoxic agents and enhance treatment. In this study, we evaluated the potential role of ABT-510 and various chemotherapeutics (cisplatin and paclitaxel) on tumor progression, angiogenesis, and the benefits of combinational treatments on tissue uptake and perfusion using an orthotopic syngeneic mouse model of EOC. Animals were treated with ABT-510 (100 mg/kg per day) alone or in combination with cisplatin (2 mg/kg per 3 days) or paclitaxel (10 mg/kg per 2 days) at 60 days after tumor induction. Radiolabeled and fluorescently labeled paclitaxel demonstrated a significant increase in tumor uptake after ABT-510 treatment. Combined treatment with ABT-510 and cisplatin or paclitaxel resulted in a significant increase in tumor cell and tumor endothelial cell apoptosis and a resultant decrease in ovarian tumor size. Combined treatment also regressed secondary lesions and eliminated the presence of abdominal ascites. The results from this study show that through vessel normalization, ABT-510 increases uptake of chemotherapy drugs and can induce regression of advanced ovarian cancer.

Epithelial–stromal interaction increases cell proliferation, survival and tumorigenicity in a mouse model of human epithelial ovarian cancer

Objective. Ovarian cancer is the fourth leading cause of cancer-related deaths among women and is among the least understood of all cancers. The objective of this study was to determine the effect of ovarian epithelial and stromal cell interaction in a mouse model of epithelial ovarian cancer (EOC) that closely resembled the human disease. Methods. A mouse model of EOC was generated by orthotopic injection of an ID8 mouse ovarian surface epithelial cell line (MOSEC) under the ovarian bursa of syngeneic mice and tissue was collected to evaluate factors contributing to the formation and development of ovarian tumors. Results. By 90 days post-injection, mice were moribund and had developed large primary ovarian tumors, secondary tumors within the peritoneal cavity, and extensive ascites fluid production. Tumors were hypervascularized and were characterized as serous epithelial carcinomatosis, which replicates the most common form of human ovarian cancer. Cells isolated from ascites fluid were more proliferative with increased expression of survival factors compared to original MOSEC cells and cells obtained from the abdomen following intraperitoneal injection. Orthotopic injection of these cells under the ovarian bursa resulted in more aggressive tumorigenesis, with mice becoming moribund at 60 days post-injection compared with 90 days post-injection with the original ID8-MOSEC cell line. Discussion. This study describes the generation of an orthotopic, syngeneic model of ovarian cancer, which replicates the phenotype of the human disease. Expression of angiogenic and proliferative factors, and the interaction of epithelial cells with the ovarian stroma are important factors in ovarian tumorigenesis.

Magnetic resonance imaging for detection and determination of tumor volume in a genetically engineered mouse model of ovarian cancer

Our laboratory developed a transgenic mouse model of spontaneous epithelial ovarian cancer (EOC) in which tumors are initiated by expression of the early region of the Simian Virus 40 (SV40) under transcriptional control of the 5’ upstream regulatory region of the Müllerian inhibiting substance type II receptor (MISIIR) gene. Female TgMISIIR-TAg-DR26 transgenic mice develop bilateral ovarian tumors with variable latency and survive an average of 152 days. In the absence of reliable methods for disease detection and evaluation of therapeutic response, preclinical studies of this transgenic mouse model of EOC would be limited to longitudinal experiments involving large numbers of animals with euthanasia as the endpoint. Therefore, a non-invasive method for detecting tumors, measuring tumor volume and calculating parameters relevant to the evaluation of therapeutic or preventive interventions (i.e., tumor growth rates, tumor initiation, tumor regression and the time for tumors to reach a given size) is required. We developed and optimized a non-invasive Magnetic Resonance Imaging (MRI) scanning protocol to obtain high resolution abdominal images that is well tolerated by mice. Superior contrast and contrast to noise ratio (CNR) was found with Gd-DTPA contrast enhanced T1-weighted sequences. Image sets in both the axial and coronal orientations for redundant measurements of normal ovary and ovarian tumor volume can be acquired in approximately 20 minutes. Accuracy of MRI-based ovary and tumor volume determinations was verified by standard volume measurements at necropsy. Serial imaging studies were performed on 41 ovarian cancer bearing TgMISIIR-TAg-DR26 transgenic mice to quantitate tumor progression over time in this model. A chemotherapy study was conducted on TgMISIIR-TAg-DR26 transgenic mice using a standard combination therapy consisting of cisplatin and paclitaxel. Our results demonstrate that MRI is well tolerated and can be repeated in serial imaging studies, permitting quantitative analysis of tumor growth and progression and response to therapeutic interventions.

Cyclooxygenase-1 Is Overexpressed in Multiple Genetically Engineered Mouse Models of Epithelial Ovarian Cancer

Cyclooxygenases-1 and -2 (Cox-1 and Cox-2) are two distinct isoforms that catalyze the conversion of arachidonic acid to prostaglandins. The role of Cox-2 in a variety of cancers is well recognized, but the contribution of Cox-1 remains much less explored. We have previously shown that human epithelial ovarian tumors have increased levels of Cox-1, but not Cox-2. We also observed that Cox-1 is highly expressed in a mouse model of epithelial ovarian cancer (EOC), which lacks p53 but overexpresses c-myc and K-ras or c-myc and Akt. More importantly, a Cox-1-selective inhibitor, SC-560, attenuates EOC growth. In the present investigation, we used various genetically engineered mouse models of EOC to determine whether Cox-1 overexpression is unique to specific genetic and oncogenic alterations or is widespread. These models include: (a) deletion of both p53 and Rb, (b) induction of the transforming region of SV40 under the control of Mullerian inhibitory substance type II receptor, or (c) activation of K-Ras in the absence of Pten locally in the ovarian surface epithelium. We found that these three models, which produce spontaneous EOC, also show up-regulated expression of Cox-1, but not Cox-2. The results provide further evidence that Cox-1 overexpression is common in various models of EOC. Thus, Cox-1 serves as a potential marker of EOC and is a possible target for the prevention and/or treatment of this deadly disease.

Cyclooxygenase-1 Is a Potential Target for Prevention and Treatment of Ovarian Epithelial Cancer

The precise genetic and molecular defects underlying epithelial ovarian cancer (EOC) remain largely unknown, and treatment options for patients with advanced disease are limited. Cyclooxygenases (COX-1 and COX-2) catalyze the conversion of arachidonic acid to prostaglandins. Whereas overwhelming evidence suggests a role for COX-2 in a variety of cancers, the contribution of COX-1 remains much less explored. The expression status of COX isoforms in ovarian cancers also remains confusing. We have previously shown that human epithelial ovarian tumors have increased levels of COX-1 but not COX-2. To more carefully examine the role of COXs in ovarian cancer, we used a mouse model of EOC in which genetic and oncogenic modifications were experimentally engineered into ovarian surface epithelial cells (OSE) thought to be the cells of origin for human EOC. These OSE cells produce tumors when allografted into host mice. Using multiple approaches, we observed that OSE cells and the tumors comprised of these cells express high levels of COX-1 but not COX-2. Prostacyclin (PGI(2)) is the major prostaglandin generated downstream of COX-1 in these cells, and SC-560, a COX-1-selective inhibitor, dramatically inhibits PGI(2) production. More importantly, SC-560 reduced the growth of tumors when OSE cells were allografted in nude female mice. In contrast, the COX-2-selective inhibitor celecoxib had little effect on tumor growth. The growth inhibitory effects of SC-560 result from reduced cell proliferation and/or accelerated apoptosis. Our results imply COX-1 as a target for the prevention and/or treatment of EOC.