Mouse Model Of Gastric Cancer Research Articles

Knockdown of hypoxia-inducible factor-1α accelerates peritoneal dissemination via the upregulation of MMP-1 expression in gastric cancer cell lines

This study was performed to clarify the role of hypoxia-inducible factor-1 α (HIF-1α) in the development of peritoneal dissemination in a xenograft mouse model of gastric cancer. HIF-1α knockdown (KD) and control (SC) gastric cancer cells, which were established using the MKN45 and MKN74 cell lines, were studied. The two paired cell lines were directly inoculated into the peritoneal cavity of nude mice. The number and the weight of disseminated nodules were compared between tumors generated from the KD and SC cells. In addition, the molecular mechanism was addressed through analysis of the expression levels of metastasis-related genes. The MKN45-KD cell line demonstrated significantly greater numbers of disseminated nodules and formed a larger tumor mass than the MKN45-SC cell line (p<0.05). MKN74-KD cells also tended to induce a greater number of nodules and to produce those with a heavier weight than the SC cells. An in vitro adhesion assay revealed differing results regarding the adhesion activity to extracellular matrix and monolayer mesothelium cells of the gastric cancer cells derived from the various parental cells. However, the expression of MMP-1 mRNA in the disseminated nodules was significantly increased in the KD cells compared to the SC cells derived from the two parental cell lines (p<0.01). An immunohisto-chemical study further demonstrated that there was stronger staining for MMP-1 in the MKN74-KD in comparison to MKN74-SC cells. Loss of HIF-1α may contribute to the development of aggressive peritoneal dissemination via the upregulation of MMP-1 in gastric cancer cells.

Antitumor effect and pharmacokinetics of intraperitoneal NK105, a nanomicellar paclitaxel formulation for peritoneal dissemination

The intraperitoneal administration of paclitaxel has been shown to be a promising treatment strategy for peritoneal malignancy. The present study evaluated the effects of intraperitoneal administration of NK105, a paclitaxel-incorporating micellar nanoparticle, which has been shown to have a remarkable effect in a mouse model of gastric cancer. Intraperitoneal NK105 significantly reduced peritoneal tumors in vivo compared with the conventional paclitaxel formulation of paclitaxel solubilized in Cremophor EL and ethanol (PTX-Cre). Moreover, intraperitoneal NK105 significantly reduced the size of subcutaneously inoculated tumors, whereas no such effect was seen with PTX-Cre. Similar systemic toxic effects were observed following the intraperitoneal administration of both NK105 and PTX-Cre. Although NK105 disappeared rapidly almost within a day from the peritoneal cavity, the paclitaxel concentration in peritoneal nodules 4 h after intraperitoneal administration was significantly higher in the NK105 group than in the PTX-Cre group (P < 0.05), whereas there were no significant differences in liver paclitaxel concentrations between the two groups. We also evaluated the pharmacokinetics following intraperitoneal administration of NK105 and PTX-Cre. Serum paclitaxel concentrations 6, 12, 24, and 48 h after the intraperitoneal administration of the drugs were significantly higher in the NK105 than the PTX-Cre group. Furthermore, the peak serum concentration was higher in the NK105 than PTX-Cre group (24 100 ± 3560 vs 108 ± 25 ng/mL, respectively; P < 0.001), as was the area under the concentration-time curve from 0 to 48 h (191 000 ± 32 100 vs 1500 ± 108 ng·h/mL, respectively; P < 0.001). Therefore, intraperitoneal chemotherapy with nanoparticulate paclitaxel NK105 may offer a novel treatment strategy for improving drug delivery in gastric cancer with peritoneal dissemination because of enhanced drug penetration into peritoneal nodules and its prolonged presence in the systemic circulation.

Abstract 1339: The role of TNF-α in PGE2-induced tumorigenesis in gastric cancer mouse model

Abstract Accumulating evidence has indicated that chronic inflammation is associated with cancer development. Mouse genetic studies have shown that the proinflammatory mediator, PGE2, plays an essential role in gastrointestinal tumorigenesis. Moreover, the proinflammatory cytokine TNF-α also plays a key role in tumorigenesis, including colitis-associated colon cancer. However, the relationship between PGE2 and TNF-α in tumorigenesis has not yet been fully investigated. We previously constructed a gastric tumor mouse model, Gan mice, that recapitulates human intestinal-type gastric cancer, in which both the PGE2 pathway and Wnt signaling are activated simultaneously in the stomach by transgenic expression of the COX-2, mPGES-1, and Wnt1 genes. The induction of the PGE2 pathway triggers an inflammatory response in the Gan mouse stomach, and the expression of proinflammatory cytokines and chemokines is induced in gastric tumor tissues. We herein examined the role of TNF-α in the PGE2-induced inflammation-associated gastric tumorigenesis by crossing Gan mice with TNF-α knockout mice. Importantly, the gastric tumorigenesis in the Gan TNF-α (-/-) compound mice was significantly suppressed in comparison to the control Gan mice. Moreover, the β-catenin level decreased significantly in the gastric tumors of Gan TNF-α (-/-) mice, suggesting a role for TNF-α in the promotion of Wnt signaling. Interestingly, inflammatory cells, including macrophages and CD4+ T cells, still infiltrated into the Gan TNF-α (-/-) mouse tumors, and the expression levels of IL-1α and IL-6 were almost the same as those in the control Gan mouse tumors. Accordingly, it is possible that TNF-α is an important tumor-promoting factor for gastric tumorigenesis, although TNF-α is not essential for PGE2-induced inflammation in the stomach. Notably, macrophages in the Gan TNF-α (-/-) mouse tumors lost their expression of the M2 marker CD206. It has been suggested that macrophages polarized to the M2(-like) type are important for tissue remodeling and tumorigenesis. It is therefore possible that TNF-α is required for macrophage polarization to a pro-tumorigenic status, which may be important for Wnt activation and gastric tumor promotion. 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 1339. doi:1538-7445.AM2012-1339

Abstract 4436: Chronic inflammation and gastric carcinogenesis: the role of CD44 variant expression

Abstract CD44 is a marker of tumor-initiating cells in gastric cancer. Recently, we have reported that a CD44 variant (CD44v) interacts with xCT, a glutamate-cystine transporter, and manifests enhanced protection against reactive oxygen species (ROS), contributing to tumor growth and resistance to chemo- or radiotherapy. Although an inflammation related metaplasia and/or dysplasia of gastric epithelium is considered as key events in gastric carcinogenesis, the cause or process of these phenomena is not known. We investigated the relationship between CD44v expression and gastric carcinogenesis, using three lineages of gastric cancer mouse models, K19-Wnt1, K19-C2mE, and K19-Wnt1/C2mE mice. We also used CD44-/- K19-Wnt1/C2mE mice and Helicobacter. felis-infected C57BL/6 mice. Chronic inflammation triggered by Helicobactor infection played a major role in CD44v related gastric carcinogenesis. CD44v expression and nuclear factor-kappa B (NF-κB) activation were promoted by inflammatory cytokines, IL-1α and TNF-α, in gastric cancer cell lines. Thus, CD44v+ tumor cell expansion induced by inflammation via NF-κB pathway might contribute to tumor development. Finally, an xCT inhibitor sulfasalazine treatment significantly suppressed tumor growth in K19-Wnt1/C2mE mice. These findings suggest a significant role of CD44v in inflammation-mediated gastric carcinogenesis via ROS defense. 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 4436. doi:1538-7445.AM2012-4436

WNT6 is a novel target gene of caveolin-1 promoting chemoresistance to epirubicin in human gastric cancer cells

Resistance to chemotherapy is a major obstacle for curative treatment of human gastric cancer (GC). However, the underlying molecular mechanisms are largely unknown. Wingless-type MMTV integration site family members (WNTs) are secreted glycoproteins involved in embryogenesis and, on inappropriate expression in the adult, in cancer. Here, we show expression of WNT6 in GC patient specimens, human GC cell lines and in a mouse model of GC. In human GC cells, WNT6 expression was enhanced by caveolin-1 (Cav1), a scaffold protein of plasma membrane caveolae. WNT6 knock-down and overexpression experiments demonstrated that WNT6 increased the resistance to apoptotic cell death induced by the anthracycline chemotherapeutics epirubicin (Epi) and doxorubicin (Dox). Epi increased the activity of the human WNT6 promoter through Cav1-dependent binding of β-catenin to the proximal WNT6 promoter. Epi increased both WNT6/Wnt6 and Cav1 expression in human GC cells and within the tumor area of a murine model of GC (CEA424-SV40 TAg). In GC patients, WNT6 expression was positively associated with the tumor stage and the nodal status, and inversely correlated with the response to ECF (Epi, cisplatin, 5-fluorouracil) chemotherapy. These results showed that WNT6 and Cav1 are upregulated by chemotherapeutics and enhance the resistance of GC cells to anthracycline drugs. Understanding the molecular mechanisms driving WNT6/Cav1-induced drug resistance will provide benefits in developing new therapies for GC.

Urinary Metabolomic Analysis of Human Gastric Cancer Mouse Models and Patients Using Gas Chromatography/Mass Spectrometry

Gastric cancer is the second cause of cancer deaths in China. To identify potential markers for screening or diagnosis of gastric cancer, we coupled xenotransplantation mouse models with a urine metabolomic approach. SGC-7901 gastric cancer cells were subcutaneously or orthotopically implanted into nude mice to establish metastasis and non-metastasis mouse models. Urine samples from mice bearing tumors or gastric cancer patients and their healthy controls were collected and subjected to gas chromatography and mass spectrometry (GC/MS) analysis. Metabolic data were analyzed using Mann-Whitney test to find urinary biomarkers for gastric cancer. Diagnostic models for gastric cancer mice and patients were constructed using principal components analysis (PCA) and validated with the area under the curve (AUC) of receiver operating characteristic (ROC) curves. The results indicated these metabolites mainly include lactic acid, serine, proline, malic acid, and fatty acids. The PCA models discriminated all gastric cancer mice or most gastric cancer patients including six of seven early stage patients, from their healthy controls with AUC value of 1.0 or 0.996, respectively. In addition, they were able to differentiate between metastatic and non-metastatic mice with AUC value of 1.0, as well as between invasive/metastatic and non-invasive cancers with AUC value of 0.982. Our data suggest that there are significant metabolic alterations during progression of gastric cancer and the potential metabolic biomarkers could be useful for screening and early diagnosis of gastric cancer progression.

Characterization of gastric cancer models from different cell lines orthotopically constructed using improved implantation techniques

To develop orthotopic gastric cancer mouse models from different cell lines and characterize the tumor features to assist further in preclinical trials and clinical treatment strategies. Human gastric cancer SGC-7901 and BGC-823 cell suspensions were injected subcutaneously into nude mice to develop solid tumors, and tumor tissue pieces were then implanted under the serous coat of the stomach. An autopsy was performed on all animals of the SGC-7901 and BGC-823 models to observe the primary tumor growth and metastases using pathological and immunohistochemical methods. Both models showed large tumors in situ resulting in pressure and infiltration of the adjacent organs. The gastric cavity became smaller, along with stenosis of the cardia or pylorus. There were biological and statistical differences between the two models. The metastasis rate in involved organs (lymph nodes, kidney, spleen, testis) was significantly higher in the BGC-823 model compared to the SGC-7901 model (P < 0.05 or P < 0.01). The median survival of the BGC-823 model was shorter than that of SGC-7901 (23 d vs 84 d, P < 0.05). Histopathologically, the primary tumor and metastatic lesions of the two models showed obvious atypia and mucus in the cytoplasm. Compared with the SGC-7901 model, BGC-823 appeared more poorly differentiated (absence of adenoid structure), had a smaller volume, and richer capillary structure. Immunohistochemical staining revealed cytokeratin 20 and epithelial membrane antigen expression was positive in the SGC-7901 tumors, while negative in BGC-823 ones. Models using the SGC-7901 and BGC-823 cell lines were established which could function in gastric cancer research on carcinogenesis mechanism and drug discovery. The two models showed different tumor behavior and the latter was more malignant than the former.

Classification OMS 2010 des tumeurs digestives : la quatrième édition

Gastric cancer is a major source of global cancer mortality with limited treatment options and poor patient survival. As our molecular understanding of gastric cancer improves, we are now beginning to recognize that these cancers are a heterogeneous group of diseases with incredibly unique pathogeneses and active oncogenic pathways. It is this molecular diversity and oftentimes lack of common oncogenic driver mutations that bestow the poor treatment responses that oncologists often face when treating gastric cancer. In this review, we will examine the treatments for gastric cancer including up-to-date molecularly targeted therapies and immunotherapies. We will then review the molecular subtypes of gastric cancer to highlight the diversity seen in this disease. We will then shift our discussion to basic science and gastric cancer mouse models as tools to study gastric cancer molecular heterogeneity. Furthermore, we will elaborate on a molecular process termed paligenosis and the cyclical hit model as key events during gastric cancer initiation that impart nondividing mature differentiated cells the ability to re-enter the cell cycle and accumulate disparate genomic mutations during years of chronic inflammation and injury. As our basic science understanding of gastric cancer advances, so too must our translational and clinical efforts. We will end with a discussion regarding single-cell molecular analyses and cancer organoid technologies as future translational avenues to advance our understanding of gastric cancer heterogeneity and to design precision-based gastric cancer treatments. Elucidation of interpatient and intratumor heterogeneity is the only way to advance future cancer prevention, diagnoses and treatment.

Combination immunotherapy and active-specific tumor cell vaccination augments anti-cancer immunity in a mouse model of gastric cancer

BackgroundActive-specific immunotherapy used as an adjuvant therapeutic strategy is rather unexplored for cancers with poorly characterized tumor antigens like gastric cancer. The aim of this study was to augment a therapeutic immune response to a low immunogenic tumor cell line derived from a spontaneous gastric tumor of a CEA424-SV40 large T antigen (CEA424-SV40 TAg) transgenic mouse.MethodsMice were treated with a lymphodepleting dose of cyclophosphamide prior to reconstitution with syngeneic spleen cells and vaccination with a whole tumor cell vaccine combined with GM-CSF (a treatment strategy abbreviated as LRAST). Anti-tumor activity to subcutaneous tumor challenge was examined in a prophylactic as well as a therapeutic setting and compared to corresponding controls.ResultsLRAST enhances tumor-specific T cell responses and efficiently inhibits growth of subsequent transplanted tumor cells. In addition, LRAST tended to slow down growth of established tumors. The improved anti-tumor immune response was accompanied by a transient decrease in the frequency and absolute number of CD4+CD25+FoxP3+ T cells (Tregs).ConclusionsOur data support the concept that whole tumor cell vaccination in a lymphodepleted and reconstituted host in combination with GM-CSF induces therapeutic tumor-specific T cells. However, the long-term efficacy of the treatment may be dampened by the recurrence of Tregs. Strategies to counteract suppressive immune mechanisms are required to further evaluate this therapeutic vaccination protocol.

Effects of combined therapy of LY294002 and SN50 on nude mice model with gastric cancer

To investigate the effect of phosphatidylinositol 3-kinase inhibitor LY294002 combined with NF-κB P65 nuclear translocation inhibitor SN50 on the tumor cell growth and apoptosis using a nude mouse model of gastric cancer. Human gastric cancer cell strain SGC7901 was transplanted subcutaneously to nude mice to establish tumor models. Model mice were randomly divided into the control group, the LY294002 treatment group, the SN50 treatment group, and the LY294002+SN50 treatment group, with 5 in each group. After being treated for 10 days, the inhibition rate of tumor growth was ascertained by measuring the size of tumor. Immunohistochemical method was used to detect the expression levels of Bcl-2, P53 and Bax proteins and transmission electron microscopy to investigate the apoptosis of tumor cells. On the 10th day after treatment, the inhibition rate of gastric cancer cellular growth in the LY294002+SN50 group was (49.2±2.5)%, which was significantly higher than that in the LY294002 group(29.4±1.5)% and SN50 group (19.7±1.6)%(P<0.05). In comparison with the other two groups, LY294002+SN50 group exhibited more severe apoptosis, with expression of Bcl-2 decreased and that of P53 and Bax increased more significantly(P<0.05). LY294002 combined with SN50 inhibits the growth of SGC7901 transplanted tumor and aggravates the apoptosis of gastric cancer cells in nude mice model.

Abstract 1761: Anti-tumor activity of trastuzumab-DM1 in combination with pertuzumab in a gastric cancer model

Abstract Objective: Anti-HER2 therapy is considered to be important for gastric cancer as well as breast cancer. In a previous study, we showed the combinational efficacy of trastuzumab and pertuzumab (an anti-HER2 antibody which binds a different HER2 domain from trastuzumab) in HER2-positive gastric cancer models (AACR, 2010). In the present study, we investigated the usefulness of a HER2 antibody-cytotoxic drug conjugate, trastuzumab-DM1 (T-DM1) as monotherapy and in combination with pertuzumab for HER2-positive gastric cancer as a novel therapy. Methods: A HER2 over-expressing human gastric cancer, NCI-N87 mouse xenograft model was used for examining the efficacy. T-DM1 was administered intravenously once every 3 weeks. Pertuzumab was administered intra-peritoneally once a week. Tumor growth inhibition (TGI) after 6 weeks of treatment was determined from tumor volume measurements. Apoptosis activity was measured with the activity of caspase 3/7. To measure ADCC activity, a real-time cell analyzer was used. Results: T-DM1 (1.25, 2.5, 5, 10, 20, 40 or 80 mg/kg) as monotherapy showed tumor growth inhibition dose-dependently in the NCI-N87 model. The maximum effective dose was 10 mg/kg and TGI on day 42 after treatment started was 113%. T-DM1 (5 mg/kg) in combination with pertuzumab (40 mg/kg) in the NCI-N87 model showed significantly potent anti-tumor activity with a TGI of 113%. Both T-DM1 and pertuzumab as monotherapies showed significant anti-tumor activity, their TGI were 79% and 47%, respectively. To examine the mechanism of action of the combination of T-DM1 and pertuzumab, we analyzed apoptosis of NCI-N87 cells after treatment with T-DM1, pertuzumab or both under stimulation of EGF or heregulin. 24 h after treatment, the combination group had significantly enhanced caspase 3/7 activity compared with the monotherapy groups. We also examined ADCC activity of these agents. Both T-DM1 and pertuzumab showed ADCC activity and that activity was enhanced by the combination. Conclusion: In this study, we showed that T-DM1 as monotherapy has significant anti-tumor activity in a mice model of HER2-positive gastric cancer. Furthermore, combining T-DM1 with pertuzumab significantly enhanced the anti-tumor activity compared with the efficacy of each agent used in monotherapy. The mechanism of this combination might be the potentiation of apoptosis induction and ADCC activity. The present study suggests that T-DM1 as monotherapy and in combination with pertuzumab would provide benefit for patients with HER2-positive gastric cancers. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1761. doi:10.1158/1538-7445.AM2011-1761

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature

CD44 Variant Regulates Redox Status in Cancer Cells by Stabilizing the xCT Subunit of System xc − and Thereby Promotes Tumor Growth

CD44 is an adhesion molecule expressed in cancer stem-like cells. Here, we show that a CD44 variant (CD44v) interacts with xCT, a glutamate-cystine transporter, and controls the intracellular level of reduced glutathione (GSH). Human gastrointestinal cancer cells with a high level of CD44 expression showed an enhanced capacity for GSH synthesis and defense against reactive oxygen species (ROS). Ablation of CD44 induced loss of xCT from the cell surface and suppressed tumor growth in a transgenic mouse model of gastric cancer. It also induced activation of p38(MAPK), a downstream target of ROS, and expression of the gene for the cell cycle inhibitor p21(CIP1/WAF1). These findings establish a function for CD44v in regulation of ROS defense and tumor growth.

Bone Marrow-Derived Myofibroblasts Contribute to the Mesenchymal Stem Cell Niche and Promote Tumor Growth

Carcinoma-associated fibroblasts (CAFs) that express α-smooth muscle actin (αSMA) contribute to cancer progression, but their precise origin and role are unclear. Using mouse models of inflammation-induced gastric cancer, we show that at least 20% of CAFs originate from bone marrow (BM) and derive from mesenchymal stem cells (MSCs). αSMA+ myofibroblasts (MFs) are niche cells normally present in BM and increase markedly during cancer progression. MSC-derived CAFs that are recruited to the dysplastic stomach express IL-6, Wnt5α and BMP4, show DNA hypomethylation, and promote tumor growth. Moreover, CAFs are generated from MSCs and are recruited to the tumor in a TGF-β- and SDF-1α-dependent manner. Therefore, carcinogenesis involves expansion and relocation of BM-niche cells to the tumor to create a niche to sustain cancer progression.

HelicoVax: Epitope-based therapeutic Helicobacter pylori vaccination in a mouse model

Helicobacter pylori is the leading cause of gastritis, peptic ulcer disease and gastric adenocarcinoma and lymphoma in humans. Due to the decreasing efficacy of anti-H. pylori antibiotic therapy in clinical practice, there is renewed interest in the development of anti-H. pylori vaccines. In this study an in silico-based approach was utilized to develop a multi-epitope DNA-prime/peptide-boost immunization strategy using informatics tools. The efficacy of this construct was then assessed as a therapeutic vaccine in a mouse model of gastric cancer induced by chronic H. pylori infection. The multi-epitope vaccine administered intranasally induced a broad immune response as determined by interferon-gamma production in ELISpot assays. This was associated with a significant reduction in H. pylori colonization compared with mice immunized with the same vaccine intramuscularly, given an empty plasmid, or given a whole H. pylori lysate intranasally as the immunogen. Total scores of gastric histological changes were not significantly different among the 4 experimental groups. These results suggest that further development of an epitope-based mucosal vaccine may be beneficial in eradicating H. pylori and reducing the burden of the associated gastric diseases in humans.

Omental milky spots in screening gastric cancer stem cells

The existence of cancer stem and progenitor cells in solid tumors has been widely postulated. However, neither the cancer stem cells nor the cancer progenitor cells have been definitively identified and functionally characterized. Here we propose a new strategy to identify and isolate gastric cancer stem cells -using omental milky spots to screen gastric cancer stem cells in peritoneal metastasis mouse models of gastric cancer. In this study, we used the property that the macrophages in omental milky spots are cytotoxic against tumor cells and so able to screen and collect cancer stem cells. Our findings suggest that macrophages in omental milky spots have not only cytotoxic properties against tumor cells but also provide a microenvironment within milky spots in which cancer stem cells are capable to survive and grow into micrometastasis. Omental milky spot become a cancer stem cell niche in this situation. Further we studied the omental milky spots for screening gastric cancer cells (OMSS-GCCs) and found that omental milky spot enriched the volume of gastric cancer stem cells. Tumors were consistently generated after an injection of 1×103 OMSS-GCCs. OMSS-GCCs high express CD133 and low express CD324. Omental milky spots are a highly efficient "natural filter" for screening gastric cancer stem cells.

Lentivirus-mediated RNA interference targeting E2F-1 inhibits human gastric cancer MGC-803 cell growthin vivo

The E2F-1 transcription factor is post-translationally modified and stabilized in response to various forms of DNA damage to regulate the expression of cell-cycle and pro-apoptotic genes. The sustained overexpression of E2F-1 is a characteristic feature of gastric cancer. In this study, we investigated the role of short hairpin RNA (shRNA) targeting E2F-1 gene on human gastric cancer MGC-803 cell growth in vivo, and preliminarily revealed the mechanism. Thus, we constructed recombinant pGCSIL-GFP-shRNA-E2F-1 lentiviral vector to knock down E2F-1 expression in human gastric cancer MGC-803 cells in vivo, and studied the effect of E2F-1 shRNA on growth of MGC-803 tumor and evaluated its treatment efficacy. Our data demonstrated that in a mouse model of established gastric cancer, intratumor injection of lentiviral shRNA targeting E2F-1 definitely decreased the endogenous E2F-1 mRNA and protein expression in MGC-803 tumor, and inhibited tumor growth and promoted tumor cells apoptosis. Moreover, we found that E2F-1 shRNA increased the expression of phosphatase and tensin homolog (PTEN), activated caspase-3 and caspase-9, and suppressed nuclear factor (NF)-κB expression in tumor tissue as determined by reverse transcription (RT)-PCR and western blotting. In summary, shRNA targeting of E2F-1 can effectively inhibits human gastric cancer MGC-803 cell growth in vivo and may be a potential therapeutic strategy for gastric cancer.

Altered Expression and Localization of Connexin32 in Human and Murine Gastric Carcinogenesis

Intercellular communication via gap junctions, composed of protein subunits called connexins (Cxs), plays a key role in controlling cell growth, differentiation and carcinogenesis. Impaired gap junctional intercellular communication has been reported in various cancers and diseases. We investigated Cx32 expression patterns and semiquantitatively assessed Cx32 expression in cancers and preneoplastic lesions. To determine if cell proliferation is correlated with Cx32 expression, we evaluated Ki67 expression in a gastric cancer mouse model. In human and mouse, normal stomach and gastric adenocarcinoma tissues were used for immunohistochemical analyses. Cx32 was detected at cell-cell (intercellular) contact points in normal cells and exhibited punctate intercellular and intracytoplasmic staining in cancer cells. The frequency of Cx32 loss of expression was significantly higher in human adenocarcinomas than in normal stomach. As tumor cells were less differentiated, Cx32 expression levels and intercellular and intracytoplasmic staining were also significantly lower. The Cx32 expression pattern in the mouse gastric cancer model was similar in several important respects to that of human. In mucous metaplasia of the mouse stomach, Cx32 was mainly expressed in the cytoplasm of epithelial cells. There was also an inverse correlation between Cx32 expression and cell proliferation in mouse tumors. However, there was no difference in the levels of Cx32 mRNA between normal and cancerous tissues. These findings suggest that altered Cx32 expression, a loss of intercellular Cx32 and a gain of intracytoplasmic Cx32 in the form of punctate "dot", plays an important role in the formation of gastric adenocarcinomas.

Abstract 1420: Inhibition of platelet-derived growth factor receptor tyrosine kinase reduces tumor growth and metastasis in an orthotopic nude mice model of human gastric cancer

Abstract Background: Recent studies have revealed that platelet-derived growth factor (PDGF) plays a role in promoting progressive tumor growth in several organs; however, whether PDGF plays a role in gastric cancer is undetermined. We examined whether inhibition of PDGF receptor (PDGF-R) tyrosine kinase signaling by imatinib affects tumor growth and metastasis of human gastric cancer cells in an orthotopic nude mouse model. Methods: TMK1 human gastric cancer cells were injected into the gastric wall of nude mice. Groups of mice (n=10 each) received saline (control), low-dose imatinib (50 mg/kg/day), high-dose imatinib (200 mg/kg/day), cancer chemotherapeutic agent irinotecan (5 mg/kg/week), or imatinib (50 mg/kg/day or 200 mg/kg/day) and irinotecan in combination for 28 days. Tumor growth and metastasis were assessed. Resected tumors were analyzed immunohistochemically. Results: Carcinoma-associated fibroblasts (CAFs) within gastric tumors expressed high levels of PDGF-R; cancer cells did not. Treatment with imatinib alone did not inhibit tumor growth and metastasis; however, treatment with irinotecan alone or combined with imatinib (single-agent therapy or relative control) significantly inhibited tumor growth. Only treatment with high-dose imatinib and irinotecan in combination inhibited lymph node and peritoneal metastasis. Immunohistochemically, only imatinib alone or in combination with irinotecan significantly decreased the stromal reaction, pericyte coverage of tumor microvessels, and microvessel area. These effects were marked in the high-dose imatinib group. Conclusion: Administration of PDGF-R tyrosine kinase inhibitor in combination with irinotecan appears to impair the progressive growth of gastric cancer cells by blockade of PDGF-R signaling pathways in CAFs and pericytes on the tumor vasculature. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1420.

Mouse model of gastric cancer and changes of T cell immune function

Objective To induce gastric cancer mouse model using 3-methylcholanthrene and N-methyl-N-nitrosourea and to assess the characteristics of the immune system during model construction. Methods BALB/c mice were used to establish gastric cancer model. Glandular stomach of the mice was sutured with 3-MCA-containing silk suture, and subsequent intragastric administration of MNU was done to induce gastric cancer. On the week 12 and 20, peripheral blood and splenocytes were tested for WBC count, and CD3~+ , CD4~+ , and CD8~+ T-cell subsets and CD4~+ CD25~+ Treg cells were assessed by flow eytometry with concurrent collection of mice stomachs for pathologic study. Data collected were then ana-lyzed statistically. Results Cancer inducing rate with multiple factors attack method was 37. 5% (6/16), and weight changes in test group [(4. 52±1.97) g] significantly decreased (P <0. 05), and the levels of Treg cells in peripheral blood [(8. 07±6. 62) %] and [spleen (6. 45±4. 13) %] both showed signifi-cant increase (P < 0. 05 ), while CD4~+/CD8~+ ratio only showed significant increase in the spleen (1.95 ±0. 62,P < 0. 05 ). Conclusion Multiple factors attack method successfully induced gastric cancer in mice. Treg cells showed high expression during progress of gastric cancer, and after tumor formation there was suppression of the immune functions. Key words: Gastric carcinoma; Flow cytometry; T-Cell