Peritoneal Dissemination Of Gastric Cancer Research Articles

Janus kinase inhibitor overcomes resistance to immune checkpoint inhibitor treatment in peritoneal dissemination of gastric cancer in C57BL/6J mice.

Cancer immunotherapy aims to unleash the immune system's potential against cancer cells, providing sustained relief for tumors responsive to immune checkpoint inhibitors (ICIs). While promising in gastric cancer (GC) trials, the efficacy of ICIs diminishes in the context of peritoneal dissemination. Our objective is to identify strategies to enhance the impact of ICI treatment specifically for cases involving peritoneal dissemination in GC. The therapeutic efficacy of anti-PD1, CTLA4 treatment alone, or in combination was assessed using the YTN16 peritoneal dissemination tumor model. Peritoneum and peritoneal exudate cells were collected for subsequent analysis. Immunohistochemical staining, flow cytometry, and bulk RNA-sequence analyses were conducted to evaluate the tumor microenvironment (TME). A Janus kinase inhibitor (JAKi) was introduced based on the pathway analysis results. Anti-PD1 and anti-CTLA4 combination treatment (dual ICI treatment) demonstrated therapeutic efficacy in certain mice, primarily mediated by CD8 + T cells. However, in mice resistant to dual ICI treatment, even with CD8 + T cell infiltration, most of the T cells exhibited anexhaustion phenotype. Notably, resistant tumors displayed abnormal activation of the Janus Kinase-Signal Transducer and Activator of Transcription (JAK-STAT) pathway compared to the untreated group, with observed infiltration of macrophages, neutrophils, and Tregs in the TME. The concurrent administration of JAKi rescued CD8 + T cells function and reshaped the immunosuppressive TME, resulting in enhanced efficacy of the dual ICI treatment. Dual ICI treatment exerts its anti-tumor effects by increasing tumor-specific CD8 + T cell infiltration, and the addition of JAKi further improves ICI resistance by reshaping the immunosuppressive TME.

Abstract 1561: Identification of tumor-associated macrophages promoting early-stage metastasis during peritoneal dissemination in gastric cancer

Abstract Peritoneal dissemination (PD) is the most frequent metastasis pattern in gastric cancer (GC) and is a significant predictor of poor prognosis. However, the mechanisms underlying PD formation remain unclear, and a PD-specific signature has not been identified. PD development is thought to be influenced by various changes occurring in GC tumor cells and immune cells present in the peritoneal cavity. We previously performed RNA-seq analysis of cell components in cytology-positive (CY1) gastric cancer ascites. The results suggested that tumor-associated macrophages (TAMs) infiltrate the abdominal cavity together with GC tumor cells and might be involved in PD development. In this study, we aimed to identify the specific TAMs responsible for promoting metastasis during the early stage of PD in GC. We demonstrated the abundant presence of TAMs in CY1 GC ascites using flow cytometry analysis and identified several genes that were significantly expressed in CY1 TAMs using single-cell RNA-seq analysis. Furthermore, we evaluated the potential of these genes to serve as therapeutic targets for the treatment of PD in GC. Citation Format: Wataru Kawase, Yukihiko Hiroshima, Itaru Hashimoto, Shizune Onuma, Mitsuhiro Furuta, Nozomu Machida, Takashi Oshima, Yohei Miyagi. Identification of tumor-associated macrophages promoting early-stage metastasis during peritoneal dissemination in gastric cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1561.

Helicobacter pylori-induced fibroblast-derived Serpin E1 promotes gastric cancer growth and peritoneal dissemination through p38 MAPK/VEGFA-mediated angiogenesis

BackgroundFibroblasts, especially cancer-associated fibroblasts (CAFs), represent the predominant stromal cell population in the tumor microenvironment and have an important function in tumorigenesis by interacting with tumor cells. However, their interaction remains elusive in an inflammatory tumor microenvironment induced by Helicobacter pylori (H. pylori).MethodsThe expression of Serpin family E member 1 (Serpin E1) was measured in fibroblasts with or without H. pylori infection, and primary gastric cancer (GC) cells. Serpin E1 knockdown and overexpression fibroblasts were generated using Serpin E1 siRNA or lentivirus carrying Serpin E1. Co-culture models of fibroblasts and GC cells or human umbilical vein endothelial cells (HUVECs) were established with direct contact or the Transwell system. In vitro functional experiments and in vivo tumorigenesis assay were employed to study the malignant behaviors of GC cells interacting with fibroblasts. ELISA was used for quantifying the levels of Serpin E1 and VEGFA in the culture supernatant. The tube formation capacity of HUVECs was assessed using a tube formation assay. Recombinant human Serpin E1 (recSerpin E1), anti-Serpin E1 antibody, and a MAPK pathway inhibitor were utilized to treat HUVECs for elucidating the underlying molecular mechanisms.ResultsSerpin E1 was predominantly expressed in gastric CAFs. H. pylori infection significantly enhanced the expression and secretion of Serpin E1 by CAFs. Both fibroblast-derived Serpin E1 and recSerpin E1 enhanced the growth, invasion, and migration of GC cells, along with increased VEGFA expression and tube formation in HUVECs. Furthermore, the co-inoculation of GC cells and fibroblasts overexpressing Serpin E1 triggered the expression of Serpin E1 in cancer cells, which facilitated together xenograft tumor growth and peritoneal dissemination of GC cells in nude mice, with an increased expression of Ki67, Serpin E1, CD31 and/or VEGFA. These processes may be mediated by Serpin E1-induced migration and p38 MAPK/VEGFA-mediated angiogenesis of HUVECs.ConclusionH. pylori infection induces Serpin E1 expression in fibroblasts, subsequently triggering its expression in GC cells through their interaction. Serpin E1 derived from these cells promotes the migration and p38 MAPK/VEGFA-mediated angiogenesis of HUVECs, thereby facilitating GC growth and peritoneal metastasis. Targeting Serpin E1 signaling is a potential therapy strategy for H. pylori-induced GC.

Inhibition of cancer cell‑platelet adhesion as a promising therapeutic target for preventing peritoneal dissemination of gastric cancer.

Platelets form complexes with gastric cancer (GC) cells via direct contact, enhancing their malignant behavior. In the present study, the molecules responsible for GC cell-platelet interactions were examined and their therapeutic application in inhibiting the peritoneal dissemination of GC was investigated. First, the inhibitory effects of various candidate surface molecules were investigated on platelets and GC cells, such as C-type lectin-like receptor 2 (CLEC-2), glycoprotein VI (GPVI) and integrin αIIbβ3, in the platelet-induced enhancement of GC cell malignant potential. Second, the therapeutic effects of molecules responsible for the development and progression of GC were investigated in a mouse model of peritoneal dissemination. Platelet-induced enhancement of the migratory ability of GC cells was markedly inhibited by an anti-GPVI antibody and inhibitor of galectin-3, a GPVI ligand. However, neither the CLEC-2 inhibitor nor the integrin-blocking peptide significantly suppressed this enhanced migratory ability. In experiments using mouse GC cells and platelets, the migratory and invasive abilities enhanced by platelets were significantly suppressed by the anti-GPVI antibody and galectin-3 inhibitor. Furthermore, in vivo analyses demonstrated that the platelet-induced enhancement of peritoneal dissemination was significantly suppressed by the coadministration of anti-GPVI antibody and galectin-3 inhibitor, and was nearly eliminated by the combined treatment. The inhibition of adhesion resulting from GPVI-galectin-3 interaction may be a promising therapeutic strategy for preventing peritoneal dissemination in patients with GC.

Abstract 635: Effects of vagus nerve signals on the development of peritoneal metastasis of murine gastric cancer

Abstract Background: Vagus nerve is the only pathway that transmits parasympathetic signals between the brain and intraabdominal organs. Vagus nerve is often resected in the process of lymph node dissection in surgery for upper gastrointestinal tract malignancies. Several studies have shown that vagus nerve controls immune cell functions in abdominal organs through so called “cholinergic anti-inflammatory pathway’, which may seriously affect the tumor progression. However, the effect of vagotomy on postoperative outcome of the patients with gastric cancer is unclear. Methods: We investigated the effects of vagotomy on the intraperitoneal immunity and the development of peritoneal dissemination of gastric cancer in murine model. C57BL/6 mice were divided into two groups and subjected to sub-diaphragmatic vagotomy with pyloroplasty (VxPP) or sham operation with vagus-nerve preservation (PP). Then, YTN16P, syngenic murine gastric cancer cell line, were injected in the peritoneal cavity of these mice and number of peritoneal metastases on the mesentery and omentum was counted after 2 weeks. Immune cells were collected from peritoneal lavage or spleen and the ratio or numbers of lymphocytes and granulocyte subsets, including large peritoneal macrophage (LPM), small peritoneal macrophage (SPM), neutrophil, eosinophil, were analyzed at several points using flow cytometry. Results: The ratios of LPM, SPM and neutrophil almost unchanged at the 2weeks after surgery. However, the expression level of F4/80 of LPM in VxPP group decreased as compared with that in PP group (Mean Fluorescent Intensity; PP: 4510 ± 3610 vs VxPP: 660 ± 300, n=3). The ratio of regulatory T cell (Treg) in the spleen decreased at 2 weeks, but recovered to the preoperative levels 3 weeks after operation. There was no significant difference in the number of metastatic nodules on the mesentery between two groups (PP: 52.9 ± 24.6 vs VxPP: 62.8 ± 18.1, p=0.25, n=13). However, the number of nodules on the omentum in VxPP group was significantly greater than that in PP group (PP:2.8 ± 1.8 vs VxPP:5.2 ± 2.4, p<0.01, n=13). The ratios of neutrophil in peritoneal lavage (PP: 17.6 ± 8.4% vs VxPP: 24.4 ± 7.6%, p=0.027, n=13) and G-MDSC/M-MDSC (PP: 1.8 ± 0.33% vs VxPP 2.77 ± 0.84%, p=0.0012, n=13) were greater, while that of LPM was less (PP:19.7 ± 17.4% vs VxPP 7.4 ± 4.1%, p<0.01, n=13) in VxPP group, those in PP group. Conclusion: Vagus nerve signal might control the frequency of macrophages and granulocytes in the peritoneal cavity, which may elicit inhibitory effect on the development of dissemination of gastric cancer to omentum. Citation Format: Yurie Futoh, Hideyo Miyato, Misaki Matsumiya, Rei Takahashi, Yuki Kaneko, Kazuya Takahashi, Yuki Kimura, Hiroko Hayakawa, Hideyuki Ohzawa, Naohiro Sata, Joji Kitayama. Effects of vagus nerve signals on the development of peritoneal metastasis of murine gastric 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 635.

FAK/IL-8 axis promotes the proliferation and migration of gastric cancer cells.

Gastric cancer (GC) is one of the most common malignancies in China and is associated with high mortality. The occurrence and development of gastric cancer are related to genetic and environmental factors. Focal adhesion kinase (FAK) is a cytoplasmic nonreceptor protein tyrosine kinase that is activated by the extracellular matrix and growth factors. FAK is highly expressed in cancer and promotes its development by regulating cancer cell proliferation, migration, and angiogenesis. The expression of IL-8 is increased in many types of malignant tumor cells and is linked to their proliferation, migration, invasion, angiogenesis, and EMT. In this study, we found FAK to be essential for the proliferation, migration, and peritoneal metastasis of gastric cancer cells. To examine the molecular regulatory mechanisms of FAK in the peritoneal dissemination of gastric cancer, we performed RNA-seq analysis of MKN-45-FAK-/- and MKN45 cells and demonstrated that IL-8 was downregulated in FAK-deficient cells. Conversely, we confirmed that IL-8 activates FAK activity. We established that IL-8 promotes the proliferation, colony formation, andmigration of gastric cancer cells that are partially mediated by FAK. Thus, we propose that an IL-8-FAK-IL-8 positive feedback loop effects the proliferation and migration of gastric cancer cells.

Nomogram to predict peritoneal dissemination in gastric cancer.

465 Background: Peritoneal metastasis is a poor prognostic factor in patients with gastric cancer and cannot be predicted, that can tailor oncological treatment. We retrospectively analyzed the clinicopathological data among 110 gastric cancer patients. Methods: Univariate logistic regression was performed with peritoneal disease as the outcome to assess potential risk factors associated with it. We found that age, gastric outlet obstruction, VEGF, Ki67, p53 and NLR ratio were significant factors associated with peritoneal disease. Using multiple logistic regression, the adjusted odds ratio and 95% confidence interval were estimated. The predictive accuracy of this regression model was estimated using ROC curve analysis and Area under the curve, sensitivity and specificity were estimated. A nomogram was plotted based on the regression coefficient for the prediction of peritoneal disease. Results: It was estimated that age less than 40 years had 0.19 (0.05, 0.79), GOO positive had 2.04 (0.69, 6.03), VEGF positive had 4.69 (1.79, 12.33), elevated Ki67 had 3.94 (1.55, 9.97), positive p53 had 0.43 (0.17, 1.11) and NLR ratio had 1.15 (0.88, 1.51) odds of having peritoneal disease from multiple logistic regression. The regression model had significant goodness of fit for the data (Hosmer-Lemeshow test P value < 0.001). On estimating the AUC from the ROC curve for the model, we found that it had a good discriminatory ability with AUC = 0.81. Sensitivity was estimated to be 71.15%, whereas specificity was estimated to be 72.41% for the model. Subsequently, a nomogram was plotted based on the regression estimated from this model. Conclusions: The nomogram could risk-stratify those at risk of developing peritoneal disease and modify therapeutic strategies like selecting patients for prophylactic intraperitoneal chemotherapy or other novel therapeutic strategies. However, the nomogram needs to be validated prospectively.

Suppression of galectin-4 attenuates peritoneal metastasis of poorly differentiated gastric cancer cells.

Peritoneal dissemination, most often seen in metastatic and/or recurrent gastric cancer, is an inoperable condition that lacks effective treatment. The use of molecular targeted drugs is also limited; therefore, identifying novel therapeutic targets and improving our understanding of this metastatic cancer are an urgent requirement. In this study, we focused on galectin-4, which is specifically expressed in poorly differentiated cells with high potential for peritoneal dissemination. We knocked out the galectin-4 gene in NUGC4 cells using CRISPR/Cas9-mediated genome editing. Proliferation and peritoneal cancer formation in knockout cells were compared with those in wild-type and galectin-4 re-expressing cells. Western blotting and proximity ligation assays were performed to identify associated molecules affected by the expression of galectin-4. The effect of galectin-4 knockdown on cell proliferation and peritoneal metastasis was studied using a specific siRNA. Expression of galectin-4 in peritoneal metastatic tumors from 10 patients with gastric cancer was examined by immunohistochemistry. Suppression of galectin-4 expression reduced proliferation and peritoneal metastasis of malignant gastric cancer cells. Galectin-4 knockout and knockdown reduced the expression of activated c-MET and CD44. Galectin-4 was found to interact with several proteins on the cell surface, including CD44 and c-MET, via its carbohydrate-binding ability. Immunohistochemistry showed galectin-4 expression in peritoneal metastatic tumor cells in all patients examined. We clarified the role of galectin-4 in the development of peritoneal dissemination of poorly differentiated gastric cancer cells. Our data highlight the diagnostic and therapeutic potential of galectin-4 in the peritoneal dissemination of gastric cancer.

Construction of a Nomogram Model for Predicting Peritoneal Dissemination in Gastric Cancer Based on Clinicopathologic Features and Preoperative Serum Tumor Markers.

BackgroundPeritoneal dissemination (PD) is the most common mode of metastasis for advanced gastric cancer (GC) with poor prognosis. It is of great significance to accurately predict preoperative PD and develop optimal treatment strategies for GC patients. Our study assessed the diagnostic potential of serum tumor markers and clinicopathologic features, to improve the accuracy of predicting the presence of PD in GC patients.MethodsIn our study, 1264 patients with GC at Fudan University Shanghai Cancer Center and Wenzhou people’s hospital from 2018 to 2020 were retrospectively analyzed, including 316 cases of PD and 948 cases without PD. All patients underwent enhanced CT scan or magnetic resonance imaging (MRI) before surgery and treatment. Clinicopathological features, including tumor diameter and tumor stage (depth of tumor invasion, nearby lymph node metastasis and distant metastasis), were obtained by imaging examination. The independent risk factors for PD were screened through univariate and multivariate logistic regression analyses, and the results were expressed with 95% confidence intervals (CIs). A model of PD diagnosis and prediction was established by using Cox proportional hazards regression model of training set. Furthermore, the accuracy of the prediction model was verified by ROC curve and calibration plots.ResultsUnivariate analysis showed that PD in GC was significantly related to tumor diameter (odds ratio (OR)=12.06, p<0.0006), depth of invasion (OR=14.55, p<0.0001), lymph node metastases (OR=5.89, p<0.0001), carcinoembryonic antigen (CEA) (OR=2.50, p<0.0001), CA125 (OR=11.46, p<0.0001), CA72-4 (OR=4.09, p<0.0001), CA19-9 (OR=2.74, p<0.0001), CA50 (OR=5.20, p<0.0001) and CA242 (OR=3.83, p<0.0001). Multivariate analysis revealed that clinical invasion depth and serum marker of CA125 and CA72-4 were independent risk factors for PD. The prediction model was established based on the risk factors using the R program. The area under the curve (AUC) of the receiver operating characteristics (ROC) was 0.931 (95% CI: 0.900–0.960), with the accuracy, sensitivity and specificity values of 90.5%, 86.2% and 82.2%, respectively.ConclusionThe nomogram model constructed using CA125, CA72-4 and depth of invasion increases the accuracy and sensitivity in predicting the incidence of PD in GC patients and can be used as an important tool for preoperative diagnosis.

Removal of small extracellular vesicles inhibits the progression of peritoneal dissemination in gastric cancer.

The prognosis of gastric cancer patients with peritoneal dissemination is extremely poor and effective treatment for peritoneal dissemination has not been established. Gastric cancer-derived small extracellular vesicles play an important role in the development of a favorable microenvironment for peritoneal metastasis and progression of peritoneal dissemination. Here, we aimed to investigate the transformation of gastric cancer cells by removing gastric cancer-derived small extracellular vesicles and to develop a novel therapy for inhibiting peritoneal dissemination. Gastric cancer cells were cultured in medium containing gastric cancer- and peritoneal mesothelium-derived small extracellular vesicles and in medium from which small extracellular vesicles were removed by ultracentrifugation. Cell function assays were performed in vitro, and the alternations in gene expression in gastric cancer cells were analyzed. The inhibitory effect of intraperitoneal lavage on peritoneal dissemination was investigated in vivo as a method to remove gastric cancer-derived small extracellular vesicles. Removal of gastric cancer-derived small extracellular vesicles suppressed the proliferative and migrative abilities of gastric cancer cells and the adhesion of gastric cancer cells to peritoneal mesothelial cells. It altered the expression of several genes related to the cell cycle and epithelial-mesenchymal transition pathways of gastric cancer cells, leading to the inhibition of gastric cancer cell growth and peritoneal dissemination in vivo. Our study provides novel insights into a novel therapy for inhibiting the peritoneal dissemination of gastric cancer by targeting gastric cancer-derived small extracellular vesicles to improve the prognosis of gastric cancer patients with peritoneal metastasis.

Targeting histone deacetylase-3 blocked epithelial-mesenchymal plasticity and metastatic dissemination in gastric cancer

Background and purposeHistone deacetylase (HDAC) inhibitors (HDIs) can modulate the epithelial-mesenchymal transition (EMT) progression and inhibit the migration and invasion of cancer cells. Emerging as a novel class of anti-cancer drugs, HDIs are attracted much attention in the field of drug discovery. This study aimed to discern the underlying mechanisms of Honokiol in preventing the metastatic dissemination of gastric cancer cells by inhibiting HDAC3 activity/expression.Experimental approachClinical pathological analysis was performed to determine the relationship between HDAC3 and tumor progression. The effects of Honokiol on pharmacological characterization, functional, transcriptional activities, organelle structure changes, and molecular signaling were analyzed using binding assays, differential scanning calorimetry, luciferase reporter assay, HDAC3 activity, ER stress response element activity, transmission electron microscopy, immune-blotting, and Wnt/β-catenin activity assays. The in vivo effects of Honokiol on peritoneal dissemination were determined by a mouse model and detected by PET/CT tomography.Key resultsHDAC3 over-expression was correlated with poor prognosis. Honokiol significantly abolished HDAC3 activity (Y298) via inhibition of NFκBp65/CEBPβ signaling, which could be reversed by the over-expression of plasmids of NFκBp65/CEBPβ. Treatments with 4-phenylbutyric acid (a chemical chaperone) and calpain-2 gene silencing inhibited Honokiol-inhibited NFκBp65/CEBPβ activation. Honokiol increased ER stress markers and inhibited EMT-associated epithelial markers, but decreased Wnt/β-catenin activity. Suppression of HDAC3 by both Honokiol and HDAC3 gene silencing decreased cell migration and invasion in vitro and metastasis in vivo.Conclusions and implicationsHonokiol acts by suppressing HDAC3-mediated EMT and metastatic signaling. By prohibiting HDAC3, metastatic dissemination of gastric cancer may be blocked.Graphical abstractConceptual model showing the working hypothesis on the interaction among Honokiol, HDAC3, and ER stress in the peritoneal dissemination of gastric cancer. Honokiol targeting HDAC3 by ER stress cascade and mitigating the peritoneal spread of gastric cancer. Honokiol-induced ER stress–activated calpain activity targeted HDAC3 and blocked Tyr298 phosphorylation, subsequently blocked cooperating with EMT transcription factors and cancer progression. The present study provides evidence to demonstrate that HDAC3 is a positive regulator of EMT and metastatic growth of gastric cancer cells. The findings here imply that overexpressed HDAC3 is a potential therapeutic target for honokiol to reverse EMT and prevent gastric cancer migration, invasion, and metastatic dissemination.• Honokiol significantly abolished HDAC3 activity on catalytic tyrosine 298 residue site. In addition, Honokiol-induced ER stress markedly inhibited HDAC3 expression via inhibition of NFκBp65/CEBPβ signaling.• HDAC3, which is a positive regulator of metastatic gastric cancer cell growth, can be significantly inhibited by Honokiol.• Opportunities for HDAC3 inhibition may be a potential therapeutic target for preventing gastric cancer metastatic dissemination.

Integration of exosomal miR-106a and mesothelial cells facilitates gastric cancer peritoneal dissemination

BackgroundMetastatic organotropism is considered the end stage of malignancy with the mechanism still have some mysteries that have not been disclosed. Although the role of miR-106a is well studied, its involvement in the formation of peritoneal metastasis transported by exosomes is less discussed. MethodsGastric cancer (GC)-derived exosomes were identified by transmission electron microscopy and the integration with peritoneal mesothelial cells (PMC) was confirmed by PKH-26 staining. Cell phenotype was assessed by EdU and flow cytometry. MiR-106a and its targets were authenticated by luciferase reporter assay. The mesothelial-to-mesenchymal transition (MMT) and extracellular matrix (ECM) degeneration were determined and morphological transformation was measured by transwell assay. Immunodeficient mouse was conducted to investigate the tumorigenesis and tumor growth. The final was tissue analysis. ResultsMiR-106a enrichment in GC-exosomes can be delivered and integrated into PMC to establish a proper pre-metastatic niche (PMN). We found that PMC could internalize exosomal-miR-106a and lead to increased apoptotic rate and decreased proliferative vitality. The direct target Smad7 and TIMP2 were proved to be effectively reduced by translocation of exosomal miR-106a. We confirmed that exosomal miR-106a was able to induce MMT and accelerate ECM through targeting Smad7 and TIMP2 to activate TGF-β pathway. Animal model investigated that the tumorigenesis and tumor growth could be influenced by exosomes, and exosomal-miR-106a could promote the formation of xenograft tumor. Tissue analysis verified the ectopic miR-106a expression in gastric cancer metastasis. ConclusionOur data suggest that exosomal miR-106a facilitates gastric cancer peritoneal dissemination by integrating PMC to destroy mesothelial barrier.

Survival analysis of a prospective multicenter observational study on surgical palliation among patients with malignant bowel obstruction caused by peritoneal dissemination of gastric cancer.

Our previous report showed that surgical palliation maintained quality of life (QOL), improved solid food intake, and had an acceptable surgical safety among patients with malignant bowel obstruction (MBO) caused by advanced gastric cancer. This study performed a survival analysis stratified by the patients' QOL to elucidate its impact on survival. Patients who underwent resection or bypass of the small intestine/colon or ileostomy/colostomy for bowel obstruction caused by peritoneal dissemination of gastric cancer were included. Validated instruments (EuroQoL-5 Dimensions) were used to assess QOL at baseline and 2weeks, 1month, and 3months following surgical palliation. Postoperative improvement in oral intake was also evaluated using the Gastric Outlet Obstruction Scoring System (GOOSS). Univariate and multivariate survival analyses were performed using baseline characteristics and changes in QOL and GOOSS scores 2weeks after surgery to determine prognostic factors. We enrolled 60 patients with a median survival time of 6.64 (95% CI 4.76-10.28) months. Patients who received postoperative chemotherapy and had lower baseline C-reactive protein (CRP) levels, higher baseline albumin levels, better baseline EuroQoL-5 Dimensions (EQ-5D) scores, and improved oral intake after palliative surgery exhibited significantly better survival. Multivariate analysis identified postoperative chemotherapy, lower baseline CRP levels, and improved oral intake as independent prognostic factors. The current study revealed that baseline QOL and postoperative QOL changes did not affect survival. Moreover, improved oral intake, lower baseline CRP levels, and postoperative chemotherapy were significant prognostic factors in patients who underwent palliative surgery for advanced gastric cancer with MBO.

Circulating methylated THBS1 DNAs as a novel marker for predicting peritoneal dissemination in gastric cancer.

ObjectivesThrombospondin 1 (THBS1) is known to play a key role in tumor metastasis, and aberrant DNA methylation is one of the mechanisms regulating THBS1. The present study investigated whether methylated THBS1 in circulating cell‐free DNA from preoperative peritoneal lavage fluid (PPLF) and peripheral blood could be used as a potential biomarker for predicting peritoneal dissemination in gastric cancer (GC) patients.MethodsThe status of THBS1 methylation was detected by quantitative methylation‐specific PCR (MSP) in tumor tissues, paired PPLF, and serum from 92 GC patients. The correlation between methylated THBS1 levels and peritoneal dissemination of GC was studied, and its diagnostic value for predicting peritoneal dissemination was clarified by the receiver operating characteristic (ROC) curve.ResultsAberrant THBS1 methylation in tumor tissues was significantly higher than that in paracancerous normal tissues (p < 0.0001). No THBS1 methylation was found in 40 healthy controls, and partial methylation was detected in 3 of 48 patients with chronic non‐atrophic gastritis. The frequency of THBS1 methylation in pairing PPLF and serum from 92 GC patients was 52.2% (48/92) and 58.7% (54/92), respectively. The results of methylated THBS1 in pairing PPLF and serum were similar to those of tumor tissues. Aberrant THBS1 methylation in tumor tissues and pairing PPLF or serum was closely related to peritoneal dissemination, tumor progression, and poor prognosis (all p < 0.0001).ConclusionCirculating methylated THBS1 DNAs in PPLF/serum may predict peritoneal dissemination, a potential poor prognostic factor for GC patients.

Abstract 2905: The relationship between peritoneal dissemination of gastric cancer &amp; exosome

Abstract Introduction: In recent years, studies of exosomes have progressed rapidly, suggesting a connection to various physiological functions and pathogenesis. The exosomes are small extracellular vesicles with a diameter of 30 to 100 nm surrounded by lipid bilayers, and are attracting attention as a new cell-cell communication medium for carrying lipids, proteins, RNA, etc. in vivo. In addition, in exosomes released by cancer cells, it contains a large number of molecules related to angiogenesis and immunosuppression. It is believed to contribute to building microenvironments suitable for cancer cell growth and promoting cancer progression. Therefore, we focused on exosomes derived from gastric cancer cells, the action of gastric cancer cells, further constructed a gastric cancer peritoneal dissemination mouse model, the action of exosomes in gastric cancer peritoneal metastasis mechanism was examined in vitro, in vivo, respectively. Method: Exosome Isolation Kit was used to extract exosomes from each gastric cancer cell line MKN-1, MKN-45, MKN-45 GFP. The determination of exosomes was carried out using Nano-drop®. Cell adhesion in exosome-treated and non-administered groups were analyzed by colony forming assay. Cell proliferation ability was analyzed using cell proliferation assay method. In vivo, we examined whether the degree of peritoneal dissemination was made using live cell imaging using fluorescent cells. MKN-45 GFP was intraperitoneally administered to BALB/c-nu mice. On days 0, 5, 10, 15, 20, groups were prepared in which 25 µg/500 µl of exosome was administered intraperitoneally and groups in which the same amount of PBS was administered intraperitoneally. In addition, MKN-45 GFP was subcutaneously injected into BALB/c-nu mice to create MKN-45 GFP tumors, which were divided into small horns and transplanted to the stomach walls of BALB/c-nu mice. In the same way as the previous method, we prepared a group in which 25 µg/500 µl exosome was intraperitoneally administered on days 0, 5, 10, 15, 20 and a group in which the same amount of PBS was intraperitoneally administered. We performed a visual comparison of transplanted cancer cells and peritoneal dissemination of cancer. Results and Discussion: The results of the colony forming assay method suggest that exosomes derived from gastric cancer cells have the effect of promoting colony formation ability of cancer cells. We evaluate the results of peritoneal seeding model mice in this study, and further proceed with the analysis of miRNA extraction and analysis from exosomes derived from gastric cancer cells, and cell adhesion of cancer cells under high concentrations of exosomes. Citation Format: Kazuya Kinoshita. The relationship between peritoneal dissemination of gastric cancer &amp; exosome [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2905.

Bufalin inhibits peritoneal dissemination of gastric cancer through endothelial nitric oxide synthase-mitogen-activated protein kinases signaling pathway.

Peritoneal dissemination threatens the survival of patients with gastric cancer (GC). Bufalin is an extract of traditional Chinese medicine, which has been proved to have anticancer effect. The target of bufalin in suppressing gastric cancer peritoneal dissemination (GCPD) and the underlying mechanism are still unclear. In this research, GC cell line MGC-803 and high-potential peritoneal dissemination cell line MKN-45P were treated with bufalin or L-NAME. Malignant biological behavior and protein level of GC cell lines were detected with MTT, wound healing, transwell, adhesion, and western blotting. Bioinformatics analysis and patient tissues were used to verify the role of endothelial nitric oxide synthase (NOS3) in GC. Mice model was used to assess the effect of bufalin and role of NOS3 in vivo. We found that bufalin inhibited the proliferation, invasion, and migration in GC cell lines. NOS3, which was an independent prognostic factor of GC patients, was predicted to be a potential target of bufalin. Further experiments proved that bufalin reduced the phosphorylation of NOS3, thereby inhibiting the mitogen-activated protein kinase (MAPK) signaling pathway, and ultimately suppressed GCPD by inhibiting EMT process. In conclusion, NOS3 was a potential therapeutic target and prognostic biomarker of GC. Bufalin could suppress GCPD through NOS3-MAPK signaling pathway, which provided more evidence support for intraperitoneal perfusion of bufalin to treat GCPD.

The Role of Open Diagnostic Peritoneal Lavage in the Evaluation of Peritoneal Cytology for Advanced Gastric Cancer: An Old Diagnostic Modality with New Usage

Background: Positive peritoneal cytology is a critical factor in prognosis. Peritoneal lavage is associated with long-term survival in patients with gastric cancer. Diagnostic peritoneal lavage (DPL) is a method for diagnosing visceral injury in trauma patients. This study aimed to investigate the usage of DPL in staging the work-up of patients with gastric cancer. Method: In this prospective study, we enrolled gastric cancer patients referring to Cancer Institute; they underwent DPL and washing specimen was sent for cytology review. After DPL, all patients underwent staging laparoscopy (SL) via the same abdominal incision. Results: DPL and SL were successful in all patients. There were six (11%) cases of peritoneal seeding discovered in SL; all of these patients had positive peritoneal cytology on DPL. Also, four patients showed positive cytology in the absence of positive SL. Thus, sensitivity, specificity, negative predictive value, positive predictive value, and accuracy of DPL were 100 % (95% CI: 54.1-100), 91.6 % (95%: 79.2-97.5), 100 % (95%CI: 85.3-100), and 60 % (95%CI: 37-79.3). The accuracy of DPL in determining the peritoneal dissemination of gastric cancer was 92.31% (95% CI: 81.5-97.9). Conclusion: DPL had an excellent ability to find peritoneal dissemination in a gastric cancer patient, which is of great value in the setting of low-resource countries.

Hyperthermic intraperitoneal chemotherapy in prevention of gastric cancer metachronous peritoneal metastases: a systematic review.

Gastric cancer progression resulting in metachronous peritoneal metastasizing is almost always associated with an adverse prognosis. This review discusses various options of preventing metachronous peritoneal metastases in radically operated gastric cancer patients. Also examined are different hyperthermic intraperitoneal chemotherapy (HIPEC) regimens employed in gastric cancer treatment, postoperative morbidity and mortality rates and long-term treatment outcomes. The authors also review their own experience of using HIPEC based on the combination of cisplatin and doxorubicin in doses of 50 mg/m2 at 42 °C for 1 h to prevent gastric cancer peritoneal dissemination. As a result, progression-free survival rose from 19.6%±5.6% to 47.1%±6.3% (Plog-rank <0.001) and dissemination-free survival-from 22.7%±6.0% to 51.9%±6.3% (Plog-rank <0.001). It is noted that the combination of the described HIPEC regimen with systemic chemotherapy helped raise metastases-free 3-year survival rate to up to 91.0%±9.0% (Plog-rank =0.025) compared with 48.6%±6.4% for patients who underwent only a combined surgery/HIPEC treatment. HIPEC is a promising combined treatment strategy for radically operated gastric cancer patients that can improve patient survival and decrease peritoneal dissemination rate. However, the number of randomized studies on adjuvant HIPEC are still insufficient for a subgroup assessment of efficacy of the given chemotherapy regimens and generation of evidence-based recommendations on the individual use of chemotherapy agents and their combinations, and HIPEC procedural techniques. Further prospective randomized studies are needed to assess the practicability of complementing HIPEC with adjuvant systemic chemotherapies.

QOL assessment after palliative surgery for malignant bowel obstruction caused by peritoneal dissemination of gastric cancer: a prospective multicenter observational study.

Patients with peritoneal dissemination of gastric cancer have poor oral intake caused by malignant bowel obstruction (MBO). Palliative surgery has often been undertaken to improve quality of life (QOL), but few prospective studies on palliative surgery in this patient population have been published. We prospectively investigated the significance of palliative surgery using patient-reported QOL measures. Patients underwent palliative surgery by small intestine/colon resection or small intestine/colon bypass or ileostomy/colostomy for MBO. The primary endpoint was change in QOL assessed at baseline, 14days, 1month, and 3months following palliative surgery using the Euro QoL Five Dimensions (EQ-5D™) questionnaire and the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire gastric cancer module (QLQ-STO22). Secondary endpoints were postoperative improvement in oral intake and surgical complications. Between April 2013 and March 2018, 63 patients were enrolled from 14 institutions. The mean EQ-5D™ utility index baseline score of 0.6 remained consistent. Gastric-specific symptoms mostly showed statistically significant improvement from baseline. Forty-two patients (67%) were able to eat solid food 2weeks after palliative surgery and 36 patients (57%) tolerated it for 3months. The rate of overall morbidity of ≥ grade III according to the Clavien-Dindo classification was 16% (10 patients) and the 30-day postoperative mortality rate was 3.2% (2 patients). In patients with MBO caused by peritoneal dissemination of gastric cancer, palliative surgery did not improve QOL while improving solid food intake, with an acceptable postoperative morbidity and mortality rate.

Inflammation-driven senescence-associated secretory phenotype in cancer-associated fibroblasts enhances peritoneal dissemination

In the tumor microenvironment, senescent non-malignant cells, including cancer-associated fibroblasts (CAFs), exhibit a secretory profile under stress conditions; this senescence-associated secretory phenotype (SASP) leads to cancer progression and chemoresistance. However, the role of senescent CAFs in metastatic lesions and the molecular mechanism of inflammation-related SASP induction are not well understood. We show that pro-inflammatory cytokine-driven EZH2 downregulation maintains the SASP by demethylating H3K27me3 marks in CAFs and enhances peritoneal tumor formation of gastric cancer (GC) through JAK/STAT3 signaling in a mouse model. A JAK/STAT3 inhibitor blocks the increase in GC cell viability induced by senescent CAFs and peritoneal tumor formation. Single-cell mass cytometry revealed that fibroblasts exist in the ascites of GC patients with peritoneal dissemination, and the fibroblast population shows p16 expression and SASP factors at high levels. These findings provide insights into the inflammation-related SASP maintenance by histone modification and the role of senescent CAFs in GC peritoneal dissemination.