Gastric Tumor Tissues Research Articles

Long non-coding RNA TTTY15 silencing inhibits gastric cancer progression by sponging microRNA-98-5p to down-regulate cyclin D2 expression

ABSTRACT Gastric cancer is the most common malignant tumor in the digestive system. However, the detection rate of early gastric cancer is low, resulting in delayed prognosis and poor outcomes. The identification of effective therapeutic targets for gastric cancer is, therefore, of profound significance. Recently, various lncRNAs have been shown to be biomarkers for different cancers. This study investigated the role of long non-coding RNA (lncRNA) TTTY15 in gastric cancer. The expression level of TTTY15, miR-98-5p, and cyclin D2 (CCND2) were evaluated by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and Western blot assay using tumor and non-tumor tissues collected from 30 patients with gastric cancer, gastric cancer cell lines (AGS, SNU-5, and NCI-N87), and the normal gastric epithelial cell line GES-1. The interaction between TTTY15 and miR-98-5p and between miR-98-5p and CCND2 were predicted by bioinformatics and then further verified by dual-luciferase and RNA pull-down analyses. Cell proliferation was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2 H-tetrazolium bromide (MTT) assay, and apoptosis was measured using flow cytometry and caspase-3 assay. The results indicate that TTTY15 and CCND2 expression increased and miR-98-5p expression decreased in gastric cancer tumor tissues and cell lines. TTTY15 knockdown inhibited gastric cancer cell proliferation but promoted apoptosis by sponging miR-98-5p, which acted as a tumor suppressor gene by reducing the expression of its target gene CCND2 in gastric cancer. In conclusion, lncRNA TTTY15 is a potential oncogene involved in gastric cancer and may be a novel therapeutic target for gastric cancer treatment.

Block of Proliferation 1 Promotes Proliferation, Invasion and Epithelial Mesenchymal Transformation in Gastric Cancer.

Background Gastric cancer (GC) is one of the leading causes of cancer-related death worldwide nowadays. Block of proliferation 1 (BOP1), a nucleolar protein involved in rRNA processing and ribosome assembly, is associated with tumor development in certain cancers of digestive system. Therefore, we hypothesized that BOP1 might play an important role in gastric cancer development. Methods Gene Expression Omnibus (GEO) database and The Cancer Genome Atlas (TCGA) were used to identify the differentially expressed genes and their clinical relevance. qPCR and western blot were performed further to examine the levels of BOP1 mRNA and protein, respectively. Cell viability, apoptosis, migration and invasion were investigated in gastric cancer cell lines with BOP1 silencing or overexpression. The epithelial mesenchymal transition (EMT) associated proteins, including E-cadherin and N-cadherin, were measured using immunoblotting. Finally, the downstream pathway of BOP1 were explored using bioinformatic analysis and qPCR. Results BOP1 was found up-regulated in gastric tumor tissues compared with paired normal tissues (P < 0.0001). Its expression was associated with more advanced pathological grades (P = 0.0006) and tumor location (P = 0.002), as well as a poor survival (HR 1.27, P = 0.015). BOP1 expression was increased in 4 kind of tumor cell lines compared with the normal group. The overexpression of BOP1 promoted cell proliferation and inhibit cell apoptosis, while silencing BOP1 showed a reversed trend. Immunoblotting results suggested that BOP enhanced N-cadherin, a mesenchymal marker, while reduced E-cadherin, an epithelial marker. Finally, bioinformatic prediction showed that the cell cycle could be a downstream pathway of BOP1. Conclusions The present study demonstrated that BOP1 contributed to the development of gastric cancer by promoting proliferation, invasion and epithelial mesenchymal transformation, which could be a biomarker or therapeutic target in GC.

Relationship between Ki-67 and CD44 expression and microvascular formation in gastric stromal tumor tissues.

BACKGROUNDA gastric stromal tumor (GST) is a mesenchymal tumor that occurs in the gastrointestinal tract; its biological characteristics are highly complex. Clinically, the severity of a GST is often evaluated by factors such as risk classification, tumor size, and mitotic figures. However, these indicators are not very accurate. Even patients classified as low risk are also at risk of metastasis and recurrence. Therefore, more accurate and objective clinical biological behavior evaluations are urgently needed.AIMTo determine the relationship between Ki-67 and CD44 expression in GSTs and microvessel formation and prognosis.METHODSEighty-six GST tissue specimens from our hospital were selected for this study. The immunohistochemical staining technique was used to detect Ki-67, CD44, and microvessel density (MVD) in the collected samples to analyze the different risk grades and mitotic figures. In addition, this approach was used to determine the differences in the expression of Ki-67 and CD44 in GST tissues with varying lesion diameters.RESULTSIn GSTs with positive expression of the Ki-67 protein, the proportions of patients with medium-to-high risk and more than five mitotic counts were 24.07% and 38.89%, respectively. In GSTs with positive expression of the CD44 protein, the proportions of patients with medium-to-high risk and more than five mitotic counts were 23.73% and 38.98%, respectively. In GSTs with negative expression of the Ki-67 protein, these values were relatively high (3.70% and 11.11%, respectively). The MVD in GSTs with positive and negative expression of the CD44 protein was 15.92 ± 2.94 and 13.86 ± 2.98/Hp, respectively; the difference between the two groups was significant (P < 0.05).CONCLUSIONKi-67 and CD44 expression in GSTs is correlated with the grade of tumor risk and mitotic figures. CD44 expression is correlated with microvessel formation in tumor tissues.

Single-cell landscape reveals active cell subtypes and their interaction in the tumor microenvironment of gastric cancer

Background: Gastric cancer remains the third most common cause of cancer-related death worldwide. The development of novel therapeutic strategies for gastric cancer requires a deep understanding of the tumor cells and microenvironment of gastric cancer.Methods: We performed the single-cell RNA sequencing (scRNA-seq) on nine untreated non-metastatic gastric cancer patients. The transcriptomic atlas and ligand-receptor-based intercellular communication networks of the single cells were characterized.Results: Here, we profiled the transcriptomes of 47,304 cells from nine patients with gastric cancer. Tregs cells were significantly enriched in the gastric tumor tissues with increased expression of immune suppression related genes, which suggest a more immunosuppressive microenvironment. We also observed the absence of separate exhausted CD8+ T cell cluster, and the low expression level of exhaustion markers PDCD1, CTLA4, HAVCR2, LAG-3, and TIGIT in those specific cells. These may serve as molecular-level evidence for the limited benefit of immunotherapy among gastric cancer patients. In addition, we found ACKR1 specifically expressed in tumor endothelial cells, associated with poor prognosis in the cohort data and potentially provided a novel target of gastric cancer treatment. Furthermore, the tight interaction between endothelial cells and fibroblast implied the important roles of fibroblast in tumor angiogenesis and the maintenance of tumor vasculature.Conclusions: In conclusion, this single-cell atlas provide understanding the cellular heterogeneity from molecular level in gastric cancer and will serve as a valuable resource for developing innovative early and companion diagnostics, as well as discovering novel targeted therapies for gastric cancer.

Fast Screening Method of Biological Samples Based on Needle Stochastic Sensors for Early Detection of Gastric Cancer

Early detection of cancer is essential for saving the life of patients. Needle stochastic sensors were proposed as new tools for the fast screening of biological samples. The design of the stochastic sensors was based on the immobilization of protoporphyrin IX in pastes of S-doped graphene. The selected biomarkers were CEA and CA19-9. High sensitivities, and wide linear concentration ranges as well as low limits of quantification were achieved. The proposed sensors were validated; student t-test shown that the proposed needle stochastic sensors can be reliably used for the pattern recognition and quantification of CEA and CA19-9 in whole blood, gastric tumor tissue, saliva, and urine.

KLK6 mediates stemness and metabolism of gastric carcinoma cells via the PI3K/AKT/mTOR signaling pathway.

Gastric cancer is a common tumor of the digestive system, which can occur in any part of the stomach. Kallikrein 6 (KLK6) is a trypsin-like serine protease and has been found to be involved in extracellular matrix remodeling, tumor invasion and nervous system plasticity. Our previous study reported that KLK6 suppressed HGC-27 gastric cancer cell growth by inhibiting epithelial-mesenchymal transition; however, the mechanism of action underlying the effect of KLK6 still remains unclear. The aim of the present study was to investigate the effect and the underlying mechanism of KLK6 on stem cell-like properties and metabolism in gastric carcinoma cells. The HGC-27 cell line was transfected with KLK6 overexpression (OV-KLK6) and interference (short hairpin-KLK6) vectors, then the transfection efficiency was confirmed using western blot analysis and reverse transcription-quantitative PCR. The percentage of CD133+ and CD44+ cells was detected using flow cytometry, while the protein expression levels of the stem-associated genes, Nanog, Oct-4, SOX2 and Notch1, the metabolic markers, hexokinase (HK)1, HK2, GLUT1, and the proteins within the PI3K signaling pathway, phosphorylated (p)-PI3K, p-AKT and p-mTOR, were determined using western blot analysis. Biochemical kits were used to measure ATP production, lactic acid content and glucose uptake. A tumorigenicity assay was performed with nude mice to detect gastric tumor volume, and the protein expression level of Oct-4, Nanog, HK1, HK2 and GLUT1, and the mRNA expression level of KLK6 was also determined in gastric tumor tissues of mice. Compared with that in the control group, KLK6 protein and mRNA expression levels were significantly decreased in the four sh-RNA groups (P<0.05). Among them, sh-RNA-3 induced the lowest KLK6 expression and was used to silence KLK6 in subsequent experiments. Compared with that in the control and negative control groups, the percentage of CD133+ and CD44+ cells, the protein expression level of Oct-4, Nanog, HK1, HK2, GLUT1, p-PI3K, p-AKT and p-mTOR, and ATP content, lactic acid production, glucose uptake and gastric tumor volume were significantly decreased by sh-KLK6 (P<0.05), whereas KLK6 overexpression induced the opposite effect (P<0.05). In conclusion, KLK6 modulated stemness properties and cell metabolic profile in gastric carcinoma cells and the mechanism may be associated with the PI3K/AKT/mTOR signaling pathway.

Hedgehog transcriptional effector GLI mediates mTOR-Induced PD-L1 expression in gastric cancer organoids.

Tumors evade immune surveillance by expressing Programmed Death-Ligand 1 (PD-L1), subsequently inhibiting CD8+ cytotoxic T lymphocyte function. Response of gastric cancer to immunotherapy is relatively low. Our laboratory has reported that Helicobacter pylori-induced PD-L1 expression within the gastric epithelium is mediated by the Hedgehog (Hh) signaling pathway. The PI3K/AKT/mTOR pathway is activated in gastric cancer and may have immunomodulatory potential. We hypothesize that Hh signaling mediates mTOR-induced PD-L1 expression. Patient-derived organoids (PDOs) were generated from gastric biopsies and resected tumor tissues. Autologous organoid/immune cell co-cultures were used to study the immunosuppressive function of MDSCs. NanoString Digital Spatial Profiling (DSP) of immune-related protein markers using FFPE slide-mounted tissues from gastric cancer patients was performed. DSP analysis showed infiltration of immunosuppressive MDSCs expressing Arg1, CD66b, VISTA and IDO1 within cancer tissues. Orthotopic transplantation of patient derived organoids (PDOs) resulted in the engraftment of organoids and the development of histology similar to that observed in the patient's tumor tissue. PDO/immune cell co-cultures revealed that PD-L1-expressing organoids were unresponsive to nivolumab in vitro in the presence of PMN-MDSCs. Depletion of PMN-MDSCs within these co-cultures sensitized the organoids to anti-PD-1/PD-L1-induced cancer cell death. Rapamycin decreased phosphorylated S6K, Gli2 and PD-L1 expression in PDO/immune cell co-cultures. Transcriptional regulation of PD-L1 by GLI1 and GLI2 was blocked by rapamycin. In conclusion, the PDO/immune cell co-cultures may be used to study immunosuppressive MDSC function within the gastric tumor microenvironment. The mTOR signaling pathway mediates GLI-induced PD-L1 expression in gastric cancer.

Beneficial Effects of Gracillin From Rhizoma Paridis Against Gastric Carcinoma via the Potential TIPE2-Mediated Induction of Endogenous Apoptosis and Inhibition of Migration in BGC823 Cells.

Tumor necrosis factor-α inducible protein-8 (TIPE2), initially recognized as a negative immune regulator, exerts an important role in suppressing the progression of numerous cancers. In our previous investigation, we found that TIPE2 expression displayed a decrease or absence in gastric tumor tissue, and the overexpression of TIPE2 suppressed the growth of gastric cancer tumors and cells, demonstrating that TIPE2 could be a potential medicinal target for gastric cancer treatment. However, it’s seldomly reported that several medicinal agents or candidates targeted TIPE2 for treating diseases, including gastric cancer. To identify the candidate targeting TIPE2 to fight against gastric cancer, several extractions from traditional natural medicinal plants with anti-tumor functions were employed to screen the active compounds according to bioassay-guided isolation. Interestingly, gracillin, a component from the ethyl acetate extraction of Rhizoma Paridis, was identified to induce the expression of TIPE2 and inhibit the cell proliferation in gastric cancer BGC-823 cells. Furthermore, the underlying mechanisms that restrain gastric cancer were evaluated by clone formation, EdU staining, flow cytometry, and other assays. Meanwhile, the role of TIPE2 in the anti-tumor effect of gracillin was elucidated via the use of siTIPE2 RNA. It was determined that gracillin could fight against gastric cancer cells by inhibiting the cell proliferation participated by the PI3K/AKT pathway and cell cycle arrest, suppressing the EMT pathway-regulating cell migration, and inducing bcl2-associated mitochondrial apoptosis. Additionally, TIPE2 maybe contribute to the benefits of gracillin. These results of the present study are an important step toward the medicinal development of gracillin, and are also of use in understanding the effect of TIPE2 as a potential tumor target.

Clinicopathological Analysis and Prognostic Assessment of TCN1 in Patients with Gastric Cancer.

Stomach cancer is the fourth most common type of cancer worldwide. TCN1 mainly encodes the vitamin B12 transporter, transcobalamin. TCN1 is a marker of gastrointestinal tumor progression, but the impact of TCN1 on survival is unclear. Gastrointestinal tumor records were reviewed and analyzed, clinicopathological data were summarized, immunohistochemical detection of TCN1 was performed again, and the protein expression in tumor tissue, non-tumor tissue, and lymph nodes was semi-quantitatively analyzed. Patients were followed up for 5 years to determine the 5-year survival rates. The strong immune reactivity of the TCN1 protein was significantly correlated with tumor invasion depth, regional lymph nodes, and a tumor diameter of >5cm (Z = -2.531 and P = .016; Z = 3.785 and P < .001; Z = 2.541 and P = .049). Kaplan-Meier survival analysis showed that the total survival time of patients in the low-expression TCN1 group was significantly longer than that in the high-expression TCN1 group (P = .001; Table 2 and Figure 5). The mean survival time of all patients was 49.774 months (95% CI: 47.871-51.676; Table 4) and the 5-year overall survival rates were 73.3, 50.8, and 34.0%, respectively. Multivariate analysis revealed that regional lymph nodes (HR = 1.253; 95% CI: 1.031-1.747, P = .012), TCN1 immune expression status (HR = 2.707; 95% CI: 1.068-1.886, P = .016), and pTNM staging (HR = 2.293; 95% CI: 1.583-3.321; P = .001) were independent risk factors for poor survival. The high expression of TCN1 in gastric tumor tissues was found to be associated with the clinicopathological factors of patients, and the high expression of TCN1 was shown to indicate a poor clinical prognosis.

The Clinical Significance and Functional Role of miR-466 in Gastric Cancer Peritoneal Metastasis.

The prognosis of metastasis gastric cancer patients remains poor and the identification of novel molecular markers will improve the management of gastric cancer patients. The present study aimed to investigate the clinical significance and functional role of miR-466 in gastric cancer peritoneal metastasis. miR-466 expression was confirmed by RT-qPCR. The biological functions were examined by MTT assay, Transwell migration, and invasion assays. The Kaplan-Meier survival curve and multivariate Cox regression analysis were used to investigate the clinical role of miR-466. The logistic regression analysis was performed to reveal the risk factors associated with peritoneal metastasis. miR-466 expression was downregulated in gastric cancer cell lines, tumor tissues, and peritoneal metastasis tissues compared with respective controls. Increased miR-466 expression inhibited proliferation, migration, and invasion of gastric cancer cells. Besides, the lower expression of miR-466 in gastric cancer patients was associated with peritoneal dissemination. Furthermore, multivariate Cox proportional hazard regression analyses demonstrated miR-466 expression level as an independent predictor of prognosis of gastric cancer. The present study provides novel evidence for the clinical and biological significance of miR-466 expression as a possible biomarker for the prognosis and identifying patients with peritoneal metastasis, as well as a potential therapeutic target in patients with gastric cancer.

Proximity Labeling, Quantitative Proteomics, and Biochemical Studies Revealed the Molecular Mechanism for the Inhibitory Effect of Indisulam on the Proliferation of Gastric Cancer Cells.

Indisulam exhibits antitumor activity against several cancer cells. Although the DCAF15-indisulam-RBM39 axis has been well documented in the inhibition of cancer cell growth, it is unknown whether RBM39 degradation alone is the mechanism of action of indisulam. Here, we verified the inhibitory effect of indisulam on the proliferation of gastric cancer cells and its dependence on DCAF15. Proximity-dependent biotin labeling with TurboID and quantitative proteomics revealed that indisulam indeed promoted the interaction between DCAF15 and RBM39. Immunoblotting and immunofluorescence also revealed that indisulam promoted the ubiquitin-mediated RBM39 degradation and RBM39 colocalized with DCAF15 in the nucleus. DCAF15 knockdown almost completely abolished the indisulam-mediated RBM39 reduction. Further knockdown of RBM39 eliminated the effect of DCAF15 on the proliferation of gastric cancer cells upon indisulam treatment. Immunoblotting of gastric tumor tissues confirmed the downregulation of RBM39 by indisulam. Database analysis unveiled that RBM39 was highly expressed in gastric cancer tissues and its high expression significantly shortened the survival time of gastric cancer patients. Taken together, we demonstrated that indisulam enhanced RBM39 ubiquitination and degradation by promoting its interaction with DCAF15, thus inhibiting the proliferation of gastric cancer cells. This work may provide valuable information for drug discovery through proteolysis targeting chimeras. MS data were deposited in ProteomeXchange (Dataset identifier: PXD024168).

RETRACTED ARTICLE: Ethaselen synergizes with oxaliplatin in tumor growth inhibition by inducing ROS production and inhibiting TrxR1 activity in gastric cancer

BackgroundOxaliplatin is one of the most commonly used chemotherapeutic agent for the treatment of various cancers, including gastric cancer. It has, however, a narrow therapeutic index due to its toxicity and the occurrence of drug resistance. Hence, it is of great significance to develop novel therapies to potentiate the anti-tumor effect and reduce the toxicity of oxaliplatin. In our previous study, we demonstrated that ethaselen (BBSKE), an inhibitor of thioredoxin reductase, effectively inhibited the growth of gastric cancer cells and promoted apoptosis in vitro. In the present study, we investigated whether BBSKE can potentiate the anti-tumor effect of oxaliplatin in gastric cancer in vivo and vitro.MethodsCellular apoptosis and ROS levels were analyzed by flow cytometry. Thioredoxin reductase 1 (TrxR1) activity in gastric cancer cells, organoid and tumor tissues was determined by using the endpoint insulin reduction assay. Western blot was used to analyze the expressions of the indicated proteins. Nude mice xenograft models were used to test the effects of BBSKE and oxaliplatin combinations on gastric cancer cell growth in vivo. In addition, we also used the combined treatment of BBSKE and oxaliplatin in three cases of gastric cancer Patient-Derived organoid (GC-PDO) to detect the anti-tumor effect.ResultsWe found that BBSKE significantly enhanced oxaliplatin-induced growth inhibition in gastric cancer cells by inhibiting TrxR1 activity. Because of the inhibition of TrxR1 activity, BBSKE synergized with oxaliplatin to enhance the production of ROS and activate p38 and JNK signaling pathways which eventually induced apoptosis of gastric cancer cells. In vivo, we also found that BBSKE synergized with oxaliplatin to suppress the gastric cancer tumor growth in xenograft nude mice model, accompanied by the reduced TrxR1 activity. Remarkably, we found that BBSKE attenuated body weight loss evoked by oxaliplatin treatment. We also used three cases of GC-PDO and found that the combined treatment of BBSKE and oxaliplatin dramatically inhibited the growth and viability of GC-PDO with increased ROS level, decreased TrxR1 activity and enhanced apoptosis.ConclusionsThis study elucidates the underlying mechanisms of synergistic effect of BBSKE and oxaliplatin, and suggests that the combined treatment has potential value in gastric cancer therapy.Supplementary InformationThe online version contains supplementary material available at 10.1186/s13046-021-02052-z.

Precise Targeting Therapy of Orthotopic Gastric Carcinoma by siRNA and Chemotherapeutic Drug Codelivered in pH-Sensitive Nano Platform.

Gastric cancer is one of the most common malignant tumors, which remains as an obstacle to human health. Nowadays, targeted nanoparticles to gastric tumor tissues, provide new strategy for improved therapy but still remain challenging. The major hurdle of targeted therapeutic nanoparticles comes from the limited enrichment and poor selectivity of therapeutic agents in in situ tumor. Herein, a pH-sensitive targeted nano platform coloaded As2 O3 and human epidermal growth factor receptor-2 (HER2)-siRNA (AH RNPs) is developed to achieve targeting therapy in orthotopic gastric carcinoma. AH RNPs can effectively prevent the degradation of siRNA and overcome the poor solubility of As2 O3 . In vitro studies show that AH RNPs could achieve synergistic inhibition of growth and metastasis on SGC7901 cells. Surprisingly, AH RNPs not only target gastric subcutaneous tumor, but also target in situ tumor, and express loaded genes in in situ tumor. Moreover, AH RNPs show excellent antitumor effect in orthotopic gastric tumor model and the anticancer mechanism is related about inhibiting the activation of ERK signal and downregulating the expression of cxc chemokine receptor 4 (CXCR4), HER2, MMP2, and MMP9 protein. This study provides a multi-functional vector for precise targeting therapy of gastric cancer, which may serve as a potential clinical application for future gastric cancer.

Abstract 433: Clinical validation of a Dickkopf-1 (DKK1) chromogenic in-situ hybridization (CISH) assay for manual and image-analysis assisted pathologist interpretation

Abstract Dickkopf-1 (DKK1) is a secreted modulator of Wnt signaling that is frequently overexpressed in tumors and associated with a poor prognosis. In this study we demonstrate an approach for clinically validating a RNAscope chromogenic in situ hybridization (CISH) assay for determining the level of DKK1 RNA in Gastric (G) and Gastroesophageal (GEJ) tumor tissues according to CLIA guidelines. This two-step process validated first the performance of the wet assay along with the ability of a pathologist to manually score the CISH signal according to a dot-based H-score paradigm, and second the ability of image analysis (IA) software (Flagship Biosciences) to unbiasedly and reproducibly quantify DKK1 staining in the same set of samples. The DKK1 CISH assay for manual scoring passed all pre-determined criteria of sensitivity, specificity, accuracy, and precision. 100% of the 40 evaluated G/GEJ tissues demonstrated acceptable staining in target tumor cells and absence in non-target cells. 100% of the evaluated tissues passed sensitivity with a broad dynamic range of signal expression across target cells, and negligible background staining. Reproducibility was measured by blinded pathology scoring of a serial subset of 12 cases, resulting in 11/12 (92%) with concordant DKK1 H-scores. Accuracy was assessed with a DKK1 qPCR assay on a 20-sample subset and a significant correlation with the H-score data was observed. The IA algorithm also passed all pre-determined criteria of sensitivity, specificity, accuracy, and precision. 36/40 samples (90%) passed analytical specificity with the IA algorithm correctly classifying staining on true cells belonging to the tumor. Failed samples were attributed to non-specific alkaline phosphatase activity, which in practice would be disregarded by the reviewing pathologist. 100% of samples passed the sensitivity criteria of appropriate cell identification and classification. IA precision compared H-scores across 3 days of staining, with 11/12 (92%) cases having concordant DKK1 H-scores and an ICC of 0.9009 (95% CI: 0.7117-0.9601). Digital H-scores were highly correlated to the validated manual H-scores of the 40-sample set (r = 0.72, p &amp;lt;0.0001). Taken together, these data demonstrate a clinically validated DKK1 RNAscope CISH laboratory-derived test (LDT) for manual and IA-assisted pathologist interpretation. The DKK1 RNAscope LDT is currently being applied as part of a phase 2 clinical study of DKN-01 in combination with tislelizumab to prospectively identify previously treated G/GEJ adenocarcinoma patients with elevated DKK1 tumor expression (NCT04363801). Citation Format: Charles Caldwell, Mike Kagey, Sofia Reitsma, Will Paces, Elizabeth Bueche, Vitria Adisetiyo, Roberto Gianani. Clinical validation of a Dickkopf-1 (DKK1) chromogenic in-situ hybridization (CISH) assay for manual and image-analysis assisted pathologist interpretation [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 433.

Kinase inhibitor screening reveals aurora-a kinase is a potential therapeutic and prognostic biomarker of gastric cancer.

Gastric cancer is one of the most common and deadly types of cancer in the world, and poor prognosis with treatment failure is widely reported in the literature. In this context, kinases have been considered a relevant choice for targeted therapy in gastric cancer. Here, we explore the antiproliferative and antimigratory effects of the AURKA inhibitor and the prognostic and therapeutic value as a biomarker of gastric cancer. A total of 145 kinase inhibitors were screened to evaluate the cytotoxic or cytostatic effects in the gastric cancer cell line. Using the Alamar Blue assay, flow cytometry, quantitative polymerase chain reaction, and observation of caspase 3/7 activity and cell migration, we investigated the antiproliferative, proapoptotic, and antimigratory effects of the AURKA inhibitor. Moreover, AURKA overexpression was evaluated in the gastric cell lines and the gastric tumor tissue. Out of the 145 inhibitors, two presented the highest antiproliferative effect. Both molecules can induce apoptosis by the caspases 3/7 pathway in addition to inhibiting cancer cell migration, mainly the AURKA inhibitor. Moreover, molecular docking analysis revealed that GW779439X interacts in the active site of the AURKA enzyme with similar energy as a well-described inhibitor. Our study identified AURKA overexpression in the gastric cancer cell line and gastric tumor tissue, revealing that its overexpression in patients with cancer is correlated with low survival. Therefore, it is feasible to suggest AURKA as a potential marker of gastric cancer, besides providing robust information for diagnosis and estimated survival of patients. AURKA can be considered a new molecular target used in the prognosis and therapy of gastric cancer.

Vitamin D modulation and microRNAs in gastric cancer: prognostic and therapeutic role.

Gastric adenocarcinoma arises after a complex interaction between the host and environmental factors. Tumor location and TNM are the tools that currently guide treatment decisions. Surgery is the only curative treatment, but relapse is common. After relapse or advanced staged disease survival is poor and systemic treatment has modestly improved survival. An association between sun exposure, vitamin D status and gastric cancer (GC) incidence and mortality has been reported. The molecular differences of the histological subtypes and the new molecular classifications account for the great heterogeneity of this disease and are the basis for the discovery of new therapeutic targets. New prognostic and predictive factors are essential and microRNAs (miRNAs) are endogenous small non-coding RNA molecules with a great potential for diagnosis, prognosis and treatment of cancer. There are hundreds of miRNAs with altered expression in tumor gastric tissue when compared to normal gastric tissue. Many of these miRNAs are associated with clinicopathological variables and survival in patients with GC. Furthermore, the expression of some of these miRNAs with prognostic importance in CG is influenced by vitamin D and others are mediators of some of the actions of this vitamin. This review aims to update the evidence on several miRNAs with prognostic value and therapeutic potential in GC, whose expression may be influenced by vitamin D or may regulate vitamin D signaling.

Silencing of circ_0078607 prevents development of gastric cancer and inactivates the ERK1/2/AKT pathway through the miR-188-3p/RAP1B axis.

The aim of this study is to explore the expression and mechanism of circ_0078607 on proliferation and apoptosis of gastric cancer. Real time PCR (RT-PCR) was performed to detect the expression of circ_0078607 in gastric cancer tumor tissues, plasma and cell lines. Cell viability was detected by cell counting Kit-8. Cell proliferation ability was assessed by cell cycle assay. The samples were analyzed by flow cytometry for the detection of apoptosis. Luciferase assay and RNA immunoprecipitation (RIP) were carried out to verify the relationship between circ_0078607 and miR-188-3p, miR-188-3p, and RAP1B. Western blot was employed to detect the protein level of RAP1B, ERK1/2 and AKT. In vivo, the effect of circ_0078607 on gastric cancer tumor growth was detected by lentivirus vector injection. Here, we found the increased level of circ_0078607 in gastric cancer tissues, gastric cancer patients plasma and cell lines. Knockdown of circ_0078607 could prevent proliferation and induce cell apoptosis in MKN-28 cells. Then we verified that circ_0078607 could interact with miR-188-3p by performed luciferase assay and RIP. Furthermore, we observed that RAP1B was a potential target of miR-188-3p. Next, we found that miR-188-3p inhibitor or overexpression of RAP1B could prevent the anti-tumor function of sh-circ_0078607. Silencing of circ_0078607 inhibited ERK1/2/AKT signal pathways via regulating miR-188-3p/RAP1B. In vivo, knockdown of circ_0078607 inhibited tumor growth. Knockdown of circ_0078607 inhibits the proliferation and induces apoptosis of gastric cancer via miR-188-3p/RAP1B signal pathway.

Patients with Helicobacter pylori-positive gastric cancer with human cytomegalovirus infection have a low tendency of advanced lymphatic metastasis in a Chinese population.

Recognized as a group I carcinogen for gastric cancer (GC) and a factor involved in the development of GC, Helicobacter pylori serves a major part in GC research. However, most studies have focused on H. pylori itself, ignoring the complicated pathogenic microbiological environment of GC and neglecting the synergistic or antagonistic effects of H. pylori with other pathogenic microorganisms. Increasing evidence has revealed that the human cytomegalovirus (HCMV) is present in several types of tumors and serves an important role in the neoplastic process of certain human malignant tumors, including GC. The aim of the present study was to explore the role of HCMV and H. pylori co-infection in GC. HCMV and H. pylori infection was analyzed in paired gastric tumor and peri-tumoral tissues from 134 (98 male and 36 female) patients using PCR. The results revealed that a total of 74 (55.2%) patients had H. pylori infection, 58 patients (43.3%) had HCMV infection, and 34 (25.4%) patients had both HCMV and H. pylori infection. Univariate and multivariate analyses demonstrated that H. pylori infection was independently associated with advanced lymphatic metastasis [P=0.007; odds ratio (OR)=3.51]. Furthermore, compared with HCMV−/H. pylori−, neither HCMV+/H. pylori− nor HCMV+/H. pylori+ were associated with metastasis, but HCMV−/H. pylori+ co-infection status was an independent risk factor for advanced lymphatic metastasis (P=0.005; OR=6.00). In conclusion, GC co-infected with HCMV and H. pylori exhibited a low tendency of lymph node metastasis. HCMV may interact with H. pylori to inhibit the process of lymphatic metastasis, and the mechanism requires further investigation.

EPRS/GluRS promotes gastric cancer development via WNT/GSK-3β/β-catenin signaling pathway.

Glutamyl-prolyl-tRNA synthetase (EPRS/GluRS) is primarily part of the multi-synthetase complex that may play a key role in cancer development. However, the biological function, molecular mechanism, and inhibitor of EPRS have not been investigated in gastric cancer (GC). Immunohistochemistry was performed to detect the expression of EPRS in human gastric tumor tissues. Knocking down of EPRS, cell-derived xenograft mouse model, and patient-derived xenograft mouse model was used to identify the biological function of EPRS. Immunoprecipitation was applied to elucidate the interaction between EPRS and SCYL2. Computer docking model and multiple in vitro and in vivo experiments were conducted to discover EPRS inhibitors. Here, we report that EPRS is frequently overexpressed in GC tissues compared to that adjacent controls and its overexpression predicts poor prognosis in GC patients. Functionally, high expression of EPRS positively co-relates with GC development both in vitro and in vivo. Mechanistically, EPRS directly binds with SCYL2 to enhance the activation of WNT/GSK-3β/β-catenin signaling pathway and the accumulation of β-catenin in the nuclear, leading to GC cell proliferation and tumor growth. Moreover, we identified that xanthoangelol (XA) and 4-hydroxyderricin (4-HD) can directly bind to EPRS to block WNT/GSK-3β/β-catenin signaling pathway. More importantly, XA and 4-HD restrain gastric cancer patient-derived xenograft tumor growth and Helicobacter pylori combined with alcohol-induced atrophic gastritis and gastric tumorigenesis. These findings unveil a promising strategy for GC prevention and therapy by targeting EPRS-mediated WNT/GSK-3β/β-catenin cascades. Moreover, XA and 4-HD may be effective reagents used for GC prevention and therapy.

Genome-Scale Analysis Identified NID2, SPARC, and MFAP2 as Prognosis Markers of Overall Survival in Gastric Cancer.

BackgroundGastric cancer is the most common gastrointestinal tumor, and the rates of recurrence and metastasis are high. Research results on molecular biomarkers used for prognosis of gastric cancer remain inconclusive. This study aimed to explore the gene expression module of gastric cancer and to determine potential prognostic biomarkers.Material/MethodsThree microarray datasets (GSE13911, GSE79973, and GSE29272) from Gene Expression Omnibus (GEO), including 206 pairs of gastric tumors and adjacent normal samples, were used for analysis of differentially expressed genes (DEGs). The 3 microarray datasets yielded 144 genes associated with the progression and prognosis of gastric cancer. After this, a risk score model was developed for result validation using an independent dataset from The Cancer Genome Atlas.ResultsThe validation of the independent dataset showed significantly increased NID2, SPARC, and MFAP2 expression in gastric tumor tissues, which were associated with poor outcomes in gastric cancer patients. Moreover, the high risk score obtained was associated with poor overall survival (HR: 1.787; 1.069–2.986; P=0.027). Subgroup analyses revealed that these significant prognostic values were detected in patients aged <65.0 years, tumors in the antrum/distal colon, grade 3 tumors, or TNM-M0 stages of cancer.ConclusionsThe findings of this study show that NID2, SPARC, and MFAP2 are upregulated in gastric tumor tissues and are significantly associated with poor overall survival. Therefore, the predictive values of the risk score model employed for the prognosis of gastric cancer could be improved by using these 3 upregulated DEGs.