Progression Of Human Gastric Cancer Research Articles

Human gastric cancer progression and stabilization of ATG2B through RNF5 binding facilitated by autophagy-associated CircDHX8

The role of circDHX8 in the interplay between autophagy and gastric cancer (GC) progression remains unclear. In this study, we investigated the mechanism underlying the role of hsa_circ_003899 (circDHX8) in the malignancy of GC. Differential expression of circRNAs between GC and normal tissues was determined using circle-seq and microarray datasets (GSE83521). These circRNAs were validated using qPCR and Sanger sequencing. The function of circDHX8 was investigated through interference with circDHX8 expression experiments using in vitro and in vivo functional assays. Western blotting, immunofluorescence, and transmission electron microscopy were used to establish whether circDHX8 promoted autophagy in GC cells. To elucidate the mechanism underlying the circDHX8-mediated regulation of autophagy, we performed bioinformatics analysis, RNA pull-down, mass spectrometry (MS), RNA immunoprecipitation (RIP), and other western Blot related experiments. Hsa_circ_0003899 (circDHX8) was identified as upregulated and shown to enhance the malignant progression in GC cells by promoting cellular autophagy. Mechanistically, circDHX8 increased ATG2B protein levels by preventing ubiquitin-mediated degradation, thereby facilitating cell proliferation and invasion in GC. Additionally, circDHX8 directly interacts with the E3 ubiquitin-protein ligase RNF5, inhibiting the RNF5-mediated degradation of ATG2B. Concurrently, ATG2B, an acetylated protein, is subjected to SIRT1-mediated deacetylation, enhancing its binding to RNF5. Consequently, we established a novel mechanism for the role of circDHX8 in the malignant progression of GC.

Abstract 1488: The FOXA2 contributes to tumor proliferation and invasion via HGF mediated ERK signaling pathway in human gastric cancer

Abstract Background: FOXA1 and FOXA2, members of the forkhead transcription factor family, are critical for epithelial differentiation in many endoderm-derived organs and important regulators in body homeostasis. It has been shown that FOXA2 plays an important role in tumorigenesis and carcinogenesis in glioma, breast cancer, hepatoma, and others. However, FOXA2 has not yet been elucidated on the tumorigenesis, proliferation, and invasion of human gastric cancer, and needs to be researched and discovered. In this study, the HGF-mediated association of FOXA2, extracellular signal-regulated protein kinase (ERK), and matrix metalloproteinase 9 (MMP9) and cancer cell proliferation and invasion were investigated in two types of human gastric cancer cell lines. Methods: In this study, cell culture, cDNA microarray analysis, western blotting, Real-time Polymerase chain reaction, Zymography, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, FOXA2 knock-down with short hairpin RNA (shRNA), chromatin immunoprecipitation assay, Standard two chamber invasion assay. Results: First, we confirmed that the expression level of FOXA2 was up-regulated by HGF (hepatocyte growth factor) in human gastric cancer cells. To identify the associated pathway of HGF-induced FOXA2 about ERK, MMP9 expression, the cells were treated with PI3K (Phosphoinositide 3-kinase) inhibitor (LY294002) and then analyzed by Western blotting. The role of FOXA2 associated with MMP9 and ERK was determined by knockdown cell of FOXA2. FOXA2-sh RNA cells showed a decreased level of MMP9 and ERK. HGF-mediated cell proliferation and invasion were decreased in FOXA2 knockdown cell. Conclusions: These results suggest that FOXA2 plays an important role in cell proliferation and metastasis in hepatocellular carcinoma, and FOXA2 may be a novel target for the prevention of progression and treatment of human gastric cancer. Citation Format: Jiyoon Jung, Sungae Koh, Kyunghee Lee. The FOXA2 contributes to tumor proliferation and invasion via HGF mediated ERK signaling pathway in human 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 1488.

PSMA1 mediates tumor progression and poor prognosis of gastric carcinoma by deubiquitinating and stabilizing TAZ

The deubiquitinating enzyme family in tumor progression play important role in intracellular protein degradation. The proteasome subunit alpha type 1 (PSMA1) has been reported to act as an oncogene in several human cancers. The present study aimed to reveal the functional significance of PSMA1 in gastric cancer (GC) progression and the underlying mechanisms. The expression of PSMA1 in human GC samples and GC cell lines was examined by western blot analysis, real-time PCR, immunohistochemistry (IHC), and in vitro ubiquitination assays and established a xenograft mouse model. We found that PSMA1 was upregulated in GC and promoted proliferation, migration and invasion in GC cells. Herein, we report transcriptional co-activator with PDZ-binding motif (TAZ) was a downstream gene of PSMA1. Mechanistically, PSMA1 directly interacted with and stabilized TAZ via deubiquitination in GC. Furthermore, we found that TAZ was the essential mediator of PSMA1-modulated oncogenic activity in vitro and in vivo. Examination of clinical samples confirmed that elevated mediators of PSMA1, concomitant with increased TAZ abundance, correlate with human GC progression. These data suggested that PSMA1 promotes GC progression and proliferation by deubiquitinating TAZ. PSMA1 promotes GC progression and proliferation regarding PSMA1-mediated deubiquitinating enzyme activity and suggest potential therapeutic targets for GC management.

USP3 promotes gastric cancer progression and metastasis by deubiquitination-dependent COL9A3/COL6A5 stabilisation

As an important regulator of intracellular protein degradation, the mechanism of the deubiquitinating enzyme family in tumour metastasis has received increasing attention. Our previous study revealed that USP3 promotes tumour progression and is highly expressed in gastric cancer (GC). Herein, we report two critical targets, COL9A3 and COL6A5, downstream of USP3, via the isobaric tags for relative and absolute quantification technique. Mechanistically, we observed that USP3 interacted with and stabilised COL9A3 and COL6A5 via deubiquitination in GC. Importantly, we found that COL9A3 and COL6A5 were essential mediators of USP3-modulated oncogenic activity in vitro and in vivo. Examination of clinical samples confirmed that elevated expression of USP3, concomitant with increased COL9A3 and COL6A5 abundance, correlates with human GC progression. These data suggest that USP3 promotes GC progression and metastasis by deubiquitinating COL9A3 and COL6A5. These findings identify a mechanism of GC metastasis regarding USP3-mediated deubiquitinating enzyme activity and suggest potential therapeutic targets for GC management.

CCNI2 promotes the progression of human gastric cancer through HDGF

BackgroundGastric cancer is a highly aggressive malignant tumor with heterogeneity and is still a global health problem. The present study aimed to investigate the role of Cyclin I-like (CCNI2) in the regulation of phenotype and tumorigenesis, as well as its underlying mechanisms.MethodThe expression profile of CCNI2 in gastric cancer was determined based on The Cancer Genome Atlas (TCGA) database and immunohistochemical staining. The effects of altered CCNI2 expression on the biological phenotypes such as proliferation, clone formation, apoptosis and migration of gastric cancer cell lines BGC-823 and SGC-7901 were investigated. Mice xenograft models were established to reveal the role of CCNI2 knockdown on tumorigenesis. The potential mechanism of CCNI2 regulating gastric cancer was preliminarily determined by RNA sequencing.ResultCCNI2 was abundantly expressed in gastric cancer and was positively correlated with pathological stage. Knockdown of CCNI2 slowed down the malignant progression of gastric cancer by inhibiting tumor cell proliferation, increasing the susceptibility to apoptosis and suppressing migration. Moreover, downregulation of CCNI2 attenuated the ability of gastric cancer cells to form tumors in mice. Additionally, there was an interaction between CCNI2 and transcription factor hepatoma-derived growth factor (HDGF) in SGC-7901 cells. Knockdown of CCNI2 alleviated the promoting effects of HDGF overexpression in gastric cancer cells.ConclusionsCCNI2 promoted the progression of human gastric cancer through HDGF, which drew further interest regarding its clinical application as a potential therapeutic target.

Activated neutrophils polarize protumorigenic interleukin-17A-producing T helper subsets through TNF-α-B7-H2-dependent pathway in human gastric cancer.

RationaleNeutrophils constitute massive cellular constituents in inflammatory human gastric cancer (GC) tissues, but their roles in pathogenesis of inflammatory T helper (Th) subsets are still unknown.MethodsFlow cytometry analysis and immunohistochemistry were used to analyze the responses and phenotypes of neutrophils in different samples from 51 patients with GC. Kaplan‐Meier plots and Multivariate analysis for the survival of patients were used by log‐rank tests and Cox proportional hazards models. Neutrophils and CD4+ T cells were purified and cultured for ex vivo, in vitro and in vivo regulation and function assays.ResultsGC patients exhibited increased tumoral neutrophil infiltration with GC progression and poor patient prognosis. Intratumoral neutrophils accumulated in GC tumors via CXCL6/CXCL8‐CXCR1‐mediated chemotaxis, and expressed activated molecule CD54 and co‐signaling molecule B7‐H2. Neutrophils induced by tumors strongly expressed CD54 and B7‐H2 in both dose‐ and time‐dependent manners, and a close correlation was obtained between the expressions of CD54 and B7‐H2 on intratumoral neutrophils. Tumor‐derived tumor necrosis factor‐α (TNF‐α) promoted neutrophil activation and neutrophil B7‐H2 expression through ERK‐NF‐κB pathway, and a significant correlation was found between the levels of TNF‐α and CD54+ or B7‐H2+ neutrophils in tumor tissues. Tumor‐infiltrating and tumor‐conditioned neutrophils effectively induced IL‐17A‐producing Th subset polarization through a B7‐H2‐dependent manner ex vivo and these polarized IL‐17A‐producing Th cells exerted protumorigenic roles by promoting GC tumor cell proliferation via inflammatory molecule IL‐17A in vitro, which promoted the progression of human GC in vivo; these effects could be reversed when IL‐17A is blocked. Moreover, increased B7‐H2+ neutrophils and IL‐17A in tumors were closely related to advanced GC progression and predicted poor patient survival.ConclusionWe illuminate novel underlying mechanisms that TNF‐α‐activated neutrophils link B7‐H2 to protumorigenic IL‐17A‐producing Th subset polarization in human GC. Blocking this pathological TNF‐α‐B7‐H2‐IL‐17A pathway may be useful therapeutic strategies for treating GC.

IL-1R2 expression in human gastric cancer and its clinical significance.

Background: Interleukin-1 receptor type II (IL-1R2), also known as CD121b, is a member of the IL-1 receptor family. IL-1R2 acts as negative regulator of the IL-1 system, modulating IL-1 availability for the signaling receptor. IL-1R2 is abnormally expressed in many human inflammatory diseases and cancers, and has important clinical significance. The present study was designed to investigate IL-1R2 expression in human gastric cancer (GC) tissues and the associated clinical implications. Methods: Immunohistochemistry was used to identify the clinical significance and prognostic value of IL-1R2 expression in GC tissues. We investigated IL-1R2 expression in GC tissues, cells, and serum using real-time PCR (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) assays. Results: IL-1R2 was highly expressed in GC tissues, and the overall survival in patients with advanced GC and high IL-1R2 expression was significantly poorer than that in patients with advanced GC and low IL-1R2 expression. Moreover, IL-1R2 mRNA levels in GC tissues and most GC cells were higher than those in para-cancer tissues and GES1 human gastric mucosal epithelial cells. The level of plasma-soluble IL-1R2 in GC patients was higher than that of the healthy control group. Conclusion: Increased IL-1R2 levels are involved in the initiation and progression of human GC, and IL-1R2 might be employed to develop immunotherapeutic approaches targeting GC.

High Expression of Interleukin-2 Receptor Subunit Gamma Reveals Poor Prognosis in Human Gastric Cancer.

Precision medicine for gastric cancer (GC) is still an unsolved issue, because most available target drugs are not specifically designed for GC. Exploring novel signaling molecules with target value for GC is in urgent need. This study aimed to reveal that interleukin-2 receptor subunit gamma (IL2RG) is such a key molecule in human GC progression. GC tissues and paracancerous gastric tissues were collected from 7 patients (5 males and 2 females) during tumor radical excision surgery. These tissues were used to identify the differentially expressed genes (DEGs) with RNA-seq and serial bioinformatics analyses including Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, gene expression profiling interactive analysis (GEPIA), and survival analysis. RT-qPCR and western blotting were performed to compare the mRNA and protein expression levels of IL2RG between GC tissues and adjacent normal gastric tissues as well as between GC cell line SGC-7901 and normal gastric epithelial cell line GES-1. Results showed striking elevations of IL2RG both in the mRNA and protein levels in GC tissues and human gastric cancer SGC-7901 cell line compared, respectively, with the adjacent normal gastric tissues and normal GES-1 cells, and higher IL2RG expression was associated with lower survival. Analyses on the GSE29272 and GSE15459 datasets from Gene Expression Omnibus verified that IL2RG was highly expressed in GC patients and was associated with poor overall survival. In addition, molecular docking showed that a small molecule, resatorvid (TAK 242), might be an inhibitor of IL2RG. We conclude that IL2RG is overexpressed in advanced GC and is associated with low survival. IL2RG may serve as a biomarker of GC progression and poor prognosis.

HHIP Overexpression Suppresses Human Gastric Cancer Progression and Metastasis by Reducing Its CpG Island Methylation.

Human hedgehog-interacting protein (HHIP), a negative regulator of hedgehog (HH) signaling pathway, has been reported to be dysregulated in many types of cancer, including gastric cancer. However, the inhibitory role of HHIP as well as the underlying molecular mechanism of HHIP regulation in gastric cancer haven’t been fully elucidated yet. In this study, we demonstrated that HHIP overexpression significantly suppressed the proliferation and invasion of AGS cells evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and transwell assays, respectively. Interestingly, methylation-specific polymerase chain reaction (MS-PCR, MSP) showed that HHIP overexpression dramatically decreased its de novo promoter methylation levels in AGS cells. Furthermore, HHIP expression was higher in adjacent non-cancerous tissue compared to matched gastric cancer tissue. High HHIP level was negatively correlated with metastasis (p = 0.035) but not local recurrence (p = 0.58). Taken together, our study suggested that HHIP can modulate gastric cancer progression and metastasis via regulation of its de novo promoter methylation levels in a feedback manner. Lower HHIP levels is positively associated with gastric cancer metastasis, which not only indicates HHIP could be served as a protective marker for gastric cancer, but also suggests restoring HHIP expression might be a potential therapeutic strategy for clinical treatment.

Ginkgolic acid (GA) suppresses gastric cancer growth by inducing apoptosis and suppressing STAT3/JAK2 signaling regulated by ROS

Gastric cancer is a frequently occurring cancer with high mortality each year worldwide. Finding new and effective therapeutic strategy against human gastric cancer is still urgently required. Ginkgolic acid (GA), a botanical drug, is extracted from the seed coat of Ginkgo biloba L. with various bioactive properties, including anti-tumor. Unfortunately, if GA has antitumor effect on human gastric cancer and the underlying molecular mechanisms have yet to be investigated. In the present study, we found that GA markedly reduced the gastric cancer cell viability. Furthermore, GA treatment led to the reduced migration ability of gastric cancer cells, which was associated with the decreased protein expression levels of Rho-associated protein kinase 1 (ROCK1), matrix metalloproteinase-2 (MMP-2), MMP-9 and α-smooth muscle actin (α-SMA). In addition, GA dose-dependently induced apoptosis in gastric cancer cells through activating Caspase-9/-3 and poly(ADP-Ribose) polymerase (PARP), which was along with the reduced Bcl-2 and Bcl-xl expression levels, and the elevated Bax and Bad levels. Consistently, Cyto-c protein expression in cytoplasm was also up-regulated by GA. Moreover, the production of reactive oxygen species (ROS) was significantly induced by GA. The activation of signal transducer and activator of transcription 3/janus kinase 2 (Stat3/JAK2) signaling pathway was inhibited by GA treatment. Intriguingly, blocking Stat3/JAK2 activation could further promote apoptosis and reduce cell viability induced by GA. However, GA-induced cell death was clearly abolished by ROS scavenger NAC, while the activation of Stat3/JAK2 signaling was restored by NAC. In vivo, GA showed effective role in reducing gastric tumor growth. Together, the findings here indicated that GA could be considered as an effective therapeutic candidate against human gastric cancer progression in future.

Magnoflorine inhibits human gastric cancer progression by inducing autophagy, apoptosis and cell cycle arrest by JNK activation regulated by ROS

The antitumor effect of magnoflorine (Mag), an alkaloid isolated from Coptidis Rhizoma, in gastric cancer (GC) cells has not been reported. In the study, Mag suppressed the proliferation of GC cells, but showed no influence on normal gastric cells. Mechanistically, Mag induced autophagy in GC cells, as evidenced by the up-regulated expression of LC3B-II and increased autophagosome formation. Furthermore, we found that Mag-triggered autophagic cell death was regulated by reactive oxygen species (ROS)-induced suppression of serine/threonine-protein kinases (AKT) signaling. What’s more, Mag treatment led to apoptosis in GC cells through enhancing cleaved Caspase-3 and PARP expressions. In addition, up-regulated expression of p27 and p21, as well as down-regulated expression of Cyclin-A and Cyclin-B1 was detected in Mag-treated GC cells, contributing to the S/G2 cell cycle arrest. Importantly, Mag incubation resulted in a significant increase in jun N-terminal kinase (JNK) phosphorylation but not p38 and ERK1/2, which was involved in the modulation of apoptosis and S/G2 phase arrest. Moreover, ROS production was highly induced by Mag treatment, and Mag-exhibited these functions was largely dependent on the generation of ROS in GC cells. Consistently, the GC cell xenograft mouse model confirmed the anti-tumor role of Mag in vivo. Collectively, these results indicated that Mag showed anti-GC effects, which could be a potential therapeutic target for GC treatment.

HOXD9 promotes the growth, invasion and metastasis of gastric cancer cells by transcriptional activation of RUFY3

BackgroundThe transcription factor HOXD9 is one of the members of the HOX family, which plays an important role in neoplastic processes. However, the role of HOXD9 in the growth and metastasis of gastric cancer (GC) remains to be elucidated.MethodsIn vitro functional role of HOXD9 and RURY3 in GC cells was determined using the TMA-based immunohistochemistry, western blot, EdU incorporation, gelatin zymography, luciferase, chromatin Immunoprecipitation (ChIP) and cell invasion assays. In vivo tumor growth and metastasis were conducted in nude mice.ResultsHOXD9 is overexpressed in GC cells and tissues. The high expression of HOXD9 was correlated with poor survival in GC patients. Functionally, HOXD9 expression significantly promoted the proliferation, invasion and migration of GC cells. Mechanically, HOXD9 directly associated with the RUFY3 promoter to increase the transcriptional activity of RUFY3. Inhibition of RUFY3 attenuated the proliferation, migration and invasiveness of HOXD9-overexpressing GC cells in vitro and in vivo. Moreover, both HOXD9 and RUFY3 were highly expressed in cancer cells but not in normal gastric tissues, with their expressions being positively correlated.ConclusionsThe evidence presented here suggests that the HOXD9-RUFY3 axis promotes the development and progression of human GC.

Chicoric acid (CA) induces autophagy in gastric cancer through promoting endoplasmic reticulum (ER) stress regulated by AMPK

Gastric cancer is one of the most common cancers leading to tumor-related deaths worldwide. Chicoric acid (CA) exhibits a variety of protective effects in different diseases. However, its role in regulating tumor progression has not been reported. Autophagy, as a conserved catabolic process, sustains cellular homoeostasis responding to stress to modulate cell fate. In the study, the effects of CA on gastric cancer were investigated. The results indicated that CA treatment markedly reduced the cell viability and induced apoptosis in gastric cancer cells, and prevented tumor growth in an established xenograft gastric cancer model. Furthermore, CA exposure significantly induced autophagy both in gastric cancer cells and tumor samples, as evidenced by the up-regulated expression of LC3II. Moreover, phosphorylated AMP-activated protein kinase (AMPK) and p70S6 kinase (p70s6k) expression were obviously promoted by CA in vitro and in vivo. Importantly, blocking AMPK activation abrogated CA-induced expression of LC3II in gastric cancer cells. In addition, endoplasmic reticulum (ER) stress in tumor samples or cells was markedly induced by CA treatment through promoting the expression of associated signals such as Parkin, protein kinase RNA-like ER kinase (PERK), activating transcription factors 4 (ATF4) and ATF6. Importantly, these effects were abolished by the inhibition of AMPK signaling. Collectively, our findings indicated that CA prevents human gastric cancer progression by inducing autophagy partly through the activation of AMPK, and represents an effective therapeutic strategy against gastric cancer development.

TNF-like ligand 1A is associated with progression and prognosis of human gastric cancer.

PurposeThis study aimed to investigate the function of TNF-like ligand 1A (TL1A) in the tumorigenesis and progression of gastric cancer (GC).MethodsRNA-seq gene expression and clinical information for GC patients were obtained from The Cancer Genome Atlas (TCGA) database. Differentially expressed genes (DEGs) between GC tissue samples and normal controls were screened with the edgeR package. Identification of gene co-expression and functional enrichment analyses were performed with Pearson’s correlation analysis and gene set enrichment analysis (GSEA), respectively. Lastly, survival analysis was performed using the Kaplan-Meier method with the log rank test.ResultsTL1A expression in GC tissue samples were significantly higher than that in normal controls (LogFC=1.07 and P=8.90E-07). Moreover, 215 genes, co-expressed with TL1A, and 21 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were obtained. Next, the miRNA-lncRNA/mRNA network, comprising 7 miRNAs, 27 lncRNAs, and 21 mRNAs, was constructed based on key genes from intersections between co-expression analysis and GSEA. In addition, survival analysis results demonstrated that TL1A (P=2.6e−07) was significantly associated with the overall survival (OS) of GC patients.ConclusionTL1A was involved in the tumorigenesis and progression of GC, and was significantly associated with the OS of GC patients.

Mucin 17 inhibits the progression of human gastric cancer by limiting inflammatory responses through a MYH9-p53-RhoA regulatory feedback loop

BackgroundMucins are key components of the mucosal barrier in the stomach that protects epithelia from carcinogenic effects of chronic inflammation. Analysis of The Cancer Genome Atlas database indicated that mucin-17 (MUC17) was more highly expressed in gastric cancer (GC) specimens, with favourable prognosis for patients. To explore the underlying mechanisms, we investigated the potential role of MUC17 in controlling chronic gastric inflammation.MethodsWe initially quantified the expression of MUC17 and inflammatory factor, as well as the association of MUC17 with survive in GC using immunohistochemistry. To establish how the inflammatory factors affect MUC17 expression, we explored luciferase reporter, chromatin immunoprecipitation (ChIP), and electrophoretic mobility shift (EMSA) assays. The role and mechanism that MUC17 plays in inflammation-induced cell proliferation was examined in AGS cells with reduced MUC17 expression and MKN45 cells overexpressing a truncated MUC17.ResultsWe found MUC17 was induced by inflammatory cytokines in GC cells via CDX1upregulation. MUC17 thus inactivated NFκB to inhibit GC cell proliferation in response to pro-inflammatory cytokines. We also revealed that the function of MUC17 was dependent on its conserved epidermal growth factor domain and on downstream sequences to enable its interaction with myosin-9, resulting in a sustained regulatory feedback loop between myosin-9, p53, and RhoA, and then activation of p38 to negatively regulate the NFκB pathway in GC cells. This mechanism was also confirmed in vivo.ConclusionsOur study demonstrates MUC17 as a GC suppressor protein which has the therapeutic potential for human GC.

MiR-9-5p Suppresses Malignant Biological Behaviors of Human Gastric Cancer Cells by Negative Regulation of TNFAIP8L3.

MicroRNA is essential for the malignant progression of human gastric cancer (GC), which is a leading cause of cancer deaths. However, the mechanism is still not so clear. In our present research, we investigated the effect of miR-9-5p in GC. We detected miR-9-5p expression in human gastric epithelial cell (GES-1) and GC cells (AGS, BGC-823, MKN-45, and MGC-803), plasma of normal or GC patients, as well as orthotopic xenograft mouse models by real-time PCR. The migration ability was detected by Transwell assays after miR-9-5p mimic or inhibitor transfection in GC cells. Our results showed that miR-9-5p expression in GC cells and plasma was significantly decreased. miR-9-5p inhibited migration of GC cells by regulating TNFAIP8L3 directly. Low expression of miR-9-5p in GC patients hardly suppressed the migration mediated by TNFAIP8L3. miR-9-5p, as a potential tumor suppressor gene, is closely related to the malignant progression of GC. Exploring the regulation between miR-9-5p and TNFAIP8L3 may provide a novel strategy for GC treatment.

The POU2F1/miR-4490/USP22 Axis Regulates Cell Proliferation and Metastasis in Gastric Cancer

Background: Gastric cancer (GC) remains the fourth most common cancer and the miRNAs can function as tumor suppressors or oncogenes.The biological role of miR-4490 has not yet been reported in the context of cancer. Methods: To explore the function of miR-4490, we performed colony formation assays, EdU incorporation assays, qRT-PCR, Western blotting, In situ hybridization (ISH), immunohistochemistry (IHC), flow-cytometry assays, ChIP and dual-luciferase reporter assays. The growth, migration, and invasive capacity of GC cells were evaluated. Findings: microRNA-4490 (miR-4490) was significantly downregulated in GC tumor samples and cell lines compared with that in normal controls and that its expression level was negatively correlated with the aggressiveness of GC. Ectopic miR-4490 expression not only reduced cell-cycle transition and proliferation but also significantly inhibited GC cell migration, invasion, and epithelial-mesenchymal transition (EMT) in vitro. Mechanistically, we confirmed that miR-4490 directly targeted USP22, which mediates the inhibition of proliferation and EMT-induced metastasis in vitro and in vivo. Moreover, we determined through luciferase and ChIP assays that the transcription factor POU2F1 directly bound to potential POU2F1 binding sites in the miR-4490 promoters and modulated their transcription. Spearman's correlation analysis showed a positive correlation between USP22 and POU2F1 and negative correlations between miR-4490 and USP22 as well as between miR-4490 and POU2F1 expression in GC tissues. Interpretation: Our findings suggest that POU2F1/miR-4490/USP22 axis plays an important role in the development and progression of human GC. Funding: National Natural Science Funds of China (Nos.81672875 and 81772964). Declaration of Interest: The authors declare that they have no competing interests. Ethical Approval: This study was reviewed and approved by the Medical Ethics Committee of Nanfang Hosital (NFEC-2017-062), Southern Medical University, Guangzhou, China. All animal studies were approved by the Institutional Animal Care and Use Committee of Committee of Nanfang Hosital.

HOXD9 Promotes the Growth, Invasion and Metastasis of Gastric Cancer Cells by Transcriptional Activation of RUFY3

Background: The transcription factor HOXD9 is one of the members of the HOX family, which plays an important role in neoplastic processes. However, the role of HOXD9 in the growth and metastasis of gastric cancer (GC) remains to be elucidated. Methods: In vitro functional role of HOXD9 and RURY3 in GC cells was determined using the TMA-based immunohistochemistry, western blot, EdU incorporation, gelatin zymography, luciferase, chromatin Immunoprecipitation (ChIP) and cell invasion assays. In vivo tumour growth and metastasis were conducted in nude mice. Findings: HOXD9 is overexpressed in GC cells and tissues. The high expression of HOXD9 was correlated with poor survival in GC patients. Functionally, HOXD9 expression significantly promoted the proliferation, invasion and migration of GC cells. Mechanically, HOXD9 directly associated with the RUFY3 promoter to increase the transcriptional activity of RUFY3. Inhibition of RUFY3 attenuated the proliferation, migration and invasiveness of HOXD9-overexpressing GC cells in vitro and in vivo. Moreover, both HOXD9 and RUFY3 were highly expressed in cancer cells but not in normal gastric tissues, with their expressions being positively correlated. Interpretation: The evidence presented here suggests that the HOXD9-RUFY3 axis promotes the development and progression of human GC. Funding Statement: This study was supported by grants from the National Natural Science Funds of China (Nos.81672875 and 81772964). The funding organization: the Guangzhou Pilot Project of the Clinical and Translational Research Center (early gastrointestinal cancer, No.7415696196402). Declaration of Interests: The authors declare that they have no conflict of interest. Ethics Approval Statement: This study was reviewed and approved by the Medical Ethics Committee of Nanfang Hospital (NFEC-2017-062), Southern Medical University, Guangzhou, China. All animal studies were approved by the Institutional Animal Care and Use Committee of Committee of Nanfang Hospital.

CDCA3 is a potential prognostic marker that promotes cell proliferation in gastric cancer.

Gastric cancer (GC) is an aggressive and highly lethal gastrointestinal cancer, with an exceedingly poor prognosis. In the present study, the carcinogenic mechanism of human GC and the role of cell division cycle‑associated 3 (CDCA3) were investigated. The expression levels of CDCA3 were investigated in GC samples and matched, peritumoral tissues by reverse transcription‑quantitative polymerase chain reaction and immunohistochemical analysis. The effects of CDCA3 on cell proliferation were explored by Cell Counting Kit‑8, colony formation, flow cytometric analysis and western blotting in vitro, and in vivo tumorigenesis in nude mice. The results demonstrated that CDCA3 expression was increased in human GC tissues compared with those in adjacent non‑tumor tissues. Evaluation of the clinicopathological significance indicated that CDCA3 was closely associated with features of GC and patients with unfavorable overall survival times. CDCA3 overexpression resulted in the stimulation of cell growth and colony formation in vitro and xenograft tumors in vivo. Conversely, knockdown of CDCA3 inhibited these effects. Furthermore, the ectopic expression of CDCA3 in SGC7901 cells consistently promoted the cell cycle transition from the G0/G1 phase to the S phase; whereas knockdown of CDCA3 in BGC823 cells blocked the transition from the G0/G1 phase. Additionally, the present study revealed that the Ras signaling pathway was involved in CDCA3‑mediated regulation of GC cell proliferation. CDCA3 activated the Ras signaling pathway to promote cell proliferation in vitro and in vivo in GC cells. Levels of CDCA3 greatly accelerated the progression of human GC. CDCA3 served as an oncogene, and may be a significant prognostic predictor and a novel therapeutic target for patients with GC.

Transcription Factor Myeloid Zinc-Finger 1 Suppresses Human Gastric Carcinogenesis by Interacting with Metallothionein 2A.

Metallothionein 2A (MT2A) suppresses the progression of human gastric cancer potentially through an "MT2A-NF-κB pathway" with unclear mechanisms. This study explored the role of a transcription factor, myeloid zinc-finger 1 (MZF1), in MT2A-NF-κB pathway and its clinical significance in gastric cancer. MZF1 expression and function in gastric cancer were investigated in vitro and in vivo. The relationship between MZF1 and MT2A was determined by gain-of-function and loss-of-function assays in gastric cancer cells and an immortalized gastric cell line GES-1. The prognostic value of MZF1 expression in association with MT2A was evaluated using IHC in two cohorts. MZF1 was epigenetically silenced in human gastric cancer cell lines and primary tumors. Overexpression of MZF1 in gastric cancer cells suppressed cell proliferation and migration, as well as the growth of xenograft tumors in nude mice. Knocking-down of MZF1 transformed GES-1 cells into a malignant phenotype characterized by increased cell growth and migration. Mechanistically, MZF1 was upregulated in both GC and GES-1 cells by MT2A ectopically expressed or induced upon treatment with a garlic-derived compound, diallyl trisulfide (DATS). MZF1 associated with MT2A was colocalized in the nuclei of GES-1 cells to target the promoter of NF-κB inhibitor alpha (NFKBIA). Clinically, MT2A and MZF1 were progressively downregulated in clinical specimens undergoing gastric malignant transformation. Downregulation of MT2A and MZF1 was significantly correlated with poorer patient prognosis. MT2A exerts its anti-gastric cancer effects by complexing with MZF1 to target NFKBIA. MT2A/MZF1 may serve as a valuable prognostic marker and a novel therapeutic target for human gastric cancer.