Regulation of epithelial-mesenchymal transition by tumor microenvironmental signals and its implication in cancer therapeutics.

Epithelial-mesenchymal transition (EMT) has been implicated in various aspects of tumor development, including tumor invasion and metastasis, cancer stemness, and therapy resistance. Diverse stroma cell types along with biochemical and biophysical factors in the tumor microenvironment impinge on the EMT program to impact tumor progression. Here we provide an in-depth review of various tumor microenvironmental signals that regulate EMT in cancer. We discuss the molecular mechanisms underlying the role of EMT in therapy resistance and highlight new therapeutic approaches targeting the tumor microenvironment to impact EMT and tumor progression.

Micropeptide CIP2A-BP encoded by LINC00665 inhibits triple-negative breast cancer progression.

TGF-ß signaling pathway plays a key role in breast cancer metastasis. Recent studies suggest that TGF-ß regulates tumor progression and invasion not only via transcriptional regulation, but also via translational regulation. Using both bioinformatics and experimental tools, we identified a micropeptide CIP2A-BP encoded by LINC00665, whose translation was downregulated by TGF-ß in breast cancer cell lines. Using TNBC cell lines, we showed that TGF-ß-activated Smad signaling pathway induced the expression of translation inhibitory protein 4E-BP1, which inhibited eukaryote translation initiation factor elF4E, leading to reduced translation of CIP2A-BP from LINC00665. CIP2A-BP directly binds tumor oncogene CIP2A to replace PP2A's B56γ subunit, thus releasing PP2A activity, which inhibits PI3K/AKT/NFκB pathway, resulting in decreased expression levels of MMP-2, MMP-9, and Snail. Downregulation of CIP2A-BP in TNBC patients was significantly associated with metastasis and poor overall survival. In the MMTV-PyMT model, either introducing CIP2A-BP gene or direct injection of CIP2A-BP micropeptide significantly reduced lung metastases and improved overall survival. In conclusion, we provide evidence that CIP2A-BP is both a prognostic marker and a novel therapeutic target for TNBC.

Mitochondria: a new intervention target for tumor invasion and metastasis.

Mitochondria, responsible for cellular energy synthesis and signal transduction, intricately regulate diverse metabolic processes, mediating fundamental biological phenomena such as cell growth, aging, and apoptosis. Tumor invasion and metastasis, key characteristics of malignancies, significantly impact patient prognosis. Tumor cells frequently exhibit metabolic abnormalities in mitochondria, including alterations in metabolic dynamics and changes in the expression of relevant metabolic genes and associated signal transduction pathways. Recent investigations unveil further insights into mitochondrial metabolic abnormalities, revealing their active involvement in tumor cell proliferation, resistance to chemotherapy, and a crucial role in tumor cell invasion and metastasis. This paper comprehensively outlines the latest research advancements in mitochondrial structure and metabolic function. Emphasis is placed on summarizing the role of mitochondrial metabolic abnormalities in tumor invasion and metastasis, including alterations in the mitochondrial genome (mutations), activation of mitochondrial-to-nuclear signaling, and dynamics within the mitochondria, all intricately linked to the processes of tumor invasion and metastasis. In conclusion, the paper discusses unresolved scientific questions in this field, aiming to provide a theoretical foundation and novel perspectives for developing innovative strategies targeting tumor invasion and metastasis based on mitochondrial biology.

CircROR1 upregulates CCNE1 expression to promote melanoma invasion and metastasis by recruiting KAT2A.

Circular RNAs (circRNAs) play important roles in cancer occurrence and progression. To explore and elucidate the clinical significance of specific circular RNA in melanoma and its potential molecular mechanism. CircROR1 expression in melanoma cells and tissues was confirmed by qRT-PCR and ISH. qRT-PCR and Western blotting were performed to measure the levels of CCNE1, KAT2A, MMP9 and TIMP2. MTT, Transwell and wound healing assays were performed to evaluate cell proliferation, invasion and metastasis. A xenograft mouse model was established to further verify the CircROR1/CCNE1 axis in vivo. RNA pull-down and RIP assays were performed to detect the direct interaction KAT2A and CircROR1. A ChIP assay was used to investigate the enrichment of H3K9ac acetylation in the CCNE1 promoter. CircROR1 was significantly upregulated in metastatic melanoma cells and tissues, promoting proliferation, invasion and metastasis in vitro and tumour growth in vivo. CircROR1 overexpression increased CCNE1 and MMP9 protein expression and decreased TIMP2 protein expression. Functional rescue assays demonstrated that CircROR1 played a role in promoting malignant progression through CCNE1. CircROR1 specifically bound to the KAT2A protein without affecting its expression. CircROR1 overexpression increased the level of H3K9ac modification in the CCNE1 promoter region by recruiting KAT2A, thus upregulating CCNE1 expression. CircROR1 upregulates CCNE1 expression through KAT2A-mediated histone acetylation. Our research confirms the critical role of CircROR1 in melanoma invasion and metastasis, and CircROR1 could serve as a potential therapeutic target for melanoma treatment.

14-3-3ε: a protein with complex physiology function but promising therapeutic potential in cancer.

Over the past decade, the role of the 14-3-3 protein has received increasing interest. Seven subtypes of 14-3-3 proteins exhibit high homology; however, each subtype maintains its specificity. The 14-3-3ε protein is involved in various physiological processes, including signal transduction, cell proliferation, apoptosis, autophagy, cell cycle regulation, repolarization of cardiac action, cardiac development, intracellular electrolyte homeostasis, neurodevelopment, and innate immunity. It also plays a significant role in the development and progression of various diseases, such as cardiovascular diseases, inflammatory diseases, neurodegenerative disorders, and cancer. These immense and various involvements of 14-3-3ε in diverse processes makes it a promising target for drug development. Although extensive research has been conducted on 14-3-3 dimers, studies on 14-3-3 monomers are limited. This review aimed to provide an overview of recent reports on the molecular mechanisms involved in the regulation of binding partners by 14-3-3ε, focusing on issues that could help advance the frontiers of this field. Video Abstract.

Integrated network pharmacology and experimental verification to explore the potential mechanism of San Ying decoction for treating triple-negative breast cancer.

Traditional Chinese medicine (TCM) has been used to treat triple-negative breast cancer (TNBC), a breast cancer subtype with poor prognosis. Clinical studies have verified that the Sanyingfang formula (SYF), a TCM prescription, has obvious effects on inhibiting breast cancer recurrence and metastasis, prolonging patient survival, and reducing clinical symptoms. However, its active ingredients and molecular mechanisms are still unclear. In this study, the active ingredients of each herbal medicine composing SYF and their target proteins are obtained from the Traditional Chinese Medicine Systems Pharmacology database. Breast cancer-related genes are obtained from the GeneCards database. Major targets and pathways related to SYF treatment in breast cancer are identified by analyzing the above data. By conducting molecular docking analysis, we find that the active ingredients quercetin and luteolin bind well to the key targets KDR1, PPARG, SOD1, and VCAM1. In vitro experiments verify that SYF can reduce the proliferation, migration, and invasion ability of TNBC cells. Using a TNBC xenograft mouse model, we show that SYF could delay tumor growth and effectively inhibit the occurrence of breast cancer lung metastasis in vivo. PPARG, SOD1, KDR1, and VCAM1 are all regulated by SYF and may play important roles in SYF-mediated inhibition of TNBC recurrence and metastasis.

Kaempferol inhibited invasion and metastasis of gastric cancer cells by targeting AKT/GSK3ß pathway based on network pharmacology and molecular docking.

This study aims to explore the mechanisms of the inhibitory effect of kaempferol on the invasion and metastasis of gastric cancer (GC) cells through network pharmacology prediction and experimental verification. It identifies core targets via PPI network analysis and finds that kaempferol binds to these targets well. In vitro experiments showed that kaempferol could inhibit the proliferation, colony formation, migration and invasion of GC cells. Western blotting indicated kaempferol may reduce AKT and GSK3ß phosphorylation, leading to lower expression of invasion-related genes SRC, MMP9, CXCR4, KDR, and MMP2. Overall, kaempferol may prevent migration and invasion of GC cells via the AKT/GSK3ß signaling pathway.

LncRNA-BC069792 suppresses tumor progression by targeting KCNQ4 in breast cancer.

BACKGROUND: Breast cancer is the most common malignant tumor that threatens women's health. Attention has been paid on the study of long- non-coding RNA (lncRNA) in breast cancer. However, the specific mechanism remains not clear. METHODS: In this study, we explored the role of lncRNA BC069792 in breast cancer. In vitro and in vivo functional experiments were carried out in cell culture and mouse models. High-throughput next-generation sequencing technology and real-time fluorescence quantitative PCR technology were used to evaluate differentially expressed genes and mRNA expression, Western blot and immunohistochemical staining were used to detect protein expression. RNA immunoprecipitation assay and dual-luciferase activity assay were used to evaluate the competing endogenous RNAs (ceRNA), and rescue and mutation experiments were used for verification. RESULTS: We found that lncRNA BC069792 was expressed at a low level in breast cancer tissues, and significantly decreased in breast cancer with high pathological grade, lymph node metastasis and high Ki-67 index groups. Moreover, BC069792 inhibited the proliferation, invasion and metastasis of breast cancer cells in vitro and in vivo. Mechanically, BC069792 acts as a molecular sponge to adsorb hsa-miR-658 and hsa-miR-4739, to up-regulate the protein expression of Potassium Voltage-Gated Channel Q4 (KCNQ4), inhibits the activities of JAK2 and p-AKT, and plays a role in inhibiting breast cancer growth. CONCLUSIONS: LncRNA BC069792 plays the role of tumor suppressor gene in breast cancer and is a new diagnostic index and therapeutic target in breast cancer.

In vivo CRISPR screens identify RhoV as a pro-metastasis factor of triple-negative breast cancer.

Metastasis is the main death reason for triple-negative breast cancer (TNBC). Thus, identifying the driver genes associated with metastasis of TNBC is urgently needed. CRISPR screens have dramatically enhanced genome editing and made it possible to identify genes associated with metastasis. In this study, we identified and explored the crucial role of ras homolog family member V (RhoV) in TNBC metastasis. Here, we performed customized in vivo CRISPR screens targeting metastasis-related genes obtained from transcriptome analysis of TNBC. The regulatory role of RhoV in TNBC was validated using gain- or loss-of-function studies in vitro and in vivo. We further conducted immunoprecipitation and LC-MS/MS to explore the metastasis mechanism of RhoV. In vivo functional screens identified RhoV as a candidate regulator involved in tumor metastasis. RhoV was frequently upregulated in TNBC and correlated with poor survival. Knockdown of RhoV significantly suppressed cell invasion, migration, and metastasis both in vitro and in vivo. In addition, we provided evidence that p-EGFR interacted with RhoV to activate the downstream signal pathway of RhoV, thereby promoting tumor metastasis. We further confirmed that this association was dependent on GRB2 through a specific proline-rich motif in the N-terminus of RhoV. This mechanism of RhoV is unique, as other Rho family proteins lack the proline-rich motif in the N-terminus.

Regulatory role of Chitinase 3-like 1 gene in papillary thyroid carcinoma proved by integration analyses of single-cell sequencing with cohort and experimental validations.

Papillary thyroid carcinoma (PTC) is one of the most common thyroid carcinomas. The gross extrathyroidal extension and extensive metastases of PTC lead to high rates of recurrence and poor clinical outcomes. However, the mechanisms underlying PTC development are poorly understood. In this study, using single-cell RNA sequencing, the transcriptome profiles of two PTC patients were addressed, including PTC1 with low malignancy and good prognosis and PTC2 with high malignancy and poor prognosis. We found that epithelial subcluster Epi02 was the most associated with the malignant development of PTC cells, with which the fold change of Chitinase 3-like 1 (CHI3L1) is on the top of the differentially expressed genes between PTC1 and PTC2 (P < 0.001). However CHI3L1 is rarely investigated in PTC as far. We then studied its role in PTC with a series of experiments. Firstly, qRT-PCR analysis of 14 PTC patients showed that the expression of CHI3L1 was positively correlated with malignancy. In addition, overexpression or silencing of CHI3L1 in TPC-1 cells, a PTC cell line, cultured in vitro showed that the proliferation, invasion, and metastasis of the cells were promoted or alleviated by CHI3L1. Further, immunohistochemistry analysis of 110 PTC cases revealed a significant relationship between CHI3L1 protein expression and PTC progression, especially the T (P < 0.001), N (P < 0.001), M stages (P = 0.007) and gross ETE (P < 0.001). Together, our results prove that CHI3L1 is a positive regulator of malignant development of PTC, and it promotes proliferation, invasion, and metastasis of PTC cells. Our study improves understanding of the molecular mechanisms underlying the progression of PTC and provides new insights for the clinical diagnosis and treatment of PTC.

A pan-cancer analysis revealing the role of LFNG, MFNG and RFNG in tumor prognosis and microenvironment.

BACKGROUND: Fringe is a glycosyltransferase involved in tumor occurrence and metastasis. However, a comprehensive analysis of the Fringe family members lunatic fringe (LFNG), manic fringe (MFNG), radical fringe (RFNG) in human cancers is lacking. METHODS: In this study, we performed a pan-cancer analysis of Fringe family members in 33 cancer types with transcriptomic, genomic, methylation data from The Cancer Genome Atlas (TCGA) project. The correlation between Fringe family member expression and patient overall survival, copy number variation, methylation, Gene Ontology enrichment, and tumor-infiltrating lymphocytes (TILs) was investigated by using multiple databases, such as cBioPortal, Human Protein Atlas, GeneCards, STRING, MSigDB, TISIDB, and TIMER2. In vitro experiments and immunohistochemical assays were performed to validate our findings. RESULTS: High expression levels of LFNG, MFNG, RFNG were closely associated with poor overall survival in multiple cancers, particularly in pancreatic adenocarcinoma (PAAD), uveal melanoma (UVM), and brain lower-grade glioma (LGG). Copy number variation analysis revealed that diploid and gain mutations of LFNG was significantly increased in PAAD and stomach adenocarcinoma (STAD), and significantly associated with the methylation levels in promoter regions. Significant differential genes between high and low expression groups of these Fringe family members were found to be consistently enriched in immune response and T cell activation pathway, extracellular matrix adhesion pathway, RNA splicing and ion transport pathways. Correlation between the abundance of tumor-infiltrating lymphocytes (TILs) and LFNG, MFNG, and RFNG expression showed that high LFNG expression was associated with lower TIL levels, particularly in PAAD. In vitro experiment by using pancreatic cancer PANC1 cells showed that LFNG overexpression promoted cell proliferation and invasion. Immunohistochemical assay in 90 PAAD patients verified the expression level of LFNG and its relationship with the prognosis. CONCLUSIONS: Our study provides a relatively comprehensive understanding of the expression, mutation, copy number, promoter methylation level changes along with prognosis values of LFNG, MFNG, and RFNG in different tumors. High LFNG expression may serve as a poor prognosis molecular marker for PAAD.

ING5 overexpression upregulates miR-34c-5p/Snail1 to inhibit EMT and invasion of lung cancer cells.

ING5 belongs to the inhibitor of growth (ING) candidate tumor suppressor family, which is involved in multiple cellular functions, such as cell cycle regulation, apoptosis, and chromatin remodelling. Previously, we reported that ING5 overexpression inhibits EMT by regulating EMT-related molecules, including Snail1, at the mRNA and protein levels. However, the mechanisms remain unclear. In the current study, we identify that ING5 overexpression induces the upregulation of miR-34c-5p. The expression levels of both ING5 and miR-34c-5p in NSCLC tissues from the TCGA database are decreased compared with that in adjacent tissues. Higher expression levels of both ING5 and miR-34c-5p predict longer overall survival (OS). Snail1 is the target gene of miR-34c-5p, as predicted by an online database, which is further verified by a dual-luciferase reporter assay. The expression level of Snail1 in NSCLC cells is markedly reduced following miR-34c-5p overexpression, leading to the inactivation of the Snail1 downstream TGF-ß/Smad3 signaling pathway. The TGF-ß signaling-specific inhibitor LY2157299 reverses the enhanced EMT, proliferation, migration, and invasion abilities induced by the miR-34c-5p inhibitor. Furthermore, tail vein injection of miR-34c-5p agomir inhibits xenografted tumor metastasis. Overall, this study concludes that miR-34c-5p, induced by ING5 overexpression, is a tumor suppressor that targets Snail1 and mediates the inhibitory effects of ING5 on the EMT and invasion of NSCLC cells. These results provide a novel mechanism mediating the antitumor effects of ING5.

A compound formulation of EGF-modified paclitaxel micelles and EGF-modified emodin micelles enhance the therapeutic effect of ovarian cancer.

Ovarian cancer is a serious threat to female health, although the incidence of it is relatively low, its mortality rate remains high due to its intense invasion and metastasis. Therefore, it is urgent to explore new treatment strategies for ovarian cancer. In this study, paclitaxel and emodin were encapsulated in different micelles, and loaded on the surface of the micelles with epidermal growth factor (EGF) as the targeting molecule, made compound formulations in proportion. In this study, EGF-modified paclitaxel micelles and EGF-modified emodin micelles were characterized, their inhibitory effects on SKOV3 cell proliferation and invasion were studied in vivo and in vitro, and its targeting ability was confirmed. The results showed that the shape, particle size, zeta potential, release rate, encapsulation rate, polydispersity index, and other physical and chemical properties of EGF-modified paclitaxel micelles plus EGF-modified emodin micelles meet the requirements, and the modification of EGF on the micelle surface could obviously improve the uptake of SKOV3 cells and inhibit the proliferation of SKOV3 cells. The compound formulation can inhibit the invasion and metastasis of ovarian cancer by inhibiting the expression of hypoxia inducible factor-α, MMP-2, MMP-9, and VE-cadherin. The in vivo studies have also showed significant pharmacodynamics results. These results indicated that EGF-modified paclitaxel micelles plus EGF-modified emodin micelles provide a new strategy for the treatment of ovarian cancer.

ETHE1 Accelerates Triple-Negative Breast Cancer Metastasis by Activating GCN2/eIF2α/ATF4 Signaling.

Triple-negative breast cancer (TNBC) is the most fatal subtype of breast cancer; however, effective treatment strategies for TNBC are lacking. Therefore, it is important to explore the mechanism of TNBC metastasis and identify its therapeutic targets. Dysregulation of ETHE1 leads to ethylmalonic encephalopathy in humans; however, the role of ETHE1 in TNBC remains elusive. Stable cell lines with ETHE1 overexpression or knockdown were constructed to explore the biological functions of ETHE1 during TNBC progression in vitro and in vivo. Mass spectrometry was used to analyze the molecular mechanism through which ETHE1 functions in TNBC progression. ETHE1 had no impact on TNBC cell proliferation and xenograft tumor growth but promoted TNBC cell migration and invasion in vitro and lung metastasis in vivo. The effect of ETHE1 on TNBC cell migratory potential was independent of its enzymatic activity. Mechanistic investigations revealed that ETHE1 interacted with eIF2α and enhanced its phosphorylation by promoting the interaction between eIF2α and GCN2. Phosphorylated eIF2α in turn upregulated the expression of ATF4, a transcriptional activator of genes involved in cell migration and tumor metastasis. Notably, inhibition of eIF2α phosphorylation through ISRIB or ATF4 knockdown partially abolished the tumor-promoting effect of ETHE1 overexpression. ETHE1 has a functional and mechanistic role in TNBC metastasis and offers a new therapeutic strategy for targeting ETHE1-propelled TNBC using ISRIB.

Triptonodiol, a Diterpenoid Extracted from Tripterygium wilfordii, Inhibits the Migration and Invasion of Non-Small-Cell Lung Cancer.

Lung cancer is the most prevalent oncological disease worldwide, with non-small-cell lung cancer accounting for approximately 85% of lung cancer cases. Tripterygium wilfordii is a traditional Chinese herb that is widely used to treat rheumatism, pain, inflammation, tumors, and other diseases. In this study, we found that Triptonodiol extracted from Tripterygium wilfordii inhibited the migration and invasion of non-small-cell lung cancer and inhibited cytoskeletal remodeling, which has not been previously reported. Triptonodiol significantly inhibited the motility activity of NSCLC at low toxic concentrations and suppressed the migration and invasion of NSCLC. These results can be confirmed by wound healing, cell trajectory tracking, and Transwell assays. We found that cytoskeletal remodeling was inhibited in Triptonodiol-treated NSCLC, as evidenced by the reduced aggregation of actin and altered pseudopod morphology. Additionally, this study found that Triptonodiol induced an increase in complete autophagic flux in NSCLC. This study suggests that Triptonodiol reduces the aggressive phenotype of NSCLC by inhibiting cytoskeletal remodeling and is a promising anti-tumor compound.

Interferon-induced transmembrane protein 2 promotes epithelial-mesenchymal transition by activating transforming growth factor-ß1/small mother against decapentaplegic 2 signaling in gastric cancer.

BACKGROUND: Gastric cancer (GC) is one of the most prevalent malignancy around the world. Primary tumor cells are enabled to invade and migrate into adjacent normal tissues to form secondary tumors. Epithelial-mesenchymal transitions (EMT) plays a pivotal role in facilitating tumor progression. Abundant evidence suggested that the transforming growth factor-ß1 (TGF-ß1) triggered the process of EMT. Nonetheless, the precise molecular mechanisms underlying EMT requires further elucidation, and there still lacks effective specific therapeutic target. In our recent research, we demonstrated that the interferon (IFN)-induced transmembrane protein 2 (IFITM2) promoted the growth and metastasis of GC. However, it remains unclear whether IFITM2 involves in TGF-ß1 mediated EMT in GC. METHODS AND RESULTS: In the present research, we investigated the functional role of IFITM2 in EMT process and TGF-ß1 signaling pathway in two GC cell lines. We noticed that silencing IFITM2 can effectively inhibit TGF-ß1 signaling mediated EMT by regulating down stream small mother against decapentaplegic (SMAD) 2/3 and transcription factors.This finding was further determined in both tumor tissues from GC patients and normal tissues adjacent to cancer. Our data demonstrated the key role of IFITM2 in TGF-ß1 signaling and EMT in GC. CONCLUSION: The findings enriched our understanding of the underlying mechanism in EMT during the progression of GC. In addition, IFITM2 would be a potential target for treating GC and other malignant tumors.

T-lymphoma invasion and metastasis 1 promotes invadopodia formation and is regulated by the PI3K/Akt signaling pathway in hepatocellular carcinoma.

At present, there are still many poorly understood aspects of the mechanisms underlying hepatocellular carcinoma (HCC) invasion and metastasis. Invadopodia are important structures for cancer cell invasion and metastasis. We determined that high T-lymphoma invasion and metastasis 1 (Tiam1) expression is associated with HCC invasion and metastasis and poor patient prognosis after surgery. Gain- and loss-of-function studies confirmed that Tiam1 promotes invadopodia formation in HCC by activating Rac1. A series of biochemical experiments confirmed that this effect is regulated by the PI3K/Akt signaling pathway. We also confirmed that PIP2 facilitates this effect. In summary, these findings reveal that Tiam1 plays an important role in invadopodia formation in HCC.

4sc-202 and Ink-128 cooperate to reverse the epithelial to mesenchymal transition in OSCC.

Treatment of oral squamous cell carcinoma remains a challenge due to a high incidence of treatment resistance, which is followed by tumor recrudescence and metastasis to the lymph nodes. Thus, it is important to explore novel inhibitors of OSCC. Here, we aimed to identify drugs that may cooperate with histone deacetylase inhibitors to reverse the EMT, inhibit EMT and cell migration and invasion, and contribute to therapeutic efficacy. We found that treatment with 4sc-202 potently reversed the EMT and thereby inhibited cell migration and invasion in vitro, in part by inducing expression of the FoxO1 tumor-suppressor gene. Furthermore, 4sc-202 also synergized with Ink-128 to inhibit tumor migration and invasion in vitro. Mechanistically, 4sc-202 induced FoxO1 expression, whereas Ink-128 promoted nuclear translocation of FoxO1. Our findings indicated that FoxO1 might reverse the EMT by interacting with Twist1 in OSCC. In conclusion, we identified an effective combination therapy involving class I histone deacetylase and mammalian target of rapamycin complex 1/2 inhibition that effectively blocked the EMT of tumor cells by upregulating FoxO1 expression to inhibit Twist1 transcription. These data have implications for developing new targets for early diagnosis and treatment of OSCC.

Poria Acid, Triterpenoids Extracted from Poria cocos, Inhibits the Invasion and Metastasis of Gastric Cancer Cells.

BACKGROUND: Poria cocos (P. cocos) is an important medicinal fungus in traditional Chinese medicine. Poria acid (PA), a triterpenoid compound, is an effective component of traditional Chinese medicine P. cocos. This experiment investigated the anti-gastric cancer biological activity of PA in vitro. METHODS: The effect of PA on the viability of gastric cancer cells was detected by the thiazolyl blue (MTT) assay. Cell adhesion assays were used to detect changes in the adhesion of cells treated after PA (0, 20, 40, and 80 µmol/L). The ability of cell invasion and migration were detected by Transwell assays and wound healing assays. A high-content imaging system was used to dynamically record the motility of the gastric cancer cells after PA (0, 20, 40, and 80 µmol/L) treatment. Western blotting was used to detect the expression of epithelial-mesenchymal transformation (EMT), invasion and migration related proteins. RESULTS: The MTT assay showed that the proliferation of gastric cancer cells was significantly inhibited after PA treatment. Cell adhesion experiments showed that the adhesion of gastric cancer cells was significantly decreased after PA treatment. Compared with the control group, the wound healing area of the gastric cancer cells treated with different concentrations of PA decreased. The Transwell assay showed that the number of gastric cancer cells passing through the cell membrane were significantly reduced after PA treatment. In addition, after PA treatment, the cells' movement distance and average movement speed were significantly lower than those of the control group. Finally, PA can significantly alter the expression of EMT-related proteins E-cadherin, N-cadherin, and Vimentin and decreased the expressions of metastasis-related proteins matrix metalloproteinase (MMP) 2, MMP-9 and tissue inhibition of matrix metalloproteinase (TIMP)1 in the gastric cancer cells. CONCLUSIONS: Triterpenoids from P. cocos have significant biological activity against gastric cancer, and the mechanism may be involved in the process of epithelial-mesenchymal transformation.

[MiR-4484 regulates the expression of integrin α 6 in gastric cancer tissues and its significance].

Objective: To investigate the relationship between the expression of integrin α 6 (ITGA6), miR-4484 and the pathologic stage of gastric cancer. Methods: Gastric cancer tissues and normal gastric mucosa tissues adjacent to cancer (>5 cm from tumor margin) of 30 patients with primary gastric cancer who underwent direct surgical resection without adjuvant therapy from June to September 2017 in West China Hospital of Sichuan University were selected. Real-time quantitative polymerase chain reaction (PCR) was used to detect the expression levels of miR-4484 and ITGA6, western blot was used to detect the expression level of ITGA6 protein, dual luciferase reporter gene was used to verify the relationship between ITGA6 and miR-4484. Spearman's correlation analysis was used to determine the relationship between miR-4484 and ITGA6 expression levels in gastric cancer tissues. Results: The expression level of ITGΑ6 in gastric cancer (32.30±13.47) was higher than that in matched normal gastric tissues (24.55±10.25, P=0.015), the area under the receiver operating characteristic (ROC) curve was 0.660 and the diagnostic sensitivity and specificity were 43.3% and 96.7%, respectively. The expression level of miR-4484 in gastric cancer (4.11±2.87) was lower than that of matched normal gastric tissues (5.75±2.80, P=0.029), the area under the ROC curve was 0.690 and the diagnostic sensitivity and specificity were 30.0% and 86.7%, respectively. The expression level of miR-4484 was negatively correlated with ITGA6 in gastric cancer tissues (r=-0.621, P<0.001). The expression level of ITGA6 protein in gastric cancer tissues (0.65±0.19) was higher than that in normal adjacent tissues (0.26±0.12, P<0.001). Compared with ITGA6 3'UTR wild-type+ miR-NC group, ITGA6 3'UTR wild-type+ miRNA mimics group had lower luciferase activity (50.69±5.10, 34.00±1.19, P<0.001), while the luciferase activity of ITGA6 3'UTR wild-type+ ASO miR-4484 group was higher than that of ITGA6 3'UTR wild-type+ miR-NC group (82.44±6.37, 50.69±5.10, P<0.001), indicated that ITGA6 was the direct target gene of miR-4484. The expression levels of miR-4484 in T1, T2, T3 and T4 (4a and 4b) gastric cancer tissues were 9.98±2.24, 5.28±2.03, 2.92±2.04 and 4.11±2.87, respectively, with statistical significance (P<0.001). The expression levels of ITGA6 in N0, N1, N2 and N3 gastric cancer tissues were 29.55±8.32, 21.71±3.75, 24.60±8.79 and 40.69±15.83, respectively, with statistical significance (P=0.022). The expression levels of miR-4484 in N0, N1, N2 and N3 gastric cancer tissues were 5.01±3.52, 5.48±2.76, 5.88±1.83 and 2.30±1.56, respectively, with statistical significance (P=0.032). The expression levels of ITGA6 in M0 and M1 gastric cancer tissues were 26.28±7.66 and 52.08±8.12, respectively, with statistical significance (P<0.001). The expression levels of miR-4484 in M0 and M1 gastric cancer tissues were 4.95±2.74 and 1.34±0.80, respectively, with statistical significance (P<0.001). Conclusions: ITGA6 is upregulated in gastric cancer tissues, while miR-4484 is downregulated in the gastric cancer group, and its expression level is related to the clinicopathological features of gastric cancer. ITGA6 is the direct target gene of miR-4484, implicates that miR-4484 may inhibit the invasion and metastasis of gastric cancer by regulating the expression of ITGA6. Both miR-4484 and ITGA6 may be the new prognostic markers and potential therapeutic targets of gastric cancer.