Metastasis Of Salivary Adenoid Cystic Carcinoma Research Articles

EN1 promotes lung metastasis of salivary adenoid cystic carcinoma by regulating the PI3K-AKT pathway and epithelial-mesenchymal transition

BackgroundEngrailed homeobox 1 (EN1) is a candidate oncogene that is epigenetically modified in salivary adenoid cystic carcinoma (SACC). We investigated the expression of EN1 in SACC tissues and cells, EN1 promoter methylation, and the role of EN1 in tumour progression in SACC.MethodsThirty-five SACC samples were screened for key transcription factors that affect tumour progression. In vitro and in vivo assays were performed to determine the viability, tumorigenicity, and metastatic ability of SACC cells with modulated EN1 expression. Quantitative methylation-specific polymerase chain reaction analysis was performed on SACC samples.ResultsEN1 was identified as a transcription factor that was highly overexpressed in SACC tissues, regardless of clinical stage and histology subtype, and its level of expression correlated with distant metastasis. EN1 promoted cell invasion and migration through epithelial-mesenchymal transition in vitro and enhanced SACC metastasis to the lung in vivo. RNA-seq combined with in vitro assays indicated that EN1 might play an oncogenic role in SACC through the PI3K-AKT pathway. EN1 mRNA levels were negatively correlated with promoter hypermethylation, and inhibition of DNA methylation by 5-aza-dC increased EN1 expression.ConclusionsThe transcription factor EN1 is overexpressed in SACC under methylation regulation and plays a pivotal role in SACC progression through the PI3K-AKT pathway. These results suggest that EN1 may be a diagnostic biomarker and a potential therapeutic target for SACC.

Hypoxia induced cell dormancy of salivary adenoid cystic carcinoma through miR-922/DEC2 axis

BackgroundHypoxia has been shown to induce cancer cells to become dormant meanwhile these cells inclined to disseminate and eventually cause metastasis. However, the molecular mechanism is still elusive. The purpose is to explore whether dormancy-associated microRNAs (DmiRs) get involved in hypoxia-induced cell dormancy of salivary adenoid cystic carcinoma (SACC). Material and MethodsThis study performed multi-perspective investigation of the biological effects of miR-922/DEC2 on SACC based on clinical samples, 2D and 3D in vitro model and nude mice in vivo model, based on our previous study of overexpression of DEC2 inducing SACC cellular dormancy. ResultsAccording to the existing microRNA array of SACC tissue, we found that miR-922 was upregulated in SACC tissue and was inversely correlated with DEC2, suggesting that miR-922 might participate in the activation of SACC cell dormancy as a DmiR. Then, we found miR-922 low SACC cells exhibited cell dormancy and a low level of fatty acid oxidation with propensity for lipid droplets accumulation through DEC2. Moreover, HIF1a downregulated the level of miR-922 to induce SACC cell dormancy. In addition, in xenografts of nude mice the inhibition of miR-922 attenuated the growth of primary tumor and the lung metastasis of SACC. ConclusionsmiR-922/DEC2 axis was necessary to hypoxia-induced cell dormancy and played an important role in the lipid metabolism reprogramming of SACC.

B-cell specific Moloney murine leukemia virus insertion site 1 contributes to invasion, metastasis, and poor prognosis in salivary adenoid cystic carcinoma

Background/purposeUpregulation of B-cell specific Moloney murine leukemia virus insertion site 1 (BMI-1) has been involved in the invasion, metastasis, and poor prognosis of many cancers. The aim of this study was to evaluate the levels and clinical significance of BMI-1 in saliva of patients with salivary adenoid cystic carcinoma (SACC), and to analyze biological function and mechanism of BMI-1 in the invasion and metastasis of SACC. Materials and methodsThe levels of BMI-1 in saliva and tumor tissues of SACC patients were determined. The correlation of salivary BMI-1 levels with clinicopathological parameters and clinical outcomes in patients with SACC was analyzed. Additionally, the effects of BMI-1 on wound-healing, transwell invasion, and epithelial-mesenchymal transition (EMT)-related protein expression in vitro as well as on tumorigenicity and experimental lung metastasis in vivo were investigated through exogenous overexpression and silencing of BMI-1 in SACC cells. ResultsBMI-1 levels increased in saliva and tumor tissues in SACC patients with invasion or metastasis. High salivary BMI-1 levels were correlated with poor TNM stage, poor overall survival, and disease-free survival. Exogenous expression of BMI-1 in SACC-83 promoted its migration and invasion, while silencing BMI-1 in SACC-LM inhibited its migration and invasion in vitro and suppressed tumorigenesis and lung metastasis in vivo. Furthermore, BMI-1 regulated the expression of EMT-related proteins in SACC. ConclusionOur study shows that BMI-1 can serve as a valuable biomarker to identify tumor invasion and metastasis in SACC, predict its prognosis, and act as a promising therapeutic target for SACC.

MiR-200b-5p inhibits tumor progression in salivary adenoid cystic carcinoma via targeting BTBD1

Salivary adenoid cystic carcinoma (SACC) is a rare malignant tumor of the salivary gland. Studies have suggested that miRNA may play a crucial role in the invasion and metastasis of SACC. This study aimed to investigate the role of miR-200b-5p in SACC progression. Reverse transcription-quantitative PCR and western blot assay were used to detect the expression levels of miR-200b-5p and BTBD1. The biological functions of miR-200b-5p were evaluated via wound-healing assays, transwell assays, and xenograft nude mice model. The interaction between miR-200b-5p and BTBD1 was assessed using luciferase assay. Results showed that miR-200b-5p was downregulated in the SACC tissues while BTBD1 was upregulated. miR-200b-5p overexpression suppressed SACC cell proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT). Bioinformatics prediction and luciferase reporter assay revealed that miR-200b-5p could directly bind to BTBD1. Besides, miR-200b-5p overexpression could rescue the tumor-promoting effect of BTBD1. miR-200b-5p inhibited tumor progression by modulating EMT-related proteins, targeting BTBD1 and inhibiting PI3K/AKT signaling pathway. Overall, our findings indicate that miR-200b-5p can suppress SACC proliferation, migration, invasion, and EMT by regulating BTBD1 and PI3K/AKT axis, providing a promising therapeutic target for SACC treatment.

Silencing of AJAP1 expression by promoter methylation activates the Wnt/β-catenin signaling pathway to promote tumor proliferation and metastasis in salivary adenoid cystic carcinoma.

Salivary adenoid cystic carcinoma (SACC) is a unique malignant tumor of the salivary gland with poor prognosis, which is not effective with chemotherapy and targeted drugs. Therefore, it is important to explore the molecular mechanism underlying SACC invasion and metastasis to develop novel therapeutic strategies and targets in clinical research. Real-time quantitative polymerase chain reaction (RT-qPCR) and western blot (WB) were performed to detect the expression of Adherens Junctions Associated Protein 1 (AJAP1). Methylation-specific PCR was used to evaluate the methylation of the AJAP1 promoter. AJAP1 was overexpressed or knocked down by lentivirus-mediated transfection. Kaplan-Meier analysis was conducted to create a survival curve and the log-rank test was used to analyze the overall survival (OS). The prognostic correlation was assessed using univariate and multivariate Cox regression analyses. Co-immunoprecipitation (Co-IP) was utilized to pull down the possible binding protein of AJAP1 and laser scanning confocal microscopy was applied to detect the subcellular localization of AJAP1, E-cadherin, and β-catenin. Cell viability, colony formation, wound healing, and Transwell invasion assays were performed to evaluate the function of AJAP1 in vitro. A subcutaneous xenograft assay in nude mice was performed to verify the function of AJAP1 in vivo. AJAP1 was downregulated in SACC tumors and was closely related to SACC lymph node/distant metastasis, which was an independent risk factor for SACC prognosis. Methylation-specific PCR confirmed that high methylation of the AJAP1 promoter was the main cause of its silencing. Overexpression or knockdown of AJAP1 in SACC cells could significantly inhibit or promote the proliferation, invasion, and metastasis of SACC cells, respectively, in both the in vitro and in vivo experiments. Mechanically, we found that AJAP1 binds to E-cadherin and β-catenin to form a complex in cytomembrane, reducing the nuclear translocation of β-catenin and blocking the Wingless/Integrated/β-catenin (Wnt/β-catenin) signaling pathway to play a suppressive role in cancer. In conclusion, these results suggest that the downregulation of AJAP1 protein expression may play a certain role in progression and metastasis of SACC. Our study indicates that AJAP1 may be a potential prognostic molecular marker and therapeutic target for SACC.

CCL25/CCR9 interaction promotes the malignant behavior of salivary adenoid cystic carcinoma via the PI3K/AKT signaling pathway.

BackgroundCC chemokine receptor 9 (CCR9), an organ-specific chemokine receptor, interacts with its exclusive ligand CCL25 to promote tumor proliferation and metastasis. However, the effect of CCR9 on salivary adenoid cystic carcinoma (SACC) malignant behavior remains unknown. This study aimed to investigate the specific molecular mechanism by which CCR9/CCL25 modulates malignant progression in SACC.MethodsImmunohistochemistry staining and RT–qPCR analyses were performed to detect the correlation of CCR9 expression and tumor progression-associated markers in SACC. In vitro, SACC cell proliferation and apoptosis were evaluated using Cell Counting Kit-8 and colon formation, and cell migration and invasion were detected by wound healing and transwell assays. Vercirnon was used as an inhibitor of CCR9, and LY294002 was used as an inhibitor of the PI3K/AKT pathway in this study. Western blot and RT–qPCR assays were carried out to measure the downstream factors of the interaction of CCL25 and CCR9. The effect of CCL25 on the development of SACC in vivo was examined by a xenograft tumor model in nude mice following CCL25, Vercirnon and LY294002 treatment.ResultsCCR9 was highly expressed in SACC compared with adjacent salivary gland tissues, and its level was associated with tumor proliferation and metastases. CCL25 enhanced cell proliferation, migration, and invasion through its interaction with CCR9 and exerted an antiapoptotic effect on SACC cells. Targeting CCR9 via Vercirnon significantly reduced the phosphorylation level of AKT induced by CCL25. CCL25/CCR9 could activate its downstream factors through the PI3K/AKT signaling pathway, such as cyclin D1, BCL2 and SLUG, thus promoting SACC cell proliferation, antiapoptosis, invasion and metastasis. The in vivo data from the xenograft mouse models further proved that CCL25 administration promoted malignant tumor progression by activating the PI3K/AKT pathway.ConclusionThe interaction of CCL25 and CCR9 promotes tumor growth and metastasis in SACC by activating the PI3K/AKT signaling pathway, offering a promising strategy for SACC treatment.

Long non-coding RNA MRPL23-AS1 suppresses anoikis in salivary adenoid cystic carcinoma in vitro.

Distant lung metastasis is the main factor that affects the survival rate of patients with salivary adenoid cystic carcinoma (SACC). Anoikis resistance is a feature of tumor cells that easily metastasize. The long non-coding RNA (lncRNA) MRPL23 antisense RNA 1 (MPRL23-AS1) is related to lung metastasis in SACC, but its role in anoikis resistance is unknown. After altering MPRL23-AS1 expression in SACC cells, anoikis resistance was detected by calcein AM/PI staining and annexin V/PI flow cytometry. The apoptosis marker activated caspase-3 and the bcl-2/bax ratio were detected by Western blotting. The relationship between MPRL23-AS1 and the promoter of the potential downstream target gene p19INK4D was identified by chromatin immunoprecipitation (ChIP)-PCR assay. p19INK4D expression in patient tissues was determined using qRT-PCR and immunohistochemistry. The functional experiments showed that MPRL23-AS1 could promote anoikis resistance in vitro. MRPL23-AS1 recruited the EZH2 to the promoter region of p19INK4D, inhibited p19INK4D expression, and promoted tumor cell anoikis resistance. p19INK4D overexpression did not affect anoikis in attached cells; however, it attenuated the anoikis resistance effect of MPRL23-AS1 in suspension cells. p19INK4D expression was significantly lower in SACC tissues than in normal tissues. The novel MRPL23-AS1/p19INK4D axis may be a potential SACC biomarker or therapeutic target.

Exosomal microRNA-23b-3p promotes tumor angiogenesis and metastasis by targeting PTEN in salivary adenoid cystic carcinoma.

MicroRNA (miR)-23b-3p is known to target various genes that are involved in cancer-related pathways. Exosomes are emerging intercellular communication agents. Exosomes secreted by cancer cells can deliver active molecules to the surrounding stromal cells, thereby influencing the recipient cells and promoting the development of cancers. However, the role of exosomal miR-23b-3p in salivary adenoid cystic carcinoma (SACC) is not yet clear. In this study, we set out to investigate the potential role of cancer-derived exosomal miR-23b-3p-related phosphatase and tensin homolog deleted on chromosome 10 in the alteration of angiogenesis and vascular permeability in SACC. We investigated the effect of exosomal miR-23b-3p on the progression of SACC. In vitro experiments indicated that exosomal miR-23b-3p led to an upregulation of vascular permeability, and reduced expression of tight junction proteins. In addition, exosomal miR-23b-3p also enhanced angiogenesis and migration. Next, the angiogenic effect of exosomal miR-23b-3p was validated in vivo, as it led to an increase in the tumor microvasculature. Furthermore, the growth rate of SACC was faster after injection of exosomes loaded with cholesterol-modified miR-23b-3p in mice. In conclusion, these results revealed that SACC cell-derived exosomes play an important role in promoting angiogenesis and local vascular microleakage of SACC by transporting miR-23b-3p, which suggests that miR-23b-3p in the exosomes may be a potential biomarker for distant metastasis of SACC. This suggests the potential of a novel therapeutic target by delivering anti-miR-23b-3p that focuses on exosomes.

Protein kinase D1 induced epithelial–mesenchymal transition and invasion in salivary adenoid cystic carcinoma via E‐cadherin/Snail regulation

Salivary adenoid cystic carcinoma (SACC) is a malignant tumor, which is characterized by a higher incidence of distant metastasis. The aim of this study was to investigate the role and mechanism of protein kinase D1 (PKD1) in regulating the epithelial-mesenchymal transition (EMT) and promotes the metastasis in SACC. We analyzed the expression of PKD1 in 40 SACC patients and different metastatic potential cell lines. Then, we investigated whether the migration and growth of SACC were regulated by PKD1 using shRNA interference or inhibition of kinase active in vitro cell. Moreover, the mechanism by which PKD1 regulates the stability of Snail protein was determined. Finally, nude mice were used to testify the function of PKD1 via tail vein injection. PKD1 was correlated with metastasis and poor prognosis of SACC patients. PKD1 inhibition attenuated proliferation, migration, invasion, and EMT of SACC cells. Conversely, kinase active PKD1 could induce EMT and promoted cell migration in human HSG cell. Furthermore, downregulation of PKD1 regulated Snail via phosphorylation at Ser-11 on Snail protein and promotion of proteasome-mediated degradation, and reduced lung metastasis in vivo. Our results suggest that PKD1 induces the EMT and promotes the metastasis, which illustrate that PKD1 may be a potential prognostic biomarker and serve as a potential therapeutic target for SACC patients.

The circular RNAs differential expression profiles in the metastasis of salivary adenoid cystic carcinoma cells.

In order to reveal circular RNAs (circRNAs) differential expression profiles and investigate the function and mechanism of circRNAs in the metastasis of salivary adenoid cystic carcinoma (SACC), microarray was used to detect differentially expressed circRNAs in SACC-83 and SACC-lung metastasis (LM) cell lines. Up-regulated circRNAs were analyzed using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses to further predict their function. Expression of candidate circRNA and microRNA (miRNA) was determined using quantitative real-time polymerase chain reaction (qRT-PCR). Constructed circRNA-miRNA-mRNA co-expression network was based on TargetScan, miRanda databases. Wound healing and transwell assays were completed to examine the effects of hsa_circRNA_001982 and miR-181a-5p on cell migration and invasion. qRT-PCR confirmed hsa_circRNA_092556, hsa_circRNA_101379, and hsa_circRNA_001982 up-regulation in SACC-LM. miR-181a-5p was down-regulated in SACC-LM and correlated with up-regulated hsa_circRNA_001982. Wound healing and transwell assays indicated that silencing hsa_circRNA_001982 inhibited the migration and invasion of the SACC-LM cells. Furthermore, over-expression of hsa_circRNA_001982 promoted the migration and invasion of SACC-83 cells. Interestingly, up-regulation or down-regulation of miR-181a-5p led to the opposite result in wound healing and transwell assays. Overall, differential expression circRNA profiles in SACC-83 and SACC-LM cells may reveal potential targets and a novel mechanism of circRNAs in the metastasis of SACC. Moreover, the interaction of hsa_circRNA_001982/miR-181a-5p is closely related to the metastasis of SACC cells.

Long Noncoding RNA MRPL23-AS1 Promotes Adenoid Cystic Carcinoma Lung Metastasis.

Lung metastasis is a major factor affecting long-term survival in patients with adenoid cystic carcinoma. Here, we showed that the long noncoding RNA (lncRNA) MRPL23 antisense RNA 1 (MRPL23-AS1) was highly expressed and correlated with lung metastasis and overall survival in patients with salivary adenoid cystic carcinoma (SACC). MRPL23-AS1 positively regulated epithelial-mesenchymal transition by forming an RNA-protein complex with enhancer of zeste homolog 2 (EZH2). MRPL23-AS1 increased the binding of EZH2 and H3K27me3 on the E-cadherin promoter region. Moreover, MRPL23-AS1 levels were higher in exosomes isolated from the blood plasma of patients with SACC, and exosomal MRPL23-AS1 affected pulmonary microvascular endothelial cells in an "exosomecrine" manner. MRPL23-AS1-enriched exosomes increased microvascular permeability and facilitated the metastasis of SACC in vivo. Collectively, these findings highlight a molecular mechanism of lung metastasis in SACC. MRPL23-AS1 may represent a biomarker and target for clinical intervention to control this intractable disease. SIGNIFICANCE: This study identifies a novel metastasis-promoting lncRNA MRPL23-AS1, which mediates the transcriptional silencing of E-cadherin through forming an RNA-protein complex with EZH2.

The NOTCH1-HEY1 pathway regulates self-renewal and epithelial-mesenchymal transition of salivary adenoid cystic carcinoma cells.

Our previous study demonstrated a close relationship between the NOTCH signaling pathway and salivary adenoid cystic carcinoma (SACC). Its receptor gene, NOTCH1, and its downstream gene, HES1, contribute to the proliferation, invasion and metastasis of SACC. Accumulating evidence supports HEY1 as another effector of the signaling pathway. The purpose of this study was to explore the effects of the NOTCH1-HEY1 pathway on the proliferation, invasion and metastasis of SACC cells. Our results verified that HEY1 is a specific molecular target of the NOTCH signaling pathway in SACC cells and that its expression in carcinoma is much higher than that in paracarcinoma tissues. The expression of NOTCH1 and HEY1 are positively correlated in the salivary adenoid cystic carcinoma tissues. NOTCH1 is significantly related to the activation of HEY1 in SACC, and that HEY1 reciprocally regulates NOTCH1 expression in SACC. HEY1 promotes cell proliferation and spheroid formation and inhibits cell apoptosis in vitro. In addition, HEY1 enhances the tumorigenicity of SACC in vivo. Furthermore, HEY1 increases cell invasion and metastasis by driving the expression of epithelial-mesenchymal transition (EMT)-related genes and MMPs. The results of this study indicate that the NOTCH1-HEY1 pathway is specifically upregulated in SACC and promotes cell proliferation, self-renewal, invasion, metastasis and the expression of EMT-related genes and MMPs. Our findings suggest that a NOTCH1-HEY1 pathway inhibitor might therefore have potential therapeutic applications in treating SACC patients by inhibiting cancer cell growth and metastasis.

Extracellular vesicles of carcinoma-associated fibroblasts creates a pre-metastatic niche in the lung through activating fibroblasts

ObjectivesCarcinoma-associated fibroblasts (CAFs) have been known to promote cancer progression by modifying the primary tumor microenvironment. We aimed to elucidate the intercellular communication between CAFs and secondary organs in salivary adenoid cystic carcinoma (SACC) metastasis.MethodsPre-metastatic and metastatic animal models of SACC were established using extracellular vesicles (EVs) from CAFs and SACC cells. Lung fibroblasts (LFs) were treated with EVs and their transcriptomic alterations were identified by RNA sequencing. ITRAQ were performed to analyze EV cargos. TC I-15 was used to inhibit EV uptake by LFs and SACC lung metastasis in vivo.ResultsHere, we show that CAF EVs induced lung pre-metastatic niche formation in mice and consequently increased SACC lung metastasis. The pre-metastatic niche induced by CAF EVs was different from that induced by SACC EVs. CAF EVs presented a great ability for matrix remodeling and periostin is a potential biomarker characterizing the CAF EV-induced pre-metastatic niche. We found that lung fibroblast activation promoted by CAF EVs was a critical event at the pre-metastatic niche. Integrin α2β1 mediated CAF EV uptake by lung fibroblasts, and its blockage by TC I-15 prevented lung pre-metastatic niche formation and subsequent metastasis. Plasma EV integrin β1 was considerably upregulated in the mice bearing xenografts with high risk of lung metastasis.ConclusionsWe demonstrated that CAF EVs participated in the pre-metastatic niche formation in the lung. Plasma EV integrin β1 might be a promising biomarker to predict SACC metastasis at an early stage. An integrated strategy targeting both tumor and stromal cells is necessary to prevent SACC metastasis.

Tumor-derived exosomes educate fibroblasts to promote salivary adenoid cystic carcinoma metastasis via NGF-NTRK1 pathway.

Salivary adenoid cystic carcinoma (SACC) is a rare head and neck malignancy characterized by unpredictable expansion, considerable perineural invasion and high risk of metastasis; however, the underlying mechanism of SACC progression remains unclear. Cancer-associated fibroblasts localized within the tumor microenvironment may promote cancer malignant transformation by enhancing tumor growth, blood vessel formation, inflammation development and metastasis occurrence. Small extracellular vesicles, including exosomes, are mediators of intercellular communication and can influence major tumor-associated pathways. The present study aimed to explore the exosome-mediated communication between SACC cells and fibroblasts. The results from confocal microscopy demonstrated that exosomes derived from the human cell line SACC-83 were internalized by the human periodontal ligament fibroblast (HPLF) cells. Following exosome internalization, HPLF cells appeared to enhance SACC-83 cell metastasis and were educated toward a protumorigenesis phenotype according to transcriptome RNA sequencing and reverse transcription-quantitative polymerase chain reaction analysis. This phenomenon included exosome-mediated stimulation of proinflammatory cytokines and nerve growth factor (NGF) secretion. Furthermore, NGF blockage reduced the enhanced SACC-83 cell invasion stimulated by the supernatant isolated from exosome-educated HPLF cells. In addition, the results reported that neurotrophic receptor tyrosine kinase 1 (NTRK1), which is the high-affinity NGF receptor, was significantly upregulated in human SACC-83 cells. These results demonstrated that SACC-83 cell-derived exosomes educated HPLF cells toward the protumorigenic phenotype via the NGF-NTRK1 pathway, which suggested that this type of exosomes may be used as a potential therapeutic target for SACC.

The MYB/miR-130a/NDRG2 axis modulates tumor proliferation and metastatic potential in salivary adenoid cystic carcinoma

Increasing evidence has emerged to suggest that N-myc downstream-regulated gene 2 (NDRG2) dysregulation participates in a number of tumor biological processes. However, the role of NDRG2 and miRNA-mediated NDRG2 regulation in salivary adenoid cystic carcinoma (SACC) progression remain unknown. Here, we determined that SACC tissues exhibited decreased level of NDRG2, which was associated with poorer rates of overall survival and distant metastasis-free survival. Silencing NDRG2 promoted SACC cell proliferation and metastasis both in vitro and in vivo. MiRNAs have been reported as vital regulators of NDRG2 expression. Based on micronome sequencing of three paired samples of SACC and normal salivary gland tissue and on an online database analysis, miR-130a was identified as a candidate miRNA that potentially regulates NDRG2. We demonstrated that the expression level of NDRG2 was dramatically reduced by exogenous miR-130a. Moreover, a luciferase assay further validated that miR-130a could degrade NDRG2 mRNA by targeting sites in the NDRG2 3′UTR. A rescue experiment suggested that NDRG2 expression could reverse the miR-130a-mediated promotion of cell proliferation and invasion. The expression of miR-130a has been reported to be regulated by certain transcription factors. In the preset study, we verified that the transcription factor MYB acted as the critical driver in SACC-upregulated miR-130a expression directly and induced NDRG2 downregulation in SACC tissues. Additionally, MYB/miR-130a activated the STAT3 and AKT pathways by downregulating NDRG2. These observations suggest that the MYB/miR-130a/NDRG2 axis, which modulates proliferation and metastasis in SACC, provides promising targets for the treatment of SACC.

Upregulation of lncRNA ADAMTS9-AS2 Promotes Salivary Adenoid Cystic Carcinoma Metastasis via PI3K/Akt and MEK/Erk Signaling.

Neurotropic infiltrative growth and distant metastasis are the main causes of death in salivary adenoid cystic carcinoma (SACC) patients. Long noncoding RNAs (lncRNAs) are involved in many human neoplasms, however, their potential roles in SACC are unclear. In our study, we found that ADAM metallopeptidase with thrombospondin type 1 motif, 9 (ADAMTS9) antisense RNA 2 (ADAMTS9-AS2) was significantly upregulated in SACC patients with metastasis and SACC-lung metastasis (LM) cells. Moreover, ADAMTS9-AS2expression was closely associated with the prognosis anddistant metastasis in SACC patients. Next, we found thatc-myc could specifically bind to the promoter of ADAMTS9-AS2 and activated its transcription. Knockdown of ADAMTS9-AS2 significantly inhibited migration and invasion of SACC cells invitro and distant lung metastasis invivo. Furthermore, ADAMTS9-AS2, which mainly expressed in the cytoplasm, shared microRNA (miRNA) response elements with Integrin α6 (ITGA6). Overexpression of ADAMTS9-AS2 competitively bound to miR-143-3p that inhibited ITGA6 from miRNA-mediated degradation, and thus it activated the activity of PI3K/Akt and MEK/Erk signaling and facilitated SACC metastasis. In summary, ADAMTS9-AS2 promotes migration and invasion in SACC by competing with miR-143-3p. This sheds a new insight into the regulation mechanism of ADAMTS9-AS2, and it provides a possible application for the SACC treatment.

Effect of simvastatin and microRNA-21 inhibitor on metastasis and progression of human salivary adenoid cystic carcinoma.

Salivary adenoid cystic carcinoma (SACC) is a common malignancy of the salivary glands. Epithelial-mesenchymal transition (EMT) and P53 signaling pathway are associated with SACC metastasis and progression. Although simvastatin (SIM) is effective against the growth of many cancer types, its side effects limit its use. microRNA-21 (miR-21) is highly expressed in a variety of tumors and has a role in promoting tumor development. Therefore, the aim of the present study was to evaluate the effect of SIM in combination with miR-21 inhibitor (miR-21i) against lung metastatic SACC cells (SACC-LM). Our results showed that miR-21i was effective in reducing the resistance of SACC-LM to SIM, resulting in SACC-LM acquisition of epithelial traits, cell migration and invasion reduction, growth inhibition and induction of apoptosis. The expression of proteins associated to metastasis and tumor progression were regulated by the combined use of SIM and miR-21i. Thus, our findings demonstrated that such combination was effective in inhibiting SACC-LM progression, suggesting that multi-target therapy against SACC might represent a potentially successful approach in clinical treatment.

IGFBP2 promotes salivary adenoid cystic carcinoma metastasis by activating the NF-κB/ZEB1 signaling pathway

Metastasis is a major cause of poor prognosis in patients suffered with salivary adenoid cystic carcinoma (SACC), in which many factors are implicated. In this study, we identified that IGFBP2, overexpressed in SACC, correlated positively with perineural invasion or metastasis and indicated worse outcome. Moreover, IGFBP2 overexpression could dramatically improve motility and invasion capacity of SACC cells in vitro. Mechanically, IGFBP2 enhanced expression of ZEB1 in a NF-κB (p65)-dependent manner and then promoted epithelial-mesenchymal transition (EMT) in SACC. In addition, IGFBP2 mutation in the nuclear localization signal could impede nuclear translocation of p65, lower ZEB1 expression, and abrogate the EMT process. In xenograft models, IGFBP2 overexpression promoted lung and liver metastases of SACC cells; while if nuclear IGFBP2 was reduced, the formation of metastases in lung and liver was weakened. Together, these results for the first time demonstrate that IGFBP2 plays an important role in invasion and metastasis of SACC through the NF-κB/ZEB1 signaling pathway and IGFBP2 may be a novel biomarker and target for SACC.

Genome-wide profiles of metastasis-associated mRNAs and microRNAs in salivary adenoid cystic carcinoma

Salivary adenoid cystic carcinoma (SACC) is often accompanied with poor prognosis due to local recurrence, distant metastasis, and perineural invasion. The mechanism involved in SACC metastasis is not yet fully understood. In this study, we profiled the expression of messenger RNA (mRNA) and microRNA (miRNA) in a SACC cell line, ACC-2, and a highly metastatic SACC cell line, ACC-M, using high-throughput sequencing. We discovered that: (1) differentially expressed (DE) mRNAs and DE miRNAs are potentially involved in SACC metastasis; (2) multiple regulatory interactions between DE miRNAs and DE mRNAs exist; and (3) miR-338–5p/3p target LAMC2 to impair motility and invasion of ACC-M and MDA-MB-231 cells. In conclusion, our study integrated the regulatory effects of miRNAs and mRNAs on SACC metastasis and provided a potential application for miRNAs in future therapeutic intervention.

BTBD7 silencing inhibited epithelial- mesenchymal transition (EMT) via regulating Slug expression in human salivary adenoid cystic carcinoma.

Epithelial-mesenchymal transition (EMT) is a complicated process that has been implicated in cancer progression and metastasis as well as the formation of many tissues and organs. BTB/POZ domain-containing protein 7 (BTBD7) is reported to regulate transcriptional factors and involved in the process of invasion and metastasis of some malignant tumors. Additionally, our preliminary studies have confirmed that BTBD7 expression was significantly correlated with Slug expression and poor prognosis of primary salivary adenoid cystic carcinoma (SACC). On this basis, this study further investigated function of BTBD7 in the invasion and metastasis of SACC in vitro, which may be a possible target of gene therapy in the future. The expression of BTBD7 and Slug were both examined in SACC-LM and SACC-83 cell lines by immunofluorescence staining. High invasive SACC-LM cells were transfected with BTBD7 siRNA and the expression levels of BTBD7 and Slug were detected in both gene and protein levels by qRT-PCR and western blot analysis. Assays were performed to survey cell migration, invasion and proliferation capabilities with BTBD7 silencing. BTBD7 and Slug proteins were detected in SACC-LM and SACC-83 cell lines. BTBD7 silencing down-regulated the expression of Slug and MMP9 meanwhile up-regulated the expression of E-cadherin in SACC-LM cells, the migration and invasion abilities of cells were obviously suppressed but with no influence on cell proliferation. BTBD7 silencing inhibited EMT through regulation of Slug expression in SACC-LM cells and might act as a potential molecular target for gene therapy of SACC.