Targeting MetaLnc9/miR-143/FSCN1 axis inhibits oxidative stress and myofibroblast transdifferentiation in oral submucous fibrosis.

Background/purpose: Persistent activation of myofibroblasts is attributed to various dysregulated biological events conferring multiple types of fibrosis diseases, including oral submucous fibrosis (OSF). Although the significance of non-coding RNAs (ncRNAs) in the occurrence of fibrosis has been appreciated, the detailed mechanisms still have not been fully elucidated. The aim of this study was to identify key dysregulated ncRNAs and elucidate their pro-fibrotic mechanisms in promoting myofibroblast activation and the pathological development of OSF. Materials and methods: Expression of non-coding RNAs and mRNAs in OSF cohort was determined using RNA sequencing and qRT-PCR. The molecular axis of pro-fibrotic ncRNAs were exploited via luciferase reporter activity assay and RNA expression rescue experiments. Functional assays, including collagen gel contraction, wound healing ability, cell migration, and reactive oxygen species (ROS) production, were conducted to assess the changes in the myofibroblastic phenotypes of primary human buccal mucosal fibroblasts. Results: Herein, we found that long non-coding RNA MetaLnc9 was upregulated in OSF specimens and positively associated with several fibrosis markers. Silencing of MetaLnc9 diminished the features of activated myofibroblasts and the production of ROS. We not only showed that MetaLnc9 functioned as a competitive endogenous RNA of microRNA (miR)-143, but also demonstrated that the pro-fibrosis effect of MetaLnc9 on myofibroblast activities was mediated by suppression of miR-143. Moreover, our data showed that fascin actin-bundling protein 1 (FSCN1) was a direct target of miR-143 and positively related to MetaLnc9. Conclusion: Upregulation of MetaLnc9 may enhance the activation of myofibroblasts by sponging miR-143 and titrating its inhibitory property on FSCN1.

The FSCN1 gene rs2966447 variant is associated with increased serum fascin-1 levels and breast cancer susceptibility.

Fascin-1 (FSCN1) is recognized as an actin-binding protein, commonly exhibits up-regulation in breast cancer (BC) and is crucial for tumor invasion and metastasis. The existence of FSCN1 gene polymorphisms may raise the potential for developing BC, and there are still no studies focusing on the relationship between the FSCN1 rs2966447 variant and BC risk in Egyptian females. Thus, we investigated the serum fascin-1 levels in BC patients and the association between the FSCN1 rs2966447 variant with its serum levels and BC susceptibility. Genotyping was conducted in 153 treatment-naïve BC females with different stages and 144 apparent healthy females by TaqMan® allelic discrimination assay, whereas serum fascin-1 level quantification was employed by ELISA. The FSCN1 rs2966447 variant demonstrated a significant association with BC susceptibility under all utilized genetic models, cancer stages and estrogen receptor negativity. Also, BC females with AT and TT genotypes had higher serum fascin-1 levels and tumor size than those with the AA genotype. Moreover, serum fascin-1 levels were significantly elevated in the BC females, notably in those with advanced-stages. Furthermore, serum fascin-1 levels were markedly positively correlated with number of positive lymph nodes as well as tumor size. Collectively, these findings revealed that the FSCN1 rs2966447 variant may be regarded as a strong candidate for BC susceptibility. Also, this intronic variant is associated with increased serum fascin-1 levels and tumor size.

N6-methyladenosine methylation on FSCN1 mediated by METTL14/IGF2BP3 contributes to human papillomavirus type 16-infected cervical squamous cell carcinoma.

Human papillomavirus (HPV) infection has been reported to be associated with N6-methyladenosine (m6A) modification in cancers. However, the underlying mechanism by which m6A methylation participates in HPV-related cervical squamous cell carcinoma (CSCC) remains largely unclear. In this study, we observed that m6A regulators methyltransferase like protein (METTL14) and insulin like growth factor 2 mRNA binding protein 3 (IGF2BP3) were upregulated in HPV-positive CSCC tissues and cell lines, and their high expression predicted poor prognosis for HPV-infected CSCC patients. Cellular functional experiments verified that HPV16 oncogenes E6/E7 upregulated the expression of METTL14 and IGF2BP3 to promote cell proliferation and epithelial mesenchymal transition of CSCC cells. Next, we found that E6/E7 stabilized fascin actin-bundling protein 1 (FSCN1) mRNA and elevated FSCN1 expression in CSCC cells through upregulating METTL14/IGF2BP3-mediated m6A modification, and FSCN1 expression was also validated to be positively associated with worse outcomes of HPV-positive CSCC patients. Finally, HPV16-positive CSCC cell lines SiHa and CaSki were transfected with knockdown vector for E6/E7 or METTL14/IGF2BP3 and overexpressing vector for FSCN1, and functional verification experiments were performed through using MTT assay, flow cytometry, wound healing assay and tumour formation assay. Results indicated that knockdown of E6/E7 or METTL14/IGF2BP3 suppressed cell proliferation, migration and tumorigenesis, and accelerated cell apoptosis of HPV-positive CSCC cells. Their tumour-suppressive effects were abolished through overexpressing FSCN1. Overall, HPV E6/E7 advanced CSCC development through upregulating METTL14/IGF2BP3-mediated FSCN1 m6A modification.

The association of FSCN1 (rs852479, rs1640233) and HOTAIR (rs920778) polymorphisms with the risk of breast cancer in Egyptian women.

BACKGROUND: Breast cancer (BC) is one of the most prevalent cancers that contribute to mortality among women worldwide. Despite contradictory findings, considerable evidence suggests that single nucleotide polymorphisms (SNPs) in the FSCN1 and HOTAIR genes may have a causative impact on the development of BC. This case-control study was conducted to evaluate the association of genotype frequency in FSCN1 rs852479, rs1640233, and HOTAIR rs920778 with susceptibility and prognosis of BC, as well as the impact of clinical stages and hormonal features. METHODS AND RESULTS: FSCN1 (rs852479, rs1640233) and HOTAIR (rs920778) were genotyped using TaqMan real-time PCR assay in 200 BC patients and 200 cancer-free controls, all representing Egyptian women. Genotypic analyses in association with clinicopathological factors and disease risk were assessed. As a result, a significant association with BC risk was observed for CC genotype frequency of FSCN1 rs852479 A > C (OR = 0.395, 95% CI 0.204-0.76, p-value = 0.005). However, no significant correlation was detected between the FSCN1 rs1640233 C > T and HOTAIR rs920778 C > T polymorphic variants and susceptibility to BC. Interestingly, CC genotype of FSCN1 rs1640233 was more likely to progress tumor size and lymph node invasion in BC cases (p-value = 0.04 and 0.02, respectively). Moreover, it was revealed that there was a non-significant correlation between the haplotype distributions of FSCN1 rs852479 and rs1640233 and the probability of BC. CONCLUSIONS: Based on the sample size and genetic characteristics of the subjects involved in the present study, our findings indicated that FSCN1 rs852479 may contribute to BC susceptibility in a sample of the Egyptian population.

Fascin actin-bundling protein 1 regulates non-small cell lung cancer progression by influencing the transcription and splicing of tumorigenesis-related genes.

Background: High mortality rates are prevalent among patients with non-small-cell lung cancer (NSCLC), and effective therapeutic targets are key prognostic factors. Fascin actin-bundling protein 1 (FSCN1) promotes NSCLC; however, its role as an RNA-binding protein in NSCLC remains unexplored. Therefore, we aimed to explore FSCN1 expression and function in A549 cells. Method: We screened for alternative-splicing events and differentially expressed genes (DEGs) after FSCN1 silence via RNA-sequencing (RNA-seq). FSCN1 immunoprecipitation followed by RNA-seq were used to identify target genes whose mRNA expression and pre-mRNA alternative-splicing levels might be influenced by FSCN1. Results: Silencing FSCN1 in A549 cells affected malignant phenotypes; it inhibited proliferation, migration, and invasion, and promoted apoptosis. RNA-seq analysis revealed 2,851 DEGs and 3,057 alternatively spliced genes. Gene ontology-based functional enrichment analysis showed that downregulated DEGs and alternatively splicing genes were enriched for the cell-cycle. FSCN1 promoted the alternative splicing of cell-cycle-related mRNAs involved in tumorigenesis (i.e., BCCIP, DLGAP5, PRC1, RECQL5, WTAP, and SGO1). Combined analysis of FSCN1 RNA-binding targets and RNA-seq data suggested that FSCN1 might affect ACTG1, KRT7, and PDE3A expression by modulating the pre-mRNA alternative-splicing levels of NME4, NCOR2, and EEF1D, that were bound to long non-coding RNA transcripts (RNASNHG20, NEAT1, NSD2, and FTH1), which were highly abundant. Overall, extensive transcriptome analysis of gene alternative splicing and expression levels was performed in cells transfected with FSCN1 short-interfering RNA. Our data provide global insights into the regulatory mechanisms associated with the roles of FSCN1 and its target genes in lung cancer.

Genetic Polymorphism in FSCN1 rs3801004 C/G and CD44 rs353639 A/C, as Prognostic Factor in Egyptian Breast Cancer Patients.

BACKGROUND: One of the main causes of cancer-related deaths is breast cancer. Fascin-1(FSCN1) is an actin-binding protein that is present in the mesenchymal, neuronal, and endothelial cells of mammals. Patients with breast cancer have been found to have FSCN1 overexpression. CD44 is crucial for the development, invasion, and tumour spread. Therefore, we aimed to investigate the role of FSCN1&CD44 gene polymorphisms in breast cancer (BC) risk and prognosis. MATERIALS & METHODS: A total of 96 BC patients and 50 controls were included in the case-control study for risk prediction. We examined the association between The SNPs on FSCN1(rs3801004) and CD44(rs353639) and BC susceptibility and clinicopathological features using a real-time PCR in a cohort of the Egyptian population.  Results: A significant association of both SNPs on FSCN1(rs3801004)C allele and CD44(rs353639)A allele and BC susceptibility(adjusted OR=4.38,95%CI:2.6-7.4,p<0.001, and adjusted OR=4.44,95%CI:2.65-7.44,p <0.001,respectively). Moreover, CC genotype in FSCN1(rs3801004) were likely to progress to developing G2&G3 and N2&N3 and stage II & stage IV, according to the TNM staging and GG+GC genotypes increased within individuals who had a positive family history of BC. Individuals who carry at least one A allele for CD44rs353639 were likely to progress developing N2 according to the TNM in BC patients. CONCLUSIONS: These findings suggest that both SNPs on FSCN1 (rs3801004) and CD44 (rs353639) affected BC susceptibility. FSCN1 (rs3801004) genetic variants may have a significant effect on BC prognosis. However, CD44 (rs353639) affected lymph node invasions in BC patients.

Targeting FSCN1 with an oral small-molecule inhibitor for treating ocular neovascularization.

BACKGROUND: Ocular neovascularization is a leading cause of blindness and visual impairment. While intravitreal anti-VEGF agents can be effective, they do have several drawbacks, such as endophthalmitis and drug resistance. Additional studies are necessary to explore alternative therapeutic targets. METHODS: Bioinformatics analysis and quantitative RT-PCR were used to detect and verify the FSCN1 expression levels in oxygen-induced retinopathy (OIR) and laser-induced choroidal neovascularization (CNV) mice model. Transwell, wound scratching, tube formation, three-dimensional bead sprouting assay, rhodamine-phalloidin staining, Isolectin B4 staining and immunofluorescent staining were conducted to detect the role of FSCN1 and its oral inhibitor NP-G2-044 in vivo and vitro. HPLC-MS/MS analysis, cell apoptosis assay, MTT assay, H&E and tunnel staining, visual electrophysiology testing, visual cliff test and light/dark transition test were conducted to assess the pharmacokinetic and security of NP-G2-044 in vivo and vitro. Co-Immunoprecipitation, qRT-PCR and western blot were conducted to reveal the mechanism of FSCN1 and NP-G2-044 mediated pathological ocular neovascularization. RESULTS: We discovered that Fascin homologue 1 (FSCN1) is vital for angiogenesis both in vitro and in vivo, and that it is highly expressed in oxygen-induced retinopathy (OIR) and laser-induced choroidal neovascularization (CNV). We found that NP-G2-044, a small-molecule inhibitor of FSCN1 with oral activity, can impede the sprouting, migration, and filopodia formation of cultured endothelial cells. Oral NP-G2-044 can effectively and safely curb the development of OIR and CNV, and increase efficacy while overcoming anti-VEGF resistance in combination with intravitreal aflibercept (Eylea) injection. CONCLUSION: Collectively, FSCN1 inhibition could serve as a promising therapeutic approach to block ocular neovascularization.

Loss of MEG3 contributes to the enhanced migration and invasion in arsenic-induced carcinogenesis through NQO1/FSCN1 pathway.

Arsenic ranks at the top among all toxic metals and poses a serious threat to human health. Inorganic arsenite and arsenate compounds have been classified as human carcinogens in various types of cancers. Maternally expressed gene 3 (MEG3), a tumor suppressor that is commonly lost in cancer, was investigated in this study for its role in the migration and invasion of arsenic-transformed cells. Our results showed that MEG3 was downregulated in both arsenic-transformed cells (As-T) and cells treated with low doses of arsenic for three months (As-treated). The analysis using TCGA dataset revealed that MEG3 expression was significantly reduced in the tumor tissues from human lung adenocarcinoma (LUAD) and lung squamous cell carcinoma (LUSC) compared to normal lung tissues. The results from the methylation-specific PCR (MSP) assay demonstrated enhanced methylation in the MEG3 promoters in both As-T and As-treated cells, indicating that increased methylation of the MEG3 promoter caused MEG3 downregulation in these cells. Moreover, As-T cells displayed increased migration and invasion and higher levels of NAD(P)H quinone dehydrogenase 1 (NQO1) and fascin actin-bundling protein 1 (FSCN1). Consistently, the results from immunohistochemistry staining showed that both NQO1 and FSCN1 are highly expressed in human lung squamous cell carcinoma tissues compared to those in normal lungs. Knockdown of MEG3 in normal BEAS-2B cells also led to increased migration and invasion, along with elevated levels of NQO1 and FSCN1. The negative regulation of MEG3 on FSCN1 was restored by NQO1 overexpression in both As-T and BEAS-2B cells. The results from immunoprecipitation assays confirmed the direct binding of NQO1 to FSCN1. Overexpression of NQO1 increased migration and invasion abilities in BEAS-2B cells, while knockdown of NQO1 by its shRNA reduced these two hallmarks of cancer. Interestingly, the reduced migration and invasion by NQO1 knockdown were restored by FSCN1. Collectively, the loss of MEG3 upregulated NQO1, which in turn stabilized FSCN1 protein through its direct binding, resulting in elevated migration and invasion in arsenic-transformed cells.

FSCN1 promotes proliferation, invasion and glycolysis via the IRF4/AKT signaling pathway in oral squamous cell carcinoma.

BACKGROUND: Oral squamous cell carcinoma (OSCC) is a disease with increasing incidence worldwide that leads to deformity and death. In OSCC, fascin actin-bundling protein 1 (FSCN1) is an oncogene involved in the tumorigenesis process. However, the functions and potential mechanisms of FSCN1 in the OSCC tumorigenesis process have not been reported thus far. METHODS: We used qRT‒PCR to detect the expression of FSCN1 in 40 paired OSCC tumor tissues (tumor) and neighboring noncancerous tissues. The role of FSCN1 was also assessed in vitro through colony formation, CCK-8, and transwell assays. Moreover, glucose consumption was detected. Western blotting was used to confirm the interaction of FSCN1, IRF4 and AKT. RESULTS: FSCN1 was remarkably overexpressed in OSCC tissues and cell lines compared to corresponding controls. In addition, colony formation, CCK-8, and transwell assays revealed a notable reduction in OSCC growth and invasion when FSCN1 was silenced. FSCN1 silencing remarkably suppressed OSCC glycolysis. Mechanistic studies showed that FSCN1 achieves its function partially by activating interferon regulatory factor 4 (IRF4) and the AKT pathway in OSCC. CONCLUSION: In conclusion, our study investigated the functions and mechanisms of the FSCN1/IRF4/AKT pathway in OSCC progression. In OSCC, FSCN1 is likely to be a biomarker and therapeutic target.

FSCN1 has a potential indication for the prognosis and regulates the migration of HNSCC.

BACKGROUND: The study of molecular markers for diagnosis and prognosis is of great clinical significance for HNSCC patients. In this study, we proposed that FSCN1 has a potential indication for prognosis and is essential for the migration of HNSCC. METHODS: We analyzed the expression and survival association of FSCN1 in HNSCC using TCGA data. We compared the expression of FSCN1 in tumors from primary and metastasis HNSCC patients using QPCR, western blotting, and immunochemistry staining. We determined the migration velocity of multiple HNSCC cell lines using a chemotaxis migration assay. We analyzed the correlation between FSCN1 expression and HNSCC cell migration. We also test the effect of FSCN1 knockdown and overexpression on HNSCC cell migration. RESULTS: FSCN1 was overexpressed in HNSCC than pair normal tissues and metastasis HNSCC than primary HNSCC. FSCN1 expression was associated with significantly poorer overall survival of HNSCC patients. FSCN1 was potentially associated with immune cell infiltration and migration-associated genes. FSCN1 level was correlated with the migration in HNSCC cell lines. Knockdown of FSCN1 reduced the migration and the overexpression of FSCN1 promoted the migration of HNSCC cell lines. CONCLUSION: FSCN1 is a potential prognostic marker and a critical biomolecule for the migration of HNSCC.

SYTL2 promotes metastasis of prostate cancer cells by enhancing FSCN1-mediated pseudopodia formation and invasion.

BACKGROUND: Metastatic prostate cancer (mPCa) has a poor prognosis with limited treatment options. The high mobility of tumor cells is the key driving characteristic of metastasis. However, the mechanism is complex and far from clarified in PCa. Therefore, it is essential to explore the mechanism of metastasis and discover an intrinsic biomarker for mPCa. METHODS: Transcriptome sequencing data and clinicopathologic features of PCa from multifarious public databases were used to identify novel metastatic genes in PCa. The PCa tissue cohort containing 102 formalin-fixed paraffin-embedded (FFPE) samples was used to evaluate the clinicopathologic features of synaptotagmin-like 2 (SYTL2) in PCa. The function of SYTL2 was investigated by migration and invasion assays and a 3D migration model in vitro and a popliteal lymph node metastasis model in vivo. We performed coimmunoprecipitation and protein stability assays to clarify the mechanism of SYTL2. RESULTS: We discovered a pseudopodia regulator, SYTL2, which correlated with a higher Gleason score, worse prognosis and higher risk of metastasis. Functional experiments revealed that SYTL2 promoted migration, invasion and lymph node metastasis by increasing pseudopodia formation in vitro and in vivo. Furthermore, SYTL2 induced pseudopodia formation by enhancing the stability of fascin actin-bundling protein 1 (FSCN1) by binding and inhibiting the proteasome degradation pathway. Targeting FSCN1 enabled rescue and reversal of the oncogenic effect of SYTL2. CONCLUSIONS: Overall, our study established an FSCN1-dependent mechanism by which SYTL2 regulates the mobility of PCa cells. We also found that the SYTL2-FSCN1-pseudopodia axis may serve as a pharmacological and novel target for treating mPCa.

The role and regulatory mechanism of FSCN1 in breast tumorigenesis and progression.

FSCN1, an actin-bundling protein, is highly expressed in almost all metastatic tumors and is associated with the poor prognosis. In breast cancer FSCN1 is highly expressed in basal-like and triple negative subgroups. There is significant progress in understanding the role of fascin in breast cancer. Studies on FSCN1 in recent years have revealed that FSCN1 not only promotes tumor migration, invasion, metastic colonization, cancer cell self-renewal and drug resistance, but also regulates glucose and lipid metabolism and mitochondrial remodeling in tumor cells. In this review, we focus on the structure and regulatory mechanism of FSCN1 in breast tumorigenesis and metastasis, and discuss the clinical value of FSCN1 with the aim to provide a direction for further research in this field.

TIMEAS, a promising method for the stratification of testicular germ cell tumor patients with distinct immune microenvironment, clinical outcome and sensitivity to frontline therapies.

PURPOSE: With the heterogeneous genetic background, prognosis prediction and therapeutic targets for testicular germ cell tumors (TGCTs) are still unclear. We defined the tumor immune microenvironment activation status (TIMEAS). METHODS: We collected a total of 314 TGCT patients from four cohorts, including a 48-case microarray. A nonnegative matrix factorization algorithm was applied to identify the "immune factor", derived the top 150 weighted genes to divide patients into immune and non-immune classes, and further separated the immune class into activated and exhausted subgroups by nearest template prediction. Tumor mutant burden, gene mutation, and copy number alteration were compared with our recently developed package "MOVICS". A random forest algorithm was performed to establish a prediction model with fewer genes. Immunohistochemistry staining was performed to identify TIMEAS in the microarray. RESULTS: We constructed the TIMEAS in the TCGA-TGCT cohort and further validated it in the GSE3218 and GSE99420 cohorts. The immune class contained the activated status of T-lymphocytes, B-lymphocytes, and macrophages, while Treg cells and the WNT/TGFß signature were more activated in the immune-suppressed subgroup. Patients in the immune-exhausted subgroup had the worst prognosis, and 22.9% of patients in the immune-activated subgroup had KRAS mutations, which might stimulate the response of the immune system and lead to a favorable prognosis. The immune-exhausted group benefited more from chemotherapy, while the immune-activated subgroup responded well to anti-PD-1/PD-L1 therapy. FSCN1 was validated as the target of the immune-exhausted microenvironment by immunohistochemistry. CONCLUSION: TIMEAS classification can separate TGCT patients; patients in the immune-activated subgroup could benefit more from anti-PD-L1 immunotherapy, and those in the immune-exhausted subgroup are more suitable for chemotherapy.

FSCN1 acts as a promising therapeutic target in the blockade of tumor cell motility: a review of its function, mechanism, and clinical significance.

Fascin actin-bundling protein 1 (FSCN1) is an actin-bundling protein that is capable of inducing membrane protrusions and plays critical roles in cell migration, motility, adhesion, and other cellular interactions. FSCN1 also plays a role in forming and stabilizing filopodia or microspikes, which assist during cell migration. Furthermore, FSCN1 is a downstream target of several microRNAs and participates in various biological processes, such as epithelial-to-mesenchymal transition and autophagy, which regulate the invasion and migration ability of cells in various cancers. Increased FSCN1 levels have been associated with enhanced migration and invasion of multiple cancers as well as poor patient prognosis. Promising results from in vitro experimental studies using docosahexaenoic acid (DHA) in breast cancer and recombinant porcine NK-lysin A in hepatocellular carcinoma have revealed that anticancer drugs targeting FSCN1 have significant potential clinical applications. This review discusses FSCN1 in terms of five aspects: structure and function, biological processes, regulatory mechanisms, clinical applications, and future prospects.

FSCN1 Promotes Esophageal Carcinoma Progression Through Downregulating PTK6 via its RNA-Binding Protein Effect.

Objective: Esophageal squamous cell carcinoma (ESCC) causes many deaths worldwide every year. Fascin actin-bundling protein 1(FSCN1) has been reported to be a promoter of ESCC via its actin-binding function, however, its new role as an RNA-binding protein (RBP) has not been investigated. Here, we explored the RBP role of FSCN1 in the development of ESCC. Methods: Whole-genome expression sequencing was performed to screen for altered genes after FSCN1 knockdown. RNA immunoprecipitation was performed to determine the target mRNA of FSCN1 as an RBP. In vitro experiments with ECA-109 and KYSE-150 and ex vivo experiments in tumor-bearing mice were performed to investigate the effects of FSCN1 and Protein Tyrosine Kinase 6 (PTK6) on ESCC progression. Results: FSCN1 could downregulate mRNA and the protein level of PTK6. The binding position of PTK6 (PTK6-T2) pre-mRNA to FSCN1 was determined. PTK6-T2 blocked the binding between FSCN1 and the pre-mRNA of PTK6, and thus reversed the promotion effect of FSCN1 on ESCC tumor progression via the AKT/GSK3ß signaling pathway. Conclusion: A novel effect of FSCN1, RBP-binding with the pre-mRNA of PTK6, was confirmed to play an important role in ESCC progression. PTK6-T2, which is a specific inhibitor of FSCN1 binding to the pre-mRNA of PTK6, could impede the development of ESCC.

LncRNA CRNDE promotes cell proliferation, migration and invasion of ovarian cancer via miR-423-5p/FSCN1 axis.

Ovarian cancer seriously threatens the health of women. LncRNA CRNDE is known to be upregulated in ovarian cancer. However, the mechanism by which CRNDE regulates the progress of ovarian cancer is largely unknown. MTT assay was applied to measure the cell viability. Colony formation assay was used to measure the cell proliferation. Cell migration was tested by wound healing, and Transwell assay was performed to detect cell invasion. In addition, the expression of miR-423-5p, CRNDE and FSCN1 were detected by RT-qPCR and western blotting, respectively. Meanwhile, dual-luciferase reporter assay and RIP assay were performed to explore the correlation between miR-423-5p and CRNDE (or FSCN1). CRNDE and FSCN1 were upregulated in ovarian cancer cells (SKOV3, CAOV-3, IGROV1, A2780 and C13K), while miR-423-5p was downregulated. Moreover, silencing of FSCN1/CRNDE significantly decreased proliferation, migration and invasion of ovarian cancer cells (SKOV3 and CI3K) via suppressing MMP-2 and MMP-9. In addition, CRNDE could sponge miR-423-5p, and FSCN1 was confirmed to be the direct target of miR-423-5p. Furthermore, CRNDE knockdown-induced inhibition of FSCN1 was notably reversed by miR-423-5p downregulation. Knockdown of CRNDE inhibited cell proliferation, migration and invasion of ovarian cancer via miR-423-5p/FSCN1 axis. Thus, CRNDE may serve a new target for ovarian cancer.

Transcriptome analysis reveals the potential biological function of FSCN1 in HeLa cervical cancer cells.

Fascin actin-bundling protein 1 (FSCN1), an actin-bundling protein associated with cell migration and invasion, is highly expressed in various tumor tissues. FSCN1 has also been reported to be a marker of increased invasive potential in cervical cancers. However, the functions of FSCN1 are still not fully understood in cervical cancers. Here, the gene expression profile of HeLa cells transfected with FSCN1 shRNA (shFSCN1) was compared with that of cells transfected with empty vector (shCtrl). The results showed that shFSCN1 extensively affected the transcription level of 5,043 genes in HeLa cells. In particular, Gene Ontology (GO) analysis showed that a large number of upregulated genes were annotated with terms including transcription regulation and DNA binding. The downregulated genes were enriched in some cancer pathways, including angiogenesis and cell adhesion. qPCR validation confirmed that FSCN1 knockdown significantly affected the expression of selected genes in HeLa cells either negatively or positively. Expression analysis in TCGA (The Cancer Genome Atlas) revealed that FSCN1 had negative correlations with several transcription factors and a positive correlation with an angiogenic factor (angiopoietin like 4, ANGPTL4) in cervical tumor tissue. In particular, validation by Western blotting showed that FSCN1 knockdown decreased the protein level of ANGPTL4. Our results demonstrated that FSCN1 is not only an actin-binding protein but also a transcriptional regulator and an angiogenic factor in cervical cancer. Thus, our study provides important insights for further study on the regulatory mechanism of FSCN1 in cervical cancer.

A novel circ_0099999/miR-330-5p/FSCN1 ceRNA crosstalk in pancreatic cancer.

BACKGROUND: Pancreatic cancer is a lethal malignancy in both sexes throughout the world. Circular RNAs (circRNAs) have been implicated in the development of pancreatic cancer by operating as competing endogenous RNAs (ceRNAs). Here, we explored circ_0099999-mediated ceRNA activity in regulating pancreatic tumorigenesis. METHODS: Ribonuclease R (RNase R) and subcellular localization assays were utilized to characterize circ_0099999. The levels of circ_0099999, microRNA (miR)-330-5p, and fascin actin-bundling protein 1 (FSCN1) were gauged by quantitative real-time PCR (qRT-PCR) and western blot. Cell proliferation, colony formation, apoptosis, migration, and invasion were evaluated by the Cell Counting Kit-8 (CCK-8), colony formation, flow cytometry, and transwell assays, respectively. The levels of glucose consumption and lactate production were determined using the assay kits. A direct relationship between miR-330-5p and circ_0099999 or FSCN1 was validated by dual-luciferase reporter assay. Tumour xenograft assays were used to analyse the role of circ_0099999 in vivo. RESULTS: Circ_0099999 was highly up-regulated in pancreatic cancer tissues and cells. Knockdown of circ_0099999 impeded cell proliferation, migration, invasion, glycolysis, and promoted apoptosis in vitro, as well as diminished tumour growth in vivo. Circ_0099999 targeted miR-330-5p, and miR-330-5p was a downstream mediator of circ_0099999 function. FSCN1 was a direct and functional target of miR-330-5p. Furthermore, circ_0099999 operated as a ceRNA for miR-330-5p to modulate FSCN1 expression. CONCLUSIONS: Our findings established a novel causal mechanism, circ_0099999/miR-330-5p/FSCN1 ceRNA crosstalk, in regulating pancreatic carcinogenesis and provided that inhibition of circ_0099999 might have therapeutic benefits in pancreatic cancer.

LncRNA LOC146880 promotes esophageal squamous cell carcinoma progression via miR-328-5p/FSCN1/MAPK axis.

We investigated the role of long non-coding RNA (lncRNA) LOC146880 in esophageal squamous cell carcinoma (ESCC). LOC146880 was significantly upregulated in ESCC tissues (n = 21) and cell lines compared to the corresponding controls. Higher LOC146880 expression correlated with poorer overall survival (OS) of ESCC patients. Moreover, CREB-binding protein (CBP) and H3K27 acetylation levels were significantly higher in the LOC146880 promoter in ESCC cell lines than in the controls. LOC146880 silencing inhibited in vitro proliferation, invasion, migration, and epithelial-mesenchymal transition of ESCC cells. LOC146880 silencing also induced G1-phase cell cycle arrest and apoptosis in ESCC cells. Bioinformatics analysis, dual luciferase reporter assays, and RNA immunoprecipitation assays showed that LOC146880 regulates FSCN1 expression in ESCC cells by sponging miR-328-5p. Moreover, FSCN1 expression correlated with activation of the MAPK signaling pathway in ESCC cells and tissues. In vivo xenograft tumor volume and liver metastasis were significantly reduced in nude mice injected with LOC146880-silenced ESCC cells as compared to those injected with control shRNA-transfected ESCC cells. These findings show that the LOC146880/miR-328-5p/FSCN1/MAPK axis regulates ESCC progression in vitro and in vivo. LOC146880 is thus a promising prognostic biomarker and potential therapeutic target in ESCC.

Effect of rs67085638 in long non-coding RNA (CCAT1) on colon cancer chemoresistance to paclitaxel through modulating the microRNA-24-3p and FSCN1.

It has been reported that rs67085638 in long non-coding RNAs (lncRNA)-CCAT1 was associated with the risk of tumorigenesis. Also, CCAT1 could affect chemoresistance of cancer cells to paclitaxel (PTX) via regulating miR-24-3p and FSCN1 expression. In this study, we aimed to investigate the effect of rs67085638 on the expression of CCAT1/miR-24-3p/FSCN1 and the response of colon cancer to the treatment of PTX. 48 colon cancer patients were recruited and grouped by their genotypes of rs67085638 polymorphism as a CC group (N = 28) and a CT group (N = 20). PCR analysis, IHC assay and Western blot, TUNEL assay and flow cytometry were conducted. LncRNA-CCAT1 and FSCN1 mRNA/protein were overexpressed, whereas miR-24-3p was down-regulated in the CT-genotyped patients and cells compared with those in the CC-genotyped patients and cells. The survival of colon cancer cells was decreased, whereas the apoptosis of colon cancer cells was increased by PTX treatment in a dose-dependent manner. MiR-24-3p was validated to target lncRNA-CCAT1 and FSCN1 mRNA, and the overexpression of CCAT1 could reduce the expression of miR-24-3p although elevating the expression of FSCN1. Knockdown of lncRNA-CCAT1 partly reversed the suppressed growth of CT-genotyped tumours. And the knockdown of lncRNA-CCAT1 partly reversed the dysregulation of lncRNA-CCAT1 and FSCN1 mRNA/protein in rs67085638-CT + NC shRNA mice. The findings of this study demonstrated that the presence of the minor allele of rs67085638 increased the expression of CCAT1 and accordingly enhanced the resistance to PTX. Down-regulation of CCAT1 significantly re-stored the sensitivity to PTX of colon cancer cells.