lncRNA Colon Cancer-associated Transcript Research Articles

Evaluation of tumor-educated platelet long non-coding RNAs (lncRNAs) as potential diagnostic biomarkers for colorectal cancer.

Cancer-derived circulating components are increasingly considered as candidate sources for non-invasive diagnostic biomarkers. This study aimed to investigate the expression of tumor-educated platelet (TEP) long non-coding RNAs (lncRNAs) in colorectal cancer (CRC) patients and determine whether it could be served as a potential tool for CRC diagnosis. Relative quantitative real-time PCR (qRT-PCR) was used to detect the expression levels of three cancer-related platelet-derived lncRNAs CCAT1, HOTTIP, and XIST in 75 CRC patients and 42 healthy controls. Quantitative data were analyzed by SPSS (IBM Corp., Armonk, NY, USA) for comparison of cancer and non-cancer individuals. The receiver operating characteristic (ROC) curve analysis was further performed to assess the diagnostic values of lncRNAs within the CRC patients. The expression levels of lncRNAs colon cancer associated transcript 1 (CCAT1) (P = 0.006) and HOXA transcript at the distal tip (HOTTIP) (P = 0.049), but not X-inactive specific transcript (XIST) (P = 0.12), were significantly upregulated in CRC patients compared to healthy individuals. However, there were no significant correlations between platelet lncRNAs and clinicopathological characteristics, including sex, age, tumor location, differentiation, and size (all at P > 0.05). The area under the ROC curve (AUC) of the lncRNA CCAT1 was 0.61 (sensitivity, 71%; specificity, 50%). TEP lncRNA CCAT1 is detectable in the circulation of CRC patients and could be considered as a potential diagnostic biomarker.

LncRNA CCAT1 participates in pancreatic ductal adenocarcinoma progression by forming a positive feedback loop with c-Myc.

Long noncoding RNAs (lncRNAs) play fundamental roles in cancer development; however, the underlying mechanisms for a large proportion of lncRNAs in pancreatic ductal adenocarcinoma (PDAC) have not been elucidated. The expression of colon cancer-associated transcript-1 (CCAT1) in PDAC specimens and cell lines was measured by quantitative real-time polymerase chain reaction (qRT-PCR). The function of CCAT1 was examined in vitro and in vivo. The interactions among CCAT1, miR-24-3p and c-Myc were determined by bioinformatics analysis, RNA immunoprecipitation (RIP), dual-luciferase reporter assay, and rescue experiments. CCAT1 was significantly increased in PDAC, positively correlated with PDAC progression and predicted a worse prognosis. Furthermore, CCAT1 enhanced Adenosine triphosphate (ATP) production to facilitate PDAC cell proliferation, colony formation and motility in vitro and tumor growth in vivo. CCAT1 may serve as an miR-24-3p sponge, thereby counteracting its repression by c-Myc expression. Reciprocally, c-Myc may act as a transcription factor to alter CCAT1 expression by directly targeting its promoter region, thus forming a positive feedback loop with CCAT1. Collectively, these results demonstrate that a positive feedback loop of CCAT1/miR-24-3p/c-Myc is involved in PDAC development, which may serve as a biomarker and therapeutic target for PDAC.

Abstract 1486: The regulatory mechanism of LncRNA CCAT1 via three-dimensional genome structure in colorectal cancer cells

Abstract Kruppel-like factor 5 (KLF5) belongs to zinc finger transcription factors and its expression is up-regulated in specific cancer types, such as gastrointestinal cancer and squamous carcinoma. It is reported that KLF5 is associated with the maintenance of stemness and essential for the integrity of intestinal epithelium and oncogenesis. Colon Cancer Associated Transcript 1 (CCAT1) is a long noncoding RNA that was initially found to be up-regulated in colorectal cancer (CRC), and studies have reported that high CCAT1 expression is associated with poor prognosis in CRC patients. In this study, we performed in vitro enChIP (engineered DNA-binding molecule-mediated chromatin immunoprecipitation)-sequencing to identify the KLF5 enhancer region which interacts with KLF5 promoter region. We successfully identified the KLF5 enhancer region in colorectal cancer cells and further found that KLF5 promoter region interacted with the transcription start site (TSS) region of CCAT1 gene, which is located at the different chromosome. Our results indicated that KLF5 protein participated in the core regulatory circuitry together with co-factors (BRD4, MED1, and RAD21), which contributes to construction of the three-dimensional genome structures consisting of KLF5 promoter, enhancer and CCAT1 TSS region. We also found that CCAT1 expression was highly correlated with KLF5 expression in CRC clinical samples and that KLF5 regulated CCAT1 expression. In conclusion, our data propose the mechanistic insight that KLF5 protein constructs the core regulatory circuitry with co-factors in the three-dimensional genome structure and coordinately regulates KLF5 and CCAT1 expression in CRC. Citation Format: Yuhki Yokoyama, Hirofumi Yamamoto. The regulatory mechanism of LncRNA CCAT1 via three-dimensional genome structure in colorectal cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1486.

LncRNA CCAT1 enhances chemoresistance in hepatocellular carcinoma by targeting QKI-5

A major reason for treatment failure of cancer is acquisition of drug resistance. The specific mechanisms underlying hepatocellular carcinoma (HCC) chemoresistance need to be fully elucidated. lncRNAs involve in drug resistance in some cancers, however, the exact functions of lncRNA colon cancer-associated transcript 1 (CCAT1) in oxaliplatin resistance in HCC are still unknown. Our study indicated that CCAT1 promoted HCC proliferation and reduced the apoptosis induced by oxaliplatin. Knockout of CCAT1 could increased chemosensitivity in vitro and in vivo. Further study found that QKI-5 was an important mediator and blocking of QKI-5/p38 MAPK signaling pathway could enhance oxaliplatin sensitivity. In conclusions, CCAT1 promoted proliferation and oxaliplatin resistance via QKI-5/p38 MAPK signaling pathway in HCC. Targeting CCAT1 in combination with chemotherapeutics may be a promising alternative to reverse drug resistance in HCC treatment.

LncRNA-CCAT1/miR-152-3p is involved in CSE-induced inflammation in HBE cells via regulating ERK signaling pathway

Emerging studies have noted that dysregulated long non-coding RNAs (lncRNAs) are implicated in the pathological processes of chronic obstructive pulmonary disease (COPD). LncRNA colon cancer-associated transcript 1 (CCAT1) plays well-defined roles in the inflammatory progression. The study aims to figure out the effect and regulatory mechanism of CCAT1 in the cigarette smoke induced inflammation in COPD. The results showed that CCAT1 was highly expressed in lung tissues of smokers with COPD compared with never-smokers without COPD. In human bronchial epithelial (HBE) cells, cigarette smoke extract (CSE) treatment led to an increase in CCAT1 expression in a dose- and time- dependent manner. Functional experiments showed that knockdown of CCAT1 amelioratedCSE-inducedinflammation. Mechanistically, CCAT1 directly targeted miR-152-3p, and miR-152-3p overexpression reversed the pro-inflammatory effects of CCAT1 on HBE cells. Subsequently, miR-152-3p was found to regulate ERK signaling pathway. PD98059, an ERK specific inhibitor, reversed miR-152-3p knockdown mediated inflammation in HBE cells. In addition, CCAT1 acted as a sponge for miR-152-3p to positively regulate ERK signaling pathway. Overall, current findings suggest that CCAT1 promoted inflammation by activating ERK signal pathway via sponging miR–152–3p in CSE–treated HBE cells. These results may provide a novel therapeutic target for alleviating cigarette smoke mediated airway inflammation.

Dexmedetomidine protects H9C2 rat cardiomyocytes against hypoxia/reoxygenation injury by regulating the long non-coding RNA colon cancer-associated transcript 1/microRNA-8063/Wnt/β-catenin axis

ABSTRACT Dexmedetomidine (Dex) protects the heart from ischemia/reperfusion (I/R) injury. The differential expression of long non-coding RNAs (lncRNAs) is associated with myocardial injury, but whether the lncRNA colon cancer-associated transcript 1 (CCAT1) is associated with Dex-mediated myocardial protection remains unclear. In this study, a hypoxia/reoxygenation (H/R) H9C2 model was established to simulate the in vitro characteristics of I/R. CCAT1 and microRNA (miR)-8063 expression levels in H/R H9C2 cells pretreated with Dex were determined via quantitative reverse transcription-polymerase chain reaction. The survival and apoptotic rates of H9C2 cells were determined via cell counting kit-8 and flow cytometry assays. Wnt3a, Wnt5a, and β-catenin protein levels were measured via western blotting. Luciferase and RNA immunoprecipitation assays were used to explore the binding relationship between miR-8063 and CCAT1. Dex pretreatment increased H/R H9C2 cell viability and CCAT1 expression, while decreasing the cell apoptosis and Wnt3a, Wnt5a, and β-catenin protein levels. Knockdown of CCAT1 abolished the protective effects of Dex on H/R H9C2 cells, and the downregulation of miR-8063 expression eliminated the effect of CCAT1 knockdown. These results revealed that CCAT1, a sponge for miR-8063, is involved in Dex-mediated H9C2 cell H/R injury by negatively targeting miR-8063 and inactivating the Wnt/β-catenin pathway. Dex protects H9C2 cells from H/R impairment by regulating the lncRNA CCAT1/miR-8063/Wnt/β-catenin axis.

LncRNA CCAT2 expression at diagnosis predicts imatinib response in chronic phase chronic myeloid leukemia patients

Long noncoding RNAs (lncRNAs) are identified as key players in the initiation, development, and prognosis of chronic myeloid leukemia (CML). Some lncRNAs are expected to serve as diagnostic biomarkers, predictors of clinical outcomes and therapeutic targets. We aimed to examine the expression of lncRNA colon cancer-associated transcript 2 (CCAT2) in CML patients, as well as to correlate CCAT2 expression with response to imatinib therapy. 43 newly diagnosed patients with chronic phase CML were included, and 30 healthy persons were selected as controls. Real-time reverse transcription PCR was performed to analyze the expression of CCAT2 in peripheral blood mononuclear cells. Our results reported for the first time the upregulated expression of CCAT2 in CML patients as compared with controls (P < 0.001). We demonstrated significant association between CCAT2 expression and therapy response at 3 months, and at 6 months (P = 0.004, and P = 0.005; respectively). Moreover, CCAT2 expression was significantly associated with spleen size (P = 0.006) and EUTOS sore (P = 0.030). LncRNA CCAT2 is highly expressed in the peripheral blood of CML patients, and the enhanced expression at diagnosis is linked to imatinib resistance. CCAT2 is expected to become a reliable molecular marker for predicting imatinib response in chronic phase CML patients.

Serum CCAT2 as a biomarker for adjuvant diagnosis and prognostic prediction of cervical cancer

Growing evidence indicates that lncRNA colon cancer-associated transcript 2 (CCAT2) is associated with cancers. However, the clinical value of CCAT2 in cervical cancer (CC) remains unclear. In this study, serum CCAT2 level was detected by real-time quantitative PCR (RT-qPCR). Carbohydrate antigen 125 (CA125) and squamous-cell carcinoma antigen (SCC) were detected by electrochemiluminescence. A receiver operating characteristic (ROC) curve was utilized to estimate the diagnostic efficiency of CCAT2. Kaplan-Meier survival analysis and univariable and multivariable analyses were performed to assess the prognostic value of CCAT2. The relative expression level of CCAT2 in primary CC patients was significantly higher than that in cervical intraepithelial neoplasias (CIN) patients and healthy controls (both P < 0.001). CCAT2 relative expression was positively correlated with tumor Federation of Gynecology and Obstetrics (FIGO) stage, SCC-Ag and lymph node metastasis (LNM) (all P < 0.05). CCAT2 expression in recurrent/metastatic CC was significantly higher compared with primary CC (P < 0.0001) or operated CC (P < 0.0001) and during follow-up, CCAT2 expression was increased before surgery and decreased significantly after surgery (P < 0.0001). Furthermore, the overall survival rate of CC patients with high CCAT2 expression group markedly decreased as compared with that of low CCAT2 expression group (P = 0.026). Univariate analyses indicated that CCAT2 was a poor prognostic factor associated with overall survival (OS). Our study indicates that CCAT2 may be valuable in complementary diagnosis and monitoring of progression and prognosis of CC patients. Combined detection of CCAT2, CA125 and SCC can greatly improve the diagnostic efficiency of primary CC.

Self-assembled fluorescent hybrid nanoparticles-mediated collaborative lncRNA CCAT1 silencing and curcumin delivery for synchronous colorectal cancer theranostics

BackgroundCancer synergistic therapy strategy in combination with therapeutic gene and small molecule drug offers the possibility to amplify anticancer efficiency. Colon cancer-associated transcript-1 (CCAT1) is a well identified oncogenic long noncoding RNA (lncRNA) exerting tumorigenic effects in a variety of cancers including colorectal cancer (CRC).ResultsIn the present work, curcumin (Cur) and small interfering RNA targeting lncRNA CCAT1(siCCAT1) were co-incorporated into polymeric hybrid nanoparticles (CSNP), which was constructed by self-assembling method with two amphiphilic copolymers, polyethyleneimine-poly (d, l-lactide) (PEI-PDLLA) and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy (polyethylene glycol) (DSPE-mPEG). Owing to the multicolor fluorescence characteristics of PEI-PDLLA, the constructed CSNP could be served as a theranostic nanomedicine for synchronous therapy and imaging both in vitro and in vivo. Resultantly, proliferation and migration of HT-29 cells were efficiently inhibited, and the highest apoptosis ratio was induced by CSNP with coordination patterns. Effective knockdown of lncRNA CCAT1 and concurrent regulation of relevant downstream genes could be observed. Furthermore, CSNP triggered conspicuous anti-tumor efficacy in the HT-29 subcutaneous xenografts model with good biosafety and biocompatibility during the treatment.ConclusionOn the whole, our studies demonstrated that the collaborative lncRNA CCAT1 silencing and Cur delivery based on CSNP might emerge as a preferable and promising strategy for synergetic anti-CRC therapy.Graphic abstract

LncRNA CCAT1 sponges miR-218-5p to promote EMT, cellular migration and invasion of retinoblastoma by targeting MTF2

Retinoblastoma (RB) is the primary neoplasms of the retina that is most common in pediatrics age. Long non-coding RNAs (lncRNAs) has been noticed for strong relation to the occurrence and progress of retinoblastoma. Previously, we have demonstrated that lncRNA colon cancer-associated transcript 1 (CCAT1) in two RB cell lines SO-RB50 and Y79 was obviously overexpressed, and notably, lncRNA CCAT1 attenuated miR-218-5p expressionand induced proliferation, cell migration and invasion. But, how lncRNA CCAT1 acts in RB development and the potential molecular mechanisms remain to be determined. In this study, the expression levels of lncRNA CCAT1 and miR-218-5p were evaluated in RB tissues by Q-PCR, which established the results in the cell lines. Further, lncRNA CCAT1 was shown to promote epithelial-to-mesenchymal transition (EMT), cellular migration and invasion of RB cells by functional analysis of downregulation and overexpression of lncRNA CCAT1 with specific siRNA and pcDNA transfection. By performing bioinformatics and dual luciferase reporter assay, we verified the direct interaction between lncRNA CCAT1 and miR-218-5p. Besides, bioinformatics analysis indicated that metal regulatory transcription factor 2 (MTF2) might be a potent novel target for miR-218-5p, which was further validated with luciferase reporter assay, Q-PCR and also Western blot analysis. Functional analysis and rescue analysis showed that lncRNA CCAT1 via competitive binding to miR-218-5p to modulate MTF2 expression thus accelerate EMT, cell migration and invasion of RB. In conclusion, here we identified the lncRNA CCAT1/miR-218-5p/MTF2 axis in RB cell lines. Our investigations on the function of lncRNA CCAT1 and the roles of the related molecules hint a novel potential target fo RB therapy.

Clinical value of lncRNA CCAT1 in serum extracellular vesicles as a potential biomarker for gastric cancer.

Long non-coding RNAs (lncRNAs) in extracellular vesicles (EVs) are considered to be novel non-invasive biomarkers for gastric cancer (GC). lncRNA colon cancer-associated transcript 1 (CCAT1) is aberrantly expressed in certain types of cancer. However, the role of EV lncRNA CCAT1 in patients with GC remains unclear. The current study aimed to assess the expression levels of lncRNA CCAT1 in the serum EVs of patients with GC and evaluate its potential clinical value. EVs were isolated from serum using a commercial kit and ultracentrifugation, and were identified by transmission electron microscopy, nanoparticle tracking analysis and western blotting. Serum EV lncRNA CCAT1 levels in patients with GC, chronic gastritis or atypical hyperplasia and healthy control subjects were detected by reverse transcription-quantitative PCR. Additionally, lncRNA CCAT1 was detected in GC and adjacent non-cancerous tissue samples. Serum EVs were successfully isolated and identified in all patients. The results revealed that serum EV lncRNA CCAT1 levels in patients with GC were significantly higher compared with those in healthy controls, patients with chronic gastritis or atypical hyperplasia (all P<0.05). Additionally, EV lncRNA CCAT1 expression levels were significantly different among various groups based on the depth of invasion, distant metastasis and the Tumor-Node-Metastasis stage. The area under the curve (AUC) value of EV lncRNA CCAT1 was 0.890 [95% confidence interval (CI), 0.826–0.937] with 79.6% sensitivity and 92.6% specificity. The combination of EV lncRNA CCAT1 and carcinoembryonic antibody produced an AUC value of 0.910 (95% CI, 0.849–0.951) with the sensitivity and specificity of 80.5 and 92.6%, respectively. In addition, lncRNA CCAT1 was determined to be stable in serum EVs. The expression levels of lncRNA CCAT1 in GC tissue were positively correlated with those in serum EVs, and high levels of lncRNA CCAT1 were associated with a low disease-free survival rate in patients with GC. The results of the present study demonstrated that serum EV lncRNA CCAT1 levels were upregulated in patients with GC compared with those healthy subjects and patients with other illnesses, and may therefore be used as a novel biomarker for this type of cancer.

Inhibiting proliferation and metastasis of osteosarcoma cells by downregulation of long non-coding RNA colon cancer-associated transcript 2 targeting microRNA-143.

Osteosarcoma is a malignant bone tumor, which has a high incidence in children and adolescents. However, the pathogenesis of osteosarcoma remains unclear. Long noncoding RNA (lncRNA) is a new potential therapeutic target and diagnostic biomarker for osteosarcoma. Hence, the present study aimed to explore the effect of lncRNA colon cancer-associated transcript (CCAT2) on osteosarcoma and its potential underlying mechanisms. For this purpose, the proliferation of osteosarcoma cells was measured using the CCK-8 assay. The scratch-wound and cell invasion assays were used to determine the migration and invasion of osteosarcoma cells, respectively. LncRNA CCAT2 and microRNA (miR)-143 binding sites were identified by the dual-luciferase reporter assay. RNA and protein expression levels were detected by reverse-transcription quantitative PCR and western blotting, respectively. Downregulation of lncRNA CCAT2 inhibited the proliferation, migration, and invasion of osteosarcoma cells. The findings also revealed that miR-143 bound directly to lncRNA CCAT2. The expression of miR-143 was upregulated by the knockdown of lncRNA CCAT2. Downregulation of the FOS-like antigen 2 was also observed after knockdown of lncRNA CCAT2. The function of lncRNA CCAT2 in osteosarcoma cells was attenuated by co-transfection with anti-miR-143 oligodeoxyribonucleotide. In conclusion, downregulation of lncRNA CCAT2 inhibited the proliferation and metastasis of osteosarcoma cells by targeting miR-143. lncRNA CCAT2 was identified as a potential target for osteosarcoma treatment.

Long Non-coding RNA CCAT1 Acts as an Oncogene and Promotes Sunitinib Resistance in Renal Cell Carcinoma.

Although sunitinib contributes to prolonging the progression-free survival of metastatic renal cell carcinoma significantly, the universal presence of resistance limits the initial response rate and restricts durable responses. The mechanisms involved in sunitinib resistance vary and need further investigation. We found long non-coding RNA (lncRNA) colon cancer-associated transcript-1 (CCAT1) overexpressed in sunitinib-resistant cells while declined in the parental cells. Moreover, lncRNA CCAT1 increased significantly in samples with resistance to sunitinib compared with those with responses to sunitinib. The reduction of CCAT1 suppressed cell growth and colony formation while triggering apoptosis. Inversely, the ectopic expression of c-Myc reversed the inhibition of cell growth and enhancement of apoptosis by the knockdown of CCAT1. We also verified that anti-apoptosis protein B-cell lymphoma 2 (Bcl-2) and myeloid cell leukemia 1 (Mcl-1) decreased along with the deregulation of CCAT1, whereas the expression of Bcl-2 and Mcl-1 restored in cells that were transfected sh-CCAT1 and c-Myc simultaneously. Apart from the in vitro experiments, we demonstrated that knockdown of CCAT1 boosted response to sunitinib by performing sunitinib-resistant ACHN mouse models. Briefly, lncRNA CCAT1 conferred renal cell carcinoma resistance to sunitinib in a c-Myc-dependent manner, providing a novel target for improvement of sunitinib therapy.

Long Noncoding RNA CCAT1 Functions as a Competing Endogenous RNA to Upregulate ITGA9 by Sponging MiR-296-3p in Melanoma.

BackgroundMelanoma is aggressive and lethal melanocytic neoplasm, and its incidence has increased worldwide in recent decades. Accumulating evidence has showed that various long noncoding RNAs (lncRNAs) participated in occurrence of malignant tumors, including melanoma. The present study was designed to investigate function of lncRNA colon cancer-associated transcript-1 (CCAT1) in melanoma.MethodsThe expression levels of CCAT1, miR-296-3p and Integrin alpha9 (ITGA9) in melanoma tissues or cells were measured using real-time quantitative polymerase chain reaction (RT-qPCR). The concentrations of glucose and lactate were measured for assessing glycolysis of melanoma cells. 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazol-3-ium bromide (MTT), flow cytometry, and transwell assays were conducted to assess proliferation, apoptosis, and migration of melanoma cells. Western blot assay was performed to measure the protein expression of ITGA9, hexokinase 2 (HK2), and epithelial–mesenchymal transition (EMT)-related proteins in melanoma tissues or cells. The relationship among CCAT1, miR-296-3p, and ITGA9 was predicted and confirmed by bioinformatics analysis, dual-luciferase reporter, and RNA immunoprecipitation (RIP) assay, respectively. A xenograft experiment was established to assess the effect of CCAT1 knockdown in vivo.ResultsCCAT1 was effectively increased in melanoma tissues and cells compared with matched controls, and deficiency of CCAT1 impeded cell glycolysis, proliferation, migration while induced apoptosis, which were abrogated by knockdown of miR-296-3p in melanoma cells. In addition, our findings revealed that ITGA9 overexpression abolished miR-296-3p overexpression-induced effects on melanoma cells. Importantly, CCAT1 regulated ITGA9 expression by sponging miR-296-3p. The results of xenograft experiment suggested that CCAT1 silencing inhibited melanoma cell growth in vivo.ConclusionLncRNA CCAT1 promoted ITGA9 expression by sponging miR-296-3p in melanoma.

Long non-coding RNA CCAT1 is overexpressed in endometrial cancer and regulates growth and transcriptome of endometrial adenocarcinoma cells.

Long non-coding RNAs (lncRNAs) play important roles in regulation of gene expression and are involved in pathogenesis of different diseases including cancer. Recent studies suggested the lncRNA Colon cancer associated transcript-1 (CCAT1) to act as putative oncogene. In this study, to elucidate the role of this lncRNA in endometrial cancer, we examined its expression in normal endometrium and type 1 endometrial cancer and knocked down its expression in endometrial cancer cell lines followed by transcriptome and pathway analyses. CCAT1 expression was examined in 100 tissue samples of normal endometrium and type 1 endometrial cancer tissues by means of RT-qPCR. Knockdown of CCAT1 expression in HEC-1B and RL95/2 endometrial cancer cells was performed by siRNA transfection. Affymetrix GeneChip arrays were used to elucidate the effect of both lncRNAs on the transcriptome of these cell lines. Median CCAT1 expression was found to be 9.3-fold higher in endometrial cancer when compared to normal endometrium (p < 0.05). In contrast to premenopausal endometrium and G1, G2 and G3 graded endometrial cancer, CCAT1 expression was nearly absent in postmenopausal tissue. Knockdown of CCAT1 by transient siRNA transfection significantly reduced proliferation of HEC-1B cancer cells in vitro by 35.5 % 6 days after transfection and notably reduced their colony formation ability. Affymetrix microarray and Ingenuity pathway analyses revealed a set of up- or down-regulated genes in transfected ERα-negative HEC-1B cells forming a network controlled by the key regulators TNF and TP53, including genes known to be involved in growth control, providing putative molecular mechanisms underlying the observed growth inhibition of HEC-1B cells. In contrast, CCAT1 knockdown in ERα-positive RL95/2 cells did not significantly affect proliferation, but resulted in down-regulation of a network of ERα target genes. Given that the lncRNA CCAT1 was found to be overexpressed in endometrial cancer, affected the growth of HEC-1B cells and the expression of growth regulatory genes, our data suggest CCAT1 to exert oncogenic functions in endometrial cancer and encourage further studies to examine to what extent this lncRNA might be a potential therapy target in this cancer entity.

Knockdown of lncRNA CCAT1 enhances sensitivity of paclitaxel in prostate cancer via regulating miR-24-3p and FSCN1

ABSTRACTDrug resistance limits the efficacy of chemotherapy in human cancers. Previous studies reported that long noncoding RNA colon cancer-associated transcript 1 (CCAT1) regulated progression of prostate cancer (PCa). However, the potential role of CCAT1 in the sensitivity of paclitaxel (PTX) in PCa and its mechanism remain largely unknown. The PTX-resistant PCa cells were established in PC3 and DU145 cells by increasing concentrations of PTX. The expressions of CCAT1, microRNA-24-3p (miR-24-3p) and fascin1 (FSCN1) were measured by quantitative real-time polymerase chain reaction. The viability and apoptosis were detected by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide assay, flow cytometry and western blot, respectively. The interaction among CCAT1, miR-24-3p and FSCN1 was explored by luciferase activity, RNA immunoprecipitation, RNA pull-down and western blot, respectively. Results showed that the expressions of CCAT1 were up-regulated and miR-24-3p was down-regulated in PCa and PTX-resistant PCa cells (PC3-TXR and DU145-TXR). Knockdown of CCAT1 or overexpression of miR-24-3p inhibited survival rate, half maximal inhibitory concentration (IC50) of PTX but increased apoptosis in PC3-TXR and DU145-TXR cells after treatment of PTX. miR-24-3p was bound to CCAT1 and its abrogation reversed knockdown of CCAT1-mediated increase of PTX sensitivity in PC3-TXR and DU145-TXR cells. Moreover, FSCN1 restoration attenuated miR-24-3p-mediated inhibition of PTX resistance. Besides, FSCN1 level was enhanced in PCa and PTX-resistant PCa cells and regulated by CCAT1 and miR-24-3p. Our data suggested interference of CCAT1 contributed to PTX sensitivity in PCa by regulating miR-24-3p and FSCN1, indicating a novel avenue for treatment of PCa through regulating chemoresistance.

LncRNA CCAT1 Acts as a MicroRNA-218 Sponge to Increase Gefitinib Resistance in NSCLC by Targeting HOXA1

Long non-coding RNA (lncRNA) colon cancer-associated transcript-1 (CCAT1) has been reported to play important roles in the development and progression of multiple human malignancies. However, the functional role and molecular mechanism of CCAT1 on gefitinib resistance in non-small cell lung cancer (NSCLC) are largely unclear. The aim of this study is to explore the roles of CCAT1 on gefitinib resistance in NSCLC and to explore the underlying mechanisms. The quantitative real-time PCR (qRT-PCR) analysis was to investigate the expression pattern of CCAT1 in gefitinib-resistant NSCLC patient tissues and cell lines, and then the effects of CCAT1 on gefitinib resistance of NSCLC in vitro and in vivo. Furthermore, bioinformatics online program predictions and luciferase reporter assay were used to validate the association of CCAT1 and miR-218 in NSCLC cells. In this study, CCAT1 was observed to be upregulated in gefitinib-resistant patient tissues and cell lines. In vitro and in vivo experiments demonstrated that CCAT1 knockdown impaired cell proliferation and promoted the gefitinib-induced cell apoptosis. Furthermore, we demonstrated that CCAT1 acts as a sponge for miR-218, and verified that HOXA1 is a novel target of miR-218. These results suggest that CCAT1 may serve as a promising therapeutic target for the treatment of epidermal growth factor receptor (EGFR) plus NSCLC patients.

The regulation and interaction of colon cancer-associated transcript-1 and miR7-5p contribute to the inhibition of SP1 expression by solamargine in human nasopharyngeal carcinoma cells.

Nasopharyngeal carcinoma (NPC) is a common head and neck malignancy with higher incidence in Southern China and Southeast Asia. Solamargine (SM), a steroidal alkaloid glycoside, has been shown to have anticancer properties. However, the underlying mechanism involved remains undetermined. In this study, we showed that SM inhibited the growth of NPC cells. Mechanistically, we found that solamargine decreased lncRNA colon cancer‐associated transcript‐1 (CCAT1) and increased miR7‐5p expression. There was a reciprocal interaction of CCAT1 and miR7‐5p. In addition, SM inhibited the expression of SP1 protein and promoter activity, which was strengthened by miR7‐5p mimics and inhibited by overexpressed CCAT1. MiR7‐5p could bind to 3'‐UTR of SP1 and attenuated SP1 gene expression. Exogenously expressed SP1 feedback resisted SM‐increased miR7‐5p expression and more importantly reversed SM‐inhibited growth of NPC cells. Finally, SM inhibited NPC tumor growth in vivo. Collectively, our results show that SM inhibits the growth of NPC cells through reciprocal regulation of CCAT1 and miR7‐5p, followed by inhibition of SP1 gene expression in vitro and in vivo. The interregulation and correlation among CCAT1, miR7‐5p and SP1, and the feedback regulatory loop unveil the novel molecular mechanism underlying the overall responses of SM in anti‐NPC.

LncRNA CCAT1 promotes autophagy via regulating ATG7 by sponging miR-181 in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is a significant clinical challenge, and the mechanisms underlying HCC pathogenesis remain incompletely understood. Colon cancer associated transcript 1 (CCAT1), is one novel long noncoding RNA (lncRNA) which is upregulated in HCC. Autophagy is a vital process in HCC progression, and it is unknown whether CCAT1 regulates autophagy in HCC. Immunofluorescence staining and transmission electron microscopy were used to analyze autophagy activity. Luciferase assay was performed to confirm miRNA-181a-5p (miR-181a-5p) bind CCAT1 and ATG7. CCAT1 levels were higher in tissue and cell lines of HCC. In function research, we found that CCAT1 facilitates HCC cell autophagy and cell proliferation. Our results show that, mechanistically, CCAT1 promotes autophagy through functioning as a sponge for miR-181a-5p, and then regulating ATG7 expression. Our findings indicate CCAT1 may play a role in regulating autophagy by sponging miR-181a-5p in HCC.

LncRNA CCAT1 promotes bladder cancer cell proliferation, migration and invasion.

ABSTRACTObjective:To study the expression patterns of long noncoding RNA (lncRNA) colon cancer-associated transcript 1 (CCAT1) and the changes in cell proliferation, apoptosis, migration and invasion induced by silencing CCAT1 in bladder cancer cells.Materials and Methods:The expression levels of CCAT1 were determined using realtime quantitative polymerase chain reaction in cancerous tissues and paired normal tissues from 34 patients with bladder cancer. The relationship between clinical characteristics and CCAT1 expression was analyzed. And then we conducted cell experiments. Bladder urothelial carcinoma cell lines T24 and 5637 cells were transfected with CCAT1 small interfering RNA (siRNA) or scramble siRNA. Cell proliferation and apoptosis changes were determined using a Cell Counting Kit-8 (CCK-8) assay and a flow cytometry assay. Migration and invasion changes were measured using a wound healing assay and a trans-well assay. microRNAs (miRNAs) were predicted by Starbase 2.0, and their differential expression levels were studied.Results:CCAT1 was significantly upregulated in bladder cancer (P < 0.05). CCAT1 upregulation was positively related to tumor stage (P = 0.004), tumor grade (P = 0.001) and tumor size (P = 0.042). Cell proliferation, migration and invasion were promoted by abnormally expressed CCAT1. miRNAs miR-181b-5p, miR-152-3p, miR-24-3p, miR-148a-3p and miR-490-3p were potentially related to the aforementioned functions of CCAT1.Conclusion:CCAT1 plays an oncogenic role in urothelial carcinoma of the bladder. In addition, CCAT1 may be a potential therapeutic target in this cancer.