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1.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 37(9): 769-774, 2021 Sep.
Artigo em Chinês | MEDLINE | ID: mdl-34533122

RESUMO

Objective To investigate the effects of nucleotide binding oligomerization domain-like receptor family caspase recruitment domain containing 3 (NLRC3) on the proliferation, migration and invasion of human colon cancer HCT116 and LoVo cells. Methods NLRC3 was knocked down in HCT116 and LoVo cells by NLRC3-specific siRNA (si-NLRC3). NLRC3 mRNA and protein expression was detected by real-time quantitative PCR and Western blotting. The proliferation ability of cancer cells was detected by CCK-8 assay; the clone formation ability was detected by clone formation assay; the invasion ability was detected by TranswellTM assay; the migration ability was detected by cell scratch healing assay. Results The transfection of si-NLRC3 down-regulated the expression of NLRC3 in HCT116 and LoVo cells. After NLRC3 knockdown, the proliferation and invasion ability of colon cancer cells were significantly strengthened and the cell migration was not significantly changed. Conclusion Knockdown of NLRC3 in HCT116 and LoVo cells can enhance cell proliferation and invasion ability, but has no effects on cancer cell migration.


Assuntos
Neoplasias do Colo , Movimento Celular/genética , Proliferação de Células/genética , Neoplasias do Colo/genética , Humanos , Peptídeos e Proteínas de Sinalização Intercelular , RNA Mensageiro , RNA Interferente Pequeno/genética
2.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 37(9): 808-814, 2021 Sep.
Artigo em Chinês | MEDLINE | ID: mdl-34533128

RESUMO

Objective To investigate the effect of lysine-specific demethylase 3A (KDM3A) on the invasion and migration of MDA-MB-231 breast cancer cells. Methods The mRNA and the protein expressions of KDM3A in MDA-MB-231 breast cancer cells and MCF-10A normal breast cells were detected by real-time quantitative PCR and Western blotting, respectively; the KDM3A level of MDA-MB-231 cells was knocked down by lentivirus infection of KDM3A short hairpin RNA (shKDM3A). The change of invasion and migration ability of MDA-MB-231 cells was detected by TranswellTM assay, and the change in the cell cycle was detected by flow cytometry. Results The expression of KDM3A in MDA-MB-231 breast cancer cells was significantly increased compared with that in MCF-10A epithelial cells; after KDM3A knockdown, the invasion and migration abilities of MDA-MB-231 cells were significantly decreased, and the cell cycle was arrested in the G0/G1 phase. Conclusion Knockdown of KDM3A inhibits the invasion and migration of MDA-MB-231 breast cancer cells and arrests the cell cycle in G0/G1 phase.


Assuntos
Neoplasias da Mama , Lisina , Neoplasias da Mama/genética , Ciclo Celular/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Feminino , Fase G1/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Histona Desmetilases com o Domínio Jumonji/genética , Invasividade Neoplásica/genética
3.
J Transl Med ; 19(1): 387, 2021 09 09.
Artigo em Inglês | MEDLINE | ID: mdl-34503528

RESUMO

OBJECTIVE: The recurrence and metastasis of nasopharyngeal cancer (NPC) may be mainly attributed to the persistence of cancer stem cells (CSCs); however, the linkage mechanism has yet to be fully elucidated. METHODS: The levels of miR-4721, FOXA1, and Nanog expression in NPC were detected by in situ hybridization and immunohistochemistry. In vivo and in vitro metastasis assays confirmed miR-4721 promotes cell migration and invasion. Tumor spheroid formation assay, side population (SP) assay, and ALDEFLUOR assay verified miR-4721 regulates cancer stem cell-like properties. Luciferase reporter assay showed that miR-4721 directly regulates FOXA1 and FOXA1 effects the promoter activity of miR-4721 and Nanog. Chromatin immunoprecipitation (ChIP) analysis and electrophoresis mobility shift assay (EMSA) revealed that FOXA1 combined the promoter region of human miR-4721 and Nanog and the possible mechanism was also analyzed. RESULTS: In this study, a new mechanism of NPC tumorigenesis related to miR-4721 was verified. We found that miR-4721, FOXA1 and Nanog control their expressions through a negative feedback loop and then activate the downstream regulator of stem cell signaling to promote the enrichment and metastasis of NPC stem cells. CONCLUSION: These findings elucidate that the feedback loop of miR-4721/FOXA1/Nanog can regulate stemness and metastasis in NPC and may provide an experimental theoretical basis for metastasis and treatment resistance in NPC.


Assuntos
MicroRNAs , Neoplasias Nasofaríngeas , Linhagem Celular Tumoral , Movimento Celular/genética , Retroalimentação , Regulação Neoplásica da Expressão Gênica , Fator 3-alfa Nuclear de Hepatócito/genética , Humanos , MicroRNAs/genética , Proteína Homeobox Nanog/genética , Neoplasias Nasofaríngeas/genética , Recidiva Local de Neoplasia , Células-Tronco Neoplásicas
4.
Medicina (Kaunas) ; 57(7)2021 Jul 14.
Artigo em Inglês | MEDLINE | ID: mdl-34356991

RESUMO

Background and Objectives: Matrix metalloproteinases (MMP) have been implicated as major determinants of tumour growth and metastasis, which are considered two of the main hallmarks of cancer. The interaction of MMP8 and other signalling molecules within and adjacent tumoral tissues, including immune cells, are rather elusive, particularly of adenocarcinoma cell type. In this study, we aimed to investigate the role of MMP8 in non-small cell lung cancer proliferation and invasiveness potential. Materials and Methods: We individually lipofected with two different single guide RNA (sgRNAs) that specifically targeted on MMP8, with CRISPR-Cas 9 protein into the cells. Results: Our results clearly indicated that the lipofection of these complexes could lead to reduced ability of A549 cells to survive and proliferate to form colonies. In addition, when compared to non-transfected cells, the experimental cell groups receiving sgRNAs demonstrated relatively decreased migration rate, hence, wider wound gaps in scratch assay. The quantitative real time-polymerase chain reaction (qRT-PCR) demonstrated significant reduction in the MAP-K, survivin and PI3-K gene expression. MMP8 might have protective roles over tumour growth and spread in our body. Conclusions: The delivery of sgRNAs targeting on the MMP8 gene could induce tumour cell death and arrest cell migratory activity.


Assuntos
Adenocarcinoma de Pulmão , Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Adenocarcinoma de Pulmão/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Humanos , Neoplasias Pulmonares/genética , Metaloproteinase 8 da Matriz , Invasividade Neoplásica , RNA Guia
5.
Biomed Res Int ; 2021: 4959381, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34337014

RESUMO

Long noncoding RNAs (lncRNAs) play nonnegligible roles in the metastasis of non-small-cell lung cancer (NSCLC). This study is aimed at investigating the biological role of lncRNA OXCT1-AS1 in NSCLC metastasis and the underlying regulatory mechanisms. The expression profiles of lncRNA OXCT1-AS1 in different NSCLC cell lines were examined. Then, the biological function of lncRNA OXCT1-AS1 in NSCLC metastasis was explored by loss-of-function assays in vitro and in vivo. Further, the protective effect of lncRNA OXCT1-AS1 on lymphoid enhancer factor 1 (LEF1) was examined using RNA pull-down and RNA immunoprecipitation assays. Additionally, the role of LEF1 in NSCLC metastasis was investigated. Results indicated that lncRNA OXCT1-AS1 expression was significantly increased in NSCLC cell lines. Functional analysis revealed that knockdown of lncRNA OXCT1-AS1 impaired invasion and migration in vitro. Additionally, the ability of lncRNA OXCT1-AS1 to promote NSCLC metastasis was also confirmed in vivo. Mechanistically, through direct interaction, lncRNA OXCT1-AS1 maintained LEF1 stability by blocking NARF-mediated ubiquitination. Furthermore, LEF1 knockdown impaired invasion and migration of NSCLC in vitro and in vivo. Collectively, these data highlight the ability of lncRNA OXCT1-AS1 to promote NSCLC metastasis by stabilizing LEF1 and suggest that lncRNA OXCT1-AS1 represents a novel therapeutic target in NSCLC.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , RNA Longo não Codificante/metabolismo , Fatores de Transcrição TCF/metabolismo , Sequência de Aminoácidos , Animais , Linhagem Celular Tumoral , Movimento Celular/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Camundongos Nus , Modelos Biológicos , Invasividade Neoplásica , Metástase Neoplásica , Estabilidade Proteica , RNA Longo não Codificante/genética , Fatores de Transcrição TCF/química , Ubiquitinação , Regulação para Cima/genética
6.
Int J Mol Sci ; 22(15)2021 Jul 27.
Artigo em Inglês | MEDLINE | ID: mdl-34360757

RESUMO

Thyroid cancer is the most common endocrine malignancy, and the characterization of the genetic alterations in coding-genes that drive thyroid cancer are well consolidated in MAPK signaling. In the context of non-coding RNAs, microRNAs (miRNAs) are small non-coding RNAs that, when deregulated, cooperate to promote tumorigenesis by targeting mRNAs, many of which are proto-oncogenes and tumor suppressors. In thyroid cancer, miR-146b-5p is the most overexpressed miRNA associated with tumor aggressiveness and progression, while the antisense blocking of miR-146b-5p results in anti-tumoral effect. Therefore, inactivating miR-146b has been considered as a promising strategy in thyroid cancer therapy. Here, we applied the CRISPR/Cas9n editing system to target the MIR146B gene in an aggressive anaplastic thyroid cancer (ATC) cell line. For that, we designed two single-guide RNAs cloned into plasmids to direct Cas9 nickase (Cas9n) to the genomic region of the pre-mir-146b structure to target miR-146b-5p and miR-146b-3p sequences. In this plasmidial strategy, we cotransfected pSp-Cas9n-miR-146b-GuideA-puromycin and pSp-Cas9n-miR-146b-GuideB-GFP plasmids in KTC2 cells and selected the puromycin resistant + GFP positive clones (KTC2-Cl). As a result, we observed that the ATC cell line KTC2-Cl1 showed a 60% decrease in the expression of miR-146b-5p compared to the control, also showing reduced cell viability, migration, colony formation, and blockage of tumor development in immunocompromised mice. The analysis of the MIR146B edited sequence shows a 5 nt deletion in the miR-146b-5p region and a 1 nt deletion in the miR-146b-3p region in KTC2-Cl1. Thus, we developed an effective CRISPR/Cas9n system to edit the MIR146B miRNA gene and reduce miR-146b-5p expression which constitutes a potential molecular tool for the investigation of miRNAs function in thyroid cancer.


Assuntos
Sistemas CRISPR-Cas , Edição de Genes , Marcação de Genes , MicroRNAs , RNA Neoplásico , Carcinoma Anaplásico da Tireoide , Neoplasias da Glândula Tireoide , Animais , Linhagem Celular , Movimento Celular/genética , Sobrevivência Celular/genética , Xenoenxertos , Humanos , Camundongos , MicroRNAs/biossíntese , MicroRNAs/genética , Transplante de Neoplasias , RNA Neoplásico/biossíntese , RNA Neoplásico/genética , Carcinoma Anaplásico da Tireoide/genética , Carcinoma Anaplásico da Tireoide/metabolismo , Carcinoma Anaplásico da Tireoide/patologia , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/metabolismo , Neoplasias da Glândula Tireoide/patologia
7.
BMC Genomics ; 22(1): 613, 2021 Aug 12.
Artigo em Inglês | MEDLINE | ID: mdl-34384362

RESUMO

BACKGROUND: Hypertrophic scar (HTS) is a fibroproliferative skin disorder characterized by excessive cell proliferation, migration, and extracellular matrix (ECM) deposition. The CUB and Sushi multiple domains 1 (CSMD1) has previously been identified as the key regulatory gene of hypertrophic scar by a large sample GWAS study. However, further research has not yet been conducted to verify this finding in other HTS patients and to determine the underlying mechanism. RESULTS: In this study, we verified that CSMD1 was downregulated in both HTS tissue and HTS-derived fibroblasts. The knockdown of CSMD1 resulted in enhanced migration and fibronectin1 (FN1) secretion in fibroblasts in vitro. In addition, the upstream and downstream regulatory mechanisms of CSMD1 were also investigated through microRNA (miRNA) databases screening and RNA-sequencing (RNA-seq) respectively. The screening of four common microRNA (miRNA) databases suggested that miR-190a-3p binds to the CSMD1 and may regulate its expression. We confirmed that miR-190a-3p directly targeted the CSMD1-3'-UTR using luciferase reporter assays. Furthermore, the overexpression of miR-190a-3p showed promotion of migratory activity and FN1 secretion in fibroblasts, resembling the effect of CSMD1 knockdown; whereas the knockdown of miR-190a-3p exerted the opposite effect. Finally, transcriptomic analysis showed activation of Janus kinase-signal transducer and activator of transcription (JAK/STAT) signaling pathway in the CSMD1 knockdown fibroblasts. CONCLUSIONS: This study has validated the conclusions of the previous GWAS study conducted in Chinese population. In vitro experiments have provided further evidence on the function of CSMD1 in the development of HTS, and have also revealed the underlying upstream and downstream regulating mechanisms. Additionally, the JAK/STAT signaling pathway identified using RNA-seq might provide a potential treatment approach, especially for HTS.


Assuntos
Cicatriz Hipertrófica , MicroRNAs/metabolismo , Movimento Celular/genética , Proliferação de Células/genética , Cicatriz Hipertrófica/genética , Cicatriz Hipertrófica/patologia , Fibroblastos , Humanos , Proteínas de Membrana , MicroRNAs/genética , Proteínas Supressoras de Tumor
8.
Gene ; 805: 145904, 2021 Dec 30.
Artigo em Inglês | MEDLINE | ID: mdl-34418470

RESUMO

Breast cancer is the second most common cause of cancer-related mortality in women. Breast cancer metastasis which usually is observed at the last stage is the major cause of breast cancer-related death. Long non-coding RNAs (lncRNAs) are member of the non-coding RNA family. It is known that lncRNAs have important functions in the genes regulation of different processes and pathways such as EMT (Epithelial mesenchymal transition), metastasis and apoptosis. Therefore, it is inevitable that lncRNAs have potential contribution for the understanding of cancer pathogenesis. lncRNA-ZEB2NAT is the natural antisense transcript of ZEB2. Herein, we investigated the effects of lncRNA-ZEB2NAT on process of EMT, metastasis and apoptosis in MCF7 and MDA-MB-231 breast cancer cells. The effect of ZEB2NAT on the expression of important genes in EMT, metastasis and apoptosis, and some protein levels was determined by qRT-PCR and western blot analysis, respectively. The effects of ZEB2NAT on cell proliferation, apoptosis, invasion and colony formation were evaluated using XTT, annexin V, invasion and colony assays, respectively. The ZEB2NAT knockdown caused anti-metastatic and apoptotic effects. The ZEB2NAT knockdown resulted in a decrease in ZEB2 and N-cadherin but an increase in E-cadherin protein levels. In addition, it was determined that ZEB2NAT knockdown significantly decreased cell proliferation and stimulated apoptosis in both cells. It was found that ZEB2NAT knockdown significantly decreased invasion and colony formation in both cells. ZEB2NAT knockdown showed anti-metastatic and apoptotic effect by affecting the important genes in both cells. These results have suggested that ZEB2NAT has an important role in EMT, metastasis and apoptosis in breast cancer and ZEB2NAT knockdown caused significant anti-cancer activities.


Assuntos
Neoplasias da Mama/genética , Homeobox 2 de Ligação a E-box com Dedos de Zinco/genética , Apoptose/genética , Neoplasias da Mama/metabolismo , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Transição Epitelial-Mesenquimal/genética , Feminino , Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Invasividade Neoplásica/genética , Metástase Neoplásica/genética , RNA Longo não Codificante/genética , Homeobox 2 de Ligação a E-box com Dedos de Zinco/metabolismo
9.
Int J Mol Sci ; 22(16)2021 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-34445288

RESUMO

Inflammatory Breast Cancer (IBC) is an aggressive form of invasive breast cancer, highly metastatic, representing 2-4% of all breast cancer cases in the United States. Despite its rare nature, IBC is responsible for 7-10% of all breast cancer deaths, with a 5-year survival rate of 40%. Thus, targeted and effective therapies against IBC are needed. Here, we proposed Lipocalin-2 (LCN2)-a secreted glycoprotein aberrantly abundant in different cancers-as a plausible target for IBC. In immunoblotting, we observed higher LCN2 protein levels in IBC cells than non-IBC cells, where the LCN2 levels were almost undetectable. We assessed the biological effects of targeting LCN2 in IBC cells with small interference RNAs (siRNAs) and small molecule inhibitors. siRNA-mediated LCN2 silencing in IBC cells significantly reduced cell proliferation, viability, migration, and invasion. Furthermore, LCN2 silencing promoted apoptosis and arrested the cell cycle progression in the G0/G1 to S phase transition. We used in silico analysis with a library of 25,000 compounds to identify potential LCN2 inhibitors, and four out of sixteen selected compounds significantly decreased cell proliferation, cell viability, and the AKT phosphorylation levels in SUM149 cells. Moreover, ectopically expressing LCN2 MCF7 cells, treated with two potential LCN2 inhibitors (ZINC00784494 and ZINC00640089) showed a significant decrease in cell proliferation. Our findings suggest LCN2 as a promising target for IBC treatment using siRNA and small molecule inhibitors.


Assuntos
Antineoplásicos/uso terapêutico , Neoplasias Inflamatórias Mamárias/tratamento farmacológico , Lipocalina-2/antagonistas & inibidores , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Neoplasias Inflamatórias Mamárias/genética , Neoplasias Inflamatórias Mamárias/patologia , Lipocalina-2/genética , Células MCF-7 , Terapia de Alvo Molecular/métodos , Invasividade Neoplásica , RNA Interferente Pequeno/farmacologia , RNA Interferente Pequeno/uso terapêutico
10.
Mol Biol (Mosk) ; 55(4): 667-675, 2021.
Artigo em Russo | MEDLINE | ID: mdl-34432784

RESUMO

MicroRNAs epigenetically regulate physiological and pathological processes. Previously, we found that miR-204-5p is expressed at low levels in melanoma cells, and an increase in its level leads to a change in proliferation, migration, and invasion of these cancer cells. Now, using bioinformatics analysis, it has been shown that the target of miR-204-5p is FOXC1 transcription factor, which is implicated in carcinogenesis. Using the luciferase reporter assay, it was found that miR-204-5p suppresses expression of the FOXC1 gene by binding to its 3' non-coding region. Transfection of small interfering RNA (siRNA) targeting FOXC1 into melanoma cells caused a decrease in miR-204-5p levels, which is consistent with the generally accepted concept of feedback regulation of miRNA expression by target genes. According to the results of the MTT test and fluorescence microscopy, the proliferation level of melanoma cells under the influence of siRNA to FOXC1 decreased 72 h after transfection. Changes in the ratio of cells by cell cycle phase were analyzed using flow cytometry. Regulatory relationships between FOXC1 and miR-204-5p, and an inhibitory effect of FOXC1 knockdown on melanoma cell proliferation were revealed. Based on the results, it can be assumed that miR-204-5p regulates proliferation of melanoma cells by affecting FOXC1 expression.


Assuntos
Melanoma , MicroRNAs , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Fatores de Transcrição Forkhead , Humanos , Melanoma/genética , MicroRNAs/genética
11.
Theranostics ; 11(16): 7658-7670, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34335956

RESUMO

SNAI1 is widely regarded as a master driver of epithelial-mesenchymal transition (EMT) and associated with breast cancer progression and metastasis. This pro-malignant role is strongly linked to posttranslational modification, especially phosphorylation, which controls its protein levels and subcellular localization. While multiple kinases are implicated in regulation of SNAI1 stability, the precise mechanism by which SNAI1 is stabilized in tumors remains to be fully elucidated. Methods: A series of in vitro and in vivo experiments were conducted to reveal the regulation of SNAI1 by Serine/Threonine Kinase 39 (STK39) and the role of STK39 in breast cancer metastasis. Results: We identified STK39, a member of Stem 20-like serine/threonine kinase family, as a novel posttranslational regulator that enhances the stability of SNAI1. Inhibition of STK39 via knockdown or use of a specific inhibitor resulted in SNAI1 destabilization. Mechanistically, STK39 interacted with and phosphorylated SNAI1 at T203, which is critical for its nuclear retention. Functionally, STK39 inhibition markedly impaired the EMT phenotype and decreased tumor cell migration, invasion, and metastasis both in vitro and in vivo. These effects were rescued by ectopic SNAI1 expression. In addition, depletion of STK39 dramatically enhanced sensitivity to chemotherapeutic agents. Conclusions: Our study demonstrated that STK39 is a key mediator of SNAI1 stability and is associated with the pro-metastatic cellular process, highlighting the STK39-SNAI1 signaling axis as promising therapeutic targets for treatments of metastatic breast cancer.


Assuntos
Neoplasias da Mama/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Fatores de Transcrição da Família Snail/metabolismo , Animais , Neoplasias da Mama/fisiopatologia , Linhagem Celular Tumoral , Movimento Celular/genética , Transição Epitelial-Mesenquimal/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Camundongos , Camundongos SCID , Invasividade Neoplásica/genética , Fosforilação , Proteínas Serina-Treonina Quinases/genética , Fatores de Transcrição da Família Snail/genética , Fatores de Transcrição/metabolismo
12.
Theranostics ; 11(16): 7640-7657, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34335955

RESUMO

Background: Since primary prostate cancer (PCa) can advance to the life-threatening metastatic PCa, exploring the molecular mechanisms underlying PCa metastasis is crucial for developing the novel targeted preventive strategies for decreasing the mortality of PCa. RNA N6-methyladenosine (m6A) is an emerging regulatory mechanism for gene expression and its specific roles in PCa progression remains elusive. Methods: Western blotting, quantitative real-time PCR and immunohistochemical analyses were used to detect target gene expression in PCa cells in vitro and prostate tissues from patients. RNA immunoprecipitation was conducted to analyze the specific binding of mRNA to the target protein. Migration and invasion assays were used to assess the migratory capacities of cancer cells. The correlation between target gene expression and survival rate of PCa patients was analyzed based the TCGA database. Results: We found that total RNA N6-methyladenosine (m6A) modification levels were markedly upregulated in human PCa tissues due to increased expression of methyltransferase like 3 (METTL3). Further studies revealed that the migratory and invasive capacities of PCa cells were markedly suppressed upon METTL3 knockdown. Mechanistically, METTL3 mediates m6A modification of USP4 mRNA at A2696, and m6A reader protein YTHDF2 binds to and induces degradation of USP4 mRNA by recruiting RNA-binding protein HNRNPD to the mRNA. Decrease of USP4 fails to remove the ubiquitin group from ELAVL1 protein, resulting in a reduction of ELAVL1 protein. Lastly, downregulation of ELAVL1 in turn increases ARHGDIA expression, promoting migration and invasion of PCa cells. Conclusions: Our findings highlight the role of METTL3 in modulating invasion and metastasis of PCa cells, providing insight into promising therapeutic strategies for hindering PCa progressing to deadly metastases.


Assuntos
Metiltransferases/genética , Neoplasias da Próstata/metabolismo , Adenosina/análogos & derivados , Adenosina/genética , Adenosina/metabolismo , Movimento Celular/genética , Proliferação de Células/genética , Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/genética , Inativação Gênica/fisiologia , Humanos , Masculino , Metiltransferases/metabolismo , Invasividade Neoplásica/genética , Metástase Neoplásica/genética , Próstata/metabolismo , Próstata/patologia , Neoplasias da Próstata/genética , RNA Mensageiro/genética , Proteínas de Ligação a RNA/metabolismo , Proteases Específicas de Ubiquitina/genética
13.
Theranostics ; 11(16): 8112-8128, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34335983

RESUMO

The coiled-coil domain containing protein members have been well documented for their roles in many diseases including cancers. However, the function of the coiled-coil domain containing 65 (CCDC65) remains unknown in tumorigenesis including gastric cancer. Methods: CCDC65 expression and its correlation with clinical features and prognosis of gastric cancer were analyzed in tissue. The biological role and molecular basis of CCDC65 were performed via in vitro and in vivo assays and a various of experimental methods including co-immunoprecipitation (Co-IP), GST-pull down and ubiquitination analysis et al. Finally, whether metformin affects the pathogenesis of gastric cancer by regulating CCDC65 and its-mediated signaling was investigated. Results: Here, we found that downregulated CCDC65 level was showed as an unfavourable factor in gastric cancer patients. Subsequently, CCDC65 or its domain (a.a. 130-484) was identified as a significant suppressor in GC growth and metastasis in vitro and in vivo. Molecular basis showed that CCDC65 bound to ENO1, an oncogenic factor has been widely reported to promote the tumor pathogenesis, by its domain (a.a. 130-484) and further promoted ubiquitylation and degradation of ENO1 by recruiting E3 ubiquitin ligase FBXW7. The downregulated ENO1 decreased the binding with AKT1 and further inactivated AKT1, which led to the loss of cell proliferation and EMT signal. Finally, we observed that metformin, a new anti-cancer drug, can significantly induce CCDC65 to suppress ENO1-AKT1 complex-mediated cell proliferation and EMT signals and finally suppresses the malignant phenotypes of gastric cancer cells. Conclusion: These results firstly highlight a critical role of CCDC65 in suppressing ENO1-AKT1 pathway to reduce the progression of gastric cancer and reveals a new molecular mechanism for metformin in suppressing gastric cancer. Our present study provides a new insight into the mechanism and therapy for gastric cancer.


Assuntos
Biomarcadores Tumorais/metabolismo , Proteínas de Ligação a DNA/metabolismo , Glicoproteínas/metabolismo , Fosfopiruvato Hidratase/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Neoplasias Gástricas/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Animais , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , China , Feminino , Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/genética , Genes Supressores de Tumor/fisiologia , Glicoproteínas/genética , Humanos , Masculino , Metformina/metabolismo , Metformina/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Oncogenes , Prognóstico , Proteínas Proto-Oncogênicas c-akt/fisiologia , Transdução de Sinais/genética , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação
14.
Int J Mol Sci ; 22(14)2021 Jul 09.
Artigo em Inglês | MEDLINE | ID: mdl-34299018

RESUMO

Connexins (Cx) form gap junctions (GJ) and allow for intercellular communication. However, these proteins also modulate gene expression, growth, and cell migration. The downregulation of Cx43 impairs endothelial cell migration and angiogenetic potential. Conversely, endothelial Cx43 expression is upregulated in an in vivo angiogenesis model relying on hemodynamic forces. We studied the effects of Cx43 expression on tube formation and proliferation in HUVECs and examined its dependency on GJ communication. Expectedly, intercellular communication assessed by dye transfer was linked to Cx43 expression levels in HUVECs and was sensitive to a GJ blockade by the Cx43 mimetic peptide Gap27. The proliferation of HUVECs was not affected by Cx43 overexpression using Cx43 cDNA transfection, siRNA-mediated knockdown of Cx43, or the inhibition of GJ compared to the controls (transfection of an empty vector, scrambled siRNA, and the solvent). In contrast, endothelial tube and sprout formation in HUVECs was minimized after Cx43 knockdown and significantly enhanced after Cx43 overexpression. This was not affected by a GJ blockade (Gap27). We conclude that Cx43 expression positively modulates the angiogenic potential of endothelial cells independent of GJ communication. Since proliferation remained unaffected, we suggest that Cx43 protein may modulate endothelial cell migration, thereby supporting angiogenesis. The modulation of Cx43 expression may represent an exploitable principle for angiogenesis induction in clinical therapy.


Assuntos
Movimento Celular/genética , Proliferação de Células/genética , Conexina 43/metabolismo , Células Endoteliais/metabolismo , Junções Comunicantes/metabolismo , Neovascularização Fisiológica/genética , Comunicação Celular/efeitos dos fármacos , Comunicação Celular/genética , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Conexina 43/genética , Conexinas/farmacologia , Células Endoteliais/efeitos dos fármacos , Expressão Gênica , Técnicas de Silenciamento de Genes , Células Endoteliais da Veia Umbilical Humana , Humanos , Técnicas In Vitro , Oligopeptídeos/farmacologia , RNA Interferente Pequeno , Reação em Cadeia da Polimerase em Tempo Real , Regulação para Cima
15.
Int J Mol Sci ; 22(14)2021 Jul 09.
Artigo em Inglês | MEDLINE | ID: mdl-34299025

RESUMO

Several studies have demonstrated that melanoma-derived extracellular vesicles (EVs) are involved in lymph node metastasis; however, the molecular mechanisms involved are not completely defined. Here, we found that EMILIN-1 is proteolyzed and secreted in small EVs (sEVs) as a novel mechanism to reduce its intracellular levels favoring metastasis in mouse melanoma lymph node metastatic cells. Interestingly, we observed that EMILIN-1 has intrinsic tumor and metastasis suppressive-like properties reducing effective migration, cell viability, primary tumor growth, and metastasis. Overall, our analysis suggests that the inactivation of EMILIN-1 by proteolysis and secretion in sEVs reduce its intrinsic tumor suppressive activities in melanoma favoring tumor progression and metastasis.


Assuntos
Movimento Celular/genética , Proliferação de Células/genética , Vesículas Extracelulares/metabolismo , Melanoma/metabolismo , Glicoproteínas de Membrana/metabolismo , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Biologia Computacional , Metástase Linfática/genética , Masculino , Espectrometria de Massas , Melanoma/genética , Melanoma/patologia , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Proteólise , RNA-Seq , Reação em Cadeia da Polimerase em Tempo Real , Estatísticas não Paramétricas , Regulação para Cima , Ensaios Antitumorais Modelo de Xenoenxerto
16.
Breast Cancer Res Treat ; 189(2): 333-345, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34241740

RESUMO

BACKGROUND: Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype that lacks targeted therapies. Patients with TNBC have a very poor prognosis because the disease often metastasizes. New treatment approaches addressing drivers of metastasis and tumor growth are crucial to improving patient outcomes. Developing targeted gene therapy is thus a high priority for TNBC patients. PEA15 (phosphoprotein enriched in astrocytes, 15 kDa) is known to bind to ERK, preventing ERK from being translocated to the nucleus and hence blocking its activity. The biological function of PEA15 is tightly regulated by its phosphorylation at Ser104 and Ser116. However, the function and impact of phosphorylation status of PEA15 in the regulation of TNBC metastasis and in epithelial-to-mesenchymal transition (EMT) are not well understood. METHODS: We established stable cell lines overexpressing nonphosphorylatable (PEA15-AA) and phospho-mimetic (PEA15-DD) mutants. To dissect specific cellular mechanisms regulated by PEA15 phosphorylation status, we performed RT-PCR immune and metastasis arrays. In vivo mouse models were used to determine the effects of PEA15 phosphorylation on tumor growth and metastasis. RESULTS: We found that the nonphosphorylatable mutant PEA15-AA prevented formation of mammospheres and expression of EMT markers in vitro and decreased tumor growth and lung metastasis in in vivo experiments when compared to control, PEA15-WT and phosphomimetic PEA15-DD. However, phosphomimetic mutant PEA15-DD promoted migration, mesenchymal marker expression, tumorigenesis, and lung metastasis in the mouse model. PEA15-AA-mediated inhibition of breast cancer cell migratory capacity and tumorigenesis was the partial result of decreased expression of interleukin-8 (IL-8). Further, we identified that expression of IL-8 was possibly mediated through one of the ERK downstream molecules, Ets-1. CONCLUSIONS: Our results show that PEA15 phosphorylation status serves as an important regulator for PEA15's dual role as an oncogene or tumor suppressor and support the potential of PEA15-AA as a therapeutic strategy for treatment of TNBC.


Assuntos
Proteínas Reguladoras de Apoptose/genética , Transição Epitelial-Mesenquimal , Neoplasias de Mama Triplo Negativas , Animais , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Interleucina-8 , Camundongos , Neoplasias de Mama Triplo Negativas/genética
17.
Mol Med Rep ; 24(4)2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34328193

RESUMO

Lung cancer is one of the most prevalent cancers in China, and its incidence and morbidity remain high due to various independent factors. Lung adenocarcinoma (ADC) is the most common type of non­small cell lung carcinoma. Circular RNA plasmacytoma variant translocation 1 (circ­PVT1) plays an oncogenic role in various types of cancer, but the specific role of circ­PVT1 in lung ADC has not yet been reported. In the present study, circ­PVT1 was knocked down in A549 cells and the cell viability, proliferation, migration and invasion were measured via MTT, colony formation, wound healing and Transwell assays, respectively. Then, the cell viability of A549 cells with circ­PVT1­knockdown or ­overexpression was detected after exposure to cisplatin (DDP). After confirming the associations among circ­PVT1, microRNA (miR)­429 and forkhead box k1 (FOXK1) using various tools and assays, the cellular functions of A549 cells treated with combined short hairpin (sh)RNA­circ­PVT1 and miR­429 inhibitor/pcDNA3.1­FOXK1 were tested again. The expression of circ­PVT1 was found to be increased in lung ADC cells, and shRNA­circ­PVT1 led to decreased cell viability, proliferation, migration and invasion. The expression of circ­PVT1 was higher in A549/DDP cells than that in A549 cells, and the activity of caspase­3 was also activated by DDP in A549/DDP cells transfected with shRNA­circ­PVT1, whereas it was inactivated by DDP in A549 cells transfected with circ­PVT1 overexpression plasmid. Furthermore, the decreased cell viability, proliferation, invasion and migration induced by shRNA­circ­PVT1 could be abated by transfection with miR­429 inhibitor and pcDNA3.1­FOXK1. In conclusion, interference of circ­PVT1 inhibits the progression of lung ADC and enhances its sensitivity to DDP via miR­429/FOXK1, which may provide a theoretical basis for the use of novel targets in the treatment of lung ADC.


Assuntos
Adenocarcinoma de Pulmão/genética , Fatores de Transcrição Forkhead/metabolismo , Neoplasias Pulmonares/genética , MicroRNAs/metabolismo , RNA Circular/genética , RNA Circular/metabolismo , Células A549 , Adenocarcinoma de Pulmão/tratamento farmacológico , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/genética , Movimento Celular/genética , Proliferação de Células/genética , Cisplatino/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Fatores de Transcrição Forkhead/genética , Técnicas de Silenciamento de Genes , Humanos , Neoplasias Pulmonares/tratamento farmacológico , MicroRNAs/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética
18.
Int J Mol Sci ; 22(12)2021 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-34208313

RESUMO

Intrahepatic cholangiocarcinoma (iCCA) is a heterogeneous bile duct cancer with a poor prognosis. Integrin αvß6 (ß6) has been shown to be upregulated in iCCA and is associated with its subclassification and clinicopathological features. In the present study, two ITGB6-knockout HuCCT1 CCA cell lines (ITGB6-ko cells) were established using the clustered regulatory interspaced short palindromic repeats (CRISPR), an associated nuclease 9 (Cas9) system, and single-cell cloning. RNA sequencing analysis, real-time polymerase chain reaction (PCR), and immunofluorescent methods were applied to explore possible downstream factors. ITGB6-ko cells showed significantly decreased expression of integrin ß6 on flow cytometric analysis. Both cell lines exhibited significant inhibition of cell migration and invasion, decreased wound-healing capability, decreased colony formation ability, and cell cycle dysregulation. RNA sequencing and real-time PCR analysis revealed a remarkable decrease in podocalyxin-like protein 2 (PODXL2) expression in ITGB6-ko cells. Colocalization of PODXL2 and integrin ß6 was also observed. S100 calcium-binding protein P and mucin 1, which are associated with CCA subclassification, were downregulated in ITGB6-ko cells. These results describe the successful generation of ITGB6-ko CCA cell clones with decreased migration and invasion and downregulation of PODXL2, suggesting the utility of integrin ß6 as a possible therapeutic target or diagnostic marker candidate.


Assuntos
Moléculas de Adesão Celular/metabolismo , Movimento Celular , Colangiocarcinoma/patologia , Técnicas de Inativação de Genes , Cadeias beta de Integrinas/genética , Sialoglicoproteínas/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Proteínas de Ligação ao Cálcio/metabolismo , Moléculas de Adesão Celular/genética , Ciclo Celular , Linhagem Celular Tumoral , Movimento Celular/genética , Colangiocarcinoma/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Cadeias beta de Integrinas/metabolismo , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica , Proteínas de Neoplasias/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Sialoglicoproteínas/genética , Ensaio Tumoral de Célula-Tronco , Regulação para Cima/genética
19.
Anticancer Res ; 41(7): 3317-3326, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34230127

RESUMO

BACKGROUND/AIM: We evaluated the impact of FosL1, a member of the activated protein-1 family, on the pathways leading to regional metastasis of head and neck squamous cell carcinoma (HNSCC). MATERIALS AND METHODS: We examined the influence of small interfering RNA (siRNA) and short heparin RNA (shRNA) mediated knockdown of FosL1 on cell migration, invasion, and proliferation in vitro as well as on regional metastasis in vivo. The prognostic significance of FosL1 was also analyzed using the Kaplan- Meier plotter using data from an HNSCC patient database. RESULTS: Down-regulation of FosL1 inhibited cell migration, invasion, and proliferation in vitro, decreased the incidence of regional metastases, and prolonged the survival of mice in vivo. We also determined that HNSCC patients with higher expression levels of FosL1 had a significantly shorter survival time than those with low expression of FosL1. CONCLUSION: FosL1 plays a crucial role in promoting cell migration, invasion, and proliferation in HNSCC.


Assuntos
Movimento Celular/genética , Proliferação de Células/genética , Invasividade Neoplásica/genética , Proteínas Proto-Oncogênicas c-fos/genética , Carcinoma de Células Escamosas de Cabeça e Pescoço/genética , Animais , Linhagem Celular Tumoral , Regulação para Baixo/genética , Humanos , Metástase Linfática/genética , Metástase Linfática/patologia , Camundongos , Camundongos Nus , Invasividade Neoplásica/patologia , Prognóstico , RNA Interferente Pequeno/genética , Carcinoma de Células Escamosas de Cabeça e Pescoço/patologia
20.
Anticancer Res ; 41(7): 3349-3361, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34230131

RESUMO

BACKGROUND/AIM: The present study investigated the oncogenic functions of TACC3 in the progression of gastric cancer (GC). MATERIALS AND METHODS: We analysed TACC3 in relation to cell growth, invasion capability, expression of epithelial-mesenchymal transition (EMT)-related markers, and ERK/Akt/cyclin D1 signaling factors. The correlation between the immunohistochemically confirmed expression of TACC3 and clinical factors was also analyzed. RESULTS: The increased proliferation and invasion of TACC3-over-expressing GC cells was accompanied by altered regulation of EMT-associated markers and activation of ERK/Akt/cyclin D1 signaling. Immunohistochemical analysis of TACC3 in human GC tissues revealed that its expression is correlated with aggressive characteristics and poor prognosis of intestinal-type GC. CONCLUSION: TACC3 contributes to gastric tumorigenesis by promoting EMT via the ERK/Akt/cyclin D1 signaling pathway. The correlation between TACC3 expression and multiple clinicopathological variables implies that its effective therapeutic targeting in GC will depend on the tumor subtype.


Assuntos
Carcinogênese/genética , Ciclina D1/genética , Transição Epitelial-Mesenquimal/genética , Sistema de Sinalização das MAP Quinases/genética , Proteínas Associadas aos Microtúbulos/genética , Proteínas Proto-Oncogênicas c-akt/genética , Neoplasias Gástricas/genética , Carcinogênese/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Transdução de Sinais/genética , Estômago/patologia , Neoplasias Gástricas/patologia
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