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BACKGROUND: Sustained activation of hepatocyte growth factor (HGF)/c-MET signaling is a major driver of hepatocellular carcinoma (HCC) progression, but underlying mechanism is unclear. ArfGAP With SH3 Domain, Ankyrin Repeat And PH Domain 2 (ASAP2) can reportedly activate GTPases and promote receptor tyrosine kinase signaling. However, the exact role of ASAP2 in HCC, especially for c-MET activation, also remains elusive. METHODS: ASAP2 expression levels in HCC tissues and cells were quantified using qRT-PCR, western blot (WB) analysis, and immunohistochemistry staining. Cell counting kit-8 (CCK-8) and colony formation assays were performed to evaluate cell proliferation rates. Flow cytometry assays were conducted to assess apoptosis rates. Wound healing and Transwell assays were performed to determine cell migration and invasion capacities. Epithelial-mesenchymal transition (EMT)-related marker expression levels were also examined. Subcutaneous implantation and tail vein injection models were applied for in vivo growth and metastasis evaluations, respectively. Bioinformatics analyses of The Cancer Genome Atlas and STRING datasets were performed to explore ASAP2 downstream signaling. Co-immunoprecipitation and Cycloheximide chasing experiments were performed to assess protein-protein interactions and protein half-life, respectively. RESULTS: ASAP2 had higher expression levels in HCC tissues than in normal liver, and also predicted poor prognosis. Knocking down ASAP2 significantly impaired cell proliferation, migration, and invasion capacities, but promoted apoptosis in HCC cells in vitro. However, overexpression of ASAP2 achieved the opposite effects. In vivo experiments confirmed that ASAP2 could promote HCC cell growth and facilitate lung metastasis. Interestingly, ASAP2 was essential for triggering EMT. Gene Set Enrichment Analysis demonstrated that c-MET signaling was greatly enriched in ASAP2-high HCC cases. Additionally, c-MET signaling activity was significantly decreased following ASAP knockdown, evidenced by reduced c-MET, p-AKT, and p-ERK1/2 protein levels. Importantly, ASAP2 knockdown effectively attenuated HGF/c-MET signaling-induced malignant phenotypes. c-MET and ASAP2 expression levels were positively correlated in our cohort. Mechanistically, ASAP2 can directly bind to CIN85, thereby disrupting its interaction with c-MET, and can thus antagonize CIN85-induced c-MET internalization and lysosome-mediated degradation. Notably, knocking down CIN85 can rescue the observed inhibitory effects caused by ASAP2 knockdown. CONCLUSIONS: This study highlights the importance of ASAP2 in sustaining c-MET signaling, which can facilitate HCC progression.
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Rapid progression is the major cause of the poor prognosis of hepatocellular carcinoma (HCC); however, the underlying mechanism remained unclear. Here, we found Calpain-2 (CAPN2), a well-established protease that accelerates tumor progression in several malignancies, is overexpressed in HCC and acts as an independent predictor for poor outcomes. Furthermore, CAPN2 promoted the proliferation and invasion of HCC, and showed a positive correlation with the levels of invasion-related markers. Mechanistically, a novel CAPN2-SRC positive regulatory loop was identified upstream of ß-catenin to prevent its ubiquitination and degradation, and subsequently promoted HCC progression: CAPN2 could proteolyze PTP1B to form a truncation of approximately 42 kDa with increased phosphatase activity, resulting in reduced SRC Y530 phosphorylation and increased SRC kinase activity; meanwhile, CAPN2 itself was a bone fide substrate of SRC that was primarily phosphorylated at Y625 by SRC and exhibited increased proteolysis activity upon phosphorylation. Interestingly, the CAPN2-SRC loop could not only restrain most of cytoplasmic ß-catenin degradation by inhibiting GSK3ß pathway, but also prevented TRIM33-induced nuclear ß-catenin degradation even in ß-catenin-mutant cells. Present study identified a CAPN2-SRC positive loop responsible for intracellular ß-catenin accumulation and signaling activation, and targeting CAPN2 protease activity might be a promising approach for preventing HCC progression.
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Calpaína , Carcinoma Hepatocelular , Neoplasias Hepáticas , beta Catenina , Quinases da Família src , Calpaína/genética , Calpaína/metabolismo , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Fatores de Transcrição/metabolismo , Via de Sinalização Wnt , beta Catenina/metabolismo , Quinases da Família src/metabolismoRESUMO
Aberrantly activated Hedgehog (Hh) pathway is critical for driving the initiation and progression of multiple types of cancers, including medulloblastoma (MB) and basal cellular carcinoma (BCC). The majority of current Hh antagonist function by targeting the transmembrane domain of the oncoprotein Smoothened (Smo), a G-protein-coupled receptor-like receptor of Hh pathway. However, the primary and acquired resistance to current Smo inhibitors raise a critical need to develop next-generation of Smo inhibitors to improve their clinical efficacy. In this study, we identify that FDA approved drug ABT-199 significantly and selectively inhibits the Hh pathway. Mechanistically, ABT-199 acts as a competitive inhibitor of oxysterol by potentially targeting the cysteine rich domain (CRD) of Smo, rather as a BH3 mimetic. ABT-199 obviously inhibits the growth of Hh-driven tumors and possesses capacity of combating the primary and acquired resistance to Smo inhibitors caused by Smo mutations. Our data reposition ABT-199 as a Smo inhibitor for treating Hh-driven tumors, especially for those bearing Smo mutations and resistant to current Smo inhibitors. Meanwhile, our findings strengthen the argument that the CRD of Smo is a promising target for developing novel Smo inhibitors with capacity of combating the resistance to Smo inhibitors.
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Antineoplásicos/uso terapêutico , Compostos Bicíclicos Heterocíclicos com Pontes/uso terapêutico , Neoplasias/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Receptor Smoothened/antagonistas & inibidores , Sulfonamidas/uso terapêutico , Animais , Antineoplásicos/metabolismo , Sítios de Ligação , Compostos Bicíclicos Heterocíclicos com Pontes/metabolismo , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Proteínas Hedgehog/metabolismo , Humanos , Hidroxicolesteróis/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Células NIH 3T3 , Neoplasias/metabolismo , Ligação Proteica , Receptor Smoothened/química , Receptor Smoothened/metabolismo , Sulfonamidas/metabolismoRESUMO
BACKGROUND: Long non-coding RNAs (lncRNAs) are crucial in the invasion, angiogenesis, progression, and metastasis of hepatocellular carcinoma (HCC). The lncRNA MYLK-AS1 promotes the growth and invasion of HCC through the EGFR/HER2-ERK1/2 signaling pathway. However, the clinical significance of MYLK-AS1 in HCC still needs to be further determined. METHODS: Bioinformatic analysis was performed to determine the potential relationship among MYLK-AS1, miRNAs and mRNAs. A total of 156 samples of normal liver and paired HCC tissues from HCC patients were used to evaluate MYLK-AS1 expression by qRT-PCR. Human HCC cell lines were used to evaluate the colony formation, cell proliferation, migration, invasion, cell cycle and apoptosis after transfection of lentiviral short-hairpin RNAs (shRNAs) targeting MYLK-AS1 or MYLK-AS1 vectors. The competitive endogenous RNA (ceRNA) mechanism was clarified using fluorescence in situ hybridization (FISH), Western blotting, qPCR, RNA binding protein immunoprecipitation (RIP), and dual luciferase reporter analysis. RESULTS: MYLK-AS1 up-regulation was detected in the HCC tumor tissues and cell lines associated with the enhancement of the angiogenesis and tumor progression. The down-regulation of MYLK-AS1 reversed the effects on angiogenesis, proliferation, invasion and metastasis in the HCC cells and in vivo. MYLK-AS1 acted as ceRNA, capable of regulating the angiogenesis in HCC, while the microRNA miR-424-5p was the direct target of MYLK-AS1. Promoting the angiogenesis and the tumor proliferation, the complex MYLK-AS1/miR-424-5p activated the VEGFR-2 signaling through E2F7, whereas the specific targeting of E2F transcription factor 7 (E2F7) by miR-424-5p, was indicated by the mechanism studies. CONCLUSIONS: MYLK-AS1 and E2F7 are closely related to some malignant clinicopathological features and prognosis of HCC, thus the MYLK-AS1/ miR-424-5p/E2F7 signaling pathway might represent a promising treatment strategy to combat HCC.
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Proteínas de Ligação ao Cálcio/genética , Carcinoma Hepatocelular/irrigação sanguínea , Fator de Transcrição E2F7/metabolismo , Neoplasias Hepáticas/irrigação sanguínea , MicroRNAs/metabolismo , Quinase de Cadeia Leve de Miosina/genética , RNA Longo não Codificante/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Progressão da Doença , Feminino , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Masculino , Pessoa de Meia-Idade , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Prognóstico , RNA Antissenso/genética , RNA Antissenso/metabolismo , Transdução de Sinais , TransfecçãoRESUMO
Background: Previous studies reported that stress-induced phosphoprotein 1 (STIP1) can be secreted by hepatocellular carcinoma (HCC) cells and is increased in the serum of HCC patients. However, the therapy-monitoring and prognostic value of serum STIP1 in HCC remains unclear. Here, we aimed to systemically explore the prognostic significance of serum STIP1 in HCC. Methods: A total of 340 HCC patients were recruited to this study; 161 underwent curative resection and 179 underwent transcatheter arterial chemoembolization (TACE). Serum STIP1 was detected by enzyme-linked immunosorbent assay (ELISA). Optimal cutoff values for serum STIP1 in resection and TACE groups were determined by receiver operating characteristic (ROC) analysis. Prognostic value was assessed by Kaplan-Meier, log-rank, and Cox regression analyses. Predictive values of STIP1 for objective response (OR) to TACE and MVI were evaluated by ROC curves and logistic regression. Results: Serum STIP1 was significantly increased in HCC patients when compared with chronic hepatitis B patients or health donors (both P < 0.05). Optimal cutoff values for STIP1 in resection and TACE groups were 83.43 and 112.06 ng/ml, respectively. High pretreatment STIP1 was identified as an independent prognosticator. Dynamic changes in high STIP1 status were significantly associated with long-term prognosis, regardless of treatment approaches. Moreover, post-TACE STIP1 was identified as an independent predictor for OR, with a higher area under ROC curve (AUC-ROC) than other clinicopathological features. Specifically, pretreatment STIP1 was significantly increased in patients with microvascular invasion (MVI), and was confirmed as a novel, powerful predictor for MVI. Conclusions: Serum STIP1 is a promising biomarker for outcome evaluation, therapeutic response assessment, and MVI prediction in HCC. Integration serum STIP1 detection into HCC management might facilitate early clinical decision making to improve the prognosis of HCC.
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Liver cancer is a lethal disease that is associated with poor prognosis. In order to identify the functionally important genes associated with liver cancer that may reveal novel therapeutic avenues, we performed integrated analysis to profile miRNA and mRNA expression levels for liver tumors compared to normal samples in The Cancer Genome Atlas (TCGA) database. We identified 405 differentially expressed genes and 233 differentially expressed miRNAs in tumor samples compared with controls. In addition, we also performed the pathway analysis and found that mitogen-activated protein kinases (MAPKs) and G-protein coupled receptor (GPCR) pathway were two of the top significant pathway nodes dysregulated in liver cancer. Furthermore, by examining these signaling networks, we discovered that FOS (Fos proto-oncogene, AP-1 transcription factor subunit), LAMC2 (laminin subunit gamma 2), and CALML3 (calmodulin like 3) were the most significant gene nodes with high degrees involved in liver cancer. The expression and disease prediction accuracy of FOS, LAMC2, CALML3, and their interacting miRNAs were further performed using a HCC cohort. Finally, we investigated the prognostic significance of FOS in another HCC cohort. Patients with higher FOS expression displayed significantly shorter time to recurrence (TTR) and overall survival (OS) compared with patients with lower expression. Collectively, our study demonstrates that FOS is a potential prognostic marker for liver cancer that may reveal a novel therapeutic avenue in this lethal disease.
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Carcinoma Hepatocelular/genética , Biologia Computacional/métodos , Neoplasias Hepáticas/genética , Proteínas Proto-Oncogênicas c-fos/genética , Fator de Transcrição AP-1/genética , Biomarcadores Tumorais/genética , Calmodulina/genética , Feminino , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Redes Reguladoras de Genes , Humanos , Laminina/genética , Masculino , Redes e Vias Metabólicas/genética , MicroRNAs/genética , Pessoa de Meia-Idade , Proteínas Quinases Ativadas por Mitógeno/genética , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Proto-Oncogene Mas , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismoRESUMO
There are some controversies about the involvement of microRNA (miR)-19a-3p in hepatocellular carcinoma (HCC) biology, even though many studies have shown that it plays an important role in cancer. In this study, we found that miR-19a-3p is usually overexpressed in HCC tissues compared with corresponding peritumorous tissues, and its expression was associated with tumor size and poor overall survival. MiR-19a-3p promoted cell proliferation significantly, and more cells were found in the S phase. In vivo, miR-19a-3p promoted liver tumor growth, and more HCC cells were found in the active cell cycle. Sequencing and bioinformatics analysis predicted that PIK3IP1 is a likely target gene of miR-19a-3p, and we next confirmed it by luciferase and rescue assays. Altogether, our data showed an important role of PIK3IP1 downregulation by miR-19a-3p in HCC progression, and the miR-19a-3p-PIK3IP1-AKT pathway may be a potential therapeutic target.
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BACKGROUND: Aberrant AKT activation contributes to cancer stem cell (CSC) traits in hepatocellular carcinoma (HCC). We previously reported that CD73 activated AKT signaling via the Rap1/P110ß cascade. Here, we further explored the roles of CD73 in regulating CSC characteristics of HCC. METHODS: CD73 expression modulations were conducted by lentiviral transfections. CD73+ fractions were purified by magnetic-based sorting, and fluorescent-activated cell sorting was used to assess differentiation potentials. A sphere-forming assay was performed to evaluate CSC traits in vitro, subcutaneous NOD/SCID mice models were generated to assess in vivo CSC features, and colony formation assays assessed drug resistance capacities. Stemness-associated gene expression was also determined, and underlying mechanisms were investigated by evaluating immunoprecipitation and ubiquitylation. RESULTS: We found CD73 expression was positively associated with sphere-forming capacity and elevated in HCC spheroids. CD73 knockdown hindered sphere formation, Lenvatinib resistance, and stemness-associated gene expression, while CD73 overexpression achieved the opposite effects. Moreover, CD73 knockdown significantly inhibited the in vivo tumor propagation capacity. Notably, we found that CD73+ cells exhibited substantially stronger CSC traits than their CD73- counterparts. Mechanistically, CD73 exerted its pro-stemness activity through dual AKT-dependent mechanisms: activating SOX9 transcription via c-Myc, and preventing SOX9 degradation by inhibiting glycogen synthase kinase 3ß. Clinically, the combined analysis of CD73 and SOX9 achieved a more accurate prediction of prognosis. CONCLUSIONS: Collectively, CD73 plays a critical role in sustaining CSCs traits by upregulating SOX9 expression and enhancing its protein stability. Targeting CD73 might be a promising strategy to eradicate CSCs and reverse Lenvatinib resistance in HCC.
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5'-Nucleotidase/genética , Carcinoma Hepatocelular/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/genética , Células-Tronco Neoplásicas/patologia , Fatores de Transcrição SOX9/genética , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proteínas Ligadas por GPI/genética , Humanos , Neoplasias Hepáticas/patologia , Células-Tronco Neoplásicas/metabolismo , Fatores de Transcrição SOX9/análiseRESUMO
BACKGROUND AND AIMS: Heat shock factor (HSF4) plays a vital role in carcinogenesis and tumour progression. However, its clinical significance implications in hepatocellular carcinoma (HCC) remained elusive. METHODS: RT-PCR and western blot were used to detect the HSF4 expression levels in HCC cells and tissues. Immunohistochemistry staining was performed on a tissue microarray containing 104 HCC patients received radical resection. In vitro effects of HSF4 on proliferation, migration and invasion were determined by colony formation and transwell assays in HCCLM3, Huh7, MHCC97L and SMMC7721 cells. Epithelial-mesenchymal transition (EMT) was identified by RT-PCR, WB and immunofluorescence in HCCLM3 and MHCC97L cells. AKT pathway activation was detected by WB and dual luciferase report system in HCCLM3 and MHCC97L cells. RESULTS: HSF4 expression was higher in primary HCC tissues derived from recurrent patients, and positively correlated with invasiveness potentials of cell lines. Clinically, patients with high HSF4 expression had significant poorer prognosis. In vitro experiments showed HSF4 silencing inhibited HCC cell proliferation, migration and invasion, whereas HSF4 overexpression had inverse effects. Moreover, silence of HSF4 induced an epithelial-like phenotype, whereas the overexpression of HSF4 resulted in a mesenchymal-like phenotype in HCC by activating AKT pathway. Further experiments showed that HSF4 could activate AKT pathway in a hypoxia-inducible factor-1α (HIF-1α) dependent, but transforming growth factor-ß (TGF-ß) independent manner. CONCLUSIONS: HSF4 is upregulated in HCC, resulting in greater proliferation, migration and invasion capacities. Moreover, high HSF4 expression is a promising predictive indicator of poor outcome after radical resection. HSF4 may promote aggressive tumour behaviour by enhancing EMT through activating AKT pathway in a HIF1α-dependent manner.
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Carcinoma Hepatocelular , Transição Epitelial-Mesenquimal , Proteínas de Choque Térmico HSP40 , Neoplasias Hepáticas , Proteínas Proto-Oncogênicas c-akt , Carcinoma Hepatocelular/genética , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de SinaisRESUMO
Using a method optimized in hepatocellular carcinoma (HCC), we established patient-derived xenograft (PDX) models with an increased take rate (42.2%) and demonstrated that FBS +10% dimethyl sulfoxide exhibited the highest tumor take rate efficacy. Among 254 HCC patients, 103 stably transplantable xenograft lines that could be serially passaged, cryopreserved and revived were established. These lines maintained the diversity of HCC and the essential features of the original specimens at the histological, transcriptome, proteomic and genomic levels. Tumor engraftment was associated with lack of encapsulation, poor tumor differentiation, large size and overexpression of cancer stem cell biomarkers, and was an independent predictor for overall survival and tumor recurrence after resection. To confirm the preclinical value of the PDX model in HCC treatment, several antitumor agents were tested in 16 selected PDX models. The results revealed a high degree of pharmacologic heterogeneity in the cohort, as well as heterogeneity to different agents in the same individual. The sorafenib responses observed between HCC patients and the corresponding PDXs were also consistent. After molecular characterization of the PDX models, we explored the predictive markers for sorafenib response and found that mitogen-activated protein kinase kinase kinase 1 (MAP3K1) might play an important role in sorafenib resistance and sorafenib response is impaired in patients with MAP3K1 downexpression. Our results indicated that PDX models could accurately reproduce patient tumors biology and could aid in the discovery of new treatments to advance in precision medicine.
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Biomarcadores Tumorais/análise , Carcinoma Hepatocelular/terapia , Neoplasias Hepáticas/terapia , Ensaios Antitumorais Modelo de Xenoenxerto , Animais , Carcinoma Hepatocelular/diagnóstico , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Quimiorradioterapia Adjuvante/métodos , Regulação para Baixo , Resistencia a Medicamentos Antineoplásicos , Feminino , Seguimentos , Perfilação da Expressão Gênica , Genômica , Hepatectomia , Humanos , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/patologia , MAP Quinase Quinase Quinase 1/metabolismo , Masculino , Pessoa de Meia-Idade , Estudo de Prova de Conceito , Estudos Prospectivos , Inibidores de Proteínas Quinases/administração & dosagem , Sorafenibe/administração & dosagem , Resultado do TratamentoRESUMO
Previous research suggests that far upstream element-binding protein 1 (FUBP1) plays an important role in various tumors including epatocellular carcinoma (HCC). However, the role of FUBP1 in liver cancer remains controversial, and the regulatory pathway by FUBP1 awaits to be determined. This study aims to identify the role of FUBP1 in HCC progression. Our result shows that the high level of FUBP1 expression in HCC predicts poor prognosis after surgery. Overexpression of FUBP1 promotes HCC proliferation, invasion, and metastasis by activating transforming growth factor-ß (TGF-ß)/Smad pathway and enhancing epithelial-mesenchymal transition (EMT) in vitro and in vivo. Inhibitor of Thrombospondin-1 (LSKL) could inhibit HCC proliferation and invasion in vitro and in vivo by blocking the activation of TGF-ß/Smad pathway mediated by thrombospondin-1 (THBS1). Our study identified the critical role of FUBP1-THBS1-TGF-ß signaling axis in HCC and provides potentially new therapeutic modalities in HCC.
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Carcinoma Hepatocelular/genética , Proteínas de Ligação a DNA/genética , Neoplasias Hepáticas/genética , Proteínas de Ligação a RNA/genética , Trombospondina 1/genética , Fator de Crescimento Transformador beta1/genética , Animais , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Xenoenxertos , Humanos , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Metástase Neoplásica , Peptídeos/farmacologia , Transdução de Sinais/efeitos dos fármacos , Proteínas Smad/genética , Trombospondina 1/antagonistas & inibidores , Análise Serial de TecidosRESUMO
Aim: To elucidate the integrative combinational gene regulatory network landscape of hepatocellular carcinoma (HCC) molecular carcinogenesis from diverse background. Materials & methods: Modified gene regulatory network analysis was used to prioritize differentially regulated genes and links. Integrative comparisons using bioinformatics methods were applied to identify potential critical molecules and pathways in HCC with different backgrounds. Results: E2F1 with its surrounding regulatory links were identified to play different key roles in the HCC risk factor dysregulation mechanisms. Hsa-mir-19a was identified as showed different effects in the three HCC differential regulation networks, and showed vital regulatory role in HBV-related HCC. Conclusion: We describe in detail the regulatory networks involved in HCC with different backgrounds. E2F1 may serve as a universal target for HCC treatment.
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Carcinoma Hepatocelular/genética , Fator de Transcrição E2F1/metabolismo , Redes Reguladoras de Genes , Neoplasias Hepáticas/genética , MicroRNAs/genética , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/mortalidade , Carcinoma Hepatocelular/virologia , Biologia Computacional , Conjuntos de Dados como Assunto , Intervalo Livre de Doença , Fator de Transcrição E2F1/antagonistas & inibidores , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Hepacivirus/isolamento & purificação , Hepacivirus/patogenicidade , Vírus da Hepatite B/isolamento & purificação , Vírus da Hepatite B/patogenicidade , Humanos , Estimativa de Kaplan-Meier , Fígado/patologia , Fígado/virologia , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/mortalidade , Neoplasias Hepáticas/virologia , MicroRNAs/metabolismo , PrognósticoRESUMO
Sorafenib, a multikinase inhibitor, is a new standard treatment for patients with advanced hepatocellular carcinoma (HCC). However, resistance to this regimen is frequently observed in clinical practice, and the molecular basis of this resistance remains largely unknown. Herein, the antitumor activity of sorafenib was assessed in 16 patient-derived xenograft (PDX) models of HCC. Gene expression analysis was conducted to identify factors that promote sorafenib resistance. Quantitative RT-PCR and immunoblotting were used to determine gene expression and activation of signaling pathways. Cell proliferation, clone formation, and transwell assays were conducted to evaluate drug-sensitivity, proliferation, and invasiveness, respectively. Kaplan-Meier analysis was used to evaluate the predictive power of biomarkers for sorafenib response. Differential gene expression analysis suggested that sorafenib resistance correlated with high karyopherin subunit alpha 3 (KPNA3) expression. Overexpression of KPNA3 in HCC cells enhanced tumor cell growth and invasiveness. Interestingly, KPNA3 was found to trigger epithelial-mesenchymal transition (EMT), a key process mediating drug resistance. On a mechanistic level, KPNA3 increased phosphorylation of AKT, which then phosphorylated ERK, and ultimately promoted TWIST expression to induce EMT and sorafenib resistance. Moreover, retrospective analysis revealed that HCC patients with low KPNA3 expression had remarkably longer survival after sorafenib treatment. Finally, we have identified a novel KPNA3-AKT-ERK-TWIST signaling cascade that promotes EMT and mediates sorafenib resistance in HCC. These findings suggest that KPNA3 is a promising biomarker for predicting patient responsiveness to sorafenib. Targeting KPNA3 may also contribute to resolving sorafenib resistance in HCC.
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BACKGROUND: Prognosis of hepatocellular carcinoma (HCC) remains poor due to high recurrence rate and ineffective treatment options, highlighting the need to better understand the mechanism of recurrence and metastasis in HCC. METHODS: We first collected messenger RNA (mRNA) expression data from 442 cases of HCC patients from The Cancer Genome Atlas (TCGA) database as well as 251 HCC patients from Zhongshan Hospital during 2009 and 2010 to analyze the expression pattern from tissue microarray (TMA) of baculoviral IAP repeat containing 3 (BIRC3). Then, we used BIRC3 gain-of-function (overexpression) and loss-of-function (knockdown) studies to examine the effect of BIRC3 on HCC cell proliferation and invasion. In addition, we also investigated the undying mechanism by which BIRC3 contributes to HCC tumor progression. Functionally, we also used a BIRC3-specific inhibitor AT-406 in HCC xenograft model to explore the potential therapeutic benefit of targeting BIRC3 in liver cancer. RESULTS: BIRC3 serves as a novel prognostic indicator for HCC patients undergoing curative resection. BIRC3 promotes HCC epithelial-mesenchymal transition (EMT), cell migration, and metastasis via upregulating MAP3K7, therefore, inducing ERK1/2 phosphorylation. The specific BIRC3 inhibitor AT-406 can inhibit HCC cell proliferation and reduce pulmonary metastases. CONCLUSION: BIRC3 induces tumor proliferation and metastasis in vitro and in vivo. BIRC3 may serve as a novel therapeutic target for liver cancer.
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Proteína 3 com Repetições IAP de Baculovírus/metabolismo , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Animais , Proteína 3 com Repetições IAP de Baculovírus/genética , Biomarcadores Tumorais/metabolismo , Western Blotting , Linhagem Celular Tumoral , Movimento Celular/genética , Movimento Celular/fisiologia , Proliferação de Células/genética , Proliferação de Células/fisiologia , Transição Epitelial-Mesenquimal/genética , Transição Epitelial-Mesenquimal/fisiologia , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo Real , Análise Serial de TecidosRESUMO
Accumulating evidence suggests that long non-coding RNA (lncRNA) plays important roles in some malignant tumors. However, the mechanism underlying how lncRNA regulates hepatocellular carcinoma (HCC) process remains largely unknown. In this study, we explored the potential role of lncRNA 00607 as a novel tumor suppressor in HCC. In this study, we examined the regulation of lncRNA 00607 by the important inflammatory cytokine tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6). We also determined the expression of LINC000607 in 159 HCC tumors and paired adjacent tissues. Effects of LINC000607 in HCC proliferation and apoptosis were examined in vitro in HCC cell lines and in vivo tumor xenografts. Furthermore, we also examine underlying mechanism by which lncRNA 00607 regulates NF-κB p65 and how LIN00607 exerts its tumor suppressor role in HCC. We found that lncRNA 00607 expression level is lower in HCC tumors compared with matched normal liver tissue, and its low expression predicts worse prognosis in HCC. Functionally, lncRNA 00607 overexpression leads to decreased HCC cell proliferation in vitro and in vivo, enhanced apoptosis and chemotherapeutic drug sensitivity. Mechanistically, lncRNA 00607 inhibits the p65 transcription by binding to the p65 promoter region, therefore contributing to increased p53 levels in HCC. Taken together, the findings of this study show that the TNF-α/IL-6-lncRNA 00607-NF-κB p65/p53 signaling axis represents a novel therapeutic avenue in cancer chemotherapy.
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Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , RNA Longo não Codificante/genética , Transdução de Sinais/genética , Fator de Transcrição RelA/genética , Proteína Supressora de Tumor p53/genética , Animais , Apoptose/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Células Hep G2 , Humanos , Masculino , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Regiões Promotoras Genéticas/genética , Transcrição Gênica/genéticaRESUMO
BACKGROUND: Liver cancer is the second leading cause of cancer-related deaths and characterized by heterogeneity and drug resistance. Patient-derived xenograft (PDX) models have been widely used in cancer research because they reproduce the characteristics of original tumors. However, the current studies of liver cancer PDX mice are scattered and the number of available PDX models are too small to represent the heterogeneity of liver cancer patients. To improve this situation and to complement available PDX models related resources, here we constructed a comprehensive database, PDXliver, to integrate and analyze liver cancer PDX models. DESCRIPTION: Currently, PDXliver contains 116 PDX models from Chinese liver cancer patients, 51 of them were established by the in-house PDX platform and others were curated from the public literatures. These models are annotated with complete information, including clinical characteristics of patients, genome-wide expression profiles, germline variations, somatic mutations and copy number alterations. Analysis of expression subtypes and mutated genes show that PDXliver represents the diversity of human patients. Another feature of PDXliver is storing drug response data of PDX mice, which makes it possible to explore the association between molecular profiles and drug sensitivity. All data can be accessed via the Browse and Search pages. Additionally, two tools are provided to interactively visualize the omics data of selected PDXs or to compare two groups of PDXs. CONCLUSION: As far as we known, PDXliver is the first public database of liver cancer PDX models. We hope that this comprehensive resource will accelerate the utility of PDX models and facilitate liver cancer research. The PDXliver database is freely available online at: http://www.picb.ac.cn/PDXliver/.
Assuntos
Bases de Dados como Assunto , Modelos Animais de Doenças , Neoplasias Hepáticas/genética , Ensaios Antitumorais Modelo de Xenoenxerto , Animais , Feminino , Humanos , Fígado/patologia , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Pessoa de Meia-Idade , MutaçãoRESUMO
Hypoxia is a critical aspect of tumor biology and has been associated with poor prognosis and resistance to traditional therapy. In the present study, differentially expressed genes and microRNAs (miRNAs/miRs) were screened for in the hepatocellular carcinoma (HCC) cell line Huh7 under hypoxic conditions. On the basis of microarray data, 11,508 mRNAs and 58 miRNAs exhibiting ≥1.5-fold change in expression under hypoxic conditions were identified. Gene Ontology (GO) and Kyoto Encyclopedia or Genes and Genomes pathway analysis revealed that the differentially expressed genes were primarily involved in cell cycle regulation, cell division, transcription and G-protein-coupled receptor signaling pathways. Using the TargetScan and miRanda software packages with the miRNA-mRNA negative expression network, differentially expressed miRNA targets were predicted. GO analysis revealed that the primary function of these miRNAs was to regulate transcription and phosphorylation. The miRNA-mRNA networks for transcription and phosphorylation were analyzed. Network analysis revealed that the key miRNAs in these networks were miR-19a, miR-34a, miR-29a, mir-196a, miR-25 and miR-1207, whose potential gene targets include DNA-binding proteins, zinc-finger proteins and transcription factors. Certain protein kinases, includingmitogen-activated protein kinase (MAPK) 1, MAPK kinase kinase4 and cyclin-dependent kinase 18, were also revealed to be present in the network. In hypoxic HCC tissue, levels of several key miRNAs implicated in the network analyses (miR-19a, miR-34a, miR-25 and miR-1207) were revealed to exhibit increased expression levels compared with the surrounding tissue. The results of the present study provide evidence that miRNAs serve an important function in transcription and phosphorylation in the hypoxic response of HCC cells.
RESUMO
The effects of long non-coding RNAs (lncRNAs) on hepatocellular carcinoma (HCC) remain largely unclear. In this study, we identified an interferon (IFN)-γ-induced LncRNA, LncRNA00364, in HCC by microarray. LncRNA00364 displays lower expression in HCC tumor samples compared to paired normal controls. Overexpression of LncRNA00364 inhibits cell proliferation, G1/S cell cycle progression and promotes apoptosis in HCC cell lines. Consistently, LncRNA00364 overexpression leads to decreased HCC tumor formation in vivo. Mechanistically, LncRNA00364 specifically binds with STAT3, resulting in inhibition of STAT3 phosphorylation and therefore leads to upregulation of IFIT2. In a clinical setting, LncRNA00364 shows an independent prognostic indicator for overall survival and cumulative recurrence in HCC patients, and correlates with IFIT2. Therefore, our study provides new insights into a novel therapeutic avenue targeting the LncRNA00364 signaling axis in HCC.