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1.
Mol Cancer Ther ; 19(1): 135-146, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31554653

RESUMO

Both the MAPK pathway and mevalonate (MVA) signaling pathway play an increasingly significant role in the carcinogenesis of colorectal carcinoma, whereas the cross-talk between these two pathways and its implication in targeted therapy remains unclear in colorectal carcinoma. Here, we identified that HMGCS1 (3-hydroxy-3-methylglutaryl-CoA synthase 1), the rate-limiting enzyme of the MVA pathway, is overexpressed in colon cancer tissues and positively regulates the cell proliferation, migration, and invasion of colon cancer cells. In addition, HMGCS1 could enhance the activity of pERK independent of the MVA pathway, and the suppression of HMGCS1 could completely reduce the EGF-induced proliferation of colon cancer cells. Furthermore, we found that trametinib, a MEK inhibitor, could only partially abolish the upregulation of HMGCS1 induced by EGF treatment, while combination with HMGCS1 knockdown could completely reverse the upregulation of HMGCS1 induced by EGF treatment and increase the sensitivity of colon cancer cells to trametinib. Finally, we combined trametinib and dipyridamole, a common clinically used drug that could suppress the activity of SREBF2 (sterol regulatory element-binding transcription factor 2), a transcription factor regulating HMGCS1 expression, and identified its synergistic effect in inhibiting the proliferation and survival of colon cancer cells in vitro as well as the in vivo tumorigenic potential of colon cancer cells. Together, the current data indicated that HMGCS1 may be a novel biomarker, and the combination of targeting HMGCS1 and MEK might be a promising therapeutic strategy for patients with colon cancer.

2.
Sheng Wu Yi Xue Gong Cheng Xue Za Zhi ; 36(2): 267-273, 2019 Apr 25.
Artigo em Chinês | MEDLINE | ID: mdl-31016944

RESUMO

To evaluate the differential expression profiles of the lncRNAs, miRNAs, mRNAs and ceRNAs, and their implication in the prognosis in clear cell renal cell carcinoma (CCRCC), the large sample genomics analysis technologies were used in this study. The RNA and miRNA sequencing data of CCRCC were obtained from The Cancer Genome Atlas (TCGA) database, and R software was used for gene expression analysis and survival analysis. Cytoscape software was used to construct the ceRNA network. The results showed that a total of 1 570 lncRNAs, 54 miRNAs, and 17 mRNAs were differentially expressed in CCRCC, and most of their expression levels were up-regulated (false discovery rate < 0.01 and absolute log fold change > 2). The ceRNA regulatory network showed the interaction between 89 differentially expressed lncRNAs and 9 differentially expressed miRNAs. Further survival analysis revealed that 38 lncRNAs (including COL18A1-AS1, TCL6, LINC00475, UCA1, WT1-AS, HOTTIP, PVT1, etc.) and 2 miRNAs (including miR-21 and miR-155) were correlated with the overall survival time of CCRCC ( P < 0.05). Together, this study provided us several new evidences for the targeted therapy and prognosis assessment of CCRCC.


Assuntos
Carcinoma de Células Renais/genética , Neoplasias Renais/genética , MicroRNAs/genética , RNA Longo não Codificante/genética , Humanos , Transcriptoma
3.
Cell Death Differ ; 26(11): 2400-2415, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30833665

RESUMO

Tumours manage to survive the ablation of mutant KRAS, despite the development of KRAS-targeted drugs. Here we describe that inhibition of mutant KRAS promotes MEK nuclear localization as an alternative mechanism of KRAS-targeted drugs resistance. Tissue microarray analysis in colon tumours shows that aberrant MEK nuclear localization is closely related to YAP levels and tumour malignancy. MEK nuclear localization could sequester ß-TrCP from cytoplasmic inactive YAP, then stabilizing YAP. Mutant KRAS restrains MEK within the cytoplasm via IQGAP1, inhibiting MEK nuclear translocation. Trametinib, an allosteric MEK inhibitor, could prevent MEK nuclear localization and subsequently promote YAP degradation. In vitro and in vivo results suggests that inhibition of MEK nuclear localization by trametinib synergizes with KRAS knockdown or deltarasin treatment in suppressing the viability of KRAS mutant colon cancer cells. Our study provides new insights into the mechanisms of resistance to KRAS ablation, and suggests novel strategies for the treatment of KRAS-mutant colon cancers.

4.
Cancer Lett ; 442: 202-212, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30429107

RESUMO

KRAS mutation is the most common type of mutation in human cancers. However, the direct pharmacological inhibition of KRAS has not been clinically successful. Trametinib (GSK1120212, Tram), a newer MEK inhibitor, inhibits RAS signaling through mitogen-activated protein kinase (MAPK) cascade suppression. The effectiveness of Tram in clinical practice is limited in KRAS mutant tumors compared to that in BRAF mutant tumors. Here, we found that Tram treatment provoked feedback activation of upstream RAS, thus causing an induction of phosphorylated MEK (pMEK) and phosphorylated ERK (pERK) rebound in KRAS mutant tumors. This failure of persistent ERK inhibition led to drug resistance. Zoledronic acid (ZA), a nitrogen-containing bisphosphonate, disrupts the biological activity of RAS by inhibiting its isoprenylation. Surprisingly, ZA overcame Tram resistance, and augmented antitumor activity was observed in KRAS mutant tumors both in vitro and in vivo. Furthermore, ZA enhanced the effect of Tram partially through the mevalonate pathway. In summary, the combination of the two FDA-approved drugs Tram and ZA may represent a novel therapeutic strategy for the treatment of KRAS mutant cancers.


Assuntos
Antineoplásicos/farmacologia , Resistencia a Medicamentos Antineoplásicos , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Mutação , Neoplasias/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas p21(ras)/genética , Piridonas/farmacologia , Pirimidinonas/farmacologia , Ácido Zoledrônico/farmacologia , Células A549 , Animais , Relação Dose-Resposta a Droga , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Feminino , Células HCT116 , Células HT29 , Humanos , Ácido Mevalônico/metabolismo , Camundongos Nus , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Neoplasias/enzimologia , Neoplasias/genética , Neoplasias/patologia , Fosforilação , Prenilação de Proteína , Transdução de Sinais/efeitos dos fármacos , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
5.
J Exp Clin Cancer Res ; 37(1): 218, 2018 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-30185207

RESUMO

BACKGROUND: ERRα, a constitutive transcription factor that regulates energy metabolism, plays an important role in the progression of various tumours. However, its role in cell survival and proliferation and its implication in targeted therapy in colon cancer remains elusive. METHODS: The expression of ERRα in colon cancer tissues and cell lines was detected by using western blotting and immunohistochemistry. A wound healing assay and a transwell assay were performed to examine the migration and invasion of the colon cancer cells. A cell viability assay, clonogenic assay, western blot assay and the dual-luciferase reporter assay were employed to study the interaction between trametinib (inhibitor of MEK) and EGF treatment. Flow cytometry, western blotting, quantitative reverse-transcription polymerase chain reaction and xenograft studies were used to identify whether the combination of trametinib and simvastatin had a synergistic effect. RESULTS: ERRα positively regulated the cell proliferation, migration and invasion of colon cancer cells, and the suppression of ERRα completely reduced the EGF treatment-induced proliferation of colon cancer cells. Further investigation showed that trametinib partially restrained the up-regulation of ERRα induced by the EGF treatment, and ERRα inhibition increased the sensitivity of colon cancer cells to trametinib. At last, we combined trametinib with simvastatin, a common clinically used drug with a new reported function of transcriptional activity inhibition of ERRα, and found that this combination produced a synergistic effect in inhibiting the proliferation and survival of colon cancer cells in vitro as well as in vivo. CONCLUSIONS: The present data indicated that ERRα acted as an oncogene in colon cancer cells, and the combined targeting of ERRα and MEK might be a promising therapeutic strategy for colon cancer treatment.


Assuntos
Neoplasias do Colo/tratamento farmacológico , Fator de Crescimento Epidérmico/genética , Piridonas/administração & dosagem , Pirimidinonas/administração & dosagem , Receptores Estrogênicos/genética , Animais , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Citometria de Fluxo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Camundongos , Inibidores de Proteínas Quinases/administração & dosagem , Transdução de Sinais/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
6.
Mol Med Rep ; 18(4): 4023-4029, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30106149

RESUMO

Dysregulation of epidermal growth factor receptor (EGFR) signaling is responsible for the resistance to EGFR tyrosine kinase inhibitors (TKIs), such as gefitinib and erlotinib, and is thereby associated with the progression of tumors in non­small cell lung cancers (NSCLCs). Immunoblotting results revealed that geranylgeranyl transferase 1 inhibitor (GGTI)­298, a geranylgeranyl transferase 1 inhibitor with potential antitumor effects, effectively inhibited the phosphorylation of EGFR and its downstream target protein kinase B (AKT). A combination of gefitinib and GGTI­298 amplified the inhibition of the EGFR­AKT signaling pathway. In addition, GGTI­298 treatment produced a synergistic effect on the inhibition of proliferation as indicated by the combination index values of <1 when combined with gefitinib in the NSCLC cell lines HCC827 and A549. These synergistic effects were also observed to induce apoptosis and migration inhibition. Further mechanistic studies demonstrated that GGTI­298 inhibited the activity of Ras homolog family member A (RhoA), and downregulation of RhoA with small interfering RNA impaired the phosphorylation of EGFR, which suggested that EGFR inhibition by GGTI­298 may be exerted mainly through RhoA mediation. These results presented a novel, promising therapeutic strategy involving a combination of two drugs for targeting EGFR signaling in lung cancer.


Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Antineoplásicos/farmacologia , Benzamidas/farmacologia , Inibidores Enzimáticos/farmacologia , Gefitinibe/farmacologia , Alquil e Aril Transferases/metabolismo , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Ciclina D1/metabolismo , Sinergismo Farmacológico , Receptores ErbB/metabolismo , Humanos , Neoplasias Pulmonares/enzimologia , Neoplasias Pulmonares/patologia , Modelos Biológicos , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteína rhoA de Ligação ao GTP/metabolismo
7.
Oncol Rep ; 40(4): 2171-2182, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30106444

RESUMO

Mutant KRAS and BRAF are associated with primary EGFR inhibitor resistance in colorectal cancer (CRC). However, other biomarkers that could predict EGFR inhibitor resistance remain elusive. In the present study, immunoblotting and cell proliferation results revealed that yes­associated protein (YAP), a downstream effector of the Hippo pathway, was positively associated with primary cetuximab resistance in CRC cells. YAP knockdown enhanced the cytotoxicity of cetuximab in CRC cells. Simvastatin, a 3­hydroxy­3­methylglutaryl­coenzyme A (HMG­CoA) reductase inhibitor of the mevalonate pathway that inhibits YAP bioactivity through nuclear translocation and total YAP expression, increased the cytotoxicity of EGFR inhibitors (cetuximab and gefitinib) against CRC cells. The combination of simvastatin and EGFR inhibitors inhibited YAP and EGFR signaling more markedly than each agent alone. Adding back geranylgeranyl pyrophosphate (GGPP), a key product of the mevalonate pathway, reversed the YAP bioactivity inhibition induced by simvastatin and the cell proliferation inhibition induced by the combination of simvastatin and EGFR inhibitors. Collectively, these results revealed that YAP may be useful in identifying cetuximab resistance in CRC and indicated that targeting of both YAP and EGFR signals may present a promising therapeutic approach for CRC.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Biomarcadores Tumorais/metabolismo , Neoplasias Colorretais/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos , Fosfoproteínas/antagonistas & inibidores , Fosfatos de Poli-Isoprenil/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Sinvastatina/farmacologia , Animais , Anticolesterolemiantes/farmacologia , Apoptose , Proliferação de Células , Cetuximab/farmacologia , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Combinação de Medicamentos , Receptores ErbB/metabolismo , Feminino , Gefitinibe , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Quinazolinas/farmacologia , Transdução de Sinais , Fatores de Transcrição , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
8.
Int J Oncol ; 53(4): 1732-1742, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30066855

RESUMO

Lung cancer is a major cause of mortality worldwide and non­small cell lung cancer (NSCLC) accounts for ~80% of all cases of lung cancer. Increasing evidence indicates that Rho family GTPase 3 (RhoE) is important in the carcinogenesis and progression of NSCLC. In addition, several studies have indicated that microRNA (miR)­200b/c is downregulated in NSCLC cells. However, the exact mechanism remains to be elucidated. In the present study, immunohistochemistry (IHC) assays were used to analyze the RhoE and epithelial­mesenchymal transition (EMT)­related proteins in NSCLC tissues. Putative target sequences of the RhoE 3' untranslated region (3'UTR) for miR­200b/c were detected using bioinformatics analysis. The mRNA expression levels of RhoE and miR­200b/c were determined by reverse transcription­quantitative polymerase chain reaction (RT­qPCR) analysis, and western blot analysis was used to detect the protein levels of RhoE in cells. The luciferase­reporter activity of the RhoE 3'UTR was detected using a dual­luciferase assay. A cell counting kit­8 assay, flow cytometry and Transwell assay were used to detect cell proliferation, cell cycle, and invasion and migration ability, respectively. The IHC assays indicated that RhoE was overexpressed in NSCLC tissues. The bioinformatics analysis revealed that the RhoE 3'UTR contained a putative target site for miR­200b/c, which was conserved across species. The results of RT­qPCR analysis showed that the mRNA expression of RhoE was overexpressed and miR­200b/200c was decreased in lung cancer tissues. The enhanced expression of miR­200b or miR­200c significantly downregulated the expression of RhoE at the mRNA and protein levels in A549 and NCI­H1299 NSCLC cells. Furthermore, luciferase assays showed that miR­200b and miR­200c directly targeted the 3'UTR of RhoE. The forced expression of miR­200b or miR­200c markedly inhibited A549 cell and NCI­H1299 cell proliferation, G0/G1 progression and cell invasion, which was consistent with the effects of RNA interference­mediated RhoE knockdown in these cells. The suppression of RhoE regulated the expression of EMT­related markers, which was consistent with the effect of miR­200b/c in NSCLC cells, and the expression of EMT­related proteins and RhoE were also correlated in the lung cancer tissues. Therefore, miR­200b and miR­200c targeted the expression of RhoE and inhibited the malignancy of NSCLC cells, and the downregulation of miR­200b and miR­200c may contribute to the high expression of RhoE in NSCLC.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/genética , MicroRNAs/metabolismo , Proteínas rho de Ligação ao GTP/genética , Regiões 3' não Traduzidas/genética , Carcinogênese/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Biologia Computacional , Regulação para Baixo , Transição Epitelial-Mesenquimal/genética , Feminino , Humanos , Pulmão/patologia , Neoplasias Pulmonares/patologia , Masculino , MicroRNAs/genética , Pessoa de Meia-Idade , Invasividade Neoplásica/genética , RNA Interferente Pequeno/metabolismo , Regulação para Cima , Proteínas rho de Ligação ao GTP/metabolismo
9.
Sheng Wu Yi Xue Gong Cheng Xue Za Zhi ; 35(1): 81-86, 2018 02 25.
Artigo em Chinês | MEDLINE | ID: mdl-29745605

RESUMO

The aim of this article is to study the regulatory feedback loop between ß-catenin and IQ motif containing GTPase activating protein 1 (IQGAP1), as well as the effect of this regulation loop in colon cancer cell proliferation. Western blot was used to detect the expression of IQGAP1 and ß-catenin after changing their expression respectively by transfection in SW1116 cells. CCK-8 cell proliferation assay was used to detect the effect of IQGAP1 involved in the proliferation of SW1116 cells promoted by ß-catenin. The results of Western blot indicated that ß-catenin could positively regulate IQGAP1, while IQGAP1 silencing could up-regulate ß-catenin, forming a negative feedback loop. The results of CCK-8 showed that IQGAP1 silencing inhibited ß-catenin-mediated proliferation in SW1116 cells. In conclusion, our research reveals a negative regulatory feedback loop between ß-catenin and IQGAP1 which has a remarkable effect on the proliferation ability of colon cancer cells.

10.
Biomed Pharmacother ; 103: 147-156, 2018 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-29649630

RESUMO

High expression levels of CD44 and YAP have been identified as poor prognostic factors in hepatocellular carcinoma (HCC). However, the mechanistic relationship between CD44 and YAP during HCC tumorigenesis remains largely unknown. To investigate the mutual regulation between standard CD44 (CD44S) and YAP1 in HCC cell lines and tissue samples, CD44S and YAP1 expression in 40 pairs of tumor samples and matched distal normal tissues from HCC patients was examined by immunohistochemical staining. High expression of either CD44S or YAP1 was associated with a younger age and worse pathology grade. In addition, high levels of CD44S and YAP1 were associated with increased vascular invasion and more severe liver cirrhosis, respectively. CD44S expression was positively correlated with YAP1 expression in these HCC tissues. In vitro experiments suggested that CD44S could positively regulate the expression of YAP1 and its target genes via the PI3K/Akt pathway in HCC cells. Moreover, CD44S is regulated by the YAP1/TEAD axis. These results reveal a novel positive feedback loop involving CD44S and YAP1, in which CD44S functions as both an upstream regulator and a downstream effector of YAP1 in HCC. This feedback loop might constitute a broadly conserved module for regulating cell proliferation and invasion during HCC tumorigenesis. Blocking this positive feedback loop that involves CD44S and YAP1 might represent a new approach for HCC treatment.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Carcinoma Hepatocelular/metabolismo , Retroalimentação Fisiológica , Receptores de Hialuronatos/metabolismo , Neoplasias Hepáticas/metabolismo , Fosfoproteínas/metabolismo , Linhagem Celular Tumoral , Citoplasma/metabolismo , Feminino , Humanos , Ácido Hialurônico/metabolismo , Masculino , Pessoa de Meia-Idade , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Fatores de Transcrição
11.
Cell Death Dis ; 9(3): 269, 2018 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-29449645

RESUMO

The epidermal growth factor receptor (EGFR) pathway and Hippo signaling play an important role in the carcinogenesis of hepatocellular carcinoma (HCC). However, the crosstalk between these two pathways and its implications in targeted therapy remains unclear. We found that the activated EGFR signaling could bypass RhoA to promote the expression of YAP(Yes-associated protein), the core effector of the Hippo signaling, and its downstream target Cyr61. Further studies indicated that EGFR signaling mainly acted through the PI3K-PDK1 (Phosphoinositide 3-kinase-Phosphoinositide-dependent kinase-1) pathway to activate YAP, but not the AKT and MAPK pathways. While YAP knockdown hardly affected the EGFR signaling. In addition, EGF could promote the proliferation of HCC cells in a YAP-independent manner. Combined targeting of YAP and EGFR signaling by simvastatin and the EGFR signaling inhibitors, including the EGFR tyrosine kinase inhibitor (TKI) gefitinib, the RAF inhibitor sorafenib and the MEK inhibitor trametinib, presented strong synergistic cytotoxicities in HCC cells. Therefore, the EGFR-PI3K-PDK1 pathway could activate the YAP signaling, and the activated EGFR signaling could promote the HCC cell growth in a YAP-independent manner. Combined use of FDA-approved inhibitors to simultaneously target YAP and EGFR signaling presented several promising therapeutic approaches for HCC treatment.

12.
Mol Med Rep ; 17(1): 1940-1946, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-29257203

RESUMO

Long noncoding RNA (lncRNA)­activated by transforming growth factor (TGF)-ß (lncRNA-ATB) was recognized as an unfavorable prognostic factor in various cancers; however, its regulatory role in gastric cancer (GC) remains elusive. The present study aimed to measure lncRNA­ATB expression in GC and to explore its involvement in GC progression. lncRNA­ATB expression levels were measured in 40 pairs of GC tissues and their normal adjacent tissues, as well as in 5 GC cell lines and a normal gastric mucosal cell line by reverse transcription-quantitative polymerase chain reaction. Knockdown experiments were performed to explore the effect of lncRNA­ATB on the cell proliferation, invasion and migration. The results demonstrated that lncRNA­ATB expression levels in GC tissues and GC cell lines were significantly higher than in the adjacent normal tissues and normal gastric mucosal cells. Further analysis of the correlation between the clinicopathological features and lncRNA­ATB expression indicated that higher expression of lncRNA­ATB was correlated with increased invasion depth, more distant metastasis and advanced tumor­node­metastasis stage. In addition, downregulated lncRNA­ATB expression suppressed cellular proliferation, invasion and migration of GC cells. In conclusion, these data suggested that lncRNA­ATB may serve as a clinical outcome predictor and potential therapeutic target in GC.


Assuntos
Regulação Neoplásica da Expressão Gênica , Invasividade Neoplásica/genética , RNA Longo não Codificante/genética , Neoplasias Gástricas/genética , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica/patologia , Neoplasias Gástricas/patologia
13.
Sci Rep ; 7: 40802, 2017 01 19.
Artigo em Inglês | MEDLINE | ID: mdl-28102288

RESUMO

Acetylcholine (ACh), known as a neurotransmitter, regulates the functions of numerous fundamental central and peripheral nervous system. Recently, emerging evidences indicate that ACh also plays an important role in tumorigenesis. However, little is known about the role of ACh in gastric cancer. Here, we reported that ACh could be auto-synthesized and released from MKN45 and BGC823 gastric cancer cells. Exogenous ACh promoted cell proliferation in a does-dependent manner. The M3R antagonist 4-DAMP, but not M1R antagonist trihexyphenidyl and M2/4 R antagonist AFDX-116, could reverse the ACh-induced cell proliferation. Moreover, ACh, via M3R, activated the EGFR signaling to induce the phosphorylation of ERK1/2 and AKT, and blocking EGFR pathway by specific inhibitor AG1478 suppressed the ACh induced cell proliferation. Furthermore, the M3R antagonist 4-DAMP and darifenacin could markedly inhibit gastric tumor formation in vivo. 4-DAMP could also significantly enhance the cytotoxic activity of 5-Fu against the MKN45 and BGC823 cells, and induce the expression of apoptosis-related proteins such as Bax and Caspase-3. Together, these findings indicated that the autocrine ACh could act through M3R and the EGFR signaling to promote gastric cancer cells proliferation, targeting M3R or EGFR may provide us a potential therapeutic strategy for gastric cancer treatment.


Assuntos
Acetilcolina/metabolismo , Receptores ErbB/metabolismo , Receptor Muscarínico M3/metabolismo , Transdução de Sinais , Neoplasias Gástricas/metabolismo , Acetilcolina/farmacologia , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Xenoenxertos , Humanos , Camundongos , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptor Muscarínico M3/agonistas , Receptor Muscarínico M3/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Neoplasias Gástricas/patologia
14.
Oncotarget ; 7(35): 56209-56218, 2016 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-27486823

RESUMO

The metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is frequently over-expressed and serves as a prognostic marker in human cancers. However, little is known about the role of MALAT1 in gastric cancer. Here, we reported that the tissue and plasma MALAT1 levels were significantly higher in gastric cancer patients with distant metastasis (P<0.01) than patients without distant metastasis and the healthy controls. In addition, high levels of plasma MALAT1 independently correlated to a poor prognosis for gastric cancer patients (hazard ratio, 0.242; 95% CI, 0.154-0.836; P=0.036; Cox regression analysis). Functional studies revealed that knockdown of MALAT1 could inhibit cell proliferation, cell cycle progression, migration and invasion, and promote apoptosis in gastric cancer cells. Furthermore, the miR-122-IGF-1R signaling correlated with the dysregulated MALAT1 expression in gastric cancer. These data suggest that MALAT1 could function as an oncogene in gastric cancer, and high MALAT1 level could serve as a potential biomarker for the distant metastasis of gastric cancer.


Assuntos
Adenocarcinoma/sangue , Biomarcadores Tumorais/metabolismo , Carcinogênese/patologia , Neoplasias Pulmonares/sangue , Oncogenes , RNA Longo não Codificante/metabolismo , Neoplasias Gástricas/patologia , Adenocarcinoma/mortalidade , Adenocarcinoma/patologia , Adenocarcinoma/secundário , Adenocarcinoma de Pulmão , Apoptose/genética , Biomarcadores Tumorais/sangue , Linhagem Celular Tumoral , Proliferação de Células , Progressão da Doença , Regulação para Baixo , Pontos de Checagem da Fase G1 do Ciclo Celular/genética , Técnicas de Silenciamento de Genes , Humanos , Neoplasias Pulmonares/mortalidade , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/secundário , MicroRNAs/metabolismo , Estadiamento de Neoplasias , Prognóstico , Interferência de RNA , RNA Longo não Codificante/sangue , RNA Longo não Codificante/genética , RNA Interferente Pequeno/metabolismo , Curva ROC , Receptores de Somatomedina/metabolismo , Análise de Regressão , Transdução de Sinais , Neoplasias Gástricas/sangue , Neoplasias Gástricas/mortalidade , Regulação para Cima
15.
Theranostics ; 6(4): 594-609, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26941850

RESUMO

Aberrant activation of the Src kinase is implicated in the development of a variety of human malignancies. However, it is almost impossible to monitor Src activity in an in vivo setting with current biochemical techniques. To facilitate the noninvasive investigation of the activity of Src kinase both in vitro and in vivo, we developed a genetically engineered, activatable bioluminescent reporter using split-luciferase complementation. The bioluminescence of this reporter can be used as a surrogate for Src activity in real time. This hybrid luciferase reporter was constructed by sandwiching a Src-dependent conformationally responsive unit (SH2 domain-Srcpep) between the split luciferase fragments. The complementation bioluminescence of this reporter was dependent on the Src activity status. In our study, Src kinase activity in cultured cells and tumor xenografts was monitored quantitatively and dynamically in response to clinical small-molecular kinase inhibitors, dasatinib and saracatinib. This system was also applied for high-throughput screening of Src inhibitors against a kinase inhibitor library in living cells. These results provide unique insights into drug development and pharmacokinetics/phoarmocodynamics of therapeutic drugs targeting Src signaling pathway enabling the optimization of drug administration schedules for maximum benefit. Using both Firefly and Renilla luciferase imaging, we have successfully monitored Src tyrosine kinase activity and Akt serine/threonine kinase activity concurrently in one tumor xenograft. This dual luciferase reporter imaging system will be helpful in exploring the complex signaling networks in vivo. The strategies reported here can also be extended to study and image other important kinases and the cross-talks among them.


Assuntos
Genes Reporter , Luciferases de Renilla/análise , Medições Luminescentes , Neoplasias/diagnóstico por imagem , Quinases da Família src/análise , Animais , Linhagem Celular Tumoral , Xenoenxertos , Luciferases de Renilla/genética , Camundongos , Análise de Sequência de DNA
16.
Eur Heart J Cardiovasc Imaging ; 17(7): 788-96, 2016 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-26341293

RESUMO

AIMS: The aim of this article is to determine the association between left atrial appendage (LAA) regional dysfunction using image-based motion-estimation computed tomography (CT) (iME) and a prior history of stroke or transient ischaemic attack (TIA) in patients with atrial fibrillation (AF). METHODS AND RESULTS: In this single-centre retrospective case-control study, among patients referred for AF ablation who underwent pre-ablation cardiac CT with retrospective ECG gating, we identified 18 patients with a prior history of stroke or TIA at the time of CT scan and 18 age- and gender-matched controls. All the patients were in sinus rhythm at the time of CT scan. Four-dimensional motion vector field was estimated from the CT images using iME. To assess myocardial deformation, area change ratio (A) and area change rate (AR) were calculated over the endocardial surface of the LAA. There was no significant difference in the baseline patient characteristics between the stroke/TIA group and the control group (67.6 ± 8.1 years old, 66.7% male, 16.7% persistent AF). LAA maximum (Amax; 23.8 ± 33.0 vs. 52.9 ± 41.2%, P = 0.02) and pre-atrial contraction area change ratio (ApreA; 13.7 ± 17.7 vs. 30.9 ± 29.2%, P = 0.04) were significantly lower in the stroke/TIA group than in the control group, respectively. The difference in LAA Amax and ApreA remained significant in multivariate analysis (P = 0.03 and P = 0.04, respectively). CONCLUSION: LAA regional dysfunction is associated with stroke/TIA in AF patients. Our results offer a basis for a prospective study to determine the role of LAA regional dysfunction by iME in predicting cerebrovascular events such as stroke or TIA.


Assuntos
Apêndice Atrial/diagnóstico por imagem , Fibrilação Atrial/diagnóstico por imagem , Tomografia Computadorizada Quadridimensional , Interpretação de Imagem Assistida por Computador , Acidente Vascular Cerebral/prevenção & controle , Idoso , Análise de Variância , Apêndice Atrial/cirurgia , Fibrilação Atrial/cirurgia , Estudos de Casos e Controles , Ablação por Cateter/métodos , Bases de Dados Factuais , Eletrocardiografia/métodos , Feminino , Seguimentos , Humanos , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Variações Dependentes do Observador , Cuidados Pré-Operatórios/métodos , Reprodutibilidade dos Testes , Estudos Retrospectivos , Medição de Risco , Resultado do Tratamento
17.
J Cancer Res Clin Oncol ; 141(4): 647-60, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25326346

RESUMO

PURPOSE: Circulating tumor cells (CTCs) have been proved to be responsible for tumor metastasis and resistant to anticancer therapies. This study aims to isolate and characterize circulating tumor cells from human gastric cancer patients, and investigate characteristic differences between gastric CTCs and gastric cancer cell lines. METHODS: We analyzed 31 cases of gastric cancer patients using anti-CD45 antibody-conjugated magnetic microbeads negative separation, combined with fluorescence activated cell sorter CD44 positive screening. Abilities of tumor formation, metastasis, invasion, migration, irradiation and drug sensitivity of CTCs and gastric cancer cell lines were detected and compared. RESULTS: Of all the 31 patients, CD44(+)/CD45(-)CTCs were isolated in 14 patients, of which 3 cases were stage IIA, 2 cases stage IIB, 2 cases stage IIIC and 7 cases stage IV. The malignant behavior was demonstrated by both clonogenetic assay and tumor xenograft in nude mice. Compared with human gastric cancer cell lines, the migration and invasion abilities of CTCs increased to 3.21-12.6-fold and 2.3-6.7-fold, respectively (all p values <0.05). In addition, the metastatic potential of CTCs is much higher in vivo than that of the control. Furthermore, CTCs were found to be relatively sensitive to FU, cisplatin and paclitaxel, but relatively resistant to irradiation, oxaliplatin, cetuximab and trastuzumab. CONCLUSIONS: CD44(+)/CD45(-) gastric CTCs were isolated and found to exhibit stronger malignant behavior when compared with human gastric cancer cell lines. Furthermore, CTCs cultured in vitro have potential implications in drug sensitivity screening for the future anticancer treatments.


Assuntos
Carcinogênese/metabolismo , Receptores de Hialuronatos/metabolismo , Células Neoplásicas Circulantes/metabolismo , Neoplasias Gástricas/metabolismo , Adulto , Idoso , Animais , Antineoplásicos/farmacologia , Western Blotting , Carcinogênese/patologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Movimento Celular/efeitos da radiação , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/efeitos da radiação , Células Cultivadas , Relação Dose-Resposta a Droga , Relação Dose-Resposta à Radiação , Feminino , Humanos , Imuno-Histoquímica , Imagem por Ressonância Magnética , Masculino , Camundongos Nus , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Células Neoplásicas Circulantes/patologia , Neoplasias Gástricas/sangue , Neoplasias Gástricas/patologia , Transplante Heterólogo , Raios X
18.
Int J Oncol ; 45(6): 2430-8, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25200917

RESUMO

Both circulating tumor cells (CTCs) and epithelial-mesenchymal transition (EMT) play an important role in invasion, migration and chemoresistant in tumor development. This study aimed to detect whether EMT occurred in human gastric CTCs and to explore the mechanism of EMT in human gastric CTCs. We analysed epithelial markers (pan-CK, E-cadherin), mesenchymal markers (N-cadherin, vimentin) EMT related miR­200s, and Akt in gastric CTCs. The impact of miR­200s on EMT, migration and invasion in CTCs was tested. We found that epithelial markers pan-CK, E-cadherin were decreased, and mesenchymal markers N-cadherin, vimentin were overexpressed in gastric CTCs. Expression of EMT related transcriptors, snail1, zeb1, twist1, were reversely correlated with miR­200s, and were positively correlated with phospho-Akt. Upregulated of miR­200s downregulated twist1 and zeb1 mRNA expression, and resulted in the supression of EMT, and impaired migration and invasion in gastric CTCs. Inhibition of p-Akt led to upregulation of miR­200s. In conclusion, gastric CTCs exhibited remarkable EMT process, and p-Akt/miR­200s signaling regulates EMT, migration and invasion in gastric CTCs.


Assuntos
Transição Epitelial-Mesenquimal/genética , MicroRNAs/biossíntese , Células Neoplásicas Circulantes/metabolismo , Proteína Oncogênica v-akt/biossíntese , Neoplasias Gástricas/genética , Biomarcadores Tumorais/biossíntese , Linhagem Celular Tumoral , Movimento Celular/genética , Regulação Neoplásica da Expressão Gênica , Humanos , MicroRNAs/genética , Invasividade Neoplásica/genética , Proteínas de Neoplasias/biossíntese , Células Neoplásicas Circulantes/patologia , Proteína Oncogênica v-akt/genética , Neoplasias Gástricas/patologia
19.
Oncol Rep ; 32(4): 1401-8, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25198583

RESUMO

Gefitinib demonstrates excellent performance in the treatment of lung adenocarcinoma patients; yet, there was no added benefit in combination with chemotherapy as reported in a phase III clinical trial. For exploring the mechanism of the failed combination therapy in lung cancer, in the present study, four therapy assessment groups, including a control group, a chemotherapy group [paclitaxel+cisplatin (TP)], a gefitinib monotherapy group (G) and a combination group[paclitaxel+cisplatin+gefitinib (TP+G)], were established in an A549 cell line and mouse xenotransplanted tumor models.By HPLC, we found that the gefitinib concentration was significantly higher in the combination group when compared to that in the G group in the non-small cell lung cancer cell line, A549 (p<0.05). Following the treatment time extension,an increased cell growth rate was observed in the combination group, while the cellular concentration of gefitinib was not decreased. The expression levels of P-IGF-1R, P-SRC and P-ERK in the fourth combination treatment group were significantly higher than levels in the fourth G treatment and control groups (p<0.05). Following downregulating of IGF-1R in the fourth combination treatment group, drug sensitivity was recovered in vitro. In the mouse model, compared with the gefitinib monotherapy group, the combination group exhibited a smaller tumor volume, lower body weight and reduced survival rate (p<0.05). Gefitinib concentrations in the serum and tumor tissues in the combination therapy group were also decreased when compared with these concentrations in the gefitinib alone group. The present study is the first to demonstrate that the decreased gefitinib concentration in serum and tumor tissues is one of the reasons resulting in the failed combination treatment (chemotherapy+gefitinib) in vivo study. Frequent use of the combination treatment in A549 lung cancer cells induced IGF-1R activation which contributed to gefitinib resistance and gave rise to the failure of the combination therapy.


Assuntos
Adenocarcinoma/tratamento farmacológico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Resistencia a Medicamentos Antineoplásicos/genética , Neoplasias Pulmonares/tratamento farmacológico , Receptores de Somatomedina/genética , Adenocarcinoma/patologia , Animais , Linhagem Celular Tumoral , Cisplatino/administração & dosagem , Gefitinibe , Humanos , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Nus , Paclitaxel/administração & dosagem , Quinazolinas/administração & dosagem , Falha de Tratamento , Carga Tumoral , Ensaios Antitumorais Modelo de Xenoenxerto
20.
Cell Signal ; 26(11): 2504-13, 2014 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25101858

RESUMO

The Hippo pathway plays an important role in both physical and pathogenesis processes. As crucial downstream effectors of Hippo pathway, YAP is inhibited by Lats1/2 through phosphorylation. However, upstream signals that regulate the Hippo pathway have been still poorly understood. Here, we found that knockdown of CD44 reduced YAP expression and nuclear localization, but nearly had no effect on its upstream effectors, Mst1 and Lats1. Downregulated CD44 expression also significantly decreased the expression of YAP downstream effectors CTGF, Cyr61 and EDN1 at mRNA level. Our next study showed that knockdown of CD44 inhibited RhoA expression, which was consistent with RhoA knockdown mediated YAP downregulation. Furthermore, we demonstrated that over expression of the constitutively active RhoA (RhoA-V14) could block the YAP expression decrease mediated by CD44 knockdown. Moreover, downregulation of CD44 significantly promoted cell apoptosis and inhibited cell proliferation, cell cycle progression and migration, which were consistent with the effects of RNAi-mediated YAP knockdown in both A549 and HepG2 cells. Overall, data are presented showing that CD44 could act through RhoA signaling to regulate YAP expression and this study also provide new insights into the regulatory mechanisms of the Hippo-YAP pathway.


Assuntos
Regulação da Expressão Gênica/fisiologia , Receptores de Hialuronatos/metabolismo , Proteínas Nucleares/biossíntese , Transdução de Sinais/fisiologia , Fatores de Transcrição/biossíntese , Proteína rhoA de Ligação ao GTP/metabolismo , Ciclo Celular/fisiologia , Proteínas de Ciclo Celular , Movimento Celular/fisiologia , Fator de Crescimento do Tecido Conjuntivo/genética , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Proteína Rica em Cisteína 61/genética , Proteína Rica em Cisteína 61/metabolismo , Células Hep G2 , Humanos , Receptores de Hialuronatos/genética , Proteínas Nucleares/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Fatores de Transcrição/genética , Proteína rhoA de Ligação ao GTP/genética
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