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1.
Int J Mol Sci ; 20(18)2019 Sep 09.
Artigo em Inglês | MEDLINE | ID: mdl-31505764

RESUMO

Wnt signaling plays a major role in bone metabolism. Advances in our understanding of secreted regulators of Wnt have yielded several therapeutic targets to stimulate osteoanabolism-the most promising of which is the Wnt inhibitor sclerostin. Sclerostin antibody recently gained approval for clinical use to treat osteoporosis, but the biology surrounding sclerostin antagonism is still incompletely understood. Numerous factors regulate the efficacy of sclerostin inhibition on bone formation, a process known as self-regulation. In previous communications we reported that the basic helix-loop-helix transcription factor Twist1-a gene know to regulate skeletal development-is highly upregulated among the osteocyte cell population in mice treated with sclerostin antibody. In this communication, we tested the hypothesis that preventing Twist1 upregulation by deletion of Twist1 from late-stage osteoblasts and osteocytes would increase the efficacy of sclerostin antibody treatment, since Twist1 is known to restrain osteoblast activity in many models. Twist1-floxed loss-of-function mice were crossed to the Dmp1-Cre driver to delete Twist1 in Dmp1-expressing cells. Conditional Twist1 deletion was associated with a mild but significant increase in bone mass, as assessed by dual energy x-ray absorptiometry (DXA) and microCT (µCT) for many endpoints in both male and female mice. Biomechanical properties of the femur were not affected by conditional mutation of Twist1. Sclerostin antibody improved all bone properties significantly, regardless of Twist1 status, sex, or endpoint examined. No interactions were detected when Twist1 status and antibody treatment were examined together, suggesting that Twist1 upregulation in the osteocyte population is not an endogenous mechanism that restrains the osteoanabolic effect of sclerostin antibody treatment. In summary, Twist1 inhibition in the late-stage osteoblast/osteocyte increases bone mass but does not affect the anabolic response to sclerostin neutralization.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Anticorpos Neutralizantes/farmacologia , Densidade Óssea , Proteínas da Matriz Extracelular/biossíntese , Fêmur/metabolismo , Osteogênese , Proteína 1 Relacionada a Twist/deficiência , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Proteínas da Matriz Extracelular/genética , Feminino , Fêmur/patologia , Deleção de Genes , Masculino , Camundongos , Camundongos Transgênicos , Osteoblastos/metabolismo , Osteoblastos/patologia , Osteócitos/metabolismo , Osteócitos/patologia , Proteína 1 Relacionada a Twist/metabolismo , Microtomografia por Raio-X
2.
Invest Ophthalmol Vis Sci ; 60(12): 3920-3929, 2019 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-31546253

RESUMO

Purpose: The continuous growth of the lens throughout life may contribute to the onset of age-related conditions in the lens (i.e., presbyopia and cataract). Volumetric growth is the result of continuous proliferation of lens epithelial cells (LECs). The driving factors controlling LEC proliferation are not well understood. This study tested the hypothesis that mechanical stretching modulates LEC proliferation. Methods: Biomechanical regulation of LEC proliferation was investigated by culturing whole porcine lenses and connective tissues ex vivo under varying physiologically relevant stretching conditions using a bespoke lens stretching device. Additionally, some lenses were treated with a YAP function inhibitor to determine the Hippo signaling pathway's role in regulating lens growth. Resulting changes in LEC labeling index were analyzed using EdU incorporation and flow cytometry for each lens. Results: LEC proliferation was found to be modulated by mechanical strain. Increasing both the magnitude of static stretching and the stretching frequency in cyclic stretching resulted in a proportional increase in the labeling indices of the LECs. Additionally, treatment with the YAP function inhibitor effectively eliminated this relationship. Conclusions: These data demonstrate that LEC proliferation is regulated in part, by the mechanotransduction of stresses induced in the lens capsule and that YAP plays an important role in mechanosensing. These results have important implications for understanding lens growth and morphogenesis. The model may also be used to identify and evaluate targets for modulating lens growth.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proliferação de Células/fisiologia , Células Epiteliais/citologia , Cristalino/citologia , Mecanotransdução Celular/fisiologia , Fatores de Transcrição/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Animais , Citometria de Fluxo , Microscopia de Fluorescência , Técnicas de Cultura de Órgãos , Fármacos Fotossensibilizantes/farmacologia , Transdução de Sinais/fisiologia , Suínos , Fatores de Transcrição/antagonistas & inibidores , Verteporfina/farmacologia
3.
EBioMedicine ; 46: 54-65, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31324603

RESUMO

BACKGROUND: Esophageal squamous cell carcinoma (ESCC) is one of the most lethal malignancies with poor prognosis. Cancer-testis genes (CTGs) have been vigorously pursued as targets for cancer immunotherapy, but the expressive patterns and functional roles of CTGs remain unclear in ESCC. METHODS: A systematic screening strategy was adopted to screen CTGs in ESCC by integrating multiple public databases and RNA expression microarray data from 119 ESCC subjects. For the newly identified ESCC prognosis-associated CTGs, an independent cohort of 118 patients with ESCC was recruited to validate the relationship via immunohistochemistry. Furthermore, functional assays were performed to determine the underlying mechanisms. FINDINGS: 21 genes were recognized as CTGs, in particular, CDCA5 was aberrantly upregulated in ESCC tissues and significantly associated with poor prognosis (HR = 1.85, 95%CI: 1.14-3.01, P = .013). Immunohistochemical staining confirmed that positive CDCA5 expression was associated with advanced TNM staging and a shorter overall survival rate (45.59% vs 28.00% for CDCA5-/+ subjects, P = 1.86 × 10-3). H3K27 acetylation in CDCA5 promoter might lead to the activation of CDCA5 during ESCC tumorigenesis. Functionally, in vitro assay of gain- and loss-of-function of CDCA5 suggested that CDCA5 could promote ESCC cells proliferation, invasion, migration, apoptosis resistance and reduce chemosensitivity to cisplatin. Moreover, in vivo assay showed that silenced CDCA5 could inhibit tumor growth. Mechanistically, CDCA5 knockdown led to an arrest in G2/M phase and changes in the expression of factors that played fundamental roles in the cell cycle pathway. INTERPRETATION: CDCA5 contributed to ESCC progression and might serve as an attractive target for ESCC immunotherapy. FUND: This work was supported by the Natural Science Foundation of Jiangsu Province (No. BK20181083 and BK20181496), Jiangsu Top Expert Program in Six Professions (No. WSW-003 and WSW-007), Major Program of Science and Technology Foundation of Jiangsu Province (No. BE2016790 and BE2018746), Jiangsu Medical Young Talent Project (No. QNRC2016566), the Program of Jiangsu Medical Innovation Team (No. CXTDA2017006), Postgraduate Research & Practice Innovation Program of Jiangsu Province (KYCX18_1487) and Jiangsu Province 333 Talents Project (No. BRA2017545).


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas de Ciclo Celular/genética , Carcinoma de Células Escamosas do Esôfago/genética , Regulação Neoplásica da Expressão Gênica , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adulto , Idoso , Animais , Antineoplásicos Imunológicos/administração & dosagem , Antineoplásicos Imunológicos/efeitos adversos , Antineoplásicos Imunológicos/uso terapêutico , Proteínas de Ciclo Celular/antagonistas & inibidores , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Carcinoma de Células Escamosas do Esôfago/tratamento farmacológico , Carcinoma de Células Escamosas do Esôfago/metabolismo , Carcinoma de Células Escamosas do Esôfago/patologia , Feminino , Perfilação da Expressão Gênica , Humanos , Imuno-Histoquímica , Masculino , Camundongos , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Transcriptoma
4.
PLoS One ; 14(7): e0217885, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31344028

RESUMO

Induced pluripotent stem cell derived cardiomyocytes (iPSC-CM) have the potential to transform regenerative cardiac medicine and the modelling of cardiac disease. This is of particular importance in the context of diabetic cardiomyopathy where diabetic individuals exhibit reduced cardiac diastolic contractile performance in the absence of vascular disease, significantly contributing towards high cardiovascular morbidity. In this study, the capacity of iPSC-CM to act as a novel cellular model of cardiomyocytes was assessed. The diabetic phenotype is characterised by insulin resistance, therefore there was a specific focus upon metabolic parameters. Despite expressing crucial insulin signalling intermediates and relevant trafficking proteins, it was identified that iPSC-CM do not exhibit insulin-stimulated glucose uptake. iPSC-CM are spontaneously contractile however contraction mediated uptake was not found to mask any insulin response. The fundamental limitation identified in these cells was a critical lack of expression of the insulin sensitive glucose transporter GLUT4. Using comparative immunoblot analysis and the GLUT-selective inhibitor BAY-876 to quantify expression of these transporters, we show that iPSC-CM express high levels of GLUT1 and low levels of GLUT4 compared to primary cardiomyocytes and cultured adipocytes. Interventions to overcome this limitation were unsuccessful. We suggest that the utility of iPSC-CMs to study cardiac metabolic disorders may be limited by their apparent foetal-like phenotype.


Assuntos
Regulação da Expressão Gênica , Transportador de Glucose Tipo 4/biossíntese , Glucose/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Miócitos Cardíacos/metabolismo , Células 3T3-L1 , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal/biossíntese , Animais , Transporte Biológico Ativo/efeitos dos fármacos , Diabetes Mellitus/metabolismo , Diabetes Mellitus/patologia , Transportador de Glucose Tipo 4/antagonistas & inibidores , Humanos , Células-Tronco Pluripotentes Induzidas/patologia , Resistência à Insulina , Camundongos , Contração Miocárdica/efeitos dos fármacos , Miócitos Cardíacos/patologia , Pirazóis/farmacologia , Quinolinas/farmacologia , Coelhos
5.
Immunohorizons ; 3(5): 172-185, 2019 05 16.
Artigo em Inglês | MEDLINE | ID: mdl-31356171

RESUMO

Cytokine IL-17A (IL-17) acts on various cell types, including epidermal keratinocytes, and induces antimicrobial peptide and chemokine production to elicit antibacterial and antifungal defense responses. Excess IL-17 leads to inflammatory skin diseases such as psoriasis. The IκB family protein IκB-ζ mediates IL-17-induced responses. However, the mechanism controlling IκB-ζ expression in IL-17-stimulated cells remains elusive. In this study, we showed that JAK kinase TYK2 positively regulates IL-17-induced IκB-ζ expression. TYK2-deficient mice showed reduced inflammation and concomitant reduction of IκB-ζ mRNA compared with wild-type mice in imiquimod-induced skin inflammation. The analysis of the IκB-ζ promoter activity using human cell lines (HaCaT and HeLa) revealed that catalytic activity of TYK2 and its substrate transcription factor STAT3, but not IL-17, is required for IκB-ζ promoter activity. In contrast, IL-17-induced signaling, which did not activate STAT3, posttranscriptionally stabilized IκB-ζ mRNA via its 3'-untranslated region. IL-17 signaling protein ACT1 was required to counteract constitutive IκB-ζ mRNA degradation by RNase Regnase-1. These results suggested that transcriptional activation by TYK2-STAT3 pathway and mRNA stabilization by IL-17-mediated signals act separately from each other but complementarily to achieve IκB-ζ induction. Therefore, JAK/TYK2 inhibition might be of significance in regulation of IL-17-induced inflammatory reactions.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Interleucina-17/metabolismo , Estabilidade de RNA , Fator de Transcrição STAT3/metabolismo , TYK2 Quinase/metabolismo , Regiões 3' não Traduzidas , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Animais , Modelos Animais de Doenças , Técnicas de Inativação de Genes , Células HeLa , Humanos , Queratinócitos/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Inibidores de Proteínas Quinases/farmacologia , Psoríase/induzido quimicamente , Psoríase/metabolismo , RNA Mensageiro/metabolismo , Ribonucleases/metabolismo , TYK2 Quinase/genética , Fatores de Transcrição/metabolismo
6.
Gene ; 712: 143956, 2019 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-31271843

RESUMO

Gastric cancer represents a common malignancy of digestive tract with high incidence and mortality. Increasing evidence suggests that the growth of gastric tumor cells relies largely on aerobic glycolysis. Currently, many potential anti-cancer candidates are derived from natural products. Here, we evaluated the effects of oleanolic acid (OA), a triterpenoid component widely found in the plants of Oleaceae family, on aerobic glycolysis and proliferation in human MKN-45 and SGC-7901 gastric cancer cells. Our results demonstrated that OA reduced the viability and proliferation of gastric cancer cells and inhibited the expression of cyclin A and cyclin-dependent kinase 2. OA blocked glycolysis in these cells evidenced by decreases in the uptake and consumption of glucose, intracellular lactate levels and extracellular acidification rate. Glycolysis inhibitor 2-deoxy-d-glucose, similar to OA, suppressed gastric cancer cell proliferation. OA also decreased the expression and intracellular activities of glycolysis rate-limiting enzymes hexokinase 2 (HK2) and phosphofructokinase 1 (PFK1). Moreover, OA downregulated the expression of hypoxia inducible factor-1α (HIF-1α) and decreased its nuclear abundance. Upregulation of HIF-1α by deferoxamine rescued OA-inhibited HK2 and PFK1. Furthermore, OA reduced the nuclear abundance of yes-associated protein (YAP) in gastric tumor cells. YAP inhibitor verteporfin, similar to OA, downregulated the expression of HIF-1α and glycolytic enzymes in gastric cancer cells; whereas overexpression of YAP abrogated all these effects of OA. Collectively, inhibition of YAP was responsible for OA blockade of HIF-1α-mediated aerobic glycolysis and proliferation in human gastric tumor cells. OA could be developed as a promising candidate for gastric cancer treatment.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Ácido Oleanólico/farmacologia , Fosfoproteínas/antagonistas & inibidores , Fosfoproteínas/metabolismo , Neoplasias Gástricas/metabolismo , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células , Sobrevivência Celular , Ciclo do Ácido Cítrico , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glicólise , Humanos , Fosforilação , Transdução de Sinais/efeitos dos fármacos , Estômago/patologia , Neoplasias Gástricas/tratamento farmacológico , Fatores de Transcrição
7.
Arterioscler Thromb Vasc Biol ; 39(7): 1343-1350, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31242037

RESUMO

Osteoporosis and cardiovascular diseases are major public health issues. Bone and cardiovascular remodeling share multiple biological markers and pathways. Medical intervention, such as using romosozumab, an antisclerostin antibody, improves the clinical outcome of osteoporosis. However, blocking sclerostin leads to Wnt (wingless/integrated) activation and participation in the cardiovascular remodeling process, which could potentially lead to adverse events. Based on the opposing roles of bisphosphonates and the Wnt pathway on endothelial dysfunction, lipid accumulation and calcification of the vessel walls, the combination of romosozumab and bisphosphonates could be a new therapeutic approach to reducing the risks of adverse cardiovascular events in romosozumab receivers. Visual Overview- An online visual overview is available for this article.


Assuntos
Alendronato/farmacologia , Anticorpos Monoclonais/efeitos adversos , Doenças Cardiovasculares/induzido quimicamente , Osteoporose/tratamento farmacológico , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Humanos , Via de Sinalização Wnt/efeitos dos fármacos
8.
Oncol Rep ; 42(2): 697-707, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31233205

RESUMO

Human non­small cell lung cancer (NSCLC) is associated with an extremely poor prognosis especially for the 40% of patients who develop brain metastasis, and few treatment strategies exist. Cucurbitacin E (CuE), an oxygenated tetracyclic triterpenoid isolated from plants particularly of the family Cucurbitaceae, has shown anti­tumorigenic properties in several types of cancer, yet the mechanism remains unclear. Yes­associated protein (YAP), a main mediator of the Hippo signaling pathway, promotes tumorigenesis, drug resistance and metastasis in human NSCLC. The present study was designed to ascertain whether CuE inhibits YAP and its downstream gene expression in the human NSCLC cell lines H2030­BrM3 (K­rasG12C mutation) and PC9­BrM3 (EGFRΔexon19 mutation), which have high potential for brain metastasis. The efficacy of CuE in suppressing brain metastasis of H2030­BrM3 cells in a murine model was also investigated. It was found that after CuE treatment in H2030­BrM3 and PC9­BrM3 cells, YAP protein expression was decreased, and YAP signaling GTIIC reporter activity and expression of the downstream genes CTGF and CYR61 were significantly (P<0.01) decreased. CuE treatment also reduced the migration and invasion abilities of the H2030­BrM3 and PC9­BrM3 cells. Finally, our in vivo study showed that CuE treatment (0.2 mg/kg) suppressed H2030­BrM3 cell brain metastasis and that mice treated with CuE survived longer than the control mice treated with 10% DMSO (P=0.02). The present study is the first to demonstrate that CuE treatment inhibits YAP and the signaling downstream gene expression in human NSCLC in vitro, and suppresses brain metastasis of NSCLC in a murine model. More studies to verify the promising efficacy of CuE in inhibiting brain metastasis of NSCLC and various other cancers may be warranted.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Neoplasias Encefálicas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Neoplasias Pulmonares/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Fatores de Transcrição/antagonistas & inibidores , Triterpenos/farmacologia , Animais , Apoptose , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/secundário , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Movimento Celular , Proliferação de Células , Feminino , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Nus , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
9.
J Exp Clin Cancer Res ; 38(1): 211, 2019 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-31118055

RESUMO

BACKGROUND: Epidermal growth factor receptor (EGFR) and epidermal growth factor receptor pathway substrate 8 (Eps8) have been widely reported to be expressed in various tumors. Eps8 is an important active kinase substrate of EGFR that directly binds to the juxtamembrane (JXM) domain of EGFR to form an EGFR/Eps8 complex. The EGFR/Eps8 complex is involved in regulating cancer progression and might be an ideal target for antitumor therapy. This study focused on the screening of small-molecule inhibitors that target the EGFR/Eps8 complex in breast cancer and non-small cell lung cancer (NSCLC). METHODS: In silico virtual screening was used to identify small-molecule EGFR/Eps8 complex inhibitors. These compounds were screened for the inhibition of A549 and BT549 cell viability. The direct interaction between EGFR and Eps8 was measured using coimmunoprecipitation (CoIP) and JXM domain replacement assays. The antitumor effects of the inhibitors were analyzed in cancer cells and xenograft models. An acute toxicity study of EE02 was performed in a mouse model. In addition, the effect of the EE02 inhibitor on the protein expression of elements downstream of the EGFR/Eps8 complex was determined by western blotting and protein chip assays. RESULTS: In this study of nearly 390,000 compounds screened by virtual database screening, the top 29 compounds were identified as candidate small-molecule EGFR/Eps8 complex inhibitors and evaluated by using cell-based assays. The compound EE02 was identified as the best match to our selection criteria. Further investigation demonstrated that EE02 directly bound to the JXM domain of EGFR and disrupted EGFR/Eps8 complex formation. EE02 selectively suppressed growth and induced apoptosis in EGFR-positive and Eps8-positive breast cancer and NSCLC cells. More importantly, the PI3K/Akt/mTOR and MAPK/Erk pathways downstream of the EGFR/Eps8 complex were suppressed by EE02. In addition, the suppressive effect of EE02 on tumor growth in vivo was comparable to that of erlotinib at the same dose. CONCLUSIONS: We identified EE02 as an EGFR/Eps8 complex inhibitor that demonstrated promising antitumor effects in breast cancer and NSCLC. Our data suggest that the EGFR/Eps8 complex offers a novel cancer drug target.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Neoplasias da Mama/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Complexos Multiproteicos/antagonistas & inibidores , Células A549 , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal/química , Animais , Apoptose/efeitos dos fármacos , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Sobrevivência Celular/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Sinergismo Farmacológico , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/química , Receptores ErbB/genética , Cloridrato de Erlotinib/farmacologia , Feminino , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Camundongos , Complexos Multiproteicos/genética , Inibidores de Proteínas Quinases/farmacologia , Transdução de Sinais/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
10.
PLoS One ; 14(5): e0216603, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31100067

RESUMO

We examined the impact of statins on Yes-associated Protein (YAP) localization, phosphorylation and transcriptional activity in human and mouse pancreatic ductal adenocarcinoma (PDAC) cells. Exposure of sparse cultures of PANC-1 and MiaPaCa-2 cells to cerivastatin or simvastatin induced a striking re-localization of YAP from the nucleus to the cytoplasm and inhibited the expression of the YAP/TEAD-regulated genes Connective Tissue Growth Factor (CTGF) and Cysteine-rich angiogenic inducer 61 (CYR61). Statins also prevented YAP nuclear import and expression of CTGF and CYR61 stimulated by the mitogenic combination of insulin and neurotensin in dense culture of these PDAC cells. Cerivastatin, simvastatin, atorvastatin and fluvastatin also inhibited colony formation by PANC-1 and MiaPaCa-2 cells in a dose-dependent manner. In contrast, the hydrophilic statin pravastatin did not exert any inhibitory effect even at a high concentration (10 µM). Mechanistically, cerivastatin did not alter the phosphorylation of YAP at Ser127 in either PANC-1 or MiaPaCa-2 cells incubated without or with neurotensin and insulin but blunted the assembly of actin stress fiber in these cells. We extended these findings with human PDAC cells using primary KC and KPC cells, (expressing KrasG12D or both KrasG12D and mutant p53, respectively) isolated from KC or KPC mice. Using cultures of these murine cells, we show that lipophilic statins induced striking YAP translocation from the nucleus to the cytoplasm, inhibited the expression of Ctgf, Cyr61 and Birc5 and profoundly inhibited colony formation of these cells. Administration of simvastatin to KC mice subjected to diet-induced obesity prevented early pancreatic acini depletion and PanIN formation. Collectively, our results show that lipophilic statins restrain YAP activity and proliferation in pancreatic cancer cell models in vitro and attenuates early lesions leading to PDAC in vivo.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Carcinoma Ductal Pancreático/prevenção & controle , Núcleo Celular/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Neoplasias Pancreáticas/prevenção & controle , Proteínas Proto-Oncogênicas p21(ras)/genética , Fatores de Transcrição/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patologia , Proliferação de Células , Ensaio de Unidades Formadoras de Colônias , Humanos , Camundongos , Mutação , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Fosforilação , Transporte Proteico , Fatores de Transcrição/metabolismo , Células Tumorais Cultivadas
11.
Arterioscler Thromb Vasc Biol ; 39(7): 1330-1342, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31092014

RESUMO

Clinical and preclinical studies over the past 3 decades have uncovered a multitude of signaling pathways involved in the initiation and progression of atherosclerosis. From these studies, signaling by proteins of the Wnt family has recently emerged as an important player in the development of atherosclerosis. Wnt signaling is characterized by a large number of ligands, receptors, and coreceptors and can be regulated at many different levels. Among Wnt modulators, the evolutionary conserved Dkk (Dickkopf) proteins, and especially Dkk-1, the founding member of the family, are the best characterized. The role of Dkks in the pathophysiology of the arterial wall is only partially understood, but their involvement in atherosclerosis is becoming increasingly evident. This review introduces recent key findings on Dkk proteins and their functions in atherosclerosis and discusses the potential importance of modulating Dkk signaling as part of a novel, improved strategy for preventing and treating atherosclerosis-related diseases. Visual Overview- An online visual overview is available for this article.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Aterosclerose/etiologia , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Animais , Apolipoproteínas E/fisiologia , Aterosclerose/tratamento farmacológico , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Calcificação Vascular/etiologia , Via de Sinalização Wnt/fisiologia , beta Catenina/fisiologia
12.
Future Oncol ; 15(13): 1535-1543, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31066301

RESUMO

YAP, acting as a crucial transcription factor in nucleus, regulates the organ size, tissue homeostasis and tumorigenesis. Dysregulation of Hippo-YAP pathway brings a significant impact on the occurrence and development of various tumor types. Moreover, regulation of YAP/TAZ far exceeds the core kinase of the Hippo pathway, and gradually opens up new therapeutic targets. For the moment, chemotherapy together with radiotherapy act as routine methods to prolong the lives of cancer patients. Seeking more effective anti-neoplastic agents seems to be the urgent problem. This brief review focuses on the research progress of YAP inhibitors as the antineoplastic targets. Small molecule inhibitors or drugs have been discovered including verteporfin, dasatinib, statins, A35, JQ1, norcantharidin, agave, MLN8237, dobutamine and peptide-based YAP inhibitors. We are trying to seek novel therapies from the relationship between known drugs and potential mechanisms.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Antineoplásicos/uso terapêutico , Terapia de Alvo Molecular , Neoplasias/tratamento farmacológico , Fosfoproteínas/antagonistas & inibidores , Humanos , Neoplasias/metabolismo , Neoplasias/patologia , Prognóstico , Fatores de Transcrição
13.
Dev Cell ; 49(3): 425-443.e9, 2019 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-31063758

RESUMO

Merlin/NF2 is a bona fide tumor suppressor whose mutations underlie inherited tumor syndrome neurofibromatosis type 2 (NF2), as well as various sporadic cancers including kidney cancer. Multiple Merlin/NF2 effector pathways including the Hippo-YAP/TAZ pathway have been identified. However, the molecular mechanisms underpinning the growth and survival of NF2-mutant tumors remain poorly understood. Using an inducible orthotopic kidney tumor model, we demonstrate that YAP/TAZ silencing is sufficient to induce regression of pre-established NF2-deficient tumors. Mechanistically, YAP/TAZ depletion diminishes glycolysis-dependent growth and increases mitochondrial respiration and reactive oxygen species (ROS) buildup, resulting in oxidative-stress-induced cell death when challenged by nutrient stress. Furthermore, we identify lysosome-mediated cAMP-PKA/EPAC-dependent activation of RAF-MEK-ERK signaling as a resistance mechanism to YAP/TAZ inhibition. Finally, unbiased analysis of TCGA primary kidney tumor transcriptomes confirms a positive correlation of a YAP/TAZ signature with glycolysis and inverse correlations with oxidative phosphorylation and lysosomal gene expression, supporting the clinical relevance of our findings.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Carcinoma de Células Renais/metabolismo , Neoplasias Renais/metabolismo , Neurofibromina 2/deficiência , Fatores de Transcrição/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/patologia , Linhagem Celular Tumoral , Glicólise , Xenoenxertos , Humanos , Neoplasias Renais/genética , Neoplasias Renais/patologia , Sistema de Sinalização das MAP Quinases , Camundongos , Camundongos SCID , Neurofibromatose 2/genética , Neurofibromatose 2/metabolismo , Neurofibromatose 2/patologia , Neurofibromina 2/genética , Neurofibromina 2/metabolismo , Fosforilação Oxidativa , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/genética
14.
Int J Mol Sci ; 20(10)2019 May 16.
Artigo em Inglês | MEDLINE | ID: mdl-31100813

RESUMO

The L-type calcium channel blocker fendiline has been shown to interfere with Ras-dependent signaling in K-Ras mutant cancer cells. Earlier studies from our lab had shown that treatment of pancreatic cancer cells with fendiline causes significant cytotoxicity and interferes with proliferation, survival, migration, invasion and anchorage independent growth. Currently there are no effective therapies to manage PDACs. As fendiline has been approved for treatment of patients with angina, we hypothesized that, if proven effective, combinatorial therapies using this agent would be easily translatable to clinic for testing in PDAC patients. Here we tested combinations of fendiline with gemcitabine, visudyne (a YAP1 inhibitor) or tivantinib (ARQ197, a c-Met inhibitor) for their effectiveness in overcoming growth and oncogenic characteristics of PDAC cells. The Hippo pathway component YAP1 has been shown to bypass K-Ras addiction, and allow tumor growth, in a Ras-null mouse model. Similarly, c-Met expression has been associated with poor prognosis and metastasis in PDAC patients. Our results presented here show that combinations of fendiline with these inhibitors show enhanced anti-tumor activity in Panc1, MiaPaCa2 and CD18/HPAF PDAC cells, as evident from the reduced viability, migration, anchorage-independent growth and self-renewal. Biochemical analysis shows that these agents interfere with various signaling cascades such as the activation of Akt and ERK, as well as the expression of c-Myc and CD44 that are altered in PDACs. These results imply that inclusion of fendiline may improve the efficacy of various chemotherapeutic agents that could potentially benefit PDAC patients.


Assuntos
Antineoplásicos/farmacologia , Carcinoma Ductal Pancreático/tratamento farmacológico , Desoxicitidina/análogos & derivados , Fendilina/farmacologia , Pirrolidinonas/farmacologia , Quinolinas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Verteporfina/farmacologia , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Carcinógenos , Ciclo Celular/efeitos dos fármacos , Proteínas de Ciclo Celular , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Desoxicitidina/farmacologia , Modelos Animais de Doenças , Humanos , Concentração Inibidora 50 , Camundongos , Metástase Neoplásica , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/patologia , Fosfoproteínas/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-met/antagonistas & inibidores
15.
World J Gastroenterol ; 25(15): 1797-1816, 2019 Apr 21.
Artigo em Inglês | MEDLINE | ID: mdl-31057295

RESUMO

Pancreatic ductal adenocarcinoma (PDAC) remains a deadly disease with no efficacious treatment options. PDAC incidence is projected to increase, which may be caused at least partially by the obesity epidemic. Significantly enhanced efforts to prevent or intercept this cancer are clearly warranted. Oncogenic KRAS mutations are recognized initiating events in PDAC development, however, they are not entirely sufficient for the development of fully invasive PDAC. Additional genetic alterations and/or environmental, nutritional, and metabolic signals, as present in obesity, type-2 diabetes mellitus, and inflammation, are required for full PDAC formation. We hypothesize that oncogenic KRAS increases the intensity and duration of the growth-promoting signaling network. Recent exciting studies from different laboratories indicate that the activity of the transcriptional co-activators Yes-associated protein (YAP) and WW-domain-containing transcriptional co-activator with PDZ-binding motif (TAZ) play a critical role in the promotion and maintenance of PDAC operating as key downstream target of KRAS signaling. While initially thought to be primarily an effector of the tumor-suppressive Hippo pathway, more recent studies revealed that YAP/TAZ subcellular localization and co-transcriptional activity is regulated by multiple upstream signals. Overall, YAP has emerged as a central node of transcriptional convergence in growth-promoting signaling in PDAC cells. Indeed, YAP expression is an independent unfavorable prognostic marker for overall survival of PDAC. In what follows, we will review studies implicating YAP/TAZ in pancreatic cancer development and consider different approaches to target these transcriptional regulators.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Carcinoma Ductal Pancreático/genética , Regulação Neoplásica da Expressão Gênica , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias Pancreáticas/genética , Fosfoproteínas/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Animais , Carcinogênese/efeitos dos fármacos , Carcinogênese/genética , Carcinogênese/patologia , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/mortalidade , Carcinoma Ductal Pancreático/patologia , Reposicionamento de Medicamentos , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Metformina/farmacologia , Metformina/uso terapêutico , Terapia de Alvo Molecular/métodos , Mutação , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/mortalidade , Neoplasias Pancreáticas/patologia , Fosfoproteínas/antagonistas & inibidores , Prognóstico , Proteínas Proto-Oncogênicas p21(ras)/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Fatores de Transcrição , Transcrição Genética/efeitos dos fármacos , Transcrição Genética/genética
16.
Neoplasia ; 21(6): 533-544, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31029033

RESUMO

Treatment options are limited for patients with triple negative breast cancer (TNBC). Understanding genes that participate in cancer progression and DNA damage response (DDR) may improve therapeutic strategies for TNBC. DAXX, a death domain-associated protein, has been reported to be critically involved in cancer progression and drug sensitivity in multiple cancer types. However, its role in breast cancer, especially for TNBC, remains unclear. Here, we demonstrated a tumor suppressor function of DAXX in TNBC proliferation, colony formation, and migration. In Mouse Xenograft Models, DAXX remarkably inhibited tumorigenicity of TNBC cells. Mechanistically, DAXX could directly bind to the promoter region of RAD51 and impede DNA damage repair, which impacted the protection mechanism of tumor cells that much depended on remaining DDR pathways for cell growth. Furthermore, DAXX-mediated inefficient DNA damage repair could sensitize BRCA-proficient TNBC cells to PARP inhibitors. Additionally, we identified that dual RAD51 and PARP inhibition with RI-1 and ABT888 significantly reduced TNBC growth both in vitro and in vivo, which provided the first evidence of combining RAD51 and PARP inhibition in BRCA-proficient TNBC. In conclusion, our data support DAXX as a modulator of DNA damage repair and suppressor of TNBC progression to sensitize tumors to the PARP inhibitor by repressing RAD51 functions. These provide an effective strategy for a better application of PARP inhibition in the treatment of TNBC.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Nucleares/genética , Poli(ADP-Ribose) Polimerase-1/genética , Rad51 Recombinase/genética , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Proteínas Supressoras de Tumor/genética , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Animais , Benzimidazóis/farmacologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Reparo do DNA/efeitos dos fármacos , Feminino , Xenoenxertos , Humanos , Camundongos , Morfolinas/farmacologia , Proteínas Nucleares/antagonistas & inibidores , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Regiões Promotoras Genéticas/efeitos dos fármacos , Pirróis/farmacologia , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/patologia , Proteínas Supressoras de Tumor/antagonistas & inibidores
17.
PLoS One ; 14(4): e0214822, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30939162

RESUMO

Cervical cancer is the second most deadly gynecological tumor worldwide. MicroRNAs (miRNAs) play very important roles in tumor oncogenesis and progression. The mechanism of post-transcription regulation of WTX gene is still unknown. A series of differential miRNAs were discovered by microarray analysis comparing three pairs of primary cervical cancer specimens and their relapsed tumors from three patients. Quantitative reverse transcriptase PCR (qRT-PCR), Western Blot (WB) and Immunohistochemistry (IHC) was used to detect the expression of miR-4524b-5p and WTX in cervical cell lines and tissues. The biological function of miR-4524b-5p and WTX was investigated through knockdown and overexpression with inhibitor/siRNA and mimic/plasmid in vitro and in vivo. In this study, we found that miR-4524b-5p is highly expressed in relapsed cervical cancer specimens. Combined in vitro and in vivo experiments, showed that miR-4524b-5p could regulate the migration and invasion ability of cervical cancer. Furthermore, we also found that miR-4524b-5p could regulate the migration and invasion of cervical cancer by targeting WTX and that WTX could regulate the expression of ß-catenin. Taken together, our data identified a miR-4524b-5p/WTX/ß-catenin regulatory axis for cervical cancer, and miR-4524b-5p may be a potential target for cervical cancer therapy.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , MicroRNAs/genética , Proteínas Supressoras de Tumor/genética , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/patologia , beta Catenina/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Animais , Linhagem Celular Tumoral , Movimento Celular/genética , Feminino , Técnicas de Silenciamento de Genes , Xenoenxertos , Humanos , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/metabolismo , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Metástase Neoplásica/genética , Metástase Neoplásica/patologia , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/metabolismo , Recidiva Local de Neoplasia/patologia , Transplante de Neoplasias , Proteínas Supressoras de Tumor/antagonistas & inibidores , Neoplasias do Colo do Útero/metabolismo
18.
Ann Vasc Surg ; 59: 244-247, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31009712

RESUMO

BACKGROUND: Inflammation and endothelial dysfunction are implicated in the onset of atherosclerosis. Inflammasome activation takes part in the pathogenesis of the atherosclerotic disease. This study investigated the influence of platelet inflammatory inhibition on the transcription of intracitosolic nucleotide-binding oligomerization domain-like receptor protein 1 (NLRP-1) inflammasome in endothelial cells. METHODS: This experimental study enrolled 10 healthy volunteers with no cardiovascular risk factors and normal results on vascular examination. They received low doses of aspirin (100 mg/day) for seven days. A venous blood sample was collected in all subjects before aspirin intake and after the experimental week. Human aortic endothelial cell (HAEC) cultures were exposed to baseline plasma and plasma from subjects after aspirin intake. NLRP-1 gene expression was analyzed in these cultures. RESULTS: HAEC cultures that were exposed to plasma from subjects at baseline showed higher expression of NLRP-1 than HAECs exposed to plasma of healthy volunteers after one week on salicilate intake (relative quantification, 1.077 ± 0.05 vs. 1.002 ± 0.06; odds ratio, 1.8; 95 confidence interval, 1.1-2.9; P < 0.01). CONCLUSIONS: Data observed in the our study indicate that in HAECs, the intracytosolic NLRP-1 expression is attenuated by the auto/paracrine platelet inhibition by aspirin, without direct platelet-endothelial cell interaction.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Anti-Inflamatórios/administração & dosagem , Proteínas Reguladoras de Apoptose/antagonistas & inibidores , Aspirina/administração & dosagem , Plaquetas/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Inflamassomos/antagonistas & inibidores , Inibidores da Agregação de Plaquetas/administração & dosagem , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Plaquetas/metabolismo , Comunicação Celular/efeitos dos fármacos , Células Cultivadas , Estudos Clínicos como Assunto , Regulação para Baixo , Células Endoteliais/metabolismo , Voluntários Saudáveis , Humanos , Inflamassomos/metabolismo , Transdução de Sinais/efeitos dos fármacos
19.
Nat Commun ; 10(1): 1464, 2019 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-30931946

RESUMO

Stem cell-derived insulin-producing beta cells (SC-ß) offer an inexhaustible supply of functional ß cells for cell replacement therapies and disease modeling for diabetes. While successful directed differentiation protocols for this cell type have been described, the mechanisms controlling its differentiation and function are not fully understood. Here we report that the Hippo pathway controls the proliferation and specification of pancreatic progenitors into the endocrine lineage. Downregulation of YAP, an effector of the pathway, enhances endocrine progenitor differentiation and the generation of SC-ß cells with improved insulin secretion. A chemical inhibitor of YAP acts as an inducer of endocrine differentiation and reduces the presence of proliferative progenitor cells. Conversely, sustained activation of YAP results in impaired differentiation, blunted glucose-stimulated insulin secretion, and increased proliferation of SC-ß cells. Together these results support a role for YAP in controlling the self-renewal and differentiation balance of pancreatic progenitors and limiting endocrine differentiation in vitro.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Diferenciação Celular/genética , Secreção de Insulina/genética , Células Secretoras de Insulina/citologia , Fosfoproteínas/genética , Células-Tronco Pluripotentes/citologia , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Diferenciação Celular/efeitos dos fármacos , Linhagem da Célula , Regulação para Baixo , Células HEK293 , Humanos , Imuno-Histoquímica , Secreção de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/efeitos dos fármacos , Fosfoproteínas/antagonistas & inibidores , Células-Tronco Pluripotentes/efeitos dos fármacos , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais , Fatores de Transcrição
20.
Drug Des Devel Ther ; 13: 1011-1022, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30992659

RESUMO

Background: KM-819 is a novel FAS-associated factor 1 (FAF1) inhibitor, and a neuroprotective agent, under clinical development for the treatment of Parkinson's disease as a disease-modifying drug. Methods: This first-in-human, single and multiple ascending dose study investigated the safety, tolerability, pharmacokinetics, and pharmacodynamics of KM-819 in healthy volunteers. Additionally, the effect of age on safety and pharmacokinetics were assessed. The starting dose was determined considering the no observed adverse effect level based on preclinical studies, and the dose escalations in subsequent cohorts were decided based on safety, tolerability, and pharmacokinetic data from previous dose cohorts. Results: After a single dose, the KM-819 plasma exposure showed a less than dose-proportional increase across a dose range of 10-400 mg. After repeated dosing, KM-819 plasma exposure increased in an approximately dose-proportional manner across the evaluated dose range (30-400 mg once daily for 7 days). The mean elimination half-life was 1.8 to 4.8 h with the lower KM-819 doses (≤30 mg), which increased to around 9 h with the higher doses (100-400 mg). When administered to the elderly population, KM-819 plasma exposure increased to 102% after a 200 mg once-daily dosing for 7 days. No clear treatment-related effects on the estimated pharmacodynamic variables were observed. Single or multiple doses of KM-819 were generally well tolerated. Conclusion: The data from this study can be used to guide rational drug dosing and choose therapeutic regimens in subsequent clinical studies.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Fármacos Neuroprotetores/efeitos adversos , Fármacos Neuroprotetores/farmacocinética , Compostos Orgânicos/farmacocinética , Doença de Parkinson/tratamento farmacológico , Administração Oral , Adulto , Relação Dose-Resposta a Droga , Esquema de Medicação , Tolerância a Medicamentos , Voluntários Saudáveis , Humanos , Masculino , Fármacos Neuroprotetores/administração & dosagem , Compostos Orgânicos/administração & dosagem , Compostos Orgânicos/química
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