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1.
Nat Commun ; 11(1): 84, 2020 01 03.
Artigo em Inglês | MEDLINE | ID: mdl-31901081

RESUMO

Areas of a junction between two types of epithelia are known to be cancer-prone in many organ systems. However, mechanisms for preferential malignant transformation at the junction areas remain insufficiently elucidated. Here we report that inactivation of tumor suppressor genes Trp53 and Rb1 in the gastric squamous-columnar junction (SCJ) epithelium results in preferential formation of metastatic poorly differentiated neoplasms, which are similar to human gastroesophageal carcinoma. Unlike transformation-resistant antral cells, SCJ cells contain a highly proliferative pool of immature Lgr5-CD44+ cells, which are prone to transformation in organoid assays, comprise early dysplastic lesions, and constitute up to 30% of all neoplastic cells. CD44 ligand osteopontin (OPN) is preferentially expressed in and promotes organoid formation ability and transformation of the SCJ glandular epithelium. OPN and CD44 overexpression correlate with the worst prognosis of human gastroesophageal carcinoma. Thus, detection and selective targeting of the active OPN-CD44 pathway may have direct clinical relevance.


Assuntos
Carcinoma de Células Escamosas/metabolismo , Junção Esofagogástrica/metabolismo , Receptores de Hialuronatos/metabolismo , Osteopontina/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Neoplasias Gástricas/metabolismo , Idoso , Idoso de 80 Anos ou mais , Animais , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/mortalidade , Carcinoma de Células Escamosas/patologia , Transformação Celular Neoplásica , Estudos de Coortes , Junção Esofagogástrica/patologia , Feminino , Humanos , Receptores de Hialuronatos/genética , Masculino , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Osteopontina/genética , Receptores Acoplados a Proteínas-G/genética , Proteína do Retinoblastoma/genética , Proteína do Retinoblastoma/metabolismo , Neoplasias Gástricas/genética , Neoplasias Gástricas/mortalidade , Neoplasias Gástricas/patologia , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
2.
Anticancer Res ; 39(11): 6291-6297, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31704859

RESUMO

BACKGROUND/AIM: Cyclins D1 and E play different roles in the cell cycle. Cyclin E promotes chromosome instability, whereas cyclin D1 regulates apoptosis of cells. This study evaluated the prognostic significance of G1 cyclins, p21 and pRb in tumor proliferation. PATIENTS AND METHODS: A total of 102 patients with colon cancer were operated on and staged according to TNM. Follow-up was 2 to 68 months (mean 38.3±16.7 months). Expression of cyclin E and D1 were evaluated using immunohistochemistry. RESULTS: Levels of cyclin E expression were correlated with cyclin D1 expression (p=0.038), p21 expression (p=0.047), and pRb expression (p=0.004). The 5-year survival rate along with prognosis of patients with advanced stage (III, IV) colon cancer and cyclin D1 positive tumors, were significantly worse (p=0.009). Statistically significant association was observed between tumor proliferative capacity Ki-67, cyclin D1 (p=0.009), pRb (p=0.031) and p21 (p=0.050). CONCLUSION: Cyclin D1 is highly expressed in advanced stage colon cancer patients, implying a potential prognostic value.


Assuntos
Neoplasias do Colo/metabolismo , Ciclina D1/metabolismo , Ciclina E/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Fase G1 , Proteínas de Neoplasias/metabolismo , Proteínas Oncogênicas/metabolismo , Proteína do Retinoblastoma/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias do Colo/química , Neoplasias do Colo/mortalidade , Neoplasias do Colo/patologia , Ciclina D1/análise , Ciclina E/análise , Inibidor de Quinase Dependente de Ciclina p21/análise , Feminino , Seguimentos , Humanos , Antígeno Ki-67/análise , Antígeno Ki-67/metabolismo , Masculino , Pessoa de Meia-Idade , Proteínas Oncogênicas/análise , Prognóstico , Proteína do Retinoblastoma/análise , Taxa de Sobrevida
3.
Artif Cells Nanomed Biotechnol ; 47(1): 4001-4011, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31588803

RESUMO

Triple-negative breast cancer (TNBC) stands for a refractory subtype, which predicts poor prognosis and has no effective therapies yet for improving it. Given the restrictions of traditional treatments, novel therapeutic strategies need excavating to alleviate the intrinsic or acquired resistance. Ribociclib, a selective CDK4/6 inhibitor, has successfully prevented cancers from deteriorating by intervening the CDK4/6-cyclin D-Rb-E2F pathway, especially for estrogen receptor-positive (ER +) breast cancer. However, there still remains limited accessibility referring to TNBC. Performing experiments on MDA-MB-231 cells, we found that LEE011 could suppress cell proliferation, and this suppression tended to be dose-dependently. Western blotting analysis presented significant decrease with the expression of CDK4/6 after LEE011 treated, and other proteins associated with this axis such as cyclin D1, p-Rb, Rb, E2F1 showed aberrant changes. Moreover, LEE011 induced G0-G1 phase cell cycle arrest, promoted cell apoptosis, and reduced cell migration in vitro. In addition, tumor growth was remarkably impeded without obvious side-effects in MDA-MB-231 xenograft models. Our research has identified that LEE011 was not completely invalid for MDA-MB-231. Considering its pivotal status in TNBC, the CDK4/6-cyclin D-Rb-E2F pathway informed us the possibility and practicality of Ribociclib (LEE011) as pharmacological intervention, but challenges warrant further validation in prospective studies.


Assuntos
Aminopiridinas/farmacologia , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Proteínas Inibidoras de Quinase Dependente de Ciclina/farmacologia , Purinas/farmacologia , Aminopiridinas/uso terapêutico , Animais , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Ciclina D1/metabolismo , Quinase 4 Dependente de Ciclina/metabolismo , Quinase 6 Dependente de Ciclina/metabolismo , Proteínas Inibidoras de Quinase Dependente de Ciclina/uso terapêutico , Fator de Transcrição E2F1/metabolismo , Feminino , Humanos , Masculino , Neoplasias Mamárias Experimentais , Camundongos , Camundongos Nus , Fosforilação/efeitos dos fármacos , Purinas/uso terapêutico , Proteína do Retinoblastoma/metabolismo , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/metabolismo , Neoplasias de Mama Triplo Negativas/patologia , Carga Tumoral/efeitos dos fármacos
4.
J Agric Food Chem ; 67(35): 9727-9737, 2019 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-31398034

RESUMO

The present study aimed to search for chicken abdominal fat deposition-related polymorphisms within RB1 and to provide functional evidence for significantly associated genetic variants. Association analyses showed that 11 single nucleotide polymorphisms (SNPs) in intron 17 of RB1, were significantly associated with both abdominal fat weight (P < 0.05) and abdominal fat percentage (P < 0.05). Functional analysis revealed that the A allele of g.32828A>G repressed the transcriptional efficiency of RB1 in vitro, through binding nuclear factor-kappa B (NF-KB) and SRY-related HMG box protein 2 (SOX2). Furthermore, RB1 mRNA expression levels in the abdominal fat tissue of individuals with the A/A genotype of g.32828A>G were lower than those of individuals with the G/G genotype. Collectively, we propose that the intronic SNP g.32828A>G of RB1 is an obesity-associated variant that directly affects binding with NF-KB and SOX2, leading to changes in RB1 expression which in turn may influence chicken abdominal fat deposition.


Assuntos
Adiposidade , Proteínas Aviárias/metabolismo , Galinhas/metabolismo , NF-kappa B/metabolismo , Proteína do Retinoblastoma/genética , Proteína do Retinoblastoma/metabolismo , Fatores de Transcrição SOX/metabolismo , Gordura Abdominal/metabolismo , Alelos , Animais , Proteínas Aviárias/genética , Sítios de Ligação , Galinhas/genética , Galinhas/crescimento & desenvolvimento , Íntrons , NF-kappa B/genética , Polimorfismo de Nucleotídeo Único , Ligação Proteica , Fatores de Transcrição SOX/genética
5.
Nucleic Acids Res ; 47(17): 9087-9103, 2019 09 26.
Artigo em Inglês | MEDLINE | ID: mdl-31400114

RESUMO

Most human cancers acquire mutations causing defects in the p53 signaling pathway. The tumor suppressor p53 becomes activated in response to genotoxic stress and is essential for arresting the cell cycle to facilitate DNA repair or to initiate apoptosis. p53-induced cell cycle-arrest is mediated by expression of the CDK inhibitor p21WAF1/Cip1, which prevents phosphorylation and inactivation of the pocket proteins RB, p130, and p107. In a hypophosphorylated state, pocket proteins bind to E2F factors forming RB-E2F and DREAM transcriptional repressor complexes. Here, we analyze the influence of RB and DREAM on p53-induced gene repression and cell-cycle arrest. We show that abrogation of DREAM function by knockout of the DREAM component LIN37 results in a reduced repression of cell-cycle genes. We identify the genes repressed by the p53-DREAM pathway and describe a set of genes that is downregulated by p53 independent of LIN37/DREAM. Most strikingly, p53-dependent repression of cell-cycle genes is completely abrogated in LIN37-/-;RB-/- cells leading to a loss of the G1/S checkpoint. Taken together, we show that DREAM and RB are key factors in the p53 signaling pathway to downregulate a large number of cell-cycle genes and to arrest the cell cycle at the G1/S transition.


Assuntos
Pontos de Checagem do Ciclo Celular/genética , Regulação da Expressão Gênica , Proteínas Interatuantes com Canais de Kv/metabolismo , Proteínas Repressoras/metabolismo , Proteína do Retinoblastoma/genética , Transativadores/fisiologia , Proteína Supressora de Tumor p53/metabolismo , Animais , Células Cultivadas , Proteína Substrato Associada a Crk/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Fatores de Transcrição E2F/genética , Fatores de Transcrição E2F/metabolismo , Fibroblastos/metabolismo , Genes cdc , Células HCT116 , Humanos , Proteínas Interatuantes com Canais de Kv/genética , Camundongos , Proteínas Repressoras/genética , Proteína do Retinoblastoma/metabolismo , Proteína p107 Retinoblastoma-Like/genética , Transativadores/genética , Transativadores/metabolismo , Proteína Supressora de Tumor p53/genética
6.
Nat Commun ; 10(1): 2939, 2019 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-31270324

RESUMO

E2F transcription factors are central regulators of cell division and cell fate decisions. E2F4 often represents the predominant E2F activity in cells. E2F4 is a transcriptional repressor implicated in cell cycle arrest and whose repressive activity depends on its interaction with members of the RB family. Here we show that E2F4 is important for the proliferation and the survival of mouse embryonic stem cells. In these cells, E2F4 acts in part as a transcriptional activator that promotes the expression of cell cycle genes. This role for E2F4 is independent of the RB family. Furthermore, E2F4 functionally interacts with chromatin regulators associated with gene activation and we observed decreased histone acetylation at the promoters of cell cycle genes and E2F targets upon loss of E2F4 in RB family-mutant cells. Taken together, our findings uncover a non-canonical role for E2F4 that provide insights into the biology of rapidly dividing cells.


Assuntos
Fator de Transcrição E2F4/metabolismo , Células-Tronco Embrionárias Murinas/metabolismo , Proteína do Retinoblastoma/metabolismo , Ativação Transcricional , Animais , Ciclo Celular , Divisão Celular , Fator de Transcrição E2F4/genética , Regulação da Expressão Gênica no Desenvolvimento , Camundongos , Células-Tronco Embrionárias Murinas/citologia , Família Multigênica , Proteína do Retinoblastoma/genética
7.
PLoS Biol ; 17(7): e3000367, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31323018

RESUMO

Human papillomaviruses (HPVs) are causative agents of various diseases associated with cellular hyperproliferation, including cervical cancer, one of the most prevalent tumors in women. E7 is one of the two HPV-encoded oncoproteins and directs recruitment and subsequent degradation of tumor-suppressive proteins such as retinoblastoma protein (pRb) via its LxCxE motif. E7 also triggers tumorigenesis in a pRb-independent pathway through its C-terminal domain, which has yet been largely undetermined, with a lack of structural information in a complex form with a host protein. Herein, we present the crystal structure of the E7 C-terminal domain of HPV18 belonging to the high-risk HPV genotypes bound to the catalytic domain of human nonreceptor-type protein tyrosine phosphatase 14 (PTPN14). They interact directly and potently with each other, with a dissociation constant of 18.2 nM. Ensuing structural analysis revealed the molecular basis of the PTPN14-binding specificity of E7 over other protein tyrosine phosphatases and also led to the identification of PTPN21 as a direct interacting partner of E7. Disruption of HPV18 E7 binding to PTPN14 by structure-based mutagenesis impaired E7's ability to promote keratinocyte proliferation and migration. Likewise, E7 binding-defective PTPN14 was resistant for degradation via proteasome, and it was much more effective than wild-type PTPN14 in attenuating the activity of downstream effectors of Hippo signaling and negatively regulating cell proliferation, migration, and invasion when examined in HPV18-positive HeLa cells. These results therefore demonstrated the significance and therapeutic potential of the intermolecular interaction between HPV E7 and host PTPN14 in HPV-mediated cell transformation and tumorigenesis.


Assuntos
Transformação Celular Neoplásica , Proteínas de Ligação a DNA/metabolismo , Proteínas Oncogênicas Virais/metabolismo , Proteínas Tirosina Fosfatases não Receptoras/metabolismo , Neoplasias do Colo do Útero/metabolismo , Sequência de Aminoácidos , Linhagem Celular , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Feminino , Células HEK293 , Células HeLa , Humanos , Modelos Moleculares , Proteínas Oncogênicas Virais/química , Proteínas Oncogênicas Virais/genética , Ligação Proteica , Domínios Proteicos , Proteínas Tirosina Fosfatases não Receptoras/química , Proteínas Tirosina Fosfatases não Receptoras/genética , Proteína do Retinoblastoma/química , Proteína do Retinoblastoma/genética , Proteína do Retinoblastoma/metabolismo , Homologia de Sequência de Aminoácidos , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/patologia
8.
Int J Radiat Oncol Biol Phys ; 105(3): 548-558, 2019 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-31271827

RESUMO

PURPOSE: Human papillomavirus negative (HPV-ve) head and neck squamous cell carcinoma (HNSCC) has a poor prognosis compared with HPV+ve HNSCCs. Expression of p16 in HPV+ve HNSCC is thought to mediate radiosensitivity via inhibition of cyclin-dependent kinase (CDK) 4/6. We used a clinically approved CDK4/CDK6 inhibitor, palbociclib, and assessed its effect on radiosensitivity in HNSCC. METHODS AND MATERIALS: The effect of palbociclib on radiosensitivity was determined in HPV-ve and HPV+ve HNSCC cell lines using colony survival assays, immunofluorescent staining of repair proteins, homologous recombination assays, cell cycle, and metaphase spread analyses. RESULTS: Only HPV-ve HNSCC cells were radiosensitized by palbociclib, which also occurred at hypoxic levels associated with radioresistance. Palbociclib led to decreased induction of BRCA1 and RAD51 after irradiation. Homologous recombination was diminished and repair of radiation-induced DNA damage was delayed in the presence of palbociclib, leading to increased chromosomal damage. Failure to repair radiation-induced damage led to cell death as a result of mitotic catastrophe. CONCLUSIONS: Here, we highlight a therapeutic strategy to improve the radiosensitivity of HPV-ve HNSCC, a patient group that has an unmet and urgent need for improved radiation therapy efficacy.


Assuntos
Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Neoplasias de Cabeça e Pescoço/radioterapia , Piperazinas/uso terapêutico , Inibidores de Proteínas Quinases/uso terapêutico , Piridinas/uso terapêutico , Tolerância a Radiação , Radiossensibilizantes/uso terapêutico , Carcinoma de Células Escamosas de Cabeça e Pescoço/radioterapia , Proteína BRCA1/metabolismo , Ciclo Celular , Morte Celular , Linhagem Celular Tumoral , Aberrações Cromossômicas/induzido quimicamente , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Reparo do DNA/efeitos dos fármacos , Proteínas de Ligação a DNA/metabolismo , Neoplasias de Cabeça e Pescoço/virologia , Recombinação Homóloga , Humanos , Proteínas de Neoplasias/metabolismo , Papillomaviridae , Fosforilação , Rad51 Recombinase/metabolismo , Proteína do Retinoblastoma/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço/virologia , Hipóxia Tumoral , Ensaio Tumoral de Célula-Tronco
9.
Mol Cell Biochem ; 460(1-2): 29-36, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31214845

RESUMO

Cardiovascular disease (CVD) is a major cause of global mortality. The proper functioning of the endothelial layer of arteries is crucial to cardiovascular health. Retinoblastoma protein (Rb), encoded by the Rb1 gene, has been shown to offer vasoprotective effects. Herein, we investigated endothelial Rb's effects on arterial function using an endothelial-specific conditional Rb1 knockout (Rb cKO) mouse model. We found that Rb deficiency reduced dihydrofolate reductase (DHFR) activity and downstream NO production in mouse aortic endothelial cells and blocked arterial vasodilation in an endothelial DHFR-dependent manner. Rb deficiency also increased phenylephrine-triggered arterial vasoconstriction, BP levels, and pathological aortic remodeling without significantly affecting prostanoid synthesis. Employing an angiotensin II (AngII)-stimulated apolipoprotein E knockout (apoE -/-) mice fed a standard, non-atherogenic diet, Rb deficiency increased aortic diameter, stimulated abdominal aortic aneurysm (AAA) development, and reduced survival. These pathological responses to Rb deficiency in AngII-stimulated apoE-/- mice were rescued by DHFR overexpression. Cumulatively, our findings reveal that endothelial Rb positively impacts arterial function by supporting vasoprotective endothelial DHFR/NO pathway activity, leading to reduced AAA development.


Assuntos
Aneurisma da Aorta Abdominal/patologia , Células Endoteliais/metabolismo , Óxido Nítrico/metabolismo , Proteína do Retinoblastoma/metabolismo , Transdução de Sinais , Tetra-Hidrofolato Desidrogenase/metabolismo , Animais , Aorta Torácica/metabolismo , Aorta Torácica/patologia , Aorta Torácica/fisiopatologia , Aneurisma da Aorta Abdominal/fisiopatologia , Artérias/metabolismo , Pressão Sanguínea , Regulação para Baixo , Células Endoteliais/patologia , Camundongos , Óxido Nítrico Sintase Tipo III/metabolismo , Prostaglandinas/metabolismo , Proteína do Retinoblastoma/deficiência , Remodelação Vascular , Vasoconstrição , Vasodilatação
10.
Nat Commun ; 10(1): 2860, 2019 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-31253784

RESUMO

Lacking targetable molecular drivers, triple-negative breast cancer (TNBC) is the most clinically challenging subtype of breast cancer. In this study, we reveal that Death Effector Domain-containing DNA-binding protein (DEDD), which is overexpressed in > 60% of TNBCs, drives a mitogen-independent G1/S cell cycle transition through cytoplasm localization. The gain of cytosolic DEDD enhances cyclin D1 expression by interacting with heat shock 71 kDa protein 8 (HSC70). Concurrently, DEDD interacts with Rb family proteins and promotes their proteasome-mediated degradation. DEDD overexpression renders TNBCs vulnerable to cell cycle inhibition. Patients with TNBC have been excluded from CDK 4/6 inhibitor clinical trials due to the perceived high frequency of Rb-loss in TNBCs. Interestingly, our study demonstrated that, irrespective of Rb status, TNBCs with DEDD overexpression exhibit a DEDD-dependent vulnerability to combinatorial treatment with CDK4/6 inhibitor and EGFR inhibitor in vitro and in vivo. Thus, our study provided a rationale for the clinical application of CDK4/6 inhibitor combinatorial regimens for patients with TNBC.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Proteínas Adaptadoras de Sinalização de Receptores de Domínio de Morte/metabolismo , Lapatinib/farmacologia , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Antineoplásicos/farmacologia , Ciclo Celular/efeitos dos fármacos , Proteínas de Ligação a DNA/genética , Proteínas Adaptadoras de Sinalização de Receptores de Domínio de Morte/genética , Receptores ErbB/antagonistas & inibidores , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Receptor ErbB-2/antagonistas & inibidores , Proteína do Retinoblastoma/genética , Proteína do Retinoblastoma/metabolismo , Neoplasias de Mama Triplo Negativas/metabolismo
11.
Cell Oncol (Dordr) ; 42(5): 705-715, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31250364

RESUMO

PURPOSE: The cyclin-dependent kinases (CDKs) CDK4 and CDK6 are important regulators of the cell cycle and represent promising targets in cancer treatment. We aimed to investigate the relevance of CDK4/6 in the development of hepatocellular carcinoma (HCC) and the potential of ribociclib, a novel orally available CDK4/6 inhibitor, as a treatment for HCC. METHODS: The effect of ribociclib was assessed in native and sorafenib-resistant HCC cell lines using viability assays, colony formation assays and FACS-based analyses. The expression of potential biomarkers of ribociclib response was assessed in cell lines and primary human hepatocytes using Western blotting. In addition, the prognostic relevance of the cyclin D-CDK4/6-retinoblastoma protein (Rb) pathway was assessed by analysing mRNA expression data from The Cancer Genome Atlas (TCGA). RESULTS: We found that ribociclib downregulated Rb and caused a profound loss of cell viability by inducing G1 cell cycle arrest in HCC cell lines exhibiting Rb-high/p16-low protein expression profiles, but not in Rb-low/p16-high cells, regardless their sensitivity to sorafenib. siRNA-based Rb silencing decreased cell proliferation, but did not diminish the sensitivity of HCC cells to ribociclib. Furthermore, we found that ribociclib synergized with sorafenib to cause cell death. mRNA analysis of primary human HCC specimens showed that CDK4 expression was correlated with patient survival and that the expression of Rb and the p16-encoding CDKN2A gene were inversely correlated. CONCLUSIONS: From our data we conclude that impairment of the cyclin D-CDK4/6-Rb pathway is a frequent feature of HCC and that it is associated with a unfavourable prognosis. We also found that ribociclib exhibits a preferential antineoplastic activity in Rb-high HCC cells. Our results warrant further investigation of Rb and p16 expression as markers of HCC sensitivity to ribociclib.


Assuntos
Aminopiridinas/farmacologia , Antineoplásicos/farmacologia , Carcinoma Hepatocelular/metabolismo , Quinase 4 Dependente de Ciclina/metabolismo , Quinase 6 Dependente de Ciclina/metabolismo , Neoplasias Hepáticas/metabolismo , Purinas/farmacologia , Proteína do Retinoblastoma/metabolismo , Apoptose/efeitos dos fármacos , Biomarcadores Tumorais , Carcinoma Hepatocelular/mortalidade , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Resistencia a Medicamentos Antineoplásicos , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Hepatócitos/efeitos dos fármacos , Humanos , Neoplasias Hepáticas/mortalidade , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Proteína do Retinoblastoma/antagonistas & inibidores , Proteína do Retinoblastoma/genética , Sorafenibe/farmacologia
12.
Cell Oncol (Dordr) ; 42(4): 477-490, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30949979

RESUMO

PURPOSE: Oral squamous cell carcinoma (OSCC) is a challenging disease to treat. Up to 50% of OSCC patients with advanced disease develop recurrences. Elucidation of key molecular mechanisms underlying OSCC development may provide opportunities to target specific genes and, thus, to improve patient survival. In this study, we examined the expression and functional role of interferon transmembrane protein 3 (IFITM3) in OSCC development. METHODS: The expression of IFITM3 in OSCC and normal oral mucosal tissues was assessed by qRT-PCR and immunohistochemistry. The role of IFITM3 in driving OSCC cell proliferation and survival was examined using siRNA-mediated gene knockdown, and the role of IFITM3 in driving cell cycle regulators was examined using Western blotting. RESULTS: We found that IFITM3 is overexpressed in more than 79% of primary OSCCs. We also found that IFITM3 knockdown led to impaired OSCC cell growth through inhibition of cell proliferation, induction of cell cycle arrest, senescence and apoptosis. In addition, we found that IFITM3 knockdown led to reduced expressions of CCND1 and CDK4 and reduced RB phosphorylation, leading to inhibition of OSCC cell growth. This information may be instrumental for the design of novel targeted therapeutic strategies. CONCLUSIONS: From our data we conclude that IFITM3 is overexpressed in OSCC and may regulate the CCND1-CDK4/6-pRB axis to mediate OSCC cell growth.


Assuntos
Carcinoma de Células Escamosas/metabolismo , Ciclina D1/metabolismo , Quinase 4 Dependente de Ciclina/metabolismo , Técnicas de Silenciamento de Genes , Proteínas de Membrana/metabolismo , Neoplasias Bucais/metabolismo , Proteínas de Ligação a RNA/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Apoptose , Carcinoma de Células Escamosas/patologia , Pontos de Checagem do Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Sobrevivência Celular , Senescência Celular , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Mucosa Bucal/patologia , Neoplasias Bucais/patologia , Fosforilação , Proteína do Retinoblastoma/metabolismo , Transdução de Sinais
13.
Nature ; 568(7753): 557-560, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30971822

RESUMO

The cell cycle is a tightly regulated process that is controlled by the conserved cyclin-dependent kinase (CDK)-cyclin protein complex1. However, control of the G0-to-G1 transition is not completely understood. Here we demonstrate that p38 MAPK gamma (p38γ) acts as a CDK-like kinase and thus cooperates with CDKs, regulating entry into the cell cycle. p38γ shares high sequence homology, inhibition sensitivity and substrate specificity with CDK family members. In mouse hepatocytes, p38γ induces proliferation after partial hepatectomy by promoting the phosphorylation of retinoblastoma tumour suppressor protein at known CDK target residues. Lack of p38γ or treatment with the p38γ inhibitor pirfenidone protects against the chemically induced formation of liver tumours. Furthermore, biopsies of human hepatocellular carcinoma show high expression of p38γ, suggesting that p38γ could be a therapeutic target in the treatment of this disease.


Assuntos
Carcinogênese/patologia , Ciclo Celular , Neoplasias Hepáticas/enzimologia , Neoplasias Hepáticas/patologia , Fígado/enzimologia , Fígado/patologia , Proteína Quinase 12 Ativada por Mitógeno/metabolismo , Idoso , Animais , Carcinogênese/efeitos dos fármacos , Carcinoma Hepatocelular/induzido quimicamente , Carcinoma Hepatocelular/patologia , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Quinases Ciclina-Dependentes/antagonistas & inibidores , Quinases Ciclina-Dependentes/metabolismo , Feminino , Hepatócitos/citologia , Hepatócitos/patologia , Humanos , Fígado/cirurgia , Neoplasias Hepáticas/induzido quimicamente , Masculino , Camundongos , Pessoa de Meia-Idade , Proteína Quinase 12 Ativada por Mitógeno/antagonistas & inibidores , Fosforilação , Piridonas/farmacologia , Proteína do Retinoblastoma/química , Proteína do Retinoblastoma/metabolismo , Homologia de Sequência , Especificidade por Substrato
14.
Asia Pac J Clin Oncol ; 15(5): e162-e166, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30990963

RESUMO

AIM: This study aimed to investigate the correlation between the expression of A-kinase anchor protein95 (AKAP95), p-retinoblastoma (phosphorylated Rb, p-Rb), cyclin D2, cyclin D3 and cyclin E2 in esophageal cancer tissues and clinicopathological indexes. METHOD: The protein expression levels of AKAP95, p-Rb, cyclin D2/3 and cyclin E2 in 40 esophageal cancer tissues were detected using immunohistochemistry, and the correlation between them was analyzed. RESULT: The percentage of p-Rb (Ser780)-, cyclin D2-, cyclin D3- and cyclin E2-positive samples was 62.50%, 70.00%, 67.50% and 60.00%, respectively. Also, the positive expression did not correlate with the histological type, histological differentiation or lymph node metastasis. The expression of AKAP95 and p-Rb (Ser780), p-Rb (Ser780) and cyclin D2 and p-Rb (Ser780) and cyclin D3 in esophageal cancer tissues was found to be correlated (P < 0.05). CONCLUSIONS: The expression of AKAP95 and p-Rb (Ser780), p-Rb(Ser780) and cyclin D2, and p-Rb (Ser780) and cyclin D3 in esophageal cancer tissue was correlated, suggesting that these proteins might play a synergistic role in cell-cycle progression. Cyclin D2/D3 and p-Rb (Ser780) were correlated whereas cyclin E2 and p-Rb (Ser780) were not, suggesting that p-Rb (Ser780) might be highly expressed and the Ser780 site of Rb protein might be phosphorylated in the early stage of the G1 phase. Ser780 was the site in the primary phosphorylation stage of several phosphorylation sites during stepwise phosphorylation (from primary to high phosphorylation).


Assuntos
Adenocarcinoma/metabolismo , Biomarcadores Tumorais/metabolismo , Carcinoma de Células Escamosas/metabolismo , Neoplasias Esofágicas/metabolismo , Proteínas de Ancoragem à Quinase A/metabolismo , Adenocarcinoma/secundário , Adenocarcinoma/cirurgia , Idoso , Carcinoma de Células Escamosas/secundário , Carcinoma de Células Escamosas/cirurgia , Ciclina D2/metabolismo , Ciclina D3/metabolismo , Ciclinas/metabolismo , Neoplasias Esofágicas/patologia , Neoplasias Esofágicas/cirurgia , Humanos , Metástase Linfática , Pessoa de Meia-Idade , Fosforilação , Prognóstico , Proteômica , Proteína do Retinoblastoma/metabolismo
15.
Nat Commun ; 10(1): 1296, 2019 03 21.
Artigo em Inglês | MEDLINE | ID: mdl-30899002

RESUMO

The dysregulation of Fbxo4-cyclin D1 axis occurs at high frequency in esophageal squamous cell carcinoma (ESCC), where it promotes ESCC development and progression. However, defining a therapeutic vulnerability that results from this dysregulation has remained elusive. Here we demonstrate that Rb and mTORC1 contribute to Gln-addiction upon the dysregulation of the Fbxo4-cyclin D1 axis, which leads to the reprogramming of cellular metabolism. This reprogramming is characterized by reduced energy production and increased sensitivity of ESCC cells to combined treatment with CB-839 (glutaminase 1 inhibitor) plus metformin/phenformin. Of additional importance, this combined treatment has potent efficacy in ESCC cells with acquired resistance to CDK4/6 inhibitors in vitro and in xenograft tumors. Our findings reveal a molecular basis for cancer therapy through targeting glutaminolysis and mitochondrial respiration in ESCC with dysregulated Fbxo4-cyclin D1 axis as well as cancers resistant to CDK4/6 inhibitors.


Assuntos
Antineoplásicos/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Neoplasias Esofágicas/tratamento farmacológico , Carcinoma de Células Escamosas do Esôfago/tratamento farmacológico , Regulação Neoplásica da Expressão Gênica , Glutamina/metabolismo , Hipoglicemiantes/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Animais , Benzenoacetamidas/farmacologia , Linhagem Celular Tumoral , Ciclina D1/genética , Ciclina D1/metabolismo , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 4 Dependente de Ciclina/genética , Quinase 4 Dependente de Ciclina/metabolismo , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/genética , Quinase 6 Dependente de Ciclina/metabolismo , Resistencia a Medicamentos Antineoplásicos/genética , Sinergismo Farmacológico , Metabolismo Energético/efeitos dos fármacos , Metabolismo Energético/genética , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/metabolismo , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago/genética , Carcinoma de Células Escamosas do Esôfago/metabolismo , Carcinoma de Células Escamosas do Esôfago/patologia , Proteínas F-Box/genética , Proteínas F-Box/metabolismo , Glutaminase/antagonistas & inibidores , Glutaminase/genética , Glutaminase/metabolismo , Glutamina/antagonistas & inibidores , Humanos , Masculino , Alvo Mecanístico do Complexo 1 de Rapamicina/genética , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Metformina/farmacologia , Camundongos , Terapia de Alvo Molecular , Fenformin/farmacologia , Proteína do Retinoblastoma/genética , Proteína do Retinoblastoma/metabolismo , Transdução de Sinais , Tiadiazóis/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
16.
Dis Model Mech ; 12(4)2019 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-30910991

RESUMO

Pancreatic cancer is one of the most lethal common cancers. The cell of origin of pancreatic ductal adenocarcinoma (PDAC) has been controversial, and recent evidence suggested acinar cells as the most probable candidate. However, the genetic alterations driving the transformation of pancreatic acinar cells in fully mature animals remain to be deciphered. In this study, lentivirus was used as a tool to introduce genetic engineering in tree shrew pancreatic acinar cells to explore the driver mutation essential for malignant transformation, establishing a novel tree shrew PDAC model, because we found that lentivirus could selectively infect acinar cells in tree shrew pancreas. Combination of oncogenic KRASG12D expression and inactivation of tumor suppressor genes Tp53, Cdkn2a and Cdkn2b could induce pancreatic cancer with full penetrance. Silencing of Cdkn2b is indispensable for Rb1 phosphorylation and tumor induction. Tree shrew PDAC possesses the main histological and molecular features of human PDAC. The gene expression profile of tree shrew PDAC was more similar to human disease than a mouse model. In conclusion, we established a novel pancreatic cancer model in tree shrew and identified driver mutations indispensable for PDAC induction from acinar cells in mature adults, demonstrating the essential roles of Cdkn2b in the induction of PDAC originating from adult acinar cells. Tree shrew could thus provide a better choice than mouse for a PDAC model derived from acinar cells in fully mature animals.


Assuntos
Células Acinares/patologia , Transformação Celular Neoplásica/patologia , Neoplasias Pancreáticas/patologia , Tupaia/fisiologia , Células Acinares/virologia , Sequência de Aminoácidos , Animais , Carcinogênese/patologia , Carcinoma Ductal Pancreático/patologia , Inibidor p16 de Quinase Dependente de Ciclina/química , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Modelos Animais de Doenças , Lentivirus/metabolismo , Masculino , Metaplasia , Camundongos , Primatas , Proteína do Retinoblastoma/metabolismo , Transdução de Sinais
17.
Biochem Biophys Res Commun ; 512(2): 331-337, 2019 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-30885433

RESUMO

Deregulated expression of microRNAs plays oncogenic or anti-oncogenic roles in various cancers. However, expression of miR-107 was not consistent among several types of cancer, and the effect of miR-107 in ovarian cancer remains unclear. In this study, we found that expression miR-107 was significantly decreased in ovarian cancer patients and in cell lines. Ectopic expression of miR-107 suppressed cell proliferation and G1 phase to S transition of cell cycle, and was associated with downregulation of cyclin E1 (CCNE1) expression. Mechanistically, CCNE1 was confirmed to be a direct target of miR-107 through the dual-luciferase reporter assay. Knockdown of CCNE1 dramatically impeded cell cycle in G1/S phase transition similarly as miR-107 overexpression did. In addition, overexpression of CCNE1 reversed the inhibition of cell proliferation induced by miR-107 overexpression. Finally, miR-107 had anti-cancer potential by suppressing tumor initiation and progression in vivo. Our finding indicates that miR-107 serves as a tumor suppressor by decreasing CCNE1 expression levels, which may provide potential therapeutic strategies in ovarian cancer treatment.


Assuntos
Ciclina E/genética , Pontos de Checagem da Fase G1 do Ciclo Celular/genética , MicroRNAs/genética , Proteínas Oncogênicas/genética , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/patologia , Animais , Linhagem Celular Tumoral , Proliferação de Células/genética , Ciclina E/antagonistas & inibidores , Ciclina E/metabolismo , Regulação para Baixo , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Genes Supressores de Tumor , Xenoenxertos , Humanos , Camundongos , Camundongos Nus , MicroRNAs/metabolismo , Proteínas Oncogênicas/antagonistas & inibidores , Proteínas Oncogênicas/metabolismo , Neoplasias Ovarianas/metabolismo , Fosforilação , Proteína do Retinoblastoma/metabolismo
18.
Cancer Sci ; 110(4): 1420-1430, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30724426

RESUMO

Inhibition of the cyclin-dependent kinase (CDK) 4/6-retinoblastoma (RB) pathway is an effective therapeutic strategy against cancer. Here, we performed a preclinical investigation of the antitumor activity of SHR6390, a novel CDK4/6 inhibitor. SHR6390 exhibited potent antiproliferative activity against a wide range of human RB-positive tumor cells in vitro, and exclusively induced G1 arrest as well as cellular senescence, with a concomitant reduction in the levels of Ser780-phosphorylated RB protein. Compared with the well-known CDK4/6 inhibitor palbociclib, orally administered SHR6390 led to equivalent or improved tumor efficacy against a panel of carcinoma xenografts, and produced marked tumor regression in some models, in association with sustained target inhibition in tumor tissues. Furthermore, SHR6390 overcame resistance to endocrine therapy and HER2-targeting antibody in ER-positive and HER2-positive breast cancer, respectively. Moreover, SHR6390 combined with endocrine therapy exerted remarkable synergistic antitumor activity in ER-positive breast cancer. Taken together, our findings indicate that SHR6390 is a novel CDK4/6 inhibitor with favorable pharmaceutical properties for use as an anticancer agent.


Assuntos
Antineoplásicos/farmacologia , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Neoplasias/metabolismo , Neoplasias/patologia , Inibidores de Proteínas Quinases/farmacologia , Animais , Apoptose/efeitos dos fármacos , Biomarcadores Tumorais , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Quinase 4 Dependente de Ciclina/metabolismo , Quinase 6 Dependente de Ciclina/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Humanos , Camundongos , Neoplasias/tratamento farmacológico , Proteína do Retinoblastoma/metabolismo , Transdução de Sinais/efeitos dos fármacos , Resultado do Tratamento , Ensaios Antitumorais Modelo de Xenoenxerto
19.
PLoS Negl Trop Dis ; 13(2): e0007163, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30789912

RESUMO

BACKGROUND: Trypanosoma cruzi is the obligate intracellular parasite that causes Chagas disease. The pathogenesis of this disease is a multifactorial complex process that involves a large number of molecules and particles, including the extracellular vesicles. The presence of EVs of T. cruzi was first described in 1979 and, since then, research regarding these particles has been increasing. Some of the functions described for these EVs include the increase in heart parasitism and the immunomodulation and evasion of the host immune response. Also, EVs may be involved in parasite adhesion to host cells and host cell invasion. METHODOLOGY/PRINCIPAL FINDINGS: EVs (exosomes) of the Pan4 strain of T. cruzi were isolated by differential centrifugation, and measured and quantified by TEM, NTA and DLS. The effect of EVs in increasing the parasitization of Vero cells was evaluated and the ED50 was calculated. Changes in cell permeability induced by EVs were evaluated in Vero and HL-1 cardiomyocyte cells using cell viability techniques such as trypan blue and MTT assays, and by confocal microscopy. The intracellular mobilization of Ca2+ and the disruption of the actin cytoskeleton induced by EVs over Vero cells were followed-up in time using confocal microscopy. To evaluate the effect of EVs over the cell cycle, cell cycle analyses using flow cytometry and Western blotting of the phosphorylated and non-phosphorylated protein of Retinoblastoma were performed. CONCLUSION/SIGNIFICANCE: The incubation of cells with EVs of trypomastigotes of the Pan4 strain of T. cruzi induce a number of changes in the host cells that include a change in cell permeability and higher intracellular levels of Ca2+ that can alter the dynamics of the actin cytoskeleton and arrest the cell cycle at G0/G1 prior to the DNA synthesis necessary to complete mitosis. These changes aid the invasion of host cells and augment the percentage of cell parasitization.


Assuntos
Vesículas Extracelulares/fisiologia , Trypanosoma cruzi/citologia , Animais , Cálcio , Linhagem Celular , Regulação da Expressão Gênica , Interações Hospedeiro-Parasita , Humanos , Microscopia Eletrônica de Transmissão , Proteína do Retinoblastoma/genética , Proteína do Retinoblastoma/metabolismo , Trypanosoma cruzi/fisiologia
20.
Oncogene ; 38(21): 4125-4141, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30700828

RESUMO

The CDK4/6 inhibitor palbociclib reduces tumor growth by decreasing retinoblastoma (RB) protein phosphorylation and inducing cell cycle arrest at the G1/S phase transition. Palbociclib in combination with anti-hormonal therapy brings significant benefit to breast cancer patients. In this study, novel combination approaches and underlying molecular/cellular mechanisms for palbociclib were explored in squamous cell lung cancer (SqCLC), the second most common subtype of non-small cell lung cancer. While approximate 20% lung patients benefit from immunotherapy, most SqCLC patients who receive platinum-doublet chemotherapy as first-line treatment, which often includes a taxane, are still in need of more effective combination therapies. Our results demonstrated enhanced cytotoxicity and anti-tumor effect with palbociclib plus taxanes at clinically achievable doses in multiple SqCLC models with diverse cancer genetic backgrounds. Comprehensive gene expression analysis revealed a sustained disruption of pRB-E2F signaling by combination that was accompanied with enhanced regulation of pleiotropic biological effects. These included several novel mechanisms such as abrogation of G2/M and mitotic spindle assembly checkpoints, as well as impaired induction of hypoxia-inducible factor 1 alpha (HIF-1α). The decrease in HIF-1α modulated a couple key angiogenic and anti-angiogenic factors, resulting in an enhanced anti-angiogenic effect. This preclinical work suggests a new therapeutic opportunity for palbociclib in lung and other cancers currently treated with taxane based chemotherapy as standard of care.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Hidrocarbonetos Aromáticos com Pontes/farmacologia , Carcinoma de Células Escamosas/tratamento farmacológico , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Fatores de Transcrição E2F/metabolismo , Neoplasias Pulmonares/tratamento farmacológico , Proteína do Retinoblastoma/metabolismo , Taxoides/farmacologia , Inibidores da Angiogênese/farmacologia , Animais , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma de Células Escamosas/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Expressão Gênica/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana , Humanos , Neoplasias Pulmonares/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/metabolismo , Piperazinas/farmacologia , Piridinas/farmacologia , Transdução de Sinais/efeitos dos fármacos
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