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1.
Bioorg Med Chem Lett ; 108: 129797, 2024 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-38759932

RESUMO

TGF-ß is an immunosuppressive cytokine and plays a key role in progression of cancer by inducing immunosuppression in tumor microenvironment. Therefore, inhibition of TGF-ß signaling pathway may provide a potential therapeutic intervention in treating cancers. Herein, we report the discovery of a series of novel thiazole derivatives as potent inhibitors of ALK5, a serine-threonine kinase which is responsible for TGF-ß signal transduction. Compound 29b was identified as a potent inhibitor of ALK5 with an IC50 value of 3.7 nM with an excellent kinase selectivity.


Assuntos
Desenho de Fármacos , Inibidores de Proteínas Quinases , Proteínas Serina-Treonina Quinases , Receptor do Fator de Crescimento Transformador beta Tipo I , Receptores de Fatores de Crescimento Transformadores beta , Tiazóis , Tiazóis/química , Tiazóis/farmacologia , Tiazóis/síntese química , Receptor do Fator de Crescimento Transformador beta Tipo I/antagonistas & inibidores , Receptor do Fator de Crescimento Transformador beta Tipo I/metabolismo , Humanos , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Relação Estrutura-Atividade , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/antagonistas & inibidores , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Estrutura Molecular , Relação Dose-Resposta a Droga
2.
Invest New Drugs ; 41(5): 751-760, 2023 10.
Artigo em Inglês | MEDLINE | ID: mdl-37702844

RESUMO

Approximately 60%-80% of patients who achieve complete remission eventually relapse after conventional chemotherapy and have poor prognoses despite the recent advances of novel anticancer agents. Continuing development of more effective novel treatments for acute myeloid leukemia (AML) is necessary. We developed (R)-WAC-224 (R-WAC), which is an anticancer quinolone, targeting topoisomerase II. This study evaluated the anti-leukemia potential of R-WAC or racemic WAC-224 (WAC) in vitro and in vivo. R-WAC significantly inhibited the human AML cell line proliferation (MV4-11, HL60, and KG1a), which was comparable to daunorubicin and cytarabine, not affected by P-glycoprotein overexpression. WAC did neither increase serum troponin-T nor decrease the crypt numbers in the small intestine, indicating WAC was less toxic than doxorubicin. R-WAC monotherapy demonstrated prolonged survival in the AML mice model and inhibited tumor growth in the MV4-11 xenograft mice model. Moreover, the combination of R-WAC and cytarabine demonstrated more active anti-leukemia effects than daunorubicin and cytarabine. Finally, R-WAC inhibited the colony-forming abilities using primary AML cells. These results indicate that R-WAC is a promising therapeutic agent for AML.


Assuntos
Leucemia Mieloide Aguda , Quinolonas , Humanos , Animais , Camundongos , Quinolonas/uso terapêutico , Sinergismo Farmacológico , Leucemia Mieloide Aguda/metabolismo , Daunorrubicina/farmacologia , Daunorrubicina/uso terapêutico , Citarabina/farmacologia , Citarabina/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Proteínas Adaptadoras de Transdução de Sinal/metabolismo
3.
Cancer Sci ; 113(2): 529-539, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-34902205

RESUMO

The emergence of tyrosine kinase inhibitors as part of a front-line treatment has greatly improved the clinical outcome of the patients with Ph+ acute lymphoblastic leukemia (ALL). However, a portion of them still become refractory to the therapy mainly through acquiring mutations in the BCR-ABL1 gene, necessitating a novel strategy to treat tyrosine kinase inhibitor (TKI)-resistant Ph+ ALL cases. In this report, we show evidence that RUNX1 transcription factor stringently controls the expression of BCR-ABL1, which can strategically be targeted by our novel RUNX inhibitor, Chb-M'. Through a series of in vitro experiments, we identified that RUNX1 binds to the promoter of BCR and directly transactivates BCR-ABL1 expression in Ph+ ALL cell lines. These cells showed significantly reduced expression of BCR-ABL1 with suppressed proliferation upon RUNX1 knockdown. Moreover, treatment with Chb-M' consistently downregulated the expression of BCR-ABL1 in these cells and this drug was highly effective even in an imatinib-resistant Ph+ ALL cell line. In good agreement with these findings, forced expression of BCR-ABL1 in these cells conferred relative resistance to Chb-M'. In addition, in vivo experiments with the Ph+ ALL patient-derived xenograft cells showed similar results. In summary, targeting RUNX1 therapeutically in Ph+ ALL cells may lead to overcoming TKI resistance through the transcriptional regulation of BCR-ABL1. Chb-M' could be a novel drug for patients with TKI-resistant refractory Ph+ ALL.


Assuntos
Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Proteínas de Fusão bcr-abl/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Animais , Antineoplásicos/farmacologia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/genética , Subunidade alfa 2 de Fator de Ligação ao Core/antagonistas & inibidores , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Mesilato de Imatinib/farmacologia , Camundongos , Mutação , Cromossomo Filadélfia , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Inibidores de Proteínas Quinases/farmacologia
4.
Cancer Sci ; 112(3): 1196-1208, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33423358

RESUMO

5-Fluorouracil (5-FU) is one of the most frequently used pharmacological agents in the treatment of colorectal cancer (CRC). Resistance to chemotherapy is a major cause of treatment failure of CRC, and it is a well known fact that cancer stem cells play a significant role in the acquisition of drug resistance. In this study, we focused on the KHDRBS3 gene that encodes KH RNA Binding Domain Containing, Signal Transduction Associated 3. We first clarified the relationship between KHDRBS3 and 5-FU resistance. We then observed higher expression levels of KHDRBS3 in KRAS-mutant organoids and cell lines in comparison with KRAS wild-type organoids and cell lines. Immunohistochemical analysis using CRC cases revealed that the prognosis of KHDRBS3-positive patients was significantly worse compared with that of KHDRBS3-negative patients. Univariate and multivariate Cox proportional hazards analyses showed that KHDRBS3 was an independent prognostic factor in patients with CRC. We determined that KHDRBS3 might play a crucial role in the acquisition of stem cell properties, such as drug resistance and spheroid/organoid formation, by regulating CD44 variant expression and the Wnt signaling pathway. In an immunodeficient mouse model, KHDRBS3-positive cells showed efficient tumor formation and formed metastatic lesions in the lungs. These results indicated that KHDRBS3 plays a crucial role in drug resistance and anchorage-independent growth by maintaining stem cell-like features in CRC cells. KHDRBS3 could be a promising candidate marker for predicting chemotherapeutic effect and prognosis in CRC patients.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Neoplasias Colorretais/terapia , Resistência a Múltiplos Medicamentos/genética , Resistencia a Medicamentos Antineoplásicos/genética , Proteínas de Ligação a RNA/metabolismo , Idoso , Idoso de 80 Anos ou mais , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Linhagem Celular Tumoral , Colectomia , Colo/patologia , Colo/cirurgia , Neoplasias Colorretais/genética , Neoplasias Colorretais/mortalidade , Neoplasias Colorretais/patologia , Feminino , Fluoruracila/farmacologia , Fluoruracila/uso terapêutico , Regulação Neoplásica da Expressão Gênica , Técnicas de Inativação de Genes , Humanos , Receptores de Hialuronatos/genética , Masculino , Camundongos , Pessoa de Meia-Idade , Mutação , Células-Tronco Neoplásicas/patologia , Organoides/efeitos dos fármacos , Organoides/metabolismo , Prognóstico , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas de Ligação a RNA/genética , Análise de Sobrevida , Via de Sinalização Wnt/genética , Ensaios Antitumorais Modelo de Xenoenxerto
5.
Haematologica ; 106(2): 483-494, 2021 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-32001531

RESUMO

Therapeutic strategies that target leukemic stem cells (LSCs) provide potential advantages in the treatment of chronic myeloid leukemia (CML). Here, we show that selective blockade of plasminogen activator inhibitor-1 (PAI-1) enhances the susceptibility of CML-LSCs to tyrosine kinase inhibitor (TKI), which facilitates the eradication of CML-LSCs and leads to sustained remission of the disease. We demonstrated for the first time that TGF-ß-PAI-1 axis was selectively augmented in CML-LSCs in the bone marrow (BM), whereby protecting CML-LSCs from TKI treatment. Furthermore, the combined administration of TKI plus a PAI-1 inhibitor, in a mouse model of CML, significantly enhanced the eradication of CML cells in the BM and prolonged the survival of CML mice. The combined therapy of imatinib and a PAI-1 inhibitor prevented the recurrence of CML-like disease in serially transplanted recipients, indicating the elimination of CML-LSCs. Interestingly, PAI-1 inhibitor treatment augmented membrane-type matrix metalloprotease-1 (MT1-MMP)-dependent motility of CML-LSCs, and the anti-CML effect of PAI-1 inhibitor was extinguished by the neutralizing antibody for MT1-MMP, underlining the mechanistic importance of MT1-MMP. Our findings provide evidence of, and a rationale for, a novel therapeutic tactic, based on the blockade of PAI-1 activity, for CML patients.


Assuntos
Proteínas de Fusão bcr-abl , Leucemia Mielogênica Crônica BCR-ABL Positiva , Animais , Humanos , Mesilato de Imatinib , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Camundongos , Células-Tronco Neoplásicas , Inibidor 1 de Ativador de Plasminogênio , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Serpina E2
6.
Gastric Cancer ; 23(5): 863-873, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32323025

RESUMO

BACKGROUND: The transcribed ultraconserved regions (T-UCRs) are a novel class of long non-coding RNAs and are involved in the development of several types of cancer. Although several different papers have described the oncogenic role of Uc.63+, there are no reports mentioning its importance in gastric cancer (GC) biology. METHODS: In this study, we evaluated Uc.63+ expression using clinical samples of GC by qRT-PCR, and also assessed the correlation between Uc.63+ expression and clinico-pathological factors. RESULTS: The upregulation of Uc.63+ was significantly correlated with advanced clinico-pathological features. Knockdown of Uc.63+ significantly repressed GC cell growth and migration, whereas overexpression of Uc.63+ conversely promoted those of GC cells. In situ hybridization of Uc.63+ revealed its preferential expression in poorly differentiated adenocarcinoma. We further conducted a microarray analysis using MKN-1 cells overexpressing Uc.63- and found that NF-κB signaling was significantly upregulated in accordance with Uc.63+ expression. CONCLUSION: Our results suggest that Uc.63+ could be involved in GC progression by regulating GC cell growth and migration via NF-κB signaling.


Assuntos
Adenocarcinoma/patologia , Biomarcadores Tumorais/metabolismo , Regulação Neoplásica da Expressão Gênica , NF-kappa B/metabolismo , RNA Longo não Codificante/genética , Neoplasias Gástricas/patologia , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Idoso , Apoptose , Biomarcadores Tumorais/genética , Estudos de Casos e Controles , Proliferação de Células , Progressão da Doença , Feminino , Humanos , Masculino , NF-kappa B/genética , Prognóstico , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Taxa de Sobrevida , Células Tumorais Cultivadas
7.
Proc Natl Acad Sci U S A ; 111(10): 3805-10, 2014 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-24567410

RESUMO

mTOR is an evolutionarily conserved kinase that plays a critical role in sensing and responding to environmental determinants. Recent studies have shown that fine-tuning of the activity of mTOR complexes contributes to organogenesis and tumorigenesis. Although rapamycin, an allosteric mTOR inhibitor, is an effective immunosuppressant, the precise roles of mTOR complexes in early T-cell development remain unclear. Here we show that mTORC1 plays a critical role in the development of both early T-cell progenitors and leukemia. Deletion of Raptor, an essential component of mTORC1, produced defects in the earliest development of T-cell progenitors in vivo and in vitro. Deficiency of Raptor resulted in cell cycle abnormalities in early T-cell progenitors that were associated with instability of the Cyclin D2/D3-CDK6 complexes; deficiency of Rictor, an mTORC2 component, did not have the same effect, indicating that mTORC1 and -2 control T-cell development in different ways. In a model of myeloproliferative neoplasm and T-cell acute lymphoblastic leukemia (T-ALL) evoked by Kras activation, Raptor deficiency dramatically inhibited the cell cycle in oncogenic Kras-expressing T-cell progenitors, but not myeloid progenitors, and specifically prevented the development of T-ALL. Although rapamycin treatment significantly prolonged the survival of recipient mice bearing T-ALL cells, rapamycin-insensitive leukemia cells continued to propagate in vivo. In contrast, Raptor deficiency in the T-ALL model resulted in cell cycle arrest and efficient eradication of leukemia. Thus, understanding the cell-context-dependent role of mTORC1 illustrates the potential importance of mTOR signals as therapeutic targets.


Assuntos
Linfopoese/fisiologia , Modelos Imunológicos , Complexos Multiproteicos/fisiologia , Células Precursoras de Linfócitos T/fisiologia , Leucemia-Linfoma Linfoblástico de Células T Precursoras/imunologia , Serina-Treonina Quinases TOR/metabolismo , Serina-Treonina Quinases TOR/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/deficiência , Animais , Proteínas de Transporte/metabolismo , Ciclo Celular/imunologia , Ciclo Celular/fisiologia , Primers do DNA , Citometria de Fluxo , Perfilação da Expressão Gênica , Immunoblotting , Imuno-Histoquímica , Alvo Mecanístico do Complexo 1 de Rapamicina , Alvo Mecanístico do Complexo 2 de Rapamicina , Camundongos , Complexos Multiproteicos/deficiência , Proteína Companheira de mTOR Insensível à Rapamicina , Proteína Regulatória Associada a mTOR , Transdução de Sinais/fisiologia , Sirolimo/farmacologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Serina-Treonina Quinases TOR/deficiência
8.
J Allergy Clin Immunol ; 138(4): 1170-1182.e9, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-26948079

RESUMO

BACKGROUND: IL-10-producing regulatory B (B10) cells potently suppress allergic diseases, such as contact hypersensitivity (CHS). Splenic B10 cells share overlapping phenotypic markers with CD5+ B1 B cells, CD1dhiCD21+CD23- marginal zone (MZ) B cells, and CD1dhiCD21+CD23+ T2-MZ precursor B cells but do not exclusively belong to either subset. OBJECTIVE: In this study we investigated the signaling mechanisms and a novel phenotypic parameter of B10 cells. METHOD: We performed microarray analysis comparing IL-10+ and IL-10- B cells. B cell-specific phosphatase and tensin homolog (PTEN)-deficient mice, which exhibit aberrant activation of the phosphatidylinositol 3-kinase (PI3K)-Akt pathway in B cells, were examined. RESULTS: Microarray analysis revealed that the PI3K-Akt pathway is important for IL-10 production in B cells. PI3K-Akt pathway inhibitors reduced B10 cell numbers in vitro. B10 cell numbers were significantly increased in B cell-specific PTEN-deficient mice. The CHS response was significantly diminished in PTEN-deficient mice. Unexpectedly, splenic B10 cells in these mice were found within the B1 B-cell subset but not within the MZ B-cell subset. In wild-type mice not only MZ B10 cells but also B1-B10 cells were identified in the spleen. In addition, these 2 B10 cell subsets were predominantly found within the CD9+CD80+ B-cell fraction. CONCLUSION: A novel splenic B1 regulatory cell subset (B1-B10 cells) was identified. Our findings show that the PI3K-Akt pathway in B cells is critical for B10 cell development and CHS response and that CD9/CD80 coexpression is a novel phenotypic parameter for both MZ-B10 and B1-B10 cells.


Assuntos
Linfócitos B/imunologia , Hipersensibilidade/enzimologia , Fosfatidilinositol 3-Quinase/imunologia , Proteínas Proto-Oncogênicas c-akt/imunologia , Animais , Linfócitos B/classificação , Linfócitos B/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Humanos , Hipersensibilidade/imunologia , Interleucina-10/metabolismo , Subpopulações de Linfócitos/efeitos dos fármacos , Subpopulações de Linfócitos/imunologia , Camundongos , Análise em Microsséries , Transdução de Sinais/efeitos dos fármacos , Baço/citologia , Baço/imunologia
9.
Rinsho Ketsueki ; 58(10): 1818-1827, 2017.
Artigo em Japonês | MEDLINE | ID: mdl-28978820

RESUMO

Leukemia stem cells (LSCs) are responsible for relapse of leukemia. LSCs maintain their self-renewal capacity, stemness properties, and therapeutic resistance in a manner dependent on their cell of origin and genetic alterations acquired during subsequent clonal evolution. Specific mechanisms of metabolic control and nutrient acquisition are implicated in the regulation of LSC survival. Recent advances in gene modification strategies in mice and in sophisticated metabolomics technologies are producing novel inquiries in LSC research performed in vivo. In this review, I examined our current knowledge on the roles of various metabolic pathways, including glucose metabolism, lipid oxidation, and an alternative route of amino acid and peptide supply, that support the maintenance of self-renewal capacity and therapeutic resistance in LSCs in vivo.


Assuntos
Leucemia/metabolismo , Células-Tronco Neoplásicas/metabolismo , Animais , Humanos , Leucemia/tratamento farmacológico , Terapia de Alvo Molecular , Oxirredução , Transdução de Sinais
10.
Cancer Sci ; 107(2): 140-8, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26583567

RESUMO

Recent strategies for treating CML patients have focused on investigating new combinations of tyrosine kinase inhibitors (TKIs) as well as identifying novel translational research agents that can eradicate CML leukemia-initiating cells (CML-LICs). However, little is known about the therapeutic benefits such CML-LIC targeting therapies might bring to CML patients. In this study, we investigated the therapeutic potential of EW-7197, an orally bioavailable transforming growth factor-ß signaling inhibitor which has recently been approved as an Investigational New Drug (NIH, USA), to suppress CML-LICs in vivo. Compared to TKI treatment alone, administration of TKI plus EW-7197 to CML-affected mice significantly delayed disease relapse and prolonged survival. Notably, combined treatment with EW-7197 plus TKI was effective in eliminating CML-LICs even if they expressed the TKI-resistant T315I mutant BCR-ABL1 oncogene. Collectively, these results indicate that EW-7197 may be a promising candidate for a new therapeutic that can greatly benefit CML patients by working in combination with TKIs to eradicate CML-LICs.


Assuntos
Compostos de Anilina/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Triazóis/farmacologia , Animais , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Humanos , Imidazóis/administração & dosagem , Camundongos , Camundongos Endogâmicos C57BL , Inibidores de Proteínas Quinases/administração & dosagem , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Piridazinas/administração & dosagem , Receptor do Fator de Crescimento Transformador beta Tipo I , Receptores de Fatores de Crescimento Transformadores beta/antagonistas & inibidores , Transfecção , Fator de Crescimento Transformador beta/antagonistas & inibidores
11.
Nature ; 463(7281): 676-80, 2010 Feb 04.
Artigo em Inglês | MEDLINE | ID: mdl-20130650

RESUMO

Chronic myeloid leukaemia (CML) is caused by a defined genetic abnormality that generates BCR-ABL, a constitutively active tyrosine kinase. It is widely believed that BCR-ABL activates Akt signalling that suppresses the forkhead O transcription factors (FOXO), supporting the proliferation or inhibiting the apoptosis of CML cells. Although the use of the tyrosine kinase inhibitor imatinib is a breakthrough for CML therapy, imatinib does not deplete the leukaemia-initiating cells (LICs) that drive the recurrence of CML. Here, using a syngeneic transplantation system and a CML-like myeloproliferative disease mouse model, we show that Foxo3a has an essential role in the maintenance of CML LICs. We find that cells with nuclear localization of Foxo3a and decreased Akt phosphorylation are enriched in the LIC population. Serial transplantation of LICs generated from Foxo3a(+/+) and Foxo3a(-/-) mice shows that the ability of LICs to cause disease is significantly decreased by Foxo3a deficiency. Furthermore, we find that TGF-beta is a critical regulator of Akt activation in LICs and controls Foxo3a localization. A combination of TGF-beta inhibition, Foxo3a deficiency and imatinib treatment led to efficient depletion of CML in vivo. Furthermore, the treatment of human CML LICs with a TGF-beta inhibitor impaired their colony-forming ability in vitro. Our results demonstrate a critical role for the TGF-beta-FOXO pathway in the maintenance of LICs, and strengthen our understanding of the mechanisms that specifically maintain CML LICs in vivo.


Assuntos
Fatores de Transcrição Forkhead/metabolismo , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismo , Animais , Antineoplásicos/uso terapêutico , Apoptose , Benzamidas , Diferenciação Celular , Linhagem Celular Tumoral , Núcleo Celular/metabolismo , Modelos Animais de Doenças , Ativação Enzimática , Proteína Forkhead Box O3 , Fatores de Transcrição Forkhead/deficiência , Fatores de Transcrição Forkhead/genética , Humanos , Mesilato de Imatinib , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Camundongos , Camundongos Endogâmicos C57BL , Células-Tronco Neoplásicas/efeitos dos fármacos , Fosforilação , Piperazinas/uso terapêutico , Inibidores de Proteínas Quinases/uso terapêutico , Transporte Proteico , Proteínas Tirosina Quinases/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo , Pirimidinas/uso terapêutico , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta/antagonistas & inibidores , Ensaio Tumoral de Célula-Tronco
12.
Genes Chromosomes Cancer ; 54(3): 142-55, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25521327

RESUMO

Homozygous deletion is a frequent mutational mechanism of silencing tumor suppressor genes in cancer. Therefore, homozygous deletions have been analyzed for identification of tumor suppressor genes that can be utilized as biomarkers or therapeutic targets for cancer treatment. In this study, to elucidate potential tumor suppressor genes involved in gastric cancer (GC), we analyzed the entire set of large homozygous deletions in six human GC cell lines through genome- and transcriptome-wide approaches. We identified 51 genes in homozygous deletion regions of chromosomes and confirmed the deletion frequency in tumor tissues of 219 GC patients from The Cancer Genome Atlas database. We evaluated the effect of homozygous deletions on the mRNA level and found significantly affected genes in chromosome bands 9p21, 3p22, 5p14, and 6q15. Among the genes in 9p21, we investigated the potential tumor suppressive effect of KLHL9. We demonstrated that ectopic expression of KLHL9 inhibited cell proliferation and tumor formation in KLHL9-deficient SNU-16 cell line. In addition, we observed that homozygous focal deletions generated truncated transcripts of TGFBR2, CTNNA1, and STXBP5. Ectopic expression of two kinds of TGFBR2-reverse GADL1 fusion genes suppressed TGF-ß signaling, which may lead to the loss of sensitivity to TGF-ß tumor suppressive activity. In conclusion, our findings suggest that novel tumor suppressor genes that are aberrantly expressed through homozygous deletions may play important roles in gastric tumorigenesis.


Assuntos
Deleção Cromossômica , Regulação Neoplásica da Expressão Gênica , Genes Supressores de Tumor , Neoplasias Gástricas/genética , Animais , Linhagem Celular Tumoral , Cromossomos Humanos Par 3 , Cromossomos Humanos Par 5 , Cromossomos Humanos Par 6 , Cromossomos Humanos Par 9 , Feminino , Humanos , Camundongos , Camundongos Nus , Proteínas Serina-Treonina Quinases/genética , RNA Mensageiro/metabolismo , Receptor do Fator de Crescimento Transformador beta Tipo II , Receptores de Fatores de Crescimento Transformadores beta/genética
13.
Nihon Rinsho ; 73(5): 784-9, 2015 May.
Artigo em Japonês | MEDLINE | ID: mdl-25985631

RESUMO

Cancer stem cells are the cellular sources of the vast majority of mature cancer cells and are reportedly responsible for the recurrence of disease following anti-cancer therapy. Recent advantages in the cancer stem cell research field have indicated that TGF-ß signaling pathway plays an essential role for the maintenance of cancer stem cells via regulating cancer stemness, quiescence, epithelial -mesenchymal transition (EMT), or therapeutic resistance in vivo. Therefore, the outcome of these investigations will hopefully be the development of novel agents that can specifically control both augmentation and suppression of TGF-ß signaling pathway in cancer stem cells, and thereby provide a novel avenue for curative cancer patient therapy.


Assuntos
Células-Tronco Neoplásicas/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismo , Antineoplásicos/uso terapêutico , Transição Epitelial-Mesenquimal , Humanos , Células-Tronco Neoplásicas/efeitos dos fármacos , Nicho de Células-Tronco
14.
Cancer Sci ; 105(4): 418-24, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24447505

RESUMO

RUNX3 is a tumor suppressor for a variety of cancers. RUNX3 suppresses the canonical Wnt signaling pathway by binding to the TCF4/ß-catenin complex, resulting in the inhibition of binding of the complex to the Wnt target gene promoter. Here, we confirmed that RUNX3 suppressed Wnt signaling activity in several gastric cancer cell lines; however, we found that RUNX3 increased the Wnt signaling activity in KatoIII and SNU668 gastric cancer cells. Notably, RUNX3 expression increased the ratio of the Wnt signaling-high population in the KatoIII cells. although the maximum Wnt activation level of individual cells was similar to that in the control. As found previously, RUNX3 also binds to TCF4 and ß-catenin in KatoIII cells, suggesting that these molecules form a ternary complex. Moreover, the ChIP analyses revealed that TCF4, ß-catenin and RUNX3 bind the promoter region of the Wnt target genes, Axin2 and c-Myc, and the occupancy of TCF4 and ß-catenin in these promoter regions is increased by the RUNX3 expression. These results suggest that RUNX3 stabilizes the TCF4/ß-catenin complex on the Wnt target gene promoter in KatoIII cells, leading to activation of Wnt signaling. Although RUNX3 increased the Wnt signaling activity, its expression resulted in suppression of tumorigenesis of KatoIII cells, indicating that RUNX3 plays a tumor-suppressing role in KatoIII cells through a Wnt-independent mechanism. These results indicate that RUNX3 can either suppress or activate the Wnt signaling pathway through its binding to the TCF4/ß-catenin complex by cell context-dependent mechanisms.


Assuntos
Subunidade alfa 3 de Fator de Ligação ao Core/genética , Neoplasias Gástricas/genética , Ativação Transcricional , Via de Sinalização Wnt/genética , Proteína Axina/metabolismo , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Linhagem Celular Tumoral , Subunidade alfa 3 de Fator de Ligação ao Core/biossíntese , Regulação Neoplásica da Expressão Gênica , Humanos , Ligação Proteica , Proteínas Proto-Oncogênicas c-myc/metabolismo , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/terapia , Fator de Transcrição 4 , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , beta Catenina/genética , beta Catenina/metabolismo
15.
Biochem Biophys Res Commun ; 450(1): 837-43, 2014 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-24960197

RESUMO

Acute myeloid leukaemia (AML) is a heterogeneous neoplastic disorder in which a subset of cells function as leukaemia-initiating cells (LICs). In this study, we prospectively evaluated the leukaemia-initiating capacity of AML cells fractionated according to the expression of a nucleolar GTP binding protein, nucleostemin (NS). To monitor NS expression in living AML cells, we generated a mouse AML model in which green fluorescent protein (GFP) is expressed under the control of a region of the NS promoter (NS-GFP). In AML cells, NS-GFP levels were correlated with endogenous NS mRNA. AML cells with the highest expression of NS-GFP were very immature blast-like cells, efficiently formed leukaemia colonies in vitro, and exhibited the highest leukaemia-initiating capacity in vivo. Gene expression profiling analysis revealed that cell cycle regulators and nucleotide metabolism-related genes were highly enriched in a gene set associated with leukaemia-initiating capacity that we termed the 'leukaemia stem cell gene signature'. This gene signature stratified human AML patients into distinct clusters that reflected prognosis, demonstrating that the mouse leukaemia stem cell gene signature is significantly associated with the malignant properties of human AML. Further analyses of gene regulation in leukaemia stem cells could provide novel insights into diagnostic and therapeutic approaches to AML.


Assuntos
Proteínas de Ligação ao GTP/genética , Predisposição Genética para Doença/genética , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Proteínas Nucleares/genética , Regiões Promotoras Genéticas/genética , Animais , Transformação Celular Neoplásica/metabolismo , Transformação Celular Neoplásica/patologia , Marcadores Genéticos/genética , Humanos , Camundongos , Camundongos Transgênicos , Prognóstico
16.
Am J Pathol ; 183(2): 592-603, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23885716

RESUMO

Nucleostemin (NS) is a nucleolar GTP-binding protein that is involved in ribosomal biogenesis and protection of telomeres. We investigated the expression of NS in human germ cell tumors and its function in a mouse germ cell tumor model. NS was abundantly expressed in undifferentiated, but not differentiated, types of human testicular germ cell tumors. NS was expressed concomitantly with OCT3/4, a critical regulator of the undifferentiated status of pluripotent stem cells in primordial germ cells and embryonal carcinomas. To investigate the roles of NS in tumor growth in vivo, we used a mouse teratoma model. Analysis of teratomas derived from embryonic stem cells in which the NS promoter drives GFP expression showed that cells highly expressing NS were actively proliferating and exhibited the characteristics of tumor-initiating cells, including the ability to initiate and propagate tumor cells in vivo. NS-expressing cells exhibited higher levels of GTP than non-NS-expressing cells. Because NS protein is stabilized by intracellular GTP, metabolic changes may contribute to abundant NS expression in the undifferentiated cells. OCT3/4 deficiency in teratomas led to loss of NS expression, resulting in growth retardation. Finally, we found that teratomas deficient in NS lost their undifferentiated characteristics, resulting in defective tumor proliferation. These data indicate that abundant expression of NS supports the undifferentiated properties of germ cell tumors.


Assuntos
Proteínas de Transporte/metabolismo , Proteínas Nucleares/metabolismo , Teratoma/metabolismo , Neoplasias Testiculares/metabolismo , Animais , Proliferação de Células , Modelos Animais de Doenças , Proteínas de Ligação ao GTP , Células Germinativas/patologia , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Proteínas de Ligação a RNA , Teratoma/patologia , Neoplasias Testiculares/patologia , Células Tumorais Cultivadas
17.
J Allergy Clin Immunol ; 131(6): 1674-82, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23534976

RESUMO

BACKGROUND: Regulatory B cells that exhibit the cell-surface CD1d(hi)CD5(+) phenotype and produce IL-10 are termed B10 cells. Although B10 cells exert potent suppressive functions in patients with various allergic and autoimmunity disorders, the precise signaling mechanisms required for B10 cell functions remain unknown. B-cell linker protein (BLNK) is an essential component of the B-cell antigen receptor signaling pathway and is required for optimal B-cell development. OBJECTIVE: We sought to elucidate the signaling pathways that are responsible for IL-10 production in B10 cells and in vivo mechanisms of how impaired B10 cell functions influence allergic and autoimmune responses. METHOD: For in vitro assays, splenic CD1d(hi)CD5(+) B cells from BLNK-deficient (BLNK(-/-)) mice were analyzed for intracellular signaling pathways and cytokine production. Contact hypersensitivity (CHS) and experimental autoimmune encephalomyelitis were examined by using BLNK(-/-) mice. RESULTS: Although the CD1d(hi)CD5(+) B-cell population was present in BLNK(-/-) mice, IL-10 production was impaired both in vitro and in vivo. BLNK(-/-) mice had exaggerated CHS and experimental autoimmune encephalomyelitis responses, which were normalized by adoptive transfer of splenic CD1d(hi)CD5(+) B cells from wild-type mice. In mice with CHS, BLNK(-/-) mice exhibited decreased B-cell and regulatory T-cell percentages and increased CD8(+) T-cell percentages in the skin and lymph nodes. In vitro BLNK was required for LPS-induced signal transducer and activator of transcription 3 phosphorylation in CD1d(hi)CD5(+) B cells. Finally, secreted IL-10 leads to autocrine expansion of IL-10-producing B cells. CONCLUSION: BLNK serves as a critical signaling component for B10 cell function by mediating IL-10 production.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Doenças Autoimunes/genética , Doenças Autoimunes/imunologia , Linfócitos B Reguladores/imunologia , Hipersensibilidade/genética , Hipersensibilidade/imunologia , Interleucina-10/biossíntese , Proteínas Adaptadoras de Transdução de Sinal/deficiência , Animais , Antígenos CD1d/metabolismo , Comunicação Autócrina , Linfócitos B Reguladores/metabolismo , Antígenos CD5/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Citocinas/biossíntese , Dermatite de Contato/genética , Dermatite de Contato/imunologia , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/genética , Encefalomielite Autoimune Experimental/imunologia , Expressão Gênica , Janus Quinases/metabolismo , Lipopolissacarídeos/imunologia , Contagem de Linfócitos , Camundongos , Camundongos Knockout , NF-kappa B/metabolismo , Fosforilação , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais
18.
Nat Med ; 12(4): 446-51, 2006 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-16565722

RESUMO

Hematopoietic stem cells (HSCs) undergo self-renewing cell divisions and maintain blood production for their lifetime. Appropriate control of HSC self-renewal is crucial for the maintenance of hematopoietic homeostasis. Here we show that activation of p38 MAPK in response to increasing levels of reactive oxygen species (ROS) limits the lifespan of HSCs in vivo. In Atm(-/-) mice, elevation of ROS levels induces HSC-specific phosphorylation of p38 MAPK accompanied by a defect in the maintenance of HSC quiescence. Inhibition of p38 MAPK rescued ROS-induced defects in HSC repopulating capacity and in the maintenance of HSC quiescence, indicating that the ROS-p38 MAPK pathway contributes to exhaustion of the stem cell population. Furthermore, prolonged treatment with an antioxidant or an inhibitor of p38 MAPK extended the lifespan of HSCs from wild-type mice in serial transplantation experiments. These data show that inactivation of p38 MAPK protects HSCs against loss of self-renewal capacity. Our characterization of molecular mechanisms that limit HSC lifespan may lead to beneficial therapies for human disease.


Assuntos
Senescência Celular/fisiologia , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Butionina Sulfoximina/farmacologia , Técnicas de Cultura de Células , Linhagem Celular , Técnicas de Cocultura , Relação Dose-Resposta a Droga , Ativação Enzimática , Inibidores Enzimáticos/farmacologia , Citometria de Fluxo , Sequestradores de Radicais Livres/farmacologia , Imidazóis/farmacologia , Imuno-Histoquímica , Camundongos , Camundongos Congênicos , Camundongos Endogâmicos C57BL , Estresse Oxidativo , Fosforilação , Piridinas/farmacologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Regulação para Cima/efeitos dos fármacos , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores
19.
Cancer Gene Ther ; 30(1): 38-50, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-35999358

RESUMO

The Philadelphia (Ph) chromosome was the first translocation identified in leukemia. It is supposed to be generated by aberrant ligation between two DNA double-strand breaks (DSBs) at the BCR gene located on chromosome 9q34 and the ABL1 gene located on chromosome 22q11. Thus, mimicking the initiation process of translocation, we induced CRISPR/Cas9-mediated DSBs simultaneously at the breakpoints of the BCR and ABL1 genes in a granulocyte-macrophage colony-stimulating factor (GM-CSF) dependent human leukemia cell line. After transfection of two single guide RNAs (sgRNAs) targeting intron 13 of the BCR gene and intron 1 of the ABL1 gene, a factor-independent subline was obtained. In the subline, p210 BCR::ABL1 and its reciprocal ABL1::BCR fusions were generated as a result of balanced translocation corresponding to the Ph chromosome. Another set of sgRNAs targeting intron 1 of the BCR gene and intron 1 of the ABL1 gene induced a factor-independent subline expressing p190 BCR::ABL1. Both p210 and p190 BCR::ABL1 induced factor-independent growth by constitutively activating intracellular signaling pathways for transcriptional regulation of cell cycle progression and cell survival that are usually regulated by GM-CSF. These observations suggested that simultaneous DSBs at the BCR and ABL1 gene breakpoints are initiation events for oncogenesis in Ph+ leukemia. (200/200 words).


Assuntos
Leucemia Mielogênica Crônica BCR-ABL Positiva , Cromossomo Filadélfia , Humanos , Proteínas de Fusão bcr-abl/genética , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Sistemas CRISPR-Cas , Translocação Genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Carcinogênese/genética
20.
EMBO J ; 27(12): 1671-81, 2008 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-18511911

RESUMO

The activation of Wnt/beta-catenin signalling has an important function in gastrointestinal tumorigenesis. It has been suggested that the promotion of Wnt/beta-catenin activity beyond the threshold is important for carcinogenesis. We herein investigated the role of macrophages in the promotion of Wnt/beta-catenin activity in gastric tumorigenesis. We found beta-catenin nuclear accumulation in macrophage-infiltrated dysplastic mucosa of the K19-Wnt1 mouse stomach. Moreover, macrophage depletion in Apc(Delta716) mice resulted in the suppression of intestinal tumorigenesis. These results suggested the role of macrophages in the activation of Wnt/beta-catenin signalling, which thus leads to tumour development. Importantly, the conditioned medium of activated macrophages promoted Wnt/beta-catenin signalling in gastric cancer cells, which was suppressed by the inhibition of tumour necrosis factor (TNF)-alpha. Furthermore, treatment with TNF-alpha induced glycogen synthase kinase 3beta (GSK3beta) phosphorylation, which resulted in the stabilization of beta-catenin. We also found that Helicobacter infection in the K19-Wnt1 mouse stomach caused mucosal macrophage infiltration and nuclear beta-catenin accumulation. These results suggest that macrophage-derived TNF-alpha promotes Wnt/beta-catenin signalling through inhibition of GSK3beta, which may contribute to tumour development in the gastric mucosa.


Assuntos
Ativação de Macrófagos , Transdução de Sinais , Neoplasias Gástricas/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Proteína Wnt1/metabolismo , Proteína da Polipose Adenomatosa do Colo/metabolismo , Animais , Linhagem Celular Tumoral , Núcleo Celular/metabolismo , Células Epiteliais/microbiologia , Quinase 3 da Glicogênio Sintase/antagonistas & inibidores , Glicogênio Sintase Quinase 3 beta , Infecções por Helicobacter/patologia , Humanos , Camundongos , Camundongos Transgênicos , Mutação/genética , NF-kappa B/metabolismo , Fosforilação , Lesões Pré-Cancerosas/enzimologia , Lesões Pré-Cancerosas/patologia , Estômago/microbiologia , Estômago/patologia , Neoplasias Gástricas/enzimologia , Neoplasias Gástricas/patologia , beta Catenina/metabolismo
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