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1.
Int J Mol Sci ; 22(13)2021 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-34201814

RESUMO

High-risk neuroblastoma (HR-NB) still remains the most dangerous tumor in early childhood. For this reason, the identification of new therapeutic approaches is of fundamental importance. Recently, we combined the conventional pharmacological approach to NB, represented by cisplatin, with fendiline hydrochloride, an inhibitor of several transporters involved in multidrug resistance of cancer cells, which demonstrated an enhancement of the ability of cisplatin to induce apoptosis. In this work, we co-administrated acetazolamide, a carbonic anhydrase isoform IX (CAIX) inhibitor which was reported to increase chemotherapy efficacy in various cancer types, to the cisplatin/fendiline approach in SKNBE2 xenografts in NOD-SCID mice with the aim of identifying a novel and more effective treatment. We observed that the combination of the three drugs increases more than twelvefold the differences in the cytotoxic activity of cisplatin alone, leading to a remarkable decrease of the expression of malignancy markers. Our conclusion is that this approach, based on three FDA-approved drugs, may constitute an appropriate improvement of the pharmacological approach to HR-NB.


Assuntos
Acetazolamida/farmacologia , Antineoplásicos/farmacologia , Bloqueadores dos Canais de Cálcio/farmacologia , Inibidores da Anidrase Carbônica/farmacologia , Cisplatino/farmacologia , Fendilina/farmacologia , Neuroblastoma/tratamento farmacológico , Animais , Apoptose , Proliferação de Células , Quimioterapia Combinada , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Neuroblastoma/patologia , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
2.
Nat Commun ; 12(1): 4245, 2021 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-34253722

RESUMO

Tuberous Sclerosis Complex (TSC) is caused by TSC1 or TSC2 mutations, resulting in hyperactivation of the mechanistic target of rapamycin complex 1 (mTORC1). Transcription factor EB (TFEB), a master regulator of lysosome biogenesis, is negatively regulated by mTORC1 through a RAG GTPase-dependent phosphorylation. Here we show that lysosomal biogenesis is increased in TSC-associated renal tumors, pulmonary lymphangioleiomyomatosis, kidneys from Tsc2+/- mice, and TSC1/2-deficient cells via a TFEB-dependent mechanism. Interestingly, in TSC1/2-deficient cells, TFEB is hypo-phosphorylated at mTORC1-dependent sites, indicating that mTORC1 is unable to phosphorylate TFEB in the absence of the TSC1/2 complex. Importantly, overexpression of folliculin (FLCN), a GTPase activating protein for RAGC, increases TFEB phosphorylation at the mTORC1 sites in TSC2-deficient cells. Overexpression of constitutively active RAGC is sufficient to relocalize TFEB to the cytoplasm. These findings establish the TSC proteins as critical regulators of lysosomal biogenesis via TFEB and RAGC and identify TFEB as a driver of the proliferation of TSC2-deficient cells.


Assuntos
Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Lisossomos/metabolismo , Proteínas Monoméricas de Ligação ao GTP/metabolismo , Biogênese de Organelas , Proteína 2 do Complexo Esclerose Tuberosa/metabolismo , Animais , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Carcinoma de Células Renais/patologia , Núcleo Celular/metabolismo , Proliferação de Células , Feminino , Regulação da Expressão Gênica , Células HEK293 , Células HeLa , Humanos , Neoplasias Renais/patologia , Lisossomos/ultraestrutura , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Fosforilação , Fosfosserina/metabolismo , Transporte Proteico , Proteínas Proto-Oncogênicas/metabolismo , Transcrição Genética , Proteína 2 do Complexo Esclerose Tuberosa/deficiência , Proteínas Supressoras de Tumor/metabolismo
3.
Magy Onkol ; 65(2): 113-120, 2021 Jun 03.
Artigo em Húngaro | MEDLINE | ID: mdl-34081759

RESUMO

The in vivo antitumor effect of two NGR sequence containing peptide-daunomycin conjugates was studied on CD13+ Kaposi's sarcoma s.c. tumor model on SCID mice, and on orthotopically developed CD13- HT-29 colon adenocarcinoma SCID mouse model. Both tumor types were positive for integrins. Significant tumor growth inhibition was observed on both tumor types by the treatment with the conjugates (Dau=Aoa-GFLGK(cyclo[KNGRE]-GG)-NH2 (1) and Dau=Aoa-GFLGK(cyclo[NleNGRE]-GG)-NH2 (2)). KS conjugate 1 with rather stable construct was more potent in tumor growth inhibition that might be explained by the CD13 receptor recognition of NGR sequence. In contrast, conjugate 2 that has propensity to rearrange isoAsp derivative showed significantly higher inhibition on CD13- HT-29 tumor model that is related to the integrin binding of isoDGR sequence. Next to the low toxic side effect of the conjugates in comparison with the free daunomycin, the positive efficiency of the conjugates was detected by the lower proliferation index and lower neovascularization of the tumor tissue.


Assuntos
Antígenos CD13 , Peptídeos Cíclicos , Animais , Linhagem Celular Tumoral , Humanos , Camundongos , Camundongos SCID , Oligopeptídeos
4.
Int J Mol Sci ; 22(10)2021 May 13.
Artigo em Inglês | MEDLINE | ID: mdl-34068008

RESUMO

A major contributing factor in triple-negative breast cancer progression is its ability to evade immune surveillance. One mechanism for this immunosuppression is through ribosomal protein S19 (RPS19), which facilitates myeloid-derived suppressor cells (MDSCs) recruitment in tumors, which generate cytokines TGF-ß and IL-10 and induce regulatory T cells (Tregs), all of which are immunosuppressive and enhance tumor progression. Hence, enhancing the immune system in breast tumors could be a strategy for anticancer therapeutics. The present study evaluated the immune response of atovaquone, an antiprotozoal drug, in three independent breast-tumor models. Our results demonstrated that oral administration of atovaquone reduced HCC1806, CI66 and 4T1 paclitaxel-resistant (4T1-PR) breast-tumor growth by 45%, 70% and 42%, respectively. MDSCs, TGF-ß, IL-10 and Tregs of blood and tumors were analyzed from all of these in vivo models. Our results demonstrated that atovaquone treatment in mice bearing HCC1806 tumors reduced MDSCs from tumor and blood by 70% and 30%, respectively. We also observed a 25% reduction in tumor MDSCs in atovaquone-treated mice bearing CI66 and 4T1-PR tumors. In addition, a decrease in TGF-ß and IL-10 in tumor lysates was observed in atovaquone-treated mice with a reduction in tumor Tregs. Moreover, a significant reduction in the expression of RPS19 was found in tumors treated with atovaquone.


Assuntos
Anti-Infecciosos/farmacologia , Apresentação do Antígeno/imunologia , Atovaquona/farmacologia , Imunossupressão , Células Supressoras Mieloides/imunologia , Linfócitos T Reguladores/imunologia , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Animais , Apoptose , Proliferação de Células , Citocinas/metabolismo , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos NOD , Camundongos SCID , Células Supressoras Mieloides/efeitos dos fármacos , Linfócitos T Reguladores/efeitos dos fármacos , Neoplasias de Mama Triplo Negativas/metabolismo , Neoplasias de Mama Triplo Negativas/patologia , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
5.
Int J Mol Sci ; 22(11)2021 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-34070369

RESUMO

Folate receptor beta (FRß) is a folate binding receptor expressed on myeloid lineage hematopoietic cells. FRß is commonly expressed at high levels on malignant blasts in patients with acute myeloid leukemia (AML), as well as on M2 polarized tumor-associated macrophages (TAMs) in the tumor microenvironment of many solid tumors. Therefore, FRß is a potential target for both direct and indirect cancer therapy. We demonstrate that FRß is expressed in both AML cell lines and patient-derived AML samples and that a high-affinity monoclonal antibody against FRß (m909) has the ability to cause dose- and expression-dependent ADCC against these cells in vitro. Importantly, we find that administration of m909 has a significant impact on tumor growth in a humanized mouse model of AML. Surprisingly, m909 functions in vivo with and without the infusion of human NK cells as mediators of ADCC, suggesting potential involvement of mouse macrophages as effector cells. We also found that TAMs from primary ovarian ascites samples expressed appreciable levels of FRß and that m909 has the ability to cause ADCC in these samples. These results indicate that the targeting of FRß using m909 has the potential to limit the outgrowth of AML in vitro and in vivo. Additionally, m909 causes cytotoxicity to TAMs in the tumor microenvironment of ovarian cancer warranting further investigation of m909 and its derivatives as therapeutic agents in patients with FRß-expressing cancers.


Assuntos
Antineoplásicos Imunológicos/farmacologia , Receptor 2 de Folato , Imunoterapia , Leucemia Mieloide Aguda , Proteínas de Neoplasias , Neoplasias Ovarianas , Animais , Células CHO , Cricetulus , Feminino , Receptor 2 de Folato/antagonistas & inibidores , Receptor 2 de Folato/imunologia , Células HL-60 , Humanos , Leucemia Mieloide Aguda/imunologia , Leucemia Mieloide Aguda/patologia , Leucemia Mieloide Aguda/terapia , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/imunologia , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/patologia , Neoplasias Ovarianas/terapia , Células THP-1 , Ensaios Antitumorais Modelo de Xenoenxerto
6.
Int J Mol Sci ; 22(11)2021 May 26.
Artigo em Inglês | MEDLINE | ID: mdl-34073327

RESUMO

Mesenchymal stromal cells (MSC) are promising candidates for regenerative therapy of the infarcted heart. However, poor cell retention within the transplantation site limits their potential. We hypothesized that MSC benefits could be enhanced through a dual-cell approach using jointly endothelial colony forming cells (ECFC) and MSC. To assess this, we comparatively evaluated the effects of the therapy with MSC and ECFC versus MSC-only in a mouse model of myocardial infarction. Heart function was assessed by echocardiography, and the molecular crosstalk between MSC and ECFC was evaluated in vitro through direct or indirect co-culture systems. We found that dual-cell therapy improved cardiac function in terms of ejection fraction and stroke volume. In vitro experiments showed that ECFC augmented MSC effector properties by increasing Connexin 43 and Integrin alpha-5 and the secretion of healing-associated molecules. Moreover, MSC prompted the organization of ECFC into vascular networks. This indicated a reciprocal modulation in the functionality of MSC and ECFC. In conclusion, the crosstalk between MSC and ECFC augments the therapeutic properties of MSC and enhances the angiogenic properties of ECFC. Our data consolidate the dual-cell therapy as a step forward for the development of effective treatments for patients affected by myocardial infarction.


Assuntos
Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais , Infarto do Miocárdio , Miocárdio , Volume Sistólico , Animais , Células Progenitoras Endoteliais/metabolismo , Células Progenitoras Endoteliais/patologia , Células Progenitoras Endoteliais/transplante , Feminino , Xenoenxertos , Masculino , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/patologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Infarto do Miocárdio/fisiopatologia , Infarto do Miocárdio/terapia , Miocárdio/metabolismo , Miocárdio/patologia
7.
Int J Mol Sci ; 22(11)2021 May 23.
Artigo em Inglês | MEDLINE | ID: mdl-34071152

RESUMO

Prostate cancer (PCa) is the second most common cancer in men, causing more than 300,000 deaths every year worldwide. Due to their superior cell-killing ability and the relative simplicity of their preparation, immunotoxin molecules have great potential in the clinical treatment of cancer, and several such molecules have been approved for clinical application. In this study, we adopted a relatively simple strategy based on a single-domain antibody (sdAb) and an improved Pseudomonas exotoxin A (PE) toxin (PE24X7) to prepare a safer immunotoxin against prostate-specific membrane antigen (PSMA) for PCa treatment. The designed anti-PSMA immunotoxin, JVM-PE24X7, was conveniently prepared in its soluble form in an Escherichia coli (E. coli) system, avoiding the complex renaturation process needed for immunotoxin preparation by the conventional strategy. The product was very stable and showed a very strong ability to bind the PSMA receptor. Cytotoxicity assays showed that this molecule at a very low concentration could kill PSMA-positive PCa cells, with an EC50 value (concentration at which the cell viability decreased by 50%) of 15.3 pM against PSMA-positive LNCaP cells. Moreover, this molecule showed very good killing selectivity between PSMA-positive and PSMA-negative cells, with a selection ratio of more than 300-fold. Animal studies showed that this molecule at a very low dosage (5 × 0.5 mg/kg once every three days) completely inhibited the growth of PCa tumors, and the maximum tolerable dose (MTD) was more than 15 mg/kg, indicating its very potent tumor-treatment ability and a wide therapeutic window. Use of the new PE toxin, PE24X7, as the effector moiety significantly reduced off-target toxicity and improved the therapeutic window of the immunotoxin. The above results demonstrate that the designed anti-PSMA immunotoxin, JVM-PE24X7, has good application value for the treatment of PCa.


Assuntos
Adenocarcinoma/tratamento farmacológico , Antígenos de Neoplasias/imunologia , Antineoplásicos Imunológicos/uso terapêutico , Glutamato Carboxipeptidase II/antagonistas & inibidores , Imunotoxinas/uso terapêutico , Terapia de Alvo Molecular , Neoplasias da Próstata/tratamento farmacológico , Anticorpos de Domínio Único/uso terapêutico , Animais , Especificidade de Anticorpos , Reações Antígeno-Anticorpo , Antígenos de Superfície/imunologia , Antineoplásicos Imunológicos/toxicidade , Linhagem Celular Tumoral , Ensaios de Seleção de Medicamentos Antitumorais , Glutamato Carboxipeptidase II/imunologia , Humanos , Imunotoxinas/toxicidade , Masculino , Dose Máxima Tolerável , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Modelos Moleculares , Conformação Proteica , Proteínas Recombinantes de Fusão/uso terapêutico , Proteínas Recombinantes de Fusão/toxicidade , Anticorpos de Domínio Único/toxicidade , Ensaios Antitumorais Modelo de Xenoenxerto
8.
Molecules ; 26(9)2021 May 10.
Artigo em Inglês | MEDLINE | ID: mdl-34068627

RESUMO

α-Trifluoromethyl chalcones were prepared and evaluated for their antiproliferative activities against androgen-independent prostate cancer cell lines as well as five additional types of human tumor cell lines. The most potent chalcone 5 showed superior antitumor activity in vivo with both oral and intraperitoneal administration at 3 mg/kg. Cell-based mechanism of action studies demonstrated that 5 induced cell accumulation at sub-G1 and G2/M phases without interfering with microtubule polymerization. Furthermore, several cancer cell growth-related proteins were identified by using chalcone 5 as a bait for the affinity purification of binding proteins.


Assuntos
Antineoplásicos/farmacologia , Chalconas/farmacologia , Neoplasias da Próstata/patologia , Receptores Androgênicos/metabolismo , Animais , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Chalconas/química , Docetaxel/farmacologia , Humanos , Masculino , Camundongos SCID , Taxoides/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
9.
J Stroke Cerebrovasc Dis ; 30(8): 105932, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34148020

RESUMO

OBJECTIVES: Bone marrow mononuclear cells (BM-MNC) show a significant therapeutic effect in combination with training even in the chronic phase of stroke. However, the mechanism of this combination therapy has not been investigated. Here, we examined its effects on brain metabolism in chronic stroke mice. MATERIALS AND METHODS: BM-MNC (1x105 cells in 100 µL of phosphate-buffered saline) were intravenously transplanted at 4 weeks (chronic stage) after the middle cerebral artery occlusion. At 3 h and 10 weeks after the administration of BM-MNC, we evaluated transcription changes of the metabolism-related genes, hypoxia inducible factor 1-α (Hif-1α), prolyl hydroxylase 3 (Phd3), pyruvate dehydrogenase kinase 1 (Pdk1), Na+/K+-ATPase (Atp1α1‒3), connexins, glucose transporters, and monocarboxylate transporters, in the brain during chronic phase of stroke using quantitative polymerase chain reaction. RESULTS: The results showed transcriptional activation of the metabolism-related genes in the contralateral cortex at 3 h after BM-MNC transplantation. Behavioral tests were performed after cell therapy, and the brain metabolism of mice with improved motor function was examined at 10 weeks after cell therapy. The therapeutic efficacy of the combination therapy with BM-MNC transplantation and training was evident in the form of transcriptional activation of ipsilateral anterior cerebral artery (ACA) cortex. CONCLUSIONS: BM-MNC transplantation combined with training for chronic stroke activated gene expression in both the ipsilateral and the contralateral side.


Assuntos
Células da Medula Óssea/metabolismo , Transplante de Medula Óssea , Encéfalo/metabolismo , Metabolismo Energético , Infarto da Artéria Cerebral Média/terapia , Condicionamento Físico Animal , Animais , Comportamento Animal , Encéfalo/fisiopatologia , Doença Crônica , Terapia Combinada , Conexinas/genética , Conexinas/metabolismo , Modelos Animais de Doenças , Metabolismo Energético/genética , Regulação da Expressão Gênica , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Infarto da Artéria Cerebral Média/genética , Infarto da Artéria Cerebral Média/metabolismo , Infarto da Artéria Cerebral Média/fisiopatologia , Masculino , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Camundongos Endogâmicos C57BL , Camundongos SCID , Atividade Motora , Recuperação de Função Fisiológica , ATPase Trocadora de Sódio-Potássio/genética , ATPase Trocadora de Sódio-Potássio/metabolismo , Transcrição Genética
10.
Int J Mol Sci ; 22(11)2021 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-34071237

RESUMO

Cell division cycle 25A (Cdc25A) is a dual-specificity phosphatase that is overexpressed in several cancer cells and promotes tumorigenesis. In normal cells, Cdc25A expression is regulated tightly, but the changes in expression patterns in cancer cells that lead to tumorigenesis are unknown. In this study, we showed that ubiquitin-specific protease 29 (USP29) stabilized Cdc25A protein expression in cancer cell lines by protecting it from ubiquitin-mediated proteasomal degradation. The presence of USP29 effectively blocked polyubiquitination of Cdc25A and extended its half-life. CRISPR-Cas9-mediated knockdown of USP29 in HeLa cells resulted in cell cycle arrest at the G0/G1 phase. We also showed that USP29 knockdown hampered Cdc25A-mediated cell proliferation, migration, and invasion of cancer cells in vitro. Moreover, NSG nude mice transplanted with USP29-depleted cells significantly reduced the size of the tumors, whereas the reconstitution of Cdc25A in USP29-depleted cells significantly increased the tumor size. Altogether, our results implied that USP29 promoted cell cycle progression and oncogenic transformation by regulating protein turnover of Cdc25A.


Assuntos
Carcinogênese/metabolismo , Regulação Neoplásica da Expressão Gênica , Proteases Específicas de Ubiquitina/metabolismo , Fosfatases cdc25/metabolismo , Animais , Apoptose , Sistemas CRISPR-Cas , Carcinogênese/genética , Ciclo Celular , Pontos de Checagem do Ciclo Celular , Proliferação de Células , Sobrevivência Celular , Transformação Celular Neoplásica , Técnicas de Silenciamento de Genes , Células HEK293 , Células HeLa , Xenoenxertos , Humanos , Masculino , Camundongos , Camundongos Nus , Camundongos SCID , Oncogenes , Ubiquitina/metabolismo , Proteases Específicas de Ubiquitina/genética , Ubiquitinação , Fosfatases cdc25/genética
11.
Int J Mol Sci ; 22(11)2021 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-34072967

RESUMO

Direct cellular interactions of MDA-MB-231cherry breast cancer cells with GFP-transduced human mesenchymal stroma/stem-like cells (MSCGFP) in a co-culture model resulted in spontaneous cell fusion by the generation of MDA-MSC-hyb5cherry GFP breast cancer hybrid cells. The proliferative capacity of MDA-MSC-hyb5 cells was enhanced about 1.8-fold when compared to the parental MDA-MB-231cherry breast cancer cells. In contrast to a spontaneous MDA-MB-231cherry induced tumor development in vivo within 18.8 days, the MDA-MSC-hyb5 cells initially remained quiescent in a dormancy-like state. At distinct time points after injection, NODscid mice started to develop MDA-MSC-hyb5 cell-induced tumors up to about a half year later. Following tumor initiation, however, tumor growth and formation of metastases in various different organs occurred rapidly within about 10.5 days. Changes in gene expression levels were evaluated by RNA-microarray analysis and revealed certain increase in dormancy-associated transcripts in MDA-MSC-hyb5. Chemotherapeutic responsiveness of MDA-MSC-hyb5 cells was partially enhanced when compared to MDA-MB-231 cells. However, some resistance, e.g., for taxol was detectable in cancer hybrid cells. Moreover, drug response partially changed during the tumor development of MDA-MSC-hyb5 cells; this suggests the presence of unstable in vivo phenotypes of MDA-hyb5 cells with increased tumor heterogeneity.


Assuntos
Neoplasias da Mama/patologia , Carcinogênese , Transformação Celular Neoplásica , Células-Tronco Mesenquimais , Microambiente Tumoral , Animais , Comunicação Celular , Divisão Celular , Fusão Celular , Linhagem Celular , Técnicas de Cocultura , Feminino , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID
12.
Aging (Albany NY) ; 13(11): 14999-15012, 2021 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-34081623

RESUMO

The ubiquitin-specific protease 8 (USP8) is a prototypic multidomain deubiquitinating enzyme with pleiotropic functions. We investigated the role of USP8 in hepatocellular carcinoma (HCC) by analyzing expression patterns of USP8 in HCC patients, and evaluating its functions and underlying signaling. Among 20 HCC patients investigated, we found that USP8 protein upregulation was a common phenomenon (17 out of 20) in HCC compared to normal liver tissue. Furthermore, the upregulation of USP8 was not associated with any clinicopathology. USP8 inhibition via genetic and pharmacological approaches resulted in growth inhibition and apoptosis induction in both sensitive and doxorubicin-resistant HCC cells. Of note, USP8 inhibition significantly enhanced doxorubicin or sorafenib's efficacy in HCC cells and mouse models. We further found that USP8 inhibition decreased levels of multiple receptor tyrosine kinases (RTKs) by ~90%, such as epidermal growth factor receptor (EGFR) and c-Met. Consistently, the downstream signaling regulated by RTKs was disrupted in HCC cells after USP8 inhibition, as shown by the decreased p-Akt, p-STAT3 and p-Raf. Our findings demonstrate that USP8 is a novel therapeutic target in HCC. Inhibiting USP8 has potential to overcome current drug resistance, particularly on HCC patients with high USP8 expression.


Assuntos
Carcinoma Hepatocelular/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos , Complexos Endossomais de Distribuição Requeridos para Transporte/antagonistas & inibidores , Neoplasias Hepáticas/tratamento farmacológico , Receptores Proteína Tirosina Quinases/metabolismo , Ubiquitina Tiolesterase/antagonistas & inibidores , Animais , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Regulação para Baixo/genética , Endopeptidases/genética , Endopeptidases/metabolismo , Complexos Endossomais de Distribuição Requeridos para Transporte/genética , Complexos Endossomais de Distribuição Requeridos para Transporte/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Masculino , Camundongos SCID , Transdução de Sinais , Ubiquitina Tiolesterase/genética , Ubiquitina Tiolesterase/metabolismo , Regulação para Cima/genética
13.
Nat Commun ; 12(1): 3834, 2021 06 22.
Artigo em Inglês | MEDLINE | ID: mdl-34158478

RESUMO

H-1 parvovirus (H-1PV) is a promising anticancer therapy. However, in-depth understanding of its life cycle, including the host cell factors needed for infectivity and oncolysis, is lacking. This understanding may guide the rational design of combination strategies, aid development of more effective viruses, and help identify biomarkers of susceptibility to H-1PV treatment. To identify the host cell factors involved, we carry out siRNA library screening using a druggable genome library. We identify one crucial modulator of H-1PV infection: laminin γ1 (LAMC1). Using loss- and gain-of-function studies, competition experiments, and ELISA, we validate LAMC1 and laminin family members as being essential to H-1PV cell attachment and entry. H-1PV binding to laminins is dependent on their sialic acid moieties and is inhibited by heparin. We show that laminins are differentially expressed in various tumour entities, including glioblastoma. We confirm the expression pattern of laminin γ1 in glioblastoma biopsies by immunohistochemistry. We also provide evidence of a direct correlation between LAMC1 expression levels and H-1PV oncolytic activity in 59 cancer cell lines and in 3D organotypic spheroid cultures with different sensitivities to H-1PV infection. These results support the idea that tumours with elevated levels of γ1 containing laminins are more susceptible to H-1PV-based therapies.


Assuntos
Parvovirus H-1/metabolismo , Laminina/metabolismo , Ácido N-Acetilneuramínico/metabolismo , Vírus Oncolíticos/metabolismo , Ligação Viral , Internalização do Vírus , Animais , Linhagem Celular Tumoral , Glioblastoma/patologia , Glioblastoma/terapia , Glioblastoma/virologia , Células HCT116 , Células HEK293 , Células HeLa , Humanos , Laminina/genética , Camundongos Endogâmicos NOD , Camundongos SCID , Terapia Viral Oncolítica/métodos , Ligação Proteica , Interferência de RNA , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
14.
Molecules ; 26(11)2021 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-34071530

RESUMO

Anti-cancer activity of catechin nanoemulsions prepared from Oolong tea leaf waste was studied on prostate cancer cells DU-145 and DU-145-induced tumors in mice. Catechin nanoemulsions composed of lecithin, Tween-80 and water in an appropriate proportion was prepared with high stability, particle size of 11.3 nm, zeta potential of -67.2 mV and encapsulation efficiency of 83.4%. Catechin nanoemulsions were more effective than extracts in inhibiting DU-145 cell growth, with the IC50 being 13.52 and 214.6 µg/mL, respectively, after 48 h incubation. Furthermore, both catechin nanoemulsions and extracts could raise caspase-8, caspase-9 and caspase-3 activities for DU-145 cell apoptosis, arresting the cell cycle at S and G2/M phases. Compared to control, catechin nanoemulsion at 20 µg/mL and paclitaxel at 10 µg/mL were the most effective in reducing tumor volume by 41.3% and 52.5% and tumor weight by 77.5% and 90.6% in mice, respectively, through a decrease in EGF and VEGF levels in serum.


Assuntos
Catequina/química , Emulsões/química , Nanopartículas/química , Folhas de Planta/metabolismo , Neoplasias da Próstata/tratamento farmacológico , Chá/química , Animais , Antineoplásicos/farmacologia , Apoptose , Caspase 8/metabolismo , Ciclo Celular , Linhagem Celular Tumoral , Endocitose , Humanos , Concentração Inibidora 50 , Lecitinas/química , Limite de Detecção , Masculino , Camundongos , Camundongos SCID , Nanotecnologia/métodos , Transplante de Neoplasias , Neoplasias Experimentais/tratamento farmacológico , Tamanho da Partícula , Polissorbatos/química , Controle de Qualidade , Solventes , Água/química
15.
Nat Commun ; 12(1): 3974, 2021 06 25.
Artigo em Inglês | MEDLINE | ID: mdl-34172737

RESUMO

Cancer stem cells (CSCs) play a critical role in invasive growth and metastasis of human head and neck squamous cell carcinoma (HNSCC). Although significant progress has been made in understanding the self-renewal and pro-tumorigenic potentials of CSCs, a key challenge remains on how to eliminate CSCs and halt metastasis effectively. Here we show that super-enhancers (SEs) play a critical role in the transcription of cancer stemness genes as well as pro-metastatic genes, thereby controlling their tumorigenic potential and metastasis. Mechanistically, we find that bromodomain-containing protein 4 (BRD4) recruits Mediators and NF-κB p65 to form SEs at cancer stemness genes such as TP63, MET and FOSL1, in addition to oncogenic transcripts. In vivo lineage tracing reveals that disrupting SEs by BET inhibitors potently inhibited CSC self-renewal and eliminated CSCs in addition to elimination of proliferating non-stem tumor cells in a mouse model of HNSCC. Moreover, disrupting SEs also inhibits the invasive growth and lymph node metastasis of human CSCs isolated from human HNSCC. Taken together, our results suggest that targeting SEs may serve as an effective therapy for HNSCC by eliminating CSCs.


Assuntos
Elementos Facilitadores Genéticos , Neoplasias de Cabeça e Pescoço/patologia , Células-Tronco Neoplásicas/efeitos dos fármacos , Carcinoma de Células Escamosas de Cabeça e Pescoço/patologia , Animais , Antineoplásicos/farmacologia , Azepinas/farmacologia , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Neoplasias de Cabeça e Pescoço/genética , Humanos , Metástase Linfática/tratamento farmacológico , Metástase Linfática/prevenção & controle , Subunidade 1 do Complexo Mediador/genética , Subunidade 1 do Complexo Mediador/metabolismo , Camundongos Endogâmicos C57BL , Camundongos SCID , NF-kappa B/genética , Células-Tronco Neoplásicas/patologia , Complexo Repressor Polycomb 1/antagonistas & inibidores , Complexo Repressor Polycomb 1/genética , Complexo Repressor Polycomb 1/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço/tratamento farmacológico , Carcinoma de Células Escamosas de Cabeça e Pescoço/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Triazóis/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
16.
Nat Commun ; 12(1): 3790, 2021 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-34145240

RESUMO

The receptor tyrosine kinase HER2 acts as oncogenic driver in numerous cancers. Usually, the gene is amplified, resulting in receptor overexpression, massively increased signaling and unchecked proliferation. However, tumors become frequently addicted to oncogenes and hence are druggable by targeted interventions. Here, we design an anti-HER2 biparatopic and tetravalent IgG fusion with a multimodal mechanism of action. The molecule first induces HER2 clustering into inactive complexes, evidenced by reduced mobility of surface HER2. However, in contrast to our earlier binders based on DARPins, clusters of HER2 are thereafter robustly internalized and quantitatively degraded. This multimodal mechanism of action is found only in few of the tetravalent constructs investigated, which must target specific epitopes on HER2 in a defined geometric arrangement. The inhibitory effect of our antibody as single agent surpasses the combination of trastuzumab and pertuzumab as well as its parental mAbs in vitro and it is effective in a xenograft model.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Antineoplásicos Imunológicos/uso terapêutico , Neoplasias da Mama/terapia , Receptor ErbB-2/antagonistas & inibidores , Receptor ErbB-2/imunologia , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais Humanizados/uso terapêutico , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Desenho de Fármacos , Feminino , Células HeLa , Humanos , Imunoglobulina G/imunologia , Imunoterapia/métodos , Células MCF-7 , Camundongos , Camundongos SCID , Trastuzumab/uso terapêutico , Ensaios Antitumorais Modelo de Xenoenxerto
17.
Nat Commun ; 12(1): 3330, 2021 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-34099664

RESUMO

Human pluripotent stem cell (hPSC)-derived pancreatic ß cells are an attractive cell source for treating diabetes. However, current derivation methods remain inefficient, heterogeneous, and cell line dependent. To address these issues, we first devised a strategy to efficiently cluster hPSC-derived pancreatic progenitors into 3D structures. Through a systematic study, we discovered 10 chemicals that not only retain the pancreatic progenitors in 3D clusters but also enhance their potentiality towards NKX6.1+/INS+ ß cells. We further systematically screened signaling pathway modulators in the three steps from pancreatic progenitors toward ß cells. The implementation of all these strategies and chemical combinations resulted in generating ß cells from different sources of hPSCs with high efficiency. The derived ß cells are functional and can reverse hyperglycemia in mice within two weeks. Our protocol provides a robust platform for studying human ß cells and developing hPSC-derived ß cells for cell replacement therapy.


Assuntos
Proteínas de Homeodomínio/metabolismo , Células Secretoras de Insulina/metabolismo , Pâncreas/metabolismo , Células-Tronco Pluripotentes/metabolismo , Animais , Diferenciação Celular/fisiologia , Linhagem Celular , Terapia Baseada em Transplante de Células e Tecidos , Diabetes Mellitus/metabolismo , Diabetes Mellitus Experimental , Proteínas de Homeodomínio/genética , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Transdução de Sinais
18.
Nat Commun ; 12(1): 3299, 2021 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-34083537

RESUMO

Bioenergetic perturbations driving neoplastic growth increase the production of reactive oxygen species (ROS), requiring a compensatory increase in ROS scavengers to limit oxidative stress. Intervention strategies that simultaneously induce energetic and oxidative stress therefore have therapeutic potential. Phenformin is a mitochondrial complex I inhibitor that induces bioenergetic stress. We now demonstrate that inflammatory mediators, including IFNγ and polyIC, potentiate the cytotoxicity of phenformin by inducing a parallel increase in oxidative stress through STAT1-dependent mechanisms. Indeed, STAT1 signaling downregulates NQO1, a key ROS scavenger, in many breast cancer models. Moreover, genetic ablation or pharmacological inhibition of NQO1 using ß-lapachone (an NQO1 bioactivatable drug) increases oxidative stress to selectively sensitize breast cancer models, including patient derived xenografts of HER2+ and triple negative disease, to the tumoricidal effects of phenformin. We provide evidence that therapies targeting ROS scavengers increase the anti-neoplastic efficacy of mitochondrial complex I inhibitors in breast cancer.


Assuntos
Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Fenformin/farmacologia , Fator de Transcrição STAT1/metabolismo , Animais , Antineoplásicos/administração & dosagem , Linhagem Celular Tumoral , Sinergismo Farmacológico , Complexo I de Transporte de Elétrons/antagonistas & inibidores , Metabolismo Energético/efeitos dos fármacos , Feminino , Glutationa/antagonistas & inibidores , Glutationa/biossíntese , Humanos , Interferon gama/administração & dosagem , Interferon gama/deficiência , Interferon gama/metabolismo , Células MCF-7 , Neoplasias Mamárias Experimentais/tratamento farmacológico , Neoplasias Mamárias Experimentais/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Camundongos SCID , NAD(P)H Desidrogenase (Quinona)/antagonistas & inibidores , NAD(P)H Desidrogenase (Quinona)/metabolismo , Naftoquinonas/administração & dosagem , Estresse Oxidativo/efeitos dos fármacos , Fenformin/administração & dosagem , Poli I-C/administração & dosagem , Espécies Reativas de Oxigênio/metabolismo , Fator de Transcrição STAT1/agonistas , Ensaios Antitumorais Modelo de Xenoenxerto
19.
Int J Mol Sci ; 22(11)2021 May 21.
Artigo em Inglês | MEDLINE | ID: mdl-34064291

RESUMO

The Arg-Gly-Asp (RGD) peptide shows a high affinity for αvß3 integrin, which is overexpressed in new tumor blood vessels and many types of tumor cells. The radiolabeled RGD peptide has been studied for cancer imaging and radionuclide therapy. We have developed a long-term tumor-targeting peptide DOTA-EB-cRGDfK, which combines a DOTA chelator, a truncated Evans blue dye (EB), a modified linker, and cRGDfK peptide. The aim of this study was to evaluate the potential of indium-111(111In) radiolabeled DOTA-EB-cRGDfK in αvß3 integrin-expressing tumors. The human glioblastoma cell line U-87 MG was used to determine the in vitro binding affinity of the radiolabeled peptide. The in vivo distribution of radiolabeled peptides in U-87 MG xenografts was investigated by biodistribution, nanoSPECT/CT, pharmacokinetic and excretion studies. The in vitro competition assay showed that 111In-DOTA-EB-cRGDfK had a significant binding affinity to U-87 MG cancer cells (IC50 = 71.7 nM). NanoSPECT/CT imaging showed 111In-DOTA-EB-cRGDfK has higher tumor uptake than control peptides (111In-DOTA-cRGDfK and 111In-DOTA-EB), and there is still a clear signal until 72 h after injection. The biodistribution results showed significant tumor accumulation (27.1 ± 2.7% ID/g) and the tumor to non-tumor ratio was 22.85 at 24 h after injection. In addition, the pharmacokinetics results indicated that the 111In-DOTA-EB-cRGDfK peptide has a long-term half-life (T1/2λz = 77.3 h) and that the calculated absorbed dose was safe for humans. We demonstrated that radiolabeled DOTA-EB-cRGDfK may be a promising agent for glioblastoma tumor imaging and has the potential as a theranostic radiopharmaceutical.


Assuntos
Quelantes/metabolismo , Glioblastoma/metabolismo , Oligopeptídeos/metabolismo , Animais , Linhagem Celular Tumoral , Compostos Heterocíclicos com 1 Anel/metabolismo , Xenoenxertos/metabolismo , Humanos , Radioisótopos de Índio/metabolismo , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Imagem Molecular/métodos , Peptídeos Cíclicos/metabolismo , Compostos Radiofarmacêuticos/metabolismo , Ratos , Distribuição Tecidual
20.
In Vivo ; 35(4): 2151-2157, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34182491

RESUMO

BACKGROUND/AIM: Osteosarcoma is the most common type of bone cancer, but current therapeutic interventions remain largely insufficient. The development of new treatment strategies is needed, and moreover, optimal rodent models are necessary for testing the efficacy of new treatment modalities of osteosarcoma. Humanized mice carry human hematopoietic and immune systems, and are considered an ideal tool to study human diseases including cancer immunology. Herein, we performed a preliminary study toward developing an in vivo bioluminescent osteosarcoma model using humanized immunodeficient (NSG) mice. MATERIALS AND METHODS: To establish the xenograft and orthotopic mouse model, NSG mice engrafted with human CD34+ hematopoietic stem cells were injected with luciferase-expressing KHOS/NP cells at two different time points. Bioluminescence images were obtained to monitor in vivo tumor growth and metastasis. Influence of the degree of human cell engraftment on tumor growth and metastatic behavior was analyzed and compared between the two groups. RESULTS: KHOS/NP-luc cells injected in humanized NSG mice formed macroscopic tumors. The percentage of human CD45+ cells in these models was similar, but the percentage of human CD45+CD3+ and their subset was higher in the late-injection group compared to that of the early-injection group. The rate of KHOS/NP tumor growth was higher in the early-injection group than in the late-injection group. In the present study, human hematopoietic cell engraftment was not influenced by KHOS/NP cell injection, but KHOS/NP osteosarcoma showed more aggressive behavior in the early-injection group than that in the late-injection group, forming larger tumor volumes and earlier metastases. CONCLUSION: The results indicated that tumor growth and progression in humanized NSG mice may have been influenced by higher levels of human cell engraftment, especially T cells. Although there exist some limitations to our study, our preliminary results can provide the basis for the development of a humanized osteosarcoma mouse model.


Assuntos
Neoplasias Ósseas , Osteossarcoma , Animais , Modelos Animais de Doenças , Células-Tronco Hematopoéticas , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Projetos Piloto
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