Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 8.110
Filtrar
1.
Methods Mol Biol ; 2265: 621-634, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33704743

RESUMO

RNA interference (RNAi) is a posttranscriptional regulatory mechanism that employs siRNA. It typically results in the degradation of a target mRNA that encodes a particular protein. Treatment with siRNA therapeutics requires the use of an effective drug delivery system to assist in delivering these therapeutics into the cytoplasm of the transfected cells. Here we describe the transfection of melanoma cancer cells with siRNA using cationic niosome nanoparticles as a delivery system. The method of niosome preparation is first introduced and is followed by complex formation with siRNA and the transfection method.


Assuntos
Melanoma , Nanopartículas , RNA Interferente Pequeno , Transfecção , Humanos , Lipossomos , Melanoma/genética , Melanoma/metabolismo , Melanoma/patologia , Melanoma/terapia , Nanopartículas/química , Nanopartículas/uso terapêutico , RNA Interferente Pequeno/química , RNA Interferente Pequeno/farmacologia
2.
Int J Nanomedicine ; 16: 1961-1976, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33727809

RESUMO

Introduction: Metastatic breast cancer seriously harms women's health and is currently the tumour type with the highest mortality rate in women. Recently, the combinatorial therapeutic approaches that integrate anti-cancer drugs and genetic agents is an attractive and promising strategy for the treatment of metastatic breast cancer. Moreover, such a combination strategy requires better drug carriers that can effectively deliver the cargo to the breast cancer cells and achieve controlled release in the cells to achieve better therapeutic effects. Methods: The tumour-targeted and redox-responsive mesoporous silica nanoparticles (MSNs) functionalised with DNA aptamers (AS1411) as a co-delivery system was developed and investigated for the potential against metastatic breast cancer. Doxorubicin (Dox) was loaded onto the MSNs, while AS1411 and a small interfering RNA (siTIE2) were employed as gatekeepers via attachment to the MSNs with redox-sensitive disulfide bonds. Results: The controlled release of Dox and siTIE2 was associated with intracellular glutathione. AS1411 mediated the targeted delivery of Dox by increasing its cellular uptake in metastatic breast cancer, ultimately resulting in a lower IC50 in MDA-MB-231 cells (human breast cancer cell line with high metastatic potency), improved biodistribution in tumour-bearing mice, and enhanced in vivo anti-tumour effects. The in vitro cell migration/invasion assay and in vivo anti-metastatic study revealed synergism in the co-delivery system that suppresses cancer cell metastasis. Conclusion: The tumour-targeted and redox-responsive MSN prepared in this study are promising for the effective delivery and controlled release of Dox and siTIE2 for improved treatment of metastatic breast cancer.


Assuntos
Neoplasias da Mama/tratamento farmacológico , Doxorrubicina/uso terapêutico , Nanopartículas/química , RNA Interferente Pequeno/administração & dosagem , Dióxido de Silício/química , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Neoplasias da Mama/patologia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Preparações de Ação Retardada/farmacologia , Preparações de Ação Retardada/uso terapêutico , Doxorrubicina/farmacologia , Portadores de Fármacos/química , Endocitose/efeitos dos fármacos , Feminino , Células HEK293 , Humanos , Camundongos Endogâmicos BALB C , Camundongos Nus , Nanopartículas/ultraestrutura , Invasividade Neoplásica , Metástase Neoplásica , Oxirredução , Porosidade , RNA Interferente Pequeno/farmacologia , Distribuição Tecidual/efeitos dos fármacos
3.
ACS Appl Mater Interfaces ; 13(5): 6109-6118, 2021 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-33497198

RESUMO

siRNA is found to effectively knock down the target gene in cells, which is considered a promising strategy for gene therapy. However, the application of siRNA is limited due to its low efficiency of the cellular uptake. Tetrahedral framework nucleic acids (tFNAs) are synthesized by four single-stranded DNAs and show multiple biological functions in recent studies, especially suitable for drug delivery. More than 60% of malignant melanomas are associated with Braf gene mutation, an attractive therapeutic target for RNA interference. In this study, we modified anti-Braf siRNA (siBraf) with tFNAs to downregulate the target gene. Meanwhile, we directly incorporated AS1411 (a DNA aptamer) to our nanostructure, which assists tFNAs to improve the cellular uptake efficacy of siBraf significantly. The results indicated that tFNAs-AS1411-siBraf exhibited more potent activity to cleave Braf mRNA than free siBraf. This study may provide a new idea for the combination therapy of siRNA and aptamers via DNA nanomaterials to achieve gene silencing.


Assuntos
Aptâmeros de Nucleotídeos/química , DNA/química , Sistemas de Liberação de Medicamentos , Melanoma/tratamento farmacológico , RNA Interferente Pequeno/farmacologia , DNA/síntese química , Inativação Gênica/efeitos dos fármacos , Humanos , Melanoma/genética , Melanoma/metabolismo , Tamanho da Partícula , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas B-raf/metabolismo , RNA Interferente Pequeno/administração & dosagem , RNA Interferente Pequeno/química , Propriedades de Superfície , Células Tumorais Cultivadas
4.
ACS Appl Mater Interfaces ; 13(5): 5999-6010, 2021 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-33506682

RESUMO

Cellular FLIP (cFLIP) is a crucial player of apoptosis-regulated pathways that is frequently overexpressed in solid cancers. To inhibit c-FLIP, pre- and post-transcriptionally, a multifunctional nanoparticle (NP) was created to deliver cFLIP-specific small interfering RNA (siRNA) into cancer cells. Specifically, Vorinostat (Vor)-loaded mesoporous silica nanoparticles (MSN) were conjugated with polyethylenimine-biotin (PB), followed by electrostatically binding with cFLIP siRNA (Vor/siR@MSN-PB). To stabilize and prolong the circulation time of nanoparticles, a bialdehyde-modified poly(ethylene glycol) (PEG) was cross-linked onto the polyethylenimine (PEI) backbone via the formation of the imine linkage (Schiff base) (Vor/siR@MSN-PB-PEG). The Schiff base is highly stable at physiological pH 7.4 but labile under slightly acidic pH conditions. In the acidic tumor microenvironment (TME), the PEG outer layer could be rapidly cleaved, resulting in the switching of the nanoparticle surface charge to positive, which specifically enhances internalization of the NPs to the biotin-positive tumor cells. Our results demonstrated the successful preparation of Vor/siR@MSN-PB-PEG NPs, in which the siRNA was effectively protected in serum and regulated the expression of cFlip, post-transcriptionally. The presence of the PEG layer resulted in high tumor accumulation and high efficacy in tumor inhibition, which was a result of the efficient cFLIP suppression. Furthermore, in the low-dose regimen of Vorinostat-the pre-transcriptional cFLIP suppressor, treatment with Vor/siR@MSN-PB-PEG NPs was found to be safe with the treated mice, indicating a promising combination regimen for cancer therapy.


Assuntos
Antineoplásicos/farmacologia , Proteína Reguladora de Apoptosis Semelhante a CASP8 e FADD/antagonistas & inibidores , Proteína Reguladora de Apoptosis Semelhante a CASP8 e FADD/genética , Nanopartículas/química , RNA Interferente Pequeno/farmacologia , Vorinostat/farmacologia , Animais , Antineoplásicos/química , Proteína Reguladora de Apoptosis Semelhante a CASP8 e FADD/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Humanos , Concentração de Íons de Hidrogênio , Camundongos , Camundongos Endogâmicos BALB C , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Tamanho da Partícula , Polietilenoglicóis/química , RNA Interferente Pequeno/química , Propriedades de Superfície , Vorinostat/química
5.
Arch Biochem Biophys ; 699: 108754, 2021 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-33450239

RESUMO

Drug resistance is one of the major challenges for treatment of hepatocellular carcinoma (HCC) with sorafenib. Our present study found that sorafenib resistant (SR) HCC cells showed epithelial-mesenchymal transition (EMT) characteristics with the downregulation of epithelial marker and upregulation of mesenchymal makers. The expression of Snail, a core factor of EMT, was increased in HCC/SR cells, while knockdown of Snail can restore sorafenib sensitivity and EMT potential of HCC/SR cells. Further, the upregulation of protein stability was responsible for the upregulation of Snail in HCC/SR cells. ATM and CSN2, which can stabilize Snail protein, were increased in HCC/SR cells. Knockdown of ATM and CSN2 can suppress the expression of Snail and increase sorafenib sensitivity of HCC/SR cells. It indicated that targeted inhibition of Snail might be helpful to overcome sorafenib resistance of HCC patients.


Assuntos
Carcinoma Hepatocelular/fisiopatologia , Resistencia a Medicamentos Antineoplásicos/fisiologia , Neoplasias Hepáticas/fisiopatologia , Fatores de Transcrição da Família Snail/metabolismo , Sorafenibe/farmacologia , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Complexo do Signalossomo COP9/genética , Complexo do Signalossomo COP9/metabolismo , Linhagem Celular Tumoral , Transição Epitelial-Mesenquimal/fisiologia , Inativação Gênica , Humanos , Estabilidade Proteica , RNA Interferente Pequeno/farmacologia , Fatores de Transcrição da Família Snail/química , Fatores de Transcrição da Família Snail/genética , Regulação para Cima/efeitos dos fármacos
6.
Anticancer Res ; 41(2): 783-794, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33517283

RESUMO

BACKGROUND/AIM: The inflammatory cytokine IL-8 and its receptor CXCR2 are key signalling pathway molecules in cancer development. We hypothesized that IL-8/CXCR2 signalling promotes tumour progression in oesophageal squamous cell carcinoma (ESCC) patients. MATERIALS AND METHODS: We examined the relationship between IL-8/CXCR2 expression and clinicopathological factors by immunohistochemistry in samples from 63 patients with resectable ESCC. The effects of IL-8/CXCR2 signalling on cell proliferation and gene expression were examined in vitro and in vivo using ESCC cell lines. RESULTS: Increased IL-8/CXCR2 signalling was associated with shorter overall survival (p<0.05) and recurrence-free survival (p<0.05) in ESCC patients. Multivariate analysis identified IL-8/CXCR2 expression as a prognostic factor for surgically treated ESCC (p<0.05). In vitro, IL-8 exposure or over-expression significantly enhanced ESCC cell proliferation. SB225002, a CXCR2-specific antagonist, and IL-8 siRNA significantly suppressed cell proliferation. CONCLUSION: IL-8/CXCR2 expression is an independent prognostic factor for surgically treated ESCC, and IL-8/CXCR2 signalling contributes to ESCC cell proliferation.


Assuntos
Neoplasias Esofágicas/cirurgia , Carcinoma de Células Escamosas do Esôfago/cirurgia , Interleucina-8/metabolismo , Receptores de Interleucina-8B/metabolismo , Regulação para Cima , Idoso , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Neoplasias Esofágicas/metabolismo , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago/metabolismo , Carcinoma de Células Escamosas do Esôfago/patologia , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Interleucina-8/antagonistas & inibidores , Interleucina-8/genética , Masculino , Camundongos , Pessoa de Meia-Idade , Transplante de Neoplasias , Compostos de Fenilureia/farmacologia , Prognóstico , RNA Interferente Pequeno/farmacologia , Receptores de Interleucina-8B/antagonistas & inibidores , Receptores de Interleucina-8B/genética , Transdução de Sinais/efeitos dos fármacos , Análise de Sobrevida
7.
Nucleic Acids Res ; 49(2): 928-953, 2021 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-33406258

RESUMO

Double-strand breaks and stalled replication forks are a significant threat to genomic stability that can lead to chromosomal rearrangements or cell death. The protein CtIP promotes DNA end resection, an early step in homologous recombination repair, and has been found to protect perturbed forks from excessive nucleolytic degradation. However, it remains unknown how CtIP's function in fork protection is regulated. Here, we show that CtIP recruitment to sites of DNA damage and replication stress is impaired upon global inhibition of SUMOylation. We demonstrate that CtIP is a target for modification by SUMO-2 and that this occurs constitutively during S phase. The modification is dependent on the activities of cyclin-dependent kinases and the PI-3-kinase-related kinase ATR on CtIP's carboxyl-terminal region, an interaction with the replication factor PCNA, and the E3 SUMO ligase PIAS4. We also identify residue K578 as a key residue that contributes to CtIP SUMOylation. Functionally, a CtIP mutant where K578 is substituted with a non-SUMOylatable arginine residue is defective in promoting DNA end resection, homologous recombination, and in protecting stalled replication forks from excessive nucleolytic degradation. Our results shed further light on the tightly coordinated regulation of CtIP by SUMOylation in the maintenance of genome stability.


Assuntos
Reparo do DNA por Junção de Extremidades/fisiologia , Replicação do DNA , Endodesoxirribonucleases/fisiologia , Processamento de Proteína Pós-Traducional , Sumoilação , Substituição de Aminoácidos , Arginina/química , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Linhagem Celular , Quinases Ciclina-Dependentes/metabolismo , Quebras de DNA de Cadeia Dupla , Reparo do DNA por Junção de Extremidades/genética , Endodesoxirribonucleases/química , Endodesoxirribonucleases/metabolismo , Genes Reporter , Instabilidade Genômica , Humanos , Lisina/química , Proteínas de Ligação a Poli-ADP-Ribose/fisiologia , Antígeno Nuclear de Célula em Proliferação/metabolismo , Proteínas Inibidoras de STAT Ativados/fisiologia , Mapeamento de Interação de Proteínas , Interferência de RNA , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , Proteínas Recombinantes de Fusão/metabolismo , Reparo de DNA por Recombinação/genética , Reparo de DNA por Recombinação/fisiologia
8.
Nucleic Acids Res ; 49(2): 805-817, 2021 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-33410907

RESUMO

Pin1 is a peptidyl-prolyl isomerase that regulates the structure and function of eukaryotic RNA polymerase II (Pol II) through interaction with the C-terminal domain (CTD) of Rpb1, the largest subunit of Pol II. We demonstrated that this function is important for cellular response to oxidative stress in the fission yeast Schizosaccharomyces pombe. In response to oxidative stress, the Atf1 transcription factor targets Sty1, the mitogen-activated protein kinase (MAPK), to specific stress-responsive promoters. Anchored Sty1 recruits Pol II through direct association with Rpb1-CTD and phosphorylates the reiterated heptad sequence at Serine 5. Pin1 binds phosphorylated CTD to promote dissociation of Sty1 from it, and directly recruits Ssu72 phosphatase to facilitate dephosphorylation of CTD for transcription elongation. In the absence of Pin1, the association of Sty1-Atf1 with Rpb1 persists on stress-responsive promoters failed to generate transcripts of the corresponding genes effectively. The identified characteristic features of the fission yeast Pin1 are conserved in humans. We demonstrated that elevated Pin1 level in cancer cells might help to sustain survival under oxidative stress generated from their altered metabolic pathways. Together, these results suggest a conserved function of Pin1 in cellular response to oxidative stress among eukaryotic cells that might have clinical implication.


Assuntos
Regulação Fúngica da Expressão Gênica , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Peptidilprolil Isomerase de Interação com NIMA/fisiologia , Estresse Oxidativo/genética , Fosfoproteínas Fosfatases/metabolismo , RNA Polimerase II/metabolismo , Proteínas de Schizosaccharomyces pombe/metabolismo , Schizosaccharomyces/metabolismo , Imunoprecipitação da Cromatina , Fosforilação , Ligação Proteica , Mapeamento de Interação de Proteínas , Processamento de Proteína Pós-Traducional , Interferência de RNA , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , Espécies Reativas de Oxigênio , Reação em Cadeia da Polimerase em Tempo Real , Proteínas Recombinantes/metabolismo , Schizosaccharomyces/genética , Transcrição Genética
9.
Carbohydr Polym ; 251: 117008, 2021 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-33142574

RESUMO

A novel folic acid mediated chitosan oligosaccharide-grafted disulfide-containing polyethylenimine copolymer-based silica nanohybrids were fabricated for co-delivering paclitaxel and P-shRNA. These nanoparticles could efficiently protect P-shRNA against degradation, and exhibited well redox-responsive P-shRNA release and pH-responsive drug release behaviors. Folic acid as the targeting head, could improve cellular uptake of nanoparticles by multidrug-resistant breast cancer cells. Moreover, these nanoparticles showed excellent delivery P-shRNA into cells and displayed high gene silencing efficiency at the targeted mRNAs to downregulate the expression of P-gp which induced up to 63% decrease. Finally, nanoparticles could completely reverse the resistance of breast cancer cells to paclitaxel and the resistance reversion index was 50.59. These results suggested that our nanoparticles could efficiently co-deliver paclitaxel and P-shRNA into cancer cells to exert its synergistic antitumor effect, and opened up a new avenue for overcoming multidrug resistance.


Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Portadores de Fármacos/química , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/farmacologia , Quitosana/análogos & derivados , Quitosana/química , Liberação Controlada de Fármacos , Ácido Fólico/química , Humanos , Concentração de Íons de Hidrogênio , Células MCF-7 , Nanopartículas/química , Oxirredução , Paclitaxel/farmacologia , Polietilenoimina/análogos & derivados , Polietilenoimina/química , RNA Interferente Pequeno/farmacologia
10.
Biochim Biophys Acta Mol Cell Res ; 1868(1): 118856, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-32931817

RESUMO

NIR, a novel INHAT, negatively regulates the transcription activity of tumor repressor p53. However, if NIR functions in the tumorigenesis dependent on the regulation of p53 remains unknown. Here, we report that NIR promotes progression of colorectal cancer (CRC) through regulating RB function. Firstly, we found that NIR expression is upregulated in the human CRC tissues and significantly associated with the poor outcome of the patients. Sequence alignment shows that NIR contains an RB-binding motif LxCxE in its INHAT-2 domain. We demonstrate that NIR interacts with RB via INHAT-2 in CRC cells and promotes RB degradation through proteasome-mediated pathway. Further, either full-length GFP-NIR or GFP-NIR-INHAT2 facilitates poly-ubiquitination of RB. In addition, NIR inhibits RB acetylation by INHAT-2, suggesting NIR might promote RB degradation through inhibiting RB acetylation. Importantly, endogenous NIR is downregulated upon DNA damage, which is consistent with the upregulation of total level and acetylation of RB. We further show that Flag-NIR inhibits DNA damage-induced RB acetylation. Thus, downregulation of NIR might contribute to maintain the cellular homeostasis under DNA damage. Consequently, depletion of NIR inhibits cell proliferation and tumor growth in mouse xenografts. Taken together, we demonstrate that NIR promotes CRC progression partially through inhibiting RB acetylation and promoting RB degradation. Targeting NIR may provide a potential therapeutic strategy for NIR-upregulated CRC patients.


Assuntos
Carcinogênese/genética , Neoplasias Colorretais/genética , Proteínas Repressoras/genética , Proteína do Retinoblastoma/genética , Proteína Supressora de Tumor p53/genética , Acetilação/efeitos dos fármacos , Idoso , Animais , Linhagem Celular Tumoral , Proliferação de Células/genética , Neoplasias Colorretais/patologia , Dano ao DNA/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Xenoenxertos , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Proteólise , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , Ubiquitinação
11.
Phytomedicine ; 80: 153369, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-33070082

RESUMO

BACKGROUND: Impairment of mitochondrial biogenesis is associated with the pathological progression of Parkinson's disease (PD). Parkin-interacting substrate (PARIS) can be ubiquitinated by parkin and prevents the repression of proliferator-activated receptor gamma coactivator-1-alpha (PGC-1α). PURPOSE: This study investigated whether the neuroprotective mechanism of carnosic acid (CA) from rosemary is mediated via the regulation of PARIS and PGC-1α by parkin. METHODS: The Western blotting and RT-PCR were used to determine protein and mRNA, respectively. To investigate the protein-protein interaction of between PARIS and ubiquitin, the immunoprecipitation assay (IP assay) was utilized. Silencing of endogenous parkin or PGC-1α was performed by using transient transfection of small interfering RNA (siRNA). RESULTS: SH-SY5Y cells treated with 6-hydroxydopamine (6-OHDA) increased PARIS protein, decreased PGC-1α protein, and reduced protein and mRNA of mitochondrial biogenesis-related genes. CA pretreatment reversed the effects of 6-OHDA. By IP assay, the interaction of PARIS with ubiquitin protein caused by CA was stronger than that caused by 6-OHDA. Moreover, knockdown of parkin attenuated the ability of CA to reverse the 6-OHDA-induced increase in PARIS and decrease in PGC-1α expression. PGC-1α siRNA was used to investigate how CA influenced the effect of 6-OHDA on the modulation of mitochondrial biogenesis and apoptosis. In the presence of PGC-1α siRNA, CA could no longer significantly reverse the reduction of mitochondrial biogenesis or the induction of cleavage of apoptotic-related proteins by 6-OHDA. CONCLUSION: The cytoprotective of CA is related to the enhancement of mitochondrial biogenesis by inhibiting PARIS and inducing PGC-1α by parkin. The activation of PGC-1α-mediated mitochondrial biogenesis by CA prevents the degeneration of dopaminergic neurons, CA may have therapeutic application in PD.


Assuntos
Abietanos/farmacologia , Mitocôndrias/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Proteínas Repressoras/metabolismo , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/metabolismo , Neurônios Dopaminérgicos/efeitos dos fármacos , Neurônios Dopaminérgicos/metabolismo , Humanos , Mitocôndrias/metabolismo , Proteínas Mitocondriais/metabolismo , Biogênese de Organelas , Oxidopamina/toxicidade , Doença de Parkinson/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , RNA Interferente Pequeno/farmacologia , Fatores de Transcrição/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo
12.
Genomics ; 113(1 Pt 1): 331-343, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33321203

RESUMO

An outbreak, caused by an RNA virus, SARS-CoV-2 named COVID-19 has become pandemic with a magnitude which is daunting to all public health institutions in the absence of specific antiviral treatment. Surface glycoprotein and nucleocapsid phosphoprotein are two important proteins of this virus facilitating its entry into host cell and genome replication. Small interfering RNA (siRNA) is a prospective tool of the RNA interference (RNAi) pathway for the control of human viral infections by suppressing viral gene expression through hybridization and neutralization of target complementary mRNA. So, in this study, the power of RNA interference technology was harnessed to develop siRNA molecules against specific target genes namely, nucleocapsid phosphoprotein gene and surface glycoprotein gene. Conserved sequence from 139 SARS-CoV-2 strains from around the globe was collected to construct 78 siRNA that can inactivate nucleocapsid phosphoprotein and surface glycoprotein genes. Finally, based on GC content, free energy of folding, free energy of binding, melting temperature, efficacy prediction and molecular docking analysis, 8 siRNA molecules were selected which are proposed to exert the best action. These predicted siRNAs should effectively silence the genes of SARS-CoV-2 during siRNA mediated treatment assisting in the response against SARS-CoV-2.


Assuntos
/terapia , Química Computacional , Desenho de Fármacos , Terapia Genética/métodos , Simulação de Acoplamento Molecular , Interferência de RNA , RNA Mensageiro/antagonistas & inibidores , RNA Interferente Pequeno/química , RNA Viral/antagonistas & inibidores , Glicoproteína da Espícula de Coronavírus/genética , Proteínas Argonauta/química , Proteínas Argonauta/genética , Composição de Bases , /virologia , Evolução Molecular , Regulação Viral da Expressão Gênica/efeitos dos fármacos , Humanos , Pandemias , Fosfoproteínas/genética , Filogenia , Dobramento de RNA , RNA Guia/química , RNA Guia/genética , RNA Mensageiro/genética , RNA Interferente Pequeno/farmacologia , RNA Interferente Pequeno/uso terapêutico , RNA Viral/genética , Alinhamento de Sequência , Termodinâmica
13.
Gene ; 772: 145358, 2021 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-33340561

RESUMO

FAM135B (family with sequence similarity 135, member B) is related to the progression of esophageal squamous cell carcinoma (ESCC). However, the role played by the gene in radiosensitivity remains unknown. Herein, we examined the relationship between FAM135B and radiosensitivity. According to the results, FAM135B is highly expressed in ESCC cells, and ESCC cells with high levels of FAM135B are resistant to irradiation. Silencing FAM135B inhibits colony formation capability and cell cycle protein expression (pP53, CDK1), promotes cell cycle arrest at the G2/M phase following irradiation. Moreover, transcriptome sequencing analysis demonstrates that FAM135B regulates downstream PI3K/Akt/mTOR signaling pathway, and western blot verifies the result. One of the mechanisms of increasing radiosensitivity by silencing FAM135B expression in ESCC cells may be achieved by regulating the PI3K/Akt/mTOR signaling pathway. Silencing FAM135B shows synergy with PI3K/Akt/mTOR pathway inhibitor (rapamycin) in increasing radiosensitivity, regulating the expression of cell cycle protein and inducing apoptosis of ESCC cells. The results indicate that FAM135B could be a potential treatment target for ESCC in management of radiosensitivity.


Assuntos
Neoplasias Esofágicas/genética , Carcinoma de Células Escamosas do Esôfago/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , RNA Interferente Pequeno/farmacologia , Regulação para Cima/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/efeitos da radiação , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/efeitos da radiação , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Neoplasias Esofágicas/terapia , Carcinoma de Células Escamosas do Esôfago/terapia , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos da radiação , Inativação Gênica , Humanos , Tolerância a Radiação/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/efeitos da radiação , Sirolimo/farmacologia
14.
Inflamm Res ; 69(12): 1215-1234, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33044562

RESUMO

OBJECTIVE AND DESIGN: Macrophages exhibit strong phenotypic plasticity and can mediate renal inflammation by polarizing into an M1 phenotype. They play a pivotal role in diabetic nephropathy (DN). Here, we have investigated the regulatory role of transforming growth factor ß-activated kinase 1-binding protein 1 (TAB1) in glycolysis and activation of macrophages during DN. METHODS: TAB1 was inhibited using siRNA in high glucose (HG)-stimulated bone marrow-derived macrophages (BMMs) and lentiviral vector-mediated TAB1 knockdown was used in streptozotocin (STZ)-induced diabetic mice. Western blotting, flow cytometry, qRT-PCR, ELISA, PAS staining and immunohistochemical staining were used for assessment of TAB1/nuclear factor-κB (NF-κB)/hypoxia-inducible factor-1α (HIF-1α), iNOS, glycolysis, inflammation and the clinical and pathological manifestations of diabetic nephropathy. RESULTS: We found that TAB1/NF-κB/HIF-1α, iNOS and glycolysis were up-regulated in BMMs under HG conditions, leading to release of further inflammatory factors, Downregulation of TAB1 could inhibit glycolysis/polarization of macrophages and inflammation in vivo and in vitro. Furthermore, albuminuria, the tubulointerstitial damage index and glomerular mesangial expansion index of STZ-induced diabetic nephropathy mice were decreased by TAB1 knockdown. CONCLUSIONS: Our results suggest that the TAB1/NF-κB/HIF-1α signaling pathway regulates glycolysis and activation of macrophages in DN.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Neuropatias Diabéticas/genética , Neuropatias Diabéticas/metabolismo , Glicólise/genética , Ativação de Macrófagos/genética , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Albuminúria/tratamento farmacológico , Albuminúria/patologia , Animais , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/metabolismo , Técnicas de Silenciamento de Genes , Subunidade alfa do Fator 1 Induzível por Hipóxia/efeitos dos fármacos , Células Mesangiais/patologia , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/efeitos dos fármacos , Nefrite Intersticial/patologia , Óxido Nítrico Sintase Tipo II/efeitos dos fármacos , RNA Interferente Pequeno/farmacologia
15.
Life Sci ; 260: 118423, 2020 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-32941896

RESUMO

AIMS: Increased expression of inhibitor of apoptosis (IAP) genes has been associated with progressive cancer and chemoresistance. Accordingly, blockade of IAPs by BV6 has resulted in ameliorative outcomes. Interleukin (IL)-6 is another important mediator involved in the growth and survival of tumor cells. Therefore, we hypothesized that simultaneous inhibition of IAPs and IL-6 could be a new promising anti-tumor treatment strategy. MATERIALS AND METHODS: In this study, we generated and characterized hyaluronate-PEG-Chitosan-Lactate (H-PCL) nanoparticles (NPs) to simultaneously deliver IL6-specific siRNA and BV6 to 4T1 (breast cancer) and CT26 (colon cancer) cells, and investigate the anti-tumor properties of this combination therapy both in vitro and in vivo. KEY FINDINGS: H-PCL NPs exhibited good physicochemical properties leading to efficient transfection of cancer cells and suppression of target molecules. Moreover, combination therapy synergistically increased apoptosis, as well as decreased cell migration, proliferation, colony formation, and angiogenesis in both 4T1 and CT26 cell lines and suppressed cancer progression in tumor-bearing mice that was associated with enhanced survival time. SIGNIFICANCE: These findings imply the effectiveness of cancer combination therapy by using H-PCL NPs loaded with anti-IL-6 siRNA and BV6.


Assuntos
Antineoplásicos/administração & dosagem , Interleucina-6/genética , Nanopartículas/administração & dosagem , Nanopartículas/química , Oligopeptídeos/administração & dosagem , RNA Interferente Pequeno/administração & dosagem , Animais , Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Embrião de Galinha/irrigação sanguínea , Embrião de Galinha/efeitos dos fármacos , Quitosana/química , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/patologia , Sistemas de Liberação de Medicamentos/métodos , Liberação Controlada de Fármacos , Feminino , Humanos , Interleucina-6/antagonistas & inibidores , Ácido Láctico/química , Espectroscopia de Ressonância Magnética , Camundongos Endogâmicos BALB C , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/patologia , Oligopeptídeos/farmacocinética , Oligopeptídeos/farmacologia , Polietilenoglicóis/química , RNA Interferente Pequeno/farmacocinética , RNA Interferente Pequeno/farmacologia , Espectroscopia de Infravermelho com Transformada de Fourier , Ensaios Antitumorais Modelo de Xenoenxerto
16.
Pharmacol Rev ; 72(4): 862-898, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32929000

RESUMO

RNA-based therapies, including RNA molecules as drugs and RNA-targeted small molecules, offer unique opportunities to expand the range of therapeutic targets. Various forms of RNAs may be used to selectively act on proteins, transcripts, and genes that cannot be targeted by conventional small molecules or proteins. Although development of RNA drugs faces unparalleled challenges, many strategies have been developed to improve RNA metabolic stability and intracellular delivery. A number of RNA drugs have been approved for medical use, including aptamers (e.g., pegaptanib) that mechanistically act on protein target and small interfering RNAs (e.g., patisiran and givosiran) and antisense oligonucleotides (e.g., inotersen and golodirsen) that directly interfere with RNA targets. Furthermore, guide RNAs are essential components of novel gene editing modalities, and mRNA therapeutics are under development for protein replacement therapy or vaccination, including those against unprecedented severe acute respiratory syndrome coronavirus pandemic. Moreover, functional RNAs or RNA motifs are highly structured to form binding pockets or clefts that are accessible by small molecules. Many natural, semisynthetic, or synthetic antibiotics (e.g., aminoglycosides, tetracyclines, macrolides, oxazolidinones, and phenicols) can directly bind to ribosomal RNAs to achieve the inhibition of bacterial infections. Therefore, there is growing interest in developing RNA-targeted small-molecule drugs amenable to oral administration, and some (e.g., risdiplam and branaplam) have entered clinical trials. Here, we review the pharmacology of novel RNA drugs and RNA-targeted small-molecule medications, with a focus on recent progresses and strategies. Challenges in the development of novel druggable RNA entities and identification of viable RNA targets and selective small-molecule binders are discussed. SIGNIFICANCE STATEMENT: With the understanding of RNA functions and critical roles in diseases, as well as the development of RNA-related technologies, there is growing interest in developing novel RNA-based therapeutics. This comprehensive review presents pharmacology of both RNA drugs and RNA-targeted small-molecule medications, focusing on novel mechanisms of action, the most recent progress, and existing challenges.


Assuntos
RNA/efeitos dos fármacos , RNA/farmacologia , Aptâmeros de Nucleotídeos/farmacologia , Aptâmeros de Nucleotídeos/uso terapêutico , Betacoronavirus , Técnicas de Química Analítica/métodos , Técnicas de Química Analítica/normas , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Infecções por Coronavirus/tratamento farmacológico , Sistemas de Liberação de Medicamentos/métodos , Desenvolvimento de Medicamentos/organização & administração , Descoberta de Drogas , Humanos , MicroRNAs/farmacologia , MicroRNAs/uso terapêutico , Oligonucleotídeos Antissenso/farmacologia , Oligonucleotídeos Antissenso/uso terapêutico , Pandemias , Pneumonia Viral/tratamento farmacológico , RNA/efeitos adversos , RNA Antissenso/farmacologia , RNA Antissenso/uso terapêutico , RNA Guia/farmacologia , RNA Guia/uso terapêutico , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/farmacologia , RNA Ribossômico/efeitos dos fármacos , RNA Ribossômico/farmacologia , RNA Interferente Pequeno/farmacologia , RNA Interferente Pequeno/uso terapêutico , RNA Viral/efeitos dos fármacos , Ribonucleases/metabolismo , Riboswitch/efeitos dos fármacos
17.
Life Sci ; 260: 118294, 2020 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-32818544

RESUMO

OBJECTIVE: To explore whether GOLPH3 regulated oxaliplatin (L-OHP) resistance of colon cancer cells via PI3K/AKT/mTOR pathway. METHODS: HCT116/L-OHP cells were divided into Blank, Control/GOLPH3 shRNA, BEZ235 (a PI3K/AKT/mTOR inhibitor), and GOLPH3 + BEZ235 groups followed by the detection with MTT, soft agar colony formation, flow cytometry and TUNEL assays. Mice bearing HCT116/L-OHP xenografts were randomized into Control, L-OHP, NC/GOLPH3 shRNA, L-OHP + NC/GOLPH3 shRNA groups. The expressions of Ki67, Caspase-3, and PI3K/AKT/mTOR pathway proteins were examined by immunohistochemistry. RESULTS: HCT116/L-OHP cells had increased GOLPH3 expression compared to HCT116 cells, which positively regulated PI3K/AKT/mTOR pathway in HCT116/L-OHP cells. BEZ235 declined IC50 of HCT116/L-OHP cells to L-OHP, decreased the expressions of ABCB1, ABCC1, ABCG2, ATP7A, ATP7B, MATE1, p-gp, MRP1 and BCRP, induced cell apoptosis, reduced cell proliferation, and arrested cells at G0/G1, which was reversed by GOLPH3 overexpression. L-OHP and GOLPH3 shRNA decreased tumor volume and reduced expression of Ki67 in tumor tissues with the increased Caspase-3. Meanwhile, the combined treatment had the better treatment effect. CONCLUSION: GOLPH3 inhibition reduced proliferation and promoted apoptosis of HCT116/L-OHP cells, and also reversed the L-OHP resistance of HCT116/L-OHP, which may be associated with the suppression of P13K/AKT/mTOR pathway.


Assuntos
Resistencia a Medicamentos Antineoplásicos/fisiologia , Proteínas de Membrana/fisiologia , Oxaliplatina/uso terapêutico , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Animais , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/patologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Células HCT116 , Humanos , Imidazóis/farmacologia , Masculino , Proteínas de Membrana/antagonistas & inibidores , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Oxaliplatina/farmacologia , Quinolinas/farmacologia , RNA Interferente Pequeno/farmacologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia
18.
Life Sci ; 259: 118199, 2020 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-32781064

RESUMO

Cellular senescence, a process whereby cells enter a state of permanent growth arrest, appears to regulate cardiac pathological remodeling and dysfunction in response to various stresses including myocardial infarction (MI). However, the precise role as well as the underlying regulatory mechanism of cardiac cellular senescence in the ischemic heart disease remain to be further determined. Herein we report an inhibitory role of Nrf2, a key transcription factor of cellular defense, in regulating cardiac senescence in infarcted hearts as well as a therapeutic potential of targeting Nrf2-mediated suppression of cardiac senescence in the treatment of MI-induced cardiac dysfunction. MI was induced by left coronary artery ligation for 28 days in mice. Heart tissues from the infarct border zone were used for the analyses. The MI-induced cardiac dysfunction was associated with increased myocardial cell senescence, oxidative stress and apoptosis in adult wild type (WT) mice. In addition, a downregulated Nrf2 activity was associated with upregulated Keap1 levels and increased phosphorylation of JAK and FYN in the infarcted border zone heart tissues. Nrf2 Knockout (Nrf2-/-) enhanced the MI-induced myocardial, cardiac dysfunction and senescence. Qiliqiangxin (QLQX), a herbal medicine which could reverse the MI-induced suppression of Nrf2 activity, significantly inhibited the MI-induced cardiac senescence, apoptosis, and cardiac dysfunction in WT mice but not in Nrf2-/- mice. These results indicate that MI downregulates Nrf2 activity thus promoting oxidative stress to accelerate cellular senescence in the infarcted heart towards cardiac dysfunction and Nrf2 may be a drug target for suppressing the cellular senescence-associated pathologies in infarcted hearts.


Assuntos
Cardiomiopatias/genética , Cardiomiopatias/patologia , Senescência Celular/genética , Infarto do Miocárdio/genética , Infarto do Miocárdio/patologia , Miocárdio/patologia , Fator 2 Relacionado a NF-E2/genética , Animais , Cardiomiopatias/diagnóstico por imagem , Ecocardiografia , Inativação Gênica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Infarto do Miocárdio/diagnóstico por imagem , Miócitos Cardíacos/metabolismo , RNA Interferente Pequeno/farmacologia , Remodelação Ventricular/fisiologia
19.
Life Sci ; 259: 118150, 2020 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-32726663

RESUMO

Conventional therapeutic methods against cancer, including chemotherapy, radiotherapy, surgery, and combination therapy, have exhibited different toxicity levels due to their unspecific mechanism of action. To overcome the challenges facing conventional cancer therapies, newly developed methods are being investigated. Significant levels of specificity, remarkable accumulation at the tumor site, limited side effects, and minimal off-target effects enable the newly synthesized nanoparticles (NPs) to become the preferred drug delivery method in anticancer therapeutic approaches. According to the literature, CD73 has a pivotal role in cancer progression and resistance to chemotherapy and radiotherapy. Therefore, CD73 has attracted considerable attention among scientists to target this molecule. Accordingly, FDA approved CDK inhibitors such as Dinaciclib that blocks CDK1, 2, 5, and 9, and exhibits significant anticancer activity. So in this study, we intended to simultaneously suppress CD73 and CDKs in cancer cells by using the folic acid (FA)-conjugated chitosan-lactate (CL) NPs loaded with anti-CD73 siRNA and Dinaciclib to control tumor progression and metastasis. The results showed that NPs could effectively transfect cancer cells in a FA receptor-dependent manner leading to suppression of proliferation, survival, migration, and metastatic potential. Moreover, the treatment of tumor-bearing mice with this combination strategy robustly inhibited tumor growth and enhanced survival time in mice. These findings imply the high potential of FA-CL NPs loaded with anti-CD73 siRNA and Dinaciclib for use in cancer treatment shortly.


Assuntos
5'-Nucleotidase/efeitos dos fármacos , Antineoplásicos/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Ácido Fólico , Nanopartículas , Compostos de Piridínio/farmacologia , RNA Interferente Pequeno/administração & dosagem , RNA Interferente Pequeno/farmacologia , 5'-Nucleotidase/antagonistas & inibidores , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Terapia Combinada , Quinases Ciclina-Dependentes/efeitos dos fármacos , Progressão da Doença , Sistemas de Liberação de Medicamentos , Sinergismo Farmacológico , Humanos , Camundongos , Metástase Neoplásica/tratamento farmacológico , Neoplasias Experimentais/tratamento farmacológico , Ensaio Tumoral de Célula-Tronco
20.
Proc Natl Acad Sci U S A ; 117(32): 19415-19424, 2020 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-32719125

RESUMO

Synthetic lethality strategies for cancer therapy exploit cancer-specific genetic defects to identify targets that are uniquely essential to the survival of tumor cells. Here we show RAD27/FEN1, which encodes flap endonuclease 1 (FEN1), a structure-specific nuclease with roles in DNA replication and repair, and has the greatest number of synthetic lethal interactions with Saccharomyces cerevisiae genome instability genes, is a druggable target for an inhibitor-based approach to kill cancers with defects in homologous recombination (HR). The vulnerability of cancers with HR defects to FEN1 loss was validated by studies showing that small-molecule FEN1 inhibitors and FEN1 small interfering RNAs (siRNAs) selectively killed BRCA1- and BRCA2-defective human cell lines. Furthermore, the differential sensitivity to FEN1 inhibition was recapitulated in mice, where a small-molecule FEN1 inhibitor reduced the growth of tumors established from drug-sensitive but not drug-resistant cancer cell lines. FEN1 inhibition induced a DNA damage response in both sensitive and resistant cell lines; however, sensitive cell lines were unable to recover and replicate DNA even when the inhibitor was removed. Although FEN1 inhibition activated caspase to higher levels in sensitive cells, this apoptotic response occurred in p53-defective cells and cell killing was not blocked by a pan-caspase inhibitor. These results suggest that FEN1 inhibitors have the potential for therapeutically targeting HR-defective cancers such as those resulting from BRCA1 and BRCA2 mutations, and other genetic defects.


Assuntos
Antineoplásicos/farmacologia , Endonucleases Flap/antagonistas & inibidores , Recombinação Homóloga/efeitos dos fármacos , Neoplasias/genética , Animais , Proteína BRCA1/deficiência , Proteína BRCA1/genética , Proteína BRCA2/deficiência , Proteína BRCA2/genética , Linhagem Celular Tumoral , Dano ao DNA/efeitos dos fármacos , Reparo do DNA/efeitos dos fármacos , Replicação do DNA/efeitos dos fármacos , Endonucleases Flap/genética , Instabilidade Genômica/genética , Humanos , Camundongos , Neoplasias/tratamento farmacológico , RNA Interferente Pequeno/farmacologia , Saccharomyces cerevisiae , Proteínas de Saccharomyces cerevisiae/genética , Bibliotecas de Moléculas Pequenas/farmacologia , Mutações Sintéticas Letais , Ensaios Antitumorais Modelo de Xenoenxerto
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...