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1.
Phytochemistry ; 191: 112903, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34384922

RESUMO

Six new Cephalotaxus alkaloids, including five cephalotaxine-type alkaloids, and one homoerythrina-type alkaloid, along with six known analogues, were isolated from the seeds of Cephalotaxus fortunei. Their structures were elucidated by combination of spectroscopic data analyses, time-dependent density functional theory (TDDFT) ECD calculation, and single-crystal X-ray diffraction. Cephalofortine B represents the first example of C-5 epi-cephalotaxine-type alkaloid. All isolated compounds were tested for cytotoxicities against HCT-116, A375, and SK-Mel-28 cell lines. Cephalofortine E showed moderate activity against HCT-116 cell line, with an IC50 value of 7.46 ± 0.77 µM.


Assuntos
Alcaloides , Antineoplásicos Fitogênicos , Cephalotaxus , Harringtoninas , Alcaloides/farmacologia , Antineoplásicos Fitogênicos/farmacologia , Harringtoninas/farmacologia , Mepesuccinato de Omacetaxina , Humanos , Estrutura Molecular , Sementes
2.
Molecules ; 26(5)2021 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-33806554

RESUMO

Harringtonolide (HO), a natural product isolated from Cephalotaxus harringtonia, exhibits potent antiproliferative activity. However, little information has been reported on the systematic structure-activity relationship (SAR) of HO derivatives. Modifications on tropone, lactone, and allyl positions of HO (1) were carried out to provide 17 derivatives (2-13, 11a-11f). The in vitro antiproliferative activity against four cancer cell lines (HCT-116, A375, A549, and Huh-7) and one normal cell line (L-02) was tested. Amongst these novel derivatives, compound 6 exhibited comparable cell growth inhibitory activity to HO and displayed better selectivity index (SI = 56.5) between Huh-7 and L-02 cells. The SAR results revealed that the tropone and lactone moieties are essential for the cytotoxic activities, which provided useful suggestions for further structural optimization of HO.


Assuntos
Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Proliferação de Células , Harringtoninas/química , Neoplasias/tratamento farmacológico , Relação Dose-Resposta a Droga , Desenho de Fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Estrutura Molecular , Relação Estrutura-Atividade
3.
Angew Chem Int Ed Engl ; 60(21): 12060-12065, 2021 05 17.
Artigo em Inglês | MEDLINE | ID: mdl-33733565

RESUMO

While numerous studies pertaining to the total synthesis of Cephalotaxus alkaloids have been reported, only two strategies have been reported to date for the successful synthesis of the C-11 oxygenated subset, due to the additional synthetic challenge posed by the remote C-11 stereocenter. Herein, we report the collective asymmetric total synthesis of C-11 oxygenated Cephalotaxus alkaloids using a chiral proline both as a starting material and as the only chirality source. A tetracyclic advanced intermediate was synthesized in a highly stereoselective manner from l-proline in 8 steps involving sequential chirality transfer steps such as a diastereoselective N-alkylation, stereospecific Stevens rearrangement and intramolecular Friedel-Crafts reaction via an unusual O-acyloxocarbenium intermediate. From a common intermediate, the asymmetric total synthesis of six C-11 oxygenated Cephalotaxus alkaloids was completed by a series of oxidation state adjustments.


Assuntos
Harringtoninas/síntese química , Alquilação , Ciclização , Estrutura Molecular , Oxirredução , Prolina/química , Estereoisomerismo
4.
Nucleic Acids Res ; 49(2): e9, 2021 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-33264395

RESUMO

There has been a surge of interest towards targeting protein synthesis to treat diseases and extend lifespan. Despite the progress, few options are available to assess translation in live animals, as their complexity limits the repertoire of experimental tools to monitor and manipulate processes within organs and individual cells. It this study, we developed a labeling-free method for measuring organ- and cell-type-specific translation elongation rates in vivo. It is based on time-resolved delivery of translation initiation and elongation inhibitors in live animals followed by ribosome profiling. It also reports translation initiation sites in an organ-specific manner. Using this method, we found that the elongation rates differ more than 50% among mouse organs and determined them to be 6.8, 5.0 and 4.3 amino acids per second for liver, kidney, and skeletal muscle, respectively. We further found that the elongation rate is reduced by 20% between young adulthood and mid-life. Thus, translation, a major metabolic process in cells, is tightly regulated at the level of elongation of nascent polypeptide chains.


Assuntos
Envelhecimento/metabolismo , Rim/metabolismo , Fígado/metabolismo , Músculo Esquelético/metabolismo , Elongação Traducional da Cadeia Peptídica , Envelhecimento/genética , Animais , Análise por Conglomerados , Cavidades Cranianas , Cicloeximida/administração & dosagem , Cicloeximida/farmacologia , Esquema de Medicação , Harringtoninas/administração & dosagem , Harringtoninas/farmacologia , Sequenciamento de Nucleotídeos em Larga Escala , Injeções Intravenosas , Cinética , Longevidade , Macrolídeos/administração & dosagem , Macrolídeos/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Órbita , Especificidade de Órgãos , Elongação Traducional da Cadeia Peptídica/efeitos dos fármacos , Iniciação Traducional da Cadeia Peptídica , Piperidonas/administração & dosagem , Piperidonas/farmacologia , Ribossomos/metabolismo , Cauda , Transcriptoma
5.
Biomolecules ; 10(11)2020 11 06.
Artigo em Inglês | MEDLINE | ID: mdl-33172112

RESUMO

Lung cancer is the major cause of cancer-associated death worldwide, and development of new therapeutic drugs is needed to improve treatment outcomes. Three-dimensional (3D) tumorspheroids offer many advantages over conventional two-dimensional cell cultures due to the similarities to in vivo tumors. We found that isoharringtonine, a natural product purified from Cephalotaxus koreana Nakai, significantly inhibited the growth of tumorspheroids with NCI-H460 cells in a dose-dependent manner and induced apoptotic cell death in our 3D cell culture system. On the other hand, A549 tumorspheroids displayed low sensitivity to isoharringtonine-induced apoptosis. Nuclear receptor subfamily 4 group A member 1 (NR4A1) is an orphan nuclear receptor known to regulate proliferation and apoptosis of cancer cells. We observed that knockdown of NR4A1 dramatically increased isoharringtonine-induced cancer cell death in A549 tumorspheroids by activating the intrinsic apoptosis pathway. Furthermore, treatment with combined isoharringtonine and iNR4A1 significantly inhibited multivulva formation in a Caenorhabditis elegans model and tumor development in a xenograft mouse model. Taken together, our data suggest that isoharringtonine is a potential natural product for treatment of non-small cell lung cancers, and inhibition of NR4A1 sensitizes cancer cells to anti-cancer treatment.


Assuntos
Apoptose/efeitos dos fármacos , Carcinogênese/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/patologia , Harringtoninas/farmacologia , Neoplasias Pulmonares/patologia , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Camundongos , Transdução de Sinais/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
6.
Molecules ; 25(18)2020 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-32906689

RESUMO

Mosquito-borne Zika virus (ZIKV) is a Flavivirus that came under intense study from 2014 to 2016 for its well-known ability to cause congenital microcephaly in fetuses and neurological Guillain-Barré disease in adults. Substantial research on screening antiviral agents against ZIKV and preventing ZIKV infection are globally underway, but Food and Drug Administration (FDA)-approved treatments are not available yet. Compounds from Chinese medicinal herbs may offer an opportunity for potential therapies for anti-ZIKV infection. In this study, we evaluated the antiviral efficacy of harringtonine against ZIKV. Harringtonine possessed anti-ZIKV properties against the binding, entry, replication, and release stage through the virus life cycle. In addition, harringtonine have strong virucidal effects in ZIKV and exhibited prophylaxis antiviral ability prior ZIKV infection. The antiviral activity also observed in the treatment against Japanese encephalitis reporter virus (RP9-GFP strain). Overall, this study demonstrated that harringtonine would be a favorable potential candidate for the development of anti-ZIKV infection therapies.


Assuntos
Antivirais/farmacologia , Harringtoninas/farmacologia , Infecção por Zika virus/virologia , Zika virus/efeitos dos fármacos , Animais , Antivirais/química , Células Cultivadas , Chlorocebus aethiops , Harringtoninas/química , Humanos , Modelos Moleculares , Conformação Molecular , Relação Estrutura-Atividade , Células Vero , Proteínas do Envelope Viral/antagonistas & inibidores , Proteínas do Envelope Viral/química , Ligação Viral/efeitos dos fármacos , Internalização do Vírus/efeitos dos fármacos , Liberação de Vírus , Replicação Viral/efeitos dos fármacos , Zika virus/genética , Infecção por Zika virus/tratamento farmacológico
7.
Hum Exp Toxicol ; 39(10): 1405-1416, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32431164

RESUMO

This study aimed to evaluate the modes of action of harringtonine (HT) and homoharringtonine (HHT) alkaloids in cell with wild (HepG2/C3A) and mutant p53 (HuH-7.5). We performed assays for cytotoxicity, genotoxicity, induction of apoptosis, cell cycle phase, and membrane integrity. Obtained data were compared with the relative expression of mRNA of genes related to proliferation, apoptosis, cell cycle control, metabolism of xenobiotics, and reticulum endoplasmic stress. The relative expression of the genes showed an increase in apoptosis-inducing mRNAs, such as TNF and BBC3, as well as a reduction in BCL2 and BAK. The mRNAs of CYP2E1 and CYP2C19 xenobiotic metabolism genes increased in both lineages, while CYP3A4 increased only in the HuH-7.5 lineage. The mRNA expression of endoplasmic reticulum (ER) stress genes (ERN1 and EIF2AK3) was shown to increase in HHT and HT treatments. A similar increase was recorded in the mRNA expression of the TRAF2 gene. The changes observed in this study support the hypothesis that ER stress was more strongly associated with TNF induction, causing cell death by apoptosis in p53 mutant cells. This result with wild and mutant p53 cells may have clinical implications in the use of these compounds.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Harringtoninas/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Proteína Supressora de Tumor p53/genética , Apoptose/efeitos dos fármacos , Carcinoma Hepatocelular/genética , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Humanos , Neoplasias Hepáticas/genética
8.
Nucleic Acids Res ; 48(10): 5201-5216, 2020 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-32382758

RESUMO

High-throughput methods, such as ribosome profiling, have revealed the complexity of translation regulation in Bacteria and Eukarya with large-scale effects on cellular functions. In contrast, the translational landscape in Archaea remains mostly unexplored. Here, we developed ribosome profiling in a model archaeon, Haloferax volcanii, elucidating, for the first time, the translational landscape of a representative of the third domain of life. We determined the ribosome footprint of H. volcanii to be comparable in size to that of the Eukarya. We linked footprint lengths to initiating and elongating states of the ribosome on leadered transcripts, operons, and on leaderless transcripts, the latter representing 70% of H. volcanii transcriptome. We manipulated ribosome activity with translation inhibitors to reveal ribosome pausing at specific codons. Lastly, we found that the drug harringtonine arrested ribosomes at initiation sites in this archaeon. This drug treatment allowed us to confirm known translation initiation sites and also reveal putative novel initiation sites in intergenic regions and within genes. Ribosome profiling revealed an uncharacterized complexity of translation in this archaeon with bacteria-like, eukarya-like, and potentially novel translation mechanisms. These mechanisms are likely to be functionally essential and to contribute to an expanded proteome with regulatory roles in gene expression.


Assuntos
Códon/metabolismo , Haloferax volcanii/genética , Haloferax volcanii/metabolismo , Biossíntese de Proteínas , Ribossomos/metabolismo , Regiões 5' não Traduzidas/genética , Códon/genética , Haloferax volcanii/efeitos dos fármacos , Harringtoninas/farmacologia , Elongação Traducional da Cadeia Peptídica/efeitos dos fármacos , Elongação Traducional da Cadeia Peptídica/genética , Iniciação Traducional da Cadeia Peptídica/efeitos dos fármacos , Iniciação Traducional da Cadeia Peptídica/genética , Biossíntese de Proteínas/efeitos dos fármacos , Pegadas de Proteínas , Fases de Leitura/genética , Ribossomos/efeitos dos fármacos , Transcriptoma/efeitos dos fármacos
9.
J Biol Chem ; 294(22): 8942-8958, 2019 05 31.
Artigo em Inglês | MEDLINE | ID: mdl-31004035

RESUMO

The endoplasmic reticulum (ER) is a nexus for mRNA localization and translation, and recent studies have demonstrated that ER-bound ribosomes also play a transcriptome-wide role in regulating proteome composition. The Sec61 translocon (SEC61) serves as the receptor for ribosomes that translate secretory/integral membrane protein-encoding mRNAs, but whether SEC61 also serves as a translation site for cytosolic protein-encoding mRNAs remains unknown. Here, using a BioID proximity-labeling approach in HEK293T Flp-In cell lines, we examined interactions between ER-resident proteins and ribosomes in vivo Using in vitro analyses, we further focused on bona fide ribosome interactors (i.e. SEC61) and ER proteins (ribophorin I, leucine-rich repeat-containing 59 (LRRC59), and SEC62) previously implicated in associating with ribosomes. We observed labeling of ER-bound ribosomes with the SEC61ß and LRRC59 BioID reporters, comparatively modest labeling with the ribophorin I reporter, and no labeling with the SEC62 reporter. A biotin pulse-chase/subcellular fractionation approach to examine ribosome exchange at the SEC61ß and LRRC59 sites revealed that, at steady state, ribosomes at these sites comprise both rapid- and slow-exchanging pools. Global translational initiation arrest elicited by the inhibitor harringtonine accelerated SEC61ß reporter-labeled ribosome exchange. RNA-Seq analyses of the mRNAs associated with SEC61ß- and LRRC59-labeled ribosomes revealed both site-enriched and shared mRNAs and further established that the ER has a transcriptome-wide role in regulating proteome composition. These results provide evidence that ribosomes interact with the ER membrane via multiple modes and suggest regulatory mechanisms that control global proteome composition via ER membrane-bound ribosomes.


Assuntos
Retículo Endoplasmático/metabolismo , Perfilação da Expressão Gênica/métodos , Ribossomos/metabolismo , Biotina/genética , Biotina/metabolismo , Citosol/metabolismo , Genes Reporter , Células HEK293 , Harringtoninas/química , Humanos , Proteínas de Membrana/química , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Polirribossomos/metabolismo , Biossíntese de Proteínas , Subunidades Proteicas/química , Subunidades Proteicas/genética , Subunidades Proteicas/metabolismo , RNA Mensageiro/metabolismo , Ribossomos/química , Canais de Translocação SEC/química , Canais de Translocação SEC/genética , Canais de Translocação SEC/metabolismo
10.
J Microbiol ; 57(1): 74-79, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30456755

RESUMO

Harringtonine (HT) and homoharringtonine (HHT), alkaloid esters isolated from the genus Cephalotaxus, exhibit antitumor activity. A semisynthetic HHT has been approved for treatment of chronic myelogenous leukemia. In addition to antileukemic activity, HT and HHT are reported to possess potent antiviral activity. In this study, we investigated the effects of HT and HHT on replication of varicella-zoster virus (VZV) in vitro. HT and HHT, but not their biologically inactive parental alkaloid cephalotaxine (CET), significantly inhibited replication of recombinant VZV-pOka luciferase. Furthermore, HT and HHT, but not CET, strongly induced down-regulation of VZV lytic genes and exerted potent antiviral effects against a VZV clinical isolate. The collective data support the utility of HT and HHT as effective antiviral candidates for treatment of VZV-associated diseases.


Assuntos
Antivirais/farmacologia , Cephalotaxus/química , Ésteres/farmacologia , Herpesvirus Humano 3/efeitos dos fármacos , Mepesuccinato de Omacetaxina/farmacologia , Extratos Vegetais/farmacologia , Antivirais/química , Ésteres/química , Harringtoninas/química , Harringtoninas/farmacologia , Herpesvirus Humano 3/genética , Herpesvirus Humano 3/fisiologia , Mepesuccinato de Omacetaxina/química , Humanos , Extratos Vegetais/química , Replicação Viral/efeitos dos fármacos
11.
Medicine (Baltimore) ; 97(39): e12102, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30278488

RESUMO

To assess treatment response and overall survival (OS) in refractory or relapsed acute myeloid leukemia (R/R AML) patients treated by different common salvage chemotherapy regimens.Medical records data from 142 R/R AML patients were reviewed in this retrospective study. Patients were treated with regimens based on the following drugs: cytarabine, granulocyte colony-stimulating factor (G-CSF), and fludarabine (FLAG) (n = 46); cytarabine and G-CSF in addition to aclarubicin or daunorubicin (CAG/DAG) (n = 30); cytarabine, G-CSF, and cladribine (CLAG) (n = 27); cytarabine, etoposide, and mitoxantrone (MEA) (n = 17); cytarabine plus idarubicin, daunorubicin, or mitoxantrone (IA/DA/MA) (n = 12); and homoharringtonine, cytarabine, and aclarubicin or daunorubicin (HAA/HAD) (n = 10).A total of 43 (35.2%) patients achieved complete remission (CR), 60 (49.2%) patients achieved overall remission rate (ORR), and 18 (14.8%) patients received allogeneic hematopoietic stem cell transplantation (allo-HSCT) after CR. Median OS was 8.0 (95% CI 6.6-9.4) months with a 1-year OS rate of (29.9 ±â€Š3.9)% and 3-year OS rate of (11.1 ±â€Š3.6)%. No difference of CR (P = .621), ORR (P = .385), and allo-HSCT (P = .537) achievement was observed among different chemotherapy regimens. Interestingly, we observed that the CLAG-based regimen did not affect CR (P = .165), while it achieved a numerically higher ORR (P = .093) and was an independent factor for prolonged OS (P = .016). No other regimens were determined to be correlated with CR, ORR, or OS.FLAG-, CAG/DAG-, CLAG-, MEA-, IA/DA/MA- and HAA/HAD-based regimens were found to achieve similar CR rates, while the CLAG-based regimen achieved numerically higher ORR rates and significant favorable OS. Therefore, CLAG-based regimens should be a prioritized treatment option for R/R AML patients.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Leucemia Mieloide Aguda/tratamento farmacológico , Terapia de Salvação/métodos , Aclarubicina/efeitos adversos , Aclarubicina/uso terapêutico , Adulto , Idoso , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Cladribina/efeitos adversos , Cladribina/uso terapêutico , Estudos de Coortes , Citarabina/efeitos adversos , Citarabina/uso terapêutico , Daunorrubicina/efeitos adversos , Daunorrubicina/uso terapêutico , Etoposídeo/efeitos adversos , Etoposídeo/uso terapêutico , Feminino , Fator Estimulador de Colônias de Granulócitos/efeitos adversos , Fator Estimulador de Colônias de Granulócitos/uso terapêutico , Harringtoninas/efeitos adversos , Harringtoninas/uso terapêutico , Humanos , Leucemia Mieloide Aguda/mortalidade , Masculino , Pessoa de Meia-Idade , Mitoxantrona/efeitos adversos , Mitoxantrona/uso terapêutico , Estudos Retrospectivos , Terapia de Salvação/efeitos adversos , Taxa de Sobrevida , Resultado do Tratamento , Vidarabina/efeitos adversos , Vidarabina/análogos & derivados , Vidarabina/uso terapêutico , Adulto Jovem
12.
BMC Complement Altern Med ; 18(1): 164, 2018 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-29788973

RESUMO

BACKGROUND: Homoharringtonine (HHT) is a natural alkaloid with potent antitumor activity, but its precise mechanism of action is still poorly understood. METHODS: We examined the effect of HHT on alternative splicing of Bcl-x and Caspase 9 in various cells using semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). The mechanism of HHT-affected alternative splicing in these cells was investigated by treatment with protein phosphatase inhibitors and overexpression of a protein phosphatase. RESULTS: Treatment with HHT downregulated the levels of anti-apoptotic Bcl-xL and Caspase 9b mRNA with a concomitant increase in the mRNA levels of pro-apoptotic Bcl-xS and Caspase 9a in a dose- and time-dependent manner. Calyculin A, an inhibitor of protein phosphatase 1 (PP1) and protein phosphatase 2A (PP2A), significantly inhibited the effects of HHT on the alternative splicing of Bcl-x and Caspase 9, in contrast to okadaic acid, a specific inhibitor of PP2A. Overexpression of PP1 resulted in a decrease in the ratio of Bcl-xL/xS and an increase in the ratio of Caspase 9a/9b. Moreover, the effects of HHT on Bcl-x and Caspase 9 splicing were enhanced in response to PP1 overexpression. These results suggest that HHT-induced alternative splicing of Bcl-x and Caspase 9 is dependent on PP1 activation. In addition, overexpression of PP1 could induce apoptosis and sensitize MCF7 cells to apoptosis induced by HHT. CONCLUSION: Homoharringtonine regulates the alternative splicing of Bcl-x and Caspase 9 through a PP1-dependent mechanism. Our study reveals a novel mechanism underlying the antitumor activities of HHT.


Assuntos
Processamento Alternativo/efeitos dos fármacos , Caspase 9/metabolismo , Harringtoninas/farmacologia , Proteína Fosfatase 1/metabolismo , Proteína bcl-X/metabolismo , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Mepesuccinato de Omacetaxina , Humanos , Camundongos
13.
Int J Mol Sci ; 19(5)2018 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-29762467

RESUMO

The cell viability and apoptosis of tumor U937 cells treated by blue light (BL) irradiation have been examined. BL irradiation can specially inhibit the proliferation and promote the apoptosis of U937 cells, relating to the production of reactive oxygen species (ROS) and the decline of mitochondrial membrane potential (ΔΨm). The apoptosis is further associated with varying downregulated B-cell lymphoma-extra large (Bcl-XL) and B-cell lymphoma 2 (Bcl-2) genes, upregulated Bcl-2-associated X (Bax) gene, the activation of caspase-3 and caspase-9, and the cleavage of poly (ADP-ribose) polymerase (PARP) by the BL irradiation process. Moreover, BL irradiation induced proliferation inhibition is higher than that treated by a common chemotherapeutic drug of homoharringtonine (HHT). When we synergize BL irradiation with HHT (BL-HHT), a higher proliferation inhibition is obtained than that treated by BL irradiation or HHT alone. These results are helpful for establishing a low toxicity and high efficiency strategy of BL irradiation for clinical treatment of acute myeloid leukemia, not limited to U937 cells.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Proliferação de Células/efeitos da radiação , Harringtoninas/farmacologia , Luz , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Caspases/genética , Caspases/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Mepesuccinato de Omacetaxina , Humanos , Potencial da Membrana Mitocondrial , Poli(ADP-Ribose) Polimerases/genética , Poli(ADP-Ribose) Polimerases/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismo , Proteína bcl-X/metabolismo
14.
Zhongguo Shi Yan Xue Ye Xue Za Zhi ; 26(2): 347-353, 2018 Apr.
Artigo em Chinês | MEDLINE | ID: mdl-29665897

RESUMO

OBJECTIVE: To investigate the effect and possible mechanism of low concentration of triptolide (TPL) combined with homoharringtonine (HHT) on the proliferation and apoptosis of KG-1α cells. METHODS: CCK-8 method was used to detect the antiproliferating effects of different concentrations of TPL and HHT single-use and combined use on KG-1α cells, and the combined index (CI) was calculated. The colony formation ability was also determined by methylcellulose colony formation assay, cell surface molecules, apoptosis rate and cell cycle changes were detected by flow cytometry. Westerrn blot was used to detect the expression of Akt signaling pathway related proteins before and after low dose TPL combined with HHT using. RESULTS: High expression of CD34 and CD123 were on KG-1a cells, which being lack expression of CD38. TPL and HHT dose-dependently inhibited the proliferation of KG-1α cells. Compared with low dosage TPL and HHT single-use groups, the cell proliferation and colony formation efficiency were lower, and the cell apoptosis rate was higher in the combined group. CI values also indicated that low concentration TPL combined with HHT possessed highly synergistic effect. After the combination of the 2 drugs, the expressions of P-Aktser473, P-Aktthr308, BCL-2, PARP and survivin protein were down-regulated and the cleavage of PARP protein was increased. CONCLUSION: Low concentration of TPL combined with HHT can synergistically inhibit KG-1α cell proliferation and induce its apoptosis through the PI3K/Akt signaling pathway and downstream protein.


Assuntos
Apoptose , Proliferação de Células , Linhagem Celular Tumoral , Diterpenos , Compostos de Epóxi , Harringtoninas , Mepesuccinato de Omacetaxina , Humanos , Fenantrenos , Fosfatidilinositol 3-Quinases
15.
Zhongguo Shi Yan Xue Ye Xue Za Zhi ; 26(2): 395-400, 2018 Apr.
Artigo em Chinês | MEDLINE | ID: mdl-29665904

RESUMO

OBJECTIVE: To explore the effects of BTZ plus HHT on proliferation and apoptosis of K562 cells, and to clarify the relationship between the mechanism inderlying the effect of BTZ plus HHT on K562 cells and BCL-2, BAX, MCL-1 proteins. METHODS: The K562 cells were divided into 4 groups by different treatment: BTZ(20 nmol/L), HHT(40 ng/ml), BTZ(20 nmol/L)+HHT(40 ng/ml) and control. The proliferation inhibition rates of K562 cells in each group were detected by using MTT, and the early apoptosis rates of K562 cells in each group were assayed by using flow cytometry with Annexin V-FITC/PI staining. The proteins level of BCL-2, BAX and MCL-1 in each group were examined by using Western blot. RESULTS: The inhibition rate of K562 cell proliferation in combined group was higher than that in BTZ, HHT alone group(P<0.01). The early apoptosis rate of K562 cells in combined group was increased significantly in comparison with BTZ and HHT alone group(P<0.05). The BCL-2 protein level of K562 cells in combined group was significantly lower than that in BTZ and HHT alone group(P<0.05). BAX protein level of K562 cells in combined group was higher than that in BTZ and HHT alone group(P<0.05). The Orders of the MCL-1 protein level of K562 cells in 4 groups were BTZ>Control>BTZ plus HHT>HHT(P<0.05 ). CONCLUSION: The combination of BTZ and HHT exerts the synergistic effect of anti-proliferative activity and induces apoptosis against K562 cells in vitro. The combination can induce apoptosis of K562 cells via suppression of BCL-2 protein and up-regulation of BAX protein. HHT can increase the sensitivity of K562 cells to BTZ by down-regulating the expression of MCL-1 protein.


Assuntos
Bortezomib/farmacologia , Harringtoninas/farmacologia , Apoptose , Proliferação de Células , Mepesuccinato de Omacetaxina , Humanos , Células K562
16.
Biomed Pharmacother ; 103: 435-442, 2018 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-29679903

RESUMO

OBJECTIVES: Breast cancer stem cells (BCSCs) contribute to breast cancer progression, relapse, and treatment resistance. Identification of the natural inhibitory components of BCSCs is therefore critical for clinical treatment. Here, we investigated whether isoharringtonine (IHT) had inhibitory effects on BCSCs in breast cancer cell lines. METHODS: HCC1806, HCC1937, and MCF7 cells were treated with IHT. The proliferation and the migration of cells were detected by MTS assay and wound healing migration assay, respectively. The proportions of BCSCs were determined by flow cytometry and tumor sphere formation assay. Using real-time quantitative polymerase chain reaction (qRT-PCR) and Western blotting, the expression of Nanog and activation of STAT3 were detected, respectively. RESULTS: Results showed that IHT inhibited the proliferation of HCC1806, HCC1937, and MCF-7 cells, and suppressed the migration of HCC1806 and HCC1937 cells in a dose-dependent manner. IHT treatment decreased the proportion of BCSCs in MCF-7, HCC1806, and HCC1937 cells. In addition, the mRNA level of Nanong was significantly downregulated after IHT treatment. We also found an inhibitory effect of IHT on STAT3 activation. CONCLUSION: IHT inhibited the proliferation, migration, and BCSC proportion of breast cancer cell lines via inhibition of the STAT3/Nanong pathway.


Assuntos
Neoplasias da Mama/metabolismo , Harringtoninas/farmacologia , Células-Tronco Neoplásicas/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/efeitos dos fármacos , Neoplasias da Mama/patologia , Movimento Celular/efeitos dos fármacos , Movimento Celular/fisiologia , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Relação Dose-Resposta a Droga , Feminino , Humanos , Células MCF-7 , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/patologia , Fator de Transcrição STAT3/antagonistas & inibidores , Transdução de Sinais/fisiologia
17.
Planta Med ; 84(14): 1038-1044, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29490384

RESUMO

Homoharringtonine (HHT), also known as omacetaxine, is a natural compound found in the genus Cephalotaxus and is a promising pharmaceutical drug used for the treatment of chronic or accelerated phase chronic myeloid leukemia. As a tool for the quantitative determination of HHT, a specific monoclonal antibody against HHT (MAb 6A1) was generated by conjugates prepared via sodium periodate-mediated oxidation. The developed indirect competitive enzyme-linked immunosorbent assay (icELISA) using MAb 6A1 was found to be highly specific and sensitive with a limit of detection for HHT of 48.8 ng/mL. Validation assays to evaluate precision and accuracy of the method were conducted by the use of intra- and inter-assay analysis, recovery test, and comparison analysis between the amounts of HHT determined by ELISA and high-performance liquid chromatography. These results revealed that the established icELISA using MAb 6A1 is specific, sensitive, and reliable enough to be applied to the qualitative analysis for HHT. Furthermore, the results of this study support the usefulness of sodium periodate as a reagent for the conjugation between Cephalotaxus alkaloids and proteins for producing specific antibodies.


Assuntos
Ensaio de Imunoadsorção Enzimática/métodos , Harringtoninas/análise , Anticorpos Monoclonais/imunologia , Cromatografia Líquida de Alta Pressão , Harringtoninas/imunologia , Mepesuccinato de Omacetaxina , Reprodutibilidade dos Testes
18.
Zhongguo Shi Yan Xue Ye Xue Za Zhi ; 26(1): 105-109, 2018 Feb.
Artigo em Chinês | MEDLINE | ID: mdl-29397826

RESUMO

OBJECTIVE: To investigate the effect of homoharringtonine (HHT) on proliferation and apoptosis of CML cell line K562 cells and to explore its possible mechanism through mTOR pathway. METHODS: K562 cells were cultured with different concentrations of HHT or in its combination with mTOR inhibitor rapamycin (RAPA) for 24 hours. The cell viability was analyzed by CCK-8 assay, the cell apoptosis was detected by flow cytometry, the expressions of BCL-6, Caspase-3 and mTOR signal pathway related proteins was assayed by Western blot, the expression of BCL-6 mRNA was determined by RT-PCR. RESULTS: The HHT inhibited proliferation and induced apoptosis of K562 cells in a concentration-dependent manner(r=0.970). With the increasing of HHT concentration, the expression level mTOR signal pathway related proteins increased(r=0.908), while the mRNA and protein expression levels of BCL-6 decreased(rmRNA=-0.961, rprotein =-0.981), as compared with the HHT alone, the combination of HHT with RAPA could down-regulate the expression of mTOR signal pathway related protein and caspase-3, and up-regulated expression of BCL-6. CONCLUSION: HHT induces apoptosis of K562 cells by inhibiting BCL-6 expression through mTOR signal pathway.


Assuntos
Apoptose , Caspase 3 , Proliferação de Células , Harringtoninas , Mepesuccinato de Omacetaxina , Humanos , Células K562 , Serina-Treonina Quinases TOR
19.
Ann Hematol ; 97(5): 865-875, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29450644

RESUMO

Homoharringtonine (HHT) is a known anti-leukemia drug that inhibits multiple myeloma (MM) cells both in vitro and in vivo. Our prior study demonstrated that the potency of HHT in MM cells was compromised significantly when myeloma cells were co-cultured with BM stromal cells. This study aimed to investigate whether PI3K/Akt inhibitor LY294002 could potentiate the antimyeloma activity of HHT against MM cells adhered to BM stromal cells and in vivo xenograft models. A co-culture system composed of MM cells and human stromal cells was employed to mimic MM cells in bone marrow niche. The inhibitory and pro-apoptotic effect of HHT and LY294002 was determined by CCK-8 assay or flow cytometry. Expression of PI3K/Akt signaling molecules and anti-apoptotic protein myeloid cell leukemia-1 (Mcl-1) was assessed by western blot analysis and/or reverse transcription real-time quantitative PCR (RT-qPCR). MM xenografts were used to evaluate antitumor effect of combined therapy with HHT and LY294002. Adhesion to BM stromal cells rendered MM cells resistant to HHT whereas silencing Mcl-1 partly reversed the resistance. LY294002 induced apoptosis in MM cells and potentiated the antimyeloma effects of HHT by inhibiting the PI3K/Akt signal pathway which was abnormally activated during adhesion. LY294002 also enhanced the antimyeloma effect of HHT in in vivo xenograft models. These findings suggest that activation of PI3K/Akt signal pathway was responsible for the resistance to HHT in MM cells adhered to stromal cells. LY294002 can potentiate the antimyeloma activity of HHT both in vitro and in vivo, which may represent a new clinical treatment in MM.


Assuntos
Adesão Celular/efeitos dos fármacos , Cromonas/administração & dosagem , Harringtoninas/administração & dosagem , Morfolinas/administração & dosagem , Mieloma Múltiplo/tratamento farmacológico , Inibidores de Fosfoinositídeo-3 Quinase , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Adulto , Idoso , Animais , Antineoplásicos Fitogênicos/administração & dosagem , Adesão Celular/fisiologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Técnicas de Cocultura , Relação Dose-Resposta a Droga , Sinergismo Farmacológico , Inibidores Enzimáticos/administração & dosagem , Feminino , Mepesuccinato de Omacetaxina , Humanos , Masculino , Camundongos , Camundongos SCID , Pessoa de Meia-Idade , Mieloma Múltiplo/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Células Estromais/efeitos dos fármacos , Células Estromais/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
20.
J Oncol Pharm Pract ; 24(3): 201-208, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29284358

RESUMO

Purpose Omacetaxine mepesuccinate ("omacetaxine") is approved by the US Food and Drug Administration for the treatment of adult patients with chronic- or accelerated-phase chronic myeloid leukemia with resistance and/or intolerance to two or more tyrosine kinase inhibitors. In May 2014, the US Food and Drug Administration approved revisions to the packaging information that included directions for home administration of reconstituted omacetaxine by patients or caregivers using syringes filled at a healthcare facility. We developed recommendations for the transport, storage, and spill-clean procedure of reconstituted omacetaxine for home and clinic administration. Methods We conducted chemical stability and microbial growth studies of reconstituted omacetaxine solution stored in vials and syringes at room temperature or refrigerated for various durations. Several shipping configurations were tested in simulated transport conditions to evaluate their ability to contain solution leakage and maintain product quality during distribution. In addition, we evaluated cleaning products and procedures for their effectiveness in removing residual omacetaxine from household surfaces after mock spills. Results Reconstituted omacetaxine showed limited degradation when refrigerated for 14 days in vials and syringes, and no microbial growth was observed for 12 days after intentional inoculation. In shipping studies, the configurations maintained prepared syringes within the recommended storage temperature range throughout transport and could contain leaks if spills occurred. In the event of an accidental spill in a home environment, effective cleaning can be achieved using household cleaning products and defined procedures. Conclusion These data provide important information regarding the safe transportation and administration of reconstituted omacetaxine in the home and clinic.


Assuntos
Antineoplásicos Fitogênicos/administração & dosagem , Antineoplásicos Fitogênicos/normas , Contaminação de Medicamentos/prevenção & controle , Harringtoninas/administração & dosagem , Harringtoninas/normas , Serviços de Assistência Domiciliar/normas , Adulto , Antineoplásicos Fitogênicos/química , Embalagem de Medicamentos/métodos , Embalagem de Medicamentos/normas , Estabilidade de Medicamentos , Armazenamento de Medicamentos/métodos , Armazenamento de Medicamentos/normas , Harringtoninas/química , Mepesuccinato de Omacetaxina , Humanos , Seringas/microbiologia , Seringas/normas , Estados Unidos , United States Food and Drug Administration
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