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1.
Phytomedicine ; 55: 191-199, 2019 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-30668429

RESUMO

BACKGROUND: Veratrum, hellebore is an important plant species of the Liliaceae family and jervine is the characteristic steroidal alkaloid constituent of Veratrum album. PURPOSE: In the current study, anti-inflammatory and antioxidant effects of jervine isolated from NH4OH-benzene extract of V. album rhizomes were investigated on CAR induced paw edema in rats. METHODS/STUDY DESIGN: In inflammatory study, 50, 100, 200 and 400  mg/kg doses of jervine, 25  mg/kg doses of DIC and IND were orally administered, and the volume of the foots were measured up to their knee arthrosis by plethismometer. After one hour of the oral administration of the all treatments, 0.1 ml of CAR solution (1%) was injected into the foot of the all rat groups and the volume of the foots were measured during 5 h after CAR injection. GPx, SOD, GR, MPO, CAT enzymes activities and GSH, LPO levels of the supernatants of paw homogenates and inflammation biomarkers such as TNF-α and IL-1ß in the rats serums were also estimated. RESULTS: According to the present results, jervine exerted 50.4-73.5% anti-inflammatory effects in carrageenan induced paw edema. Inflammation biomarkers such as TNF-α, IL-1ß and MPO that increased by CAR injection were suppressed by the administrations of all doses of jervine, IND and DIC. In all paw tissues, LPO levels as indicator of oxidative tissue damage were found to be high in CAR-treated group and it was found to be decreased in all doses of jervine. CONCLUSION: Jervine, DIC and IND reduced the negative effects of CAR due to increasing effects on the SOD, CAT, GSH, GPx and GR antioxidants.


Assuntos
Anti-Inflamatórios/farmacologia , Antioxidantes/farmacologia , Alcaloides de Veratrum/farmacologia , Animais , Anti-Inflamatórios/administração & dosagem , Antioxidantes/metabolismo , Carragenina/toxicidade , Avaliação Pré-Clínica de Medicamentos/métodos , Edema/induzido quimicamente , Edema/tratamento farmacológico , Enzimas/metabolismo , Inflamação/tratamento farmacológico , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Extratos Vegetais/química , Ratos Sprague-Dawley , Rizoma/química , Fator de Necrose Tumoral alfa/metabolismo , Veratrum/química , Alcaloides de Veratrum/administração & dosagem , Alcaloides de Veratrum/isolamento & purificação
2.
ACS Nano ; 12(10): 9881-9893, 2018 10 23.
Artigo em Inglês | MEDLINE | ID: mdl-30231203

RESUMO

Pancreatic ductal adenocarcinoma (PDAC) remains one of the most difficult cancers to treat. It is refractory to most existing therapies, including immunotherapies, due to the presence of an excessive desmoplastic stroma, which restricts penetration of drugs and cytotoxic CD8+ T cells. Stromal modulation has shown promising results in the enhancement of immune checkpoint blockade treatment in PDAC. We demonstrate here effective stromal modulation by a polymeric micelle-based nanoformulation to codeliver a sonic hedgehog inhibitor (cyclopamine, abbreviated as CPA) and a cytotoxic chemotherapy drug (paclitaxel, abbreviated as PTX). The formulation, M-CPA/PTX, modulated the PDAC stroma by increasing the intratumoral vasculature density, which then promoted the tumor infiltration by cytotoxic CD8+ T cells without depletion of tumor-restraining α-smooth muscle action-positive fibroblasts and type I collage in the stroma. The combination of M-CPA/PTX and the PD-1 checkpoint blockade significantly prolonged animal survival in an orthotopic murine PDAC model as well as a genetically engineered mouse model of PDAC. The superior antitumor efficacy was mediated by enhanced tumor infiltration of CD8+ T cells without concomitant infiltration of suppressive regulatory T cells or myeloid-derived suppressor cells and by the coordinated action of PTX and interferon-gamma. Our results demonstrate that stroma-modulating nanoformulations are a promising approach to potentiate immune checkpoint blockade therapy of pancreatic cancer.


Assuntos
Antineoplásicos/farmacologia , Carcinoma Ductal Pancreático/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Paclitaxel/farmacologia , Neoplasias Pancreáticas/tratamento farmacológico , Células Estromais/imunologia , Alcaloides de Veratrum/farmacologia , Animais , Antineoplásicos/administração & dosagem , Carcinoma Ductal Pancreático/imunologia , Proliferação de Células/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/imunologia , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Humanos , Injeções Intravenosas , Camundongos , Camundongos Endogâmicos C57BL , Paclitaxel/administração & dosagem , Neoplasias Pancreáticas/imunologia , Células Estromais/patologia , Alcaloides de Veratrum/administração & dosagem
3.
Appl Biochem Biotechnol ; 186(2): 371-383, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29637395

RESUMO

Diabetes mellitus, which is the result of autoimmune destruction of the insulin-producing ß cells, occurs by loss of insulin-secreting capacity. The insufficient source of insulin-producing cells (IPCs) is the major obstacle for using transplantation as diabetes treatment method. The present study suggests a method to form islet-like clusters of IPCs derived from mouse embryonic stem cells (mESCs). This protocol consists of several steps. Before starting this protocol, embryoid bodies (EBs) should be cultured in suspension in conditioned medium of isolated mouse pancreatic islet in combination with activing A to be induced. Then differentiated mESCs were replaced with dishes supplemented with basic fibroblast growth factor (bFGF). Next, bFGF was withdrawn, and cyclopamine and noggin were added. Then the cells were treated with B27, nicotinamide, and islet-conditioned medium for maturation. mESCs, as the control group, were cultured without any treatment. An enhanced expression of pancreatic-specific genes was detected by qRT-PCR and immunofluorescence in the differentiated mESCs. The differentiated mESCsco express other markers of pancreatic islet cells as well as insulin. This method exhibited higher insulin generation and further improvement in IPCs protocol that may result in an unlimited source of ES cells suitable for transplantation. The results indicated that conditioned medium, just as critical components of the stem cell niche associated with other factors, had high potential to differentiate mESCs into IPCs.


Assuntos
Células-Tronco Embrionárias/metabolismo , Proteínas de Homeodomínio/metabolismo , Ilhotas Pancreáticas/metabolismo , Transativadores/metabolismo , Animais , Proteínas de Transporte/administração & dosagem , Diferenciação Celular , Técnicas de Cocultura , Meios de Cultivo Condicionados , Células-Tronco Embrionárias/citologia , Fator 2 de Crescimento de Fibroblastos/metabolismo , Insulina/biossíntese , Ilhotas Pancreáticas/citologia , Masculino , Camundongos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais , Alcaloides de Veratrum/administração & dosagem
4.
Pharm Res ; 35(1): 17, 2018 01 05.
Artigo em Inglês | MEDLINE | ID: mdl-29305793

RESUMO

PURPOSE: The aim of this study was to determine whether co-administration of hedgehog (Hh) pathway inhibitor cyclopamine (CYP) and microtubule stabilizer docetaxel (DTX) as polymer-drug conjugates, methoxy poly(ethylene glycol)-block-poly(2-methyl-2-carboxyl-propylenecarbonate-graft-dodecanol-graft-cyclopamine) (P-CYP) and methoxy poly(ethylene glycol)-block-poly(2-methyl-2-carboxyl-propylene carbonate-graft-dodecanol-graft-docetaxel) (P-DTX) could synergistically inhibit orthotopic pancreatic tumor growth in NSG mice. METHODS: P-DTX and P-CYP were synthesized from mPEG-b-PCC through carbodiimide coupling reaction and characterized by 1H-NMR. The micelles were prepared by film hydration and particle size was measured by dynamic light scattering (DLS). Cytotoxicity, apoptosis and cell cycle analysis of P-DTX and P-CYP were evaluated in MIA PaCa-2 cells. In vivo efficacy of P-DTX and P-CYP were evaluated in NSG mice bearing MIA PaCa-2 cells derived orthotopic pancreatic tumor. RESULTS: P-CYP and P-DTX self-assembled into micelles of <90 nm and their combination therapy efficiently inhibited the proliferation of MIA PaCa-2 cells, induced apoptosis and cell cycle arrest at M-phase more efficiently than P-CYP and P-DTX monotherapies. Furthermore, the combination therapy of P-CYP and P-DTX significantly reduced Hh component expression compared to P-CYP alone as determined by Western blot analysis. Lastly, the combination therapy induced greater inhibition of orthotopic pancreatic tumor growth in NSG mice compared to their monotherapies. CONCLUSION: Combination of polymer conjugated anticancer drug (P-DTX) with polymer conjugated Hh inhibitor (P-CYP) enhanced pancreatic cancer cell killing, apoptosis as well as in vivo tumor growth inhibition with no obvious toxicities.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Neoplasias Pancreáticas/tratamento farmacológico , Polímeros/química , Taxoides/farmacologia , Alcaloides de Veratrum/farmacologia , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/química , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Docetaxel , Portadores de Fármacos , Liberação Controlada de Fármacos , Ouriços-Cacheiros/metabolismo , Humanos , Camundongos , Micelas , Microtúbulos/metabolismo , Metástase Neoplásica , Neoplasias Pancreáticas/patologia , Espectroscopia de Prótons por Ressonância Magnética , Taxoides/administração & dosagem , Taxoides/química , Alcaloides de Veratrum/administração & dosagem , Alcaloides de Veratrum/química
5.
Neurochem Int ; 110: 38-48, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28887094

RESUMO

There is a pressing need of developing approaches for delayed post-stroke therapy for patients who fail to receive thrombolysis within the narrow time window. Neuroprotection of Salvianolic Acids for Injection (SAFI) for cerebral ischemia-reperfusion injury in acute phase has been well documented. The current study was to determine the influence of SAFI at the subacute phase after stroke in mice, and to elucidate the underlying mechanisms. Adult male C57BL/6 mice were subjected to permanent occlusion of the distal middle cerebral artery (dMCAO), followed by daily intraperitoneal injection of SAFI 24 h after stroke for 14 days. Motor behavior was measured by neurological function evaluations weekly, and proliferation, migration, survival and differentiation of neural progenitor cells (NPCs) were examined with immunohistochemistry. Sonic hedgehog (Shh) inhibitor cyclopamine (CYC) was injected to determine the involvement of Shh pathway in the therapeutic effects of SAFI. The results showed that SAFI led to dramatic brain functional improvement, elevated NPCs proliferation, and prompted long-term survival of newborn neurons in the subventricular zone (SVZ). Up-regulation of Shh, Ptch and nuclear translocation of Gli1 were observed in the peri-infarct region, accompanied with robust production of Brain derived neurotrophic factor (BDNF) and Nerve growth factor (NGF). Simultaneous administration with CYC strikingly attenuated the beneficial outcomes of SAFI as well as abolished SAFI induced BDNF and NGF production. Collectively, our study demonstrated SAFI significantly promoted long-term functional recovery and neurogenesis, which might be dependent on Shh signaling mediated BDNF and NGF production. Therefore, SAFI might serve as a potential clinically translatable therapy during recovery stage after stroke.


Assuntos
Alcenos/administração & dosagem , Medicamentos de Ervas Chinesas/administração & dosagem , Proteínas Hedgehog/antagonistas & inibidores , Neurogênese/efeitos dos fármacos , Polifenóis/administração & dosagem , Recuperação de Função Fisiológica/efeitos dos fármacos , Acidente Vascular Cerebral/tratamento farmacológico , Animais , Proteínas Hedgehog/metabolismo , Injeções Intraperitoneais , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurogênese/fisiologia , Distribuição Aleatória , Recuperação de Função Fisiológica/fisiologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Acidente Vascular Cerebral/metabolismo , Alcaloides de Veratrum/administração & dosagem
6.
Oncotarget ; 8(26): 42495-42509, 2017 Jun 27.
Artigo em Inglês | MEDLINE | ID: mdl-28477008

RESUMO

Glioblastoma multiforme (GBM) is the most common and deadly brain cancer, characterized by its aggressive proliferation to adjacent tissue and high recurrence rate. We studied the efficacy and related mechanisms of the combination of cyclopamine (Cyp, a Sonic-hedgehog pathway (Shh) inhibitor) and temozolomide (TMZ, the clinically most used chemotherapeutic agent) in anti-GBM treatment. The micellarized Cyp (MCyp) showed better performance than Cyp solution in inhibiting GBM cells proliferation (3.77-fold against U87 MG cells and 3.28-fold against DBTRG-05MG cells) and clonogenity (1.35-fold against U87 MG cells and 2.17-fold against DBTRG-05MG cells), and preferred behavior of inhibiting cell invasion, colony formation through attenuated Gli1 expression. In addition, combination of MCyp and TMZ exhibited synergistic cytotoxicity, correlating with their ability in inducing apoptosis and eliminating neurospheres formation, and the combination of TMZ was accompanied with the enhanced blockage of Shh pathway. The optimal ratio of MCyp combined to TMZ was 1:20. So we proposed to use TMZ to kill tumor parenchyma and MCyp as the cancer stem cells inhibitor to resist tumor recurrence. These findings demonstrated that combination of TMZ with micellarized Cyp is a promising strategy for exerting different functions of drugs for tumor treatment.


Assuntos
Neoplasias Encefálicas/metabolismo , Dacarbazina/análogos & derivados , Glioblastoma/metabolismo , Micelas , Alcaloides de Veratrum/administração & dosagem , Proteína GLI1 em Dedos de Zinco/metabolismo , Apoptose/efeitos dos fármacos , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Dacarbazina/administração & dosagem , Sinergismo Farmacológico , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glioblastoma/tratamento farmacológico , Glioblastoma/genética , Glioblastoma/patologia , Humanos , Transdução de Sinais/efeitos dos fármacos , Temozolomida , Ensaio Tumoral de Célula-Tronco , Proteína GLI1 em Dedos de Zinco/genética
7.
Nanomedicine ; 13(2): 391-401, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-27520724

RESUMO

Repeated treatments with chemotherapeutic agent(s) fail due to cancer stem cells (CSCs) and chemoresistance regulated by microRNAs (miRNA) whose expression alters owing to dysfunctional signaling pathways including Hedgehog (Hh) signaling. We previously demonstrated the combination of Hh inhibitor cyclopamine (CYP) and paclitaxel (PTX) effectively inhibit PTX-resistant cells and side population, a cell fraction rich in CSCs. In this study, we synthesized mPEG-b-PCC-g-PTX-g-DC (P-PTX) and mPEG-b-PCC-g-CYP-g-DC (P-CYP) polymer-drug conjugates, which they self-assembled into micelles. The combination of P-PTX and P-CYP alleviated PTX resistance and suppressed tumor colony formation. Further, combination therapy inhibited Hh signaling and up-regulated tumor suppressor miRNAs. We established orthotopic prostate tumor in nude mice and there was significant tumor growth inhibition in the group treated with the combination therapy of P-PTX and P-CYP compared with monotherapy. In conclusion, this combination therapy of P-PTX and P-CYP has the potential to treat chemoresistant prostate cancer.


Assuntos
Antineoplásicos/administração & dosagem , Nanoconjugados , Paclitaxel/administração & dosagem , Neoplasias da Próstata/tratamento farmacológico , Alcaloides de Veratrum/administração & dosagem , Animais , Linhagem Celular Tumoral , Sistemas de Liberação de Medicamentos , Humanos , Masculino , Camundongos , Camundongos Nus , Micelas , Polímeros/uso terapêutico
8.
Biomaterials ; 103: 12-21, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27376555

RESUMO

The dense extracellular matrix in pancreatic ductal adenocarcinoma dramatically reduces the penetration and efficacy of nanotherapeutics. Disruption of the tumor extracellular matrix may help improve the distribution and efficacy of nanotherapeutics in pancreatic cancer. In this study, we tested whether cyclopamine, a special inhibitor of the hedgehog signaling pathway with powerful anti-fibrotic activity, could promote the penetration and efficacy of nanotherapeutics in pancreatic cancer. It was shown that cyclopamine disrupted tumor extracellular fibronectins, decompressed tumor blood vessels, and improved tumor perfusion. Furthermore, cyclopamine improved the accumulation and intratumoral distribution of i.v.-administered fluorescence indicator-labeled nanoparticles. Finally, cyclopamine also significantly improved the tumor growth inhibition effect of i.v.-injected nanotherapeutics in pancreatic tumor xenograft mouse models. Thus, cyclopamine may have great potential to improve the therapeutic effects of nanomedicine in patients with pancreatic cancer.


Assuntos
Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/patologia , Nanopartículas/uso terapêutico , Paclitaxel/administração & dosagem , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/metabolismo , Alcaloides de Veratrum/administração & dosagem , Animais , Antineoplásicos Fitogênicos/administração & dosagem , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Sinergismo Farmacológico , Camundongos , Camundongos Nus , Neoplasias Pancreáticas/patologia , Distribuição Tecidual , Resultado do Tratamento
9.
Biomed Chromatogr ; 30(9): 1515-22, 2016 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26972867

RESUMO

A simple and sensitive high-performance liquid chromatography coupled with hybrid triple quadrupole-linear ion trap mass spectrometry (Q-trap-MS) method was developed and validated for the determination of veratramine, the major bioactive and neurotoxic component in Veratrum nigrum L. Veratramine and the internal standard (IS) were separated with a Waters Symmetry C18 column and eluted with a gradient mobile phase system containing acetonitrile and 0.1% aqueous formic acid. The analysis was performed by using positive electrospray ionization mode with multiple reaction monitoring (MRM). Transition ions of m/z 410.2 → 295.2 for veratramine and m/z 426.1 → 113.8 for the IS were monitored. The method was validated with a good linearity in the range of 1-1000 ng/mL and lower limit of quantification of 1 ng/mL. The precision (CV) of intra- and inter-day ranged from 3.92 to 7.29%, while the accuracy (bias) intra- and inter-day were between -4.78 and 1.65%. The recovery, stability and matrix effect were within the acceptable ranges. Five metabolites of veratramine, including four hydroxylated and one sulfated metabolites, were tentatively identified using predictive MRM-information dependent acquisition-enhanced product ion mode (predictive MRM-IDA-EPI). The developed method was successfully applied to the pharmacokinetic and metabolic study of veratramine in mice after oral administration of veratramine. Copyright © 2016 John Wiley & Sons, Ltd.


Assuntos
Cromatografia Líquida/métodos , Espectrometria de Massas em Tandem/métodos , Alcaloides de Veratrum/farmacocinética , Administração Oral , Animais , Limite de Detecção , Camundongos , Padrões de Referência , Alcaloides de Veratrum/administração & dosagem
10.
Biomed Mater ; 11(2): 025011, 2016 Mar 23.
Artigo em Inglês | MEDLINE | ID: mdl-27007569

RESUMO

Ischemic stroke is often associated with loss of cortical neurons leading to various neurological deficits. A cell replacement based on stem cell transplantation to repair the damaged brain requires the generation of specific neuronal subtypes. Recently, induced pluripotent stem cells have been used to generate various subtypes of neurons in vitro for transplantation in stroke-damaged brains. However, whether these cells can be primed as neuronal precursors to become cortical projection neurons by means of biomaterials releasing differentiation factors is not known. Here, we report that microspheres of biodegradable poly(ester-amide) composed of adipic acid, L-phenyl-alanine and 1,4-butanediol, loaded with differentiation factors, can be used to fate human induced pluripotent stem cell-derived long-term expandable neuroepithelial-like stem cells to cortical projection neurons. The three factors, Wnt3A, BMP4 and cyclopamine, were released from loaded microspheres over at least one month following biphasic dynamic time course, promoting cortical differentiation of the cells in vitro. Microspheres did not evoke significant inflammatory response after transplantation into intact rodent brain. Our study shows the potential of biodegradable polymer microspheres to promote neuronal differentiation by continuous release of factors, thereby creating the appropriate microenvironment. This new strategy may improve the efficacy of stem cell-based therapeutic approaches.


Assuntos
Células-Tronco Pluripotentes Induzidas/citologia , Neurônios/citologia , Implantes Absorvíveis , Animais , Materiais Biocompatíveis/química , Proteína Morfogenética Óssea 4/administração & dosagem , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Córtex Cerebral/citologia , Sistemas de Liberação de Medicamentos , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/transplante , Teste de Materiais , Microesferas , Células-Tronco Neurais/citologia , Células-Tronco Neurais/efeitos dos fármacos , Células-Tronco Neurais/transplante , Neurogênese/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Poliésteres/química , Ratos , Ratos Sprague-Dawley , Acidente Vascular Cerebral/terapia , Alcaloides de Veratrum/administração & dosagem , Proteína Wnt3A/administração & dosagem
11.
AAPS J ; 18(2): 432-44, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26791530

RESUMO

Veratramine, a major alkaloid from Veratrum nigrum L., has distinct anti-tumor and anti-hypertension effects. Our previous study indicated that veratramine had severe toxicity toward male rats. In order to elucidate the underling mechanism, in vivo pharmacokinetic experiments and in vitro mechanistic studies have been conducted. Veratramine was administrated to male and female rats intravenously via the jugular vein at a dose of 50 µg/kg or orally via gavage at 20 mg/kg. As a result, significant pharmacokinetic differences were observed between male and female rats after oral administration with much lower concentrations of veratramine and 7-hydroxyl-veratramine and higher concentrations of veratramine-3-O-sulfate found in the plasma and urine of female rats. The absolute bioavailability of veratramine was 0.9% in female rats and 22.5% in male rats. Further experiments of veratramine on Caco-2 cell monolayer model and in vitro incubation with GI content or rat intestinal subcellular fractions demonstrated that its efficient passive diffusion mediated absorption with minimal intestinal metabolism, suggesting no gender-related difference during its absorption process. When veratramine was incubated with male or female rat liver microsomes/cytosols, significant male-predominant formation of 7-hydroxyl-veratramine and female-predominant formation of veratramine-3-O-sulfate were observed. In conclusion, the significant gender-dependent hepatic metabolism of veratramine could be the major contributor to its gender-dependent pharmacokinetics.


Assuntos
Anti-Hipertensivos/farmacocinética , Caracteres Sexuais , Alcaloides de Veratrum/farmacocinética , Administração Intravenosa , Administração Oral , Animais , Anti-Hipertensivos/administração & dosagem , Células CACO-2 , Relação Dose-Resposta a Droga , Feminino , Humanos , Masculino , Microssomos Hepáticos/efeitos dos fármacos , Microssomos Hepáticos/metabolismo , Ratos , Ratos Sprague-Dawley , Distribuição Tecidual/efeitos dos fármacos , Distribuição Tecidual/fisiologia , Alcaloides de Veratrum/administração & dosagem
12.
Biochem Biophys Res Commun ; 470(1): 144-149, 2016 Jan 29.
Artigo em Inglês | MEDLINE | ID: mdl-26773496

RESUMO

Microphthalmia-associated transcription factor (MITF) is a key regulator of differentiation of melanocytes and retinal pigment epithelial cells, but it also has functions in non-pigment cells. MITF consists of multiple isoforms, including widely expressed MITF-A and MITF-H. In the present study, we explored the potential role played by the Hedgehog signaling on MITF expression in two common types of primary liver cancer, using human cholangiocarcinoma cell lines, the KKU-100 and HuCCT1, along with the HepG2 human hepatocellular carcinoma cell line. Importantly, cholangiocarcinoma is characterized by the activated Hedgehog signaling. Here we show that MITF-A mRNA is predominantly expressed in all three human liver cancer cell lines examined. Moreover, cyclopamine, an inhibitor of the Hedgehog signalling, increased the expression levels of MITF proteins in HuCCT1 and HepG2 cells, but not in KKU-100 cells, suggesting that MITF expression may be down-regulated in some liver cancer cases.


Assuntos
Carcinoma Hepatocelular/metabolismo , Colangiocarcinoma/metabolismo , Proteínas Hedgehog/metabolismo , Neoplasias Hepáticas/metabolismo , Fator de Transcrição Associado à Microftalmia/metabolismo , Alcaloides de Veratrum/administração & dosagem , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Células Hep G2 , Humanos , Transdução de Sinais/efeitos dos fármacos
13.
Pancreas ; 45(3): 370-5, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26390428

RESUMO

OBJECTIVES: In mouse models of pancreatic cancer, IPI-926, an oral Hedgehog inhibitor, increases chemotherapy delivery by depleting tumor-associated stroma. This multicenter phase Ib study evaluated IPI-926 in combination with FOLFIRINOX (5-fluorouracil, leucovorin, irinotecan, oxaliplatin) in patients with advanced pancreatic cancer. METHODS: Patients were treated with once-daily IPI-926 plus FOLFIRINOX. A 3 + 3 dose escalation design was used, with cohort expansion at the maximum tolerated dose. A subset of patients underwent perfusion computed tomography to assess changes in tumor perfusion. RESULTS: The maximum tolerated dose was identified 1 dose level below standard FOLFIRINOX. Common treatment-related adverse events included liver function test abnormalities, neuropathy, nausea/vomiting, and diarrhea. Objective response rate was high (67%), and patients receiving IPI-926 maintenance showed further declines in CA19-9 levels even after FOLFIRINOX discontinuation. Treatment did not result in consistent increases in tumor perfusion. The study closed early when a separate phase II trial of IPI-926 plus gemcitabine indicated detrimental effects of this combination. CONCLUSIONS: This is the first study to demonstrate the feasibility of using FOLFIRINOX as the chemotherapeutic backbone in a clinical trial design. Although robust antitumor activity and acceptable safety were observed with the addition of IPI-926 to this regimen, future development of Hedgehog inhibitors in pancreatic cancer seems unlikely.


Assuntos
Adenocarcinoma/tratamento farmacológico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias Pancreáticas/tratamento farmacológico , Adenocarcinoma/metabolismo , Idoso , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Antígeno CA-19-9/metabolismo , Camptotecina/administração & dosagem , Camptotecina/efeitos adversos , Camptotecina/análogos & derivados , Diarreia/induzido quimicamente , Relação Dose-Resposta a Droga , Estudos de Viabilidade , Feminino , Fluoruracila/administração & dosagem , Fluoruracila/efeitos adversos , Proteínas Hedgehog/metabolismo , Humanos , Irinotecano , Estimativa de Kaplan-Meier , Leucovorina/administração & dosagem , Leucovorina/efeitos adversos , Masculino , Pessoa de Meia-Idade , Náusea/induzido quimicamente , Doenças do Sistema Nervoso/induzido quimicamente , Compostos Organoplatínicos/administração & dosagem , Compostos Organoplatínicos/efeitos adversos , Oxaliplatina , Neoplasias Pancreáticas/metabolismo , Transdução de Sinais/efeitos dos fármacos , Resultado do Tratamento , Alcaloides de Veratrum/administração & dosagem , Alcaloides de Veratrum/efeitos adversos , Alcaloides de Veratrum/farmacocinética , Vômito/induzido quimicamente
14.
Nanoscale ; 7(18): 8607-18, 2015 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-25898852

RESUMO

Cancer stem cells (CSCs) have the ability to transform into bulk cancer cells, to promote tumor growth and establish tumor metastasis. To effectively inhibit tumor growth and prevent metastasis, treatments with conventional chemotherapy drugs should be combined with CSC targeted drugs. In this study, we describe the synthesis and characterization of a new amphiphilic polymer, hyaluronic acid-cystamine-polylactic-co-glycolic acid (HA-SS-PLGA), composed of a hydrophobic PLGA head and a hydrophilic HA segment linked by a bioreducible disulfide bond. With a double emulsion method, a nano delivery system was constructed to deliver doxorubicin (DOX) and cyclopamine (CYC, a primary inhibitor of the hedgehog signaling pathway of CSCs) to both a CD44-overexpressing breast CSC subpopulation and bulk breast cancer cells and allow an on-demand release. The resulting drug-loaded NPs exhibited a redox-responsive drug release profile. Dual drug-loaded particles potently diminished the number and size of tumorspheres and HA showed a targeting effect towards breast CSCs. In vivo combination therapy further demonstrated a remarkable synergistic anti-tumor effect and prolonged survival compared to mono-therapy using the orthotopic mammary fat pad tumor growth model. The co-delivery of drug and the CSC specific inhibitor towards targeted cancer chemotherapeutics provides an insight into anticancer strategy with facile control and high efficacy.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Neoplasias da Mama/tratamento farmacológico , Preparações de Ação Retardada/administração & dosagem , Ácido Hialurônico/química , Nanocápsulas/química , Células-Tronco Neoplásicas/efeitos dos fármacos , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Protocolos de Quimioterapia Combinada Antineoplásica/química , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Preparações de Ação Retardada/química , Difusão , Doxorrubicina/administração & dosagem , Feminino , Interações Hidrofóbicas e Hidrofílicas , Ácido Láctico/química , Células MCF-7 , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Nanocápsulas/ultraestrutura , Células-Tronco Neoplásicas/patologia , Oxirredução , Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Resultado do Tratamento , Alcaloides de Veratrum/administração & dosagem , Alcaloides de Veratrum/química
15.
J Control Release ; 202: 40-8, 2015 Mar 28.
Artigo em Inglês | MEDLINE | ID: mdl-25637565

RESUMO

Cyclopamine (CPA), a potent inhibitor of the Hedgehog pathway, has produced promising anticancer results in a number of preclinical studies. CPA has also been found to enhance tumor response to radiation therapy. However, CPA is water insoluble. A drug delivery system suitable for systemic administration of CPA is needed before CPA can be considered for clinical translation. We hypothesized that CPA solubilized in a liquid-lipid nanoparticle system (CPA-LLP) for intravenous injection would have desirable pharmacokinetic properties and increased anticancer efficacy. We further hypothesized that CPA-LLP would enhance the response of tumor cells to targeted radiotherapy delivered selectively through intratumoral injection of lutetium-177 bound to core-crosslinked polymeric micelles (CCPM-(177)Lu). We tested the combination therapy in 4T1 murine breast cancer and Miapaca-2 human pancreatic adenocarcinoma models. The results showed that CPA-LLP had higher antitumor cytotoxicity than free CPA (IC50 values [mean±SEM]: 2.7±0.2µM vs. 11.3±1.2µM against 4T1 cells; 1.8±0.2 vs. 17.1±1.26µM against Miapaca-2 cells; p<0.0001). In both cell lines, CPA-LLP resulted in significantly lower clonogenicity than free CPA (p<0.05). Moreover, in both cell lines, CPA-LLP significantly enhanced the cell response to CCPM-(177)Lu radiotherapy as measured by clonogenic assay (p<0.05). In 4T1 and Miapaca-2 mouse xenograft models, the combination of CPA-LLP and CCPM-(177)Lu delayed tumor growth more than either monotherapy did alone. In the 4T1 tumor model, tumor size at 16days after treatment was significantly smaller with the combination therapy than with all the other treatments. In the Miapaca-2 model, the combination therapy resulted in the highest rate of mouse survival and prevented tumor relapse. In conclusion, the combination of CPA-LLP and CCPM-(177)Lu was an effective strategy for treating breast and pancreatic cancer and deserves further investigation.


Assuntos
Antineoplásicos/administração & dosagem , Lutécio/administração & dosagem , Neoplasias Mamárias Experimentais/terapia , Nanopartículas/administração & dosagem , Neoplasias Pancreáticas/terapia , Radioisótopos/administração & dosagem , Alcaloides de Veratrum/administração & dosagem , Animais , Antineoplásicos/farmacocinética , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Quimiorradioterapia , Terapia Combinada , Feminino , Humanos , Lutécio/farmacocinética , Lutécio/uso terapêutico , Neoplasias Mamárias Experimentais/metabolismo , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Radioisótopos/farmacocinética , Radioisótopos/uso terapêutico , Distribuição Tecidual , Carga Tumoral/efeitos dos fármacos , Alcaloides de Veratrum/farmacocinética , Alcaloides de Veratrum/uso terapêutico
16.
J Chromatogr Sci ; 53(7): 1092-9, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25547283

RESUMO

Veratramine (VAM) is the major lipid-soluble alkaloid existing in Veratrum nigrum L. that has been demonstrated to exert neurotoxic effects. To better understand the potential mechanism of neurotoxicity of VAM, VAM-induced DNA damage was measured in the cerebellum and cerebral cortex of mice after a 7-day repetitive oral dose by using single-cell gel electrophoresis (comet assay). A method based on high-performance liquid chromatography-electrospray ionization tandem mass spectrometry was developed for the determination of VAM and its in vivo and in vitro metabolites, to establish the potential correlation between metabolites and neurotoxicity. In vitro experiment was carried out using rat liver microsomes, whereas the in vivo study was conducted on rats at a single dose of 3 mg/kg. The results showed that VAM caused DNA damage in the cerebellum and cerebral cortex of mice in a dose-dependent manner. Phenyl mono-oxidation, sulfate conjugation and phenyl di-oxidation were proposed to be the main in vivo metabolic pathways of VAM, whereas the major in vitro metabolic pathways were phenyl mono-oxidation, hydroxylation and methylation. Phenyl-oxidation reaction was likely to be associated with reactive oxygen species production, leading to the DNA damage in the mouse brain.


Assuntos
Encéfalo/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Alcaloides de Veratrum/metabolismo , Alcaloides de Veratrum/toxicidade , Animais , Encéfalo/metabolismo , Cromatografia Líquida de Alta Pressão/métodos , Masculino , Camundongos , Microssomos Hepáticos/efeitos dos fármacos , Microssomos Hepáticos/metabolismo , Ratos , Espectrometria de Massas por Ionização por Electrospray/métodos , Veratrum/química , Alcaloides de Veratrum/administração & dosagem , Alcaloides de Veratrum/isolamento & purificação
17.
AAPS PharmSciTech ; 16(1): 98-107, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25190362

RESUMO

The relapse of cancer after first line therapy with anticancer agents is a common occurrence. This recurrence is believed to be due to the presence of a subpopulation of cells called cancer stem cells in the tumor. Therefore, a combination therapy which is susceptible to both types of cells is desirable. Delivery of this combinatorial approach in a nanoparticulate system will provide even a better therapeutic outcome in tumor targeting. The objective of this study was to develop and characterize nanoparticulate system containing two anticancer agents (cyclopamine and paclitaxel) having different susceptibilities toward cancer cells. Both drugs were entrapped in glyceryl monooleate (GMO)-chitosan solid lipid as well as poly(glycolic-lactic) acid (PLGA) nanoparticles. The cytotoxicity studies were performed on DU145, DU145 TXR, and Wi26 A4 cells. The particle size of drug-loaded GMO-chitosan nanoparticles was 278.4 ± 16.4 nm with a positive zeta potential. However, the PLGA particles were 234.5 ± 6.8 nm in size with a negative zeta potential. Thermal analyses of both nanoparticles revealed that the drugs were present in noncrystalline state in the matrix. A sustained in vitro release was observed for both the drugs in these nanoparticles. PLGA blank particles showed no cytotoxicity in all the cell lines tested, whereas GMO-chitosan blank particles showed substantial cytotoxicity. The types of polymer used for the preparation of nanoparticles played a major role and affected the in vitro release, cytotoxicity, and uptake of nanoparticles in the all the cell lines tested.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Preparações de Ação Retardada/administração & dosagem , Preparações de Ação Retardada/síntese química , Nanocápsulas/química , Células-Tronco Neoplásicas/efeitos dos fármacos , Neoplasias da Próstata/tratamento farmacológico , Protocolos de Quimioterapia Combinada Antineoplásica/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Difusão , Composição de Medicamentos/métodos , Estabilidade de Medicamentos , Humanos , Masculino , Nanocápsulas/administração & dosagem , Nanocápsulas/ultraestrutura , Células-Tronco Neoplásicas/patologia , Paclitaxel/administração & dosagem , Paclitaxel/química , Tamanho da Partícula , Neoplasias da Próstata/patologia , Resultado do Tratamento , Alcaloides de Veratrum/administração & dosagem , Alcaloides de Veratrum/química
18.
Oncotarget ; 5(22): 11681-94, 2014 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-25373737

RESUMO

Glioblastoma are highly aggressive brain tumors with poor prognosis. While various dysregulation of signaling pathways in gliomas have been described, the identification of biomarkers and therapy targets remains an important task for novel diagnostic and therapeutic approaches. Here we described that the Suppressor of fused (also known as Sufu) is significantly down-regulated in high-grade gliomas, correlating with a poor prognosis. We demonstrated that ectopic expression of Sufu inhibited cell proliferation, invasion and vasculogenic mimicry. In addition, overexpression of Sufu reduced Gli reporter gene transcription activity and prevented Gli1 nuclear accumulation, whereas knockdown of Sufu reversed these effects. Furthermore, overexpressed Sufu sensitized glioblastoma to Temozolomide and Cyclopamine. Thus, Sufu is potential tumor suppressor and therapeutic target in glioblastoma.


Assuntos
Neoplasias Encefálicas/metabolismo , Núcleo Celular/metabolismo , Glioma/metabolismo , Proteínas Repressoras/metabolismo , Fatores de Transcrição/metabolismo , Adulto , Animais , Linhagem Celular Tumoral , Proliferação de Células , Dacarbazina/administração & dosagem , Dacarbazina/análogos & derivados , Progressão da Doença , Regulação para Baixo , Feminino , Glioma/tratamento farmacológico , Humanos , Masculino , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Invasividade Neoplásica , Transplante de Neoplasias , Prognóstico , RNA Interferente Pequeno/metabolismo , Temozolomida , Resultado do Tratamento , Alcaloides de Veratrum/administração & dosagem , Proteína GLI1 em Dedos de Zinco
19.
Mol Cancer Ther ; 13(5): 1259-69, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24634412

RESUMO

Hedgehog (Hh) pathway inhibition in cancer has been evaluated in both the ligand-independent and ligand-dependent settings, where Hh signaling occurs either directly within the cancer cells or within the nonmalignant cells of the tumor microenvironment. Chondrosarcoma is a malignant tumor of cartilage in which there is ligand-dependent activation of Hh signaling. IPI-926 is a potent, orally delivered small molecule that inhibits Hh pathway signaling by binding to Smoothened (SMO). Here, the impact of Hh pathway inhibition on primary chondrosarcoma xenografts was assessed. Mice bearing primary human chondrosarcoma xenografts were treated with IPI-926. The expression levels of known Hh pathway genes, in both the tumor and stroma, and endpoint tumor volumes were measured. Gene expression profiling of tumors from IPI-926-treated mice was conducted to identify potential novel Hh target genes. Hh target genes were studied to determine their contribution to the chondrosarcoma neoplastic phenotype. IPI-926 administration results in downmodulation of the Hh pathway in primary chondrosarcoma xenografts, as demonstrated by evaluation of the Hh target genes GLI1 and PTCH1, as well as inhibition of tumor growth. Chondrosarcomas exhibited autocrine and paracrine Hh signaling, and both were affected by IPI-926. Decreased tumor growth is accompanied by histopathologic changes, including calcification and loss of tumor cells. Gene profiling studies identified genes differentially expressed in chondrosarcomas following IPI-926 treatment, one of which, ADAMTSL1, regulates chondrosarcoma cell proliferation. These studies provide further insight into the role of the Hh pathway in chondrosarcoma and provide a scientific rationale for targeting the Hh pathway in chondrosarcoma.


Assuntos
Condrossarcoma/metabolismo , Proteínas Hedgehog/metabolismo , Receptores Acoplados a Proteínas-G/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Alcaloides de Veratrum/farmacologia , Proteínas ADAMTS , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/farmacologia , Calcinose/tratamento farmacológico , Calcinose/genética , Calcinose/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Condrossarcoma/genética , Condrossarcoma/patologia , Modelos Animais de Doenças , Proteínas da Matriz Extracelular/genética , Humanos , Camundongos , Receptor Smoothened , Carga Tumoral/efeitos dos fármacos , Alcaloides de Veratrum/administração & dosagem , Ensaios Antitumorais Modelo de Xenoenxerto
20.
Cancer ; 120(4): 537-47, 2014 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-24151134

RESUMO

BACKGROUND: During development, the Hedgehog pathway plays important roles regulating the proliferation and differentiation of chondrocytes, providing a template for growing bone. In this study, the authors investigated the components of dysregulated Hedgehog signaling as potential therapeutic targets for osteosarcoma. METHODS: Small-molecule agonists and antagonists that modulate the Hedgehog pathway at different levels were used to investigate the mechanisms of dysregulation and the efficacy of Hedgehog blockade in osteosarcoma cell lines. The inhibitory effect of a small-molecule Smoothened (SMO) antagonist, IPI-926 (saridegib), also was examined in patient-derived xenograft models. RESULTS: An inverse correlation was identified in osteosarcoma cell lines between endogenous glioma-associated oncogene 2 (GLI2) levels and Hedgehog pathway induction levels. Cells with high levels of GLI2 were sensitive to GLI inhibition, but not SMO inhibition, suggesting that GLI2 overexpression may be a mechanism of ligand-independent activation. In contrast, cells that expressed high levels of the Hedgehog ligand gene Indian hedgehog (IHH) and the target genes patched 1 (PTCH1) and GLI1 were sensitive to modulation of both SMO and GLI, suggesting ligand-dependent activation. In 2 xenograft models, active autocrine and paracrine, ligand-dependent Hedgehog signaling was identified. IPI-926 inhibited the Hedgehog signaling interactions between the tumor and the stroma and demonstrated antitumor efficacy in 1 of 2 ligand-dependent models. CONCLUSIONS: The current results indicate that both ligand-dependent and ligand-independent Hedgehog dysregulation may be involved in osteosarcoma. It is the first report to demonstrate Hedgehog signaling crosstalk between the tumor and the stroma in osteosarcoma. The inhibitory effect of IPI-926 warrants additional research and raises the possibility of using Hedgehog pathway inhibitors as targeted therapeutics to improve treatment for osteosarcoma.


Assuntos
Neoplasias Ósseas/genética , Osteossarcoma/tratamento farmacológico , Osteossarcoma/genética , Fatores de Transcrição/genética , Adolescente , Adulto , Antineoplásicos/administração & dosagem , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Criança , Feminino , Proteínas Hedgehog/genética , Proteínas Hedgehog/metabolismo , Humanos , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Masculino , Terapia de Alvo Molecular , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Osteossarcoma/etiologia , Osteossarcoma/patologia , Receptores Acoplados a Proteínas-G/genética , Receptores Acoplados a Proteínas-G/metabolismo , Transdução de Sinais/efeitos dos fármacos , Receptor Smoothened , Fatores de Transcrição/metabolismo , Alcaloides de Veratrum/administração & dosagem , Ensaios Antitumorais Modelo de Xenoenxerto , Proteína GLI1 em Dedos de Zinco , Proteína Gli2 com Dedos de Zinco
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