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1.
ChemMedChem ; 10(7): 1149-52, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25951302

RESUMO

Inhibition of adenosine A2A receptors has been shown to elicit a therapeutic response in preclinical animal models of Parkinson's disease (PD). We previously identified the triazolo-9H-purine, ST1535, as a potent A(2A)R antagonist. Studies revealed that ST1535 is extensively hydroxylated at the ω-1 position of the butyl side chain. Here, we describe the synthesis and evaluation of derivatives in which the ω-1 position has been substituted (F, Me, OH) in order to block metabolism. The stability of the compounds was evaluated in human liver microsomes (HLM), and the affinity for A(2A)R was determined. Two compounds, (2-(3,3-dimethylbutyl)-9-methyl-8-(2H-1,2,3-triazol-2-yl)-9H-purin-6-amine (3 b) and 4-(6-amino-9-methyl-8-(2H-1,2,3-triazol-2-yl)-9H-purin-2-yl)-2-methylbutan-2-ol (3 c), exhibited good affinity against A(2A)R (Ki =0.4 nM and 2 nM, respectively) and high in vitro metabolic stability (89.5% and 95.3% recovery, respectively, after incubation with HLM for two hours).


Assuntos
Adenosina/análogos & derivados , Receptor A2A de Adenosina/metabolismo , Triazóis/metabolismo , Adenosina/química , Adenosina/metabolismo , Relação Dose-Resposta a Droga , Humanos , Ligantes , Microssomos Hepáticos/química , Microssomos Hepáticos/metabolismo , Estrutura Molecular , Relação Estrutura-Atividade , Triazóis/química
2.
Eur J Pharmacol ; 761: 353-61, 2015 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-25936513

RESUMO

Antagonism of the adenosine A2A receptor represents a promising strategy for non-dopaminergic treatment of Parkinson׳s disease (PD). Previously, the adenosine A2A receptor antagonist ST1535 was shown to possess potential beneficial effects in animal models of PD. Two metabolites of ST1535, namely ST3932 and ST4206, were tested in vitro to assess their affinity and activity on cloned human A2A adenosine receptors, and their metabolic profile. Additionally, ST3932 and ST4206 were investigated in vivo in animal models of PD following oral/intraperitoneal administration of 10, 20 and 40mg/kg using ST1535 as a reference compound. ST3932 and ST4206 displayed high affinity and antagonist behaviour for cloned human adenosine A2A receptors. The Ki values for ST1535, ST3932 and ST4206 were 8, 8 and 12nM, respectively, and their IC50 values on cyclic AMP were 427, 450 and 990nM, respectively. ST1535, ST3932 and ST4206 antagonized (orally) haloperidol-induced catalepsy in mice, potentiated (intraperitoneally) the number of contralateral rotations induced by l-3,4-dihydroxyphenylalanine (l-DOPA) (3mg/kg) plus benserazide (6mg/kg) in 6-Hydroxydopamine hydrobromide (6-OHDA)-lesioned rats, and increased mouse motor activity by oral route. Thus, ST3932 and ST4206, two ST1535 metabolites, show a pharmacological activity similar to ST1535, both in vitro and in vivo, and may be regarded as an interesting pharmacological alternative to ST1535.


Assuntos
Adenina/análogos & derivados , Antagonistas do Receptor A2 de Adenosina/farmacologia , Antiparkinsonianos/farmacologia , Gânglios da Base/efeitos dos fármacos , Atividade Motora/efeitos dos fármacos , Transtornos Parkinsonianos/tratamento farmacológico , Receptor A2A de Adenosina/efeitos dos fármacos , Triazóis/farmacologia , Adenina/administração & dosagem , Adenina/metabolismo , Adenina/farmacologia , Antagonistas do Receptor A2 de Adenosina/administração & dosagem , Antagonistas do Receptor A2 de Adenosina/metabolismo , Administração Oral , Animais , Antiparkinsonianos/administração & dosagem , Antiparkinsonianos/metabolismo , Gânglios da Base/metabolismo , Gânglios da Base/fisiopatologia , Ligação Competitiva , Catalepsia/induzido quimicamente , Catalepsia/prevenção & controle , AMP Cíclico/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Células HEK293 , Haloperidol , Humanos , Injeções Intraperitoneais , Ligantes , Masculino , Camundongos , Oxidopamina , Transtornos Parkinsonianos/induzido quimicamente , Transtornos Parkinsonianos/metabolismo , Transtornos Parkinsonianos/fisiopatologia , Ligação Proteica , Ratos Sprague-Dawley , Receptor A2A de Adenosina/genética , Receptor A2A de Adenosina/metabolismo , Transfecção , Triazóis/administração & dosagem , Triazóis/metabolismo
3.
Eur J Med Chem ; 80: 8-35, 2014 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-24763360

RESUMO

Many known 5-HT7 ligands contain either a serotonin-like or an arylpiperazine structure that, in published SAR studies, are generally supposed to bind the same receptor pocket. Conversely, we explored the hypothesis that two such moieties can co-exist in the same ligand, binding to different pockets. We thus designed and synthesized a set of compounds including both a 5-hydroxyindol-3-ylethyl and a 1-arylpiperazine moieties connected by a short linker. The compounds were tested for their affinity for human 5-HT7 serotonin receptor. We further prepared a novel series of 5-HT7 ligands, where the 5-hydroxyindol-3-ylethyl moiety was bioisosterically replaced by a 3-hydroxyanilinoalkyl one. Among the newly synthesized compounds, potent ligands at the 5-HT7 receptor, behaving as antagonists in functional tests, were identified, even if they showed limited subtype selectivity. Docking studies within a model of the 5-HT7 receptor showed that the binding site can actually accommodate both moieties, with the serotonin-like one in the putative orthosteric site and the arylpiperazine one occupying an accessory pocket. The present results demonstrate that it is possible to devise and develop new 5-HT7 ligands merging two privileged structures in the same molecule.


Assuntos
Materiais Biomiméticos/química , Materiais Biomiméticos/metabolismo , Desenho de Fármacos , Piperazinas/química , Receptores de Serotonina/metabolismo , Serotonina/química , Materiais Biomiméticos/síntese química , Humanos , Ligantes , Simulação de Acoplamento Molecular , Conformação Proteica , Receptores de Serotonina/química , Relação Estrutura-Atividade
4.
J Med Chem ; 56(13): 5456-63, 2013 Jul 11.
Artigo em Inglês | MEDLINE | ID: mdl-23789814

RESUMO

The synthesis and preliminary in vitro evaluation of five metabolites of the A2A antagonist ST1535 (1) are reported. The metabolites, originating in vivo from enzymatic oxidation of the 2-butyl group of the parent compound, were synthesized from 6-chloro-2-iodo-9-methyl-9H-purine (2) by selective C-C bond formation via halogen/magnesium exchange reaction and/or palladium-catalyzed reactions. The metabolites behaved in vitro as antagonist ligands of cloned human A2A receptor with affinities (Ki 7.5-53 nM) comparable to that of compound 1 (Ki 10.7 nM), thus showing that the long duration of action of 1 could be in part due to its metabolites. General behavior after oral administration in mice was also analyzed.


Assuntos
Adenina/análogos & derivados , Antagonistas do Receptor A2 de Adenosina/farmacologia , Doença de Parkinson/prevenção & controle , Receptor A2A de Adenosina/metabolismo , Triazóis/farmacologia , Adenina/síntese química , Adenina/metabolismo , Adenina/farmacologia , Antagonistas do Receptor A2 de Adenosina/síntese química , Antagonistas do Receptor A2 de Adenosina/metabolismo , Animais , Comportamento Animal/efeitos dos fármacos , Ligação Competitiva , Células CHO , Cricetinae , Cricetulus , Células HEK293 , Humanos , Camundongos , Modelos Químicos , Estrutura Molecular , Doença de Parkinson/metabolismo , Ensaio Radioligante , Receptor A2A de Adenosina/genética , Fatores de Tempo , Triazóis/síntese química , Triazóis/metabolismo
5.
J Med Chem ; 56(3): 1247-61, 2013 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-23281824

RESUMO

A systematic modification of the caffeinyl core and substituents of the reference compound (E)-8-(3-chlorostyryl)caffeine led to the 9-deazaxanthine derivative (E)-6-(4-chlorostyryl)-1,3,5,-trimethyl-1H-pyrrolo[3,2-d]pyrimidine-2,4-(3H,5H)-dione (17f), which acts as a dual human A(2a) antagonist/MAO-B inhibitor (K(i)(A(2A)) = 260 nM; IC(50)(MAO-B) = 200 nM; IC(50)(MAO-A) = 10 µM) and dose dependently counteracts haloperidol-induced catalepsy in mice from 30 mg/kg by the oral route. The compound is the best balanced A(2A) antagonist/MAO-B inhibitor reported to date, and it could be considered as a new lead in the field of anti-Parkinson's agents. A number of analogues of 17f were synthesized and qualitative SARs are discussed. Two analogues of 17f, namely 18b and 19a, inhibit MAO-B with IC(50) of 68 and 48 nM, respectively, being 5-7-fold more potent than the prototypical MAO-B inhibitor deprenyl (IC(50) = 334 nM).


Assuntos
Cafeína/análogos & derivados , Inibidores da Monoaminoxidase/síntese química , Inibidores da Monoaminoxidase/farmacologia , Xantinas/química , Cafeína/síntese química , Cafeína/química , Cafeína/farmacologia , Células HEK293 , Humanos , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Simulação de Acoplamento Molecular , Inibidores da Monoaminoxidase/química
6.
Eur J Pharmacol ; 661(1-3): 8-14, 2011 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-21549693

RESUMO

5-HT(6) receptor is one of the most recently cloned serotonin receptors, and it might play important roles in Alzheimer's disease, depression, and learning and memory disorders. Availability of only very few 5-HT(6) receptor agonists, however, does not allow examining their contribution in psychopharmacological processes. Therefore, a new 5-HT(6) receptor agonist, ST1936, was synthesized. ST1936 binds to human 5-HT(6) receptors with good affinity (K(i)=28.8 nM). ST1936 also exhibited some moderate binding affinity for 5HT(2B), 5HT(1A), 5HT(7) receptors and adrenergic α receptors. ST1936 behaved as a full 5-HT(6) agonist on cloned cells and was able to increase Ca(2+) concentration, phosphorylation of Fyn kinase, and regulate the activation of ERK1/2 that is a downstream target of Fyn kinase. These effects were completely antagonized by two 5-HT(6) receptor antagonists, SB271046 and SB258585. The other 5-HT(6) receptor agonist, WAY181187 also increased Fyn kinase activity. These results suggest that both ST1936 and WAY181187 mediate 5-HT(6) receptor-dependent signal pathways, such as cAMP, Fyn and ERK1/2 kinase, as specific agonists.


Assuntos
Cálcio/metabolismo , AMP Cíclico/metabolismo , Etilaminas/farmacologia , Indóis/farmacologia , Proteínas Quinases/metabolismo , Receptores de Serotonina/genética , Receptores de Serotonina/metabolismo , Agonistas do Receptor de Serotonina/farmacologia , Linhagem Celular Tumoral , Clonagem Molecular , Ativação Enzimática/efeitos dos fármacos , Etilaminas/metabolismo , Humanos , Indóis/metabolismo , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-fyn/metabolismo , Transdução de Sinais/efeitos dos fármacos , Tiazóis/farmacologia , Triptaminas/farmacologia
7.
Pharmacol Biochem Behav ; 98(2): 169-72, 2011 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-21192969

RESUMO

5-HT(6) receptors are relatively recently-discovered receptors. After an uncertain beginning, where results were ambiguous, findings are now apparently more congruent. Nevertheless, discrepancies still exist. The aim of the present manuscript is to point out some of these discrepancies, in order to reflect on the current status of the field of the 5-HT(6) receptor neuropharmacology, and where the field should move next. Examples of 5-HT(6) receptor ligand-induced changes in behavior, neurochemistry and binding highlight areas where discrepancies remain and further experimental attention is needed. Possible methodological as well as conceptual issues underlying the inconsistencies are considered in an effort to increase awareness of the need for more thorough consideration of these aspects in future research.


Assuntos
Receptores de Serotonina/efeitos dos fármacos , Receptores de Serotonina/fisiologia , Antagonistas da Serotonina/farmacologia , Animais , Comportamento Animal/efeitos dos fármacos , Comportamento Animal/fisiologia , Encéfalo/efeitos dos fármacos , Encéfalo/fisiologia , Humanos , Ligantes , Antagonistas da Serotonina/farmacocinética , Agonistas do Receptor de Serotonina/farmacologia
9.
Cancer Res ; 70(5): 1804-13, 2010 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-20145150

RESUMO

Angiogenesis is one of the crucial events for cancer development and growth. Two members of the vascular endothelial growth factor (VEGF) family, VEGF-A and placental growth factor (PlGF), which are able to heterodimerize if coexpressed in the same cell, are both required for pathologic angiogenesis. We have generated a PlGF1 variant, named PlGF1-DE in which the residues Asp72 and Glu73 were substituted with Ala, which is unable to bind and activate VEGF receptor-1 but is still able to heterodimerize with VEGF. Here, we show that overexpression in tumor cells by adenoviral delivery or stable transfection of PlGF1-DE variant significantly reduces the production of VEGF homodimer via heterodimerization, determining a strong inhibition of xenograft tumor growth and neoangiogenesis, as well as significant reduction of vessel lumen and stabilization, and monocyte-macrophage infiltration. Conversely, the overexpression of PlGF1wt, also reducing the VEGF homodimer production comparably with PlGF1-DE variant through the generation of VEGF/PlGF heterodimer, does not inhibit tumor growth and vessel density compared with controls but induces increase of vessel lumen, vessel stabilization, and monocyte-macrophage infiltration. The property of PlGF and VEGF-A to generate heterodimer represents a successful strategy to inhibit VEGF-dependent angiogenesis. The PlGF1-DE variant, and not PlGF1wt as previously reported, acts as a "dominant negative" of VEGF and is a new candidate for antiangiogenic gene therapy in cancer treatment.


Assuntos
Neoplasias Pulmonares/irrigação sanguínea , Proteínas de Membrana/metabolismo , Neoplasias Ovarianas/irrigação sanguínea , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Animais , Linhagem Celular Tumoral , Dimerização , Feminino , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/terapia , Proteínas de Membrana/biossíntese , Proteínas de Membrana/genética , Camundongos , Camundongos Nus , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Neovascularização Patológica/terapia , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/terapia , Ligação Proteica , Transfecção , Fator A de Crescimento do Endotélio Vascular/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Arterioscler Thromb Vasc Biol ; 30(3): 426-35, 2010 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-20056909

RESUMO

OBJECTIVE: The beneficial effect of the natural compound propionyl-l-carnitine (PLC) on intermittent claudication in patients with peripheral arterial disease is attributed to its anaplerotic function in ischemic tissues, but inadequate information is available concerning action on the vasculature. METHODS AND RESULTS: We investigated the effects of PLC in rabbit hind limb collateral vessels after femoral artery excision, mouse dorsal air pouch, chicken chorioallantoic membrane, and vascular cells by angiographic, Doppler flow, and histomorphometrical and biomolecular analyses. PLC injection accelerated hind limb blood flow recovery after 4 days (P<0.05) and increased angiographic quadriceps collateral vascularization after 7 days (P<0.001) Histomorphometry confirmed the increased vascular area (P<0.05), with unchanged intramuscular capillary density. PLC-induced dilatative adaptation, and growth was found associated with increased inducible nitric oxide synthase and reduced arterial vascular endothelial growth factor and intracellular adhesion molecule-1 expression. PLC also increased vascularization in air pouch and chorioallantoic membrane (P<0.05), particularly in large vessels. PLC increased endothelial and human umbilical vascular endothelial cell proliferation and rapidly reduced inducible nitric oxide synthase and NADPH-oxidase 4-mediated reactive oxygen species production in human umbilical vascular endothelial cells; NADPH-oxidase 4 also regulated NF-kappaB-independent intracellular adhesion molecule-1 expression. CONCLUSIONS: Our results provided strong evidence that PLC improves postischemic flow recovery and revascularization and reduces endothelial NADPH-oxidase-related superoxide production. We recommend that PLC should be included among therapeutic interventions that target endothelial function.


Assuntos
Vasos Sanguíneos/fisiologia , Carnitina/análogos & derivados , Endotélio Vascular/metabolismo , NADPH Oxidases/metabolismo , Neovascularização Fisiológica/efeitos dos fármacos , Fluxo Sanguíneo Regional/efeitos dos fármacos , Superóxidos/metabolismo , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Carnitina/farmacologia , Proliferação de Células/efeitos dos fármacos , Embrião de Galinha , Membrana Corioalantoide/irrigação sanguínea , Circulação Colateral/efeitos dos fármacos , Modelos Animais de Doenças , Endotélio Vascular/efeitos dos fármacos , Membro Posterior/irrigação sanguínea , Humanos , Molécula 1 de Adesão Intercelular/metabolismo , Isquemia/fisiopatologia , Camundongos , NADPH Oxidase 4 , Neovascularização Fisiológica/fisiologia , Coelhos , Fluxo Sanguíneo Regional/fisiologia , Fator A de Crescimento do Endotélio Vascular/metabolismo
11.
Front Psychiatry ; 1: 22, 2010.
Artigo em Inglês | MEDLINE | ID: mdl-21423433

RESUMO

Adenosine A(2A) receptors seem to exist in typical (more in striatum) and atypical (more in hippocampus and cortex) subtypes. In the present study, we investigated the affinity of two adenosine A(2A) receptor antagonists, ST1535 [2 butyl -9-methyl-8-(2H-1,2,3-triazol 2-yl)-9H-purin-6-xylamine] and KW6002 [(E)-1,3-diethyl-8-(3,4-dimethoxystyryl)-7-methyl-3,7-dihydro-1H-purine-2,6,dione] to the "typical" and "atypical" A(2A) binding sites. Affinity was determined by radioligand competition experiments in membranes from rat striatum and hippocampus. Displacement of the adenosine analog [(3)H]CGS21680 [2-p-(2-carboxyethyl)phenethyl-amino-5'-N-ethylcarbox-amidoadenosine] was evaluated in the absence or in the presence of either CSC [8-(3-chlorostyryl)-caffeine], an adenosine A(2A) antagonist that pharmacologically isolates atypical binding sites, or DPCPX (8-cyclopentyl-1,3-dipropylxanthine), an adenosine A(1) receptor antagonist that pharmacologically isolates typical binding site. ZM241385 [84-(2-[7-amino-2-(2-furyl) [1,2,4]-triazol[2,3-a][1,3,5]triazin-5-yl amino]ethyl) phenol)] and SCH58261 [(5-amino-7-(ß-phenylethyl)-2-(8-furyl)pyrazolo(4,3-e)-1,2,4-triazolo(1,5-c) pyrimidine], two other adenosine A(2A) receptor antagonists, which were reported to differently bind to atypical and typical A(2A) receptors, were used as reference compounds. ST1535, KW6002, ZM241385 and SCH58261 displaced [(3)H]CGS21680 with higher affinity in striatum than in hippocampus. In hippocampus, no typical adenosine A(2A) binding was detected, and ST1535 was the only compound that occupied atypical A(2A) adenosine receptors. Present data are explained in terms of heteromeric association among adenosine A(2A), A(2B) and A(1) receptors, rather than with the presence of atypical A(2A) receptor subtype.

12.
J Med Chem ; 51(9): 2708-21, 2008 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-18396857

RESUMO

With the aim of understanding the influence of fluorine on the double bond of the cis-stilbene moiety of combretastatin derivatives and encouraged by a preliminary molecular modeling study showing a different biological environment on the interaction site with tubulin, we prepared, through various synthetic approaches, a small library of compounds in which one or both of the olefinic hydrogens were replaced with fluorine. X-ray analysis on the difluoro-CA-4 analogue demonstrated that the spatial arrangement of the molecule was not modified, compared to its nonfluorinated counterpart. SAR analysis confirmed the importance of the cis-stereochemistry of the stilbene scaffold. Nevertheless, some unpredicted results were observed on a few trans-fluorinated derivatives. The position of a fluorine atom on the double bond may affect the inhibition of tubulin polymerization and cytotoxic activity of these compounds.


Assuntos
Antineoplásicos/síntese química , Bibenzilas/síntese química , Flúor , Estilbenos/síntese química , Animais , Antineoplásicos/química , Antineoplásicos/farmacologia , Bibenzilas/química , Bibenzilas/farmacologia , Biopolímeros , Bovinos , Linhagem Celular Tumoral , Células Cultivadas , Cristalografia por Raios X , Ensaios de Seleção de Medicamentos Antitumorais , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , Humanos , Microcirculação/citologia , Microtúbulos/efeitos dos fármacos , Microtúbulos/metabolismo , Microtúbulos/ultraestrutura , Estereoisomerismo , Estilbenos/química , Estilbenos/farmacologia , Relação Estrutura-Atividade , Tubulina (Proteína)/metabolismo
13.
Am J Pathol ; 169(2): 643-54, 2006 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-16877362

RESUMO

Placenta growth factor (PlGF), a member of the vascular endothelial growth factor family, plays an important role in adult pathological angiogenesis. To further investigate PlGF functions in tumor growth and metastasis formation, we used transgenic mice overexpressing PlGF in the skin under the control of the keratin 14 promoter. These animals showed a hypervascularized phenotype of the skin and increased levels of circulating PlGF with respect to their wild-type littermates. Transgenic mice and controls were inoculated intradermally with B16-BL6 melanoma cells. The tumor growth rate was fivefold increased in transgenic animals compared to wild-type mice, in the presence of a similar percentage of tumor necrotic tissue. Tumor vessel area was increased in transgenic mice as compared to controls. Augmented mobilization of endothelial and hematopoietic stem cells from the bone marrow was observed in transgenic animals, possibly contributing to tumor vascularization. The number and size of pulmonary metastases were significantly higher in transgenic mice compared to wild-type littermates. Finally, PlGF promoted tumor cell invasion of the extracellular matrix and increased the activity of selected matrix metalloproteinases. These findings indicate that PlGF, in addition to enhancing tumor angiogenesis and favoring tumor growth, may directly influence melanoma dissemination.


Assuntos
Melanoma/patologia , Metástase Neoplásica/patologia , Proteínas da Gravidez/genética , Animais , Movimento Celular , Células Cultivadas , Relação Dose-Resposta a Droga , Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Células-Tronco Hematopoéticas/citologia , Humanos , Metaloproteinase 1 da Matriz/genética , Metaloproteinase 9 da Matriz/genética , Melanoma/irrigação sanguínea , Melanoma/genética , Camundongos , Camundongos Transgênicos , Necrose , Invasividade Neoplásica , Neovascularização Patológica , Fator de Crescimento Placentário , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Células Tumorais Cultivadas
14.
J Med Chem ; 49(11): 3143-52, 2006 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-16722633

RESUMO

We studied the anticancer activity of a series of new combretastatin derivatives with B-ring modifications. The structure-activity relationship (SAR) information confirmed the importance of cis-stereochemistry and of a phenolic moiety in B-ring. We selected the benzo[b]thiophene and benzofuran combretastatin analogues 11 (ST2151) and 13 (ST2179) and their phosphate prodrugs (29 and 30) for their high antitumor activity in in vitro and in vivo models. Cell exposure to IC50 of 11, 13, and CA-4 led to the arrest of various cell types in the G2/M phase of the cell cycle and induction of apoptosis. Mainly, 11 and 13 induced the formation of multinucleated cells with abnormal chromatin distribution, with only a minimal effect on the microtubule organization, with respect to CA-4. Interestingly, both the pharmacokinetic profile of 29 and its in vivo antitumor effect and those of 30, active even after oral administration, suggest additional pharmacological differences between these compounds and CA-4P.


Assuntos
Antineoplásicos/síntese química , Bibenzilas/síntese química , Estilbenos/síntese química , Animais , Antineoplásicos/farmacocinética , Antineoplásicos/farmacologia , Benzofuranos/síntese química , Benzofuranos/farmacocinética , Benzofuranos/farmacologia , Bibenzilas/farmacocinética , Bibenzilas/farmacologia , Ligação Competitiva , Biopolímeros , Linhagem Celular Tumoral , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/ultraestrutura , Colchicina/química , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/ultraestrutura , Endotélio Vascular/citologia , Feminino , Humanos , Camundongos , Camundongos Nus , Microtúbulos/efeitos dos fármacos , Microtúbulos/ultraestrutura , Organofosfatos/síntese química , Organofosfatos/farmacocinética , Organofosfatos/farmacologia , Pró-Fármacos/síntese química , Pró-Fármacos/farmacocinética , Pró-Fármacos/farmacologia , Estereoisomerismo , Estilbenos/farmacocinética , Estilbenos/farmacologia , Relação Estrutura-Atividade , Tiofenos/síntese química , Tiofenos/farmacocinética , Tiofenos/farmacologia , Tubulina (Proteína)/química , Tubulina (Proteína)/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Bioorg Med Chem ; 14(1): 169-80, 2006 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-16214345

RESUMO

A small library of cyclic RGD pentapeptide mimics incorporating stereoisomeric 5,6- and 5,7-fused bicyclic lactams was synthesized. This library was found to contain high-affinity ligands for the alpha(v)beta3 integrin. The aim of this study was to investigate activity, selectivity, and structure of these ligands in order to identify new specific alpha(v)-integrin antagonists that could be evaluated as tumor angiogenesis inhibitors. In vitro screening, including receptor-binding assays to purified alpha(v)beta3, alpha(v)beta5, and alpha5beta1 integrins, and platelet aggregation assay, revealed ST1646 as a potent, highly selective alpha(v)beta3/alpha(v)beta5 integrin antagonist. Structure determination of the cyclic RGD pentapeptide mimics performed by a combination of NMR spectroscopy, and molecular mechanics and dynamics calculations showed a strong dependence of the RGD cyclopeptide conformation on lactam ring size and stereochemistry. ST1646 revealed the highest ability within the library to adopt the proper RGD orientation required for binding to the alpha(v)beta3 integrin, as deduced from the recently solved crystal structure of the extracellular segment of integrin alpha(v)beta3 in complex with a cyclic pentapeptide ligand.


Assuntos
Aminoácidos/química , Integrinas/efeitos dos fármacos , Oligopeptídeos/química , Oligopeptídeos/farmacologia , Animais , Adesão Celular/efeitos dos fármacos , Fibronectinas/metabolismo , Cobaias , Integrinas/metabolismo , Espectroscopia de Ressonância Magnética , Inibidores da Agregação Plaquetária/química , Inibidores da Agregação Plaquetária/farmacologia , Conformação Proteica , Relação Estrutura-Atividade , Vitronectina/metabolismo
16.
Mol Cancer Ther ; 4(11): 1670-80, 2005 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-16275988

RESUMO

The aim of the present study was to identify specific alpha(v)beta3/alpha(v)beta5 integrin antagonists active on tumor-induced angiogenesis. To this purpose, in vitro integrin-binding assays were used to screen a library of conformationally constrained bicyclic lactam Arg-Gly-Asp-containing pseudopeptides. The results identified ST1646 as a high-affinity specific ligand for alpha(v)beta3 and alpha(v)beta5 integrins with negligible interacting with alpha5beta1 integrin. In all the assays, ST1646 was equipotent to or more potent than the well-characterized integrin antagonists c(RGDfV) and cyclo(Arg-Gly-Asp-d-Phe-[NMe]Val) (EMD121974). In the chorioallantoic membrane assay, topical administration of ST1646 was able to prevent the angiogenic responses elicited by recombinant fibroblast growth factor-2 or vascular endothelial growth factor. In addition, systemic administration of ST1646 in mice exerted a significant antiangiogenic activity on neovascularization triggered by mammary carcinoma MDA-MB435 cells implanted s.c. in a dorsal air sac via a (Millipore Filter Corporation, Bedford, MA) chamber. Moreover, ST1646 delivery via an osmotic pump inhibited the growth and vascularization of tumor xenografts originating from the injection of alpha(v)beta3/alpha(v)beta5-expressing human ovarian carcinoma cells in nude mice. In agreement with the biochemical and pharmacologic studies, Monte Carlo/Stochastic Dynamics simulation showed that the bicyclic scaffold in ST1646 forced the compound to assume a preferred conformation superimposable to the X-ray conformation of alpha(v)beta3-bound EMD121974. Accordingly, computer-docking studies indicated that the ST1646-alpha(v)beta3 integrin complex maintains the ligand-receptor distances and interactions observed in the crystalline EMD121974-alpha(v)beta3 integrin complex. Taken together, these observations indicate that ST1646 represents a dual alpha(v)beta3/alpha(v)beta5 integrin antagonist with interesting biochemical and biological features to be tested in cancer therapy.


Assuntos
Integrina alfaVbeta3/antagonistas & inibidores , Integrinas/antagonistas & inibidores , Oligopeptídeos/química , Peptídeos Cíclicos/farmacologia , Receptores de Vitronectina/antagonistas & inibidores , Animais , Antineoplásicos/farmacologia , Arginina/química , Ácido Aspártico/química , Bovinos , Adesão Celular , Linhagem Celular Tumoral , Proliferação de Células , Galinhas , Cristalografia por Raios X , Relação Dose-Resposta a Droga , Endotélio Vascular/citologia , Fator 2 de Crescimento de Fibroblastos/metabolismo , Glicina/química , Cobaias , Humanos , Concentração Inibidora 50 , Integrinas/metabolismo , Ligantes , Camundongos , Camundongos Nus , Microcirculação , Modelos Químicos , Modelos Moleculares , Conformação Molecular , Método de Monte Carlo , Transplante de Neoplasias , Neovascularização Patológica , Peptídeos Cíclicos/química , Agregação Plaquetária , Ligação Proteica , Conformação Proteica , Proteínas Recombinantes/química , Processos Estocásticos , Fator A de Crescimento do Endotélio Vascular/metabolismo , Vitronectina/química
17.
J Gene Med ; 6(9): 992-1002, 2004 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-15352072

RESUMO

BACKGROUND: In mouse models of retinopathy of prematurity (ROP) inhibitors of vascular endothelial growth factor (VEGF) functions administered systemically completely block retinal neovascularization. In contrast, selective ocular VEGF depletion has achieved an approx. 50% inhibition of retinal neovascular growth. It is unclear whether a more complete inhibition of new blood vessel development can be obtained with an anti-VEGF therapy localized to the eye. Therefore, the objective of the present study was to determine the effect of local anti-VEGF therapy in a different animal model which closely mimics human ROP. METHODS: Rats were exposed to alternating cycles of high and low levels of oxygen for 14 days immediately after birth; thereafter, they were intravitreally injected with an adenoviral vector expressing a secreted form of the VEGF receptor flt-1 (Ad.sflt), which acts by sequestering VEGF. Contralateral eyes were injected with the control vector carrying the reporter gene expressing beta-galactosidase (Ad.betaGal). RESULTS: At the peak of retinal neovascular growth, i.e. post-natal day 21 (P21), we observed up to 97.5% decrease in retinal neovascularization in animals injected with Ad.sflt. At the end of observation (P28), no significant difference in retinal vessel number was detected in both oxygen-injured and normoxic Ad.sflt-treated retinas compared with untreated or Ad.betaGal-treated retinas. CONCLUSION: Adenoviral-mediated sflt-1 gene transfer induces a near-complete inhibition of ischemia-induced retinal neovascularization in rats without affecting pre-existing retinal vessels.


Assuntos
Terapia Genética/métodos , Neovascularização Retiniana/prevenção & controle , Retinopatia da Prematuridade/terapia , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/uso terapêutico , Adenoviridae/genética , Animais , Modelos Animais de Doenças , Vetores Genéticos/uso terapêutico , Humanos , Recém-Nascido , Ratos , Retina/patologia , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/biossíntese , Corpo Vítreo/metabolismo
18.
J Biol Chem ; 279(42): 43929-39, 2004 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-15272021

RESUMO

Placenta growth factor (PlGF) belongs to the vascular endothelial growth factor (VEGF) family and represents a key regulator of angiogenic events in pathological conditions. PlGF exerts its biological function through the binding and activation of the seven immunoglobulin-like domain receptor Flt-1, also known as VEGFR-1. Here, we report the first detailed mutagenesis studies that provide a basis for understanding molecular recognition between PlGF-1 and Flt-1, highlighting some of the residues that are critical for receptor recognition. Mutagenesis analysis, performed on the basis of a structural model of interaction between PlGF and the minimal binding domain of Flt-1, has led to the identification of several PlGF-1 residues involved in Flt-1 recognition. The two negatively charged residues, Asp-72 and Glu-73, located in the beta3-beta4 loop, are critical for Flt-1 binding. Other mutations, which bring about a significant decrease in PlGF binding activity, are Gln-27, located in the N-terminal alpha-helix, and Pro-98 and Tyr-100 on the beta6 strand. The mutation of one of the two glycosylated residues of PlGF, Asn-84, generates a PlGF variant with reduced binding activity. This indicates that, unlike in VEGF, glycosylation plays an important role in Flt-1 binding. The double mutation of residues Asp-72 and Glu-73 generates a PlGF variant unable to bind and activate the receptor molecules on the cell surface. This variant failed to induce in vitro capillary-like tube formation of primary endothelial cells or neo-angiogenesis in an in vivo chorioallantoic membrane assay.


Assuntos
Proteínas da Gravidez/metabolismo , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Alantoide/fisiologia , Sequência de Aminoácidos , Animais , Sítios de Ligação , Linhagem Celular , Embrião de Galinha , Córion/fisiologia , DNA Complementar/genética , Feminino , Ácido Glutâmico , Glutamina , Humanos , Dados de Sequência Molecular , Mutagênese , Mutagênese Sítio-Dirigida , Placenta , Fator de Crescimento Placentário , Gravidez , Proteínas da Gravidez/química , Estrutura Secundária de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Transfecção
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