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1.
Hypertens Pregnancy ; 41(2): 99-106, 2022 May.
Artigo em Inglês | MEDLINE | ID: mdl-35249426

RESUMO

Placental growth factor (PlGF), a member of the vascular endothelial growth factor family of proteins regulating angiogenesis, has been shown to have acute vasodilatory effects on human resistance arteries. However, the acute hemodynamic effects of PlGF in vivo are not known. The aim of this study was to report acute cardiovascular changes induced by recombinant human PlGF administered intravenously in male Guinea Pigs with implanted telemeters. PlGF decreased mean arterial blood pressure by 10-20% within minutes. The magnitude of reduction was similar at three dose levels; however, the duration of relative hypotension was dose-dependent. Blood pressure reduction resulted in a compensatory increase in heart rate, or reflex tachycardia. To rule out any direct effect on the heart, PlGF was tested in the ex vivo Langendorff heart preparation, and no cardiac changes were found. Together these results suggest that the PlGF-related changes in blood pressure are largely mediated by its actions in the vasculature.


Assuntos
Hipotensão , Fator A de Crescimento do Endotélio Vascular , Animais , Feminino , Cobaias , Hemodinâmica , Humanos , Masculino , Neovascularização Patológica/metabolismo , Placenta/metabolismo , Fator de Crescimento Placentário/metabolismo , Gravidez , Fator A de Crescimento do Endotélio Vascular/metabolismo
2.
J Hypertens ; 38(11): 2295-2304, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32618892

RESUMO

OBJECTIVES: Although epidemiological studies have shown that obesity is associated with increased incidence of hypertension during pregnancy, the mechanisms linking these two comorbidities are not as well studied. Previous investigations detected lower levels of the anti-hypertensive and pregnancy-related factor, placental growth factor (PlGF), in obese hypertensive pregnancies. Therefore, we examined whether obese hypertensive pregnant rats have reduced PlGF and whether increasing its levels by administering recombinant human (rh)PlGF reduces their blood pressure. METHODS: We utilized a genetic model of obesity characterized to be heavier, hypertensive and fertile, namely rats having heterozygous deficiency of the melanocortin-4 receptor (MC4R-def). RESULTS: MC4R-def obese rats had lower circulating levels of PlGF than wild-type lean controls at gestational day 19. Also, assessment of the PlGF receptor, Flt-1, in the vasculature showed that its levels were reduced in aorta and kidney glomeruli but increased in small mesenteric arteries. Chronic intraperitoneal administration of rhPlGF from gestational day 13-19 significantly increased circulating PlGF levels in both obese and lean rats, but reduced blood pressure only in the obese pregnant group. The rhPlGF treatment did not alter maternal body and fat masses or circulating levels of the adipokines, leptin and adiponectin. In addition, this treatment did not impact average foetal weights but increased placental weights regardless of obese or lean pregnancy. CONCLUSION: PlGF is reduced in MC4R-def obese hypertensive pregnant rats, which is similar to findings in obese hypertensive pregnant women, while increasing its levels with exogenous rhPlGF reduces their blood pressure.


Assuntos
Pressão Sanguínea/efeitos dos fármacos , Hipertensão/metabolismo , Obesidade/metabolismo , Fator de Crescimento Placentário , Proteínas Recombinantes , Animais , Feminino , Humanos , Fator de Crescimento Placentário/administração & dosagem , Fator de Crescimento Placentário/farmacologia , Gravidez , Ratos , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/farmacologia
3.
Hypertension ; 67(4): 740-7, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26831193

RESUMO

Preeclampsia is a pregnancy-specific disorder of new-onset hypertension. Unfortunately, the most effective treatment is early delivery of the fetus and placenta. Placental ischemia appears central to the pathogenesis of preeclampsia because placental ischemia/hypoxia induced in animals by reduced uterine perfusion pressure (RUPP) or in humans stimulates release of hypertensive placental factors into the maternal circulation. The anti-angiogenic factor soluble fms-like tyrosine kinase-1 (sFlt-1), which antagonizes and reduces bioavailable vascular endothelial growth factor and placental growth factor (PlGF), is elevated in RUPP rats and preeclampsia. Although PlGF and vascular endothelial growth factor are both natural ligands for sFlt-1, vascular endothelial growth factor also has high affinity to VEGFR2 (Flk-1) causing side effects like edema. PlGF is specific for sFlt-1. We tested the hypothesis that PlGF treatment reduces placental ischemia-induced hypertension by antagonizing sFlt-1 without adverse consequences to the mother or fetus. On gestational day 14, rats were randomized to 4 groups: normal pregnant or RUPP±infusion of recombinant human PlGF (180 µg/kg per day; AG31, a purified, recombinant human form of PlGF) for 5 days via intraperitoneal osmotic minipumps. On day 19, mean arterial blood pressure and plasma sFlt-1 were higher and glomerular filtration rate lower in RUPP than normal pregnant rats. Infusion of recombinant human PlGF abolished these changes seen with RUPP along with reducing oxidative stress. These data indicate that the increased sFlt-1 and reduced PlGF resulting from placental ischemia contribute to maternal hypertension. Our novel finding that recombinant human PlGF abolishes placental ischemia-induced hypertension, without major adverse consequences, suggests a strong therapeutic potential for this growth factor in preeclampsia.


Assuntos
Hipertensão Induzida pela Gravidez/tratamento farmacológico , Isquemia/complicações , Placenta/irrigação sanguínea , Pré-Eclâmpsia/tratamento farmacológico , Proteínas da Gravidez/farmacologia , Prenhez , Análise de Variância , Animais , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Hipertensão Induzida pela Gravidez/etiologia , Isquemia/tratamento farmacológico , Estresse Oxidativo/fisiologia , Placenta/metabolismo , Fator de Crescimento Placentário , Circulação Placentária/efeitos dos fármacos , Circulação Placentária/fisiologia , Insuficiência Placentária , Pré-Eclâmpsia/etiologia , Gravidez , Proteínas da Gravidez/metabolismo , Ratos , Ratos Sprague-Dawley , Valores de Referência , Medição de Risco
4.
Structure ; 19(8): 1097-107, 2011 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-21827946

RESUMO

The anti-VEGF receptor 2 antibody IMC-1121B is a promising antiangiogenic drug being tested for treatment of breast and gastric cancer. We have determined the structure of the 1121B Fab fragment in complex with domain 3 of VEGFR2, as well as the structure of a different neutralizing anti-VEGFR2 antibody, 6.64, also in complex with VEGFR2 domain 3. The two Fab fragments bind at opposite ends of VEGFR2 domain 3; 1121B directly blocks VEGF binding, whereas 6.64 may prevent receptor dimerization by perturbing the domain 3:domain 4 interface. Mutagenesis reveals that residues essential for VEGF, 1121B, and 6.64 binding are nonoverlapping among the three contact patches.


Assuntos
Inibidores da Angiogênese/química , Anticorpos Monoclonais/química , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/química , Motivos de Aminoácidos , Sequência de Aminoácidos , Anticorpos Monoclonais Humanizados , Especificidade de Anticorpos , Sítios de Ligação de Anticorpos , Cristalografia por Raios X , Ensaio de Imunoadsorção Enzimática , Modelos Moleculares , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Ligação Proteica , Estrutura Quaternária de Proteína , Estrutura Terciária de Proteína , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Ramucirumab
5.
Transl Oncol ; 3(5): 318-25, 2010 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-20885894

RESUMO

Microtubules are a well-validated target for anticancer therapy. Molecules that bind tubulin affect dynamic instability of microtubules causing mitotic arrest of proliferating cells, leading to cell death and tumor growth inhibition. Natural antitubulin agents such as taxanes and Vinca alkaloids have been successful in the treatment of cancer; however, several limitations have encouraged the development of synthetic small molecule inhibitors of tubulin function. We have previously reported the discovery of two novel chemical series of tubulin polymerization inhibitors, triazoles (Ouyang et al. Synthesis and structure-activity relationships of 1,2,4-triazoles as a novel class of potent tubulin polymerization inhibitors. Bioorg Med Chem Lett. 2005; 15:5154-5159) and oxadiazole derivatives (Ouyang et al. Oxadiazole derivatives as a novel class of antimitotic agents: synthesis, inhibition of tubulin polymerization, and activity in tumor cell lines. Bioorg Med Chem Lett. 2006; 16:1191-1196). Here, we report on the anticancer effects of a lead oxadiazole derivative in vitro and in vivo. In vitro, IMC-038525 caused mitotic arrest at nanomolar concentrations in epidermoid carcinoma and breast tumor cells, including multidrug-resistant cells. In vivo, IMC-038525 had a desirable pharmacokinetic profile with sustained plasma levels after oral dosing. IMC-038525 reduced subcutaneous xenograft tumor growth with significantly greater efficacy than the taxane paclitaxel. At efficacious doses, IMC-038525 did not cause substantial myelosuppression or peripheral neurotoxicity, as evaluated by neutrophil counts and changes in myelination of the sciatic nerve, respectively. These data indicate that IMC-038525 is a promising candidate for further development as a chemotherapeutic agent.

6.
Arthritis Rheum ; 60(4): 1145-51, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19333919

RESUMO

OBJECTIVE: To investigate whether agonist anti-platelet-derived growth factor receptor alpha (anti-PDGFRalpha) antibodies are present in the serum of patients with systemic sclerosis (SSc; scleroderma). METHODS: Sera were obtained from healthy subjects and scleroderma patients. An electrochemiluminescence binding assay was performed for detection of serum autoantibodies to PDGFRalpha, PDGFRbeta, epidermal growth factor receptor (EGFR), and colony-stimulating factor receptor 1 (CSFR1). Serum immunoglobulin was purified by protein A/G chromatography. To assess Ig agonist activity, PDGFRalpha-expressing cells were incubated with pure Ig and the level of receptor phosphorylation determined in an enzyme-linked immunoassay, as well as by Western blotting. Ig agonist activity was also assessed in a mitogenic assay and by MAP kinase activation in a PDGFRalpha-expressing cell line. RESULTS: Sera from 34.3% of the healthy subjects and 32.7% of the SSc patients contained detectable autoantibodies to PDGFRalpha and PDGFRbeta, but not EGFR or CSFR1. Purified Ig from these sera was shown to retain PDGFR binding activity and, at 200-1,000 microg/ml, exhibited no agonist activity in a cell-based PDGFRalpha phosphorylation assay and did not stimulate a mitogenic response or MAP kinase activation in a PDGFRalpha-expressing cell line. Two purified Ig samples that were unable to bind PDGFRalpha did exhibit binding activity to a nonglycosylated form of PDGFRalpha. CONCLUSION: Although approximately one-third of sera from scleroderma patients contained detectable autoantibodies to PDGFR, these antibodies were not specific to scleroderma, since they were also detected in a similar percentage of samples from normal subjects. PDGFRalpha agonist activity was not demonstrated when purified Ig from these sera was tested in cell-based assays.


Assuntos
Especificidade de Anticorpos , Autoanticorpos/imunologia , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/imunologia , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Escleroderma Sistêmico/imunologia , Adulto , Idoso , Autoanticorpos/sangue , Autoanticorpos/farmacologia , Linhagem Celular , Feminino , Humanos , Sistema de Sinalização das MAP Quinases/imunologia , Masculino , Pessoa de Meia-Idade , Mitógenos/imunologia , Fosforilação/imunologia , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/agonistas , Receptor beta de Fator de Crescimento Derivado de Plaquetas/imunologia
7.
Bioorg Med Chem ; 17(2): 731-40, 2009 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-19101155

RESUMO

A series of arylphthalazine derivatives were synthesized and evaluated as antagonists of VEGF receptor II (VEGFR-2). IM-094482 57, which was prepared in two steps from commercially available starting materials, was found to be a potent inhibitor of VEGFR-2 in enzymatic, cellular and mitogenic assays (comparable activity to ZD-6474). Additionally, 57 inhibited the related receptor, VEGF receptor I (VEGFR-1), and showed excellent exposure when dosed orally to female CD-1 mice.


Assuntos
Ftalazinas/farmacocinética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Administração Oral , Animais , Disponibilidade Biológica , Feminino , Isoquinolinas/síntese química , Isoquinolinas/farmacocinética , Camundongos , Camundongos Endogâmicos , Ftalazinas/administração & dosagem , Ftalazinas/síntese química , Piperidinas , Quinazolinas , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores
8.
Structure ; 16(2): 216-27, 2008 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-18275813

RESUMO

Therapeutic anticancer strategies that target and inactivate the epidermal growth factor receptor (EGFR) are under intense study in the clinic. Here we describe the mechanism of EGFR inhibition by an antibody drug IMC-11F8. IMC-11F8 is a fully human antibody that has similar antitumor potency as the chimeric cetuximab/Erbitux and might represent a safer therapeutic alternative. We report the X-ray crystal structure of the Fab fragment of IMC-11F8 (Fab11F8) in complex with the entire extracellular region and with isolated domain III of EGFR. We compare this to our previous study of the cetuximab/EGFR interaction. Fab11F8 interacts with a remarkably similar epitope, but through a completely different set of interactions. Both the similarities and differences in binding of these two antibodies have important implications for the development of inhibitors that could exploit this same mechanism of EGFR inhibition.


Assuntos
Anticorpos Monoclonais/química , Antineoplásicos/química , Receptores ErbB/química , Sequência de Aminoácidos , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/uso terapêutico , Anticorpos Monoclonais Humanizados , Antineoplásicos/imunologia , Antineoplásicos/uso terapêutico , Sítios de Ligação , Cetuximab , Cristalografia por Raios X , Epitopos/química , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/imunologia , Glicosilação , Fragmentos Fab das Imunoglobulinas/química , Modelos Moleculares , Dados de Sequência Molecular , Mutação , Estrutura Terciária de Proteína
9.
Am J Physiol Endocrinol Metab ; 292(3): E964-76, 2007 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-17132826

RESUMO

We generated three fully human monoclonal antibody antagonists against fibroblast growth factor receptor-1 (FGFR1) that potently block FGF signaling. We found that antibodies targeting the c-splice form of the receptor (FGFR1c) were anorexigenic when administered intraperitoneally three times weekly to mice, resulting in rapid, dose-dependent weight loss that plateaued (for doses>4 mg/kg) at 35-40% in 2 wk. Animals appeared healthy during treatment and regained their normal body weights and growth trajectories upon clearance of the antibodies from the bloodstream. Measurements of food consumption and energy expenditure indicated that the rapid weight loss was induced primarily by decreased energy intake and not by increased energy expenditure or cachexia and was accompanied by a greater reduction in fat than lean body mass. Hypophagia was not caused through malaise or illness, as indicated by absence of conditioned taste aversion, pica behavior, and decreased need-induced salt intake in rats. In support of a hypothalamic site of action, we found that, after intraperitoneal injections, anti-FGFR1c (IMC-A1), but not a control antibody, accumulated in the median eminence and adjacent mediobasal hypothalamus and that FGFR1c is enriched in the hypothalamus of mice. Furthermore, a single intracerebroventricular administration of 3 microg of IMC-A1 via the 3rd ventricle to mice caused an approximately 36% reduction in food intake and an approximately 6% weight loss within the ensuing 24 h. Our data suggest that FGF signaling through FGFR1c may play a physiological role in hypothalamic feeding circuit and that blocking it leads to hypophagia and weight loss.


Assuntos
Anticorpos Monoclonais/farmacologia , Ingestão de Alimentos/efeitos dos fármacos , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/antagonistas & inibidores , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/imunologia , Redução de Peso/efeitos dos fármacos , Animais , Anticorpos Monoclonais/efeitos adversos , Especificidade de Anticorpos , Composição Corporal/efeitos dos fármacos , Sistema Nervoso Central/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Feminino , Macaca fascicularis , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Isoformas de Proteínas/imunologia , Ratos , Ratos Sprague-Dawley
10.
Curr Med Chem ; 13(18): 2101-11, 2006.
Artigo em Inglês | MEDLINE | ID: mdl-16918340

RESUMO

Heparanase is an endo-beta-D-glucuronidase that degrades heparan sulfate glycosaminoglycan side chains of the proteoglycans in extracellular matrix and basement membrane. Heparanase enzymatic activity is important in the promotion of tumor angiogenesis, primary tumor growth, invasion, and metastasis. Expression of heparanase in many tumor types conversely correlates with prognosis. Much progress has been made in studying the regulation of heparanase expression, processing and activation. The interaction between heparanase and its substrate heparan sulfate has been well characterized. The fact that heparanase was identified as the single predominant heparan sulfate-degrading enzyme in human cancer sparked considerable interest in developing heparanase inhibitors for potential therapeutic applications. Recent progress in drug development led to several classes of heparanase inhibitors, including chemically modified natural products, small molecule inhibitors, and antibodies. Some of these inhibitors have demonstrated potent activities to inhibit tumor angiogenesis, tumor progress, or tumor metastasis. A leading compound, PI-88, is currently being evaluated in clinical phase II trials in patients with melanoma, liver, or lung cancers. This review summarizes the recent progress in heparanase biochemical research and the development of heparanase antagonists as novel anti-cancer therapeutics.


Assuntos
Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Glucuronidase/antagonistas & inibidores , Glucuronidase/fisiologia , Neoplasias/tratamento farmacológico , Biomarcadores/análise , Desenho de Fármacos , Glucuronidase/análise , Humanos , Metástase Neoplásica , Neoplasias/patologia , Neovascularização Patológica , Nitrogênio/química , Oxigênio/química , Sulfatos/química
11.
Bioorg Med Chem Lett ; 16(2): 404-8, 2006 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-16246551

RESUMO

A novel class of N-(4-{[4-(1H-benzoimidazol-2-yl)-arylamino]-methyl}-phenyl)-benzamides are described as inhibitors of the endo-beta-glucuronidase heparanase. Among them are N-(4-{[4-(1H-benzoimidazol-2-yl)-phenylamino]-methyl}-phenyl)-3-bromo-4-methoxy-benzamide (15h), and N-(4-{[5-(1H-benzoimidazol-2-yl)-pyridin-2-ylamino]-methyl}- phenyl)-3-bromo-4-methoxy-benzamide (23) which displayed good heparanase inhibitory activity (IC(50) 0.23-0.29 microM), with the latter showing oral exposure in mice.


Assuntos
Benzamidas/farmacologia , Benzimidazóis/farmacologia , Inibidores Enzimáticos/farmacologia , Glucuronidase/antagonistas & inibidores , Administração Oral , Animais , Benzamidas/administração & dosagem , Benzamidas/química , Benzimidazóis/administração & dosagem , Benzimidazóis/química , Configuração de Carboidratos , Sequência de Carboidratos , Desenho de Fármacos , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/administração & dosagem , Inibidores Enzimáticos/química , Técnicas In Vitro , Camundongos , Modelos Animais , Dados de Sequência Molecular , Estrutura Molecular , Peso Molecular , Relação Estrutura-Atividade
12.
Bioorg Med Chem Lett ; 16(2): 409-12, 2006 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-16246560

RESUMO

A novel class of 1-[4-(1H-benzoimidazol-2-yl)-phenyl]-3-[4-(1H-benzoimidazol-2-yl)-phenyl]-ureas are described as potent inhibitors of heparanase. Among them are 1,3-bis-[4-(1H-benzoimidazol-2-yl)-phenyl]-urea (7a) and 1,3-bis-[4-(5,6-dimethyl-1H-benzoimidazol-2-yl)-phenyl]-urea (7d), which displayed good heparanase inhibitory activity (IC(50) 0.075-0.27 microM). Compound 7a showed good efficacy in a B16 metastasis model.


Assuntos
Carbanilidas/farmacologia , Inibidores Enzimáticos/farmacologia , Glucuronidase/antagonistas & inibidores , Neoplasias Pulmonares/tratamento farmacológico , Melanoma Experimental/tratamento farmacológico , Animais , Carbanilidas/síntese química , Carbanilidas/classificação , Linhagem Celular Tumoral , Desenho de Fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/classificação , Técnicas In Vitro , Melanoma Experimental/secundário , Camundongos , Estrutura Molecular , Peso Molecular , Metástase Neoplásica/tratamento farmacológico , Relação Estrutura-Atividade
14.
Bioorg Med Chem Lett ; 15(23): 5154-9, 2005 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-16198562

RESUMO

A novel triazole-containing chemical series was shown to inhibit tubulin polymerization and cause cell cycle arrest in A431 cancer cells with EC(50) values in the single digit nanomolar range. Binding experiments demonstrated that representative active compounds of this class compete with colchicine for its binding site on tubulin. The syntheses and structure-activity relationship studies for the triazole derivatives are described herein.


Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Triazóis/química , Triazóis/farmacologia , Moduladores de Tubulina/química , Moduladores de Tubulina/farmacologia , Antineoplásicos/síntese química , Humanos , Microtúbulos/efeitos dos fármacos , Estrutura Molecular , Relação Estrutura-Atividade , Triazóis/síntese química , Moduladores de Tubulina/síntese química , Células Tumorais Cultivadas
15.
Cancer Cell ; 7(4): 301-11, 2005 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-15837620

RESUMO

Recent structural studies of epidermal growth factor receptor (EGFR) family extracellular regions have identified an unexpected mechanism for ligand-induced receptor dimerization that has important implications for activation and inhibition of these receptors. Here we describe the 2.8 angstroms resolution X-ray crystal structure of the antigen binding (Fab) fragment from cetuximab (Erbitux), an inhibitory anti-EGFR antibody, in complex with the soluble extracellular region of EGFR (sEGFR). The sEGFR is in the characteristic "autoinhibited" or "tethered" inactive configuration. Cetuximab interacts exclusively with domain III of sEGFR, partially occluding the ligand binding region on this domain and sterically preventing the receptor from adopting the extended conformation required for dimerization. We suggest that both these effects contribute to potent inhibition of EGFR activation.


Assuntos
Anticorpos Monoclonais/química , Receptores ErbB/química , Modelos Moleculares , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais Humanizados , Complexo Antígeno-Anticorpo/química , Antineoplásicos/química , Antineoplásicos/imunologia , Sítios de Ligação/genética , Ligação Competitiva , Cetuximab , Cristalografia por Raios X , Fator de Crescimento Epidérmico/química , Epitopos/química , Epitopos/genética , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/imunologia , Humanos , Fragmentos Fab das Imunoglobulinas/química , Fragmentos Fab das Imunoglobulinas/imunologia , Fragmentos Fab das Imunoglobulinas/farmacologia , Mutação/genética , Ligação Proteica , Estrutura Quaternária de Proteína , Estrutura Terciária de Proteína/efeitos dos fármacos , Agregação de Receptores/efeitos dos fármacos , Proteínas Recombinantes/química , Proteínas Recombinantes/imunologia , Fator de Crescimento Transformador alfa/química
16.
Mol Cancer Ther ; 4(3): 369-79, 2005 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-15767546

RESUMO

Platelet-derived growth factor receptor alpha (PDGFRalpha) is a type III receptor tyrosine kinase that is expressed on a variety of tumor types. A neutralizing monoclonal antibody to human PDGFRalpha, which did not cross-react with the beta form of the receptor, was generated. The fully human antibody, termed 3G3, has a Kd of 40 pmol/L and blocks both PDGF-AA and PDGF-BB ligands from binding to PDGFRalpha. In addition to blocking ligand-induced cell mitogenesis and receptor autophosphorylation, 3G3 inhibited phosphorylation of the downstream signaling molecules Akt and mitogen-activated protein kinase. This inhibition was seen in both transfected and tumor cell lines expressing PDGFRalpha. The in vivo antitumor activity of 3G3 was tested in human glioblastoma (U118) and leiomyosarcoma (SKLMS-1) xenograft tumor models in athymic nude mice. Antibody 3G3 significantly inhibited the growth of U118 (P=0.0004) and SKLMS-1 (P <0.0001) tumors relative to control. These data suggest that 3G3 may be useful for the treatment of tumors that express PDGFRalpha.


Assuntos
Anticorpos Monoclonais/química , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Animais , Becaplermina , Bioensaio , Linhagem Celular Tumoral , Relação Dose-Resposta Imunológica , Citometria de Fluxo , Humanos , Cinética , Ligantes , Sistema de Sinalização das MAP Quinases , Camundongos , Camundongos Nus , Camundongos Transgênicos , Transplante de Neoplasias , Fosforilação , Fator de Crescimento Derivado de Plaquetas/química , Ligação Proteica , Proteínas Serina-Treonina Quinases/metabolismo , Estrutura Terciária de Proteína , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-akt , Proteínas Proto-Oncogênicas c-sis , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/imunologia , Fatores de Tempo , Transfecção
17.
Blood ; 105(11): 4337-44, 2005 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-15701713

RESUMO

VE-cadherin is an adhesion molecule localized at the adherens junctions of endothelial cells. It is crucial for the proper assembly of vascular structures during angiogenesis and maintaining vascular integrity. We have studied 3 monoclonal antibodies (mAbs) against murine VE-cadherin that inhibit angiogenesis and tumor growth. Two of these, BV13 and 10G4, also disrupted normal vessels, resulting in severe vascular leakage, whereas the third, E4G10, did not. The goal of the current report was to identify the epitope of E4G10 and distinguish it from those of the disruptive mAbs. We mapped the epitope of E4G10 to within the first 10 amino acids of mature VE-cadherin and demonstrated that conserved tryptophan residues in this sequence are required for VE-cadherin-mediated trans-adhesion. The disruptive mAbs target a different epitope within amino acids 45 to 56, which structural homology modeling suggests is not involved in trans-adhesion. From our studies, we hypothesize that E4G10 can only bind the neovasculature, where VE-cadherin has not yet engaged in trans-adhesion and its epitope is fully exposed. Thus, E4G10 can inhibit junction formation and angiogenesis but is unable to target normal vasculature because its epitope is masked. In contrast, BV13 and 10G4 bind an epitope that is accessible regardless of VE-cadherin interactions, leading to the disruption of adherens junctions. Our findings establish the immediate N-terminal region of VE-cadherin as a novel target for inhibiting angiogenesis.


Assuntos
Anticorpos Monoclonais/farmacologia , Caderinas/imunologia , Epitopos , Neoplasias/tratamento farmacológico , Neovascularização Patológica/prevenção & controle , Inibidores da Angiogênese , Animais , Antígenos CD , Antígenos de Neoplasias/imunologia , Permeabilidade Capilar/efeitos dos fármacos , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Endotélio Vascular , Mapeamento de Epitopos , Junções Intercelulares/efeitos dos fármacos , Camundongos , Neoplasias/irrigação sanguínea , Neoplasias/patologia , Neovascularização Patológica/tratamento farmacológico , Triptofano
18.
Blood ; 105(3): 1303-9, 2005 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-15471949

RESUMO

Although widespread skeletal dissemination is a critical step in the progression of myeloma, little is known regarding mechanisms that control metastasis of this cancer. Heparanase-1 (heparanase), an enzyme that cleaves heparan sulfate chains, is expressed at high levels in some patients with myeloma and promotes metastasis of some tumor types (eg, breast, lymphoma). Using a severe combined immunodeficient (SCID) mouse model, we demonstrate that enhanced expression of heparanase by myeloma cells dramatically up-regulates their spontaneous metastasis to bone. This occurs from primary tumors growing subcutaneously and also from primary tumors established in bone. Interestingly, tumors formed by subcutaneous injection of cells metastasize not only to bone, but also to other sites including spleen, liver, and lung. In contrast, tumors formed by injection of cells directly into bone exhibit a restricted pattern of metastasis that includes dissemination of tumor to other bones but not to extramedullary sites. In addition, expression of heparanase by myeloma cells (1) accelerates the initial growth of the primary tumor, (2) increases whole-body tumor burden as compared with controls, and (3) enhances both the number and size of microvessels within the primary tumor. These studies describe a novel experimental animal model for examining the spontaneous metastasis of bone-homing tumors and indicate that heparanase is a critical determinant of myeloma dissemination and growth in vivo.


Assuntos
Osso e Ossos/patologia , Divisão Celular/fisiologia , Glucuronidase/metabolismo , Mieloma Múltiplo/patologia , Metástase Neoplásica/patologia , DNA Complementar/análise , Glucuronidase/genética , Humanos , Proteínas Recombinantes , Transfecção , Células Tumorais Cultivadas
19.
Hum Antibodies ; 13(3): 81-90, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-15598988

RESUMO

Recombinant protein production in plants such as corn is a promising means to generate high product yields at low comparable production cost. The anti-EGFR monoclonal antibody C225, cetuximab, is a well-characterized receptor antagonist antibody recently approved for the treatment of refractory colorectal cancer. We initiated a study to test and compare the functional activity of glycosylated and aglycosylated C225 produced in stable transgenic corn seed. Both corn antibodies were shown to be functionally indistinguishable from mammalian-derived C225 in demonstrating high-affinity binding to the EGF receptor, blocking of ligand-dependent signaling, and inhibiting cell proliferation. In addition, consistent with cetuximab, both corn antibodies possessed strong anti-tumor activity in vivo. Acute dose primate pharmacokinetic studies, however, revealed a marked increase in clearance for the glycosylated corn antibody, while the aglycosylated antibody possessed in vivo kinetics similar to cetuximab. This experimentation established that corn-derived receptor blocking monoclonal antibodies possess comparable efficacy to mammalian cell culture-derived antibody, and offer a cost effective alternative to large-scale mammalian cell culture production.


Assuntos
Anticorpos Monoclonais/biossíntese , Anticorpos Monoclonais/farmacologia , Antineoplásicos/isolamento & purificação , Antineoplásicos/farmacologia , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/imunologia , Zea mays/genética , Zea mays/imunologia , Animais , Anticorpos Monoclonais/farmacocinética , Anticorpos Monoclonais Humanizados , Citotoxicidade Celular Dependente de Anticorpos , Antineoplásicos/farmacocinética , Cetuximab , Feminino , Humanos , Técnicas In Vitro , Cinética , Macaca fascicularis , Masculino , Camundongos , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/terapia , Plantas Geneticamente Modificadas , Ligação Proteica , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/isolamento & purificação , Proteínas Recombinantes de Fusão/farmacologia , Transplante Heterólogo
20.
Blood ; 104(4): 1137-44, 2004 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-15105287

RESUMO

FMS-like tyrosine kinase 3 (FLT3), a class III receptor tyrosine kinase, is expressed at high levels in the blasts of approximately 90% of patients with acute myelogenous leukemia (AML). Internal tandem duplications (ITDs) in the juxtamembrane domain and point mutations in the kinase domain of FLT3 are found in approximately 37% of AML patients and are associated with a poor prognosis. We report here the development and characterization of a fully human anti-FLT3 neutralizing antibody (IMC-EB10) isolated from a human Fab phage display library. IMCEB10 (immunoglobulin G1 [IgG1], kappa) binds with high affinity (KD=158 pM) to soluble FLT3 in enzyme-linked immunosorbent assay (ELISA) and to FLT3 receptor expressed on the surfaces of human leukemia cell lines. IMC-EB10 blocks the binding of FLT3 ligand (FL) to soluble FLT3 in ELISA and competes with FL for binding to cell-surface FLT3 receptor. IMC-EB10 treatment inhibits FL-induced phosphorylation of FLT3 in EOL-1 and EM3 leukemia cells and FL-independent constitutive activation of ITD-mutant FLT3 in BaF3-ITD and MV4;11 cells. Activation of the downstream signaling proteins mitogen-activated protein kinase (MAPK) and AKT is also inhibited in these cell lines by antibody treatment. The antibody inhibits FL-stimulated proliferation of EOL-1 cells and ligand-independent proliferation of BaF3-ITD cells. In both EOL-1 xenograft and BaF3-ITD leukemia models, treatment with IMC-EB10 significantly prolongs the survival of leukemia-bearing mice. No overt toxicity is observed with IMC-EB10 treatment. Taken together, these data demonstrate that IMC-EB10 is a specific and potent inhibitor of wild-type and ITD-mutant FLT3 and that it deserves further study for targeted therapy of human AML.


Assuntos
Anticorpos Antineoplásicos/farmacologia , Leucemia/tratamento farmacológico , Proteínas Proto-Oncogênicas/imunologia , Receptores Proteína Tirosina Quinases/imunologia , Animais , Anticorpos Antineoplásicos/uso terapêutico , Ligação Competitiva , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Fragmentos Fab das Imunoglobulinas/farmacologia , Fragmentos Fab das Imunoglobulinas/uso terapêutico , Masculino , Proteínas de Membrana , Camundongos , Camundongos Endogâmicos NOD , Camundongos Nus , Mutação , Neoplasias Experimentais/tratamento farmacológico , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Proteínas Proto-Oncogênicas/genética , Receptores Proteína Tirosina Quinases/antagonistas & inibidores , Receptores Proteína Tirosina Quinases/genética , Transdução de Sinais/efeitos dos fármacos , Taxa de Sobrevida , Sequências de Repetição em Tandem , Tirosina Quinase 3 Semelhante a fms
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