RESUMO
The interaction of acute myeloid leukaemic (AML) blasts with the bone marrow (BM) microenvironment is a major determinant governing disease progression and resistance to treatment. The constitutive expression of E-selectin in the vascular compartment of BM, a key endothelial cell factor, directly mediates chemoresistance via E-selectin ligand/receptors. Despite the success of hypomethylating agent (HMA)-containing regimens to induce remissions in older AML patients, the development of primary or secondary resistance is common. We report that following treatment with 5-azacitidine, promoter regions regulating the biosynthesis of the E-selectin ligands, sialyl Lewis X, become further hypomethylated. The resultant upregulation of these gene products, in particular α(1,3)-fucosyltransferase VII (FUT7) and α(2,3)-sialyltransferase IV (ST3GAL4), likely causes functional E-selectin binding. When combined with the E-selectin antagonist uproleselan, the adhesion to E-selectin is reversed and the survival of mice transplanted with AML cells is prolonged. Finally, we present clinical evidence showing that BM myeloid cells from higher risk MDS and AML patients have the potential to bind E-selectin, and these cells are more abundant in 5-azacitidine-non-responsive patients. The collective data provide a strong rationale to evaluate 5-azacitidine in combination with the E-selectin antagonist, uproleselan, in this patient population.
Assuntos
Azacitidina , Selectina E , Leucemia Mieloide Aguda , Síndromes Mielodisplásicas , Humanos , Selectina E/metabolismo , Leucemia Mieloide Aguda/tratamento farmacológico , Animais , Síndromes Mielodisplásicas/tratamento farmacológico , Camundongos , Azacitidina/farmacologia , Azacitidina/uso terapêutico , Idoso , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Feminino , Antígeno Sialil Lewis X , Masculino , Fucosiltransferases , Pessoa de Meia-IdadeRESUMO
BACKGROUND: Alongside the need to develop more effective and less toxic immunosuppression, the shortage of human organs available for organ transplantation is one of the major hurdles facing the field. Research into xenotransplantation, as an alternative source of organs, has unveiled formidable challenges. Porcine lungs perfused with human blood rapidly sequester the majority of circulating neutrophils and platelets, which leads to inflammation and organ failure within hours, and is not significantly attenuated by genetic modifications to the pig targeted to diminish antibody binding and complement and coagulation cascade activation. METHODS: Here, we model the interaction of freshly isolated human leukocytes with xenotransplanted vasculature under physiologic flow conditions using microfluidic channels coated with porcine endothelial cells. Both isolated human neutrophils and whole human blood were perfused over transgenic pig aortic endothelial cells that had been activated with rhTNF-α or rhIL-4 using the BioFlux system. Novel compounds GMI-1271 and rPSGL1.Fc were tested as E- and P- selectin antagonists, respectively. Cellular adhesion and rolling events were tracked using FIJI (imageJ). RESULTS: Porcine endothelium activated with either rhTNF-α or rhIL-4 expressed high amounts of selectins, to which isolated human neutrophils readily rolled and tethered. Both E-and P-selectin antagonism significantly reduced the number of neutrophils rolling and rolling distance in a dose-dependent manner, with near total inhibition at higher doses (P < .001). Similarly, with whole human blood, selectin blocking compounds exhibited dose-dependent inhibition of prevalent leukocyte adhesion and severe endothelial injury (Untreated: 394 ± 97 PMNs/hpf, 57 ± 6% loss EC; GMI1271+rPSGL1.Fc: 23 ± 9 PMNs/hpf, 8 ± 6% loss EC P < .01). CONCLUSIONS: Selectin blockade may be useful as part of an integrated strategy to prevent neutrophil-mediated organ xenograft injury, especially during the early time points following reperfusion.
Assuntos
Selectina E/metabolismo , Células Endoteliais/imunologia , Leucócitos/imunologia , Selectina-P/metabolismo , Animais , Animais Geneticamente Modificados , Adesão Celular/fisiologia , Humanos , Neutrófilos/imunologia , Suínos , Transplante Heterólogo/métodosRESUMO
BACKGROUND: This study evaluated E-selectin inhibition with GMI-1271 (Uproleselan [GMI]) alone and in combination with the standard of care low-molecular-weight heparin (LMWH) to improve vein recanalization, decrease vein wall inflammation and protect against adverse bleeding in a primate model. We sought to examine this novel treatment of venous thrombosis. METHODS: Using a well-documented primate animal model, iliac vein thrombosis was induced by balloon occlusion of the iliac vein for 6 hours. Starting on day 2 after thrombosis, animals began treatment in two phases. In phase one, nontreated controls received no treatment (n = 5) vs animals treated with the E-selectin inhibitor GMI, 25 mg/kg, subcutaneous (SC), once daily (n = 4) for 21 days (previously published data). In phase two, animals were treated with GMI plus a combination of LMWH 1.5 mg/kg or 40 mg (GMI + LMWHc) SC once daily (n = 8) for 19 days; and animals treated with LMWH 1.5 mg/kg or 40 mg (LMWHc) SC once daily (n = 6) for 19 days. Animals were evaluated by magnetic resonance venography for vein recanalization and inflammation by gadolinium extravasation, duplex ultrasound, coagulation tests (thromboelastography, bleeding time, prothrombin time, activated partial thromboplastin time, fibrinogen) and complete blood count at baseline, days 2, 7, 14, and 21 at euthanasia. Statistical analysis included using unpaired t test with Welch's correction for direct comparisons and one-way analysis of variance for comparison between the groups. RESULTS: Percent vein recanalization by magnetic resonance venography was highest in the GMI alone group followed by GMI + LMWHc, both significantly different from control. On ultrasound examination, animals treated with GMI alone had no decrease in open vein lumen by day 21, whereas decreases were observed in groups GMI + LMWHc (-26%), LMWHc (-27%), and controls (-80%). Vein wall inflammation decreased significantly in all treated groups. Intimal fibrosis and intimal thickness was best preserved in the GMI alone group. An analysis of total vein wall collagen revealed a trend in all treatment groups of decreasing vein wall collagen. No clinically significant bleeding events were noted in any group. The LMWH groups trended to have prolonged coagulation test values, whereas E-selectin inhibition with GMI did not cause clinically significant changes in coagulation measures. CONCLUSIONS: Treatment with E-selectin inhibition results in improved vein recanalization, a decrease in vein wall inflammation and vein wall intimal thickness and fibrosis, with no changes in markers of coagulation. E-selectin inhibition with GMI alone is superior to E-selectin inhibition combined with LMWH, LMWH alone, and no treatment in this deep vein thrombosis model of iliac vein thrombosis.
Assuntos
Anticoagulantes/uso terapêutico , Selectina E/antagonistas & inibidores , Glicolipídeos/uso terapêutico , Heparina de Baixo Peso Molecular/uso terapêutico , Trombose Venosa/tratamento farmacológico , Animais , PapioRESUMO
OBJECTIVE: There is an inter-relationship between thrombosis and inflammation. Previously, we have shown the importance of P-selectin in thrombogenesis and thrombus resolution in many preclinical animal models. The role of E-selectin has been explored in rodent models and in a small pilot study of clinical calf vein deep venous thrombosis. The purpose of this study was to determine the role of E-selectin in thrombosis in a primate model of proximal iliac vein thrombosis, a model close to the human condition. METHODS: Iliac vein thrombosis was induced with a well-characterized primate model. Through a transplant incision, the hypogastric vein and iliac vein branches were ligated. Thrombus was induced by balloon occlusion of the proximal and distal iliac vein for 6 hours. The balloons were then deflated, and the primates recovered. Starting on postocclusion day 2, animals were treated with the E-selectin inhibitor GMI-1271, 25 mg/kg subcutaneously, once daily until day 21 (n = 4). Nontreated control animals received no treatment (n = 5). All animals were evaluated by magnetic resonance venography (MRV); evaluation of vessel area by ultrasound, protein analysis, hematology (complete blood count), and coagulation tests (bleeding time, prothrombin time, activated partial thromboplastin time, fibrinogen, and thromboelastography) were performed at baseline, day 2, day 7, day 14, and day 21 with euthanasia. In addition, platelet function and CD44 expression on leukocytes were determined. RESULTS: E-selectin inhibition by GMI-1271 significantly increased vein recanalization by MRV vs control animals on day 14 (P < .05) and day 21 (P < .0001). GMI-1271 significantly decreased vein wall inflammation by MRV with gadolinium vein wall enhancement vs control also on day 14 (P < .0001) and day 21 (P < .0001). The thromboelastographic measure of clot strength (maximum amplitude) showed significant decreases in animals treated with GMI-1271 vs controls at day 2 (P < .05) and day 7 (P < .05). Animals treated with GMI-1271 had significant vessel area increase by day 21 vs controls (P < .05) by ultrasound. Vein wall intimal thickening (P < .001) and intimal fibrosis (P < .05) scores were significantly decreased in GMI-1271-treated animals vs controls. Importantly, no significant differences in hematology or coagulation test results were noted between all groups, suggesting that E-selectin inhibition carries no bleeding potential. GMI-1271 did not affect platelet function or aggregation or CD44 expression on leukocytes. In addition, no episodes of bleeding were noted in either group. CONCLUSIONS: This study suggests that E-selectin modulates venous thrombus progression and that its inhibition will increase thrombus recanalization and decrease vein wall inflammation, without affecting coagulation. The use of an E-selectin inhibitor such as GMI-1271 could potentially change how we treat deep venous thrombosis.
Assuntos
Anti-Inflamatórios/farmacologia , Selectina E/antagonistas & inibidores , Fibrinolíticos/farmacologia , Glicolipídeos/farmacologia , Veia Ilíaca/efeitos dos fármacos , Trombose Venosa/tratamento farmacológico , Animais , Modelos Animais de Doenças , Selectina E/metabolismo , Veia Ilíaca/diagnóstico por imagem , Veia Ilíaca/metabolismo , Papio , Transdução de Sinais , Trombose Venosa/diagnóstico por imagem , Trombose Venosa/metabolismoRESUMO
BACKGROUND: A critical component of disease progression in rheumatoid arthritis (RA) involves neovascularization associated with pannus formation. 2-methoxyestradiol (2ME2) is a naturally occurring molecule with no known physiologic function, although at pharmacologic concentrations it has antiproliferative and antiangiogenic activities. We investigated the impact of orally administered 2ME2 on the initiation and development of proliferative synovitis using the anti-collagen monoclonal antibodies (CAIA) model. METHODS: Severe polyarticular arthritis was induced in Balb/c female mice by administration of 2 mg of a monoclonal antibody cocktail intravenously into the tail vein of mice. Twenty-four hours following monoclonal antibody administration, mice were injected with 25 microg of LPS (E. coli strain 0111:B4) via the intraperitoneal route. Treatment with 2ME2 (100, 75, 50, 25, 10, 1 mg/kg, p.o., daily), or vehicle control began 24 hrs following LPS challenge and continued to day 21. Hind limbs were harvested, sectioned and evaluated for DMARD activity and general histopathology by histomorphometric analysis and immunohistochemistry (vWF staining). In a separate study, different dosing regimens of 2ME2 (100 mg/kg; q.d. vs q.w. vs q.w. x 2) were evaluated. The effect of treatment with 2ME2 on gene expression of inflammatory cytokines and angiogenic growth factors in the joint space was evaluated 5 and 14 days after the induction of arthritis. RESULTS: Mice treated with 2ME2 beginning 24 hours post anti-collagen monoclonal antibody injection, showed a dose-dependent inhibition in mean arthritic scores. At study termination (day 21), blinded histomorphometric assessments of sectioned hind limbs demonstrated decreases in synovial inflammation, articular cartilage degradation, pannus formation, osteoclast activity and bone resorption. At the maximal efficacious dosing regimen (100 mg/kg/day), administration of 2ME2 resulted in total inhibition of the study parameters and prevented neovascularization into the joint. Examination of gene expression on dissected hind limbs from mice treated for 5 or 14 days with 2ME2 showed inhibition of inflammatory cytokine message for IL-1beta, TNF-alpha, IL-6 and IL-17, as well as the angiogenic cytokines, VEGF and FGF-2. CONCLUSION: These data demonstrate that in the CAIA mouse model of RA, 2ME2 has disease modifying activity that is at least partially attributable to the inhibition of neovascular development. Further, the data suggests new mechanistic points of intervention for 2ME2 in RA, specifically inhibition of inflammatory mediators and osteoclast activity.
Assuntos
Artrite Reumatoide/tratamento farmacológico , Vasos Sanguíneos/efeitos dos fármacos , Estradiol/análogos & derivados , Articulações/efeitos dos fármacos , Neovascularização Patológica/tratamento farmacológico , 2-Metoxiestradiol , Inibidores da Angiogênese/farmacologia , Inibidores da Angiogênese/uso terapêutico , Proteínas Angiogênicas/antagonistas & inibidores , Proteínas Angiogênicas/metabolismo , Animais , Anticorpos Monoclonais/toxicidade , Antirreumáticos/farmacologia , Antirreumáticos/uso terapêutico , Artrite Reumatoide/metabolismo , Artrite Reumatoide/fisiopatologia , Vasos Sanguíneos/patologia , Vasos Sanguíneos/fisiopatologia , Reabsorção Óssea/tratamento farmacológico , Reabsorção Óssea/patologia , Cartilagem Articular/efeitos dos fármacos , Cartilagem Articular/patologia , Cartilagem Articular/fisiopatologia , Citocinas/efeitos dos fármacos , Citocinas/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Estradiol/farmacologia , Estradiol/uso terapêutico , Feminino , Articulações/patologia , Articulações/fisiopatologia , Camundongos , Camundongos Endogâmicos BALB C , Neovascularização Patológica/patologia , Neovascularização Patológica/fisiopatologia , Membrana Sinovial/irrigação sanguínea , Membrana Sinovial/efeitos dos fármacos , Membrana Sinovial/fisiopatologia , Sinovite/tratamento farmacológico , Sinovite/patologia , Sinovite/fisiopatologia , Fatores de Tempo , Resultado do TratamentoRESUMO
Clinical studies using the microtubule-targeting agent 2-methoxyestradiol (2ME2; Panzem) in cancer patients show that treatment is associated with clinical benefit, including prolonged stable disease, complete and partial responses, and an excellent safety profile. Studies have shown that 2ME2 is metabolized by conjugation at positions 3 and 17 and oxidation at position 17. To define structure-activity relationships for these positions of 2ME2 and to generate metabolically stable analogues with improved anti-tubulin properties, a series of analogues was generated and three lead analogues were selected, ENMD-1198, ENMD-1200, and ENMD-1237. These molecules showed improved metabolic stability with >65% remaining after 2-h incubation with hepatocytes. Pharmacokinetic studies showed that oral administration of the compounds resulted in increased plasma levels compared with 2ME2. All three analogues bind the colchicine binding site of tubulin, induce G(2)-M cell cycle arrest and apoptosis, and reduce hypoxia-inducible factor-1alpha levels. ENMD-1198 and ENMD-1200 showed improved in vitro antiproliferative activities. Significant reductions in tumor volumes compared with vehicle-treated mice were observed in an orthotopic breast carcinoma (MDA-MB-231) xenograft model following daily oral treatment with all compounds (ANOVA, P < 0.05). Significantly improved median survival time was observed with ENMD-1198 and ENMD-1237 (200 mg/kg/d) in a Lewis lung carcinoma metastatic model (P < 0.05). In both tumor models, the high-dose group of ENMD-1198 showed antitumor activity equivalent to that of cyclophosphamide. ENMD-1198 was selected as the lead molecule in this analogue series and is currently in a phase I clinical trial in patients with refractory solid tumors.
Assuntos
Antineoplásicos/farmacologia , Estrenos/farmacologia , Microtúbulos/efeitos dos fármacos , Moduladores de Tubulina/farmacologia , 2-Metoxiestradiol , Administração Oral , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Antineoplásicos/farmacocinética , Apoptose/efeitos dos fármacos , Ligação Competitiva/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Colchicina/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais , Estradiol/análogos & derivados , Estradiol/química , Estrenos/administração & dosagem , Estrenos/química , Estrenos/farmacocinética , Fase G2/efeitos dos fármacos , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Concentração Inibidora 50 , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mitose/efeitos dos fármacos , Ratos , Análise de Sobrevida , Tubulina (Proteína)/metabolismo , Moduladores de Tubulina/administração & dosagem , Moduladores de Tubulina/química , Moduladores de Tubulina/farmacocinéticaRESUMO
Metastatic castration resistant prostate cancer (mCRPC) relapses due to acquired resistance to docetaxel-based chemotherapy and remains a major threat to patient survival. In this report, we tested the effectiveness of a dual CXCR4/E-selectin antagonist, GM-I1359, in vitro and in vivo, as a single agent or in combination with docetaxel (DTX). This agent was compared to the single CXCR4 antagonist, CTCE-9908, and E-selectin antagonist, GMI-1271. Here we demonstrate that CXCR4 antagonism reduced growth and enhanced DTX treatment in PCa cell lines as well as restored DTX effectiveness in DTX-resistant cell models. The efficacy of dual antagonist was higher respect to those observed for single CXCR4 antagonism. GM1359 impacted bone marrow colonization and growth in intraventricular and intratibial cell injection models. The anti-proliferative effects of GMI-1359 and DTX correlated with decreased size, osteolysis and serum levels of both mTRAP and type I collagen fragment (CTX) in intra-osseous tumours suggesting that the dual CXCR4/E-selectin antagonist was a docetaxel-sensitizing agent for bone metastatic growth. Single agent CXCR4 (CTCE-9908) and E-selectin (GMI-1271) antagonists resulted in lower sensitizing effects compared to GMI-1359. These data provide a biologic rationale for the use of a dual E-selectin/CXCR4 inhibitor as an adjuvant to taxane-based chemotherapy in men with mCRPC to prevent and reduce bone metastases.
Assuntos
Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/secundário , Docetaxel/administração & dosagem , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Bibliotecas de Moléculas Pequenas/administração & dosagem , Animais , Neoplasias Ósseas/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Docetaxel/farmacologia , Sinergismo Farmacológico , Selectina E/antagonistas & inibidores , Glicolipídeos/administração & dosagem , Glicolipídeos/farmacologia , Humanos , Masculino , Camundongos , Células PC-3 , Peptídeos/administração & dosagem , Peptídeos/farmacologia , Neoplasias de Próstata Resistentes à Castração/metabolismo , Receptores CXCR4/antagonistas & inibidores , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: Hepatocellular carcinoma (HCC) represents a highly vascularized tumor entity and the process of angiogenesis is essential for the growth of HCC. Importantly, the pro-angiogenic transcription factors HIF-1alpha and STAT3 have been implicated in HCC progression, thus representing interesting targets for molecular targeted therapy. We hypothesized that therapeutic inhibition of HIF-1alpha could be achieved by using a novel tubulin-binding agent (ENMD-1198). ENMD-1198 is an analog of 2-methoxyestradiol (2ME2) with antiproliferative and antiangiogenic activity. METHODS: The human HCC cell lines HUH-7 and HepG2 were used for experiments. Effects of ENMD-1198 on constitutive and inducible (hypoxia, growth factors) activation of signaling cascades, including HIF-1alpha and STAT3, were investigated by Western blotting. Changes in VEGF expression were determined by real-time PCR. Effects of ENMD-1198 on cancer cell migration and invasion were evaluated in in vitro-assays. The growth-inhibitory effects of ENMD-1198 (200 mg/kg/day) were determined in a subcutaneous tumor model (HUH-7). RESULTS: ENMD-1198 inhibited the phosphorylation of MAPK/Erk, PI-3K/Akt and FAK. Moreover, activation of HIF-1alpha and STAT3 was dramatically reduced by ENMD-1198, which resulted in lower VEGF mRNA expression (P < 0.05). In addition, tumor cell migratory and invasive properties were significantly inhibited (P < 0.05, for both). In vivo, treatment with ENMD-1198 led to a significant reduction in tumor growth, tumor vascularization, and numbers of proliferating tumor cells (P < 0.05 for all). CONCLUSION: The novel microtubule destabilizing agent ENMD-1198 is suitable for inhibiting HIF-1alpha and STAT3 in human HCC cells and leads to reduced tumor growth and vascularization in vivo. Hence, inhibition of HIF-1alpha and STAT3 could prove valuable for therapy of hepatocellular carcinoma.
Assuntos
Antineoplásicos/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Estrenos/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Neovascularização Patológica/tratamento farmacológico , Moduladores de Tubulina/farmacologia , Inibidores da Angiogênese/farmacologia , Carcinoma Hepatocelular/sangue , Carcinoma Hepatocelular/irrigação sanguínea , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/antagonistas & inibidores , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Neoplasias Hepáticas/sangue , Neoplasias Hepáticas/irrigação sanguínea , Neoplasias Hepáticas/patologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator de Transcrição STAT3/antagonistas & inibidores , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Ativação Transcricional/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/biossíntese , Fator A de Crescimento do Endotélio Vascular/genéticaRESUMO
Many human adenocarcinomas can be killed in vitro by targeted cytotoxic T-lymphocytes (CTL); however, major histocompatibility complex (MHC)-restrictions are typically required. The MUC1 antigen is common in many human adenocarcinomas, and is associated with a variable number of tandem repeats. It has been proposed that antigens with such repeated epitopes may be vulnerable to cytotoxic T-lymphocyte killing without MHC-restriction. Therefore, it is possible that MUC1-expressing malignant cells may be killed by targeted cytotoxic T-lymphocyte in the absence of MHC-restriction. In this study, a human MUC1-expressing murine mammary carcinoma cell line was used to determine if cytotoxic T-lymphocyte killing of MUC1-expressing adenocarcinoma cells requires MHC-restriction. Specifically, MUC1-stimulated human mononuclear cells (M1SMC) were observed to kill human MUC1-transfected, MUC1-expressing murine mammary carcinoma cells, but not the mock-transfected, non-MUC1-expressing murine mammary carcinoma cells. Furthermore, the killing was blocked by antibody to MUC1, indicating MUC1-specific killing. In conclusion, cytotoxic T-lymphocyte killing of MUC1-expressing adenocarcinoma cells can be MHC-unrestricted.
Assuntos
Adenocarcinoma/imunologia , Citotoxicidade Imunológica , Leucócitos Mononucleares/imunologia , Complexo Principal de Histocompatibilidade/imunologia , Neoplasias Mamárias Animais/imunologia , Mucina-1/imunologia , Adenocarcinoma/metabolismo , Animais , Anticorpos Antineoplásicos/imunologia , Anticorpos Antineoplásicos/metabolismo , Células Apresentadoras de Antígenos/imunologia , Antígenos de Neoplasias/biossíntese , Antígenos de Neoplasias/imunologia , Antígenos de Neoplasias/metabolismo , Linfócitos T CD4-Positivos/imunologia , Epitopos , Feminino , Humanos , Epitopos Imunodominantes/imunologia , Ativação Linfocitária , Neoplasias Mamárias Animais/metabolismo , Camundongos , Mucina-1/química , Transgenes , Células Tumorais CultivadasAssuntos
Selectina E , Leucemia Mieloide Aguda , Humanos , Tirosina Quinase 3 Semelhante a fms , Leucemia Mieloide Aguda/tratamento farmacológico , Ligantes , Mutação , Inibidores de Proteínas Quinases/farmacologia , Receptores CXCR4/genética , Transdução de Sinais , Sistema de Sinalização das MAP QuinasesRESUMO
PURPOSE: Endostatin is the first endogenous angiogenesis inhibitor to enter clinical trials. Laboratory investigations with endostatin have indicated broad antitumor activity coupled with remarkably low toxicity. A phase I trial of recombinant human endostatin was designed to evaluate toxicity and explore biologic effectiveness in patients with refractory solid tumors. PATIENTS AND METHODS: Endostatin was administered as a 1-hour intravenous infusion given daily for a 28-day cycle. A starting dose of 30 mg/m2 was explored with subsequent dose escalations of 60, 100, 150, 225, and 300 mg/m2. Assessment of serum pharmacokinetics was performed on all 21 patients. Western blot assay and mass spectroscopy were employed to evaluate endostatin metabolism. Circulating levels of endogenous proangiogenic growth factors were examined. Tumor and tumor blood supply were imaged by dynamic computed tomography (CT), magnetic resonance imaging, ultrasound, and positron emission tomography. RESULTS: Endostatin given on this schedule was essentially free of significant drug-related toxicity. Two transient episodes of grade 1 rash were observed. No clinical responses were observed. Endostatin pharmacokinetics were linear with dose, and serum concentrations were achieved that are associated with antitumor activity in preclinical models. No aggregate effect on circulating proangiogenic growth factors were seen, although several patients exhibited persistent declines in vascular endothelial growth factor levels while enrolled in the study. A few patients demonstrated changes in their dynamic CT scans suggestive of a decline in microvessel density, although overall, no consistent effect of endostatin on tumor vasculature was seen. CONCLUSION: Endostatin given daily as a 1-hour intravenous infusion was well tolerated without dose-limiting toxicity at doses up to 300 mg/m2.
Assuntos
Inibidores da Angiogênese/farmacocinética , Colágeno/farmacocinética , Neoplasias/metabolismo , Neovascularização Patológica/metabolismo , Fragmentos de Peptídeos/farmacocinética , Adulto , Idoso , Inibidores da Angiogênese/farmacologia , Western Blotting , Colágeno/farmacologia , Diagnóstico por Imagem , Relação Dose-Resposta a Droga , Endostatinas , Fatores de Crescimento Endotelial/sangue , Feminino , Fator 2 de Crescimento de Fibroblastos/sangue , Humanos , Infusões Intravenosas , Peptídeos e Proteínas de Sinalização Intercelular/sangue , Linfocinas/sangue , Masculino , Pessoa de Meia-Idade , Neoplasias/irrigação sanguínea , Neoplasias/diagnóstico , Neovascularização Patológica/diagnóstico , Fragmentos de Peptídeos/farmacologia , Proteínas Recombinantes , Distribuição Tecidual , Fator A de Crescimento do Endotélio Vascular , Fatores de Crescimento do Endotélio VascularRESUMO
PURPOSE: To perform a phase I trial of recombinant human endostatin (rhEndostatin; EntreMed, Rockville, MD) given as a daily 20-minute intravenous (IV) injection in adult patients with refractory solid tumors. PATIENTS AND METHODS: The daily dose was increased from 15 to 240 mg/m(2) by a factor of 100% in cohorts of three patients. In the absence of dose-limiting toxicity, uninterrupted treatment was continued until the tumor burden increased by more than 50% from baseline. Correlative studies included dynamic contrast-enhanced magnetic resonance imaging of tumor blood flow, urinary vascular endothelial growth factor and basic fibroblast growth factor levels, rhEndostatin serum pharmacokinetics, and monitoring of circulating antibodies to rhEndostatin. RESULTS: There were no notable treatment related toxicities among 15 patients receiving a total of 50 monthly cycles of rhEndostatin. One patient with a pancreatic neuroendocrine tumor had a minor response and two patients showed disease stabilization. Linearity in the pharmacokinetics of rhEndostatin was indicated by dose-proportionate increases in the area under the curve for the first dose and the peak serum concentration at steady state. Daily systemic exposure to rhEndostatin in patients receiving 240 mg/m(2)/d was approximately 50% lower than that provided by the therapeutically optimal dose in preclinical studies. CONCLUSION: rhEndostatin administered as a 20-minute daily IV injection at doses up to 240 mg/m(2) showed no significant toxicities. Evidence of clinical benefit was observed in three patients. Due to high variability between the peak and trough serum concentrations associated with the repeated short IV infusion schedule, daily serum drug levels only briefly exceeded concentrations necessary for in vitro antiangiogenic effects.
Assuntos
Inibidores da Angiogênese/uso terapêutico , Colágeno/uso terapêutico , Neoplasias/tratamento farmacológico , Fragmentos de Peptídeos/uso terapêutico , Adulto , Idoso , Inibidores da Angiogênese/efeitos adversos , Inibidores da Angiogênese/farmacocinética , Colágeno/efeitos adversos , Colágeno/farmacocinética , Creatinina/metabolismo , Endostatinas , Fatores de Crescimento Endotelial/urina , Feminino , Fator 2 de Crescimento de Fibroblastos/urina , Humanos , Infusões Intravenosas , Linfocinas/urina , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Neoplasias/patologia , Fragmentos de Peptídeos/efeitos adversos , Fragmentos de Peptídeos/farmacocinética , Proteínas Recombinantes/uso terapêutico , Fatores de Tempo , Distribuição Tecidual , Fator A de Crescimento do Endotélio Vascular , Fatores de Crescimento do Endotélio VascularRESUMO
PURPOSE: Current combination treatment strategies in malignancy are designed to evaluate the use of cytotoxic drugs and antiangiogenic agents. Endostatin, a fragment of collagen XVIII, specifically inhibits proliferation, migration, and differentiation of endothelial cells in vitro as well as angiogenesis and tumor progression in in vivo models. In this study, we determine the antitumor effect of rhEndostatin administered alone or in combination with Adriamycin against established orthotopic murine mammary carcinoma. EXPERIMENTAL DESIGN: Mice bearing orthotopically established DA-3 mammary adenocarcinoma tumors received varying doses of rhEndostatin alone and in combination with Adriamycin to assess tumor growth inhibition. Additional studies of this in vivo combination included a determination of Adriamycin-induced cardiotoxicity and in vitro effects on human umbilical vein endothelial cell proliferation and cord formation. RESULTS: For single-agent activity, optimal tumor growth inhibition was observed after s.c. administration of 50 mg/kg/day rhEndostatin or 5 mg/kg Adriamycin injected i.v. every 4 days. Combination of Adriamycin with optimal or suboptimal doses of rhEndostatin resulted in synergistic inhibition of DA-3 tumor growth. Importantly, unlike other antiangiogenic agents, rhEndostatin did not exacerbate the cardiotoxicity of Adriamycin. The synergistic interaction between rhEndostatin and Adriamycin was also observed in vitro for inhibition of human umbilical vein endothelial cell proliferation and inhibition of cord formation. CONCLUSIONS: These data suggest that the synergy observed with rhEndostatin in combination with Adriamycin is exerted at the level of the endothelial cell and can result in enhanced tumor growth inhibition. The potential benefit of Adriamycin used in combination with rhEndostatin is being considered for clinical evaluation.
Assuntos
Adenocarcinoma/tratamento farmacológico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Endotélio Vascular/efeitos dos fármacos , Neoplasias Mamárias Experimentais/tratamento farmacológico , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Animais , Divisão Celular/efeitos dos fármacos , Modelos Animais de Doenças , Progressão da Doença , Doxorrubicina/administração & dosagem , Sinergismo Farmacológico , Endostatinas/administração & dosagem , Feminino , Humanos , Técnicas In Vitro , Neoplasias Mamárias Experimentais/metabolismo , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Recombinantes/uso terapêuticoRESUMO
PURPOSE: A clinical study was performed to evaluate the pharmacokinetics (PK) and toxicity of three dose levels of the angiogenesis inhibitor recombinant human (rh) angiostatin when administered twice daily by s.c. injection. EXPERIMENTAL DESIGN: Eligible patients had cancer not amenable to standard treatments. Three groups of 8 patients received 7.5, 15, or 30 mg/m(2)/day divided in two s.c. injections for 28 consecutive days followed by a 7-day washout period. PK assessment was done at days 1 and 28. Thereafter, in absence of toxicity or a 100% increase in tumor size, treatment was continued without interruption. RESULTS: Median age was 53 years (range, 43-75), male:female ratio 10:14, Eastern Cooperative Oncology Group performance 0-1. At the range of doses evaluated, serum PK of all 24 of the patients showed linear relation between dose and area under the curve (0- infinity) and C(max) (reached after 2 h). Thirteen of 24 patients developed erythema at injection sites (11 patients, CTC grade 1; 2 patients, CTC grade 2) without pain or itching, spontaneously resolving within 2-3 weeks of treatment. Two patients went off study after developing hemorrhage in brain metastases, and 2 patients developed deep venous thrombosis. No other relevant treatment-related toxicities were seen, even during prolonged treatment. A panel of coagulation parameters was not influenced by rhAngiostatin treatment. Long-term (>6 months) stable disease (<25% growth of measurable uni- or bidimensional tumor size) was observed in 6 of 24 patients. Five patients received rhAngiostatin treatment for >1 year (overall median time on treatment 99 days). CONCLUSIONS: Long-term twice-daily s.c. treatment with rhAngiostatin is well tolerated and feasible at the selected doses, and merits additional evaluation. Systemic exposure to rhAngiostatin is within the range of drug exposure that has biological activity in preclinical models.
Assuntos
Angiostatinas/farmacocinética , Angiostatinas/uso terapêutico , Antineoplásicos/uso terapêutico , Neoplasias/tratamento farmacológico , Proteínas Recombinantes/farmacocinética , Proteínas Recombinantes/uso terapêutico , Adulto , Idoso , Angiostatinas/efeitos adversos , Formação de Anticorpos , Antineoplásicos/efeitos adversos , Antineoplásicos/farmacocinética , Eritema/induzido quimicamente , Feminino , Humanos , Masculino , Taxa de Depuração Metabólica , Pessoa de Meia-Idade , Neoplasias/irrigação sanguínea , Neoplasias/patologia , Proteínas Recombinantes/efeitos adversosRESUMO
Using a nonviral, electroporation-based gene transfection approach, we demonstrate the efficient and consistent transfection of two poorly immunogenic tumor cell lines: B16F10 melanoma and renal carcinoma (RENCA). Three genes, IL-12, angiostatin (AS), and an endostatin:angiostatin fusion protein (ES:AS) were subcloned into a DNA plasmid containing EBNA1-OriP, which was then transfected into B16F10 and RENCA cells. Significant levels of protein were secreted into the culture supernatants of transfected cells in vitro. Transfected tumor cells were injected subcutaneously into mice. All the three transgenes were capable of significantly delaying and reducing the formation of primary B16F10 and RENCA tumors, as well as B16F10 lung metastases. By day 11 post-injection, all control mice that received either mock-transfected or empty vector DNA-transfected B16F10 tumor cells had developed large primary tumors. In contrast, mice that received IL-12-transfected B16F10 cells did not develop appreciable tumors until day 17, and these were significantly smaller than controls. Similar results were observed for the RENCA model, in which only one of the IL-12 mice had developed tumors out to day 31. Expression of AS or ES:AS also significantly delayed and reduced primary tumors. Overall, ES:AS was more effective than AS alone. Furthermore, 25% of the AS mice and 33% of the ES:AS mice remained tumor-free at day 17, by which point all control mice had significant tumors. Mouse survival rates also correlated with the extent of tumor burden. Importantly, no lung metastases were detected in the lungs of mice that had received either AS or ES:AS-transfected B16F10 tumor cells and significantly fewer metastases were found in the IL-12 group. The consistency of our transfection results highlight the feasibility of directly electroporating tumor cells as a means to screen, identify, and validate in vivo potentially novel antiangiogenic and/or antineoplastic genes.
Assuntos
Carcinoma de Células Renais/metabolismo , Regulação Neoplásica da Expressão Gênica , Neoplasias Renais/metabolismo , Melanoma/metabolismo , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Angiostatinas/biossíntese , Angiostatinas/genética , Animais , Carcinoma de Células Renais/irrigação sanguínea , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/patologia , Divisão Celular/genética , Linhagem Celular Tumoral , Clonagem Molecular , Eletroporação , Endostatinas/biossíntese , Endostatinas/genética , Antígenos Nucleares do Vírus Epstein-Barr/biossíntese , Antígenos Nucleares do Vírus Epstein-Barr/genética , Terapia Genética , Vetores Genéticos , Interleucina-12/biossíntese , Interleucina-12/genética , Neoplasias Renais/irrigação sanguínea , Neoplasias Renais/genética , Neoplasias Renais/patologia , Neoplasias Pulmonares/secundário , Masculino , Melanoma/irrigação sanguínea , Melanoma/genética , Melanoma/patologia , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/genética , Transfecção , Vírus/genéticaRESUMO
Targeting angiogenesis to inhibit tumor development is now considered a valid approach to disease modulation. Recently, a number of laboratories have focused their research on the development of cancer vaccines that target modulators of angiogenesis. In this review we describe a number of novel vaccines that target mediators of angiogenesis and inhibit tumor progression in preclinical models.
Assuntos
Inibidores da Angiogênese/imunologia , Inibidores da Angiogênese/uso terapêutico , Vacinas Anticâncer/uso terapêutico , Neoplasias/terapia , Animais , Humanos , Neoplasias/prevenção & controleRESUMO
PURPOSE: Tumor vascular density may provide a prognostic indicator of metastatic potential or survival. The purpose of this study was to develop 99mTc-ethylenedicysteine-endostatin (99mTc-EC-endostatin) for the evaluation of anti-angiogenesis therapy. METHOD: 99mTc-EC-endostatin was prepared by conjugating ethylenedicysteine (EC) to endostatin, followed by adding pertechnetate and tin chloride. Radiochemical purity was > 95%. In vitro cell viability, affinity and TUNEL assays were performed. Tissue distribution and planar imaging of radiolabeled endostatin were determined in tumor-bearing rats. To assess anti-angiogenic treatment response, rats were treated with endostatin, paclitaxel and saline, followed by imaging with 99mTc-EC-endostatin. Tumor response to endostatin therapy in tumor-bearing animal models was assessed by correlating tumor uptake dose with microvessel density, VEGF, bFGF and IL-8 expression during endostatin therapy. RESULTS: In vitro cell viability and TUNEL assays indicated no marked difference between EC-endostatin and endostatin. Cellular uptake assay suggests that endostatin binds to endostatin receptor. Biodistribution of 99mTc-EC-endostatin in tumor-bearing rats showed increased tumor-to-tissue count density ratios as a function of time. Tumor uptake (%ID/g) of 99mTc-EC-endostatin was 0.2-0.5. Planar images confirmed that the tumors could be visualized clearly with 99mTc-EC-endostatin. The optimal time for imaging using radiolabeled endostatin was 2 hrs. 99mTc-EC-endostatin could assess treatment response. There was a correlation between tumor uptake and cellular targets expression. CONCLUSION: The results indicate that it is feasible to use 99mTc-EC-endostatin to assess efficiency of anti-angiogenesis therapy.
Assuntos
Inibidores da Angiogênese/uso terapêutico , Colágeno/uso terapêutico , Cisteína/análogos & derivados , Cisteína/uso terapêutico , Neoplasias Mamárias Experimentais/irrigação sanguínea , Neovascularização Patológica/tratamento farmacológico , Fragmentos de Peptídeos/uso terapêutico , Inibidores da Angiogênese/farmacocinética , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Colágeno/farmacocinética , Cisteína/farmacocinética , Endostatinas , Fatores de Crescimento Endotelial/metabolismo , Feminino , Fator 2 de Crescimento de Fibroblastos/metabolismo , Marcação In Situ das Extremidades Cortadas , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Interleucina-8/metabolismo , Linfocinas/metabolismo , Neoplasias Mamárias Experimentais/diagnóstico por imagem , Neovascularização Patológica/metabolismo , Paclitaxel/farmacologia , Fragmentos de Peptídeos/farmacocinética , Cintilografia , Ratos , Ratos Endogâmicos F344 , Tecnécio/farmacocinética , Tecnécio/uso terapêutico , Células Tumorais Cultivadas/efeitos dos fármacos , Células Tumorais Cultivadas/metabolismo , Células Tumorais Cultivadas/efeitos da radiação , Fator A de Crescimento do Endotélio Vascular , Fatores de Crescimento do Endotélio VascularRESUMO
ENMD-2076 is a novel orally active, small molecule kinase inhibitor with a mechanism of action involving several pathways key to tumor growth and survival: angiogenesis, proliferation, and the cell cycle. ENMD-2076 has selective activity against the mitotic kinase Aurora A, as well as kinases involved in angiogenesis (VEGFRs, FGFRs). ENMD-2076 inhibited the growth in vitro of a wide range of human solid tumor and hematopoietic cancer cell lines with IC(50) values ranging from 0.025 to 0.7 µmol/L. ENMD-2076 was also shown to induce regression or complete inhibition of tumor growth in vivo at well-tolerated doses in tumor xenograft models derived from breast, colon, melanoma, leukemia, and multiple myeloma cell lines. Pharmacodynamic experiments in vivo showed that in addition to inhibiting Aurora A, single doses of ENMD-2076 had sustained inhibitory effects on the activation of Flt3 as well as the angiogenic tyrosine kinases, VEGFR2/KDR and FGFR1 and 2. ENMD-2076 was shown to prevent the formation of new blood vessels and regress formed vessels in vivo at doses equivalent to those that gave substantial activity in tumor xenograft models. These results indicate that ENMD-2076 is a well-tolerated, orally active multitarget kinase inhibitor with a unique antiangiogenic/antiproliferative profile and provides strong preclinical support for use as a therapeutic for human cancers. Several phase 1 studies involving ENMD-2076 have been recently completed, and the compound is currently being evaluated in a phase 2 clinical trial in patients with platinum-resistant ovarian cancer.
Assuntos
Inibidores da Angiogênese/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Pirazóis/farmacologia , Pirimidinas/farmacologia , Inibidores da Angiogênese/química , Animais , Aurora Quinase A , Aurora Quinases , Linhagem Celular Tumoral , Proliferação de Células , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Nus , Camundongos SCID , Inibidores de Proteínas Quinases/química , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Pirazóis/química , Pirimidinas/química , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
OBJECTIVE: Rheumatoid arthritis (RA) pannus may be dependent on angiogenesis and several critical growth factors including vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF). 2-Methoxyestradiol (2ME2), an endogenous metabolite with low estrogen receptor affinity, has both antiangiogenic and antiproliferative activity. 2ME2 was assessed in the rat collagen-induced arthritis (CIA) model to determine if it could prevent or involute established synovitis. METHODS: Rats were immunized on Day 0 with collagen and randomized to a vehicle control or two 2ME2 prevention arms. In additional studies, multiple parallel treatment arms were initiated at Day 10 after arthritis onset. RESULTS: 2ME2 in preventive protocols at 30 or 100 mg/kg significantly delayed the onset and reduced the severity of clinical and radiographic CIA. In established CIA, oral 2ME2 at 50 mg/kg/bid, 100 mg/kg/day, and 300 mg/kg/day reduced severity compared to vehicle controls. Efficacy of 2ME2 delivery by osmotic pumps at 60 mg/kg/day was equivalent to 300 mg/kg/day by daily gavage. The 3 oral treatment protocols all significantly reduced radiographic scores in a dose-dependent fashion, with the greatest benefit at 300 mg/kg. 2ME2 showed marked suppression of synovial gene expression of proangiogenic bFGF and VEGF, with parallel reduction of synovial blood vessels. Serum antibody levels to native type II collagen were not reduced, suggesting that 2ME2 did not influence humoral immunity. CONCLUSION: Our results indicate that 2ME2 may represent a novel agent for the treatment of inflammatory autoimmune diseases such as RA.