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1.
Blood ; 119(25): 6128-35, 2012 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-22529289

RESUMO

Deficiency of a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13 (ADAMTS13), a VWF-cleaving protease, is the key factor in the pathogenesis of thrombotic thrombocytopenic purpura (TTP), a life-threatening thrombotic microangiopathy. It is well established that ADAMTS13 deficiency results in elevated plasma levels of ultra-large VWF multimers (ULVWF), which are prone to induce platelet aggregation; however, the actual trigger of TTP development remains uncertain. Here we describe a new animal model in which some TTP-like symptoms can be triggered in ADAMTS13 knockout mice by challenge with 2000 units/kg body weight of recombinant human VWF containing ULVWF multimers. Animals rapidly showed clinical symptoms and developed severe thrombocytopenia. Schistocytosis, a decrease in hematocrit, and elevated serum lactate dehydrogenase levels were observed. The heart was identified as the most sensitive target organ with rapid onset of extensive platelet aggregation in the ventricles and myocardial necrosis. Prophylactic administration of 200 units/kg recombinant human ADAMTS13 protected ADAMTS13 knockout mice from developing TTP. Therapeutic administration of 320 units/kg rhADAMTS13 reduced the incidence and severity of TTP findings in a treatment interval-dependent manner. We therefore consider this newly established mouse model of thrombotic microangiopathy highly predictive for investigating the efficacy of new treatments for TTP.


Assuntos
Proteínas ADAM/uso terapêutico , Modelos Animais de Doenças , Metaloendopeptidases/genética , Camundongos Knockout , Púrpura Trombocitopênica Trombótica/tratamento farmacológico , Púrpura Trombocitopênica Trombótica/genética , Proteínas ADAM/administração & dosagem , Proteína ADAMTS13 , Animais , Peso Corporal/efeitos dos fármacos , Relação Dose-Resposta a Droga , Feminino , Hematócrito , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Contagem de Plaquetas , Púrpura Trombocitopênica Trombótica/sangue , Púrpura Trombocitopênica Trombótica/patologia , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/uso terapêutico , Resultado do Tratamento
2.
Semin Thromb Hemost ; 36(5): 522-8, 2010 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-20632249

RESUMO

Von Willebrand factor (VWF) is cleaved by the plasma metalloprotease ADAMTS13 ( A Disintegrin and Metalloproteinase with Thrombo Spondin repeats, number 13) that regulates the hemostatic activity of VWF by limiting its multimeric size in the human system. In vitro and ex vivo studies have shown that human recombinant VWF (rVWF) is virtually resistant to the proteolytic activity of murine ADAMTS13. In contrast, rabbit and cynomolgus ADAMTS13 is able to cleave human rVWF. These findings were consistent with in vivo results showing distinct pharmacological behavior of human rVWF depending on the cleaving capacity of ADAMTS13 present in the species tested. Studies were performed using three mouse strains (ADAMTS13 deficient, C57BL/6J [wild type], VWF deficient), rats, rabbits, and cynomolgus monkeys. All animals were infused once with different doses of human rVWF and, in addition, 14 daily doses were given to rats and cynomolgus monkeys. Exaggerated pharmacological effects were observed in mice, with the ADAMTS13 knockout mouse being the most sensitive strain. Similar findings with decreased incidence and severity were seen in normal C57BL/6J mice and also in VWF-deficient mice, where they were least pronounced. In rats, exaggerated pharmacological effects were observed only after 14 doses. Rabbits and cynomolgus monkeys showed no exaggerated pharmacological effects. These differences between species and between mouse strains suggest that the efficiency of ADAMTS13 to cleave rVWF determines the severity of clinical, laboratory and pathohistological findings. These observations highlight the importance of evaluating species' suitability for the generation of meaningful preclinical data for determining the therapeutic safety margins for human patients. Only animals with a sufficient rVWF cleavage capacity by endogenous ADAMTS13 (rabbits and cynomolgus monkeys) are considered appropriate animal models for preclinical evaluation of the rVWF product.


Assuntos
Proteínas ADAM/metabolismo , Fator de von Willebrand/farmacologia , Animais , Humanos , Proteínas Recombinantes/farmacologia
3.
Eur J Pharmacol ; 820: 206-216, 2018 Feb 05.
Artigo em Inglês | MEDLINE | ID: mdl-29274331

RESUMO

New therapeutic agents are needed to overcome the toxicity and suboptimal efficacy observed in current treatment of glomerulonephritis (GN). BaxB01 is a fully human monoclonal antibody targeting a disease-related immunologically distinct isoform of Macrophage migration Inhibitory Factor (MIF), designated oxidized MIF (oxMIF) and locally expressed in inflammatory conditions. We report the pharmacokinetic profile of BaxB01, and its dose and exposure-related disease-modifying activity in experimentally induced rat GN. BaxB01 bound to rat oxMIF with high affinity and reduced rat macrophage migration in vitro. After intravenous administration in rats, BaxB01 demonstrated favorable pharmacokinetics, with a half-life of up to nine days. Disease modification was dose-related (≥ 10mg/kg) as demonstrated by significantly reduced proteinuria and diminished histopathological glomerular crescent formation. Importantly, a single dose was sufficient to establish an exposure-related, anti-inflammatory milieu via amelioration of glomerular cellular inflammation. Pharmacodynamic modeling corroborated these findings, consistently predicting plasma exposures that were effective in attenuating both anti-inflammatory activity and reducing loss of kidney function. This pharmacologic benefit on glomerular function and structure was sustained during established disease, while correlation analyses confirmed a link between the antibody's anti-inflammatory activity and reduced crescent formation in individual rats. Finally, safety assessment in rats showed that the experimental therapeutic was well tolerated without signs of systemic toxicity or negative impact on kidney function. These data define therapeutically relevant exposures correlated with mechanism-based activity in GN, while toxicological evaluation suggests a large therapeutic index and provides evidence for achieving safe and effective exposure to a MIF isoform-directed therapeutic in nephritis-associated disease.


Assuntos
Anticorpos Monoclonais/farmacocinética , Anticorpos Monoclonais/uso terapêutico , Glomerulonefrite/tratamento farmacológico , Glomerulonefrite/imunologia , Fatores Inibidores da Migração de Macrófagos/imunologia , Terapia de Alvo Molecular , Segurança , Animais , Anticorpos Monoclonais/efeitos adversos , Anticorpos Monoclonais/imunologia , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Progressão da Doença , Feminino , Glomerulonefrite/metabolismo , Humanos , Glomérulos Renais/efeitos dos fármacos , Glomérulos Renais/patologia , Masculino , Monócitos/citologia , Monócitos/efeitos dos fármacos , Isoformas de Proteínas/imunologia , Ratos
4.
Haematologica ; 91(10): 1336-42, 2006 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-17018382

RESUMO

BACKGROUND AND OBJECTIVES: Homozygous beta-thalassemia patients may develop iron overload even if untransfused, due to inappropriately high intestinal iron absorption. Reduction of hepcidin synthesis has been reported both in patients and in animal models. We have measured liver hepcidin and other iron gene transcripts in two different mouse models of beta-thalassemia at different ages. DESIGN AND METHODS: Mice Hbb(th/th), characterized by spontaneous homozygous deletion of the major b1 globin gene were studied at 2 and 8 months. Mice Hbb(th/3+), characterized by the heterozygous deletion of b1 and b2 globin genes were studied at 4 and 10 months. Hematologic data were obtained and iron overload estimated by Perls' staining of the liver. Expression of liver hepcidin, Tfr2, Hjv, Fpn and Hfe RNA was assessed by real-time polymerase chain reaction. Levels of serum cytokines (interleukin-6, IL-1beta, IL-10, granulocyte-macrophage colony-stimulating factor) levels were assayed by enzyme-linked immunosorbent assay. RESULTS: Hemoglobin, hematocrit and mean corpuscular volume were significantly reduced in both beta-thalassemia models, more significantly in Hbb(th/3+), which have the greater, age-dependent, iron overload. Hepcidin RNA was not increased despite iron overload in both strains. Fpn RNA was increased and Tfr2 was decreased in older animals. Inflammatory cytokine levels were striking variable and unrelated to hepcidin levels. INTERPRETATION AND CONCLUSIONS: Although anemia is reported to inhibit hepcidin expression, normal hepcidin synthesis was maintained in both thalassemic models studied. However, hepcidin levels were inappropriate for the body iron, especially in Hbb(th/3+) 10-month-old animals. As we previously reported in wild type mice after parenteral iron overload, Tfr2 is reduced and Fpn RNA increased in thalassemic mice. Inflammatory cytokines did not play a major role in increasing hepcidin levels or in modifying iron homeostasis in this study.


Assuntos
Peptídeos Catiônicos Antimicrobianos/biossíntese , Peptídeos Catiônicos Antimicrobianos/genética , Fígado/metabolismo , Talassemia beta/genética , Talassemia beta/metabolismo , Animais , Modelos Animais de Doenças , Feminino , Regulação da Expressão Gênica , Hepcidinas , Ferro/metabolismo , Sobrecarga de Ferro/genética , Sobrecarga de Ferro/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Camundongos Knockout , Camundongos Mutantes
5.
Expert Rev Clin Pharmacol ; 8(2): 163-77, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25660348

RESUMO

Nonacog gamma is a new recombinant factor IX to treat factor IX deficiency. It is indicated for control of bleeding episodes, perioperative management and routine prophylaxis to prevent or reduce the frequency of bleeding episodes in adults and children with hemophilia B. Nonacog gamma was first approved in the USA in June 2013 under the trade name RIXUBIS followed by market approvals in Australia and the EU in 2014, and marketing authorization decision is pending in Japan. Nonacog gamma is derived from a recombinant Chinese hamster ovary cell line using a state of the art biotechnological manufacturing process. Recombinant factor IX is produced by Baxter's protein-free fermentation technology, which was first developed for ADVATE. The product is purified and formulated in the absence of any human or animal-derived protein. Nonacog gamma was characterized both in comprehensive in vitro and in vivo non-clinical studies as well as in an extensive clinical trial program.


Assuntos
Fator IX/uso terapêutico , Hemofilia B/tratamento farmacológico , Hemostáticos/uso terapêutico , Adulto , Animais , Células CHO , Criança , Cricetinae , Cricetulus , Humanos , Proteínas Recombinantes/uso terapêutico
6.
Int J Hematol ; 98(5): 525-32, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-24061775

RESUMO

Baxter has developed a new recombinant factor IX (rFIX) drug product (BAX326) for treating patients with hemophilia B, or congenital FIX deficiency. An extensive preclinical program evaluated the pharmacokinetics, efficacy, and safety of BAX326 in different species. The efficacy of BAX326 was tested in three mouse models of primary pharmacodynamics: tail-tip bleeding, carotid occlusion, and thrombelastography. The pharmacokinetics was evaluated after a single intravenous bolus injection in mice, rats, and macaques. Toxicity was assessed in rats and macaques, safety pharmacology in rabbits and macaques, and immunogenicity in mice. BAX326 was shown to be efficacious in all three primary pharmacodynamic studies (P ≤ 0.0076). Hemostatic efficacy was dose related and similar for the three lots tested. Pharmacokinetic results showed that rFIX activity and rFIX antigen concentrations declined in a bi-phasic manner, similar to a previously licensed rFIX product. BAX326 was well tolerated in rabbits and macaques at all dose levels; no thrombogenic events and no adverse clinical, respiratory, or cardiovascular effects occurred. BAX326 was also shown to have a similar immunogenicity profile to the comparator rFIX product in mice. These results demonstrate that BAX326 has a favorable preclinical safety and efficacy profile, predictive of a comparable effect to that of the previously licensed rFIX in humans.


Assuntos
Fator IX/farmacologia , Hemofilia B/tratamento farmacológico , Proteínas Recombinantes/farmacologia , Animais , Tempo de Sangramento , Coagulação Sanguínea/efeitos dos fármacos , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Fator IX/administração & dosagem , Fator IX/efeitos adversos , Hemofilia B/sangue , Humanos , Macaca , Masculino , Camundongos , Coelhos , Ratos , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/efeitos adversos , Tromboelastografia , Resultado do Tratamento
7.
Eur J Pharm Sci ; 47(5): 979-87, 2012 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-23046836

RESUMO

As to the alignment of "Horizon 2020", ir is a more integrated approach to European science policy than expressed in the proposals previously drafted, and specifically considers: (i) promoting excellence in Science, (ii) establishing a sound industrial leadership and (iii) expressing an ambition to address current and future societal challenges. In this respect, the quest for a knowledge-based economy in Europe should result in proposals for industrial and employment policies that will consolidate the major European advantages in the biomedical, healthcare and pharmaceutical sectors. Horizon 2020 also provides the possibility of adopting a more flexible and simplified management route to drive European research through innovation, research and development. What should be additionally considered? Unmet medical needs, under pressure from demographic changes, await the generation of new medicines and health technologies which will evolve into a driver for a unified European policy. We believe that this should be focused on harnessing pharmaceutical knowledge for clinical use, as part of a response to accommodate patient needs and economic growth based on a robust, scientific approach. The bolder ambition for European research is to unlock key bottlenecks currently undermining European competitiveness. The historical lack of an appropriate business/innovation environment with reduced access to adequate risk finance instruments has severed the path for economic growth and industrial development. These issues are of critical importance and a solution is urgently needed to foster translation from the university to the healthcare sector through the generation and support of start-ups, spin-offs, university-industry consortia, and other platforms, which support translational research. The ultimate goal is implementation of holistic programmes: the 'bench to bedside' paradigm of medicines and other healthcare products. The European Research Council supports the basic biomedical research programmes of long term importance for development of medicines; however, fundamental research initiatives on medicines development will not be competitive in such an environment. In order to strengthen the long term outlook, we must foster innovative research within the university sector, EUFEPS proposes that a fund for such research be set up within Horizon 2020, which would be open for individual research groups and which would include Public-Public Partnerships (complementing already existing Public-Private Partnerships). How do we look for implementation? There is an established research agenda for medicines research that is globally focused, and which incorporates a cooperative model between universities and industry, facilitating integration of complex technologies. Regulatory Science will play an important role in this integration. This agenda uses tools arising from systems approaches (including discovery with systems biology and also systems pharmacology) and has the potential for providing better knowledge management, as well as technological innovation (including manufacturing). It also addresses the drive towards personalised medicines and can, with support from both public and private sectors, foster translation of knowledge to new technologies and from that, to new medicinal products and complex integrated systems. This is a part of a strategy capable of solving unmet medical needs, which would increase the quality of life of patients suffering from chronic and debilitating diseases. The instruments to allow the development of a research agenda should strengthen existing partnerships such as the IMI-JU model; allow for the creation of European-network infrastructures that can bring together existing competences with adequate European coordination, thus promoting advanced training and continuous professional development for the pharmaceutical sciences. This will be the cornerstone of a knowledge management strategy, providing education and training for healthcare professionals and scientists. A key role for EUFEPS is to help the research community to embrace these new holistic policies applied to the spectrum of pharmaceutical, medical and cognate sciences.


Assuntos
Pesquisa Biomédica , Descoberta de Drogas , Indústria Farmacêutica , Europa (Continente) , Parcerias Público-Privadas , Universidades
10.
Am J Physiol Lung Cell Mol Physiol ; 291(3): L457-65, 2006 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-16603592

RESUMO

Nitric oxide (NO) is a potential new therapeutic agent for sickle cell disease (SCD). We investigated the effects of NO donor on hypoxia-induced acute lung injury that occurs when transgenic sickle cell SAD mice are exposed to chronic hypoxia, a model for lung vasoocclusive sickle cell events. In wild-type and SAD mice, intraperitoneal injection of S-nitrosoalbumin (NO-Alb) produced no significant hematologic changes under room air conditions, whereas it induced mild temporary hypotension and inhibition of platelet aggregation. NO-Alb administration (300 mg/kg ip twice a day, equivalent to 7.5 microM NO) in wild-type and SAD mice exposed to 46 h of hypoxia (8% oxygen) followed by 2 h of normoxia resulted in 1) reduction of the hypoxia-induced increase in blood neutrophil count, 2) prevention of hypoxia-induced increased IL-6 and IL-1beta levels in bronchoalveolar lavage, 3) reduction of the lung injury induced by hypoxia-reoxygenation, 4) prevention of thrombus formation, and 5) prevention of hypoxia-induced increase of lung matrix metalloproteinase-9 gene expression. These effects provide new insights into the possible use of NO-Alb in the treatment of acute lung injury in SCD.


Assuntos
Anemia Falciforme/complicações , Hipóxia/complicações , Compostos Nitrosos/farmacologia , Pneumopatia Veno-Oclusiva/prevenção & controle , Traumatismo por Reperfusão/complicações , Soroalbumina Bovina/farmacologia , Animais , Animais Geneticamente Modificados , Modelos Animais de Doenças , Avaliação de Medicamentos , Feminino , Hemodinâmica/efeitos dos fármacos , Masculino , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Compostos Nitrosos/uso terapêutico , Agregação Plaquetária/efeitos dos fármacos , Soroalbumina Bovina/uso terapêutico
11.
Arzneimittelforschung ; 54(2): 95-101, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-15038458

RESUMO

To determine whether the beneficial effects of catecholamines on the variables of hemorrhagic hypovolemic shock are augmented by coadministration of alpha1-acid glycoprotein during resuscitation, alpha1-acid glycoprotein (200 mg/kg), a placebo formulation or Ringer's solution was infused in a rat model of hemorrhagic hypovolemic shock for 1 h concomitantly with either norepinephrine (CAS 51-40-1; 0.1, 0.3, 1 microg x kg(-1) x min(-1)) or dopamine (CAS 62-31-7; 5, 10, 15 microg x kg(-1) x min(-1)). Resuscitation with norepinephrine or dopamine alone was continued for a further 4 h. Mean arterial blood pressure, cardiac output, stroke volume, heart rate and total peripheral vascular resistance were measured during the entire 5-h period. The combination of dopamine or norepinephrine with alpha1-acid glycoprotein more effectively restored mean arterial blood pressure and cardiac output than analogous combinations with placebo formulation or Ringer's solution. So co-administration with alpha1-acid glycoprotein considerably augments the beneficial effects of catecholamines on the main variables of hemorrhagic hypovolemic shock.


Assuntos
Catecolaminas/farmacologia , Orosomucoide/farmacologia , Choque Hemorrágico/tratamento farmacológico , Animais , Pressão Sanguínea/efeitos dos fármacos , Débito Cardíaco/efeitos dos fármacos , Cardiotônicos/uso terapêutico , Dopamina/uso terapêutico , Relação Dose-Resposta a Droga , Combinação de Medicamentos , Hidratação , Frequência Cardíaca/efeitos dos fármacos , Humanos , Masculino , Norepinefrina/uso terapêutico , Ratos , Ratos Sprague-Dawley , Ressuscitação , Choque Hemorrágico/fisiopatologia , Volume Sistólico/efeitos dos fármacos , Resistência Vascular/efeitos dos fármacos , Vasoconstritores/uso terapêutico
12.
Semin Thromb Hemost ; 28(2): 215-26, 2002 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-11992244

RESUMO

We used a canine and a murine model of von Willebrand disease (vWD) to study the in vivo effects of recombinant von Willebrand factor (vWF). Two preparations were used: (1) a fully processed mature vWF; this was achieved by coexpression of furin. (2) A preparation containing unprocessed pro-vWF, the propeptide still covalently linked to mature vWF. Both preparations induced an increase in canine and murine factor VIII:C (FVIII), which was sustained even when vWF antigen had been removed from the circulation. vWF multimers were analyzed in the plasma samples after infusion using ultra high-resolution 3% agarose gels to allow the separation of homoforms and heteroforms of the vWF polymers. Administration of pro-vWF to dogs with severe vWD resulted in the removal of the propeptide and maturation of vWF in the circulation, indicating that the propeptide cleavage from unprocessed vWF can occur extracellularly. This suggests that the vWF propeptide, besides being derived from the Weibel-Palade bodies of endothelial cells after stimulation, can also be cleaved by pro-vWF in plasma. Using a murine model of vWD, the involvement of the low-density lipoprotein receptor-related protein (LRP) in the clearance of FVIII was established. The low levels of FVIII observed in the absence of vWF are due to an enhanced clearance of FVIII by binding to LRP and removal from the circulation through endocytosis. Administration of the receptor-associated protein (RAP) as a recombinant fusion protein to vWF knockout mice significantly improved the in vivo recovery of recombinant FVIII and the survival time of otherwise rapidly cleared FVIII.


Assuntos
Técnicas de Sonda Molecular , Doenças de von Willebrand/tratamento farmacológico , Fator de von Willebrand/farmacologia , Animais , Dimerização , Modelos Animais de Doenças , Cães , Fator VIII/efeitos dos fármacos , Fator VIII/metabolismo , Humanos , Infusões Parenterais , Proteína Associada a Proteínas Relacionadas a Receptor de LDL , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade , Camundongos , Camundongos Knockout , Precursores de Proteínas/administração & dosagem , Precursores de Proteínas/metabolismo , Precursores de Proteínas/farmacocinética , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/farmacologia , Relação Estrutura-Atividade , Doenças de von Willebrand/metabolismo , Fator de von Willebrand/administração & dosagem
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