RESUMO
INTRODUCTION: MC710, a 1:10 protein weight ratio mixture of plasma-derived activated factor VII (FVIIa) and factor X (FX), is a novel bypassing agent for haemostasis in haemophilia patients with inhibitors. We evaluated the haemostatic efficacy and safety of one to two administrations of MC710 in 21 joint, muscle, and subcutaneous bleeding episodes in 14 male patients, in a multi-centre, open-label, non-randomized clinical trial. METHODS: Subjects were intravenously administered one or two doses of 60 or 120 µg kg-1 MC710 (as FVIIa) once or twice (to a maximum of 180 µg kg-1 ) over up to five bleeding episodes per subject. The haemostatic efficacy of MC710 was determined for each episode by investigator evaluation, using changes in visual analogue scale (VAS) for pain relief, and/or knee joint or muscle circumference for swelling reduction, and range of motion (ROM) for improvement of joint mobility. RESULTS: In 21 treatments for bleeding episodes, 19 were rated "excellent" or "effective" 8 h after the last treatment. VAS significantly decreased over time, and ROM significantly improved over time compared with the values before treatment. One mild adverse reaction, decreased blood potassium, and two serious adverse events, both knee joint bleeding, were observed within 1 week after first administration, with no significant effect on safety. Furthermore, diagnostic markers did not show any signs of disseminated intravascular coagulation (DIC). CONCLUSION: These results show that MC710 has sufficient haemostatic efficacy and safety, and can be used as a potential bypassing agent to control bleeding in haemophilia patients with inhibitors.
Assuntos
Fator VIIa/uso terapêutico , Fator X/uso terapêutico , Hemofilia A/tratamento farmacológico , Adolescente , Adulto , Humanos , Masculino , Adulto JovemRESUMO
MC710, a mixture of plasma-derived activated factor VII and factor X at a protein weight ratio of 1:10, is a novel bypassing agent for haemostasis in haemophilia patients with inhibitors. In a Phase II trial, we evaluated the haemostatic efficacy and safety of single doses of MC710, and investigated pharmacokinetic and pharmacodynamic parameters in nine joint bleeding episodes in six male haemophilia patients with inhibitors. This trial was a multi-centre, open-label, non-randomized study of two doses (60 and 120 µg kg(-1) as FVIIa dose), allowing the re-administration of different MC710 dosages to the same subjects. Haemostatic efficacy was assessed by evaluating reduction in pain and swelling, as well as increase in range of motion in a bleeding joint. The results of the study showed that in nine bleeding episodes, seven treatments were rated as 'excellent' or 'effective' according to investigator's rating system of efficacy at 8 h after administration. No serious or severe adverse events were observed after administration; furthermore, measurement of several diagnostic markers revealed no signs or symptoms of disseminated intravascular coagulation (DIC). The haemostatic potential of MC710 was confirmed at doses of 60 and 120 µg kg(-1) in this trial. MC710 is thus expected to be a safe and efficacious novel bypassing agent for controlling bleeding in haemophilia patients with inhibitors.
Assuntos
Inibidores dos Fatores de Coagulação Sanguínea/sangue , Coagulantes/uso terapêutico , Fator VIIa/uso terapêutico , Fator X/uso terapêutico , Hemofilia A/tratamento farmacológico , Hemofilia B/tratamento farmacológico , Adolescente , Adulto , Coagulantes/farmacocinética , Esquema de Medicação , Quimioterapia Combinada , Fator VIIa/farmacocinética , Fator X/farmacocinética , Meia-Vida , Hemorragia/prevenção & controle , Humanos , Masculino , Tempo de Tromboplastina Parcial , Tempo de Protrombina , Adulto JovemRESUMO
Severe heritable protein C (PC) deficiency is quite rare, although heterozygous PROC mutation is the second leading cause of genetic predisposition to thrombosis in Japanese adults. The aim of the study was to search the optimal management, the paediatric onset and outcomes of PC deficiency were characterized in Japan. The genetic study, postmarketing survey of activated PC(aPC) concentrate (Anact(®)C) and intensive review in Japan for 20 years enabled the analysis of the disease onset, genotype, treatment and prognosis. Symptomatic PC deficiency was determined in 27 Japanese children. All but two patients presented within 16 days after birth (three prenatal and six neonatal onsets). Postnatal-onset cases had normal growth at full-term delivery. Of the 27 patients, 19 suffered intracranial thrombosis or haemorrhage (ICTH) (three foetal hydrocephalies), 16 developed purpura fulminans (PF) and 10 had both at the first presentation. ICTH preceded PF in both affected cases. Low PC activities of 18 mothers and/or 12 fathers indicated 20 inherited PC deficiencies (2 homozygotes, 11 compound heterozygotes and 7 heterozygotes) and seven unidentified causes of PC deficiency. Nine of 11 patients studied had PROC mutations. Four unrelated patients (50%) carried PC nagoya (1362delG). No PC-deficient parents had experienced thromboembolism. Of the 18 patients with aPC therapy, two died and eight evaluable survivors had neurological sequelae. This first comprehensive study of paediatric PC deficiency suggested that perinatal ICTH was the major presentation, occurring earlier than neonatal PF. PC nagoya was prevalent in paediatric, but not adult, patients in Japan. Early maternal screening and optimal PC therapy are required for newborns at risk of PC deficiency.
Assuntos
Deficiência de Proteína C/tratamento farmacológico , Proteína C/uso terapêutico , Adolescente , Anticoagulantes/uso terapêutico , Criança , Pré-Escolar , Feminino , Genótipo , Heterozigoto , Homozigoto , Humanos , Lactente , Recém-Nascido , Japão , Masculino , Proteína C/genética , Deficiência de Proteína C/genética , Deficiência de Proteína C/patologia , Púrpura Fulminante/tratamento farmacológico , Púrpura Fulminante/patologia , Trombose/tratamento farmacológico , Trombose/patologia , Resultado do Tratamento , Tromboembolia Venosa/tratamento farmacológico , Tromboembolia Venosa/patologiaRESUMO
We reported the results of a clinical pharmacological study of MC710 (a mixture of plasma-derived FVIIa and FX) in haemophilia patients with inhibitors during a non-haemorrhagic state. This report provides the results of a clot waveform analysis (CWA) and thrombin generation test (TGT) using blood samples obtained in this study. CWA and TGT were conducted using blood samples obtained from a pharmacokinetic and pharmacodynamic study in which MC710 (five dose rates: 20, 40, 80, 100 and 120 µg kg(-1)) was compared with NovoSeven (120 µg kg(-1)) and FEIBA (two dose rates: 50 and 75 U kg(-1)) as control drugs in 11 haemophilia patients with inhibitors without haemorrhagic symptoms. CWA showed that MC710 provided significantly greater improvement than the control drugs in activated partial thromboplastin time (APTT) at 80 µg kg(-1); maximum clot velocity and maximum clot acceleration were more enhanced by MC710 than by control drugs. TGT revealed that MC710 significantly shortened the initiation time of thrombin generation in comparison to FEIBA and induced greater thrombin generation potency than NovoSeven. It was not clear whether or not MC710 caused significant dose-dependent changes in the two measurements; however, differences between MC710 and the control drugs were clarified. MC710 was confirmed to have superior coagulation activity and thrombin productivity and is expected to have superior bypassing activity.
Assuntos
Coagulação Sanguínea/efeitos dos fármacos , Fator VIIa/farmacologia , Fator X/farmacologia , Hemofilia A/tratamento farmacológico , Hemofilia B/tratamento farmacológico , Adolescente , Adulto , Inibidores dos Fatores de Coagulação Sanguínea/sangue , Testes de Coagulação Sanguínea/métodos , Estudos Cross-Over , Relação Dose-Resposta a Droga , Quimioterapia Combinada , Hemofilia A/sangue , Hemofilia B/sangue , Humanos , Japão , Masculino , Trombina/metabolismo , Adulto JovemRESUMO
MC710, a combined product of plasma-derived activated factor VII (FVIIa) and factor X (FX) at a protein weight ratio of 1:10, is a novel bypassing agent for haemostasis in haemophilia patients with inhibitors. In this study, pharmacokinetic (PK), pharmacodynamic (PD) parameters and safety of single doses of MC710 were investigated in 11 male haemophilia patients with inhibitors in a non-bleeding state. This was a multi-centre, open-labelled, non-randomized, active controlled crossover, dose-escalation study of five doses (20-120 µg kg(-1) of FVIIa) with re-administration of different MC710 dosages to the same subjects. The active controls were NovoSeven (120 µg kg(-1)) and/or FEIBA (50 and 75 U kg(-1)) which were used to compare PD parameters. The area under the curve (AUC) and maximum plasma concentration (C(max)) of MC710 active ingredients increased dose-dependently within the range of 20 and 120 µg kg(-1). After administration of MC710, activated partial thromboplastin time (APTT) was dose-dependently improved and prothrombin time (PT) was shortened to approximately 6 s at 10 min, and APTT improvement and PT shortening effects were maintained until 12 h after administration of MC710 at all doses. No serious or severe adverse event was observed after administration of MC710; furthermore, several diagnostic marker values and those changes did not indicate any signs of disseminated intravascular coagulation (DIC). These results suggest that MC710 would have haemostatic potential equal to or greater than NovoSeven and FEIBA and was be tolerable when given at doses up to 120 µg kg(-1).
Assuntos
Fator VIIa/farmacologia , Fator X/farmacologia , Hemofilia A/tratamento farmacológico , Adolescente , Adulto , Área Sob a Curva , Coagulação Sanguínea/efeitos dos fármacos , Estudos Cross-Over , Relação Dose-Resposta a Droga , Quimioterapia Combinada/métodos , Fator VIIa/farmacocinética , Fator X/farmacocinética , Humanos , Masculino , Tempo de Tromboplastina Parcial , Tempo de Protrombina , Adulto JovemRESUMO
Studies conducted in European and North American countries have demonstrated that various factors including races affect the frequency of inhibitor formation in haemophilia patients. The present study was undertaken to analyse factors affecting the incidence of inhibitor formation in Japanese haemophilia A and B patients. Analytical data were retrospectively collected from haemophilia A and B patients born after 1988, the year when monoclonal antibody-purified factor VIII products were first marketed in Japan. Various data were collected from 184 patients (153 cases of haemophilia A; 31 cases of haemophilia B). The sample size of haemophilia B cases was too small to reveal any significant differences between the inhibitor formation group and the inhibitor-free group in any of background variables. For patients with haemophilia A, on the other hand, univariate analysis identified the severity of haemophilia and a positive family history of inhibitor development as risk factors for the formation of inhibitors. In analyses of the clotting factor products used, the incidence of inhibitor formation did not differ significantly between the group treated with plasma-derived products (29.7%) and the group treated with recombinant products (25.0%). When background variables were compared, age was higher in the group treated with plasma-derived products but none of the other background variables differed between the two groups. These results suggest that in Japanese haemophilia patients, the type of clotting factor preparations used for therapy has not influenced the incidence of inhibitor formation.
Assuntos
Coagulantes/imunologia , Fator IX/imunologia , Fator VIII/imunologia , Hemofilia A/imunologia , Hemofilia B/imunologia , Isoanticorpos/imunologia , Adolescente , Adulto , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/uso terapêutico , Criança , Pré-Escolar , Coagulantes/uso terapêutico , Fator IX/uso terapêutico , Fator VIII/uso terapêutico , Feminino , Hemofilia A/tratamento farmacológico , Hemofilia B/tratamento farmacológico , Humanos , Japão , Masculino , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/uso terapêutico , Estudos Retrospectivos , Fatores de Risco , Adulto JovemRESUMO
We conducted a questionnaire survey of haemophilia treaters participating in the Fourth Seminar on Regular Replacement Therapy (sponsored by Baxter Bioscience, 4 March 2006) to clarify the current status (up to January 2006) of replacement therapy for haemophilia. The haemophilia treaters including medical doctor, nurse belonged to 48 institutions located in the 23 prefectures of Japan. Topics included age at the initiation of regular replacement therapy (prophylaxis), and expected future situation of patients who are currently receiving prophylaxis. Data were collected from 1267 patients with haemophilia A and 273 patients with haemophilia B who had been treated at the represented institutions. Of these haemophilia A and B patients, 23% and 16% had received a prophylactic treatment regimen respectively. A breakdown of each disease by severity demonstrated that of the patients with severe haemophilia A and B patients, 27% and 18% of patients received a prophylaxis treatment, compared to 17% and 19% of patients with moderate type, and 1% and 3% of patients with mild type respectively. Of those severe haemophilia A and B patients receiving prophylaxis, the percentage of primary prophylaxis, which means prophylaxis begins under 2 years of age, was still small for 24% and 29% respectively. However, approximately half of the patients received prophylaxis during the age of 2-14 years, which suggests that secondary prophylaxis is widely spread in the age group in Japan. Problems in introduction of prophylaxis include difficulty in peripheral venous access, a lack of understanding of the therapy by the caregiver. In addition, the fear of inhibitor development, as well as the psychological anxiety in paediatric patients, was also mentioned as barriers to initiating and continuing prophylaxis.
Assuntos
Hemartrose/prevenção & controle , Hemofilia A/tratamento farmacológico , Hemofilia B/tratamento farmacológico , Adolescente , Fatores Etários , Criança , Pré-Escolar , Uso de Medicamentos/estatística & dados numéricos , Fator IX/uso terapêutico , Fator VIII/uso terapêutico , Pesquisas sobre Atenção à Saúde , Hemartrose/etiologia , Hemofilia A/complicações , Hemofilia B/complicações , Humanos , Lactente , Japão , Masculino , Padrões de Prática Médica/estatística & dados numéricos , Índice de Gravidade de Doença , Adulto JovemRESUMO
Biogenic polyamines, putrescine, spermidine, and spermine, are ubiquitous cellular cations and exert multiple biological functions. Polyamine analogues mimic biogenic polyamines at macromolecular level but are unable to substitute for natural polyamines and maintain cell proliferation, indicating biomedical applications. The mechanistic differences in DNA binding mode between natural and synthetic polyamines have not been explored. The aim of this study was to examine the interaction of calf thymus DNA with three polyamine analogues, 1,11-diamino-4,8-diazaundecane (333), 3,7,11,15-tetrazaheptadecane x 4 HCl (BE-333), and 3,7,11,15,19-pentazahenicosane x 5 HCl (BE-3333), using FTIR, UV-visible, and CD spectroscopy. Polyamine analogues bind with guanine and backbone PO2 group as major targets in DNA, whereas biogenic polyamines bind to major and minor grooves as well as to phosphate groups. Weaker interaction with DNA was observed for analogues with respect to biogenic polyamines, with K(333) = 1.90 (+/-0.5) x 10(4) M(-1), K(BE-333) = 6.4 (+/-1.7) x 10(4) M(-1), K(BE-3333) = 4.7 (+/-1.4) x 10(4) M(-1) compared to K(Spm) = 2.3 (+/-1.1) x 10(5) M(-1), K(Spd) = 1.4 (+/-0.6) x 10(5) M(-1), and K(Put) = 1.02 (+/-0.5) x 10(5) M(-1). A partial B- to A-DNA transition was also provoked by analogues. These data suggest distinct differences in the binding of natural and synthetic polyamines with DNA.
Assuntos
Antineoplásicos/farmacologia , DNA/química , Poliaminas/química , Animais , Antineoplásicos/química , Bovinos , Dicroísmo Circular , Humanos , Concentração de Íons de Hidrogênio , Cinética , Modelos Químicos , Neoplasias/tratamento farmacológico , Fosfatos/química , Espectrofotometria/métodos , Espectroscopia de Infravermelho com Transformada de FourierRESUMO
The aim of this study was to investigate the enhancing effect of polyamines on intestinal absorption of fluorescein isothiocyanate-labeled dextran (MW 4400, FD-4) in the in situ loop study and in vivo oral absorption study. Absorption of FD-4 from the jejunum was significantly enhanced by 5 mM spermine without serious membrane damage in the jejunum. An in vivo oral absorption study was also performed, and plasma FD-4 levels increased significantly after co-administration of 30 mM spermine. In the in vitro transport studies with Caco-2 cells, prolonged incubation with spermine resulted in a gradual decrease in transepithelial electrical resistance. This finding suggests that the absorption-enhancing mechanism of spermine partly includes opening the tight junctions of the epithelium via the paracellular route. These results indicate that excess oral ingestion of polyamines may have widespread health effects via the modulation of the intestinal epithelial barrier function.
Assuntos
Dextranos/metabolismo , Fluoresceína-5-Isotiocianato/análogos & derivados , Absorção Intestinal/efeitos dos fármacos , Espermina/farmacologia , Animais , Células CACO-2 , Impedância Elétrica , Fluoresceína-5-Isotiocianato/metabolismo , Humanos , Íleo/metabolismo , Jejuno/efeitos dos fármacos , Jejuno/metabolismo , Jejuno/patologia , Ratos , Ratos Sprague-DawleyRESUMO
The safety and efficacy of Kogenate, a recombinant factor VIII (rFVIII) preparation for the treatment of bleeding episodes, were studied in a 123-patient meta-analysis population of previously treated patients (PTPs), including 15 enrolled in the registration Phase III trial (PTP-I group), 93 from the post-marketing special investigation (PTP-II group), and 15 from short-term special investigations in surgery or tooth extraction (SI group). These patients (82 severe, 31 moderate, 9 mild, and 1 unknown), aged 11 months to 72 years, were enrolled in 28 centers in Japan. Blood samples taken at the baseline and at 3, 6, 9, 12, 18, and 24 months after the introduction of Kogenate were evaluated for FVIII inhibitor antibodies, antibodies formed against trace proteins derived from the rFVIII production process, and for general changes in laboratory test results. Mean exposure to Kogenate was 1103 days in PTP-I, 86 days in PTP-II, 27 days in patients in surgery, and 2 days in patients with tooth extraction. Assessment of FVIII inhibitor activity was conducted in 115 of the 123 patients by means of the Bethesda assay. Twelve patients were found to have a low titer of FVIII inhibitor (0.5-3.0 BU/mL) prior to any administration of Kogenate, and 103 were inhibitor-negative at the baseline. Among this latter group, 3 patients (2.9%) tested inhibitor-positive, with titers ranging from 1.2 to 2.1 BU/mL, with 4 patients below 1.0 BU/mL. One patient in the 11 PTPs investigated (PTP-I) developed antibodies against baby hamster kidney protein and mouse immunoglobulin G, but these findings were transient and asymptomatic. Hemostasis was achieved (markedly effective or effective) in 3666 of the 3855 bleeding episodes (95.1%) observed in 108 patients. Only 1 infusion was necessary in 3790 (98.3%) of these episodes. These data indicate that Kogenate is safe and very effective for the treatment of bleeding in PTPs with hemophilia A.
Assuntos
Contaminação de Medicamentos , Fator VIII/efeitos adversos , Hemofilia A/sangue , Hemorragia/sangue , Adolescente , Adulto , Idoso , Animais , Autoanticorpos/sangue , Autoanticorpos/imunologia , Inibidores dos Fatores de Coagulação Sanguínea/sangue , Inibidores dos Fatores de Coagulação Sanguínea/imunologia , Criança , Pré-Escolar , Ensaios Clínicos Fase II como Assunto , Ensaios Clínicos Fase III como Assunto , Cricetinae , Cães , Fator VIII/administração & dosagem , Fator VIII/imunologia , Seguimentos , Hemofilia A/tratamento farmacológico , Hemofilia A/imunologia , Hemorragia/tratamento farmacológico , Hemorragia/imunologia , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Lactente , Pessoa de Meia-Idade , Fatores de TempoRESUMO
We encountered a 2-year-old boy with acquired haemophilia, which rarely occurs in children, who was complicated with nephrotic syndrome. In mid-August 2001, he was diagnosed to have nephrotic syndrome based on the presence of massive proteinuria and hypoalbuminaemia. Activated partial thromboplastin time (APTT) was normal at 42.4 s at that time. After starting prednisone administration of 2 mg kg(-1) day(-1), the proteinuria disappeared immediately. However, in early October the same year, subcutaneous ecchymosis and intramuscular bleeding occurred for no apparent reason, and from the examination results his APTT was 106.4 s, factor VIII (FVIII) activity was <1%, and the anti-FVIII inhibitor titre was 6.9 BU ml(-1). As a result, he was diagnosed to have acquired haemophilia. The anti-nuclear antibody and anti-phospholipid antibody were negative. With recombinant activated FVII, haemostasis was obtained, and after administering three courses of steroid pulse therapy (methyl prednisolone: 20 mg kg(-1) day(-1) x 3 days), the anti-FVIII inhibitory activity disappeared. An analysis of the immunological and coagulation properties of his FVIII autoantibodies showed the anti-FVIII inhibitory activity to be mediated by IgG(1) antibody. In other words, his FVIII inhibitor was a Th1 dominant and it provided a good response to treatment. These findings correlate with those of previous reports. The patient thereafter frequently demonstrated a recurrence of nephrotic syndrome. As a result, he is presently being managed with cyclosporine. However, no recurrence of the anti-FVIII titre has been observed.
Assuntos
Glucocorticoides/administração & dosagem , Hemofilia A/tratamento farmacológico , Metilprednisolona/administração & dosagem , Síndrome Nefrótica/tratamento farmacológico , Autoanticorpos/análise , Pré-Escolar , Fator VII/uso terapêutico , Fator VIII/imunologia , Fator VIIa , Hemofilia A/complicações , Hemofilia A/imunologia , Humanos , Imunoglobulina G/imunologia , Masculino , Síndrome Nefrótica/complicações , Pulsoterapia/métodos , Proteínas Recombinantes/uso terapêutico , Resultado do TratamentoRESUMO
The safety and efficacy of a recombinant factor VIII (rFVIII) preparation (Kogenate) for the treatment of bleeding episodes was studied in previously untreated patients (PUPs) with severe, moderate, and mild hemophilia A. Patient peripheral blood samples taken at baseline and at 3, 6, 9, 12, 18, and 24 months after the first infusion were evaluated for FVIII inhibitor antibodies by the Bethesda assay, for antibodies formed against trace proteins derived from the rFVIII production process, and for general changes in laboratory test results. Samples for general laboratory testing were also drawn every 6 months after the first 24 months. Hemostatic efficacy was assessed by physicians, and adverse events were recorded throughout the study period. Forty-three PUPs (30 with FVIII:C <1%; 10 with FVIII:C 1%-5%; and 3 with FVIII:C >5%) aged 3 months to 32 years were enrolled at 33 centers in Japan. Patients were studied for a mean of 51 months (range, 11-80 months), and the mean exposure time was 83 days (range, 2-571 days). The incidence of occurrence of FVIII inhibitors was 34.9% (high responders [> or = 10 Bethesda U/mL], 11.6%; low responders [0.5-<10 Bethesda U/mL], 23.3%). The median cumulative exposure time of inhibitor detection was 12 days, indicating inhibitor development at an early stage after the start of infusion of this preparation. Hemostasis was achieved with a single dose of Kogenate in 94.8% of the 951 bleeding episodes recorded in the study. Transient increases in antibodies against baby hamster kidney proteins and antimouse immunoglobulin G were observed in 14.0% and 18.6% of patients, respectively. Anti-rFVIII seroconversion was observed in 18.6% of patients and only in patients with inhibitor antibodies. Antibody responses to trace proteins were not correlated with drug-related adverse events with the exception of FVIII activity inhibition in PUPs with anti-rFVIII seroconversion. These data indicate that Kogenate is safe and effective for the treatment of bleeding in PUPs with hemophilia A.
Assuntos
Fator VIII/uso terapêutico , Hemofilia A/tratamento farmacológico , Adolescente , Adulto , Criança , Pré-Escolar , Fator VIII/antagonistas & inibidores , Fator VIII/imunologia , Hemorragia/etiologia , Hemorragia/prevenção & controle , Hemostasia/efeitos dos fármacos , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Lactente , Isoanticorpos/biossíntese , Isoanticorpos/sangue , Japão , Masculino , Vigilância de Produtos Comercializados , Resultado do TratamentoRESUMO
Polyamines such as spermidine and spermine are abundant in living cells and are believed to aid in the dense packaging of cellular DNA. DNA condensation is a prerequisite for the transport of gene vectors in living cells. To elucidate the structural features of polyamines governing DNA condensation, we studied the collapse of lambda-DNA by spermine and a series of its homologues, H2N(CH2)3NH(CH2)n=2-12NH(CH2)3NH2 (n = 4 for spermine), using static and dynamic light scattering techniques. All polyamines provoked DNA condensation; however, their efficacy varied with the structural geometry of the polyamine. In 10 mM sodium cacodylate buffer, the EC50 values for DNA condensation were comparable (4 +/- 1 microM) for spermine homologues with n = 4-8, whereas the lower and higher homologues provoked DNA condensation at higher EC50 values. The EC50 values increased with an increase in the monovalent ion (Na+) concentration in the buffer. The slope of a plot of log [EC50(polyamine4+)] against log [Na+] was approximately 1.5 for polyamines with even number values of n, whereas the slope value was approximately 1 for compounds with odd number values of n. Dynamic light scattering measurements showed the presence of compact particles with hydrodynamic radii (Rh) of about 40-50 nm for compounds with n = 3-6. Rh increased with further increase in methylene chain length separating the secondary amino groups of the polyamines (Rh = 60-70 nm for n = 7-10 and >100 nm for n = 11 and 12). Determination of the relative binding affinity of polyamines to DNA using an ethidium bromide displacement assay showed that homologues with n = 2 and 3 as well as those with n > 7 had significantly lower DNA binding affinity compared to spermine and homologues with n = 5 and 6. These data suggest that the chemical structure of isovalent polyamines exerts a profound influence on their ability to recognize and condense DNA, and on the size of the DNA condensates formed in aqueous solution.
Assuntos
DNA Viral/efeitos dos fármacos , Poliaminas/farmacologia , Bacteriófago lambda/efeitos dos fármacos , Bacteriófago lambda/genética , Sítios de Ligação , DNA Viral/química , Luz , Concentração Osmolar , Espalhamento de Radiação , Espermina/farmacologiaAssuntos
Úlcera da Perna/etiologia , Úlcera da Perna/cirurgia , Deficiência de Proteína C/complicações , Transplante de Pele/métodos , Adulto , Humanos , Masculino , Prognóstico , Deficiência de Proteína C/diagnóstico , Índice de Gravidade de Doença , Resultado do Tratamento , Cicatrização/fisiologiaRESUMO
Since the first patient with antithrombin deficiency was reported, various hereditary thrombophilia have been discovered. However, we experienced a family line of multiple thrombosis in which known hereditary thrombophilia were all refuted. Case 1 died of inferior vena cava thrombosis at the age of 56 days. Case 2, the elder sister of Case 1, developed deep vein thrombosis of the left leg at age 2. She was started on warfarin but contracted deep vein thrombosis of the right leg at the age of 7. In the family of these cases there have been another five cases of thrombosis, spanning three generations, giving a total of seven cases. Six of the cases developed at an early age, below 50 years. Antithrombin, protein C, protein S, heparin cofactor II, soluble thrombomodulin, plasminogen, alpha 2 plasminogen inhibitor, and tissue factor pathway inhibitor were measured but there were no abnormalities, nor was there any resistance to activated protein C. The onset of thrombosis in this family is becoming younger with the passing of generations, and clinical symptoms have been showing a worsening tendency.
Assuntos
Trombose/genética , Fatores Etários , Pré-Escolar , Feminino , Humanos , Lactente , Masculino , Trombose Venosa/genéticaRESUMO
The purpose of the present study is to answer the question of whether there is a species difference in the effects of a pharmacological dose of dehydroepiandrosterone (DHEA) on the enzymes that participate in oleic acid (18:1) formation in the liver. Feeding a diet containing 0.5% (w/w) DHEA for 14 days markedly increased the activities of acyl-coenzyme A (CoA) synthetase, palmitoyl-CoA chain elongase and stearoyl-CoA desaturase in the liver of rats and mice. These enzyme activities, however, were not changed by DHEA in guinea pigs. The treatments of rats and mice with DHEA markedly increased proportions of 18:1 in hepatic lipids, especially phosphatidylcholine (selectively at C-2 position), triacylglycerol and cholesterol ester. DHEA caused no significant changes in acyl compositions of hepatic lipids of guinea pigs. The levels of DHEA or dehydroepiandrosterone sulfate (DHEAS) were markedly increased in serum and livers by DHEA administration to rats, mice and guinea pigs. High correlations were observed between hepatic levels of DHEA or DHEAS and stearoyl-CoA desaturase activities in rats. These results indicate that there are species differences in the inducing effects of DHEA or DHEAS on hepatic formation of 18:1 and that guinea pigs lack the machinery to induce the enzymes.
Assuntos
Desidroepiandrosterona/farmacologia , Fígado/efeitos dos fármacos , Fígado/enzimologia , Ácido Oleico/biossíntese , Acetiltransferases/metabolismo , Administração Oral , Animais , Desidroepiandrosterona/administração & dosagem , Desidroepiandrosterona/sangue , Sulfato de Desidroepiandrosterona/metabolismo , Indução Enzimática/efeitos dos fármacos , Cobaias , Metabolismo dos Lipídeos , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos , Microssomos Hepáticos/enzimologia , Ácido Oleico/sangue , Palmitoil Coenzima A/metabolismo , Ratos , Ratos Wistar , Especificidade da Espécie , Estearoil-CoA Dessaturase/metabolismo , Fatores de TempoRESUMO
We studied the formation of triplex DNA in the purine-pyrimidine-rich promoter site sequence of cyclin D1, located at -116 to -99 from the transcription initiation site, with a molecular beacon comprised of a G-rich 18-mer triplex forming oligodeoxyribonucleotide. Formation of triplex DNA was monitored by enhanced fluorescence of the beacon, due to the weakening of fluorescence energy transfer, upon its binding to the target duplex. Electrophoretic mobility shift assay confirmed triplex DNA formation by these oligonucleotides. In low salt buffer (10 mM Na(+)), triplex DNA formation was not observed in the absence of a ligand such as spermine. At room temperature (22 degrees C), the equilibrium association constant (K(a)) calculated in the presence of 1 microM spermine and 10 mM Na(+) was 3.2 x 10(8) M(-1). The K(a) value was 1.0 x 10(9) M(-1) in the presence of 150 mM Na(+), and it increased by 10-fold by the addition of 1 mM spermine. Delta H, Delta S, and Delta G of triplex DNA formation, calculated from the temperature dependence of K(a) in the range of 20--45 degrees C, were -35.9 kcal/mol, -77 cal/(mol.K), and -13 kcal/mol, respectively, in the presence of 150 mM NaCl. The corresponding values were -52.9 kcal/mol, -132.5 cal/(mol.K), and -13.4 kcal/mol in the presence of 150 mM NaCl and 1 mM spermine. Structurally related polyamines exerted different degrees of triplex DNA stabilization, as determined by binding constant measurements. Comparison of spermine versus hexamine showed a 17-fold increase in the equilibrium association constant, whereas bis(ethyl) derivatization lead to a 4-fold decrease of this value. In the absence of added duplex and polyamines, the molecular beacon dissociated with a melting temperature of 67 degrees C. Thermodynamic parameters of beacon melting were calculated from the melting curve, and the Delta H, Delta S, and Delta G values were 37.8 kcal/mol, 112 cal/(mol.K), and 4.4 kcal/mol, respectively. These results demonstrate that molecular beacons can be used for the direct determination of the equilibrium association constants and thermodynamic parameters of triplex DNA formation in the presence of ligands such as polyamines.
Assuntos
Ciclina D1/genética , Sondas de DNA/metabolismo , DNA/metabolismo , Corantes Fluorescentes/metabolismo , Conformação de Ácido Nucleico , Sondas de Oligonucleotídeos/metabolismo , Regiões Promotoras Genéticas , Ciclina D1/metabolismo , DNA/genética , Sondas de DNA/síntese química , Eletroforese em Gel de Poliacrilamida , Corantes Fluorescentes/síntese química , Humanos , Sondas de Oligonucleotídeos/síntese química , Desnaturação Proteica , Cloreto de Sódio , Espectrometria de Fluorescência , Espermina/metabolismo , Temperatura , TermodinâmicaRESUMO
A multicenter and open-labeled clinical trial of human recombinant factor VIIa (rFVIIa) was conducted in Japanese patients with severe hemophilia A or B with inhibitors. The trial consisted of 2 parts. In study 1, the pharmacokinetics, pharmacodynamics, and safety of a single dose of 120 microg/kg of rFVIIa were investigated in 8 patients. In the subsequent study 2, the hemostatic effect and safety of rFVIIa were evaluated during a 24-week period in 10 patients. In study 1, the mean maximum FVII-coagulant activity (FVII:C) was found to occur after 10 minutes; activity then decreased rapidly and returned to the baseline within 24 hours after a single intravenous infusion of rFVIIa. The mean half-life of FVII:C was 3.5 hours. The activated partial thromboplastin time and prothrombin time in the patients were immediately shortened but returned to the baseline within 24 hours after dosing. In study 2, 86 microg/kg to 120 microg/kg of rFVIIa (mean, 97 microg/kg) was administered 1 to 85 times to 10 patients. A total of 58.0% (91/157) of bleeding episodes were treated excellently or effectively, with 5 (3.2%) ineffective episodes. There was no apparent trend in the relationship of the hemostatic effect with bleeding sites, mean dose, or number of injections. The efficacy rate, however, was significantly higher (90.0%) in bleeding episodes treated within 3 hours than in those treated at longer intervals (31.0%). No treatment-related adverse events were observed, and there was no evidence of antibody formation to rFVIIa. In conclusion. rFVIIa is an effective and well-tolerated option for treatment of bleeding episodes in hemophilia patients with inhibitors.
Assuntos
Fator VIIa/farmacologia , Hemofilia A/tratamento farmacológico , Adolescente , Adulto , Qualidade de Produtos para o Consumidor , Fator VIIa/administração & dosagem , Fator VIIa/efeitos adversos , Fator VIIa/farmacocinética , Hemofilia A/imunologia , Humanos , Isoanticorpos/sangue , Japão , Pessoa de Meia-Idade , Proteínas Recombinantes/efeitos adversos , Proteínas Recombinantes/farmacocinética , Proteínas Recombinantes/farmacologia , Equivalência TerapêuticaRESUMO
To determine whether the fibrinolytic system is related to the occurrence of cardiac complication in Kawasaki disease, we measured tissue plasminogen activator, plasminogen activator inhibitor-1, and related factors in the plasma of children with Kawasaki disease. Patients (mean age, 1.8 years) were classified into patients with cardiac complication (n = 9) and no complication (n = 14) groups echocardiographically. They underwent single, high-dose, intravenous-gamma-globulin infusion therapy. Blood was drawn just before and the day after the single high-dose intravenous gamma-globulin infusion therapy (acute phase), and at early and late convalescent phases. Leukocytosis was normalized immediately after the single, high-dose, intravenous gamma-globulin infusion therapy. C-reactive protein and fibrinogen were increased in the acute phase and normalized by convalescent phases. D-dimer fraction of fibrin degradation products changed in a similar manner. Tissue plasminogen activator and plasminogen activator inhibitor-1 were increased in acute phase. The tissue plasminogen activator/plasminogen activator inhibitor-1 ratio was lower in the complication group than in the no complication group throughout the observation period (0.095 versus 0.208 after single, high-dose, intravenous gamma-globulin infusion therapy, p = 0.006; 0.094 versus 0.183 at late convalescent phase, p = 0.024). A low tissue plasminogen activator/plasminogen activator inhibitor-1 ratio can predict the propensity for cardiac complication, and can help the physician to decide whether additional therapies are necessary in acute phase Kawasaki disease.
Assuntos
Cardiopatias/sangue , Síndrome de Linfonodos Mucocutâneos/sangue , Inibidor 1 de Ativador de Plasminogênio/análise , Ativador de Plasminogênio Tecidual/análise , Doença Aguda , Biomarcadores , Proteína C-Reativa/análise , Pré-Escolar , Convalescença , Feminino , Produtos de Degradação da Fibrina e do Fibrinogênio/análise , Fibrinogênio/análise , Cardiopatias/diagnóstico por imagem , Cardiopatias/etiologia , Humanos , Imunoglobulinas Intravenosas/uso terapêutico , Lactente , Contagem de Leucócitos , Masculino , Síndrome de Linfonodos Mucocutâneos/complicações , Síndrome de Linfonodos Mucocutâneos/terapia , Valor Preditivo dos Testes , Sensibilidade e Especificidade , UltrassonografiaRESUMO
The recombinant full-length FVIII product Kogenate has been reformulated using sucrose (rFVIII-FS) instead of human serum albumin as a stabiliser in purification and formulation. The in vivo recovery, haemostatic efficacy, and safety of rFVIII-FS were investigated in 20 previously treated patients with severe or moderate haemophilia A for > or = 24 weeks. In vivo recoveries of 73.5 +/- 16.3%, 78.4 +/- 16.1%, and 82.8 +/- 23.9% after the initial infusion of 50 IU kg(-1) rFVIII-FS and at weeks 12 and 24, respectively, showed no significant changes over time. A total of 1115 infusions (mean dose 24.1 +/- 8.4 IU kg(-1)) were included in the analysis of haemostatic efficacy. One (80.5%) or two (8.2%) infusions achieved adequate haemostasis in 88.7% of all bleeding episodes, and haemostatic efficacy was judged 'excellent' or 'good' in 749 of 764 episodes (98.0%). The haemostatic efficacy was judged as 'excellent' or 'good' in 924 of 1115 (82.9%) infusions. Twenty-one adverse events were observed in 12 patients in the total 1541 infusions included in the safety analysis. Causality with respect to rFVIII-FS could not be ruled out in three events in one HIV-negative patient: elevated CD4(%), decreased CD8(%), and elevated CD4/CD8 ratio. No FVIII inhibitor development was observed in any patient. ELISA assay testing for antibodies to rFVIII, baby hamster kidney cell (BHK) protein, and murine IgG were all negative. These results show that rFVIII-FS is a safe and effective for long-term treatment of patients with haemophilia A.