RESUMO
PURPOSE: Sorafenib is a multikinase inhibitor that targets the MAPK pathway and is currently used for the treatment of hepatocellular and renal carcinoma. Recently, it has been shown that sorafenib is also cytotoxic to multiple myeloma (MM) cells. Here, we have further analyzed the mechanism of sorafenib-induced death in MM cells. METHODS: Cell death induced by sorafenib in MM cell lines and in plasma cells from MM patients was evaluated by analysis of gene expression by RT-MLPA and quantitative PCR, protein levels and functionality by Western blot and flow cytometry and gene silencing with siRNA. RESULTS: Cell death was characterized by phosphatidylserine exposure, ΔΨm loss, cytochrome c release and caspase activation, hallmarks of apoptosis. DL50 at 24 h ranged from 6 to 10 µM. Ex vivo treatment with 20 µM sorafenib induced apoptosis in around 80 % myeloma cells from six multiple myeloma patients. Sorafenib induced caspase-dependent degradation of Bcl-xL and Mcl-1 proteins, destabilizing the mitochondria and speeding up the development of apoptosis. Sorafenib treatment increased levels of Puma at mRNA and protein level and gene silencing with siRNA confirmed a relevant role for Puma in the induction of apoptosis. Co-treatment with the pan-caspase inhibitor Z-VAD-fmk prevented cell death to a variable degree depending on the cell line. In RPMI 8226 cells, Z-VAD-fmk prevented most of sorafenib-induced death. However, death in MM.1S was only prevented by co-incubation with both Z-VAD-fmk and the RIP1K inhibitor necrostatin-1, indicating that under conditions of inefficient caspase activation, sorafenib induces death by necroptosis. CONCLUSION: Our results demonstrate a key role for Puma in the triggering of sorafenib-induced apoptosis and that this drug can also induce death by necroptosis in multiple myeloma cells.
Assuntos
Antineoplásicos/farmacologia , Proteínas Reguladoras de Apoptose/metabolismo , Apoptose/efeitos dos fármacos , Mitocôndrias/patologia , Mieloma Múltiplo/tratamento farmacológico , Mieloma Múltiplo/patologia , Niacinamida/análogos & derivados , Compostos de Fenilureia/farmacologia , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Reguladoras de Apoptose/antagonistas & inibidores , Proteínas Reguladoras de Apoptose/genética , Western Blotting , Inibidores de Caspase/farmacologia , Caspases/química , Proliferação de Células/efeitos dos fármacos , Citometria de Fluxo , Humanos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Mieloma Múltiplo/metabolismo , Necrose , Niacinamida/farmacologia , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sorafenibe , Células Tumorais CultivadasRESUMO
The h-R3 is a humanized growth factor receptor monoclonal antibody (mAb) in development for the treatment of head and neck tumours in which malignant cells overexpress the Epidermal Growth Factor receptor. The present study was designed to evaluate the toxicity of repeated intravenous doses of the h-R3 mAb in a relevant species demonstrated by the avidin-biotin-peroxidase immunohistochemical (IHC) technique in skin biopsy samples from three Cercopithecus aethiops sabaeus monkeys (green monkeys). Additionally, 18 green monkeys were daily intravenously treated during 14 consecutive days. Monkeys were distributed into three experimental groups with three animals of each sex in each group. Group I received saline solution and served as control group; group II received 2.85 mg/kg of h-R3 mAb; and group III received 11.4 mg/kg of the h-R3 mAb. During the study there were no deaths, neither pathological clinical signs, or variations in the corporal weight curve. The electroneurophysiological and sanguine chemistry results did not evidence alterations related to the assay substance. Areas of haematomas, haemorrhages and inflammation, probably related with the administration procedure, were observed at the administration zones of all animals; this fact could also explain the increase in the neutrophil count of all animals at the end of the study. The electrocardiography study showed that in the 14 days of the study one female monkey, from the higher dose group, shifted its cardiac axis from +60 degrees to + 120 degrees; this finding could be interpreted as a right ventricular elongation due to the relative high daily administered volume. It is concluded that doses up to 11.4 mg/kg of h-R3, intravenously administered during 14 consecutive days to Cercopithecus aethiops sabaeus monkeys do not produce considerable toxic effects in the studied system.
Assuntos
Anticorpos Monoclonais/toxicidade , Antineoplásicos/toxicidade , Receptores ErbB/imunologia , Animais , Anticorpos Monoclonais/administração & dosagem , Antineoplásicos/administração & dosagem , Chlorocebus aethiops , Relação Dose-Resposta a Droga , Eletrofisiologia , Receptores ErbB/metabolismo , Feminino , Hematoma/etiologia , Hematoma/patologia , Hemorragia/etiologia , Hemorragia/patologia , Técnicas Imunoenzimáticas , Injeções Intravenosas/efeitos adversos , Masculino , Pele/efeitos dos fármacos , Pele/metabolismo , Testes de ToxicidadeRESUMO
BACKGROUND: Blomia tropicalis is a mite that is one of the most important causes of allergic sensitization in studies of tropical and subtropical regions. Nonetheless, there are no published studies of the sensitivity and specificity of B. tropicalis extracts. METHOD: An open, non-randomized study was carried out in 205 asthmatic children and in 205 non-asthmatic children, both groups with an age range of 1 to 14 years. Duplicate skin prick tests were made using allergen extracts standardized by biological units and positive and negative controls. The test was considered positive when the average diameter of the wheal was > or = 3 mm and negative when it was < 3 mm. The validity of the test was estimated by calculating sensitivity, specificity, and the positive and negative predictive values. RESULTS: The skin prick test was positive in 82.92% of the asthmatic children and in 10.7% of the non-asthmatic children, yielding a sensitivity of 82.9%, 95% CI 76.9; 87.7 and a specificity of 89.3%, 95% IC 84.0; 93.0. The positive predictive value was 88.5%, 95% IC 83.0; 92.5 and the negative predictive value was 83.9%, 95% IC 78.2; 88.4. CONCLUSIONS: Blomia tropicalis is an important cause of bronchial asthma among the children of the northern Camagüey region.
Assuntos
Alérgenos , Asma/etiologia , Hipersensibilidade Imediata/diagnóstico , Ácaros/imunologia , Alérgenos/isolamento & purificação , Animais , Asma/epidemiologia , Criança , Pré-Escolar , Cuba/epidemiologia , Feminino , Humanos , Hipersensibilidade Imediata/complicações , Lactente , Masculino , Valor Preditivo dos Testes , Sensibilidade e Especificidade , Testes Cutâneos , Extratos de TecidosRESUMO
BACKGROUND: There are no antecedents in Cuba of studies developed to prove the efficacy of an allergy test to Blomia tropicalis despite of being one of the main mites which cause this ailment which affects more than 2,200,000 people. OBJECTIVES: Evaluate the allergenic extract Blomia tropicalis elaborated at the Centro Nacional de Biopreparados (BioCen), in the prick skin test, as a diagnose of allergy to this allergen which exists in all the tropical and subtropical countries. MATERIAL AND METHODS: We selected 50 patients with a positive clinic history of allergy to house dust and 50 healthy persons. Al the persons selected were skin prick tested with dilution's of the extracts (20,000 UB/mL) and positive and negative control. We measured the average diameter of the wheal and the perpendicular diameter, then we calculated the average. We also calculated size of the wheal in the duplicated, which constituted the size of the reaction. We estimated the validity of the test calculating the sensitivity and specificity. We used the EPITABLE program to do this. RESULTS: The skin prick tested positive in 78% of the patients and 6% in the healthy persons, for a sensitivity of 78% the interval of confidence (IC) 63.7-88.0 and a specificity of 94%, IC 82.5-98.4 the predictive negative value 81%, IC 68.2-89.7. CONCLUSIONS: The allergenic extract of Blomia tropicalis elaborated in BioCen is very sharp to test the allergy to this mite.
Assuntos
Ácaros e Carrapatos/imunologia , Alérgenos , Hipersensibilidade/diagnóstico , Adolescente , Adulto , Animais , Poeira/efeitos adversos , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
BACKGROUND: In persons older than one year, respiratory allergies can be studied from the etiologic viewpoint using skin test with specific allergens. MATERIAL AND METHOD: The objective of this research is to make an assessment of the skin reactivity to mites Dermatophagoides pteronyssinus, Dermatophagoides siboney and Blomia tropicalis in 76 patients with a diagnosis of allergic and nasal polyps which had been treated in Otorhinolaryngologie and sent to the service of allergologie in Esmeralda Camagüey. Every patient received a skin puncture test twice. Fifteen minutes later the hives began to appear and we measured the longer diameter and the perpendicular diameter of every hive and we calculated an average. We also took the average size of the hives in the second test, considering them positive when the medium diameter was < 3 mm and negative when it was < 3 mm. In those patients suffering from rhinitis the greatest positive frequency was for those with Dermatophagoides pteronyssinus and the greatest negative frequency to Dermatophagoides siboney. In patients with nasal polyps there was a positive predominance (p < 0.05) to Dermatophagoides pteronyssinus and a negative predominance (p < 0.05) to Dermatophagoides siboney. So, we many conclude that the mite Dermatophagoides pteronyssinus causes more positive reactions than Dermatophagoides siboney and Blomia tropicalis in those patients with allergic rhinitis and nasal polyps.