RESUMO
The aim of this study was to evaluate the anti-tumor activity of Amblyomin-X, a serine protease Kunitz-type inhibitor. Amblyomin-X induced tumor mass regression and decreased number of metastatic events in a B16F10 murine melanoma model. Alterations on expression of several genes related to cell cycle were observed when two tumor cell lines were treated with Amblyomin-X. PSMB2, which encodes a proteasome subunit, was differentially expressed, in agreement to inhibition of proteasomal activity in both cell lines. In conclusion, our results indicate that Amblyomin-X selectively acts on tumor cells by inducing apoptotic cell death, possibly by targeting the ubiquitin-proteasome system.
Assuntos
Antineoplásicos/uso terapêutico , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Ixodidae/química , Melanoma Experimental/tratamento farmacológico , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas e Peptídeos Salivares/uso terapêutico , Inibidores de Serina Proteinase/uso terapêutico , Ubiquitina/metabolismo , Animais , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Apoptose/efeitos dos fármacos , Proteínas de Artrópodes , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Modelos Animais de Doenças , Ixodidae/genética , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Metástase Neoplásica , Complexo de Endopeptidases do Proteassoma/efeitos dos fármacos , Glândulas Salivares/química , Proteínas e Peptídeos Salivares/química , Proteínas e Peptídeos Salivares/isolamento & purificação , Inibidores de Serina Proteinase/química , Inibidores de Serina Proteinase/isolamento & purificaçãoRESUMO
Bothrops insularis is a threatened snake endemic to Queimada Grande Island, southern coast of São Paulo, Brazil, and the occurrence of sexual abnormalities in males, females and intersexes (females with functional ovaries and rudimentary hemipenis) has been reported in this population. The aim of this study was to identify ontogenetic shifts in protease expression of offspring of captive-bred B. insularis. Three neonates from a single litter were maintained at the facilities of Laboratory of Herpetology, Institute Butantan, for 41 months. The snakes were individually milked and venoms were analyzed both by SDS-PAGE, under reducing conditions, and for biochemical activities. The venoms from the mother and from a pool of adult specimens were used as references. In regard to the electrophoretic patterns, common bands were identified mainly between 14 and 50 kDa among snakes. The occurrence of proteolytic activity was noticed predominantly between 27 and 45 kDa in zymograms. Inhibitory assays with 1,10-phenantroline (10 mM) and PMSF (5 mM) showed that venoms possessed both metalloproteases and serine proteases. Venoms of young specimens showed a higher coagulant activity than those of adults, especially upon factors X and II. All venoms presented fibrino(geno)lytic activity, degrading Aalpha and Bbeta chains of fibrinogen, and lysing fibrin plate. These findings can reflect important individual, ontogenetic and sexual differences on venom composition and are likely correlated with diet habits of this species.
