RESUMO
We characterized an anti-cancer fusion protein consisting of anthrax lethal factor (LF) and the catalytic domain of Pseudomonas exotoxin A by (i) mutating the N-terminal amino acids and by (ii) reductive methylation to dimethylate all lysines. Dimethylation of lysines was achieved quantitatively and specifically without affecting binding of the fusion protein to PA or decreasing the enzymatic activity of the catalytic moiety. Ubiquitination in vitro was drastically decreased for both the N-terminally mutated and dimethylated variants, and both appeared to be slightly more stable in the cytosol of treated cells. The dimethylated variant showed greatly reduced neutralization by antibodies to LF. The two described modifications offer unique advantages such as increased cytotoxic activity and diminished antibody recognition, and thus may be applicable to other therapeutic proteins that act in the cytosol of cells.
Assuntos
ADP Ribose Transferases/genética , Antígenos de Bactérias/genética , Toxinas Bacterianas/genética , Exotoxinas/genética , Mutação , Proteínas Recombinantes de Fusão/metabolismo , Proteínas Recombinantes de Fusão/toxicidade , Fatores de Virulência/genética , ADP Ribose Transferases/química , Animais , Antígenos de Bactérias/química , Antineoplásicos , Toxinas Bacterianas/química , Linhagem Celular , Cricetinae , Citosol/metabolismo , Epitopos/genética , Epitopos/imunologia , Epitopos/metabolismo , Exotoxinas/química , Humanos , Cinética , Espectrometria de Massas , Metilação , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Estabilidade Proteica , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Ubiquitinação , Fatores de Virulência/química , Exotoxina A de Pseudomonas aeruginosaRESUMO
Saponins are plant glycosides with favorable anti-tumorigenic properties. Several saponins inhibit tumor cell growth by cell cycle arrest and apoptosis with IC50 values of up to 0.2 microM. We discuss diverse groups of saponins (dioscins, saikosaponins, julibrosides, soy saponins, ginseng saponins and avicins) investigated in relation to tumor therapy and focus on cellular and systemic mechanisms of tumor cell growth inhibition both in vitro and in vivo. The review also describes saponins in combination with conventional tumor treatment strategies, which result in improved therapeutic success. Some combinations of saponins and anti-tumorigenic drugs induce synergistic effects with potentiated growth inhibition.
