RESUMO
Linear cyclen-based polyamine (LCPA, M(w) = 7392, M(w)/M(n) = 1.19) as a novel non-viral gene vector was designed and synthesized from 1,7-diprotected 1,4,7,10-tetraazacyclododecane (cyclen), bis(beta-hydroxylethyl)amine and epichlorohydrin. Agarose gel retardation and fluorescent titration using ethidium bromide showed the good DNA-binding ability of LCPA. It could retard pDNA at an N/P ratio of 4 and form polyplexes with sizes around 250-300 nm from an N/P ratio of 10 to 60 and relatively lower zeta-potential values (< +3 mV) even at the N/P ratio of 60. The cytotoxicity of LCPA assayed by MTT is much lower than that of 25 kDa PEI. In vitro transfection against A549 and 293 cells showed that the transfection efficiency of LCPA/DNA polyplexes is close to that of 25 kDa PEI at an N/P ratio of 10-15, indicating that the new material could be a promising non-viral polycationic reagent for gene delivery.
Assuntos
Técnicas de Transferência de Genes , Compostos Heterocíclicos/química , Poliaminas/química , Poliaminas/metabolismo , Soluções Tampão , Linhagem Celular , Ciclamos , DNA/metabolismo , Eletroforese em Gel de Ágar , Epicloroidrina/química , Humanos , Concentração de Íons de Hidrogênio , Indicadores e Reagentes/síntese química , Indicadores e Reagentes/química , Indicadores e Reagentes/metabolismo , Indicadores e Reagentes/toxicidade , Tamanho da Partícula , Plasmídeos/genética , Poliaminas/síntese química , Poliaminas/toxicidade , TransfecçãoRESUMO
A novel linear poly[1,7-bis(mercaptoacetyl)-1,4,7,10-tetraazacyclododecane] (PBMAC) containing macrocyclic polyamine was synthesized through oxidation of 1,7-bis(mercaptoacetyl)-1,4,7,10-tetraazacyclododecane (BMAC) and characterized. Gel electrophoresis experiments showed that PBMAC can promote the DNA cleavage more efficiently than its monomer under physiological conditions without any thiol additives. Fluorescence quenching and DNA melting experiments demonstrated that PBMAC has stronger binding ability with DNA than that of monomer.
Assuntos
Compostos Aza/síntese química , Cicloparafinas/síntese química , Clivagem do DNA/efeitos dos fármacos , Dissulfetos/química , Polímeros/química , Compostos Aza/química , Cicloparafinas/química , Corantes Fluorescentes/química , Poliaminas/química , Polímeros/síntese química , Polímeros/metabolismo , Temperatura de TransiçãoRESUMO
The novel linear polymer of a macrocyclic polyamine copper (II) complex, which has many cyclen groups linked by epichlorohydrin, has been synthesized as a DNA cleavage agent. The structure of the polymer 3 was identified by 1HNMR and IR and its molecular weight was measured by GPC. The result of agarose gel electrophoresis assay showed that Cu-(II) complex 4 could act as a powerful catalyst for the cleavage of plasmid DNA under physiological conditions.
Assuntos
Cobre/química , DNA/química , Plasmídeos/metabolismo , Poliaminas/química , Polímeros/síntese química , Cobre/metabolismo , Ciclamos , DNA/metabolismo , Clivagem do DNA , Epicloroidrina/química , Compostos Heterocíclicos/química , Modelos Moleculares , Plasmídeos/químicaRESUMO
This study provided an experimental evidence for the use of cyclen (1, 4, 7, 10-tetraazacyclododecane)-based polymer for gene delivery. The interesting interaction of the polymer with plasmid DNA was studied by using fluorescence titration, circular dichroism spectra, agarose gel electrophoresis and atomic force microscopy. It was found that polyplex was formed between the polycation and plasmid DNA. The results demonstrated that the cyclen-based polymer could act as non-viral gene vector with relatively low cytotoxicity.