Block and graft copolymers of N-(2-hydroxypropyl)methacrylamide (HPMA) with 2-(trimethylammonio)ethyl methacrylate were synthesised and used for preparation of polyelectrolyte complexes with calf thymus DNA intended for targeted delivery of genes in vivo. In this study the effects of the speed of component mixing, total concentration of polymers, ionic strength of solvents, copolymer structure and content of HPMA in the copolymers on parameters of the polyelectrolyte complexes was investigated. Static and dynamic light scattering methods were used as a main tool for characterising these complexes. The presence of HPMA units in the polycation had no significant effect on its ability to form complexes with DNA, but did affect molecular parameters and aggregation (precipitation) of the complexes. The size of the complexes increases whereas their molecular weight decreases with increasing content of HPMA units. The density of the complexes decreases with increasing HPMA content independently of the copolymer structure. In order to prepare stable DNA complexes containing single DNA molecule, the following rules should be observed: (1) copolymers should have a content of HPMA units higher than 40%; (2) the DNA concentrations in solutions should be kept below 4 x 10(-5) g/ml and (3) both components should be mixed together in deionized water. The stability of the complexes against precipitation in 0.15 M NaCl and the resistance of the complexed DNA to the action of nucleases was also studied. Whereas DNA complexes of all copolymers showed very good nuclease stability, the presence of a sufficiently high content of HPMA is necessary for their good stability in 0.15 M NaCl. The investigation of the stability and the interaction of DNA complexes in aqueous solutions of serum albumin and dilute human blood serum revealed adsorption of biomacromolecules on DNA complexes accompanied by significant changes in the zeta-potential which finally resulted in formation of a "protein layer" and in undesirable precipitation of DNA complexes. In in vitro transfection experiments, the transfection efficiency of DNA complexes with copolymers was always higher than that of the cationic homopolymer slightly increasing with increasing content of HPMA in the copolymers but being about 10-100-times lower than the complexes DNA-poly(L-lysine. In the cytoplasmic injections, it was observed that DNA complexes produced greater gene expression than a direct microinjection of free DNA. The block copolymer complexes were also found to be more efficient than the corresponding simple polycation complexes. In the nuclear microinjection, precisely the opposite behaviour was observed.

• MeSH
• akrylamidy chemická syntéza   chemie   MeSH
• chemické jevy MeSH
• deoxyribonukleasy metabolismus   MeSH
• DNA aplikace a dávkování   chemie   MeSH
• dospělí MeSH
• fyzikální chemie MeSH
• kationty MeSH
• kyseliny karboxylové chemie   MeSH
• lidé MeSH
• methakryláty chemická syntéza   chemie   MeSH
• oocyty metabolismus   MeSH
• polyethylenglykoly MeSH
• polymery chemická syntéza   chemie   MeSH
• polyvinyly chemická syntéza   chemie   MeSH
• radiační rozptyl MeSH
• rozpustnost MeSH
• světlo MeSH
• technika přenosu genů přístrojové vybavení   MeSH
• transfekce MeSH
• Xenopus MeSH
• zvířata MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• klinické zkoušky MeSH
• práce podpořená grantem MeSH
• Názvy látek
• 2-(trimethylammonio)ethyl methacrylate MeSH Prohlížeč
• akrylamidy MeSH
• deoxyribonukleasy MeSH
• DNA MeSH
• kationty MeSH
• kyseliny karboxylové MeSH
• methakryláty MeSH
• N-(2-hydroxypropyl)methacrylamide MeSH Prohlížeč
• poly(vinylamine) MeSH Prohlížeč
• polyethylenglykoly MeSH
• polymery MeSH
• polyvinyly MeSH

Biophysical properties of polycation/DNA complexes designed for gene delivery were studied with respect to the conditions of their preparation, chemical structure and molecular weight of the polycations involved. The polycations used included a variety of cationic polymers and copolymers containing primary and tertiary amino or quaternary ammonium groups. It was found that the molecular weight and the size of these polyelectrolyte complexes (PECs) increase with increasing temperature and pH of the buffer. By decreasing the molecular weight of polycations used for PEC formation, the complexes become unstable towards coagulation in aqueous solution at lower pH. The self-assembly of DNA with low-molecular-weight polycations in water provides PECs with the lowest molecular weight, smallest size and the lowest density but their stability in NaCl solutions is very poor. Despite the complexity of the multistep transfection process, a direct correlation between the transfection efficiency in vitro and the stability of the complexes in NaCl solutions and coagulation in 0.15 M NaCl solution was found. DNA complexes with polycations containing primary amino groups showed the best stability in saline solutions and also the best transfection activity. PECs formed by polycations with quaternary ammonium groups were the least resistant to destruction by the added salt and provided the lowest activity in transfection assays. The highest transfection activity was found for DNA complexes formed with a statistical copolymer containing primary and tertiary amines.

• MeSH
• chemické jevy MeSH
• chlorid sodný MeSH
• DNA genetika   farmakokinetika   MeSH
• Escherichia coli MeSH
• fyzikální chemie MeSH
• genetické vektory genetika   farmakokinetika   MeSH
• lékové transportní systémy metody   MeSH
• melanom experimentální genetika   metabolismus   MeSH
• mikroskopie atomárních sil MeSH
• molekulová hmotnost MeSH
• myši MeSH
• nádorové buňky kultivované MeSH
• polyaminy chemická syntéza   farmakokinetika   MeSH
• polyelektrolyty MeSH
• radiační rozptyl MeSH
• roztoky MeSH
• stabilita léku MeSH
• světlo MeSH
• transfekce metody   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• chlorid sodný MeSH
• DNA MeSH
• polyaminy MeSH
• polycations MeSH Prohlížeč
• polyelektrolyty MeSH
• roztoky MeSH