RESUMO
Over the past 10 years, polyvalent DNA-gold nanoparticle (DNA-GNP) conjugate has been demonstrated as an efficient, universal nanocarrier for drug and gene delivery with high uptake by over 50 different types of primary and cancer cell lines. A barrier limiting its in vivo effectiveness is limited resistance to nuclease degradation and nonspecific interaction with blood serum contents. Herein we show that terminal PEGylation of the complementary DNA strand hybridized to a polyvalent DNA-GNP conjugate can eliminate nonspecific adsorption of serum proteins and greatly increases its resistance against DNase I-based degradation. The PEGylated DNA-GNP conjugate still retains a high cell uptake property, making it an attractive intracellular delivery nanocarrier for DNA binding reagents. We show that it can be used for successful intracellular delivery of doxorubicin, a widely used clinical cancer chemotherapeutic drug. Moreover, it can be used for efficient delivery of some cell-membrane-impermeable reagents such as propidium iodide (a DNA intercalating fluorescent dye currently limited to the use of staining dead cells only) and a diruthenium complex (a DNA groove binder), for successful staining of live cells.
Assuntos
DNA/química , Substâncias Intercalantes/química , Nanopartículas Metálicas/química , Polietilenoglicóis/química , Propídio/química , Antibióticos Antineoplásicos/química , Antibióticos Antineoplásicos/toxicidade , Sobrevivência Celular/efeitos dos fármacos , Complexos de Coordenação/química , Complexos de Coordenação/metabolismo , DNA/metabolismo , Desoxirribonuclease I/química , Desoxirribonuclease I/metabolismo , Doxorrubicina/química , Doxorrubicina/toxicidade , Portadores de Fármacos/química , Difusão Dinâmica da Luz , Ouro/química , Células HeLa , Humanos , Substâncias Intercalantes/metabolismo , Microscopia Confocal , Propídio/metabolismo , Rutênio/químicaRESUMO
The DNA binding and cellular localization properties of a new luminescent heterobimetallic Ir(III) Ru(II) tetrapyridophenazine complex are reported. Surprisingly, in standard cell media, in which its tetracationic, isostructural Ru(II) Ru(II) analogue is localized in the nucleus, the new tricationic complex is poorly taken up by live cells and demonstrates no nuclear staining. Consequent cell-free studies reveal that the Ir(III) Ru(II) complex binds bovine serum albumin, BSA, in Sudlow's Site I with a similar increase in emission and binding affinity to that observed with DNA. Contrastingly, in serum-free conditions the complex is rapidly internalized by live cells, where it localizes in cell nuclei and functions as a DNA imaging agent. The absence of serum proteins also greatly alters the cytotoxicity of the complex, where high levels of oncosis/necrosis are observed due to this enhanced uptake. This suggests that simply increasing the lipophilicity of a DNA imaging probe to enhance cellular uptake can be counterproductive as, due to increased binding to serum albumin protein, this strategy can actually disrupt nuclear targeting.