Improving cellular uptake and in vivo tumor suppression efficacy of liposomal oligonucleotides by urea as a chemical penetration enhancer.

BACKGROUND: Liposomes are among the most widely used carriers for the delivery of antisense oligonucleotides (AsODNs) to intracellular targets. Although different strategies have been employed, the question of how to improve liposomal uptake and enhance the release of AsODN into cytoplasm still remains to be answered with respect to the use of a safe, easy and economic method. In the present study, the possibility of enhancing such processes at cellular and animal levels using urea as a penetration enhancer was investigated. METHODS: To perform this investigation, a cationic liposome containing an AsODN against protein kinase (PKC)-α was prepared, and the effect of urea on its cellular internalization and the related sequence-specific inhibition of gene expression in human lung adenocarcinoma A549 cells were investigated by flow cytometry and the reverse transcriptase-polymerase chain reaction, respectively. In in vivo studies, a xenograft lung tumor was established in nude mice by A549 cells and the enhancement effect of urea toward the effects of liposomal AsODN on tumor growth was investigated. RESULTS: Cellular studies revealed that urea treatment increases liposomal uptake and the release of AsODN into the cytoplasm by approximately 40%. Sequence-specific inhibition of target gene PKC-α expression was also increased by approximately two-fold by urea at 200-300 nM AsODN. In animal studies, urea significantly decreased the tumor volume (approximately 40%) and increased its doubling time from approximately 13 days to 17 days. CONCLUSIONS: Urea, and possibly other membrane fluidizers, could be regarded as penetration enhancers for liposomal AsODN delivery and may improve the therapeutic effect of these gene-therapy vectors at both cellular and animal levels.

9-nitrocamptothecin polymeric nanoparticles: cytotoxicity and pharmacokinetic studies of lactone and total forms of drug in rats.

The objective of this study was to evaluate the cytotoxicity and pharmacokinetics of total and lactone forms of 9-nitrocamptothecin (9-NC), an effective antineoplastic drug, after intravenous injection of drug incorporated into poly (DL-lactic-glycolic acid) nanoparticles (NPs). Drug-loaded NPs (9-NC.NP) were prepared by the nanoprecipitation method and examined for particle characteristics and in-vitro release in phosphate buffered saline. The best formulation showed a narrow size with an average diameter of 207+/-26 nm and a drug loading of more than 33.5%. The drug release profile showed a sustained 9-NC release up to 160 h. For a pharmacokinetic study, the concentration of 9-NC as the lactone form (9-NC.lac) and as the total of the lactone and carboxylate forms (9-NC.tot) in plasma was determined by using reverse-phase high performance liquid chromatography after intravenous administration of 9-NC.NP and a control solution to cannulated Wistar rats. In-vitro cytotoxic activity of 9-NC.NP and control solution was evaluated on the human ovarian cancer cell line (A2780sn) by MTT cell cytotoxicity assay. Results of in-vivo studies showed that NP encapsulation markedly increased the plasma concentration of both lactone and total forms of 9-NC compared with free drug. In comparison with free drug, NPs resulted in 3.63-fold and 5.40-fold increases in area under the plasma concentration-versus-time curve (AUC(0-infinity)) for lactone and total forms of 9-NC, respectively. The values of mean residence time and elimination half-life (T(1/2)) were also significantly higher for NPs than for free drug. The in-vitro cytotoxicity study revealed that the IC50 value of NPs decreased 10-fold compared with the drug solution. Prepared NPs described here were considered potentially useful in both stabilizing and delivering 9-NC and enhancing the efficacy of this drug for cancer treatment for which high drug retention in the body, protection from the drug-active lactone form, and gradual drug release appeared to be related.