Abstract
Purpose: In this study the effectiveness of encapsulating of 5-azacytidine into the lipid nanoparticles was investigated and in vitro effect of encapsulated 5-azacytidine studied on MCF-7 cell lines Methods: 5-azacytidine -loaded solid lipid nanoparticles were produced by double emulsification (w/o/w) method by using stearic acid as lipid matrix, soy lecithin and poloxamer 407 as surfactant and co-surfactant respectively. Particle size, zeta potential, surface morphology, entrapment efficiency and kinetic of drug release were studied. In vitro effect of 5-azacytidine on MCF-7 cell line studied by MTT assay, DAPI staining, Rhodamine B relative uptake, and also Real time RT-PCR was performed for studying difference effect of free and encapsulated drug on expression of RARß2 gene. Results: The formulation F5 with 55.84±0.46 % of entrapment efficiency shows zero order kinetic of drug release and selected for in vitro studies; the cytotoxicity of free drug and encapsulated drug in 48 h of incubation have significant difference. DAPI staining shows morphology of apoptotic nucleus in both free and encapsulated drug, Rhodamine B labeled SLNs show time dependency and accumulation of SLNs in cytoplasm. Real time qRT-PCR doesn't show any significant difference (p>0.05) in expression of RARß2 gene in both cells treated with free or encapsulated drug. Conclusion: The results of the present study indicated that the entrapment of 5-azacytidine into SLNs enhanced its cytotoxicity performance and may pave a way for the future design of a desired dosage form for 5-azacytidine.
Highlights
5-azacytidine was first synthesized in 19631 as an effective anti-cancer agent for the treatment of leukemia.[2,3,4] In 2004, the FDA approved the application of 5-azacytidine in treating myelodysplastic syndrome as the first drug effective in epigenetic therapy by inhibiting DNA methyltransferase.[5]
In vitro effect of 5-azacytidine on MCF-7 cell line studied by MTT assay, DAPI staining, Rhodamine B relative uptake, and Real time RT-PCR was performed for studying difference effect of free and encapsulated drug on expression of RARß2 gene
The results of the present study indicated that the entrapment of 5-azacytidine into SLNs enhanced its cytotoxicity performance and may pave a way for the future design of a desired dosage form for 5-azacytidine
Summary
5-azacytidine was first synthesized in 19631 as an effective anti-cancer agent for the treatment of leukemia.[2,3,4] In 2004, the FDA approved the application of 5-azacytidine in treating myelodysplastic syndrome as the first drug effective in epigenetic therapy by inhibiting DNA methyltransferase.[5]. The second strategy is to encapsulate 5-azacytidine in nanoparticles to protect it from enzymatic deamination This concept makes drug uptake independent of transporters by nanoparticle endocytosis into the targeted cells.[16] Nanoparticulate carriers are introduced for passive drug targeting to tumor tissues through the enhanced permeability and retention (EPR) effect.[17,18,19] EPR implies that nanoparticles tend to accumulate in tumor tissue much more than they do in normal tissue. Reported drawbacks of colloidal carriers such as liposomes, nanosponges, microemulsions and nanoemulsions, polymeric nanoparticles, and nanocapsules are burst drug release, physical and chemical instability during storage,[20,21,22,23] difficulty in industrial fabrication, the presence of organic solvents applied in the production of these systems, some limitations in polymer toxicity[24,25] and many more too numerous to mention. The cytotoxicity and uptake of the nanoparticulate system were investigated through MTT assay, DAPI staining, Rhodamine B labeling, and Real Time Quantitative Reverse Transcription PCR investigation on the expression of the retinoic acid receptor β2 (RARß2) gene of MCF-7 (human breast adenocarcinoma cell line)
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