Studies On Nanostructured Lipid-Polymer Hybrid Carriers Of Mitoxantrone Hydrochloride

In this study, novel nanostructured lipid-polymer hybrid carriers (NLPCs) were developed using mitoxantrone (MTO) as a model drug to enhance the oral bioavailability and antitumor activity via overcoming the multidrug resistance (MDR). Meanwhile the multicomponent co-delivery system were established by simultaneous encapsulating the anticancer drug MTO, BCRP inhibitor ciclosporin (CsA) and Bcl-2 inhibitor (sodium glycocholate, GcNa) into NLPCs and then modifying by PEG-PE (PEG-MTO-CsA-GcNA-NLPCs) to obtain synergetic antitumor activity and reversion of MDR. While MTO and BCRP inhibitor CsA were simultaneously encapsulated into NLPCs (MTO-CsA-NLPCs) to further enhance the oral bioavailabilty of MTO.The in-vitro and in-vivo characteristics of the prepared NLPCs were investigated systematically including pharmaceutical properties, pharmacokinetics, tissue distribution behaviors, cytotoxicity, celluar uptake and endocytosis mechanisms.In the pre-formulation study, the partition coefficient of MTO in Compritol 888 ATO was significantly increased by anionic polymers through the electrostatic interactions with cationic MTO.On the basis of this observation, the negative polymers of dextran sodium sulfate (DS) and carrageenin were introduced into solid-liquid mixed lipids, respectively, to prepare two kinds of NLPCs for intravenous and oral drug delivery.The prepared MTO-NLPCs and MTO-CsA-NLPCs by emulsification-ultrasonic method showed excellent performance of spherical shape and small particle size(134.3 nm and 144.0nm).The incorporation of anionic polymer carrageenin significantly enhanced the encapsulation efficiency of MTO in lipids up to 95.8% and 93.3% by electrostatic interaction. Superior sustained-release characteristics were found in MTO-NLPCs and MTO-CsA-NLPCs because the cumulative release of MTO was 79.6% and 85.2% at 48 h, respectively, and burst-release was effectively avoided.The prepared intravenous formulations MTO-NLPCs and PEG-MTO-CsA-GcNa-NLPCs showed excellent performance of spherical shape and small particle size(130.3 nm and 125.0 nm).The incorporation of anionic polymer DS significantly enhanced the encapsulation efficiency of MTO in MTO-NLPCs and PEG-MTO-CsA-GcNa-NLPCs up to 97.4% and 97.8% by electrostatic interaction. Superior sustained-release characteristics were found in MTO-NLPCs and PEG-MTO-CsA-GcNa-NLPCs because the cumulative release of MTO was 86.9% and 85.7% at 72 h, respectively, and burst-release was effectively avoided.A sensitive and selective LC-MS/MS method was developed for the determination of MTO in biological specimen. Pharmacokinetic behaviors of MTO-NLPCs after oral and intravenous administration in rats were investigated. The results showed that MTO-NLPCs had higher AUC and MRT than drug solution, and showed obvious effect in prolonging the drug circulation time. Tissue distribution of MTO-NLPCs after intravenous administration in mice was investigated. The results demonstrated that compared with drug solution, MTO-NLPCs changed the tissue distribution characteristics obviously. MTO-NLPCs had a higher drug concentration in plasma, spleen and brain, while decreased the distribution of MTO in heart and kidney.To investigate the MDR reversal effect and clarify the possible mechanism of NLPCs, the studies on cytotoxicity, cellular uptake and uptake mechanism were conducted in MCF-7 and MCF-7/MX cells, respectively. The cytotoxicity investigations by MTT method indicated that two NLPCs could partly overcome MTO resistance, while the MTO-CsA-NLPCs and PEG-MTO-CsA-GcNa-NLPCs could completely reverse the MDR in drug-resistant tumor cells of MCF-7/MX cells. The cellular uptake efficiency of two NLPCs were significantly decreased at 4℃and pre-incubation with endocytosis inhibitors of sodium azide and chlorpromazine implying that the NLPCs entered into the MCF-7/MX cells by an endocytosis process mediated by clathrin. The endocytosis inhibitor of indomethacin had no effect on the cellular uptake efficiency of NLPCs indicating possible non-caveolae-mediated endocytosis of NLPCs.The endocytosis process of NLPCs can bypass the efflux of MTO mediated by BCRP, which enhances the accumulation of MTO in resistant cells and overcomes the MDR.