Thiolated Nanostructured Lipid Carriers For Improving Oral Absorption Of Docetaxel

Oral delivery is the most convenient administration route in clinic,and easily accepted by patients.Since most of the anticancer drugs,classified as a BCS ? drug,exhibits poor aqueous solubility,poor intestinal permeability,and,in particular,the low bioavailability obtained after oral administration,Therefore,it is very important to develop effective drug delivery system to improve the bioavailability of anticancer drugs.Lipid-based carrier is widely used in oral drug delivery owing to it's excellent biocompatibility.In this dissertation,we tried to improve the oral bioavailability of docetaxel by nanostructured lipid carriers.In addition,thiolated polymers had better mucoadhesion and enhancement of drug permeation,we considered to further enhance the oral bioavailability of docetaxel by thiols modification of nanostructured lipid carriers.In this study,docetaxel was selected as a model drug,docetaxel-loaded nanostructured lipid carriers(DNLCs)were prepared by emulsification-ultrasonication.The types and amounts of solid lipids,liquid lipids and emulsifiers were screened and optimized.The physicochemical properties,gastrointestinal stability and in vitro release profile of DNLCs were investigated.Differential scanning calorimetry(DSC)demonstrated that the drug was present in a amorphous or molecular state.FTIR analysis suggested that the interaction of DTX and lipids involved hydrogen bonding and hydrophobic interactions.The stability of DNLCs in the gastrointestinal(GI)tract was investigated using simulated GI fluids,which indicated that nanostructured lipid carriers exhibited good stability in GI.The in vitro release behavior of DNLCs was performed using dynamic dialysis method.The formulation DNLCs exhibited sustained drug release compared with docetaxel solution,and the drug released from DNLCs was mainly followed by Fickian diffusion.The absorption site of DNLCs in the GI tract was investigated by intragastrical and intraduodenal administration to rats,the results indicated that intestine was the main absorption site of docetaxel,and the drug absorption in the intestine was markedly improved by nano structured lipid carriers.After oral administration,UPLC-MS/MS method was used to study the pharmacokinetics of DNLCs and docetaxel solution,the absolute bioavailability of DNLCs was 9.05%,which was 4.31-fold of docetaxel solution.Utilizing an everted intestinal rings the uptake mechanism for DNLCs was investigated.Clathrin,caveolae and macropinocytosis were involved in the endocytosis pathway of DNLCs.In order to elucidate the intestinal transport pathway,rats were treated with the lymphatic transport inhibitor,cycloheximide.The AUC0-12 h of DNLCs was significantly reduced about 73%,suggesting that DNLCs were transported into systemic circulation via portal vein and intestinal lymphatic pathway.The intestine cells morphology was observed by TEM,and chylomicrons were formed in the cytoplasm,it further indicated that DNLCs could be transported by lymphatic route.Histological study showed that no significant changes in the structures of the intestine exposed to blank NLCs and the NLCs could be considered as safe for oral drug administration purposes.In order to prolong the residence time of nanostructured lipid carriers in the GI tract,cysteine was introduced in the surface of the DNLCs,and PEG2000-monostearate(PEG2000-MSA)as a linker.Thiolated PEG2000-monostearate(Cy-PEG-MSA)was synthesized using PEG2000-MSA and L-cysteine mediated by 4-Nitrophenyl chloroformate.The structure of Cy-PEG-MSA were characterized by 1H-NMR and FTIR.The amount of thiol groups on Cy-PEG-MSA was quantified using Ellman's reagent and thiol groups content was 299.2±14.0 ?mol/g.Thiolated nanostructured lipid carriers were prepared by emulsification-ultrasonication combined with the post-insertion method.The XPS results showed that the cysteine-unmodified NLCs(uNLCs)did not have any sulfur on the surface of the nanoparticles,but the cNLCs had sulfur on their surface.The in vitro release behavior of different formulations(docetaxel solution,uNLCs and cNLCs)were performed using dynamic dialysis method.The drug release was not interfered by cysteine and the in vitro release of DTX from uNLCs and cNLCs corresponds to the Ritger-Peppas model.Mucoadhesion studies of uNLCs and cNLCs were performed by mucus glycoprotein assay.Conjugation of cysteine on the uNLCs significantly improved their mucoadhesion.The absorption of different formulations(docetaxel solution,uNLCs and cNLCs)in the different intestinal segment(duodenum,jejunum and ileum)was examined by the in situ single-pass intestine perfusion method.The absorption rate(Ka)and effective permeability(Peff)were markedly higher than those of unmodified NLCs and DTX solution in total intestinal segments.The results obtained indicated that thiolated NLCs can improve the docetaxel intestinal permeability.In vivo imaging system captured pictures also showed that cNLCs not only increased intestinal absorption but also improved accumulation in blood.The methodology of everted sacs was used to investigate the uptake way of cNLCs into intestine.The absorption of cNLCs was controlled by clathrin-mediated,caveolae-mediated,macropinocytosis and caveolae-and clathrin-independent endocytosis.The in vivo pharmacokinetics of different formulations were investigated by gavage,the AUC0-24 h of cNLCs was 1533.00±100.91 ng/mL·h,which was 12.3-fold,and 1.64-fold higher than docetaxel solution and uNLCs,respectively.The results indicated that thiolated NLCs could significantly improve the oral absorption of docetaxel.