A new kind of biodegradable pH-sensitive drug delivery system was developed via chitosan-albumin conjugate hydrogel. Through changing the feeding modes of reactants, two types of hydrogels（comb-type and reticular-type） were synthesized by amidation reactions between 6-O-succinoylated N-phthaloyl chitosan and albumin. The structures and morphologies of the hydrogels were characterized by SEM. And their water swelling capacity, drug loading and releasing properties at different pH values were also investigated. It was found that the comb-type of hydrogels with looser space construction had better water swelling ratio（more than 400% of its original mass） than the reticular-type of ones did（about 180% of its original mass）. In vitro release experiments of Rifampicin show that the hydrogels provided the controlled release of the entrapped drug for more than 50 h. The drug release rates of both types of hydrogels under acidic condition were lower than those under neutral or basic condition. The introduction of albumin not only improved the hydrophilicity of chitosan, but also provided the possibility of the carrier system combining other biologically active materials more easily to fulfill the delivery and therapy functions.
Two kinds of paclitaxel（PTX） conjugate micelles,of which one contained 25%（mass fraction） PTX [M（PTX）] and the other contained 22.5%（mass fraction） of PTX and 1.4%（mass fraction） of folate（FA）[FA-M（PTX）],were prepared for cell apoptosis and anti-tumor activity evaluation on U14 cervical cancer mouse models in comparison with 0.9%（mass fraction） saline（control） and equivalent Taxol.Seven days after tail intravenous injection of the drugs,the mice were sacrificed to measure the tumor masses.The average tumor masses were 4.26,2.89,2.63,and 2.17 g for the control,Taxol,M（PTX） and FA-M（PTX） groups,respectively.The inhibition rates of tumor growth calculated for the three drug groups were 32%,38% and 49%,respectively.Flow cytometry（FC） analysis and terminal deoxynucleotidyl transferase（TdT）-mediated deoxyuridine triphosphate（dUTP） nick end labeling（TUNEL） assay were conducted on the cancer tissues.The cell apoptosis rates based on the FC data and the TUNEL data were 20%,31%,37%,42%,and 10%,22%,26%,34%,respectively,both showing statistically significant differences（P〈0.05） between three drug groups and the control group,and between the FA-M（PTX） group and the other two drug groups.In conclusion,the composite FA-M（PTX） micelles can be used for U14 cervical cancer treatment.
To further enhance the transfection efficiency of a micelleplex system based on monomethoxy poly（ethylene glycol）-block-poly（e-caprolactone）-block-poly（L-lysine） （MPEG-b-PCL-b-PLL）, cholesterol （Chol） moieties are attached to the e-termini of PLL segments to obtain MPEG-b-PCL-b-PLL/Chol. The structure and morphology of the copolymer are studied by IH-NMR, TEM and DLS （dynamic light scattering）. The cytotoxicity, cell uptake, endosomal release and mRNA knockdown are studied by MTT （3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide） assay, flow cytometry, CLSM （confocal laser scanning microscopy） and RT-PCR （real-time polymerase chain reaction）. The results show that compared to their precursor MPEG-b-PCL-b-PLL, the cholesterol-grafted copolymer shows significantly lower toxicity, more rapid cellular endocytosis and endosome escape, and consequently displays enhanced siRNA transfection efficiency even at a lower N/P ratio. These improvements are ascribed to enhanced interaction of the cholesterol moieties with both cellular membrane and endosomal membrane. Moreover, effect of the PLL block length is examined. The final conclusion is that long enough PLL segments and incorporation of proper fraction of cholesterol onto the PLL segments benefit the enhancement of siRNA transfection efficiency.
Enhanced permeation and retention（EPR） targeting effect of rhodamine B labeled PEG-b-P（LA-co-DHP） [PEG：poly（ethylene glycol）;LA：L-lactide;DHP：2,2-dihydroxylmethyl-propylene carbonate] micelles（RhB-micelles） was observed in H22 liver cancer bearing mice.The RhB-micelles were prepared by conjugating rhodamine B with the DHP units of amphiphilic block copolymer PEG-b-P（LA-co-DHP） followed by subsequent self-assembling of the conjugate.The parent copolymer PEG-b-P（LA-co-DHP） was synthesized by ring-opening copolymerization of LA and DHP with PEG as macroinitiator and diethyl zinc（ZnEt2） as catalyst.The micelles have a spherical shape and the average diameter is ca.50 nm by TEM（transmission electron microscope） or 80 nm by DLS（dynamic light scattering）.Their in vitro cell uptake experiment by CLSM（confocal laser scanning microscopy） and flow cytometry showed preferential internalization of micelles by MCF-7 human breast cancer cells to free RhB.The in vivo tests by live animal imaging and ex vivo excised organ imaging showed that after vena tail injection,free RhB molecules were distributed in the whole body through the circulation system and then gradually metabolized and excreted and there was no preferential partition in tumor bed from the beginning to the end.But the RhB-micelles were preferentially distributed to the tumor bed so that their concentration（fluorescent intensity） in tumor bed got the level of the liver at a certain time point between 1 and 6 h and reached a maximum relative intensity at around 12 h,indicating an obvious EPR effect of RhB-micelles in H22 liver cancer.
LIU Tong-junLIU ShiHU Xiu-liSHENG Shi-houHUANG Yu-binJING Xia-bin
