Pluronic/TPGS Nanoliposomes Containing Folic Acid Targeting As An Anti-breast Cancer Drug Carrier

Breast cancer is the most common cancer in women,and chemotherapy has an irreplaceable place in the treatment of breast cancer.However,many chemotherapeutic drugs suffer from low solubility and serious toxic side effects,so the delivery of chemotherapeutic drugs by nano drug delivery systems has become a hot research topic in recent years.Liposomes are a more maturely studied drug carrier,and surface modification of liposomes can improve their poor stability and weak targeting defects.Pluronic is an amphiphilic surfactant,D-α-tocopheryl polyethylene glycol 1000 succinate(TPGS)is an excellent emulsifier,and folic acid(FA)is a water-soluble vitamin commonly used as a targeting ligand.There are few studies on the co-modification of liposomes by Pluronic,TPGS and folic acid,and the effects of co-modification on the physicochemical properties,carrier properties and targeting behavior of liposomes are still unknown.In this thesis,liposomes with different Pluronic F87 to TPGS mass ratios were constructed to explore the changes in their physicochemical and structural properties and then optimize the matrix composition of liposomes.On this basis,FA-F87 was synthesized chemically and curcumin was used as a model drug to prepare F87/TPGS curcumin liposomes containing folic acid targeting(cur-FA-F87/TPGS-Lps)to investigate its carrier properties and its targeting behavior at the cellular level.The main findings are as follows:(1)Liposomes with different Pluronic F87 to TPGS mass ratios were constructed using a thin film dispersion method combined with dynamic highpressure microfluidization(DHPM).The results of the characterization using laser particle size measurement and transmission electron microscopy showed that the particle size of cur-F87/TPGS-Lps(40-60 nm)with TPGS addition was smaller than that of cur-F87-Lps(95.9 nm),and the particle size decreased gradually with increasing the proportion of TPGS,and both had good dispersion and a spherical morphology.The encapsulation rate was highest at a Pluronic F87 to TPGS mass ratio of 1:2.In vitro release studies showed that both cur-F87-Lps and curF87/TPGS-Lps exhibited a slow release after the initial phase of abrupt release,and the higher the proportion of TPGS,the lower the cumulative release rate of curcumin.The storage stability experiments showed that cur-F87/TPGS-Lps(1:2)and curF87/TPGS-Lps(1:4)had good stability during storage at 4 ℃.In the antioxidant activity study,both cur-F87-Lps and cur-F87/TPGS-Lps showed a concentrationdependent scavenging ability of DPPH radicals,and cur-F87/TPGS-Lps had a stronger scavenging ability of DPPH radicals compared to cur-F87-Lps.By comparing the particle size,drug encapsulation rate,stability and antioxidant activity of the obtained curcumin liposomes,we finally determined that the ratio of Pluronic F87 to TPGS was 1:2 to obtain curcumin liposomal drug carriers with small particle size,high encapsulation rate and good stability.(2)FA-F87 was synthesized by dehydration condensation,and the successful linkage of FA-F87 was determined by nuclear magnetic resonance technique,and the folic acid content of 36.35 % was determined using UV spectrophotometer.F87-Lps,FA-F87-Lps,F87/TPGS-Lps and FA-F87/TPGS-Lps were prepared by film dispersion method combined with DHPM,and the particle size and distribution of liposomes were determined by dynamic light scattering method.The results showed that the particle size distribution of these four liposomes was relatively uniform.In addition,the particle size of F87-Lps and F87/TPGS-Lps increased after FA modification,and the particle size of F87-Lps and FA-F87-Lps decreased after TPGS modification.Breast cancer MCF-7 cells overexpressing folate receptor were selected as a cell model and the cytotoxicity of liposomes was studied by MTT method.The results showed that liposomes F87-Lps and FA-F87-Lps without TPGS addition were not toxic to cancer cells,indicating that these two liposomes have good biocompatibility.In contrast,the liposomes F87/TPGS-Lps and FA-F87/TPGSLps with TPGS addition exhibited some toxicity.(3)Cur-F87-Lps,cur-FA-F87-Lps,cur-F87/TPGS-Lps and cur-FA-F87/TPGSLps were prepared by a thin film dispersion method combined with DHPM.The results of the characterization using laser particle size measurement and transmission electron microscopy showed that the four curcumin liposomes had a relatively uniform particle size distribution and a spherical morphology.In addition,the particle sizes of cur-F87-Lps and cur-F87/TPGS-Lps were increased by FA modification,and the particle sizes of cur-F87-Lps and cur-FA-F87-Lps were decreased by TPGS modification.MCF-7,an over-expressed folate receptor,was selected as a cell model and the cytotoxicity assay of curcumin liposomes was studied by MTT method.The results showed that cur-FA-F87-Lps and cur-FAF87/TPGS-Lps containing folic acid were better targeted to MCF-7 cells compared with cur-F87-Lps and cur-F87/TPGS-Lps without folic acid.In addition,curF87/TPGS-Lps and cur-FA-F87/TPGS-Lps with TPGS were more toxic to MCF-7cells compared to cur-F87-Lps and cur-FA-F87-Lps without TPGS addition.And the toxicity of these four curcumin liposomes to MCF-7 cells was cur-FA-F87/TPGSLps > cur-F87/TPGS-Lps > cur-FA-F87-Lps > cur-F87-Lps,indicating that curcumin liposomes containing both folic acid and TPGS modifications could jointly enhance the cytotoxicity to MCF-7.Finally,the intracellular distribution of curcumin liposomes after uptake by cells was observed by fluorescence characterization assay using the autofluorescence property of curcumin.The results showed that curcumin liposomes were mainly present in the cytoplasm of MCF-7 cells after uptake by the cells.