Drug Delivery System Loading SiRNA And Chemotherapy

Breast cancer is the most common cancer of women,keeping the sustained faster growth of incidence and mortality than other cancers.We designed and synthesized siRAN to knockdown 17β-HSD which occupies a key position for estrogen-dependent breast DNAbinding chemotherapy drug,has been co-encapsulated in a liposome-polycation-DNA nanoparticle drug delivery system(DDS).We design the DDS armed with EPPT,a peptide motif targeting above 90%breast cancer cells,and modified with MCP(a matrix metalloproteinases-sensitive peptide)and GALA(a pH-senstive peptide.Above combined strategies act like a relay-race to co-delivery siRNA and DOX and provide an efficient strategy for reduction of estrogen levels and a chemotherapy.The study could provide theoretical and experimental basis of intelligent materials and innovation for the cancer treatment.Furthermore if the most efficient gene silencing activity and chemotherapy could be achieved,these combina tions are very promising in clinical applications.Firstly,functional material EPPT-PEG2000-DSPE was synthesized and evaluated.Solidphase peptide synthesis was used for the synthesis of EPPT-cys,which was one of the starting material the structure identified by MS,HPLC et al.Then the product EPPT-PEG2000-DSPE was synthesized by conjugating bifunctional PEG with DSPE and EPPT-cys.The purity test was perfoemed.Secondly,different nano-drug delivery systems were prepared:(1)Liposome-Polycation-DNA(LPD)delivery systems:To improve the efficiency in vivo we have successfully developed a core/shell type of nanoparticle formulation,called liposome-polycation-DNA complex(LPD).The formulation was optimized using size and distribution and encapsulation as evaluation.The results showed the optimized formulation protamine/(siRNA/calf thymus DNA)weight of ratio was 0.8,and a PEGylation degree of liposomes 10 mol%.(2)siRNA-loaded nanoparticles with dendritic poly-L-lysine:we further investigate siRNA delivery with dendritic dendrigraft poly-L-lysine(G5)in parallel.The formulation was also optimized using similar evaluation parameters.The results showed that the optimized formulation was characterized by a G5/siRNA mole ratio 0.3.(3)Dox-loaded liposomes:The novel liposomal drug delivery system was developed using film hydration.The results shown that the encapsulation efficiency was 90%and the drug loading was up to 98%.The development of Dox-liposomes could be applied in applied in the proof-of-concept tests for intelligent targeting system and combined administration.(4)Fluorescent-loaded Liposomes:Method of "film hydration" was used in preparation process.The particle size and distribution of liposomes were controlled in the end.Two different fluorescent-loaded liposomes,5-Fluorescein(5-FAM)and Dioctadecyltetramethyl indo tricarbocyanine iodide(Dir)were successfully developed for following in vitro and in vivo targeting ability tests.Finally,the proof-of-concept tests of intelligent targeting system were performed.The in vitro growth inhibition of the intelligent targeting system that had pH-sensitive and enzymesensitive functional materials on 4T1 breast cancer cells model was used to investigate by MTT assay.Meanwhile,the in vivo pharmacodynamics study of intelligent targeting system that had pH-sensitive and enzyme-sensitive functional materials were investigated on 4T1 breast cancer cells model.The results showed that comparing with the control group,the liposome drug delivery system had enhanced inhibition effect on breast cancer.In summary,in this study we firstly synthesized targeting peptide EPPT-PEG2000-DSPE for development of intelligent liposomes.Several breast tumor nano-drug delivery systems were developed.The optimization and development of siRNA-loaded deliveries,i.e.LPD and DGL-G5 were conducted as a focus to improve the siRNA in vivo efficiency.The proof-ofconcept tests of intelligent targeting system showed it had enhanced inhibition effect on breast cancer.Although the targeting ability tests for EPPT and pharmacodynamics study of combined siRNA with DOX are on going.We hope our study had advanced effect in breast cancer therapy.