Construction And Evaluation Of Pore-Forming Peptide Modified Doxorubicin Liposomes

Doxorubicin(DOX)is a member of the anthracycline antibiotics with abroad-spectrum anti-tumor activity,but it is highly toxic to normal cells and diseased cells.Improving the target activity of DOX and reducing it toxicity is the key to use of DOX in anti-tumor therapy.Liposomes are bilayer vesicles.As a new kind of drug delivery systems,liposomes have the advantages of improving drug targeting and reducing toxic effect.Appropriate targeted modification of liposomes can further enhance the anti-tumor effect and reduce toxic effect.Matrix metalloproteinases(MMPs)is an enzyme overexpress in tumor tissues which can degrade various protein components in etracellular matrix(ECM).ECM is the histological barrier of tumor cell invasion,so MMPs play? key role in tumor invasion and metastasis.Pore-forming peptide is a linear cationic channel peptide that can effectively penetrate the lipid membrane and self-assemble in the lipid membrane to form pores,thereby enabling release of small molecules in the membrane.MMP-sensitive pore-forming peptide modifcation of the liposomes could signifcantly improving the selective accumulation of drug in tumor cells and increasing the effectiveness of drug treatment.The DOX liposomes were prepared by ammonium sulphate gradients method and their particle size and encapsulation efficiency were characterized.Functional polymer were synthesized by the reaction between the sulfhydryl groups of pore-forming peptide and the maleimide groups of PEG.Pore-forming peptide modifed DOX liposome system was successfully constructed by the interaction between the functional polymer and the DOX-loaded liposomes.Pore-forming peptide modifed DOX liposomes are responsive to MMPs.In this paper,the enzyme-sensitive properties of the liposome carrier was evaluated by the introduction of MMPs.Part one Preparation and characterization of lipid carrierObjective: Preparation and characterization of saturated phospholipid lipid carriers and unsaturated phospholipid lipid carriers.Methods: The DOX liposomes were prepared by thin film dispersion combined with ammonium sulphate gradient method.The particle size distribution and the entrapment efficiency of the drug loaded liposomes were investigated.Results: In this research,two kinds of liposome carriers(group of unsaturated phospholipid and group of saturated phospholipid)were prepared.The prepared liposomes had good appearance,and the particle size showed a normal single peak distribution.The mean diameter of the liposomes were between 100 nm and 110 nm,with an entrapment efficiency more than 90%and PDI less than 0.3.Conclusions: In this study,the liposome prepared has a high encapsulation efficiency and the preparation method is simple and with a good reproducibility.Part two To investigation of the permeability of lipid carrierObjective: To determine the permeability of pore-forming peptide to lipid carrier by release test.Methods: By examining the diffusion of doxorubicin solution under different conditions,the best condition for the release test were selected,and then the permeability of the pore-forming peptide to lipid carriers were examined by the release test.Results: The permeability test results showed that all pore-forming peptide used have different permeability to liposome carrier,and within a certain range,there is a positive correlation between the molar percentage of the pore-forming peptides in the lipid material and the permeability of the liposome.Conclusions: All pore-forming peptides used in this study have good permeability to liposome carrier.Part three Evaluation of enzyme-sensitive properties of the lipid carrierObjective: To establish a method for evaluating the enzyme-sensitive properties of the liposome carrier as well as preliminary evaluating the enzyme-sensitive properties of the liposome.Methods: Functional polymer were synthesized by the Michael reaction between the sulfhydryl groups of pore-forming peptide and the maleimide groups of PEG.We constructed the MMPs-sensitive DOX-loaded liposome by the interaction between the functional polymer and the DOX-loaded liposome.In this study,the enzyme-sensitive properties of the liposome carrier was evaluated by the introduction of MMPs.Results: We optimize the enzyme activation condition that can be used to study the enzyme-sensitive properties of the liposome.A series of functional polymer were synthesized by the reaction between pore-forming peptide and different types of PEG,and DOX liposomal was decorated with different functional polymer to construct the pore-forming peptide modified DOX liposomes.Finally,a method for evaluating the enzyme-sensitive properties of lipid carriers was established.As demonstrated by the assay via this method,the carrier can suffer from an enzyme-sensitive drug release.Conclusions: By examining the leakage of DOX in the lipid carrier under the action of collagenase,we evaluated the enzyme-sensitive properties of the liposome carrier.The results showed that the carrier has obvious enzyme-sensitive properties.