Design And Evaluation Of ATB^0,+Transporter-Mediated Active Targeting Liposomes For Tumor Therapy

In view of the short half-life of small molecule anticancer drugs and the lack of selectivity to tumor tissues,which may lead to undesirable therapeutic effects and severe systemic toxicity,amino acid molecules are used as modified groups to target the amino acid transporters highly expressed in tumor cells.A series of amphiphilic polymers(AA-PS)with different amino acid modified PEG were designed and synthesized.Using docetaxel(DTX)as the model drug,the active-targeting liposomes were prepared by film dispersion method using the synthesized with amino acid-modified polymer AA-PS and PS-Lips/DTX were prepared as control.Their pharmacological properties and in vitro release characteristics were investigated.The cellular uptake mechanisms and anti-tumor activity,as well as tissue distributions were also studied.The anti-tumor effect and safety of the active targeting liposomal drug delivery system were evaluated,and the mechanisms of their transporter-mediated targeting action was described in detail and analyzed by computional methods.The full text of the study includes the following parts:Three kinds of amino acids,aspartic acid,glycine and lysine,were selected as reaction substrates to synthesize amino acid modified polymer AA-PS by esterification with stearylated polyethylene glycol.And the structure of the polymer was confirmed by 1H-NMR.DTX was used as a model drug to be encapsulated in the liposomal drug delivery system.The factors affecting the preparation were investigated and the preparation process of the formulation was optimized.The distribution of liposomes was uniform with a spherical shape of 95?108 nm,possessing high encapsulation efficiency and drug loading.The preparations also showed good sustained-release properties in vitro and high plasma compatibility.Immunohistochemistry and western blot were used to detect the expression of ATB0,+in tumor cell line HepG2 and normal fibroblast L929 cells.It was confirmed that ATB0,+transporter were differentially expressed in tumor tissues and normal tissues.The results of cell uptake showed that the targeting liposomes modified by amino acids could significantly increase the uptake of liposomes in HepG2 cells.The results of MTT assay showed that modified liposomes could enhance the growth inhibition of HepG2 cells,and activate more caspase-related proteins to induce apoptosis of tumor cells.The mechanisms of the uptake was determined by the specific binding of the amino acid substrates and ATB0,+in the process of the transporter-mediated internalization.The uptake of asparate-and lysine-modified liposomes in HepG2 cells was 1.5 and 1.9 times higher than that of the non-targeted liposomes after 5 h incubation.In this paper,the molecular docking experiment and molecular dynamics simulation were first used to simulate the energy change of small molecule substrate and transporter during the recognition process to analyze the transporter-mediated endocytosis.The In vivo tissue distribution of LPS-Lips and PS-Lips in tumor-bearing mice was studied by small animal In vivo imaging.The results showed that LPS-Lips/DiR could significantly increase the drug distribution in tumor tissue and reduce cardiotoxicity and hepatotoxicity through EPR effect by long circulation and active targeting.The tumor growth was significantly inhibited in LPS-Lips/DTX treated group,compared with that in PS-Lips/DTX and Taxotere?treated groups.The inhibitory rate of LPS-Lips/DTX group was 74.1%,which was 1.5 times and 2.0 times higher than that of non-targeting PS-Lips/DTX group and Taxotere? group.In addition,survival status of mice was also improved due to the low side effects of LPS-Lips/DiR.These studies provide first evidences that ATB0,+can be used as a novel and effective target for drug delivery system in tumor cells using chemically modified liposomes for loading with chemotherapeutics and targeting them for the transporter-mediated endocytosis.As ATB0,+is highly upregulated in several cancers,this approach holds potential for tumor-selective delivery of drugs to treat these cancer types.