Effect of Hydrophilicity/Hydrophobicity of Linker in Gal/GalNAc-containing Ligands on Liver Targeting of Its Modified Liposomes
NIE Hua1, LIU Bao-wen1, ZHAO Ying1, YE Xiao-ling1, HE Ze-he1, YUAN Si-ying1, LIU Xiao-min2*
1. Medical College of Jiaying University, Meizhou 514031, China; 2. Institute of Hakka Medicinal Biological Resources, Medical College of Jiaying University, Meizhou 514031, China
Abstract：OBJECTIVE To investigate the effect of the hydrophilic/hydrophobicity of linker in the Gal/GalNAc-cholesterol(CHS)ligand and its modified liposome surface charge on the affinity of asialoglycoprotein receptor(ASGPR), so as to provide theoretical guidance and experimental basis for construction of high-efficiency Gal/GalNAc ligand-modified drug delivery vector for targeting liver cancer cells. METHODS The enzymatic method was adopted to synthesize Gal/GalNAc-CHS ligands with hydrophobic linker:CHS-C8-GalNAc, CHS-C8-Gal, CHS-C8-Lac, and Gal/GalNAc-CHS ligand with hydrophilic linker:CHS-DIO-GalNAc.Among them, CHS-DIO-GalNAc was reported for the first time. Hydrogenated soya phosphatide, cholesterol and various ligands were mixed in a molar ratio(6∶3∶1), and 1% distearoyl phosphatidylglycerole sodium salt(DSPG-Na) was added to the total lipid mass if anionic liposomes were prepared. Then, ligand-modified liposomal doxorubicin(DOX) was prepared by thin film dispersion-high pressure extrusion-ammonium sulfate gradient method. With mice as the experimental subjects, the liposomal DOX was administered by tail vein, and the distribution characteristics of four kinds of Gal/GalNAc ligand-modified liposomal DOX in blood and liver tissues of mice were compared. RESULTS CHS-DIO-GalNAc was identified as the target product by MS, NMR and HMBC.In addition, the yield was > 90%. The obtained liposomes had rounded appearance, and clear edges, with the particle sizes ranging from 60 nm to 75 nm, and Zeta potential ranging from -6 mv to +5 mV(-4 mV to -17 mV for anionic liposomes added with DSPG-Na), polydispersity index<0.1, encapsulation rate > 98% and leak rate<3%.According to the results of biological distributions experiments in mice, the elimination rate in blood and the accumulation rate in liver tissue of liposomal DOX modified by CHS-DIO-GalNAc were significantly higher than those modified by CHS-C8-GalNAc, CHS-C8-Gal and CHS-C8-Lac.Moreover, the competitive inhibition experiment of ASGPR in vivo, proved that the liver high uptake rate of liposomal DOX modified by CHS-DIO-GalNAc or CHS-C8-GalNAc ligand was the active endocytosis mediated by ASGPR on the surface of liver parenchymal cell membrane. CONCLUSION The stronger the hydrophilicity of the linker in the Gal/GalNAc ligand structure is, the higher the recognition efficiency of ASGPR for Gal/GalNActarget modified on the liposomes surface becomes.At the same time, the addition of negatively charged phospholipids to liposomes can synergistically enhance the affinity of ASGPR for the liposomes modified with Gal/GalNAc ligands.The results of the study will provide beneficial and helpful guide to the design of Gal/GalNAc ligand with high affinity for ASGPR.
聂华, 刘宝雯, 赵莹, 叶小玲, 何泽和, 袁思颖, 刘小敏. Gal/GalNAc配体中连接臂的亲水/疏水性对其修饰脂质体肝靶向性的影响[J]. 中国药学杂志, 2023, 58(1): 65-76.
NIE Hua, LIU Bao-wen, ZHAO Ying, YE Xiao-ling, HE Ze-he, YUAN Si-ying, LIU Xiao-min. Effect of Hydrophilicity/Hydrophobicity of Linker in Gal/GalNAc-containing Ligands on Liver Targeting of Its Modified Liposomes. Chinese Pharmaceutical Journal, 2023, 58(1): 65-76.
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2023, Vol. 58 
