Reconstituted apolipoprotein b lipoparticle, a preparation method and uses thereof

Abstract

The present invention provides a method for preparing a reconstituted apolipoprotein B lipoparticle and the method comprises steps of (a) dissolving an apolipoprotein B and a lipid in a first buffer containing 2 M to 8 M urea and 1 wt % to 15 wt % amphiphilic compounds to form a mixture; and (b) dialyzing the mixture against a second buffer containing 0 M to 2M urea and 0 wt % to 0.5 wt % amphiphilic compounds for 1 to several times for lowering concentrations of the urea and the amphiphilic compounds in the mixture. The present invention further provides an apolipoprotein B lipoparticle and a use for the production of an apolipoprotein B lipoparticle used for delivering a hydrophobic substance.

What is claimed is: 
     
         1 . A method for preparing a reconstituted apolipoprotein B lipoparticle, comprising steps of:
 a) dissolving apolipoprotein B and lipid in a first buffer containing 1 wt % to 15 wt % amphiphilic compound and 2M to 8M urea to form a mixture; and   b) dialyzing the mixture against a second buffer for 1 to 5 times to reduce the concentrations of the urea and the amphiphilic compound in the mixture, wherein, in the second buffer, the concentration of urea ranges from 0 to 2 M and the concentration of the amphiphilic compound is ranges from 0 to 0.5 wt %.   
     
     
         2 . The method of  claim 1 , further comprising steps of dissolving a hydrophobic substance in a buffer which has the same composition as the first buffer, in the step a), followed by filtering out the buffer having the hydrophobic substances. 
     
     
         3 . The method of  claim 2 , wherein the hydrophobic substance is selected from a contrast medium or a lipophilic drug. 
     
     
         4 . The method of  claim 3 , wherein the contrast mediums is at least one selected from the group consisting of superparamagnetic iron oxide nanoparticle, Mangafodipir, Gadoxetic acid, Gadopentetic acid, Gadobenic acid, Gadoteric acid and Vistarem. 
     
     
         5 . The method of  claim 3 , wherein the lipophilic drug is at least one selected from the group consisting of tatra-O-methyl nordihydroguaiaretic acid (M4N), paclitaxel and doxorubincin. 
     
     
         6 . The method of  claim 1 , wherein the pH value of the first buffer is 11. 
     
     
         7 . The method of  claim 1 , wherein the pH value of the second buffer ranges from 8.8 to 11. 
     
     
         8 . The method of  claim 1 , wherein the apolipoprotein B is selected from natural apolipoprotein B or synthetic reconstituted apolipoprotein B. 
     
     
         9 . The method of  claim 8 , wherein the apolipoprotein B is at least one selected from the group consisting of apolipoprotein B-100, B-29 and B-48. 
     
     
         10 . The method of  claim 1 , wherein the lipid is at least one selected from the group consisting of phospholipid, cholesterol and triglyceride. 
     
     
         11 . The method of  claim 1 , wherein the amphiphilic compound is selected from non-ionic surfactant or bile acid. 
     
     
         12 . The method of  claim 11 , wherein the non-ionic surfactant is Triton X-100. 
     
     
         13 . The method of  claim 11 , wherein the bile acid is deoxycholic acid. 
     
     
         14 . A reconstituted apolipoprotein B lipoparticle, comprising an outer layer composed of apolipoprotein B and lipid and an inner core, wherein the outer layer comprises at least one of apolipoprotein B and the inner core comprises at least one of a hydrophobic substance. 
     
     
         15 . The reconstituted apolipoprotein B lipoparticle of  claim 14 , wherein the lipoprotein B is selected from natural apolipoprotein B or synthetic apolipoprotein B. 
     
     
         16 . The reconstituted apolipoprotein B lipoparticle of  claim 15 , wherein the apolipoprotein B is at least one selected from the group consisting of apolipoprotein B-100, B-29 and B-48. 
     
     
         17 . The reconstituted apolipoprotein B lipoparticle of  claim 14 , wherein the lipid is at least one selected from the group consisting of phospholipid, cholesterol and triglyceride. 
     
     
         18 . The reconstituted apolipoprotein B lipoparticle of  claim 14 , wherein the hydrophobic substance is selected from a contrast medium or a lipophilic drug. 
     
     
         19 . The reconstituted apolipoprotein B lipoparticle of  claim 18 , wherein the contrast medium is at least one selected from the group consisting of superparamagnetic iron oxide nanoparticles, Mangafodipir, Gadoxetic acid, Gadopentetic acid, Gadobenic acid, Gadoteric acid and vistarem. 
     
     
         20 . The reconstituted apolipoprotein B lipoparticle of  claim 18 , wherein the lipophilic drug is at least one selected from the group consisting of tatra-O-methyl nordihydroguaiaretic acid (M4N), paclitaxel, and doxorubincin.