Study On Recombinant Human Interferon α Loaded Long-acting Injectable Microspheres

Interferon-α(IFN-α) is a protein with extensive antiviral, antiproliferative andimmunomodulatory biological activity. It was currently widely used to treat variousviral diseases and many cancers,especially to chronic hepatitis B, viral disease in giantcell and blood source tumor. Ordinarily it is administered intramuscularly andsubcutaneouly. Due to its short circulation half life(5-7 hours), it need to beadministered frequently, which may bring inconvenience to patients. This study isdesigned to use poly(lactic-co glycolic acid) as carrier materials which is degradablein vivo to encapsulate IFN-α,and to control the in vitro release of IFN-αthroughinside pores in microspheres and the erosion and degradation of polymers. The study first begun with the preparation of PLGA microspheres by doubleemulsion (w/o/w) technique. Single-factor experiment was designed to investigate theinfluential factors of the size and encapsulation efficiency of microspheres. Theinfluential factors include stirring rate when preparing double emulsion, polyvinylglycolide(PVA) concentration, PLGA concentration, the osmotic pressure of innerwater phase. It suggested that stirring rate, PVA concentration, PLGA concentration,the osmotic pressure of inner water phase have significant effect on the appearanceand particle size of microspheres. The appearance and particle size are importantfactors to control the release of microspheres. The smaller of microspheres, the moresignificant burst effect of microspheres. The encapsulation efficiency are usuallyhigh(more than 80%),and other factors didn't have significant effect on it exceptPLGA concentration and PVA concentration in our test. It was also found that osmoticpressure of inner water phase could significantly affect the size, appearance andrelease behavior of microspheres. After desalination of inner water phase, the particlesize was smaller; the number of microspheres decreased; the density of microspheresincreased; burst effect of microspheres significantly decreased. According to previous studies, there were plenty of influential factors whenpreparing microspheres by double emulsion method, and due to the water/oil interfaceformed using this technique, protein may be damaged and inactivated. Therefore,solid in oil in oil(s/o/o) method was then studied to prepare IFN-α loaded PLGAmicrospheres. First Zinc-complexed IFN- α microparticles were formulatedsuccessfully which can be redissolved and whose electrophoretic andchromatographic behavior were the same as IFN- α original solution. Fine 3中英文摘要microparticles of IFN-αand its complex with zinc were prepared by phase separationmethod which were then encapsulated into microspheres. PLGA concentration,stirring rate, IFN-α microparticle size, additives in oil phase were investigated toevaluate their effects on particle size, appearance and encapsulation efficiency ofmicrospheres. It was found in our test that PLGA concentration, stirring rate andadditives such as mannitol had significant effects on the appearance and particle sizeof microspheres. Since protein is insoluble in oil phase, encapsulation efficiency ofmicrospheres prepared by s/o/o method was usually very high(more than 90%). Therelease behavior of microspheres prepared by this method can be affectedsignificantly after formulation of Zinc-complexed IFN-α or addition of additivessuch as mannitol and poloxamer 188(F-68), especially burst effect. IFN- α concentration in release medium was determined byBCA(Bicinchoninic acid) protein determination method and the in vitro releasebehavior of microspheres was studied. In vitro test suggested IFN-α loadedmicrospheres had significant long-acting effect in 30 days prepared by both w/o/wand s/o/o methods. In accordance with previous studies, the release behavior ofmicrospheres made by PLGA with molecular weight of...