Microencapsulation of PEGylated adenovirus within PLGA microspheres for enhanced stability and gene transfection efficiency
Microencapsulation of PEGylated adenovirus within PLGA microspheres for enhanced stability and gene transfection efficiency
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DOI：
10.1007/s11095-007-9441-y
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发表时间：
2007-12-01
影响因子：
3.7
通讯作者：
Park, Tae Gwan
中科院分区：
文献类型：
--
作者：
Mok, Hyejung;Park, Ji Won;Park, Tae Gwan
目的。为了提高聚乳酸-乙醇酸共聚物(PLGA)微球的稳定性和基因转染活性，用聚乙二醇(PEG)对编码腺病毒(ADV)的绿色荧光蛋白(GFP)进行表面修饰。在病毒表面制备了一系列具有不同聚乙二醇化浓度的聚乙二醇化腺病毒(PEGylated Adv，PEGADV)，并测定了每一种聚乙二醇腺病毒的GFP表达效率。通过施加高剪切均质过程或暴露在低pH条件下，比较了裸露ADV和聚乙二醇腺病毒的物理稳定性。采用水包油(W/O/W)复乳和溶剂挥发的方法，将裸露的ADV或PEGADV微囊包裹在PLGA微球中。测定了10天内PLGA微球的体外累积ADV和聚乙二醇腺病毒释放曲线。定量检测GFP对HeLa细胞的转染率，并用巨噬细胞分析包裹在PLGA微球中的ADV和PEGADV免疫应答的相对程度。在模拟W/O/W配方条件下，聚乙二醇化ADV的物理稳定性比裸ADV有很大提高，如在高剪切均质化过程中暴露在水/有机界面上。在低pH条件下，聚乙二醇腺病毒比阿昔洛韦更稳定。ADV和PEG-AD均以类似的持续方式从PLGA微球中释放。然而，当ADV和聚乙二醇腺病毒包裹的微球转染HeLa细胞时，聚乙二醇腺病毒微球的GFP基因转染率高于腺病毒微球。聚乙二醇腺病毒微球对巨噬细胞的天然免疫应答程度也有所降低。聚乙二醇化的ADV在PLGA微球中比裸露的ADV更安全，因为它们在苛刻的处方条件和微球的酸性微环境下具有更高的物理稳定性，从而提高了基因转染率。
Purpose. Green fluorescent protein (GFP) encoding adenovirus (ADV) was surface modified with polyethylene glycol (PEG) for microencapsulation within poly(lactic-co-glycolic acid) (PLGA) microspheres with the aim of improving stability and gene transfection activity.Methods. A series of PEGylated ADV (PEG-ADV) with different PEG seeding densities on the viral surface was prepared and the GFP expression efficiency of each PEG-ADV in the series determined. The physical stabilities of naked ADV and PEG-ADV were comparatively evaluated by exerting a high shear homogenization process or by exposure to low pH. Naked ADV or PEG-ADV was microencapsulated within PLGA microspheres using a water-in-oil-in-water (W/O/W) double emulsion and solvent evaporation method. In vitro cumulative ADV and PEG-ADV release profiles from PLGA microspheres were determined over a 10-day period. GFP transfection efficiencies into HeLa cells were quantified, and the relative extent of the immune response for ADV and PEG-ADV encapsulated within PLGA microspheres was analyzed using macrophage cells.Results. The physical stability of PEGylated ADV was greatly enhanced relative to that of naked ADV under the simulated W/O/W formulation conditions, such as exposure to an aqueous/organic interface during high shear-stressed homogenization. PEG-ADV was also more stable than ADV at low pH. ADV and PEG-AD were both released from PLGA microspheres similarly in a sustained fashion. However, when the ADV and PEG-ADV encapsulated microspheres transfected into HeLa cells, PEG-ADV microspheres demonstrated a higher GFP gene transfection efficiency than ADV microspheres. The PEG-ADV microspheres also exhibited a reduced extent of innate immune response for macrophage cells.Conclusions. PEGylated ADV could be more safely microencapsulated within PLGA microspheres than naked ADV due to their enhanced physical stability under the harsh formulation conditions and acidic microenvironmental conditions of the microsphere, thereby increasing gene transfection efficiency.