Enhanced growth inhibition of hepatic multicellular tumor spheroids by lactosylated poly(ethylene glycol)-siRNA conjugate formulated in PEGylated polyplexes
Enhanced growth inhibition of hepatic multicellular tumor spheroids by lactosylated poly(ethylene glycol)-siRNA conjugate formulated in PEGylated polyplexes
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DOI：
10.1002/cmdc.200700076
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发表时间：
2007-09-01
期刊：
影响因子：
3.4
通讯作者：
Kataoka, Kazunori
中科院分区：
文献类型：
--
作者：
Oishi, Motoi;Nagasaki, Yukio;Kataoka, Kazunori
聚乙二醇化复合物（lac-PEG化聚合复合物），由聚（L-赖氨酸）和乳糖化聚抑制RecQL I基因产物的（乙二醇）-小干扰RNA缀合物显示出对多细胞HuH-7球状体的明显生长抑制（人肝癌细胞系）长达21天（IC 50 = 6 nM）;该系统用作模拟肿瘤体内生物学的体外三维（3D）模型。由此制备的聚乙二醇化复合物具有约110 nm的尺寸，在其外围具有成簇的乳糖部分作为表达无唾液酸糖蛋白受体的HuH-7细胞的靶向配体。相比之下，观察到OligofectAMINE/siRNA（cotionic lipoplex）对HuH-7球状体几乎没有生长抑制作用，即使lipoplex对常规单层培养的HuH-7细胞显示出比lac-PEG化聚合复合物更强的生长抑制作用。FITC标记的共轭物中的lac-PEG化的复合物显示出顺利渗透到HuH-7球状体相比，在lipoplexes，如通过共聚焦荧光扫描显微镜观察。这表明，约100 nm的小尺寸和由于非离子和亲水性乳糖基化PEG层而减少的非特异性相互作用有助于PEG化多聚复合物顺利渗透到球状体内部，最终促进其摄取到组成球状体的细胞中。在用PEG化的多聚复合物处理的HuH-7球状体中也观察到指示程序性细胞死亡的细胞凋亡，揭示了所观察到的生长抑制确实是由RecQL 1 siRNA的RNAi诱导的。这些数据表明，智能聚乙二醇化复合物确实可以渗透到体内3D肿瘤块的多个细胞层中，通过RNAi发挥治疗作用。
PEGylated polyplexes (lac-PEGylated polyplexes) composed of poly(L-lysine) and lactosylated poly(ethylene glycol)-small interfering RNA conjugate, which inhibits the RecQL I gene product, were revealed to show an appreciable growth inhibition of multicellular HuH-7 spheroids (human hepatocarcinoma cell lines) for up to 21 days (IC50 = 6 nM); this system used as an in vitro three-dimensional (3D) model mimicking the in vivo biology of tumors. The PEGylated polyplexes thus prepared had a size of approximately 110 nm with clustered lactose moieties on their periphery as targeting ligands for the asiologlycoprotein-receptor-expressing HuH-7 cells. In contrast, OligofectAMINE/siRNA (cotionic lipoplex) was observed to have almost no growth-inhibitory effect against HuH-7 spheroids, even though the lipoplex showed a stronger growth-inhibitory effect than the lac-PEGylated polyplexes on conventional monolayer-cultured HuH-7 cells. The FITC-tagged conjugate in the lac-PEGylated polyplexes showed smooth penetration into the HuH-7 spheroids compared with that in the lipoplexes, as observed by confocal fluorescence-scanning microscopy. This indicates that the small size of approximately 100 nm and the reduced nonspecific interaction due to the nonionic and hydrophilic lactosylated PEG layer contributes to the smooth penetration of the PEGylated polyplexes into the spheroid interior, eventually facilitating their uptake into the cells composing the spheroids. Cellular apoptosis indicating programmed cell death was also observed in the HuH-7 spheroids treated with the PEGylated polyplexes, revealing that the observed growth inhibition was indeed induced by the RNAi of the RecQL1 siRNA. These data suggest that the smart PEGylated polyplexes can indeed penetrate into the multiple cell layers of 3D tumor masses in vivo, exerting therapeutic effects through the RNAi.