利用卵泡刺激素肽偶联的纳米颗粒系统进行靶向基因沉默可提高其在体外卵巢透明细胞癌中的特异性和疗效。
Targeted gene silencing using a follicle-stimulating hormone peptide-conjugated nanoparticle system improves its specificity and efficacy in ovarian clear cell carcinoma in vitro.
机构信息
Obstetrics and Gynecology Hospital, Fudan University, Shanghai 200011, People's Republic of China.
出版信息
J Ovarian Res. 2013 Nov 20;6(1):80. doi: 10.1186/1757-2215-6-80.
BACKGROUND
RNA interference technology has shown high therapeutic potential for cancer treatment. However, serum instability, poor tissue permeability and non-specific uptake of short interfering RNA (siRNA) limit its administration in vivo. To overcome these limitations and improve the specificity for ovarian cancer, we developed a targeted nanoparticle delivery system for siRNA. This system included follicle-stimulating hormone (FSH) β 33-53 peptide as a targeting moiety that specifically recognized FSH receptor (FSHR) expressed on ovarian cancer cells. Growth regulated oncogene α (gro-α) has been reported to be involved in ovarian cancer development and progression. Thus, siRNA targeted to gro-α was used as an antitumor drug in this delivery system.
METHODS
FSH β 33-53 peptide-conjugated gro-α siRNA-loaded polyethylene glycol (PEG)-polyethylenimine (PEI) nanoparticles (FSH33-G-NP) were prepared and characterized by gel retardation assay and transmission electron microscopy. Particle size and zeta potential were determined. Expression of gro-α mRNA and protein was detected by real-time quantitative RT-PCR, immunocytochemistry and enzyme-linked immunosorbent assay. The proliferation, migration and invasion of the ovarian clear cell carcinoma cell line ES-2 were evaluated by cell counting kit-8 assay, cell scratch assay and transwell migration assay.
RESULTS
A siRNA sequence that is effective in silencing gro-α expression was obtained and loaded into the targeted delivery system. Compared with gro-α siRNA-loaded nanoparticles without FSH peptide modification (G-NP), FSH33-G-NP significantly down-regulated gro-α expression in ES-2 cells at mRNA and protein levels. Consequently, the aggressive biological behaviors of ES-2 cells, including proliferation, migration and invasion, were suppressed after silencing gro-α expression, and the addition of the FSH β 33-53 peptide enhanced the suppressive effects.
CONCLUSIONS
This study indicated that a FSHR-mediated delivery system could mediate the highly selective delivery of siRNA into ovarian cancer cells and that silencing gro-α expression could be a potential choice for ovarian cancer treatment.
背景
RNA 干扰技术在癌症治疗方面显示出很高的治疗潜力。然而，血清不稳定性、组织通透性差和短干扰 RNA（siRNA）的非特异性摄取限制了其在体内的应用。为了克服这些限制并提高对卵巢癌的特异性，我们开发了一种用于 siRNA 的靶向纳米颗粒递药系统。该系统包括卵泡刺激素（FSH）β 33-53 肽作为靶向部分，其特异性识别卵巢癌细胞上表达的 FSH 受体（FSHR）。生长调节癌基因α（gro-α）已被报道参与卵巢癌的发生和发展。因此，靶向 gro-α 的 siRNA 被用作该递药系统中的抗肿瘤药物。
方法
制备 FSHβ 33-53 肽偶联 gro-α siRNA 负载的聚乙二醇（PEG）-聚亚乙基亚胺（PEI）纳米颗粒（FSH33-G-NP），并通过凝胶阻滞实验和透射电子显微镜进行表征。测定粒径和 Zeta 电位。通过实时定量 RT-PCR、免疫细胞化学和酶联免疫吸附试验检测 gro-α mRNA 和蛋白的表达。通过细胞计数试剂盒-8 检测、细胞划痕实验和 Transwell 迁移实验评估卵巢透明细胞癌细胞系 ES-2 的增殖、迁移和侵袭。
结果
获得了有效沉默 gro-α 表达的 siRNA 序列，并将其载入靶向递药系统。与未用 FSH 肽修饰的 gro-α siRNA 负载的纳米颗粒（G-NP）相比，FSH33-G-NP 可显著下调 ES-2 细胞中 gro-α 的 mRNA 和蛋白表达。因此，沉默 gro-α 表达后，ES-2 细胞的侵袭、迁移和侵袭等侵袭性生物学行为受到抑制，而添加 FSHβ 33-53 肽则增强了这种抑制作用。
结论
本研究表明，FSHR 介导的递药系统可以介导 siRNA 高度选择性地递送至卵巢癌细胞，沉默 gro-α 表达可能是卵巢癌治疗的一种潜在选择。
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