Folate receptor-targeted nanoparticle delivery of HuR-RNAi suppresses lung cancer cell proliferation and migration.
Folate receptor-targeted nanoparticle delivery of HuR-RNAi suppresses lung cancer cell proliferation and migration.
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DOI：
10.1186/s12951-016-0201-1
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发表时间：
2016-06-21
影响因子：
10.2
通讯作者：
Ramesh R
中科院分区：
文献类型：
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作者：
Muralidharan R;Babu A;Amreddy N;Basalingappa K;Mehta M;Chen A;Zhao YD;Kompella UB;Munshi A;Ramesh R
人抗原R （HuR）是一种RNA结合蛋白，在包括肺癌在内的许多人类癌症中过度表达，并已被证明可以调节几种癌蛋白的表达。此外，HuR在癌细胞中的过度表达与预后不良和治疗抵抗有关。因此，我们假设在癌细胞中靶向抑制HuR可能会抑制几种由HuR调节的癌蛋白，从而产生有效的抗癌效果。为了验证我们的假设，在本研究中，我们研究了叶酸受体-α (FRA)靶向DOTAP：胆固醇脂质纳米颗粒携带HuR siRNA （HuR- fnp）对人肺癌细胞的作用。我们在FRA过表达的人H1299肺癌细胞系中检测了hr - fnp的治疗效果，并与FRA低表达或不表达的正常肺成纤维细胞（CCD16）进行了比较。理化性质研究表明，hhr - fnp粒径为303.3 nm，表面带正电荷（+4.3 mV）。凝胶阻滞和血清稳定性实验表明，FNPs可以有效地保护siRNA免受快速降解。与CCD16细胞相比，H1299细胞的FNP摄取明显更高，表明受体剂量效应。在H1299细胞中的竞争性抑制研究结果表明，HuR-FNPs通过fr介导的内吞作用被有效地内化。生物学研究表明，hr - fnp而非C-FNP（对照siRNA）诱导H1299细胞G1期细胞周期阻滞和凋亡，导致显著的生长抑制。此外，hhr - fnp对H1299细胞的细胞毒性明显高于对CCD16细胞的细胞毒性。H1299细胞活力降低与HuR mRNA和蛋白表达显著降低相关。此外，在hr - fnp处理的H1299细胞中，观察到hr调节的癌蛋白（cyclin D1、cyclin E和Bcl-2）表达减少，p27肿瘤抑制蛋白表达增加，而在c - fnp处理的细胞中则没有。最后，与C-FNP相比，hur - fnp处理的H1299细胞的细胞迁移明显受到抑制。我们的研究结果表明，HuR是肺癌治疗的分子靶点，使用HuR- fnp抑制其在体外产生显著的治疗效果。
Human antigen R (HuR) is an RNA binding protein that is overexpressed in many human cancers, including lung cancer, and has been shown to regulate the expression of several oncoproteins. Further, HuR overexpression in cancer cells has been associated with poor-prognosis and therapy resistance. Therefore, we hypothesized that targeted inhibition of HuR in cancer cells should suppress several HuR-regulated oncoproteins resulting in an effective anticancer efficacy. To test our hypothesis, in the present study we investigated the efficacy of folate receptor-α (FRA)-targeted DOTAP:Cholesterol lipid nanoparticles carrying HuR siRNA (HuR-FNP) against human lung cancer cells. The therapeutic efficacy of HuR-FNP was tested in FRA overexpressing human H1299 lung cancer cell line and compared to normal lung fibroblast (CCD16) cells that had low to no FRA expression. Physico-chemical characterization studies showed HuR-FNP particle size was 303.3 nm in diameter and had a positive surface charge (+4.3 mV). Gel retardation and serum stability assays showed that the FNPs were efficiently protected siRNA from rapid degradation. FNP uptake was significantly higher in H1299 cells compared to CCD16 cells indicating a receptor-dose effect. The results of competitive inhibition studies in H1299 cells demonstrated that HuR-FNPs were efficiently internalized via FRA-mediated endocytosis. Biologic studies demonstrated HuR-FNP but not C-FNP (control siRNA) induced G1 phase cell-cycle arrest and apoptosis in H1299 cells resulting in significant growth inhibition. Further, HuR-FNP exhibited significantly higher cytotoxicity against H1299 cells than it did against CCD16 cells. The reduction in H1299 cell viability was correlated with a marked decrease in HuR mRNA and protein expression. Further, reduced expression of HuR-regulated oncoproteins (cyclin D1, cyclin E, and Bcl-2) and increased p27 tumor suppressor protein were observed in HuR-FNP-treated H1299 cells but not in C-FNP-treated cells. Finally, cell migration was significantly inhibited in HuR-FNP-treated H1299 cells compared to C-FNP. Our results demonstrate that HuR is a molecular target for lung cancer therapy and its suppression using HuR-FNP produced significant therapeutic efficacy in vitro.