Synthesis and characterization of glycol chitosan DNA nanoparticles for retinal gene delivery.
Synthesis and characterization of glycol chitosan DNA nanoparticles for retinal gene delivery.
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DOI：
10.1002/cmdc.201300371
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发表时间：
2014-01
期刊：
影响因子：
3.4
通讯作者：
Naash, Muna I.
中科院分区：
文献类型：
--
作者：
Mitra, Rajendra N.;Han, Zongchao;Merwin, Miles;Al Taai, Muhammed;Conley, Shannon M.;Naash, Muna I.
鉴于单基因眼病的数量和非单基因退行性眼病的数量，基因治疗已被认为是一种治疗方法，DNA的有效治疗递送策略的发展是一个关键的研究目标。在这里，我们生成，表征和评估由乙二醇壳聚糖（GCS）和质粒DNA（pDNA）组成的非病毒纳米颗粒。我们表明，这些颗粒是稳定的，不聚集在盐水中，是耐DNA酶，并有一个250 nm的流体动力学直径。我们进一步表明，这些纳米颗粒中的质粒保持其正确的构象，可以在细胞内释放和表达。为了确定这些NP是否适合眼内使用，将携带普遍表达的CBA-eGFP表达盒的pDNA压缩并视网膜下注射到成年WT白化病小鼠中。在注射后（PI）第14天，我们观察到仅在用GCS NP处理的眼睛中的视网膜色素上皮（RPE）中有大量GFP表达，但在未压实的pDNA或媒介物（盐水）处理的眼睛中没有。我们没有观察到总体视网膜毒性的迹象，并且在PI-30天时，GCS NP-、pDNA-或媒介物处理的眼睛之间的视网膜电图功能没有差异。这些结果表明，随着进一步的发展，GCS NP可能是我们现有的用于治疗RPE相关疾病的遗传疗法的有用补充。
Given the number of monogenic ocular diseases and the number of non-monogenic degenerative ocular diseases for which gene therapy has been considered as a treatment, the development of effective therapeutic delivery strategies for DNA is a critical research goal. Here we generate, characterize, and evaluate non-viral nanoparticles composed of glycol chitosan (GCS) and plasmid DNA (pDNA). We show that these particles are stable, do not aggregate in saline, are resistant to DNases, and have a hydrodynamic diameter of ∼250 nm. We further show that the plasmid in these NPs maintains its proper conformation and can be released and expressed inside the cell. To determine whether these NPs would be suitable for intraocular use, pDNA carrying the ubiquitously expressed CBA-eGFP expression cassette was compacted and subretinally injected into adult WT albino mice. At post-injection (PI) day 14, we observe substantial GFP expression exclusively in the retinal pigment epithelium (RPE) in eyes treated with GCS NPs but not in uncompacted pDNA or vehicle (saline) treated eyes. We observe no signs of gross retinal toxicity and at PI-30 days, there is no difference in electroretinogram function between GCS NP-, pDNA-, or vehicle-treated eyes. These results suggest that with further development GCS NPs may be a useful addition to our available repertoire of genetic therapies for the treatment of RPE-associated diseases.
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