Institute of Biomedical Engineering, Department of Chemical Engineering, École Polytechnique, P.O. Box 6079, Station Centre-ville, Montréal, Québec, Canada.
Eur J Pharm Sci. 2012 Jan 23;45(1-2):138-49. doi: 10.1016/j.ejps.2011.10.029. Epub 2011 Nov 9.
Glucagon like peptide 1 (GLP-1), a blood glucose homeostasis modulating incretin, has been proposed for the treatment of type 2 diabetes mellitus (T2DM). However, native GLP-1 pharmacokinetics reveals low bioavailability due to degradation by the ubiquitous dipeptydil peptidase IV (DPP-IV) endoprotease. In this study, the glucosamine-based polymer chitosan was used as a cationic polymer-based in vitro delivery system for GLP-1, DPP-IV resistant GLP-1 analogues and siRNA targeting DPP-IV mRNA. We found chitosans to form spherical nanocomplexes with these nucleic acids, generating two distinct non-overlapping size ranges of 141-283 nm and 68-129 nm for plasmid and siRNA, respectively. The low molecular weight high DDA chitosan 92-10-5 (degree of deacetylation, molecular weight and N:P ratio (DDA-Mn-N:P)) showed the highest plasmid DNA transfection efficiency in HepG2 and Caco-2 cell lines when compared to 80-10-10 and 80-80-5 chitosans. Recombinant native GLP-1 protein levels in media of transfected cells reached 23 ng/L while our DPP-IV resistant analogues resulted in a fivefold increase of GLP-1 protein levels (115 ng/L) relative to native GLP-1, and equivalent to the Lipofectamine positive control. We also found that all chitosan-DPP-IV siRNA nanocomplexes were capable of DPP-IV silencing, with 92-10-5 being significantly more effective in abrogating enzymatic activity of DPP-IV in media of silenced cells, and with no apparent cytotoxicity. These results indicate that specific chitosan formulations may be effectively used for the delivery of plasmid DNA and siRNA in a combination therapy of type 2 diabetes.
胰高血糖素样肽 1（GLP-1）是一种调节血糖稳态的肠促胰岛素，已被提议用于治疗 2 型糖尿病（T2DM）。然而，由于广泛存在的二肽基肽酶 IV（DPP-IV）内肽酶的降解，天然 GLP-1 的药代动力学显示出低生物利用度。在这项研究中，基于葡聚糖的聚合物壳聚糖被用作 GLP-1、DPP-IV 抗性 GLP-1 类似物和针对 DPP-IV mRNA 的 siRNA 的基于阳离子聚合物的体外递送系统。我们发现壳聚糖与这些核酸形成球形纳米复合物，分别产生质粒和 siRNA 的两个不同且不重叠的大小范围，分别为 141-283nm 和 68-129nm。与 80-10-10 和 80-80-5 壳聚糖相比，低分子量高 DDA 壳聚糖 92-10-5（脱乙酰度、分子量和 N:P 比（DDA-Mn-N:P））在 HepG2 和 Caco-2 细胞系中显示出最高的质粒 DNA 转染效率。转染细胞培养基中的重组天然 GLP-1 蛋白水平达到 23ng/L，而我们的 DPP-IV 抗性类似物使 GLP-1 蛋白水平增加了五倍（115ng/L），与 Lipofectamine 阳性对照相当。我们还发现，所有壳聚糖-DPP-IV siRNA 纳米复合物都能够沉默 DPP-IV，92-10-5 能够更有效地阻断沉默细胞培养基中 DPP-IV 的酶活性，并且没有明显的细胞毒性。这些结果表明，特定的壳聚糖制剂可有效地用于 2 型糖尿病联合治疗中的质粒 DNA 和 siRNA 的递送。
