Physicochemical characterization of siRNA-peptide complexes
Physicochemical characterization of siRNA-peptide complexes
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DOI：
10.1002/btpr.13
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发表时间：
2008-07-01
影响因子：
2.9
通讯作者：
Chen, P.
中科院分区：
文献类型：
--
作者：
Law, Maggie;Jafari, Mousa;Chen, P.
短干扰RNA（siRNA）触发RNA干扰（RNAi），其中互补mRNA被降解，导致编码蛋白质的沉默。期望递送载体增加siRNA的溶液稳定性并改善其细胞摄取以克服其快速酶促降解和低转染效率。在这项研究中，精氨酸-9（R9），一种来自HIV 1达特蛋白的细胞穿透肽，被研究作为siRNA的潜在载体。结缔组织生长因子（CTGF）编码siRNA由于其治疗乳腺癌的治疗潜力而被使用。通过紫外维斯光谱和圆二色性（CD）研究了R9与siRNA的相互作用。通过动态光散射（DLS）测定siRNA-R9复合物的流体动力学直径，并通过测量电泳迁移率获得复合物的Zeta电位。还使用UV-vis光谱定量盐添加的影响。siRNA和R9容易通过分子缔合形成复合物/聚集体，伴随着表面电荷随着肽浓度的增加而变化，在siRNA饱和时达到类似于1 μ m的最大流体动力学直径。从UV/维斯光谱和CD测定的R9与siRNA的最高结合比为103：1，从DLS测定的最高结合比为39.1：1（分别对应于2.2：1（+/-）和84：1的电荷比）。结合率的差异可能是因为吸收和光散射之间的信号贡献的差异。本文所述的CTGF siRNA-R9复合物的物理化学表征表明，可以使用各种方法来控制siRNA-肽复合物的性质，这为用肽载体配制siRNA治疗剂提供了基础。
Short interfering RNAs (siRNAs) trigger RNA interference (RNAi), where the complementary mRNA is degraded, resulting in silencing of the encoded protein. A delivery carrier is desired to increase the solution stability of siRNA and improve its cellular uptake to overcome its rapid enzymatic degradation and low transfection efficiency. In this study, Arginine-9 (R9), a cell-penetrating peptide derived from the HIV 1 Tat protein, was investigated as a potential carrier for siRNAs. A connective tissue growth factor (CTGF) encoding siRNA was used because of its therapeutic potential of treating breast cancer. The interaction between R9 and siRNA was studied by UV/vis spectroscopy and circular dichroism (CD). The hydrodynamic diameter of the siRNA-R9 complexes was determined by dynamic light scattering (DLS), and the Zeta potential of the complexes was obtained by measuring the electrophoretic mobility. The effect of salt addition is also quantified using UV-vis spectroscopy. The siRNA and R9 readily formed complexes/aggregates through molecular association, accompanying a change in surface charge with increasing peptide concentration, reaching a maximum hydrodynamic diameter of similar to 1 mu m at siRNA saturation. The highest binding ratio of R9 to siRNA determined from the UV/vis spectra and CD is 103:1 and 39.1:1 from DLS (corresponds to charge ratios of 2.2:1 (+/-) and 84:1, respectively). The difference in binding ratio is possibly because of the difference in signal contribution between absorption and light scattering. The physicochemical characterization of CTGF siRNA-R9 complexes presented here have shown that various methods call be used to control the properties of the siRNA-peptide complexes, which provide a basis for the formulation of siRNA therapeutics with peptide carriers.