Gaziova Zuzana, Baumann Volker, Winkler Anna-Maria, Winkler Johannes
University of Vienna, Department of Medicinal Chemistry, Althanstraße 14, 1090 Vienna, Austria.
University of Vienna, Department of Medicinal Chemistry, Althanstraße 14, 1090 Vienna, Austria.
Bioorg Med Chem. 2014 Apr 1;22(7):2320-6. doi: 10.1016/j.bmc.2014.02.004. Epub 2014 Feb 20.
The therapeutic application of siRNA suffers from poor bioavailability caused by rapid degradation and elimination. The covalent attachment of PEG is a universal concept to increase molecular size and enhance the pharmacokinetic properties of biomacromolecules. We devised a facile approach for attachment of PEG molecules with a defined molecular weight, and successful purification of the resulting conjugates. We directly conjugated structurally defined PEG chains with twelve ethylene glycol units to the 3'-terminal hydroxyl group of both sense and antisense strands via an aminoalkyl linker. The conjugates were easily purified by HPLC and successful PEGylation and molecule integrity were confirmed by ESI-MS. The evaluation of in vitro gene knockdown of two different targets in MCF-7 breast cancer cells showed stable pharmacologic activity when combined with a standard transfection reagent. Sense strand PEGylation even increased the silencing potency of a CRCX4-siRNA which had modest activity in its wild-type form. The results indicate that PEG chains at the 3'-terminus of both strands of siRNA are well tolerated by the RNAi effector. The attachment of short, chemically defined PEG chains is a feasible approach to improve the pharmacokinetic properties of siRNA, and can be combined with other targeted and untargeted delivery vehicles.
由于快速降解和清除，小干扰RNA（siRNA）的治疗应用存在生物利用度差的问题。聚乙二醇（PEG）的共价连接是一种通用的方法，可增加分子大小并增强生物大分子的药代动力学性质。我们设计了一种简便的方法来连接具有确定分子量的PEG分子，并成功纯化了所得的缀合物。我们通过氨基烷基接头将具有十二个乙二醇单元的结构确定的PEG链直接连接到正义链和反义链的3'-末端羟基上。通过高效液相色谱法（HPLC）很容易纯化缀合物，并通过电喷雾电离质谱（ESI-MS）确认了成功的聚乙二醇化和分子完整性。在MCF-7乳腺癌细胞中对两个不同靶标的体外基因敲低评估表明，与标准转染试剂联合使用时具有稳定的药理活性。正义链聚乙二醇化甚至提高了CRCX4-siRNA的沉默效力，其野生型形式的活性适中。结果表明，RNA干扰效应器对siRNA两条链3'-末端的PEG链耐受性良好。连接短的、化学定义的PEG链是改善siRNA药代动力学性质的一种可行方法，并且可以与其他靶向和非靶向递送载体结合使用。
