The delivery of small interfering RNA (siRNA) and its therapeutic usage as an anti-cancer agent requires a carrier system for selective internalization into the cytosol of tumor cells. We prepared folate-bearing formulations by first complexing siRNA with the novel azido-functionalized sequence-defined cationizable lipo-oligomer 1106 (containing two cholanic acids attached to an oligoaminoamide backbone in T-shape configuration) into spherical, ∼100–200 nm sized lipopolyplexes, followed by surface-functionalization with various folate-conjugated DBCO-PEG agents. Both the lipo-oligomer and the different defined shielding and targeting agents with mono- and bis-DBCO and varying PEG length were generated by solid phase supported synthesis. A bivalent DBCO surface agent with a PEG₂₄ spacer was identified as the optimal formulation in terms of nanoparticle size, folate receptor (FR) targeting, cellular uptake and gene silencing in vitro. Notably, near-infrared fluorescence bioimaging studies showed that double-click incorporation of bivalent DBCO-PEG₂₄ resulted in siRNA/1106/DBCO₂-ss₂-PEG₂₄-FolA lipopolyplexes with extended biodistribution and intratumoral delivery in a subcutaneous FR-positive leukemia mouse model. Intravenous administration of analogous therapeutic siRNA lipopolyplexes (directed against the kinesin spindle motor protein EG5) mediated tumoral EG5 mRNA knockdown by ∼60% and, in combination with the novel antitubulin drug pretubulysin, significantly prolonged survival of aggressive leukemia bearing mice without noticeable side effects.

小干扰RNA（siRNA）的递送及其作为抗癌剂的治疗用途需要一种载体系统，用于选择性内化到肿瘤细胞的细胞质中。我们首先通过将siRNA与新型叠氮基官能化的序列定义的可阳离子化的脂寡聚体1106（含有两个以T型构型连接到寡氨基酰胺骨架上的胆酸相连）的siRNA络合制成球形，约100-200 nm大小的脂质多聚体，然后用各种叶酸偶联的DBCO-PEG试剂进行表面官能化。通过固相支持的合成，既生成了脂寡聚物，又定义了具有单-和双-DBCO和变化的PEG长度的不同定义的屏蔽和靶向剂。具有PEG _24的二价DBCO表面剂在纳米颗粒大小，叶酸受体（FR）靶向，细胞摄取和体外基因沉默方面，spacer被确定为最佳制剂。值得注意的是，近红外荧光生物成像研究表明，双击并入二价DBCO-PEG _24会导致siRNA / 1106 / DBCO ₂ -ss ₂ -PEG ₂₄ -FolA在皮下FR阳性白血病小鼠模型中具有扩展的生物分布和肿瘤内递送的脂多聚体。静脉内施用类似治疗性siRNA脂蛋白复合物（针对驱动蛋白纺锤体运动蛋白EG5）介导的肿瘤EG5 mRNA敲低约60％，并与新型抗微管蛋白药物pretubulysin组合使用，可显着延长侵袭性白血病小鼠的存活率，而无明显副作用。