Hu Yunping, Singh Ravi, Deng Zhiyong, Mintz Akiva, Hsu Wesley
J Biomed Nanotechnol. 2016 Oct;12(10):1952-61. doi: 10.1166/jbn.2016.2236.
Recent evidence suggests that brachyury is a crucial molecular driver in the initiation and propagation of chordoma. However, no small molecules have been used to specifically target brachyury. Short hairpin RNA (shRNA) has therapeutic promise for the genetic treatment of cancer, but the usage of shRNA therapeutics is limited by obstacles related to effective delivery into the nuclei of target cancer cells due to their inherent sensitivity to nucleases and large polyanionic characteristics. To overcome instability and low transfection efficiency, liposome-protamine-DNA (LPD) nanoparticles were synthesized and investigated as a non-viral carrier of shRNA targeting brachyury in chordoma cells. The size, zeta potential, affinity and transfection efficiency of LPD-shRNA complexes were characterized, and their biological functions in chordoma cells were evaluated. The transfection efficiency of LPD-shRNA was significant higher than naked shRNA. LPD delivered brachyury shRNA into chordoma cells and inhibited brachyury expression, induced apoptosis, upregulated the epithelial biomarker, E-cadherin, downregulated the mesenchymal biomarker, Snail and Slug, and suppressed cell growth. These data indicate that LPD might be a promising non-viral carrier for shRNA in gene targeted therapy of chordoma.
最近的证据表明，短尾相关转录因子（brachyury）是脊索瘤发生和发展的关键分子驱动因素。然而，目前尚无小分子可特异性靶向短尾相关转录因子。短发夹RNA（shRNA）在癌症基因治疗方面具有治疗前景，但由于其对核酸酶的固有敏感性和较大的聚阴离子特性，shRNA疗法的应用受到有效递送至靶癌细胞核相关障碍的限制。为了克服不稳定性和低转染效率，合成了脂质体-鱼精蛋白-DNA（LPD）纳米颗粒，并将其作为靶向脊索瘤细胞中短尾相关转录因子的shRNA的非病毒载体进行研究。对LPD-shRNA复合物的大小、ζ电位、亲和力和转染效率进行了表征，并评估了它们在脊索瘤细胞中的生物学功能。LPD-shRNA的转染效率显著高于裸shRNA。LPD将短尾相关转录因子shRNA递送至脊索瘤细胞，抑制短尾相关转录因子表达，诱导细胞凋亡，上调上皮生物标志物E-钙黏蛋白，下调间充质生物标志物Snail和Slug，并抑制细胞生长。这些数据表明，LPD可能是脊索瘤基因靶向治疗中一种有前景的shRNA非病毒载体。
