9R穿膜肽可增强P53抗肿瘤效果

Anti-tumor efficacy of P53 with 9R cell-penetrating peptides

摘要:

为解决P53蛋白难以进入细胞内部发挥治疗作用的瓶颈难题。将p53基因融合插入带有9个精氨酸作为穿膜肽的表达载体中表达融合蛋白CPPs-P53，并与没有穿膜肽的P53蛋白进行比较，利用Western blotting 方法检测蛋白的表达情况，MTT及Annexin V/PI双染法检测细胞生长抑制率及细胞凋亡率。Western blotting 检测表明已成功在原核表达系统中表达融合蛋白CPPs-P53和P53蛋白，且蛋白纯度均已达到90％以上；MTT检测表明，P53蛋白对肿瘤细胞的生长虽有一定的抑制作用，但融合蛋白CPPs-P53与之相比，对肿瘤细胞生长的抑制效果显著增强，细胞生长抑制率有明显的提升，并且细胞生长抑制率呈现剂量依赖性；Annexin V/PI双染检测细胞凋亡情况也表明P53虽可以在一定程度上诱导肿瘤细胞的凋亡，但与P53蛋白相比较，融合蛋白CPPs-P53诱导的凋亡细胞明显增加，凋亡率是P53蛋白的2~3倍。由此说明在抑制肿瘤细胞的生长和诱导细胞凋亡方面，CPPs-P53比没有穿膜肽的P53蛋白的效果更显著

Abstract:

To enhance the penetration of P53 into tumor cells by fusion it with the cell penetrating peptide 9R. The fusion gene of 9R-p53 was cloned into the expression vector. The fusion protein, CPPs-P53, was expressed and purified. We detected the rate of cell growth inhibition and apoptosis by MTT and Annexin-V-FITC/PI double stained method respectively for measuring its effect on tumor cells. CPPs-P53 and P53 were successfully expressed and purified, the purity of both proteins reached up to 90%. MTT assay showed that the cell growth inhibition by CPPs-P53 was more efficient than P53, and the rate of cell growth inhibition is dose-dependent. The apoptosis experiment showed that P53 could induce apoptosis of tumor cells. Compared with the P53, CPPs-P53 had a more significant effect in inducing cell apoptosis (**P<0.01). The CPPs-P53 shows more significant effects than P53 in inhibiting cell growth and inducing apoptosis on tumor cells