硫修饰脂质体包裹VEGF反义寡核苷酸对肺癌血管生成及转移的抑制作用

Inhibitory effects of phosphorothioate-modified antisense VEGF oligodeoxynucleotides formulated in cationic liposome on angiogenesis and metastasis of lung cancer

目的:探讨硫修饰脂质体包裹的VEGF反义寡核苷酸对肺癌血管生成和转移的抑制作用。方法:将硫修饰脂质体包裹的VEGF反义寡核苷酸(ASODN)、正义寡核苷酸(SODN)、错义寡核苷酸(MODN)加入培养的Lewis肺癌细胞中,采用免疫组织化学方法检测肺癌细胞中VEGF蛋白表达,观察经ODN处理的条件培养基对牛主动脉内皮细胞增殖的影响。另外,复制小鼠Lewis肺癌模型40只,随机分为对照组、VEGF ASODN组、VEGF SODN组及VEGF MSODN组,每组10只,接种肿瘤细胞24小时内,给予脂质体、VEGF ASODN、VEGF SODN、VEGF MSODN皮下注射,每周2次,连续4周。检测皮下肿瘤的变化和肺转移率,并用免疫组织化学方法检测肿瘤组织微血管密度(MVD),超声检测肿瘤组织PS、RI变化。结果:ASODN能够下调肺癌细胞中VEGF蛋白的表达,经ASODN处理的条件培养基能显著抑制牛主动脉内皮细胞的增殖。ASODN组小鼠肿瘤生长及肺转移受到显著抑制,MVD、PS、RI与其它各组相比有明显差异(P<0.01)。结论:硫修饰脂质体包裹的VEGF反义寡核苷酸能够显著抑制肺癌血管生成,进而抑制肿瘤生长及转移。

Objective：The study aims to investigate whether phosphorothioate-modified antisense vascular endothelial growth factor oligodeoxynucleotides（ VEGF ASODN）micro-encapsulated in cationic liposome could inhibit angiogenesis and metastasis of lung cancer. Methods： The phosphorothioate-modify VEGF ASODN, VEGF SODN （ sense oligodeoxynucleotides ） and VEGF MODN （ mismatch oligonucleotides）micro-encapsulated in cationic liposome were added into in vitro culture of Lewis lung carcinoma（ LLC ） cells, respectively. Expression of VEGF protein was detected by immunohistochemistry staining. And the inhibitory effects on bovine aortic endothelial cell proliferation by conditioned media obtained from LLC cells treated with ODN were observed. 40 mice of Lewis lung cancer models were established and subsequently were randomized into 4 groups ： control group, ASODN group, SODN group, and MSODN group. Liposome, ASODN, SODN or MODN were given by twice a week for 4 weeks, respectively. The weights of subcutaneous tumors were measured. The rates of lung metastasis were detected, while the microvessel density（MVD） in tumor mass was detected by imuunohistochemistry staining. Peak systolic flow velocity （PS） and resistance index （RI） were measured with a sonographic scanner. Results：ASODN downregulated the expression of VEGF in LLC cells at the level of protein in vitro. The conditioned media obtained from LLC cells treated with ASODN significantly inhibited the proliferation of bovine aortic endothelial cells. ASODN significantly suppressed the growth of subcutaneous tumors and lung metastasis. MVD, PS and RI of VEGF ASODN group were remarkably different from those of other 3 groups （ P 〈 0. 01 ）. Conclusion： The study demonstrated that the phosphorothioate-modified ASODN could significantly suppress the growth and metastasis of lung cancer by inhibiting angiogenesis.