Rodríguez Mercè, Coma Sílvia, Noé Véronique, Ciudad Carlos J
Department of Biochemistry, School of Pharmacy, University of Barcelona, Spain.
Antisense Nucleic Acid Drug Dev. 2002 Oct;12(5):311-25. doi: 10.1089/108729002761381294.
The development and the effect of immunoliposomes directed against human breast cancer cells overexpressing p185/HER2 are described. These immunoliposomes carry an antisense oligonucleotide directed toward the translational start site of dihydrofalate reductase (DHFR) RNA, which causes high cytotoxicity. To prepare the immunoliposomes, we followed two methodologies based on the high affinity between streptavidin and biotin and the use of biotinylated antibodies. In the first approach, the streptavidin molecule is covalently attached to the phospholipid DOPE, which is mixed with the cationic liposome DOTAP complexed with the antisense oligonucleotide. The second approach, which is much easier to perform, involves the binding of streptavidin to antibody and oligonucleotide, both biotinylated, and the latter complexed with DOTAP. The formation of the intermediary complexes of this immunoliposome was studied sequentially by gel electrophoresis. The uptake of the oligonucleotide carried by the immunoliposome was monitored by flow cytometry and confocal microscopy. As a model, we used SKBR3 cells that overexpress p185. The full immunoliposomes were more toxic than the antisense oligonucleotide in the absence of the antibody, thus increasing the sensitivity of the treatment.
本文描述了针对过表达p185/HER2的人乳腺癌细胞的免疫脂质体的开发及其效果。这些免疫脂质体携带一种针对二氢叶酸还原酶（DHFR）RNA翻译起始位点的反义寡核苷酸，具有高细胞毒性。为制备免疫脂质体，我们基于链霉亲和素与生物素之间的高亲和力以及使用生物素化抗体，采用了两种方法。在第一种方法中，链霉亲和素分子共价连接到磷脂DOPE上，DOPE与与反义寡核苷酸复合的阳离子脂质体DOTAP混合。第二种方法操作起来要容易得多，它涉及链霉亲和素与生物素化的抗体和寡核苷酸结合，且寡核苷酸与DOTAP复合。通过凝胶电泳依次研究了这种免疫脂质体中间复合物的形成。通过流式细胞术和共聚焦显微镜监测免疫脂质体携带的寡核苷酸的摄取情况。作为模型，我们使用了过表达p185的SKBR3细胞。在没有抗体的情况下，完整的免疫脂质体比反义寡核苷酸毒性更大，从而提高了治疗的敏感性。
