Cellular dynamics of EGF receptor-targeted synthetic viruses
Cellular dynamics of EGF receptor-targeted synthetic viruses
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DOI：
10.1038/sj.mt.6300176
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发表时间：
2007-07-01
影响因子：
12.4
通讯作者：
Braeuchle, Christoph
中科院分区：
文献类型：
--
作者：
de Bruin, Karla;Ruthardt, Nadia;Braeuchle, Christoph
表皮生长因子受体(EGFR)在高比例的人类肿瘤中过度表达。在肿瘤基因治疗中，EGFR是非病毒载体组织特异性靶向的一个有吸引力的治疗靶点。在这项研究中，我们分析和比较了EGFR靶向和非靶向复合体对EGFR过度表达的Huh7细胞内化的影响。用流式细胞仪和单粒子示踪法评价药物的摄取动力学和内化动力学。我们的结果清楚地表明，与非靶向颗粒相比，EGFR靶向可以导致更快、更有效的内化。5分钟后，50%的EGFR靶向复合体被内化，而非靶向复合体即使在20分钟后也只达到约20%的内在化。此外，单粒子跟踪揭示了内化过程的三相动力学，这通常被观察到与靶标无关的复合体。I期以缓慢的、肌动蛋白细胞骨架介导的颗粒漂移运动为特征，包括内化过程。在II期，颗粒速度增加，胞浆内扩散正常且受限。第三阶段的特点是沿微管快速活跃的转运。通过EGFR靶向受体介导的内吞作用的多个复合体，导致I期缩短和强烈加速内化。
The epidermal growth factor receptor (EGFR) is overexpressed on a high percentage of human carcinomas. EGFR is an attractive therapeutic target for tissue-specific targeting by non-viral vectors in cancer gene therapy. In this study we analyzed and compared the effects of EGFR-targeted and untargeted polyplexes in respect to internalization into EGFR overexpressing HuH7 cells. Uptake kinetics and internalization dynamics were evaluated by flow cytometry and single-particle tracking. Our results clearly show that EGFR targeting leads to faster and more efficient internalization compared with untargeted particles. After 5 minutes 50% of the EGFR-targeted polyplexes were internalized, whereas untargeted polyplexes reached only approximately 20% internalization even after 20 minutes. In addition, single-particle tracking revealed a three-phase dynamics of the internalization process, and this was generally observed for polyplexes independent of targeting. Phase I was characterized by slow, actin cytoskeleton-mediated movement of the particles with drift, and included the internalization process. During phase II particles displayed increased velocities with normal and confined diffusion in the cytoplasm. Phase III was characterized by fast active transport along microtubules. Targeting of polyplexes for receptor-mediated endocytosis by the EGFR resulted in shortening of phase I and strongly accelerated internalization.