Targeting heat shock proteins on cancer cells: Selection, characterization, and cell-penetrating properties of a peptidic GRP78 ligand
Targeting heat shock proteins on cancer cells: Selection, characterization, and cell-penetrating properties of a peptidic GRP78 ligand
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DOI：
10.1021/bi060264j
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发表时间：
2006-08-08
期刊：
影响因子：
2.9
通讯作者：
Janda, Kim D.
中科院分区：
文献类型：
--
作者：
Kim, Youngsoo;Lillo, Antonietta M.;Janda, Kim D.
肽配体可用于特异性细胞靶向和将有效载荷递送到靶细胞中。在这里，我们描述了使用全细胞淘选对人黑色素瘤细胞系Me 6652/4的环肽噬菌体展示库的池的筛选。该策略导致选择环状13-mer Pep 42，CTVALPGGYVRVC，其显示相对于参考细胞系Me 6652/56优先内化到黑素瘤细胞系Me 6652/4中。这种易位是一种受体介导的过程，不需要静电相互作用，也不涉及转移到溶酶体区室。Pep 42的细胞受体被鉴定为葡萄糖调节蛋白78（GRP 78）的表面膜形式，GRP 78是热休克蛋白家族的成员，也是恶性癌细胞的标志物。通过激光共聚焦显微镜监测Pep 42-量子点缀合物的细胞摄取和细胞内运输，并观察内质网内的共定位。Pep 42的摄取可被针对所鉴定的受体的单克隆抗体阻断。此外，Pep 42显示出特异性靶向表达GRP 78的癌细胞。通过流式细胞术评价Pep 42-Taxol缀合物的体外细胞毒性，其中缀合物显示诱导细胞凋亡，并且在Me 6652/4细胞中更有效地促进程序性细胞死亡。总之，所呈现的数据表明，环肽Pep 42可能是设计用于选择性杀死恶性癌细胞的药物缀合物的构建中的有力工具。
Peptidic ligands can be used for specific cell targeting and the delivery of payloads into the target cell. Here we describe the screening of a pool of cyclic peptide phage display libraries using whole-cell panning against human melanoma cell line Me6652/4. This strategy resulted in the selection of the cyclic 13-mer Pep42, CTVALPGGYVRVC, which showed preferential internalization into melanoma cell line Me6652/4 versus the reference cell line Me6652/56. This translocation is a receptor-mediated process that does not require electrostatic interactions nor does it involve transfer to the lysosomal compartment. The cellular receptor for Pep42 was identified as the surface membrane form of glucose-regulated protein 78 (GRP78), a member of the heat shock protein family and a marker on malignant cancer cells. The cellular uptake and intracellular trafficking of Pep42-Quantum Dot conjugates was monitored by confocal laser microscopy, and colocalization within the endoplasmic reticulum was observed. The uptake of Pep42 could be blocked by a monoclonal antibody against the identified receptor. Furthermore, Pep42 was shown to target specifically GRP78-expressing cancer cells. The in vitro cytotoxicity of a Pep42-Taxol conjugate was evaluated by flow cytometry wherein the conjugate was shown to induce apoptosis and was more effective in promoting programmed cell death in Me6652/4 cells. In summary, the data presented suggest that cyclic peptide Pep42 might be a powerful tool in the construction of drug conjugates designed to selectively kill malignant cancer cells.