Systematic evolution of a DNA aptamer binding to rat brain tumor microvessels - Selective targeting of endothelial regulatory protein pigpen
Systematic evolution of a DNA aptamer binding to rat brain tumor microvessels - Selective targeting of endothelial regulatory protein pigpen
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DOI：
10.1074/jbc.m100347200
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发表时间：
2001-05-11
影响因子：
4.8
通讯作者：
Schluesener, H
中科院分区：
文献类型：
--
作者：
Blank, M;Weinschenk, T;Schluesener, H
肿瘤微血管在结构和代谢功能上与正常血管不同，并且新血管生成与内皮蛋白表达的定量和定性变化相关。这些分子可以作为区分肿瘤血管系统与正常大脑血管系统的分子地址。我们已经应用了系统进化配体指数富集（SELEX）对转化的内皮细胞作为一个复杂的目标，选择单链DNA配体（适体）的功能作为组织学标记，以检测大鼠实验性胶质瘤，致命的脑肿瘤，是高度血管化的微血管。通过基于荧光的方法（流式细胞术和荧光显微术）分析SELEX选择程序以及随后的去卷积-SELEX。在分析的25种适体中，选择了一种适体，其选择性结合大鼠脑胶质母细胞瘤的微血管，但不结合正常大鼠脑的脉管系统，包括瘤周区域。通过配体介导的磁性DNA亲和纯化，从内皮细胞中分离适体III.1的分子靶蛋白。该蛋白质通过质谱鉴定为小鼠猪圈的大鼠同源物，小鼠猪圈是一种不广为人知的内皮蛋白，其表达与体外从静止表型到血管生成表型的转变平行。由于新血管生成（新血管的形成）是肿瘤发展的关键特征，因此所提出的适体可用作探针来分析胶质母细胞瘤的病理性血管生成，结果表明，pigpen在实验性大鼠脑胶质母细胞瘤的肿瘤微血管中高表达，可能在抑制脑肿瘤的血供，从而促进肿瘤生长中起重要作用。
Tumor microvessels differ in structure and metabolic function from normal vasculature, and neoangiogenesis is associated with quantitative and qualitative changes in expression of endothelial proteins. Such molecules could serve as molecular addresses differentiating the tumor vasculature from those of the normal brain. We have applied Systematic Evolution of Ligands by EXponential enrichment (SELEX) against transformed endothelial cells as a complex target to select single-stranded DNA-ligands (aptamers) that function as histological markers to detect microvessels of rat experimental glioma, a fatal brain tumor that is highly vascularized. Both the SELEX selection procedure as well as subsequent deconvolution-SELEX were analyzed by fluorescence based methods (flow cytometry and fluorescence microscopy), Of 25 aptamers analyzed, one aptamer was selected that selectively bound microvessels of rat brain glioblastoma but not the vasculature of the normal rat brain including peritumoral areas. The molecular target protein of aptamer III.1 was isolated from endothelial cells by ligand-mediated magnetic DNA affinity purification. This protein was identified by mass spectrometry as rat homologue of mouse pigpen, a not widely known endothelial protein the expression of which parallels the transition from quiescent to angiogenic phenotypes in vitro, Because neoangiogenesis, the formation of new blood vessels, is a key feature of tumor development, the presented aptamer can be used as a probe to analyze pathological angiogenesis of glioblastoma, The presented data show that pigpen is highly expressed in tumor microvessels of experimental rat brain glioblastoma and may play an important role in warranting blood supply, thus growth of brain tumors.