Preparation of a cathepsin D sensitive near-infrared fluorescence probe for imaging
Preparation of a cathepsin D sensitive near-infrared fluorescence probe for imaging
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DOI：
10.1021/bc990052h
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发表时间：
1999-09-01
影响因子：
4.7
通讯作者：
Weissleder, R
中科院分区：
文献类型：
--
作者：
Tung, CH;Bredow, S;Weissleder, R
多种蛋白酶在发病过程中过表达或活化，并代表治疗药物的重要靶点。我们以前已经表明，在近红外荧光（NIRF）光谱中敏感的光学成像探针可用于酶活性的体内成像。在目前的研究中，我表明，这些探针可以被设计为对特定酶具有特异性，例如，已知在许多肿瘤中过表达的组织蛋白酶D。NIR花青荧光染料作为光学报告分子，连接到对组织蛋白酶D具有特异性的11个氨基酸肽序列的氨基末端。随后将肽连接到合成的接枝共聚物上，用于有效的肿瘤递送。多个荧光染料的紧密空间接近导致结合态的荧光淬灭。在体外测试期间，在切割后观察到350倍的信号放大。使用用人组织蛋白酶D稳定转染的啮齿动物肿瘤细胞系的细胞培养实验证实了细胞内的酶特异性活化。该序列而不是乱序对照序列在体外显示酶特异性。我们的结论是，可激活的NIRF光学探针，可以合成潜在的探测生物体中的特定酶。
A variety of proteases are overexpressed or activated during pathogenesis and represent important targets for therapeutic drugs. We have previously shown that optical imaging probes sensitive in the near-infrared fluorescence (NIRF) spectrum can be used for in vivo imaging of enzyme activity. In the current study, me show that these probes can be designed with specificity for specific enzymes, for example, cathepsin D which is known to be overexpressed in many tumors. A NIR cyanine fluorochrome served as the optical reporter and was attached to the amino terminal of an 11 amino acid peptide sequence with specificity for cathepsin D. The peptides were subsequently attached to a synthetic graft copolymer for efficient tumoral delivery. The close spatial proximity of the multiple fluorochromes resulted in quenching of fluorescence in the bound state. A 350-fold signal amplification was observed post cleavage during in vitro testing. Cell culture experiments using a rodent tumor cell line stably transfected with human cathepsin D confirmed enzyme specific activation within cells. This sequence but not a scrambled control sequence showed enzyme specificity in vitro. We conclude that activatable NIRF optical probes can be synthesized to potentially probe for specific enzymes in living organisms.