Design and Synthesis of Near-Infrared Peptide for in Vivo Molecular Imaging of HER2.
Design and Synthesis of Near-Infrared Peptide for in Vivo Molecular Imaging of HER2.
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DOI：
10.1021/acs.bioconjchem.5b00565
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发表时间：
2016-02-17
影响因子：
4.7
通讯作者：
Wang TD
中科院分区：
文献类型：
--
作者：
Joshi BP;Zhou J;Pant A;Duan X;Zhou Q;Kuick R;Owens SR;Appelman H;Wang TD
我们报道了HER2细胞外结构域特异性肽的开发、表征和验证。该探针化学被开发用于分子成像，通过使用结构模型来选择氨基酸的最佳组合，从而最大限度地提高与HER2结构域3独特的疏水和亲水性相互作用的可能性。利用fmoc介导的固相合成技术，鉴定了序列KSPNPRF，并通过GGGSK连接剂与FITC或Cy5.5偶联，以证明该化学结构具有用不同荧光团标记的灵活性。通过改变构象刚性间隔段和移动c端上的疏水和亲水氨基酸，形成了一个混乱的序列进行控制。我们在体外结肠直肠癌细胞的敲除和竞争实验中验证了HER2的肽特异性，并测量了kd = 21 nM的结合亲和力和k = 0.14 min−1 （7.14 min）的时间常数。我们在结直肠癌的临床前模型中使用这种肽局部或静脉给药，以证明自发性腺瘤的特异性摄取，并显示近红外荧光实时体内成像的可行性。我们在人类近端结肠标本的免疫荧光研究中使用这种肽来评估无柄锯齿状和散发性腺瘤的特异性。改进的可视化可用于内镜指导组织活检和检测癌前病变，否则将错过。我们设计的针对HER2特异性的肽很有希望在早期癌症检测的分子成像方法中进行临床翻译。
We report the development, characterization, and validation of a peptide specific for the extracellular domain of HER2. This probe chemistry was developed for molecular imaging by using a structural model to select an optimal combination of amino acids that maximize the likelihood for unique hydrophobic and hydrophilic interactions with HER2 domain 3. The sequence KSPNPRF was identified and conjugated with either FITC or Cy5.5 via a GGGSK linker using Fmoc-mediated solid-phase synthesis to demonstrate fiexibility for this chemical structure to be labeled with different fluorophores. A scrambled sequence was developed for control by altering the conformationally rigid spacer and moving both hydrophobic and hydrophilic amino acids on the C-terminus. We validated peptide specificity for HER2 in knockdown and competition experiments using human colorectal cancer cells in vitro, and measured a binding affinity of kd = 21 nM and time constant of k = 0.14 min−1 (7.14 min). We used this peptide with either topical or intravenous administration in a preclinical model of colorectal cancer to demonstrate specific uptake in spontaneous adenomas and to show feasibility for real time in vivo imaging with near-infrared fluorescence. We used this peptide in immunofluorescence studies of human proximal colon specimens to evaluate specificity for sessile serrated and sporadic adenomas. Improved visualization can be used endoscopically to guide tissue biopsy and detect premalignant lesions that would otherwise be missed. Our peptide design for specificity to HER2 is promising for clinical translation in molecular imaging methods for early cancer detection.