识别循环肿瘤细胞的类肽纳米片层研究

Circulating Tumor Cells (CTC) are tumor cells existing in peripheral blood. CTC detection can help diagnostics and prognostics of tumor. Currently antibodies against the specific surface marker proteins are used as CTC capture reagents. However, use of antibodies has been hampered by their poor stability and high production cost. We propose to develop the nanosheets self-assembled by amphiphilic peptoids which display high density of affinity peptide loops to recognize CTC. EpCAM and HER2, the two breast cancer surface markers, are chosen as the targets. First, combinatorial peptide libraries against extracellular domains are designed and synthesized. Candidate affinity peptides are screened out. Then affinity of candidate peptides is characterized by surface plasmon resonance imaging. Last, the high affinity peptide sequences are inserted to the amphiphilic sheet-forming peptoid segments to form the peptoid-peptide-peptoid hybrid molecules. Peptoid nanosheets are produced by a hierarchical mechanism, first involving adsorption of amphiphilic peptoids into a monolayer at the air-water interface, followed by lateral compression of the monolayer into an ordered, irreversible peptoid bilayer (nanosheet) with the hydrophobic residues inside and the hydrophilic residues outside. During the process, peptoid strands align and pack tightly with neighboring strands to form the stable bilayer sheet, at the mean time the hydrophilic peptide inside the hybrid molecule is pushed into the shape of a loop in the aqueous phase and displayed on both sides of the nanosheet. The project will study the self-assembly condition to display the peptide loops with the best conformation and density to bind the surface marker proteins with multivalency in order to replace the antibodies for CTC detection.

对外周血中循环肿瘤细胞（CTC）的检测可以对肿瘤的诊断和预后提供帮助。目前对CTC的检测多利用抗体，然而抗体成本高、稳定性差，影响了检测方法的应用性。本项目探索利用类肽分子自组装形成纳米片层使其在表面展示高密度亲和多肽环来特异识别CTC。以乳腺癌CTC表面标志物蛋白EpCAM和HER2为模型。首先设计组合多肽文库初筛出候选多肽,然后进行高通量的亲和力分析,最后将亲和多肽序列插入两亲性类肽序列中形成类肽-多肽-类肽杂合体分子。在气-液交界面两亲性类肽先排列成单层，当受到侧向挤压时, 类肽链会整齐排列并且不可逆折叠形成稳定的双层结构，疏水部分被包裹其中，亲水部分暴露在外面。杂合体分子中的亲和多肽链在组装过程中因其亲水性被挤入水相形成闭合环状。本项目将研究使多肽环以最优的构象和密度在纳米片层表面展示的自组装条件，以多价态与CTC表面蛋白结合，代替抗体用于CTC的检测。

循环肿瘤细胞（CTC）是存在于外周血中的肿瘤细胞，对CTC的检测可以对肿瘤的诊断和预后提供帮助。目前对CTC的检测多利用特异识别CTC表面标记物蛋白的抗体，然而抗体成本高、稳定性差，影响了检测方法的应用性。本项目探索利用两亲性类肽分子自组装形成纳米片层，同时在表面展示高密度亲和多肽环来特异识别CTC。以乳腺癌CTC表面标志物蛋白EpCAM和HER2为模型。首先针对抗原表位设计并合成组合多肽文库，利用高通量多肽合成仪和原位筛选微流控芯片初筛出候选多肽。然后利用表面等离子体共振成像技术进行高通量的亲和力分析。最后将亲和多肽序列插入两亲性类肽序列中形成类肽-多肽-类肽杂合体分子。在气-液交界面两亲性类肽先排列成单层，当受到侧向挤压时,类肽会整齐排列并不可逆折叠形成纳米片层，疏水部分被包裹其中，亲水部分暴露在外面,且相邻类肽携带相反电荷形成稳定的双层结构。杂合体分子中的亲和多肽链在组装过程中因其亲水性被挤入水相形成闭合环状展示在片层表面。本项目将研究该新型纳米生物材料的自组装条件，以期使多肽环以最优的构象和密度在纳米片层表面展示，以多价态与CTC表面蛋白结合，代替抗体用于CTC的检测。在本项目支持下共发表SCI论文12篇，申请发明专利11项。