一种抗cea粘附体，在人胰腺癌组织切片中显示高清晰度染色，并在体内选择性靶向肿瘤

IF 5 2区医学 Q2 Medicine

Translational Oncology Pub Date : 2025-08-28 DOI:10.1016/j.tranon.2025.102512

Johan Nilvebrant , Carlos Fernández Moro , Eleftherios Papalanis , Masih Ostad Novin , Haozhong Ding , Ruonan Li , Maryam Oroujeni , Arun Selvam , Béla Bozóky , Torbjörn Gräslund , Timea Szekerczes , Tatiana Sandalova , Hugh Salter , Adnane Achour , Vladimir Tolmachev , Mikael Björnstedt , Per-Åke Nygren

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引用次数: 0

摘要

我们报道了针对高度糖基化的人癌胚抗原相关粘附分子5 （CEACAM5， CEA）的小的非免疫球蛋白粘附体亲和蛋白的开发和表征，以及它们在人胰腺癌样本的免疫组织化学（IHC）分析和体内肿瘤成像中的应用。利用组合蛋白工程技术，从一个由58个氨基酸组成的三螺旋束蛋白结构域构建的大型噬菌体展示文库中，共鉴定出19个独特的cea抗体。分子模型表明，所有富集的克隆共享一个结合表面，有几个聚集的色氨酸残基与CEA N1结构域以苯丙氨酸残基为中心的疏水斑块相互作用。其中一个被命名为C9的变体在生物传感器分析中表现出最高的亲和力，并被重组为在大肠杆菌中表达的15 kDa的同型二聚体。对生物素化形式C9-C9-Bio在匹配的冷冻和福尔马林固定石蜡包埋（FFPE）人类胰腺癌样本（n = 7）上的免疫组化性能进行了评估。与临床级单克隆抗体II-7和CEA31以及多克隆试剂相比，C9-C9-Bio显示出高灵敏度的CEA检测，背景染色最少。包括类内相关性和Bland-Altman评估在内的统计分析显示，C9-C9-Bio与FFPE组织样本中的两种单克隆抗体之间具有良好的一致性。此外，99mTc[Tc]标记的C9-C9构建物在体外显示出cea依赖于人类癌细胞系的结合，并且在作为体内成像示踪剂研究时，选择性地结合到小鼠表达cea的BxPC3异种移植物上，允许在4小时后可视化肿瘤。总之，这些发现强调了易于产生的结合cea的C9粘附体在各种临床应用中的潜在用途，包括免疫结构和医学成像，以及作为指导各种治疗方式的靶向片段来治疗表达cea的肿瘤。

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An anti-CEA affibody showing high-definition staining in human pancreatic cancer tissue sections and selective tumor targeting in vivo

We report development and characterization of small non-immunoglobulin affibody affinity proteins directed to the highly glycosylated human carcinoembryonic antigen-related adhesion molecule 5 (CEACAM5, CEA), and their use in immunohistochemical (IHC) analyses of human pancreatic cancer samples and for in vivo tumor imaging. A total of nineteen unique anti-CEA affibodies were identified from large phage display libraries constructed using combinatorial protein engineering of a small 58 amino acid three-helix bundle protein domain. Molecular modeling suggested that all enriched clones share a binding surface with several clustered tryptophan residues interacting with a hydrophobic patch in the N1 domain of CEA centered around a phenylalanine residue. One variant, designated as C9, exhibited the highest affinity in biosensor analyses and was reformatted into a 15 kDa homodimer expressed in Escherichia coli. The biotinylated form, C9-C9-Bio, was evaluated for its IHC performance on matched frozen and formalin-fixed, paraffin-embedded (FFPE) sections of human pancreatic cancer samples (n = 7). Compared to clinical-grade monoclonal antibodies II-7 and CEA31, as well as a polyclonal reagent, C9-C9-Bio demonstrated highly sensitive CEA detection with minimal background staining. Statistical analyses including intraclass correlation and Bland-Altman assessments revealed excellent agreement between C9-C9-Bio and the two monoclonal antibodies in FFPE tissue samples. Further, a ^99mTc[Tc]-labeled C9-C9 construct showed CEA-dependent binding to human cancer cell lines in vitro, and selectively bound to CEA-expressing BxPC3 xenografts in mice when investigated as a tracer for in vivo imaging, allowing for a visualization of tumors after four hours. In summary, these findings highlight the potential use of the easily produced CEA-binding C9 affibody for various clinical applications, including IHC and medical imaging, and as a targeting moiety for directing various therapeutic modalities to CEA-expressing tumors.

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来源期刊

Translational Oncology ONCOLOGY-

CiteScore

8.40

自引率

2.00%

发文量

314

审稿时长

54 days

期刊介绍： Translational Oncology publishes the results of novel research investigations which bridge the laboratory and clinical settings including risk assessment, cellular and molecular characterization, prevention, detection, diagnosis and treatment of human cancers with the overall goal of improving the clinical care of oncology patients. Translational Oncology will publish laboratory studies of novel therapeutic interventions as well as clinical trials which evaluate new treatment paradigms for cancer. Peer reviewed manuscript types include Original Reports, Reviews and Editorials.