蛋白酶切靶点融合物（PROteolysis Targeting Chimeras，PROTAC）在降解包括难以药物靶向的蛋白在内的多个癌基因蛋白方面展现出巨大的治疗潜力。PROTAC是一种双功能分子，其中一部分与靶蛋白结合，而另一部分则招募蛋白降解机制。随着PROTAC领域的不断进展，我们通过开发基于抗体的蛋白酶切靶点融合物（ABTAC）的组合方法，探索了一种可能有效降解促进癌症增殖和进展的重要癌基因蛋白——表皮生长因子受体（EGFR）。本研究的目标是合成和表征一种降解EGFR的ABTAC，用于治疗非小细胞肺癌（NSCLC）。使用赖氨酸共轭和铜自由的齐式炔烃-酸剂环加成（CuAAC）点击化学反应对西妥昔单抗（Cetuximab）和泼尼松胺（一种E3连接酶招募配体）进行了连接。通过反相液相色谱和质谱的分析表征，确认将五个E3连接酶抑制剂分子/抗体进行了共轭。与西妥昔单抗相比，ABTAC在HCC827（对EGFR敏感）和H1650（对EGFR耐药）细胞中的IC50下降了近10-30倍。多细胞3D球体试验强烈提示，与对照组和单独使用抗体相比，ABTAC诱导了显著的细胞凋亡，并抑制了细胞增殖。圆二色谱和表面等离子体共振（SPR）证实，在共轭后，抗体的结构和受体结合效力发生了轻微变化。版权 © 2023. Elsevier B.V.出版。

PROteolysis Targeting Chimeras (PROTACs) showed tremendous therapeutic potential in degrading several oncoproteins including undruggable proteins. PROTACs are bifunctional molecules where one-part binds to target protein while the other end recruits protein degradation machinery. With the unveiling advancements in the field of PROTACs, we explored a combinatorial approach by developing antibody-based PROTAC (ABTAC) which may effectively degrade one of the key oncoprotein driving proliferation and progression of cancer - Epidermal growth factor receptor (EGFR). The objective of current research was to synthesize and characterize an EGFR degrading ABTAC for the treatment of non-small cell lung cancer (NSCLC). Cetuximab and pomalidomide (E3 ligase recruiting ligand) were conjugated using lysine conjugation and copper free azide-alkyne cycloaddition (CuAAC) click chemistry. Analytical characterization using reverse-phase liquid chromatography and mass spectrometry suggested conjugation of five E3-ligase inhibitor molecules/antibody. Nearly 10-30 folds reduction in IC50 was observed with ABTAC in HCC827 (EGFR sensitive) and H1650 (EGFR resistant) cells compared to cetuximab. Multicellular 3D spheroid assay strongly suggested that ABTAC induced significant apoptosis and also inhibited cell proliferation compared to control and antibody alone. Circular dichroism and surface plasmon resonance (SPR) confirmed minor alterations in the structure and receptor binding efficacy of the antibody post-conjugation.Copyright © 2023. Published by Elsevier B.V.