协同增效的PEG化二肽连接子实现稳定高载药ADC的构建

Synergistic PEGylated Dipeptide Linker Enables Stable High-Drug-Load ADCs

关键词:  抗体偶联药物  聚乙二醇化连接子  高抗体偶联比  药代动力学

Key words:  Antibody-drug conjugate  PEGylated linker  High drug-to-antibody ratio  Pharmacokinetics

摘要:

目的 旨在通过理性设计新型连接子，构建具有高抗体偶联比且稳定性优异的抗体偶联药物(Antibody-Drug Conjugates, ADC)。方法 设计并合成了一种新型ADC药物DS001。通过合理构建聚乙二醇(Polyethylene glycol，PEG)修饰的缬氨酸-赖氨酸连接子[VK-(mPEG24)-PAB]，采用定点偶联技术将八个单甲基奥瑞他汀E (Monomethyl auristatin E，MMAE)分子精准连接至靶向Trop2的抗体hRs7上，从而获得药物抗体比(Drug-to-Antibody Ratio, DAR)为8的均一偶联物，得到DS001—[hRs7-VK-(mPEG24)-PAB-MMAE]。结果 体外实验表明，DS001能保持与Trop2抗原的高亲和力结合，并表现出强效且特异的细胞毒性，其半数抑制浓度(median inhibition concentration, IC50)值达纳摩尔级。同时，DS001在血浆中显示出显著延长的稳定性。经优化的连接子有效缓解了由疏水性驱动的聚集效应，成功实现了DAR 8的稳定构建。结论 本研究开发的PEG化VK连接子策略，能够有效克服高载药量ADC中常见的聚集与稳定性难题。DS001在保持高效抗肿瘤活性的同时，具备优异的药代动力学特性。

Abstract:

OBJECTIVE This study aimed to rationally design a novel linker to construct an Antibody-Drug Conjugates (ADC) with high DAR and excellent stability. METHODS A new ADC drug, DS001, was designed and synthesized. By rationally constructing a polyethylene glycol (PEG)-modified valine-lysine linker [VK-(mPEG24)-PAB-MMAE], eight Monomethyl auristatin E (MMAE) molecules were precisely conjugated to the anti-Trop2 antibody hRs7 via site-specific conjugation technology, resulting in a homogeneous conjugate with a Drug-to-Antibody Ratio (DAR) of 8. RESULTS In vitro experiments demonstrated that DS001 maintained high binding affinity to the Trop2 antigen and exhibited potent and specific cytotoxicity, with an median inhibition concentration (IC??) in the nanomolar range. Additionally, DS001 showed significantly extended stability in plasma. The optimized linker effectively mitigated hydrophobicity-driven aggregation, enabling the stable construction of an ADC with DAR 8. CONCLUSION The PEGylated VK linker strategy developed in this study effectively addresses the common challenges of aggregation and instability in high-drug-load ADCs. DS001 retains potent antitumor activity while demonstrating favorable pharmacokinetic properties, offering a promising design approach for the development of next-generation high-DAR ADCs.