The tissue-specific targeted delivery and efficient cellular uptake of siRNAs are the main obstacles to their clinical application. Antibody-siRNA-conjugates (ARCs) can deliver siRNA by exploiting the targeting property of antibodies like antibody-drug conjugates (ADCs). However, the effective conjugation of antibodies and siRNAs and the release of siRNAs specifically at target sites have posed challenges to the development of ARCs. In this study, the successful conjugation of antibodies and siRNAs was achieved using a multifunctional peptide as a linker, composed of a cell-penetrating peptide (CPP) and a substrate peptide (SP), which is highly expressed in solid tumors. The resulting antibody-multifunctional peptide (SP-CPP)-siRNA system delivered the siRNA to target tumor cells by the specific binding of the antibody. Once the enzymes on the tumor cell surface hydrolyzed the substrate peptide linker, siRNA-CPP was released from ARCs. The released siRNA-CPP entered the targeted cells via the cellular penetrating ability of CPP, resulting in improved siRNA-mediated gene silencing efficiency, verified both in vitro and in vivo. After intravenous administration, the designed ARCs achieved approximately 66.7% EGFP (Enhanced Green Fluorescent Protein) downregulation efficiency in nude mice xenografted with the HCT116-EGFP tumor model. The proposed system provides a prospective choice for ARC production and the safe and efficient delivery of siRNAs.

siRNAs 的组织特异性靶向递送和有效细胞摄取是其临床应用的主要障碍。抗体-siRNA-偶联物(ARC) 可以通过利用抗体-药物偶联物(ADC) 等抗体的靶向特性来传递siRNA。然而，抗体和siRNAs的有效结合以及siRNAs在靶位点的特异性释放对ARCs的发展提出了挑战。在这项研究中，使用多功能肽作为接头实现了抗体和 siRNA 的成功结合，该肽由细胞穿透肽 (CPP) 和底物肽 (SP) 组成，在实体瘤中高度表达。由此产生的抗体-多功能肽（SP-CPP）-siRNA系统通过抗体的特异性结合将siRNA递送至靶向肿瘤细胞。一旦肿瘤细胞表面的酶水解底物肽接头，siRNA-CPP就会从ARC中释放出来。释放的siRNA-CPP通过CPP的细胞穿透能力进入靶细胞，提高了siRNA介导的基因沉默效率，验证了两者体外和体内。静脉给药后，设计的 ARC 在异种移植 HCT116-EGFP 肿瘤模型的裸鼠中实现了大约 66.7% 的 EGFP（增强型绿色荧光蛋白）下调效率。所提出的系统为 ARC 生产和 siRNA 的安全和有效交付提供了一个前瞻性选择。