蛋白水解靶向嵌合体（PROTAC）被认为是下一个重磅炸弹疗法。然而，由于其固有的局限性，PROTAC 的功效经常因有限的组织渗透，特别是其作用位点的细胞内化不足而受到损害。本文基于超 pH 敏感和酶敏感纳米技术，在构建了细胞周期蛋白依赖性激酶 4 的 PROTAC 和6（CDK4/6）。结直肠癌（CRC）对许多治疗方法甚至免疫检查点阻断都几乎没有反应，因此被选为本研究的肿瘤模型。结果发现，PSRN 可以在循环中维持纳米结构（40 nm），并通过增强的渗透和保留效应在肿瘤中有效积累。然后，它们响应酸性肿瘤微环境而解离成单聚体（<10nm），促进肿瘤渗透和细胞内化。最终，在组织蛋白酶 B 裂解后，CDK4/6 降解 PROTAC 在细胞内释放。重要的是，PSRN 导致靶蛋白在体外和体内的降解增强。 CDK4/6 的降解还通过上调癌细胞中程序性细胞死亡配体 1 (PD-L1) 的表达以及抑制肿瘤微环境中的调节性 T 细胞增殖来增强免疫检查点阻断的功效。通过与 α-PD-1 组合，在 CT26 肿瘤模型中很好地实现了增强的抗肿瘤效果。总体而言，我们的研究验证了 PROTAC 精确细胞内递送的重要性，并为 CRC 的靶向联合治疗引入了一种有前景的治疗策略。© 2024。作者。

PROteolysis TArgeting Chimeras (PROTACs) have been considered the next blockbuster therapies. However, due to their inherent limitations, the efficacy of PROTACs is frequently impaired by limited tissue penetration and particularly insufficient cellular internalization into their action sites. Herein, based on the ultra-pH-sensitive and enzyme-sensitive nanotechnology, a type of polymer PROTAC conjugated and pH/cathepsin B sequential responsive nanoparticles (PSRNs) are deliberately designed, following the construction of the PROTAC for Cyclin-dependent kinase 4 and 6 (CDK4/6). Colorectal cancer (CRC) which hardly responds to many treatments even immune checkpoint blockades was selected as the tumor model in this study. As a result, PSRNs were found to maintain nanostructure (40 nm) in circulation and efficiently accumulated in tumors via enhanced permeation and retention effect. Then, they were dissociated into unimers (<10 nm) in response to an acidic tumor microenvironment, facilitating tumor penetration and cellular internalization. Eventually, the CDK4/6 degrading PROTACs were released intracellularly following the cleavage of cathepsin B. Importantly, PSRNs led to the enhanced degradation of target protein in vitro and in vivo. The degradation of CDK4/6 also augmented the efficacy of immune checkpoint blockades, through the upregulation of programmed cell death-ligand 1 (PD-L1) expression in cancer cells and the suppression of regulatory T cells cell proliferation in tumor microenvironment. By combination with α-PD-1, an enhanced anti-tumor outcome is well achieved in CT26 tumor model. Overall, our study verifies the significance of precise intracellular delivery of PROTACs and introduces a promising therapeutic strategy for the targeted combination treatment of CRC.© 2024. The Author(s).