Li Enze, Wen Liewei, Yin Caiyun, Wang Nan, Yang Suleixin, Feng Wenzheng, Chen Meiwan
State Key Laboratory of Mechanism and Quality Research of Chinese Medicine, Institute of Chinese Medical Sciences, University of Macau, Macau SAR, China.
Guangdong Provincial Key Laboratory of Tumor Interventional Diagnosis and Treatment, Zhuhai People's Hospital (The Affiliated Hospital of Beijing Institute of Technology), Beijing Institute of Technology, Zhuhai 519088, China.
J Control Release. 2025 Sep 24;388(Pt 1):114262. doi: 10.1016/j.jconrel.2025.114262.
Cuproptosis as a copper-dependent cell death modality driven by pathological aggregation of lipoylated proteins and proteotoxic stress resulting from destabilization of iron-sulfur (FeS) cluster proteins, represents a promising therapeutic strategy for cancer. However, the therapeutic efficacy of cuproptosis can be compromised by the intrinsic compensatory mechanisms within cancers, particularly low intracellular copper ion concentration and protective autophagy, which facilitates cellular adaptation and survival under stress. To overcome this limitation, a self-amplifying cuproptosis nanoregulator (SHK-Cu/PF/pATG5@HA, abbreviated as SC/TpA@HA) for CD44-targeted delivery is developed that integrate shikonin‑copper (SHK-Cu) coordination complexes with CRISPR/Cas9 plasmids targeting ATG5, condensed by fluorinated polyethyleneimine (PF)-condensed to enhance cancer therapy. Briefly, Cu is released from the dissociated SHK-Cu complex upon intracellular GSH activation induces dihydrolipoamide S-acetyltransferase (DLAT) oligomerization and reduces FeS cluster proteins, triggering tricarboxylic acid (TCA) cycle collapse and irreversible mitochondrial damage. Concurrently, CRISPR/Cas9-mediated ATG5 knockout prevents autophagosome formation, creating an autophagic flux trap that accumulates copper-damaged mitochondria. Notably, such mitochondrial dysfunction as induced by copper overload combined with impaired cellular damage clearance from autophagy blockade elevates cuproptosis. In addition, the immunogenic cell death through cuproptosis in cancer cells, as validated by the exposure of calreticulin and the extracellular release of HMGB1, triggers a potent anti-tumor immune response. This response is enhanced through autophagy inhibition, as ATG5 deletion blocks the downstream signaling of copper-activated Unc-51-like autophagy activating kinase 1/2 (ULK1/2), ultimately amplifying cytotoxic T lymphocyte infiltration. Therefore, this dual intervention through copper overload and autophagy blockade potentiates both cuproptosis and anti-tumor immune effect, representing an innovative strategy of cuproptosis treatment.
铜死亡作为一种由脂酰化蛋白的病理性聚集和铁硫（FeS）簇蛋白不稳定导致的蛋白毒性应激所驱动的铜依赖性细胞死亡方式，是一种很有前景的癌症治疗策略。然而，癌症内部的固有补偿机制，特别是细胞内铜离子浓度低和保护性自噬，可能会损害铜死亡的治疗效果，而保护性自噬有助于细胞在应激状态下适应和存活。为了克服这一限制，开发了一种用于靶向CD44递送的自扩增铜死亡纳米调节剂（SHK-Cu/PF/pATG5@HA，简称为SC/TpA@HA），它将紫草素-铜（SHK-Cu）配位复合物与靶向ATG5的CRISPR/Cas9质粒整合在一起，由氟化聚乙烯亚胺（PF）凝聚以增强癌症治疗效果。简而言之，细胞内谷胱甘肽激活后，SHK-Cu复合物解离释放出铜，诱导二氢硫辛酰胺S-乙酰转移酶（DLAT）寡聚化，并使FeS簇蛋白减少，触发三羧酸（TCA）循环崩溃和不可逆的线粒体损伤。同时，CRISPR/Cas9介导的ATG5基因敲除可防止自噬体形成，形成一个自噬通量陷阱，使铜损伤的线粒体积累。值得注意的是，铜过载诱导的线粒体功能障碍与自噬阻断导致的细胞损伤清除受损相结合，会增强铜死亡。此外，通过钙网蛋白的暴露和HMGB1的细胞外释放验证，癌细胞中由铜死亡引起的免疫原性细胞死亡会触发强烈的抗肿瘤免疫反应。这种反应通过自噬抑制得到增强，因为ATG5缺失会阻断铜激活的Unc-51样自噬激活激酶1/2（ULK1/2）的下游信号传导，最终放大细胞毒性T淋巴细胞浸润。因此，这种通过铜过载和自噬阻断的双重干预增强了铜死亡和抗肿瘤免疫效应，代表了一种创新的铜死亡治疗策略。
