Novel Pt@PCN-Cu-induced cuproptosis amplifies αPD-L1 immunotherapy in pancreatic ductal adenocarcinoma through mitochondrial HK2-mediated PD-L1 upregulation

Background: Copper accumulation triggers mitochondrial-driven cell death, known as Cuproptosis, offering a promising mechanism for targeted Cancer therapy. Recent studies have highlighted the critical role of intratumoral copper levels in regulating the expression of programmed cell death ligand-1 (PD-L1), suggesting that copper-induced Cuproptosis not only enhances Cancer cell death but may also amplify the effects of anti-PD-L1 antibodies (αPD-L1). However, in tumors where monotherapy with αPD-L1 shows limited efficacy, particularly in pancreatic ductal adenocarcinoma (PDAC), the role of copper-induced Cuproptosis in enhancing αPD-L1 treatment efficacy and its underlying mechanisms remain unclear. Meanwhile, inadequate tumor drug accumulation and glycolysis significantly restrict the efficacy of Cuproptosis. To address these challenges, we have synthesized a novel nanozyme, Pt@PCN-Cu, designed to stabilize intracellular copper accumulation and effectively induce Cuproptosis. Additionally, we aim to determine whether this strong induction of Cuproptosis can synergize with αPD-L1 to enhance Cancer therapy, ultimately paving the way for novel strategies to improve PDAC treatment.

Methods: Pt@PCN-Cu was synthesized via a one-pot method, and its therapeutic potential was assessed in combination with αPD-L1 for the treatment of PDAC. Initially, the material's properties were characterized, and its efficient cellular uptake was confirmed. Anti-tumor efficacy was evaluated by inducing Cuproptosis in PDAC cell lines and xenograft models. RNA Sequencing (RNA-seq) was utilized to identify key regulators involved in the modulation of PD-L1 expression by Cuproptosis. Lastly, the therapeutic efficacy of Pt@PCN-Cu combined with αPD-L1 was evaluated in vivo, focusing on tumor growth inhibition and immune modulation within the tumor microenvironment (TME).

Results: Pt@PCN-Cu demonstrates excellent physicochemical properties and remarkable cascade catalytic activity, providing a solid foundation for further in vitro and in vivo studies. In vitro, Pt@PCN-Cu efficiently transports copper and induces Cuproptosis primarily through mitochondrial dysfunction. Mechanistic studies show that Pt@PCN-Cu triggers the dissociation of Hexokinase 2 (HK2) from mitochondria, leading to a reduction in HK2 activity. This decline in HK2 activity impairs glycolysis, a metabolic pathway essential for tumor energy metabolism, which in turn results in elevated PD-L1 levels. In vivo, Pt@PCN-Cu demonstrates excellent safety and accumulates at the tumor site in a subcutaneous PDAC mouse model, inducing Cuproptosis. Moreover, the combination of Pt@PCN-Cu with αPD-L1 further enhanced its therapeutic efficacy and effectively reprogrammed the immunosuppressive TME.

Conclusion: This study presents strong evidence confirming the safety and therapeutic potential of Pt@PCN-Cu in PDAC treatment. Importantly, Pt@PCN-Cu not only induces Cuproptosis but also significantly enhances antitumor efficacy in combination with αPD-L1 by regulating PD-L1 expression through HK2 modulation. These findings underscore a more effective and innovative approach for treating PDAC.

Cuproptosis; HK2; Immunotherapy; Nanozyme; PD-L1; Tumor microenvironment.