Targeting G6PD with cinobufagin induces metabolic crisis and apoptosis in DLBCL

The pentose phosphate pathway plays a critical role in cancer metabolism by fueling nucleotide synthesis and maintaining redox balance. Glucose-6-phosphate dehydrogenase (G6PD), the rate-limiting enzyme of the PPP, represents a promising yet underexploited therapeutic target. Its role and the feasibility of its pharmacologic inhibition in diffuse large B-cell lymphoma (DLBCL) remain unclear. We analyzed G6PD expression and activity in DLBCL patients. A natural product library was screened to identify G6PD inhibitors. The lead compound, cinobufagin, was characterized using molecular docking, enzymatic assays, transcriptomics, metabolomics, and in vitro and in vivo functional studies. G6PD was overexpressed and hyperactive in DLBCL, correlating with poor patient survival. Genetic knockdown of G6PD impaired DLBCL growth in vitro and in vivo. High-throughput screening identified cinobufagin as a direct, high-affinity G6PD binder and inhibitor. Cinobufagin treatment suppressed G6PD activity, leading to a dual metabolic crisis: depletion of nucleotide precursors and NADPH, accompanied by ROS accumulation. Multi-omics profiling confirmed the concerted suppression of nucleotide synthesis and redox metabolism, and activation of stress-response pathways. Consequently, cinobufagin induced DNA damage, cell cycle arrest, and apoptosis in DLBCL cells. In a xenograft mouse model using human DLBCL cells, cinobufagin significantly inhibited tumor growth. This study establishes G6PD as a therapeutically relevant metabolic target in DLBCL and identifies cinobufagin as a direct natural G6PD inhibitor. By elucidating the mechanism through which G6PD inhibition triggers a synthetically lethal metabolic crisis, our work provides a rational strategy for targeting DLBCL metabolism.