Metabolism meets glycosylation: PGM3 as a novel therapeutic target in human diseases

Metabolic rewiring is a hallmark of cancer, enabling tumor cells to sustain their growth, maintain redox balance, and resist therapy. Among the pathways integrating nutrient availability with cellular function, the hexosamine biosynthetic pathway (HBP) acts as a central nutrient-sensing hub, linking glucose, glutamine, and acetyl-CoA to the synthesis of UDP-N-acetylglucosamine (UDP-GlcNAc), the donor substrate for N- and O-linked glycosylation. These processes regulate protein folding, receptor stability, signaling, and genome integrity, translating metabolic status into functional proteome changes. The present PhD project involved a multidisciplinary approach, combining metabolomics, glycoproteomics, molecular biology, and pharmacological inhibition, to dissect the role HBP in both physiological and cancer contexts. Using immune and pancreatic ductal adenocarcinoma (PDAC) models, the study investigated how modulation of PGM3 activity, key HBP enzyme, shapes immune regulation, redox homeostasis, proteome stability, and DNA repair mechanisms. Our results demonstrate that PGM3 acts as a pivotal regulator balancing cellular homeostasis and malignant adaptation. Its deficiency due to isomorphic mutations leads to congenital disorders of glycosylation, characterized by severe SCID-like immunodeficiency and milder HIES-like phenotypes, accompanied by a marked loss of naïve CD4⁺ T-cells and an altered pattern of CD4⁺ T-cell differentiation. Conversely, PGM3 overexpression in KRAS-driven PDAC supports glycosylation-dependent signaling and promotes survival under metabolic stress. Pharmacological inhibition of PGM3 with the competitive inhibitor FR054 disrupted HBP flux, depleted UDP-GlcNAc levels, and impaired both N- and O-glycosylation. This, in turn, induced endoplasmic reticulum (ER) stress, activated the unfolded protein response (UPR), and weakened antioxidant response. FR054 treatment enhanced cellular glutamine dependence, which was subsequently exploited to assess synthetic lethality with erastin. The combined treatment resulted in pronounced lipid peroxidation and increased ferroptotic cancer cell death. Moreover, PGM3 inhibition sensitizes PDAC cells to gemcitabine (GEM), a genotoxic agent, by interfering with O-glycosylation processes that regulate factors involved in DNA repair mechanisms under replicative stress conditions. FR054 in combination with GEM interfered with the DNA damage response (DDR) activated by GEM, leading to sustained γH2AX accumulation, impaired ATR-Chk1 and ATM-Chk2 signaling, and persistent RPA2-Ser8 phosphorylation. These defects reduced checkpoint activation and homologous recombination (HR) repair efficiency, generating a “BRCAness-like” phenotype even in HR-proficient PDAC cells, increasing sensitivity to GEM. Overall, these findings establish PGM3 as a central metabolic checkpoint linking PTMs, redox balance, and genome stability. Its inhibition undermines tumor metabolic resilience, creates synthetic lethality with ferroptosis inducers, and restores sensitivity to DNA-damaging therapies. The development of improved FR054-like inhibitors further supports PGM3 as a promising target for combination treatments in aggressive cancers such as PDAC.