NANP Targeting Radio-sensitizes Glioblastoma through TNFR1 Sialylation-Driven Mesenchymal Shift

GBM remains a highly devasting brain tumor mainly due to the resistance to radiotherapy (RT), the most impactful first-line treatment for GBM. Our clonal evolution study reveals no dominant RT-resistant clones to target for recurrence prevention, leading us for a genome-wide CRISPR screening for radio-sensitizing targets in GBM. The screening highlighted DNA damage response genes, particularly in the non-homologous end-joining (NHEJ) pathway, such as PRKDC and NHEJ1, as potent radio-sensitizers. Additionally, NANP (N-acylneuraminate-9-phosphatase), a critical enzyme in the sialic acid synthetic pathway, was identified as a novel RT-sensitizing target associated with patient outcomes. NANP knockdown induces increased G2/M arrest and apoptosis following radiation, with γH2AX staining and comet assays indicating greater DNA damage. NANP deficiency impairs DNA repair, favoring error-prone NHEJ over homologous recombination (HR). Mechanistically, NANP modulates the sialylation of TNFR1, thereby influencing NF-kB signaling and the mesenchymal (MES) state of glioma stem cells (GSCs), which consequently affects RT sensitivity. Intracranial orthotopic xenograft experiments validate the function of NANP in vivo. In summary, our findings identify NANP as a novel radio-sensitizing target, dependent on TNFR1 sialylation and MES shift, providing a basis for developing innovative RT sensitizers for GBM.