IGSF3 drives acquired temozolomide resistance in glioblastoma through ASNS-dependent asparagine biosynthesis.

Glioblastoma (GBM) is the most aggressive primary malignancy of the central nervous system, and acquired resistance to temozolomide (TMZ) is a major cause of tumor recurrence and poor clinical outcomes. However, the tumor-intrinsic drivers and metabolic adaptations underlying this process remain incompletely understood. In this study, we integrated transcriptomic data from established TMZ-sensitive and TMZ-resistant models with public bulk and single-nucleus sequencing datasets, and validated candidate genes using clinical specimens, cell-based drug-sensitivity assays, xenograft models, metabolic measurements, luciferase reporter assays, chromatin immunoprecipitation, and small-molecule binding assays. Immunoglobulin superfamily member 3 (IGSF3) was consistently upregulated in TMZ-resistant cells, resistant xenografts, and recurrent GBM samples, and was mainly enriched in malignant glioma cells. Functionally, IGSF3 overexpression promoted resistance to TMZ-related treatment, whereas IGSF3 knockdown restored drug sensitivity. Mechanistically, IGSF3 was detected in the nucleus, where it enhanced the activity of the asparagine synthetase (ASNS) promoter. In addition, IGSF3 was found to be enriched at the ASNS promoter, thereby increasing ASNS expression and asparagine production, reduced reactive oxygen species accumulation, preserved glutathione redox balance, and limited DNA damage induced by treatment. ASNS gain- and loss-of-function rescue experiments showed that ASNS was required for the pro-resistance effect of IGSF3. Finally, Tucatinib bound to IGSF3, suppressed the IGSF3-ASNS pathway, and enhanced TMZ efficacy in vitro and in vivo. These findings identify the IGSF3-ASNS axis as a tumor-intrinsic metabolic adaptation that drives acquired TMZ resistance in GBM and suggest a potential therapeutic strategy for recurrent GBM.