Glutaredoxin 1 promotes sorafenib resistance in renal cell carcinoma through ferroptosis suppression: Integrative bioinformatics analysis and experimental validation

Background: Sorafenib resistance has limited the therapeutic efficacy in advanced renal cell carcinoma (RCC). Glutaredoxin 1 (GRX1, also known as GLRX), a redox-regulating oxidoreductase belonging to the thioredoxin superfamily, may mediate this resistance through Ferroptosis suppression, but the underlying mechanisms remain incompletely understood.

Methods: GRX1 expression and sorafenib sensitivity were analyzed using The Cancer Genome Atlas-Kidney Renal Clear Cell Carcinoma (TCGA-KIRC, n = 601), Gene Expression Omnibus (GEO, n = 14), and Genomics of Drug Sensitivity in Cancer (GDSC, 752 cell lines) datasets. Functional experiments including lentiviral-mediated GRX1 knockdown (shRNA) and overexpression were performed in human RCC cell lines 786-O and Caki-1 and their sorafenib-resistant derivatives established through chronic drug exposure. The expression of ferroptosis-related proteins, including Glutathione Peroxidase 4 (GPX4), solute carrier family 7 member 11 (SLC7A11), and acyl-CoA synthetase long-chain family member 4 (ACSL4), was assessed by Western blotting. Ferroptosis was further evaluated by measuring lipid Reactive Oxygen Species (ROS), malondialdehyde (MDA), reduced glutathione (GSH), and intracellular Fe2 + levels, and confirmed by pharmacological rescue with ferroptosis-specific inhibitors. BALB/c nude mouse xenograft models (n = 6 per group, four treatment arms per cell line) validated findings in vivo. Three machine learning algorithms, linear regression (LR), random forest (RF, 500 trees), and elastic net (EN, α=0.5), were constructed for exploratory prediction of sorafenib sensitivity.

Results: GRX1 was significantly upregulated in RCC and correlated with advanced stage and grade. A treatment-dependent paradox was observed: high GRX1 was associated with better overall survival in TCGA-KIRC but promoted sorafenib resistance in functional experiments. GRX1 knockdown enhanced sorafenib sensitivity and increased ferroptotic responses, with decreased GPX4 and SLC7A11, increased ACSL4, elevated lipid ROS, MDA, and Fe2 + , and depleted GSH, whereas overexpression conferred resistance. Pharmacological rescue by ferrostatin-1 and deferoxamine, but not by Apoptosis or Necroptosis inhibitors, identified Ferroptosis as the dominant cell death mechanism. In sorafenib-resistant cells, GRX1 was elevated 2.5-3.1-fold, and its knockdown restored sensitivity in vitro and reduced final xenograft tumor weights by approximately 55% in sh-GRX1+sorafenib groups versus sh-NC+sorafenib controls. Random Forest showed modest but reproducible exploratory predictive performance for sorafenib sensitivity (R² = 0.199).

Conclusion: The treatment-dependent role of GRX1, which is protective in untreated tumors but can promote sorafenib resistance in treated cancers, establishes it as a critical ferroptosis-suppressing mediator and a potential therapeutic target for overcoming sorafenib resistance in RCC.