Targeting CPT‑1‑mediated fatty acid oxidation causes radiation sensitization in glioblastoma

Background: Glioblastoma (GBM) is the most common primary brain tumor in adults and has a median survival of less than two  years despite surgery, chemotherapy, and radiotherapy. A subpopulation of glioma stem‑like cells (GSCs) survives radiation and repopulates the tumor. Recent studies have shown that GSCs rely heavily on mitochondrial fatty‑acid oxidation (FAO) for ATP production, NAD⁺ regeneration, and redox balance. Carnitine‑palmitoyl‑transferase‑1 (CPT‑1) catalyzes the rate limiting step of FAO by shuttling long‑chain fatty acids into the mitochondrial matrix. We therefore hypothesized that pharmacologic inhibition of CPT‑1 will limit GSC metabolism and sensitize GSCs to radiation. Methods: Human GBM cell line U‑118 and a patient‑derived GSC line (GNS144) were treated for 72 h with perhexiline (5 µM), etomoxir (10 µM), or vehicle control.Cells then received a single fraction of 0, 2, 4, 6, or  8 Gy. Cell viability was measured 48 h later. Sphere formation assays were performed 7-14 days after radiation; sphere number and mean diameter were recorded. Immunoblotting was performed to evaluate for stem cell markers CD44, Nestin, and Vimentin. All experiments were performed in triplicate; statistical significance was assessed by two‑way ANOVA with Tukey post‑hoc test (p < 0.05). Results: Perhexiline + radiation and etomoxir + radiation produced a dose dependent decline in cell viability compared with radiation alone (p < 0.001). Combination treatment reduced sphere number by 55-70 % across the 4-8 Gy range (p ≤ 0.005) and lowered mean sphere diameter by 30 % (p ≤ 0.005). Immunoblotting analysis showed a 2‑fold reductions in CD44, Nestin, and Vimentin expression in the drug + radiation arms versus radiation alone (p < 0.01). Conclusions: These results suggest that inhibition of CPT‑1‑mediated FAO with perhexiline or etomoxir enhances radiation induced cell death, suppresses sphere forming capacity, and downregulates key GSC markers in both standard and patient-derived GBM models. These findings should be investigated further and suggest that CPT‑1 blockade could be integrated with current standard of care regimens to overcome stem cell driven radiation resistance.