DMG-50. Dual Targeting of Mitochondrial and Lipid Metabolism in Diffuse Midline Glioma: Trifluoperazine and Atorvastatin Combination Therapy

Diffuse Midline Gliomas (DMGs) are highly aggressive pediatric brain tumors with limited treatment options and poor prognosis. Although targeting oxidative phosphorylation (OXPHOS) has shown therapeutic promise, DMGs exhibit significant metabolic plasticity, adapting and reprogramming energy production under therapeutic pressure. We observed that trifluoperazine (TFP), an antipsychotic drug, disrupts mitochondrial respiration in DMG cells and induces oxidative stress. In response, these cells engage metabolic reprogramming, upregulating cholesterol biosynthesis and accumulating lipid droplets to maintain membrane integrity, energy balance, and survival. We hypothesized that co-administration of TFP with atorvastatin, an HMG-CoA reductase inhibitor, would block this compensatory mechanism, driving DMG cells into metabolic collapse and cell death. In vitro, TFP treatment reduced the viability of DMG cell lines over 72 hours (n = 5; IC50: 4.1–7.1 μM). Super-resolution microscopy demonstrated mitochondrial fragmentation, while flow cytometry showed loss of mitochondrial membrane potential and oncreased reactive oxygen species. WB analysis confirmed decreased expression of mitochondrial proteins (NDUFA12A, COXIV) and upregulation of stress marker ATF4. Seahorse assays, metabolomics, and lipid analyses indicated disruption of OXPHOS and TCA cycle, reduced free fatty acid levels, along with lipid droplet accumulation. Transcriptomic profiling revealed upregulation of cholesterol biosynthesis (SC5D, DHCR7) and lipid storage (PPARG, PLIN1) genes, alongside downregulation of fatty acid oxidation (CPT1A). Collectively, these findings highlight cellular adaptation to mitochondrial stress, as DMG cells shift from classical oxidative metabolism toward heightened lipid and cholesterol pathways. Although atorvastatin alone had limited efficacy in vitro, its combination with TFP displayed strong synergy, further reducing cell viability. In vivo, TFP monotherapy significantly extended survival in DMG PDX models (68 vs. 56 days, p = 0.02). Ongoing studies suggest that combining TFP with atorvastatin will additionally enhance survival by targeting critical mitochondrial and lipid metabolic dependencies. This dual-targeting strategy - simultaneously disrupting OXPHOS and cholesterol biosynthesis - leverages FDA-approved agents, offering a rapidly translatable therapeutic approach that may overcome this metabolic reprogramming and improve outcomes for children with DMGs.