DDDR-54. Mitochondrial Protease LonP1 Drives Resistance to Proteasome Inhibitors in Glioblastoma and Offers a Novel Therapeutic Target

Abstract Proteasome inhibitors like bortezomib and marizomib although demonstrated encouraging results in preclinical studies, neither drug achieved success in clinical trials for the treatment of glioblastoma. This discrepancy between laboratory findings and clinical outcomes highlights the substantial challenge posed by drug resistance in glioblastoma, which is driven by factors such as intratumoral heterogeneity, the blood-brain barrier, and tumor microenvironment adaptations. Growing evidence implicates the mitochondrial protease LonP1 as a key mediator of proteasome inhibitors resistance. Resistance to proteasome inhibitors in glioblastoma (GBM) remains a major therapeutic challenge. This study primarily investigates the mechanistic role of LonP1 in compensating for proteasome inhibition and evaluates dual targeting of LonP1 and the proteasome as a novel therapeutic strategy in malignant glioblastoma treatment. Using in vitro and in vivo GBM models, we demonstrate that LonP1 overexpression (e.g., lentiviral mediated overexpression of LonP1) compensates for proteasome inhibition by degrading misfolded proteins and maintaining mitochondrial proteostasis, while also promoting resistance via mitochondrial metabolic reprogramming. Genetic or pharmacological inhibition of LonP1 (e.g., lentiviral mediated inducible knockdown of LonP1 or BT317, a dual LonP1/proteasome inhibitor) synergizes with proteasome inhibitors, restoring sensitivity through ROS accumulation, impaired oxidative phosphorylation, and suppression of glioma stem cell (GSC) survival. Multi-omics analysis reveals that LonP1-driven mitochondrial dysfunction correlates with LonP1 overexpression, marked by increasing mtDNA copy number and TCA cycle induction. Clinical data further associate high LonP1 expression with poor prognosis in proteasome inhibitors-resistant GBM. Conversely, genetic or pharmacological inhibition of LonP1 restores proteasome inhibitors sensitivity, synergistically inducing apoptosis via ROS accumulation and impaired mitochondrial metabolism. These findings establish LonP1 as a critical central driver of proteasome inhibitors resistance and provide a rationale for co-targeting LonP1 and the proteasome to overcome therapeutic resistance in cancer and advocate for dual targeting strategies to improve therapeutic outcomes.