Blood–Brain Barrier-Permeable, Reactive Oxygen Species-Producing, and Mitochondria-Targeting Nanosystem Amplifies Glioblastoma Therapy

Gemcitabine (GTB), a clinically approved nucleoside analogue for cancer treatment, faces therapeutic limitations due to rapid enzymatic deactivation by cytidine deaminase (CDA) in tumor microenvironments. Over 90% of systemically administered GTB undergoes catalytic conversion to inactive 2′-deoxy-2′,2′-difluorouracil metabolites through CDA-mediated deamination. To address this pharmacological challenge, we developed a multifunctional codelivery nanosystem through strategic engineering of reactive oxygen species (ROS)-generating, mitochondria-targeting CPUL1-TPP (CT) nanoaggregates. These self-assembling CT/GTB complexes were further optimized with DSPE-MPEG2k (DP) and Angiopep-2-conjugated DSPE-MPEG2k (Ang-DP) to create blood–brain barrier (BBB)-penetrating Ang-DP@CT/GTB nanoparticles, enhancing both physiological stability and low-density lipoprotein receptor-related protein 1 (LRP1)-mediated glioma targeting. Comparative analyses revealed that Ang-DP@CT/GTB nanoparticles significantly enhanced GTB’s antiglioblastoma efficacy compared to free drug administration in both in vitro and in vivo models. Mechanistic investigations demonstrated that the nanosystem upregulates heme oxygenase-1 (HO-1), subsequently downregulating CDA expression to mitigate GTB metabolism. This coordinated molecular modulation prolongs GTB’s therapeutic activity while leveraging the ROS-generating capacity of CT components for synergistic tumor suppression. The BBB-permeable codelivery platform exemplifies a rational design paradigm for multifunctional carrier-free pure nanodrugs (PNDs), demonstrating how clinical drug reformulation can overcome inherent pharmacokinetic limitations. This nanotechnology-driven approach provides critical insights for optimizing chemotherapeutic performance through metabolic pathway regulation and targeted delivery engineering.