Metformin Restores Mitochondrial Function and Neurogenesis in <i>POLG</i> Patient‐Derived Brain Organoids

Mitochondrial dysfunction and impaired neurogenesis are central to mitochondrial DNA polymerase (POLG)-related disorders, yet therapeutic options remain limited. Here, patient-derived induced pluripotent stem cell (iPSC)-based cortical organoids are used to model POLG-associated neurodegeneration and assess the therapeutic potential of metformin. Single-cell RNA-seq reveals distinct vulnerabilities in dopaminergic, glutamatergic, and GABAergic neuronal subtypes, with dopaminergic neurons exhibiting the most severe loss and mitochondrial transcriptomic deficits. Metformin treatment (250 µm, 2 months) significantly restores neuronal identity, subtype-specific gene expression, and mitochondrial function. Functional assays demonstrate improved mitochondrial membrane potential (TMRE), increased mitochondrial mass (MTG, MTDR), and reduced oxidative stress (MitoSOX, BAX/cleaved caspase 3). Notably, mitochondrial DNA (mtDNA) copy number and the expression of mitochondrial replisome proteins (POLG, POLG2) are upregulated, indicating enhanced mitochondrial genome maintenance. Calcium measurement confirms improved neuronal excitability. Untargeted metabolomics further reveals metformin-induced metabolic reprogramming, including enrichment of the tricarboxylic acid (TCA) cycle, amino acid metabolism, and redox-related pathways. Together, these findings demonstrate that metformin enhances mitochondrial integrity and neural function across multiple neuronal subtypes and offer mechanistic insights into its potential as a treatment for POLG-related disorders.