Divisato, G.; Tozza, S.; Cascone, E.; Polishchuk, E.; Zizolfi, M. C.; Giannino, E.; Marsella, F.; Di Girolamo, D.; Menale, C.; Perone, L.; Gianfico, P.; Cuda, G.; Bucci, C.; Maiuri, P.; Polishchuk, R.; Manganelli, F.; Parisi, S.

Biallelic mutations in the sorbitol dehydrogenase (SORD) gene have been identified as one of the most common causes of autosomal-recessive Charcot Marie Tooth disease type 2 (CMT2) and distal hereditary neuropathy, collectively referred to as SORD deficiency. These mutations result in loss of sorbitol dehydrogenase activity, a key enzyme in the polyol pathway that metabolizes glucose, leading to marked accumulation of sorbitol in patient derived fibroblasts. However, the mechanisms by which SORD dysfunction drives axonal degeneration remain poorly understood, and robust in vitro models of human SORD deficient motor neurons (MNs) are still lacking. To address this gap, we established a human in vitro model of SORD deficiency by generating induced pluripotent stem cells (iPSCs) from fibroblasts affected individual carrying biallelic SORD mutations (SORDc.757delG/c.316_425+165del), and unaffected heterozygous carriers (SORDc.757delG/wt and SORDwt/c.316_425+165del). These iPSCs were subsequently differentiated into motor neuron progenitors (MNPs) and MNs. Comprehensive analysis of SORD-deficient human cells, including fibroblasts, MNPs, and MNs, revealed pronounced structural and functional abnormalities in the mitochondrial compartment, characterized by mitochondrial fragmentation and increased proton leak. Importantly, fibroblasts derived from two additional unrelated patients carrying the SORD mutation (SORDc.757delG/ c.757delG) further confirmed that SORD deficiency is associated with a mitochondrial phenotype. At the molecular level, SORD deficiency led to upregulation of aldose reductase (AR), another key enzyme of the polyol pathway, resulting in disruption of cellular redox homeostasis and increased oxidative stress. Consistent with these alterations, MNs derived from CMT2/SORD patients exhibited clear neurodegenerative features, including severe defects in neurite branching and synaptic architecture, ultimately impairing neuronal connectivity. Notably, pharmacological inhibition of AR effectively rescued both mitochondrial dysfunction and neuronal structural defects, supporting the targeting of AR as a promising therapeutic strategy for polyol pathway associated neuropathies as CMT2/SORD and diabetic neuropathy.