LARGE-SCALE PHENOTYPIC SCREENING IDENTIFIES MODULATORS OF NEURONAL REGENERATION FOR SPINAL CORD REPAIR

Spinal cord injury (SCI) severely limits natural regeneration by disrupting axonal connectivity and triggering a cascade of secondary damage. Despite the identification of numerous molecular targets involved in neural repair, few pharmacological strategies have integrated multi-pathway modulation with clinical applicability. To address this challenge, we performed large-scale phenotypic screening to identify clinically available compounds capable of enhancing the resilience and regenerative capacity of neurons under injury-related conditions. Following exposure of rat primary cortical neurons to oxidative stress, we screened a comprehensive library comprising approved compounds and late-stage clinical candidates. High-content imaging and quantitative morphometric analysis assessed neuronal survival, axonal outgrowth, and cytoskeletal integrity, identifying multiple candidate molecules that consistently promoted neuronal survival and stimulated axonal outgrowth across a range of concentrations. Functional annotation analysis revealed convergence onto pathways associated with inflammation regulation, oxidative stress control, and neuromodulatory signaling, suggesting a coordinated mechanism supporting structural recovery. Selected lead compounds were validated through multi-dose analysis and pathway-centric profiling, confirming reproducible improvements in neuronal morphology and network stability in vitro. To assess translational applicability, representative lead compounds were investigated in spinal cord contusion model of rats, achieving sustained release at the lesion site via local hydrogel-based delivery. Histological and cytological analyses revealed favorable cellular responses. Furthermore, functional assessments demonstrated significant motor recovery, validating the therapeutic potential of the identified candidates. Collectively, this research demonstrates that phenotype-based drug repositioning can systematically identify multi-target modulators of neuroregeneration, providing an expandable framework integrating in vitro discovery with targeted in vivo delivery strategies for spinal cord recovery.