Modeling spastic paraplegia 4 with corticospinal motor neuron-enriched cortical organoids reveals genotype-phenotype and HDAC6-targetable pathology

Spastic paraplegia 4 (SPG4), the most common form of hereditary spastic paraplegia, causes progressive gait deficiency due to corticospinal tract degeneration. SPG4 results from mutations in the SPAST gene, which encodes spastin, a microtubule-severing AAA-ATPase. To dissect genotype-phenotype relationships, we generated isogenic human induced pluripotent stem cell lines carrying either an SPAST missense (SPAST^WT/C448Y) or truncation (SPAST^WT/S245X) mutation and differentiated them into corticospinal motor neuron-enriched cortical organoids. These models revealed mutation-specific patterns of aberrant neuronal activity, microtubule hypoacetylation, and axonal degeneration. We identified mutant M1-spastin-induced hyperactivation of histone deacetylase 6 (HDAC6), a major tubulin deacetylase, as the key pathogenic culprit. Pharmacological inhibition of HDAC6 with tubastatin A restored microtubule acetylation and rescued axonal degeneration in organoids, with corresponding improvements in corticospinal tract integrity and gait defects in SPG4 transgenic mice. Our study uncovers HDAC6 hyperactivation as a targetable mechanism for SPG4 and verifies human organoids as a platform for therapeutic discovery.