EXPLOITING GABAA RECEPTORS IN A GENE THERAPY APPROACH FOR FOCAL EPILEPSY

Gene therapy is increasingly being explored as an alternative therapeutic strategy for individuals affected by drug-resistant focal epilepsy. Traditional chemogenetic approaches rely on the expression of engineered receptors that do not respond to endogenous ligands but are selectively activated by synthetic compounds, raising concerns regarding receptor immunogenicity and the safety profile of the exogenous ligands. In this work, we propose a novel chemogenetic strategy based on the enhancement of endogenous γ-aminobutyric acid type A receptor (GABAAR) expression, aiming to strengthen GABA-mediated inhibitory signaling and increase sensitivity to clinically established GABAergic antiseizure drugs (ASDs) within epileptogenic cortical regions. To achieve this, we developed a third-generation lentiviral vector encoding two GABAAR subunits, which was shown to effectively promote subunit expression and receptor assembly selectively in excitatory neurons. Functional and pharmacological characterization of the expressed receptors was performed in collaboration with the laboratory of Prof. Eleonora Palma, by using microtransplantation of membranes from vector-injected mice into Xenopus oocytes. This approach revealed an increased sensitivity to phenobarbital, attributable to overexpression of native GABAAR subunit composition. In vivo evaluation of this therapeutic strategy is currently ongoing in collaboration with Dr. Rob Wykes (University College London) using the tetanus toxin model of focal neocortical epilepsy. Preliminary results show a flattening of seizure progression over time in treated animals. These data require further investigation with larger experimental cohorts. Overall, this study introduces a novel chemogenetic approach that enhances the pharmacological response of endogenous inhibitory receptors without the need for synthetic receptor–ligand systems, potentially offering a safer and more translationally relevant therapeutic strategy for focal epilepsy.