Enhancing KCC2 function reduces interictal activity and prevents seizures in mesial temporal lobe epilepsy

The neuronal K/Cl cotransporter KCC2 regulates the transmembrane chloride gradient, which controls the efficacy of GABAergic signaling. In mesial temporal lobe epilepsy (mTLE) and other neurological disorders, reduced KCC2 expression or function can result in depolarizing GABA signaling, which is thought to contribute to pathological activity and seizures. Therefore, restoring chloride homeostasis represents a promising therapeutic strategy. We investigated the mechanisms and antiseizure effects of two small molecules, prochlorperazine (PCPZ) and CLP-257, that have been identified as potential KCC2 enhancers. We found that both compounds enhance KCC2 function and clustering in cortical neurons while reducing its membrane diffusion, without altering canonical regulatory phosphorylation. CLP-257 also selectively increased extrasynaptic, but not synaptic, GABAA receptor-mediated currents. Using in vitro recordings from resected brain tissue of patients with drug-resistant mTLE and in vivo recordings from a mouse model, we show that PCPZ and CLP-257 (or its prodrug CLP-290) effectively suppressed spontaneous epileptiform activity in both models. These findings reveal that PCPZ and CLP-257 act as genuine KCC2 enhancers and provide experimental evidence of the therapeutic potential of such compounds for treating drug-resistant mTLE. Significance statementA major challenge in treating epilepsy is the high percentage of patients with drug-resistant forms, like mesial temporal lobe epilepsy (mTLE). This study investigates a therapeutic strategy by targeting the neuronal KCC2 transporter, which is often dysfunctional in epilepsy. Our findings identify two compounds, prochlorperazine and CLP-257, that enhance KCC2 function by promoting its clustering on the cell membrane, a previously uncharacterized mechanism. Importantly, these compounds effectively reduce spontaneous epileptiform activity in human brain tissue from mTLE patients and significantly reduce seizure frequency in a mouse model. This work provides a critical proof-of-concept for activating KCC2 as a viable therapeutic approach for drug-resistant epilepsy.