MECHANOSENSITIVE ION CHANNEL DYSFUNCTION IN NEURODEVELOPMENT: <EM >TMEM63B</EM> VARIANTS DRIVE SEVERE EPILEPTIC ENCEPHALOPATHY AND NEURODEGENERATION

TMEM63B is a mechanosensitive cation channel that plays a crucial role in cell volume regulation and mammalian water homeostasis. While its paralogs, TMEM63A and TMEM63C, have been implicated in neurodevelopmental disorders, the role of TMEM63B in human disease remains largely unexplored. We recently identified a cohort of 17 patients with severe, drug-resistant developmental and epileptic encephalopathy (DEE) associated with progressive neurodegenerative brain changes, all carrying heterozygous variants in the TMEM63B gene.
To investigate the underlying pathophysiological mechanisms, we expressed selected TMEM63B variants in Neuro2A cells and generated induced pluripotent stem cells from two patients, which were differentiated into neural progenitor cells (NPCs) and mature cortical neurons. Mechanosensory function and neuronal network activity were assessed using hypotonic stress and controlled mechanical stimulation with dual-laser optical tweezers, combined with calcium imaging, patch-clamp electrophysiology, and multi-electrode array recordings.
Heterologous expression of TMEM63B variants resulted in constitutive inward leak currents under isotonic conditions and impaired calcium signaling in response to hypotonic stress. Patient-derived NPCs displayed significantly reduced mechanically evoked calcium responses, confirming defective mechanosensation. In mature neurons, TMEM63B variants led to reduced intrinsic excitability but increased spontaneous excitatory synaptic activity, consistent with compensatory synaptic scaling. Notably, seizure-like events were observed exclusively in patient-derived neuronal networks.
Together, these findings demonstrate that TMEM63B variants disrupt mechanosensitive signaling and alter neuronal excitability, providing a cellular basis for the DEE phenotype. Importantly, these patient-specific neural models represent a valuable platform for identifying novel therapeutic targets and support precision medicine approaches for drug-resistant DEE.