Modelling human brain circuitry in patients with KCNQ2-Developmental Epileptic Encephalopathy using induced pluripotent stem cells
Stewart, Rachel
Stewart, Rachel
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Publication Date
2023-10-16
Type
Thesis
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Abstract
Mutations (pathogenic variants) in KCNQ2, which encode the voltage-dependent K+ channel Kv7.2 (responsible for neuronal M-current) can cause developmental epileptic encephalopathy (DEE), a disorder presenting with severe early-onset seizures and impaired neurodevelopment. Treatment of this disorder is very difficult because the effect of KCNQ2 mutations on neurodevelopment is still largely unknown. In this study, we generated patient-specific iPSCs from 4 patients harbouring different KCNQ2-DEE loss-of-function, mutations: R213Q, F261L, A265T and A294V, and 4 sibling controls which were subsequently differentiated into excitatory cortical neurons to model the disease in vitro. We performed multi-electrode array (MEA) recordings of cortical neurons with the KCNQ2-DEE pathogenic variants F261L, A265T and A294V and found that for each mutation, neurons were hyperexcitable and displayed a burst-suppression firing pattern that is reminiscent of the interictal electroencephalography pattern seen in patients (burst suppression). We chose to focus on the pathogenic variant A265T for further functional analysis and found that A265T neurons displayed functional enhancement of Na+ channels and Ca2+ activated K+ channels while also exhibited altered spontaneous Ca2+ transients via Ca2+ imaging experiments. We showed that the Na+ channel blocker carbamazepine, and the K+ channel opener retigabine, were capable of reducing the phenotype in patient neurons, albeit not to the level of controls. Our data shows for the first time that patient-specific iPSC-derived cortical neurons harbouring the pathogenic variants F261L, A265T and A294V are hyperexcitable and display an irregular firing pattern compared to sibling controls. Moreover, we have generated a patient-specific disease model which is invaluable for drug testing and discovery with great potential for precision medicine.
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Publisher
NUI Galway