Uncovering mechanisms of CHD2-associated epilepsy using human cortical organoids

University Of Texas San Antonio, San Antonio TX

Investigators

Abstract

PROJECT SUMMARY/ABSTRACT Mutations in CHD2 gene are associated with developmental and epileptic encephalopathy (DEE), a severe form of childhood epilepsy. CHD2 belongs to the chromodomain helicase DNA binding (CHD) family of ATP dependent chromatin remodelers known to play a critical role in neurodevelopment via chromatin organization and gene regulation; and is the only one among 9 other CHD (CHD1-CHD9) family proteins that causes a brain restricted phenotype suggesting its non-redundant role in neurodevelopmental disorders. Studies in mice have shown that knockdown of CHD2 in embryonic cortex decreases the amplification of radial glial cells (RGCs) and promotes the generation of intermediate progenitor cells (IPs). Moreover, Chd2+/- mice, the only known heterozygous mouse line to date to mimic CHD2 haploinsufficiency in human, exhibits reduced number of GABAergic interneurons, disrupts cell proliferation in the developing forebrain and displays a shift in neuronal excitability. Additionally, a study using 2D culture of medial ganglionic eminence (MGE) like progenitors and cortical interneurons derived from human embryonic stem cells (hESCs) has shown that CHD2 deficiency impairs interneuron development and alters its functions. Though these studies have been critical towards our fundamental understanding of the role of CHD2 in neurodevelopment, neither mice nor 2D neuronal cultures represent the full breadth of human brain development and leave a critical gap in knowledge about the human specific cellular and molecular mechanisms associated with CHD2 mutation. To fill this gap, we propose to use patient derived induced pluripotent stem cells (iPSCs) differentiated into human cortical organoids (hCO). hCO closely mimic the development of human forebrain and is an innovative model system to study patient specific cellular and molecular mechanisms underlying a disease-causing mutation. Based on our preliminary data using single cell RNA sequencing (scRNA-seq) in hCO, we hypothesize that CHD2 mutation leads to defects in cortical progenitor proliferation and differentiation during neurodevelopment contributing to altered cortical circuitry and epilepsy. To test our hypothesis, we will 1) determine the cellular impact of CHD2 mutation in cortical development and epilepsy using immunohistochemistry (Aim 1) and 2) delineate the molecular mechanisms of CHD2 mutation in neurodevelopment and epilepsy by performing multiome analysis (scRNA-seq + scATAC-seq) (Aim 2). The proposed study will not only increase our understanding of the childhood epilepsy and related neurodevelopmental disorders but also provide greater insight into novel in vitro cortical organoid models to study a number of pathogenic variants. To our knowledge, this is the first time that patient derived iPSCs differentiated into cortical organoids have been proposed to study CHD2 mutation in humans, and in our model system, we will be able to identify previously unknown cellular and molecular mechanisms underlying CHD2 associated epilepsy paving the way for development of better therapeutic intervention strategies.