The Role of STRADA in Inhibitory Neuron Migration and Corticogenesis

PROJECT SUMMARY Malformations of cortical development (MCDs) are a major cause of pediatric epilepsy that are notoriously difficult to treat. Polyhydramnios, megalencephaly, and symptomatic epilepsy (PMSE) is a particularly severe MCD characterized by multifocal intractable epilepsy, profound neurocognitive impairment, and a high early mortality rate. PMSE is caused by a homozygous loss of STRADA, an inhibitor of the mechanistic target of rapamycin (mTOR) pathway, with a mutation carrier frequency of ~4% in the Old Order Mennonite population of North America. My earlier work demonstrated that the loss of Strada in PMSE models impairs excitatory neuron migration during cortical development, through disrupting cytoskeletal organization (actin polymerization), in an mTOR-dependent manner. Our pilot clinical trial using the mTOR inhibitor rapamycin showed initial promise as the first effective drug to control PMSE seizures. However, seizures returned to their baseline frequency over time, prompting a search for alternative mechanisms for treatment and further studies to determine STRADA’s role in corticogenesis. I have observed that both PMSE patient brain and Strada knockout (KO) mouse brain exhibit a notable lack of GABAergic inhibitory neurons (INs) in cortex and hippocampus, suggestive of an IN migratory disorder. INs are critical for the modulation of excitatory:inhibitory tone, and their dysfunction can yield hyperexcitable neural networks, promoting epilepsy. Thus, defining the mechanism of STRADA’s function in IN migration, and the mTOR dependence of this function, remains a critical gap in understanding its role in corticogenesis and pioneering therapeutic strategies. I hypothesize that STRADA drives IN migration through coordination of cytoskeletal dynamics via mTOR-dependent (cofilin and actin polymerization) and -independent (SAD kinases and tubulin polymerization) mechanisms, a process which is disrupted early in PMSE, resulting in a paucity of inhibitory neurons in cortex and hippocampus, and predisposing these patients to severe epilepsy. To test this hypothesis, in Aim 1, I will determine the effect of Strada loss on IN migration during brain development in vivo, using both ubiquitous Strada KO and focal IN progenitor cell Strada KO in mouse models and quantifying the consequent distribution of INs. In Aim 2, I will define the effect of STRADA loss in vitro on patient-derived IN migration, assess phosphorylation of nodes along STRADA, mTOR, and SAD kinase signaling pathways, and quantify polymerization of actin and tubulin to determine the degree of STRADA’s impact on cytoskeletal organization. I will determine the mTOR dependence of STRADA’s role in IN migration, using drugs and plasmid expression to target various nodes in the pathway, for mechanistic elucidation and treatment strategy. My objective is to utilize the training opportunities in this K08 project, and the mentorship of MCD experts Drs. Peter Crino and Louis Dang, to develop a comprehensive mechanistic understanding of STRADA’s function in brain development. I will use this to establish a program innovating targeted therapies for PMSE and related disorders, with a goal of early, even fetal, intervention to prevent epilepsy and neurocognitive dysfunction.