Autism human neural precursor cells exhibit common defects in neurite outgrowth, cell migration, and mTOR signaling
Creators: name: Prem, Smrithi
Publisher: name: No Publisher Supplied
Year: 2018

description: Autism spectrum disorders (ASD) are a group of developmental disorders characterized by deficits in social interaction and communication and the presence of repetitive and restrictive behaviors. Despite high prevalence (1:68), and large social and economic impacts, uncovering the mechanisms that contribute to ASD and finding therapeutics for treatment of the disorder has been thwarted by our inability to directly study human neurons, the limitations of animal models, and disorder heterogeneity. Recent studies have found that ASD risk genes converge onto the cerebral cortex of the developing mid-fetal brain (8-24 weeks old). During this time, neural precursor cells (NPCs) in the developing brain are undergoing proliferation, migration, and differentiation to form neurons and the normal cytoarchitecture of the brain. Indeed, post-mortem and imaging studies in humans with ASD have uncovered structural changes that are suggestive of alterations in these basic developmental processes. However, genetic, imaging, and post-mortem analyses cannot give mechanistic insight into the alterations found in the ASD brain. For the past few decades, studies to uncover the mechanisms underpinning ASD  have primarily been conducted in rodent models. However, rodent models, due to differences in physiology and genetics, are unable to truly recapitulate human neurodevelopment. Moreover, rodent models cannot be utilized to study the vast majority of cases of ASD which are idiopathic or polygenic. Therefore, to really understand ASD, we need to study developmental processes like proliferation, migration, and differentiation in human neural cells derived from individuals with idiopathic ASD. Yet, until the discovery of induced pluripotent stem cell (iPSC) technology such studies were impossible.  iPSCs are stem cells reprogrammed from mature somatic tissues (like white blood cells) that have the capability of forming almost any cell in the body including neurons. As iPSCs retain the genetic signature of the individuals from whom they are derived, we can for the first time, study the development and function of neural cells of people with genetically complex neuropsychiatric disorders. The three iPSC studies of idiopathic ASD published thus far have uncovered common defects in synapse formation, dendritic spines, and neuronal activity in ASD neurons. Yet, despite studies indicating the importance of early neurodevelopment in ASD pathogenesis, most iPSC studies have focused on post-mitotic differentiated neurons and have largely neglected study of early developmental processes in NPCs. Therefore, the goals of my studies were to assess neurite outgrowth, migration, and signaling pathways in neural precursor cells derived from 6 individuals with ASD. Our cohort consists of three individuals with idiopathic ASD (I-ASD) and their unaffected brothers (Sib) as controls, 3 individuals with 16p11.2 deletion (16pDel) and ASD and unaffected controls from the NIMH. Fascinatingly, despite the heterogeneity of ASD, all 6 patients in our cohort had reductions in neurite outgrowth and cell migration when compared to unaffected individuals. On the other hand, we were able to define distinct autism NPC “subgroups” by using developmentally relevant extracellular factors (EFs) such as serotonin (5-HT), PACAP, and nerve growth factor (NGF). Specifically, treatment with EFs stimulated neurite outgrowth and cell migration in both unaffected patients and 16pdel NPCs, whereas they failed to elicit neurite or migration responses in I-ASD. Further studies revealed that NPCs derived from all the ASD NPCs, both idiopathic and 16pDel, showed dysregulations in the mTOR pathway. Two individuals (I-ASD-1 and I-ASD-3) had lower mTOR pathway activity characterized by reductions in P-AKT and P-S6 while the other four individuals (I-ASD-2 &amp; the 3 16pDel patients) showed higher mTOR pathway activity as characterized by higher levels of P-S6. Thus, molecular subtyping of patients was accomplished by characterizing the levels of mTOR pathway components. As signaling pathways have been shown to regulate neurodevelopmental processes, I wanted to manipulate and “normalize” mTOR pathway activity in these patients to see if neurodevelopmental phenotypes could be rescued. Thus, I selected 1 patient from each group (under vs overactive mTOR) and applied agonist and antagonist drugs. In I-ASD-1, where mTOR pathways were underactive, the use of AKT agonist sc-79 rescued the neurite outgrowth, migration, and EF response defects seen in these NPCs. Conversely, application of AKT inhibitor MK-2206 to Sib-1 NPCs, led to reductions in neurite outgrowth and migration and abolished EF responses in this unaffected individual! In I-ASD-2, where mTOR pathway activity was higher, application of MK-2206 successfully increased neurite outgrowth and migration to the level of Sib NPCs. These studies show that the mTOR pathway is a critically important regulator of neurodevelopmental processes and that targeting this altered pathway could rescue developmental defects seen in our ASD patient NPCs. In conclusion, by studying human neural precursor cells derived from patients with ASD, I discovered that alterations in early developmental processes that are essential to building the brain are commonly altered. Moreover, by utilizing EFs and studying mTOR signaling, I was able to determine different subtypes of ASD. Lastly, common aberrations in the mTOR pathway were also observed in our ASD patients. By targeting these mTOR abnormalities, I successfully reversed the phenotypes seen in our NPCs. Thus, by using patient derived NPCs, I have demonstrated the utility of this model in the study, categorization, and potentially even treatment of autism spectrum disorders. Future studies can help elucidate how alterations in NPCs correlate with patient phenotypes and whether drugs that can rescue phenotypes in vitro could ultimately be made into targeted therapeutics for patients.