Periodic Reporting for period 3 - COSYN (Comorbidity and Synapse Biology in Clinically Overlapping Psychiatric Disorders)
Reporting period: 2019-01-01 to 2020-06-30
Using these samples as a starting point, we have set out to understand comorbidity by comparing symptom and syndrome overlap with novel neurobiological criteria and to elucidate mechanisms of comorbidity using neurobiology for the major genomic clue of synaptic dysfunction to unravel the cellular mechanisms of comorbidity.
We have generated novel neuronal cell models by using micro-lithography to generate micro-islands of support cells (glia cells) to allow single neurons to grow and form synapses onto themselves in culture (autapses). We have optimized and characterized these cultures in terms of in vitro maturation and synapse formation. Many in vitro differentiation and maturation procedures have been compared to find the optimal in vitro model and be able to generate novel neurobiological criteria and to elucidate mechanisms of comorbidity using these criteria. We have used patch clamp physiology to characterize synapse maturation and function in vitro and to standardize procedures of systematic analysis of synapses from the selected patients. Furthermore, we have introduced modern genome editing (CRISPR/Cas9) to correct genetic variants in the patient cell lines or to (re-)create the same mutations in a standard (non-patient) cell line. We have established a quality control pipe-line with exome sequencing of all patient in vitro cell lines to identify any confounding genetic rearrangements that regularly emerge during re-programming of the primary (skin) cells and gene editing. Only cell models that pass the stringent QC (approximately 50%) are used for cellular analysis. These novel cellular models provide unique opportunities to discover synaptic deficits associated with these rare genetic variants of strong effect and to build future cell models for compound screening and therapy design. In addition to these novel models, COSYN also exploits conventional cell culture models for high content screening (cellomics) and synapse recordings.
For the existing cell lines, multiple advanced neuroscience platforms have been used to evaluate an extensive set of molecular and cellular parameters, and to identify alterations in synaptic biology. The analyses on 22q11 samples, and comparison to 22q11 mouse models, are now completed and the consortium is working on new cell models for comorbidity, especially for ID and ASD.
COSYN partners are now completing the analysis of the first genome editing projects (CRISPR/Cas9), especially for STXBP1 and UNC13A. Mutations in these genes cause ‘SNAREopathies’, a complex mix of comorbid symptoms, prominently those related to ID and ASD. The extensive synaptic analyses provide new insights into the molecular functions of these genes and possible differences between human synapses and previously characterize rodent model synapses. These studies also form the basis for the analyses of engineered mutations in STXBP1, precisely mirroring the mutations detected in some of our patient samples. In this way COSYN will be able to perform side-by-side comparisons between patient-own and engineered cell models.
COSYN is a crucial next step in “decoding” the genetic findings via intensive focus on the clinical and molecular comorbidities of ID, autism, and schizophrenia.