Multiplexed profiling of GPCR activities by combining split TEV assays and EXT-based barcoded readouts

G protein-coupled receptors (GPCRs) are the largest class of cell surface receptors and are implicated in the physiological regulation of many biological processes. The high diversity of GPCRs and their physiological functions make them primary targets for therapeutic drugs. For the generation of novel compounds, however, selectivity towards a given target is a critical issue in drug development as structural similarities between members of GPCR subfamilies exist. Therefore, the activities of multiple GPCRs that are both closely and distantly related to assess compound selectivity need to be tested simultaneously. Here, we present a cell-based multiplexed GPCR activity assay, termed GPCRprofiler, which uses a beta-arrestin recruitment strategy and combines split TEV protein-protein interaction and EXT-based barcode technologies. This approach enables simultaneous measurements of receptor activities of multiple GPCR-ligand combinations by applying massively parallelized reporter assays. In proof-of-principle experiments covering 19 different GPCRs, both the specificity of endogenous agonists and the polypharmacological effects of two known antipsychotics on GPCR activities were demonstrated. Technically, normalization of barcode reporters across individual assays allows quantitative pharmacological assays in a parallelized manner. In summary, the GPCRprofiler technique constitutes a flexible and scalable approach, which enables simultaneous profiling of compound actions on multiple receptor activities in living cells

The multimodal Munich Clinical Deep Phenotyping study to bridge the translational gap in severe mental illness treatment research

Introduction: Treatment of severe mental illness (SMI) symptoms, especially negative symptoms and cognitive dysfunction in schizophrenia, remains a major unmet need. There is good evidence that SMIs have a strong genetic background and are characterized by multiple biological alterations, including disturbed brain circuits and connectivity, dysregulated neuronal excitation-inhibition, disturbed dopaminergic and glutamatergic pathways, and partially dysregulated inflammatory processes. The ways in which the dysregulated signaling pathways are interconnected remains largely unknown, in part because well-characterized clinical studies on comprehensive biomaterial are lacking. Furthermore, the development of drugs to treat SMIs such as schizophrenia is limited by the use of operationalized symptom-based clusters for diagnosis. Methods: In line with the Research Domain Criteria initiative, the Clinical Deep Phenotyping (CDP) study is using a multimodal approach to reveal the neurobiological underpinnings of clinically relevant schizophrenia subgroups by performing broad transdiagnostic clinical characterization with standardized neurocognitive assessments, multimodal neuroimaging, electrophysiological assessments, retinal investigations, and omics-based analyzes of blood and cerebrospinal fluid. Moreover, to bridge the translational gap in biological psychiatry the study includes in vitro investigations on human-induced pluripotent stem cells, which are available from a subset of participants. Results: Here, we report on the feasibility of this multimodal approach, which has been successfully initiated in the first participants in the CDP cohort; to date, the cohort comprises over 194 individuals with SMI and 187 age and gender matched healthy controls. In addition, we describe the applied research modalities and study objectives. Discussion: The identification of cross-diagnostic and diagnosis-specific biotype-informed subgroups of patients and the translational dissection of those subgroups may help to pave the way toward precision medicine with artificial intelligence-supported tailored interventions and treatment. This aim is particularly important in psychiatry, a field where innovation is urgently needed because specific symptom domains, such as negative symptoms and cognitive dysfunction, and treatment-resistant symptoms in general are still difficult to treat

Studying and modulating schizophrenia-associated dysfunctions of oligodendrocytes with patient-specific cell systems

Abstract Postmortem studies in patients with schizophrenia (SCZ) have revealed deficits in myelination, abnormalities in myelin gene expression and altered numbers of oligodendrocytes in the brain. However, gaining mechanistic insight into oligodendrocyte (OL) dysfunction and its contribution to SCZ has been challenging because of technical hurdles. The advent of individual patient-derived human-induced pluripotent stem cells (hiPSCs), combined with the generation of in principle any neuronal and glial cell type, including OLs and oligodendrocyte precursor cells (OPCs), holds great potential for understanding the molecular basis of the aetiopathogenesis of genetically complex psychiatric diseases such as SCZ and could pave the way towards personalized medicine. The development of neuronal and glial co-culture systems now appears to enable the in vitro study of SCZ-relevant neurobiological endophenotypes, including OL dysfunction and myelination, with unprecedented construct validity. Nonetheless, the meaningful stratification of patients before the subsequent functional analyses of patient-derived cell systems still represents an important bottleneck. Here, to improve the predictive power of ex vivo disease modelling we propose using hiPSC technology to focus on representatives of patient subgroups stratified for genomic and/or phenomic features and neurobiological cell systems. Therefore, this review will outline the evidence for the involvement of OPCs/OLs in SCZ in the context of their proposed functions, including myelination and axon support, the implications for hiPSC-based cellular disease modelling and potential strategies for patient selection