<> "The repository administrator has not yet configured an RDF license."^^ . <> . . "Target Discovery for Pro-Resolution Therapies in Inflammatory \r\nDiseases Using CRISPR and Small Molecule Screening in iPS-Derived Macrophage"^^ . "Immune-mediated inflammatory diseases (IMIDs) are characterized by chronic non￾resolving inflammation leading to progressive tissue damage. IMIDs are commonly \r\naccompanied by diverse comorbidities and include a large set of diseases, such as systemic \r\nlupus erythematosus, rheumatoid arthritis and multiple sclerosis. Current therapies cannot cure \r\nIMIDs and primarily rely on suppressing autoimmune reactions and inflammation. The \r\ndiseases commonly flare upon therapy cessation, indicating a need for lifelong anti￾inflammatory and immunosuppressive therapies. Thus, developing drugs that enhance the pro￾resolution in IMIDs could facilitate remission maintenance and boost tissue protection. Subsets \r\nof the tissue-resident macrophages (TRM) were shown to drive the pro-resolution and tissue￾protective programs during tissue inflammation. However, the inaccessibility of primary TRM \r\nfrom human tissues/organs affected by IMIDs, calls for the development of in vitro\r\nmacrophage models that resemble the phenotypes and functions of protective TRM subsets.\r\nThus, using a TRM in vitro model combined with CRISPR-Cas9 and tool molecules could \r\nhelp to identify molecular targets driving TRM differentiation/function Thereby accelerating \r\nthe discovery of remission-promoting therapeutics in IMIDs.\r\nTo study TRM differentiation, I implemented and further fine-tuned different iPSC-derived \r\nTRM-like models and an infiltrating macrophage model, including iMACs, iMicroglia and \r\niMonocytes. I characterized these models for their yield, viability, target phenotypic features, \r\nfunctions and robustness. My experiments hinted that the iMonocyte protocol may be more \r\ndependent on iPSC lines used in differentiation. iMAC and iMicroglia protocols provided a \r\ngood yield (3.5-9.5*106\r\nand 3.5-4.7*106 per plate, respectively) and viability (92-96% and 94-\r\n97%, respectively). Moreover, iMACs can be harvested multiple times from one differentiation \r\nrun, while maintaining a similar phenotype. Furthermore, I assessed iMACs and iMicroglia for \r\ntheir amenability for a candidate drug screen and a pooled CRISPR-Cas9 knockout screen with \r\na customized gRNA library of ~700 gene targets. I tested doxycycline-inducible and stably \r\nexpressed Cas9. My experiments showed a difficulty in random Cas9 integration under \r\nantibiotic resistance and inferred an advantage in using Cas9 co-expressed with fluorescent \r\nprotein. Moreover, iMAC model turned out to be difficult to use with the CRISPR-Cas9 \r\nsystem, but iMicroglia with shorter differentiation performed better. However, iMACs \r\nremained a highly reproducible and robust model that can be incorporated into TRM in vitro\r\nstudies using tool molecules.\r\nBased on these results, I performed drug perturbation on iMACs and CRISPR experiments \r\non iMicroglia. The differentiation CRISPR-Cas9 screen at the time of the thesis submission \r\nwas submitted for sequencing and is yet to elucidate genes involved in TRM \r\ndifferentiation/function in iMicroglia. Additionally, I tested the effects of known (Cytochalasin \r\nD) and candidate (splicing inhibitors) drugs on iMAC surface marker phenotype and \r\nefferocytosis (apoptotic cell clearance). Splicing inhibitors Herboxidiene (GEX1A) and \r\nPladienolide B (PladB) targeting SF3B1 subunit of the spliceosome, inhibited iMAC \r\nefferocytosis. \r\nThus, iMAC platform combined with phenotypic and functional macrophage readouts can \r\nrepresent not only a drug screening system but also a new hypothesis-generating tool, \r\nfacilitating new knowledge about macrophage biology and macrophage therapeutic targeting \r\nin IMIDs and possibly cancer. Finally, the pooled CRISPR screen is yet to define genes \r\ninvolved in TRM differentiation/function is ongoing in iMicroglia, which will be followed by \r\nvalidation studies of top hits (beyond my PhD thesis)."^^ . "2026" . . . . . . . "Tomasz"^^ . "Stadler"^^ . "Tomasz Stadler"^^ . . . . . . "Target Discovery for Pro-Resolution Therapies in Inflammatory \r\nDiseases Using CRISPR and Small Molecule Screening in iPS-Derived Macrophage (PDF)"^^ . . . "Target Discovery for Pro-Resolution Therapies in Inflammatory \r\nDiseases Using CRISPR and Small Molecule Screening in iPS-Derived Macrophage (Other)"^^ . . . . . . "Target Discovery for Pro-Resolution Therapies in Inflammatory \r\nDiseases Using CRISPR and Small Molecule Screening in iPS-Derived Macrophage (Other)"^^ . . . . . . "Target Discovery for Pro-Resolution Therapies in Inflammatory \r\nDiseases Using CRISPR and Small Molecule Screening in iPS-Derived Macrophage (Other)"^^ . . . . . . "Target Discovery for Pro-Resolution Therapies in Inflammatory \r\nDiseases Using CRISPR and Small Molecule Screening in iPS-Derived Macrophage (Other)"^^ . . . . . . "Target Discovery for Pro-Resolution Therapies in Inflammatory \r\nDiseases Using CRISPR and Small Molecule Screening in iPS-Derived Macrophage (Other)"^^ . . . . . "HTML Summary of #37256 \n\nTarget Discovery for Pro-Resolution Therapies in Inflammatory \nDiseases Using CRISPR and Small Molecule Screening in iPS-Derived Macrophage\n\n" . "text/html" . . . "000 Allgemeines, Wissenschaft, Informatik"@de . "000 Generalities, Science"@en . . . "500 Naturwissenschaften und Mathematik"@de . "500 Natural sciences and mathematics"@en . .