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Abstract:
Enhancing the efficacy of cytokine-based therapies by systematically tuning how an agonist associates its receptor is emerging as a powerful new concept in drug discovery. De novo protein design can serve an excellent tool for tailoring novel cytokine receptor agonists with altered binding affinities and geometries. Here, we report the design and characterization of agonists that tune the granulocyte-colony stimulating factor receptor (G-CSFR) activity, which is central for the proliferation and granulocytic differentiation of hematopoietic stem cells. Selected design agonists were deployed to study the impact of varying the binding affinity and dimerization geometry on receptor association, downstream signaling, and cellular response. Hence, we achieved design agonists with altered signaling specificities that are hyper-thermostable, engage the receptor with tight affinity, and bias granulopoietic differentiation over triggering proliferation. Furthermore, the design agonists modulate the kinetics and amplitudes of signal transduction pathways, yielding differential gene expression patterns. Thus, unlike G-CSF, the design agonists could achieve selective activation of hematopoietic stem cell functions with minimal undesired immunomodulatory effects. These findings demonstrate the potential of dissecting the complexity of G-CSFR signaling, outcompeting the native ligand, and therefore open ways for more specific therapeutic applications.
