Mid-sized GDNF Mimics For Neural Regeneration

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Project Summary Repair of traumatic injuries relies upon glial cell line-derived neurotrophic factor (GDNF), and related extracellular cytokines collectively called GDNF family ligands (GFLs). GFLs interact with solubilized forms of the GDNF-family receptors (sGFRα1–4) forming complexes which then can bind and activate NCAM (nuclear cell adhesion molecule) and RET (REarranged on Transfection) receptors leading to intracellular signaling and a range of responses conducive to neuronal connectivity. GFLs have been tested in animals and in clinical trials. However, they have poor in vivo stabilities, unfavorable tissue permeation characteristics, and are expensive to prepare with batch-to-batch reproduciblity. Gene therapy approaches have also been attempted, but these are extremely risky because continued expression leads to uncontrollable growth post therapy. Few small molecule mimics of GFL•GFRα interface regions have been reported in the literature. This is surprising because appropriate small molecules could cause conformational changes in sGFRαs transforming them into NCAM/RET agonists which may communicate between cells (trans-signaling) to trigger valuable responses for repair of the peripheral nervous system after trauma. Preliminary studies feature design, synthesis, and testing of two mimics of the GDNF loop which is responsible for most if the GFL•GFRα interface interaction (ie the interface “hot loop”). These loop mimics bind GFRα1 (best so far Kd 240 nM), and are currently being tested in cellular models for repair of traumatic injuries to the peripheral nervous system (PNS). This application is to optimize these initial leads and test them more extensively. Year 1 will focus on on design, syntheses, and GFRα1-binding affinities for similar “cyclo-organopeptide hot loop mimics” by the PI (10 – 20 compounds). Free loop mimics with superior GFRα binding affinities, and samples of ones covalently anchored to hyaluronic acid supports (which mimic the media around synapses), will be selected for Aim 2. The PI is an expert on design and synthesis of growth factor hot loop mimics; he will oversee that part of the work closely. In year 2 the emphasis will shift to testing the best hot-loop mimics identified at that time in 2D and 3D-cellular models for PNS recovery from traumatic injury. Active compounds will also be assayed to test if they cause intracellular activation of NCAM and/or RET. That work will be overseen by Professor Sakiyama, the subcontractor on this application, who has extensive experience with GFLs and supported GFLs, particularly GDNF, tested 2D and 3D cellular assays for neurite outgrowth on sensory and motor neurons. She is an expert in neuronal repair.