Glaucoma, a leading cause of blindness worldwide (70 million patients), is managed medically by treating the symptom of increased intraocular pressure (IOP), but 10% of patients still go blind. IOP is controlled in the anterior segment of the eye, which contains the trabecular meshwork (TM) extracellular matrix, the anatomical pathway for drainage of aqueous humor fluid. The TM tissue is diseased in most forms of glaucoma; loss of TM homeostasis leads to elevated IOP. Hereditary open angle glaucoma, affecting ~3 million young patients, is caused by mutations in myocilin, a protein highly expressed in the TM. Since 3/2011, studies funded from R01EY021205 have changed the paradigm for anti-glaucoma therapeutics by laying the molecular foundation for approaches that target the disease process, which are now being pursued in academia and industry. We clarified molecular details of the toxic gain-of-function pathogenic mechanism in which mutant myocilin accumulates in the endoplasmic reticulum (ER) of TM cells, leading to TM cell death and an accelerated timeline for vision loss. Studies from R01EY021205 (a) contributed fundamental knowledge of myocilin structure, (b) discovered a counter-productive interaction between myocilin and the ER-resident Hsp90 chaperone Grp94, and (c) characterized myocilin misfolding as amyloid. Wild-type and many different myocilin variants harbor a misfolding propensity; thus, proteostasis issues identified in familial myocilin- associated glaucoma are likely at play in many more patients. Amyloid formation by myocilin places glaucoma alongside more well-studied amyloid diseases like Alzheimer and SOD-1 dependent amyotrophic lateral sclerosis, yet our comprehension of the role of amyloid in glaucoma is in its infancy. Our current objective is to better understand molecular aspects of myocilin fibrilization, focused on the relevant olfactomedin (OLF) domain. Our multidisciplinary team will (a) clarify initiation of aggregation by studying solution structures of wild-type and selected OLF variants, as well as corresponding multi-length scale dynamics, using hydrogen-deuterium exchange mass spectrometry and nuclear magnetic resonance (NMR) structure and relaxation methods (Wade Van Horn, Co-I) (b) compare the end-point structures of selected OLF aggregates to known amyloids by solid state NMR (Anant Paravastu, Co- I) and evaluate cytotoxicity of intermediate aggregates and (c) evaluate common allele full-length myocilin variants for experimental hallmarks of pathogenicity. The expected outcome is a better understanding of the myocilin misfolding process at the molecular level, including molecular determinants of pathogenicity, to enable novel modalities for studying, diagnosing, and treating myocilin-associated glaucoma. More broadly, continued structure/dysfunction studies of myocilin will not only contribute to our understanding of glaucoma and its role in the TM, but will also extend our comprehension of the many other OLF domains, which are implicated broadly in physiology and diseases.

Public Health Relevance

Our long-term goal is to develop a new therapy for glaucoma, a prevalent eye disease characterized by increased intraocular pressure, neurodegeneration of the retina, and vision loss. We are focusing on myocilin, an extracellular matrix protein involved in regulating eye pressure; mutations in myocilin lead to an early-onset, inherited form of glaucoma. We are studying disease-causing mutants to understand the molecular mechanisms of pathogenesis, which, in turn, is guiding our efforts develop a new therapeutic that treats the underlying cause of the disease.

National Institute of Health (NIH)
National Eye Institute (NEI)
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Macromolecular Structure and Function B Study Section (MSFB)
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Liberman, Ellen S
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Georgia Institute of Technology
Schools of Arts and Sciences
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Patterson-Orazem, Athéna C; Hill, Shannon E; Fautsch, Michael P et al. (2018) Epitope mapping of commercial antibodies that detect myocilin. Exp Eye Res 173:109-112
Huard, Dustin J E; Crowley, Vincent M; Du, Yuhong et al. (2018) Trifunctional High-Throughput Screen Identifies Promising Scaffold To Inhibit Grp94 and Treat Myocilin-Associated Glaucoma. ACS Chem Biol 13:933-941
Wang, Yiming; Gao, Yuan; Hill, Shannon E et al. (2018) Simulations and Experiments Delineate Amyloid Fibrilization by Peptides Derived from Glaucoma-Associated Myocilin. J Phys Chem B 122:5845-5850
Huard, Dustin Je; Lieberman, Raquel L (2018) Progress toward development of a proteostasis drug for myocilin-associated glaucoma. Future Med Chem 10:1391-1393
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Goldenzweig, Adi; Goldsmith, Moshe; Hill, Shannon E et al. (2016) Automated Structure- and Sequence-Based Design of Proteins for High Bacterial Expression and Stability. Mol Cell 63:337-346

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