CRISPR-mediated Inhibition of TGF-β2 in the Trabecular Meshwork

Project Summary

In primary open-angle glaucoma (POAG) patients, the increase of eye pressure is due to damage in the trabecular meshwork (TM), a specialized tissue that functions as a safety valve. Many studies including ours have shown that cell factors, including TGF-β2 are elevated in the POAG TM, and their elevation induces glaucomatous damages. So one key question is how these factors are elevated in the glaucomatous TM (GTM). Our recent publication showed that there is an excessive modification of DNA associated proteins in the GTM, and this modification plays an important role in the determination of the level of TGF-beta2 in the GTM. Therefore, one potential therapeutic strategy for treating POAG would be lowering TGF-β2 by manipulation of DNA-associated protein modification.

Recently, a novel technology called CRISPR interference (CRISPRi) enables researchers to achieve DNA-associated protein modification for individual cell factors. The CRISPRi system consists of an enzyme (dCas9-KRAB) and an RNA molecule.  The RNA molecule would guide the enzyme to the desired location in TM cells and restore abnormal DNA-associated protein modification to normal status. This system is suitable for POAG because it may correct the pathology in the TM. We will use TM cells, perfusion cultured human eyes, and mice to study the efficiency of the CRISPRi system. Upon completion of this project, we expect to have determined whether the dCAS9-KRAB system is a useful tool/therapy for treating glaucoma.