Abstract
Glaucoma is a neurodegenerative disease characterized by loss of retinal ganglion cells (RGCs) and their
axons. Damage from glaucoma is permanent and may cause irreversible blindness. Glaucoma is a major
public health problem in the United States and worldwide, where it is the leading cause of irreversible
blindness. Four classic risk factors have been identified for the most common type of glaucoma, primary open
angle glaucoma (POAG): increasing age, race / ethnicity, family history, and increased intraocular pressure
(IOP). Currently, all treatments for glaucoma slow or halt disease by targeting one risk factor, increased IOP.
However, IOP-lowering therapies fail to prevent vision loss in many patients. Thus, glaucoma patients
desperately need new, more effective therapies that target other aspects of glaucoma beyond elevated IOP,
such as the mechanisms underlying family history of glaucoma. Unfortunately, most genetic risk associated
with glaucoma remains poorly understood. Most cases of POAG have a complex genetic basis and involve
many risk factors. To date, >127 risk factor loci have been identified with genome-wide association studies
(GWAS's). In each of these loci, several single nucleotide polymorphisms (SNPs), have been associated with
risk for POAG and the effect of these SNPs on the nearest gene has been presumed to be the source of risk.
However, little progress has been made in precisely defining these loci, including the causative SNP(s),
gene(s), or mechanism(s). Very few of the disease-associated SNPs the >127 glaucoma risk loci are in coding
sequence. Consequently, we hypothesize that the functional SNPs in each locus confer glaucoma risk by
increasing or decreasing transcript levels of effector genes in the locus. To test our hypothesis, we have
prioritized three loci for detailed studies (chr 4p14, chr 21q21.3, and chr 17q21.3), which each containing an
exemplar candidate for being the effector gene (APBB2, APP, MAPT), which are all expressed in RGCs and all
share links with neurodegeneration associated with Alzheimer disease. To test whether these are indeed the
effectors and study their mechanisms, we propose a complementary set of experiments using human, mouse,
and molecular approaches. In Specific Aim 1 we will use genotyped human donor retinal tissue to identify how
transcript and protein levels for the exemplar candidates are altered in association with the high-risk allele at
each locus (using scRNAseq, IHC, and ELISA), as well as assess other transcriptomic changes across the
locus (and genome). We will also use BiT-STARR-seq, to identify SNPs in each locus capable of changing
transcription. In Specific Aim 2 we will use AAV2 constructs in mice to manipulate expression levels of the
exemplar candidate genes and study how dysregulation of these genes influences RGC health. With
completion of these experiments, we will advance understanding the basic biology of glaucoma risk factors
identified by GWAS. We will also identify gene regulation and relevant biological pathways associated with
familial glaucoma risk, which are key to the development of new sight-saving therapies.