Viewing Blindness In A New Genetic Light

Don’t be fooled by its name, exfoliation syndrome has nothing to do with the removal of dead skin cells. Instead, the syndrome is characterised by the build-up of abnormal proteins aggregates in the eye’s anterior chamber, causing glaucoma and vision loss.

Despite affecting up to 70 million people worldwide in 2010, the origins of the condition remain elusive. Though researchers have used genome-wide association studies (GWAS) to probe for predictive genetic markers, the results have so far been inconclusive.

To demystify the disease and pinpoint exfoliation syndrome’s genetic cause, an international collaborative team led by researchers from Agency for Science, Research and Technology’s (A*STAR) Genome Institute of Singapore (GIS),  Bioprocessing Technology Institute (BTI) and Singapore Eye Research Institute (SingHealth) turned to a more focused approach: whole exome sequencing.

Unlike GWAS which spans the whole genome, exome sequencing examines only the exome, or the genome’s protein-coding regions. In this feature, study co-author Khor Chiea Chuen, Senior Principal Investigator at GIS, details how the team finally found the missing genetic link for exfoliation syndrome.

Narrowing the Sequencing Scope

For all its complexity, the nucleotide sequences that make up our genome can be classified as either exons or introns. Exons are composed of the segments of DNA that are directly translated into proteins; meanwhile, introns do not directly code for proteins but are essential for gene regulation.

While previous GWAS efforts had identified common noncoding variants associated with exfoliation syndrome, it was tricky to determine exactly which genes were affected by these variants. In contrast, though rare, protein-coding genetic variants can directly implicate the disease-causing gene.

Indeed, previous studies have shown that the exome accounts for around 1.1 percent of the whole genome but contains over 80 percent of disease-related variants1, leading the researchers to adopt exome sequencing instead. Though researchers look at fewer genes using exome sequencing, they do so at a greater resolution and with higher chance of actionability—making the technique key for precision medicine.

“Whereas GWAS only scans the surface of the human genome using array-based technology, exome sequencing directly interrogates all possible genetic variation in the exome,” explained Khor. “Because of this depth, the technique can detect instances of protein-changing genetic variants occurring in just a single person.”

Finding the Needle in the Haystack

To identify the rare genetic variants linked to exfoliation syndrome, Khor and collaborators embarked on a wide-ranging global study. Sequencing the exomes of individuals with and without exfoliation syndrome across three continents between 1999 and 2019, they identified 415,871 rare variants across 18,753 genes predicted to impair protein function.

Amidst the sea of genes, one stood out: CYP39A1. “CYP39A1 emerged as the gene with the strongest statistical evidence of enrichment for rare variants in affected individuals compared to unaffected individuals,” noted Khor.

In healthy individuals, CYP39A1 codes for a protein that converts cholesterol into its derivatives. However, those with exfoliation syndrome had mutations resulting in the loss of CYP39A1 function. This finding suggests that those with the mutation were unable to metabolise cholesterol effectively, resulting in its build up in the eye.

“For the past 20 years, researchers thought that exfoliation syndrome was a disease involving proteins. We are the first to show that cholesterol also plays a crucial role,” said Khor. “The key breakthrough was when we stained cholesterol within the exfoliative material, confirming that it is an integral part of the disease process.”

As the first team to demonstrate the link between cholesterol and exfoliation syndrome, their discovery opens the door to treatments specifically targeting CYP39A1. Currently, exfoliation syndrome is treated using medication to lower blood pressure or laser surgery, both of which do not target the disease’s origins2.

But by developing treatments that retard the formation of exfoliative material, clinicians can address the root of the problem and advance precision medicine—showing that identifying specific variants can someday result in targeted treatments suitable for each individual.

“This study showed that it is indeed possible to perform functional genomic studies in Singapore,” added Khor. Moving forward, the team is looking to develop a high throughput screen for identifying different CYP39A1 variants, potentially paving the way for assessing exfoliation syndrome risk and diagnosing the disease before it manifests.

References:

1. What is exome sequencing [Online]. https://sapac.illumina.com/techniques/sequencing/dna-sequencing/targeted-resequencing/exome-sequencing.html

2. Diagnosis and management of pseudoexfoliation glaucoma [Online].  https://www.aao.org/eyenet/article/diagnosis-management-of-pseudoexfoliation-glaucoma