Developing micro-RNAs as biomarkers for Primary Open Angle Glaucoma

Abstract

 

Purpose

 

Primary open angle glaucoma (POAG) often goes undetected, highlighting the need for novel diagnostic or treatment response biomarkers. A family of ~2500 noncoding microRNAs (miRNAs) function as key molecular regulators by repressing their target gene. miRNA-based therapeutics are promising strategies to treat and detect various disorders. Recent studies illustrated presence of ~500 miRNAs in aqueous humor (AqH). Limited data is available on differential miRNA levels in AqH from patients with various subtypes of POAG, or on the correlation between miRNAs in AqH and plasma. We aimed to identify miRNAs as potential POAG biomarkers in AqH and plasma.

 

Methods

 

AqH samples from 3 cataract patients (control), 3 POAG patients with peripheral visual field loss (PVFL), and 3 patients with early paracentral visual field loss, a POAG subtype associated with vascular dysregulation (ePaVFL), were analyzed on the Qiagen MIHS-3218Z platform (~1800 miRNAs). Subsequently, AqH samples from 4 control and 3 PVFL patients were analyzed on a custom-built array (CMIHS-02263) containing 372 miRNAs that were differentially expressed, highly abundant, or minimally variable on MIHS-3218Z. Finally, plasma and AqH samples from 3 control and 3 ePaVFL patients were analyzed on the Qiagen miFinder384HC platform. Supervised clustering by RandomForests detected best classifying miRNAs, and a linear mixed effects model controlling for “subject” as a random factor tested correlation in plasma versus AqH miRNA profiles.

 

Results

 

~800 unique miRNAs in 5ml AqH were identified on the MIHS-3218Z platform. Supervised clustering separated the three sample types and identified two miRNAs among the top classifying input variables (Fig. 1A,D). Similarly, CMIHS-02263 separately clustered control versus PVFL, using five top classifying miRNAs (Fig. 1B,E). Abundance of all assayed miRNAs was correlated between AqH and plasma in the same subjects (P<0.0001). Two input miRNAs were sufficient to completely separate the two sample types (Fig. 1C,F).

 

Conclusions

 

Our pilot study suggests that miRNAs are abundant in AqH from cataract and POAG patients and may be valuable diagnostic biomarkers for various subtypes of POAG. In addition, levels of miRNAs in plasma, a more readily accessible biomarker source than AqH, appear to correlate with AqH levels and allowed us to distinguish between cataract patients and ePaVFL patients.