Surveying the Tear Proteome to Stratify Glaucoma Patients

SCIEX describes how mass spec is extending precision medicine to eye disease

Aaron Hudson, PhD
Aaron Hudson, PhD
vice president and general manager of global marketing, SCIEX

Glaucoma is called the silent thief of sight because in its most common form, there are usually no symptoms until the disease has insidiously progressed to the point of irreparable optic nerve damage and irreversible sight loss.1,2 If left untreated, the initial loss of peripheral vision will continue to become tunnel vision, which will then shrink down until all sight is lost.

“The thought of being blind sent me into a deep depression.”3

“Looking back, I could find out that there were many times, and a couple of auto accidents, in which I didn’t see cars coming from the left or the right side—and that was a consequence of losing that peripheral vision. But you never know that when you don’t know you have any disease.”4

“I find myself, since I’m half-blind, constantly worrying about: what if I lost the vision in my right eye? Because that would change everything about my life. It would change my dreams, it would change my relationships, it would change everything.”5

These quotes, all shared by glaucoma patients, dramatize how people who lose their vision also lose peace of mind. Even eye diseases that do not impair vision can be devastating.

“After getting dry eyes, I became very frustrated and almost depressed for a while. It’s hard to deal with.”6

The most common form of glaucoma, open-angle glaucoma, is also the most mysterious. Although a strong hereditary component has been implicated, the underlying disease mechanisms remain largely a mystery.2

Biomarkers in tears

To unravel the mystery and to identify biomarkers for the diagnosis and stratification of patients for precision medicines, researchers in Finland and Singapore are working together to analyze the proteome of tears from individuals with eye disease. Specifically, the researchers are using advanced analytics techniques with liquid chromatography (LC) triple time-of-flight (TOF) mass spectrometry (MS) and SWATH Acquisition to rapidly obtain complete data on tear samples from individual patients.

Tear fluid is especially useful because it is a more accessible and less complex body fluid than serum or plasma, and sampling is much less invasive. Using the SWATH Acquisition method enables the capture of a wealth of information from each sample in one go, meaning that researchers can go back to interrogate their data time and time again as more information emerges about the biology of the eye and tears. The advantage of the MS method is that it allows researchers to process samples from individual patients quickly, sensitively, and precisely, eliminating the need to pool samples.7–11 With MS, it is possible to analyze the proteomic profiles of individual patients, even in large clinical trials. Eventually, it may bring proteomic analysis to clinical practice. It has the precision needed to achieve precision/stratified therapy.7,11

Clinical studies

In one clinical study, LC-MS and the TripleTOF system were used to evaluate the expression levels of proteins in tears between patients with glaucoma. People with glaucoma are prone to getting concomitant ocular surface disease, such as dry eye disease. During a year-long study of patients with glaucoma who were experiencing dry eye symptoms, the researchers were able to identify protein biomarkers that predicted which patients would benefit most from a switch of eye drop medication from one with preservatives to one without preservatives.7

Using SWATH Acquisition, the researchers discovered that the dry eye symptoms of patients with increased levels of proinflammatory proteins and decreased levels of protective proteins improved more after the medication switch than those of other patients. The study went on to define three subpopulations based on these and other biomarkers: a group that did not respond to the medication switch, a group that had moderate improvement in symptoms in response to the switch, and a group that benefitted the most from the medication switch.7

Another study using the TripleTOF system with SWATH Acquisition to examine the proteomic expression of tears has found proteins that could be used as biomarkers to stratify patients with dry eye disease, identifying those who would benefit most from treatment with flourometholone, as opposed to polyvinyl alcohol.8 Similarly, studies using SWATH Acquisition have revealed proteins that may be potential biomarkers for predicting progression to severe thyroid eye disease in patients with autoimmune thyroid disease,9 whereas other proteins have been discovered that may be indicative of aging and the effects of aging in eye tissues and functions.10

A wider view

Tears are also being analyzed to understand a host of other eye diseases and infections, such as diabetic retinopathy, peripheral ulcerative keratitis, aniridia, ocular allergies, and trachoma.12 Research continues apace to better understand the physiology and pathophysiology of the eye and eye diseases, particularly over time, as many eye diseases occur more often with older age.

The increasingly common utilization of advanced analytical technologies such as MS to better interrogate biological samples from individual patients and healthy controls means that we are getting ever closer to the identification and use of biomarkers to predict and diagnose disease, as well as to monitor patient responses to therapeutic agents, marking progress in the field of predictive, preventive, and personalized medicine, both in general and in terms of addressing eye disease. Precision medicine promises to revolutionize healthcare for many people, not only those with eye disease but also individuals with other diseases, such as cancer and cardiovascular disease.


1. University of Utah Health. Glaucoma: The Silent Thief of Sight.
2. National Eye Institute, National Institutes of Health. Glaucoma: The ‘Silent Thief’ Begins to Tell Its Secrets. 01/12/14.
3. Nättinen J, Jylhä A, Aapola U, et al. Patient Stratification in Clinical Glaucoma Trials Using the Individual Tear Proteome. Sci. Rep. 2018; 8: Article 12038.
4. Glaucoma Australia. My Glaucoma Story. Victoria’s Story.
5. Glaucoma Research Foundation. Art Takahara: Learning about Glaucoma
6. Glaucoma Research Foundation. Personal Story: Hannah Eckstein.
7. Cook N, Mullins A, Gautam R, et al. Evaluating Patient Experiences in Dry Eye Disease Through Social Media Listening Research. Ophthalmol. Ther. 2019; 8(3): 407–20.
8. Nättinen J, Jylhä A, Aapola U, et al. Topical fluorometholone treatment and desiccating stress change inflammatory protein expression in tears. Ocul. Surf. 2018; 16: 84–92.
9. Chng CL, Seah LL, Yang M, et al. Tear Proteins Calcium Binding Protein A4 (S100A4) and Prolactin Induced Protein (PIP) are Potential Biomarkers for Thyroid Eye Disease. Sci. Rep. 2018; 8: Article 16936.
10. Nättinen J, Jylhä A, Aapola U, et al. Age‑Associated Changes in Human Tear Proteome. Clin. Proteomics 2019; 16: 11.
11. Jylhä A, Nättinen J, Aapola U, et al. Comparison of iTRAQ and SWATH in a Clinical Study with Multiple Time Points. Clin. Proteomics 2018; 15: 24.
12. Hagan S, Martin E, Enríquez-de-Salamanca A. Tear Fluid Biomarkers in Ocular and Systemic Disease: Potential Use for Predictive, Preventive and Personalised Medicine. EPMA J. 2016; 7: 15.

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