Meibomian gland dysfunction is suppressed via selective inhibition of immune responses by topical LFA-1/ICAM antagonism with lifitegrast in the allergic eye disease (AED) model

Abstract

Purpose: The etiology of meibomian gland dysfunction (MGD) is incompletely understood, despite being a common ophthalmic condition and an area of unmet medical need. It is characterized by an insufficiency in glandular provision of specialized lipids (meibum) to the tear film and is a major cause of dry eye. Work in the allergic eye disease (AED) mouse model has revealed an immunopathogenic role in MGD causation, now raising interest in the applicability of immunomodulatory therapies. As such, we herein ask whether inhibition of lymphocyte function associated antigen (LFA)-1/intracellular adhesion molecules (ICAM)-1 signaling via topical lifitegrast administration has a therapeutic effect on MGD in AED mice.

Methods: Mice were induced with AED by i.p. injection of ovalbumin (OVA) mixed with alum and pertussis toxin, followed 2 weeks later by once daily topical OVA challenges for 7 days. Mice were treated topically with 5% lifitegrast ophthalmic solution or vehicle (PBS) 30 min prior to challenge. We developed a clinical ranking method to assess MGD severity, and also scored clinical allergy. Conjunctivae and draining lymph nodes were collected for flow cytometry.

Results: Topical lifitegrast significantly inhibited clinical MGD severity, which was associated with diminished pathogenic T_H17 cell and neutrophil numbers in the conjunctiva. No significant change in conjunctival T_H2 cells or eosinophils, and only marginal differences in ocular allergy were observed.

Conclusions: In AED mice, lifitegrast inhibited MGD severity marked by a reduction in select immune populations in the conjunctiva. Our findings warrant future examination of lifitegrast in the treatment of patients with forms of MGD.

Keywords: Dry eye; Lifitegrast; MGD; Meibomian gland dysfunction; Neutrophils; T(H)17.

Fig. 1.. Clinical MG Orifice Rank Assesses Severity of Meibomian Gland Plugging

A clinical ranking method was developed to quantify apparent differences in the severity of MG plugging based on the size and appearance of MG orifice at the posterior lid margin. Representative slit lamp images for each numerical clinical rank above and corresponding description below each image.

Fig. 2.. Topical Lifitegrast Administration Reduces Clinical MGD Plugging

Clinical severity of MG plugging was ranked after treatment and challenge in AED and naïve mice. (A) Rank distribution of individual MG orifices across all naïve mice and AED mice on day 7 of treatment with lifitegrast or vehicle. (B) Average rank of orifices per eye on day 7 of treatment and allergen challenge comparing naïve (n = 17), AED vehicle (n = 21), and AED lifitegrast (n = 22). Data were collected across 4 independent experiments and plotted as means ± SEM with individual counts. Two-way ANOVA was performed with Tukey’s post hoc test (*P < 0.05, **P < 0.01, ***P < 0.001, n.s. indicates non-significance).

Fig. 3.. Clinical Ocular Allergy is Marginally Reduced with Topical Lifitegrast Administration

Clinical disease was scored by summing the individual scores of tearing, lid edema, chemosis and conjunctival redness at (A) 20 min and (B) 6 h each day after allergen challenge. Lifitegrast or vehicle was administered at 30 min prior to allergen challenge. Day 7 conjunctival allergy score at (C) 20 min and (D) 6 h was compared between AED vehicle and AED lifitegrast treatment. Data were collected across four independent experiments with naïve (n = 12), AED vehicle (n = 21), and AED lifitegrast (n = 22) mice and is represented as means ± SEM. Two-way ANOVA with Tukey’s post hoc test (A,B) and Mann-Whitney test (C,D) were performed, respectively (*P < 0.05, **P < 0.01, ***P < 0.001, n.s. indicates non-significance).

Fig. 4.. Topical Lifitegrast Administration Reduced Pathogenic T_H17 responses but not T_H2 in Conjunctiva

(A) Representative flow cytometry gating strategy for conjunctival T cells (CD45⁺CD90⁺CD3⁺CD4⁺). Representative flow contour plots and quantification of the number of (B) T_H1 (CD4⁺Tbet⁺), (C) T_H2 (CD4⁺ GATA-3⁺), (D) T_H17 (CD4⁺RoRγt⁺), and (E) IFN-γ producing T_H17 (CD4⁺RORγt⁺T-bet⁺) conjunctival cells (n = 6 mice per group). Data represented as means ± SEM with individual values. One-way ANOVA was performed with Tukey’s post-hoc test. (*P < 0.05, **P < 0.01, ***P < 0.001, n.s. indicates non-significance).

Fig. 5.. Topical Lifitegrast Administration Reduced Conjunctival PMN recruitment, but not Eosinophils

(A) Representative flow cytometry gating strategy for conjunctival leukocytes (CD45⁺), PMNs (CD45⁺Ly6G⁺), and eosinophils (CD45⁺Siglec-F⁺). (B) Quantification of number of leukocytes in the conjunctiva. (C) Representative contour plots and quantification of number of eosinophils and PMNs from naïve (n = 6), AED vehicle (n = 7), and AED Lifitegrast (n = 9) conjunctiva. Data across 3 independent experiments were plotted as means ± SEM with individual values. One-way ANOVA was performed with Tukey’s post-hoc test. (*P < 0.05, **P < 0.01, ***P < 0.001, n.s. indicates non-significance).