1. Introduction

Non-infectious posterior uveitis (NIU) is defined as inflammation of the uveal tract. Approximately 10–15% of blindness affecting young and middle-age people in the West is caused by uveitis ^[1]. Therapies for posterior uveitis are limited to corticosteroids, cyclosporine and tumour necrosis factor (TNF)-blocking agents as well as off-label use of methotrexate, azathioprine and mycophenolates ^[2][3]. However, serious side effects (e.g., glaucoma, cataracts, liver failure and immunosuppression) can arise from long-term use of these treatments ^[4]. Therefore, there is an urgent need for new therapeutic interventions which are more specific, and which cause fewer side effects. To achieve this goal, an improved understanding of disease pathogenesis is needed in particular for the identification of key mediators that may be therapeutic targets.

Experimental Autoimmune Uveitis (EAU) is an animal model of NIU which is induced by CD4⁺T cell immune responses to retinal antigens. The main features of EAU are retinal and/or choroidal inflammation, retinal vasculitis, photoreceptor destruction and loss of vision, all of which resemble pathological features in human NIU. The model is well-established and commonly used for investigating fundamental mechanisms of retinal immunopathogenesis. EAU can be induced in many species but is most commonly induced in mice and rats by active immunisation with retinal antigens: interphotoreceptor binding protein (IRBP) for mice, and retinal soluble antigen (SAg) for rats ^[5][6][7].

Whilst angiogenesis and ischemia are not a common feature of uveitis, vascular inflammation, tissue damage and retinal complications are involved in vision loss ^[8][9]. Neovascularisation is a major problem in ocular diseases affecting the retina. The underlying stress leading to the induction of new vessel growth is mainly a result of ischemia but also inflammation, which causes the induction of angiogenic factors, including vascular endothelial growth factor A (VEGF-A), by local tissue resident cells ^[10]. VEGF-A has been detected in intraocular fluids during EAU and in some cases of NIU ^[11][12][13]. In this review, VEGF-A is referred to as VEGF unless indicated otherwise.

VEGF expression in inflammatory or ischemic conditions has been of great interest due to the recent success of therapeutic targeting of angiogenic pathways with anti-VEGF antibodies in ocular vascular diseases ^[14][15]. However, based on experimental approaches, a high level of VEGF does not always lead to angiogenesis and new vessel formation ^[16]. The reason might be that additional microenvironmental conditions/mediators are needed for angiogenesis. Such mediators may include cytokines, growth factors and bioactive lipids which all have the potential to alter tissue homeostasis, especially in the immune-privileged posterior segment in which the ocular microenvironment provides protection against retinal immune insults. Hence, there is a need to understand the retinal vascular pathways involved in uveitis and in EAU progression.

Leukotrienes (LT) have been shown to be involved in VEGF expression and in inducing angiogenesis whilst causing vascular inflammation ^[17][18][19]. Recent findings have demonstrated clinical efficacy of targeting the high-affinity LTB₄ receptor (BLT1) pathway in EAU by applying a BLT1 antagonist ^[20] and by capturing LTB₄ which prevented tissue damage and retinal complications ^[21] and suggest that the LTB₄ pathway is a promising new therapeutic target in intraocular inflammatory diseases.

2. Retinal Damage Associated with Disease Severity and Potential Treatment

2.1. Retinal Damage Associated with Disease Severity

Neovascularisation at the site of inflammation fuels the ongoing inflammatory process. In several chronic diseases affecting the retina (e.g., diabetic retinopathy, retinal vein occlusion, AMD and chorioretinal vein occlusion), neovascularisation is a major problem. Pathological retinal neovascularisation is characterised by leaky and tuft-like vessels, which are associated with retinal exudates and haemorrhage, leading to retinal damage, retinal detachment, or both ^[22]. The underlying stress leading to the induction of new vessel growth is mainly ischemia, which causes the induction of VEGF by local cells. However, the impact of inflammation cannot be ruled out since VEGF has been detected in ocular Behçet disease and uveitic macular oedema ^[11][14]. Besides the hypoxia-regulated growth factors, other non-oxygen-regulated growth factors acting in this function include the Tie (tyrosine kinase with immunoglobulin-like and EGF-like domain)1-Tie2 receptors, the Tie2 ligand angiopoietin 2. Several cytokines, hormones, and growth factors also regulate VEGF gene expression, leading to the release of VEGF ^[23]. VEGF is the most specific mitogenic factor for vascular endothelial cells. Phenotypical modifications in retinal endothelial cells during EAU progression have been proposed to increase the level of angiogenic factors including VEGF ^[24]. Its role in the pathogenesis of uveitic complications such as choroidal-retinal neovascularisation has been described ^[25] and is further supported by the proven efficacy of intravitreal anti-VEGF therapies in uveitis ^[14][15]. It has been demonstrated that a prolonged duration of the inflammatory condition involves cytokines and IL-6 and TNFα are strongly associated with retinal damage and disease severity ^[26].

2.2 Potential Treatment Approaches in Targeting LT Pathways in Retinal Vasculitis

Finding and targeting those pathways having the highest impact on disease outcomes is always a huge challenge. The LTB₄ pathway is clearly active in numerous disease states including inflammatory diseases and remains a target of interest for therapeutic drug development, especially where new therapeutic indications are supported by more definitive mechanistic biology. Drugs targeting the LTB₄ pathway are classified into 2 broad classes: antagonists of the known LTB₄ receptors (BLT1 and BLT2) and inhibitors of the enzymes responsible for generation of LTB₄ (e.g., TA4H and 5-LOX) ^[27].

Some of the BLT receptor antagonists which have reached phase 2 clinical trials are Etalocib (LY293111; Lilly), Amelubant (BIIL 284; Boehringer Ingelheim), Moxilubant (CGS-25019C, LTB-019; Novartis) and CP-105696 (Pfizer). The target diseases have been chronic obstructive pulmonary disease (COPD), asthma, rheumatoid arthritis, cystic fibrosis and cancer. Antagonist LY293111 in asthmatic patients showed that it led to a significant reduction in the number of neutrophils in BAL fluid, but failed to improve respiratory function or airway reactivity after allergen challenge ^[28][29][30]. However, a study examining the role of the BLT1 antagonist, CP-105696, in monkeys showed that the compound inhibited both LTB₄-mediated neutrophil chemotaxis and upregulation of CD11b⁺ cells ^[31]. At least six LTA₄H inhibitors of the current generation have entered clinical trials and reached phase 2 studies. Of the LTA4H inhibitors, Ubenimex (Bestatin; Nippon-Kayaku, Eiger Biopharmaceuticals), Acebilustat (CTX-4430 Celtaxsys) and Tosedostat (CHR-2797; CTI Biopharma) have been applied in asthma, myelodysplastic syndrome, solid tumours, active cystic fibrosis, acne vulgari, pulmonary arterial hypertension (PAH), atopic dermatitis and allergic conjunctivitis ^[31][28].

The well-known 5-LOX inhibitor, Zileuton, is already in clinical use for the maintenance treatment of asthma and has been tried in other inflammatory/allergic diseases including atopic dermatitis and allergic conjunctivitis. Zileuton inhibits VEGF-Induced angiogenesis, expression of cell adhesion molecules (VCAM-1, ICAM-1), TNFα secretion and production of NO ^[18], the latter being required for efficient angiogenesis which is synthesised by endothelial NOS. NO is produced from bone marrow-derived macrophages stimulated with IFNγ and TNFα and is thought to be produced similarly in vivo ^[32]. The anti-angiogenic effect of Zileuton might also relate to the activation of the large-conductance Ca2⁺-activated K⁺ (BK) channel, resulting in activation of pro-apoptotic signalling cascades. Another possible explanation is based on studies which demonstrated that Zileuton was effective in inhibiting biosynthesis of multiple AA metabolites, including 12- and 15-hydroxyeicosatetranolic acid (12-, 15-HETE) in a hamster cheek pouch model of carcinogenesis. 12-HETE is involved in mediating VEGF-induced angiogenesis ^[18]. Therefore, there is a possibility that the anti-angiogenic effect of Zileuton comes from preventing the 12-LOX, and not the 5-LOX, pathway thereby blocking the production of 12-HETE, the metabolite of the 12-LOX pathway ^[28].

A new therapeutic approach could be by using more selective inhibitors of LTB₄ and BLT1 interactions in affected tissues/organs. Nomacopan is one example of a compound that demonstrates functional selectivity for LTB₄ and shows a promising reduction in disease severity in a preclinical uveitis model, a mouse model of Bullous Pemphigoid ^[33] and is now in phase 2 clinical study for bullous pemphigoid.