ophthalmic drug discovery: novel targets and mechanisms … · test visual function, ......

Visual impairment is one of the most feared disabilities. The vast majority of information we acquire from the environment is dependent on our ability to see. Loss of sight affects activities of daily living and substantially reduces quality of life. The leading causes of visual impairment are age-related, and cause damage to the retina and optic nerve. These causes include age-related macular degeneration (AMD), diabetic retinopathy and glaucoma (FIGS 1,2; TABLE 1).

Blindness and visual impairment represent a substan-tial burden to the health-care system. The total annual economic impact of major visual disorders among Americans aged 40 years or older has been estimated at US$35.4 billion1. As the lifespan of our population con-tinues to increase and advances in health care improve longevity, there will be an increasing number of people who are at risk of developing visual impairment, and the economic impact of visual impairment will continue to grow.

The unique features of the eye provide both benefits and challenges for drug discovery and delivery (FIG. 1;

FIG. 2b). In contrast to other parts of the central nerv-ous system, the eye is clinically accessible because it is located outside the cranium. With new non-invasive diagnostic technologies, such as optical coherence tomography, the retina and optic nerve can be clearly visualized and their macro- and microstructures evalu-ated in situ (FIG. 2a). The accessibility of these tissues also

facilitates drug delivery using methods such as eye drops or local injections. Local delivery and exposure to a drug minimizes systemic toxic effects and therefore increases the therapeutic index.

In addition, there are many modalities available to test visual function, thus allowing easy assessment of drug effectiveness. However, despite these many bene-fits there are also challenges related to ocular barriers. For example, ocular barriers impede drug transport and lower the efficacy of many drugs. Such barriers include the blood–ocular barriers, blood flow, lymphatic clear-ance and tear dilution2.

During the past decade, there have been considerable advances in the understanding of the pathogenesis and genetics of ophthalmic diseases. Although there were no effective treatments for wet AMD and retinal vascular diseases 10 years ago, effective therapies such as anti-angiogenic agents now exist. By contrast, latanoprost was launched a decade ago for the treatment of glaucoma (and soon became the first-line treatment) but no new classes of glaucoma drugs have emerged since then.

Various drugs that act by inhibiting complement activation and angiogenic pathways are currently under development for the treatment of AMD, and neuropro-tective drugs are being investigated for both AMD and glaucoma3–6. However, the effective treatment of ocular diseases remains an unmet medical need, and a vigor-ous effort to develop new drugs and therapies is now

1Department of Ophthalmology and Molecular Medicine Research Center, State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu 610041, China.2Department of Ophthalmology and Shiley Eye Center, University of California San Diego, La Jolla, California 92093, USA. 3Institute for Genomic Medicine, University of California San Diego, La Jolla, California 92093, USA.4Department of Nanoengineering, University of California San Diego, La Jolla, California 92093, USA.Correspondence to K.Z. e‑mail: [email protected]:10.1038/nrd3745 Published online 15 June 2012 Corrected online 29 June 2012

Age-related macular degeneration(AMD). A disease process that is characterized by the degeneration of photoreceptor cells in the macula, leading to loss of central vision.

Ophthalmic drug discovery: novel targets and mechanisms for retinal diseases and glaucomaKang Zhang1,2,3, Liangfang Zhang3,4 and Robert N. Weinreb2

Abstract | Blindness affects 60 million people worldwide. The leading causes of irreversible blindness include age-related macular degeneration, retinal vascular diseases and glaucoma. The unique features of the eye provide both benefits and challenges for drug discovery and delivery. During the past decade, the landscape for ocular drug therapy has substantially changed and our knowledge of the pathogenesis of ophthalmic diseases has grown considerably. Anti-angiogenic drugs have emerged as the most effective form of therapy for age-related macular degeneration and retinal vascular diseases. Lowering intraocular pressure is still the mainstay for glaucoma treatment but neuroprotective drugs represent a promising next-generation therapy. This Review discusses the current state of ocular drug therapy and highlights future therapeutic opportunities.

R E V I E W S

NATURE REVIEWS | DRUG DISCOVERY VOLUME 11 | JULY 2012 | 541

© 2012 Macmillan Publishers Limited. All rights reserved

Nature Reviews | Drug Discovery

Retina (DR, RD)

Choroid (blood vessel layer under the retina)

Lens(cataract)

Cornea(keratitis)

Pupil

Optic nerve:goes to thebrain (glaucoma)

Light

Macula (AMD, DME)

Diabetic retinopathyA microvascular complication of type 2 diabetes that is characterized by increased vascular permeability in the initial stages of disease.

GlaucomaA progressive disease that is characterized by optic disc cupping and peripheral visual field loss.

Retinal pigment epithelium(RPE). A monolayer of pigmented cells in the retina, located between the photoreceptor cells and choroid.

MaculaA highly pigmented area near the centre of the retina that is responsible for detailed central vision.

Geographic atrophyAn advanced form of age-related macular degeneration that is characterized by the atrophy of retinal pigment epithelium and photoreceptor cells.

Choroidal neovascularization(CNV). An advanced form of age-related macular degeneration that is characterized by the growth of abnormal blood vessels into the subretinal space.

underway. In this Review we discuss the current state of ocular drug therapy, highlight innovations within drug discovery and delivery, and discuss potential concepts for the future treatment of retinal diseases and glaucoma.

Ocular diseasesAge-related macular degeneration. AMD is defined as an abnormality of the retinal pigment epithelium (RPE) that leads to degeneration of the overlying photorecep-tor in the macula and consequent loss of central vision7,8. AMD usually occurs in individuals over 50 years of age, and is the most common cause of vision loss in elderly individuals throughout the developed world9–15. AMD represents a major public health burden, with both eco-nomic and social consequences. It is estimated that over 9 million people in the United States suffer from inter-mediate or advanced forms of AMD16.

The aetiology and pathophysiology of AMD are poorly understood. Histologically, the disease is char-acterized by the accumulation of membranous debris underneath the RPE basement membrane, which forms drusen. Drusen are small, yellowish extracellular deposits of lipid, cellular debris and protein, including comple-ment components, anaphylatoxins, modulators12,17–20,21–26 and high-temperature requirement A serine peptidase 1 (HTRA1)27. The formation of drusen can be caused by RPE dysfunction or by a change in the composition or permeability (to nutrients) of Bruch’s membrane.

Therefore, early AMD is characterized by drusen (FIG. 2c) and hyper- or hypopigmentation of the RPE without loss of vision. Advanced AMD leads to loss of vision and can be classified into two categories: geographic atrophy, which is characterized by atrophy of the RPE (FIG. 2d); and choroidal neovascularization (CNV; also known as wet AMD) (FIG. 2e), which is due to the abnor-mal growth of blood vessels.

It has been suggested that one or more of the follow-ing key processes is involved in AMD (FIG. 3): oxidative damage; lipofuscin accumulation and impaired activity

or function of the RPE; increased apoptosis; abnormal immune system activation28; senescent loss of homeo-static control29; or abnormalities in Bruch’s membrane. Recent evidence suggests that there may be crosstalk and interaction among these pathways30. For example, oxidative stress, abnormal complement activation or inflammatory pathways may lead to RPE apoptosis or abnormal angiogenesis26,30,31.

Vascular endothelial growth factor A (VEGFA) has been implicated in CNV and increased vascular permea-bility that results in loss of vision32. VEGFA is a 46 kDa homodimeric glycoprotein that was first identified in vascularized tumours32–34. It is produced by many ocular cell types in response to hypoxia and has multiple func-tions in the eye. It stimulates endothelial cell growth, pro-motes vascular permeability and induces dissociation of tight junction components.

Animal models of AMD have contributed to the under-standing of disease pathogenesis and the development of treatments. Current models with features of dry AMD include monkeys with naturally occurring drusen, mice that have been genetically manipulated or have had high exposure to environmental risk factors for AMD (for example, a high-fat diet, exposure to cigarette smoke, and so on), and mice that have been immunized with carboxyethylpyrrole-modified mouse serum albumin35. Animal models of wet AMD include primates and rodents with laser-induced CNV, animals with mechani-cally induced CNV, rats that have been transfected with angiogenic factors as well as transgenic and knockout mice. Genetically modified mice are the only animal models developed to date that recapitulate the features of both dry and wet AMD35–40.

Ranibizumab (Lucentis; Genentech/Roche), beva-cizumab (Avastin; Genentech/Roche) and aflibercept (Eylea; Regeneron Pharmaceuticals) — all anti-VEGF agents — are currently the most common therapies for neovascular AMD. Other less commonly used treat-ments include photodynamic therapy, pegaptanib (Macugen; Eyetech) and retinal laser photocoagulation. Prophylactic treatments include oral vitamins and anti-oxidants (comprising vitamin C, vitamin E, β-carotene, zinc and copper), and are used in patients who are at a high risk of developing advanced AMD41.

Diabetic retinopathy and retinal vein-occlusive diseases. Diabetic retinopathy is the leading cause of blindness and visual impairment in working-age individuals. In the United States, 4.1 million individuals have diabetic retinopathy and, of these, approximately 900,000 have sight-threatening retinopathy42. These rates are expected to double by 2025. Retinal vein-occlusive diseases include central retinal vein occlusions, hemiretinal vein occlusions and branch retinal vein occlusions. Retinal vein-occlusive diseases affect approximately 100,000 individuals in the United States.

Diabetic retinopathy and retinal vein-occlusive dis-eases result in loss of vision through the following major mechanisms: retinal vascular leakage and exudation, which results in macular oedema; and retinal ischaemia and secondary neovascularization, which can lead to

Figure 1 | Human eye structure and diseases. Schematic diagram of the anatomy of the eye and associated diseases. AMD, age-related macular degeneration; DME, diabetic macular oedema; DR, diabetic retinopathy; RD, retinal degeneration.

R E V I E W S

542 | JULY 2012 | VOLUME 11 www.nature.com/reviews/drugdisc


Intraocular pressure(IOP). The fluid pressure inside the eye, which is determined by the production and

drainage of aqueous humour.

Aqueous humourA clear, watery fluid produced by the ciliary epithelium that fills the anterior and posterior chambers of the eye.

vitreous haemorrhage and retinal detachment. VEGFA has been implicated in increasing retinal vascular per-meability and leakage, which leads to macular oedema and ocular neovascularization34,43. Current treatments for diabetic macular oedema or macular oedema arising from retinal vein-occlusive diseases include anti-VEGF therapies, focal laser therapy and steroids. Laser pan-retinal photocoagulation is the main therapy for neo-vascularization in proliferative diabetic retinopathy and retinal vein-occlusive diseases.

Glaucoma. Glaucomas are a group of progressive optic neuropathies that are characterized by a slow and pro-gressive degeneration of retinal ganglion cells (RGCs) and their axons, resulting in a distinct appearance of the optic disc (FIG. 2f) and a concomitant pattern of vision loss6. Characteristic changes to the optic nerve in glau-coma include the enlargement and elongation of the optic nerve cup, the thinning and eventual notching of the neuroretinal rim, asymmetry in cup size between the two eyes and disc haemorrhages. Changes in the visual field with perimetric testing include scotomas that corre-spond to local and/or diffuse loss of nerve fibres within the retina. These characteristic changes consist of arcuate sco-tomas, nasal steps and paracentral scotomas. The disease involves the entire visual pathway, including the brainstem and the visual cortex — not just the eye44.

It has been estimated that glaucoma will affect more than 80 million individuals worldwide by 2020, with at least 6–8 million individuals becoming bilaterally blind45. Glaucoma is also the leading global cause of irre-versible blindness46 and is perhaps the most prevalent of all neurodegenerative diseases.

The biological basis of glaucoma is not fully under-stood, and the factors contributing to its progression are currently not well characterized. Intraocular pressure (IOP) is the only proven treatable risk factor, and lower-ing IOP is the main approach for reducing disease pro-gression. Impaired ocular blood flow is also thought to be a risk factor47 but the evidence for this is conflicting, perhaps largely owing to the absence of an accurate and reproducible test of ocular blood flow or the optic nerve microcirculation. With or without treatment, glaucoma can progress through a continuum48 that is initiated by an acceleration of RGC apoptosis, with the disease initially being asymptomatic but eventually resulting in blindness in some individuals, particularly those who are inadequately treated6.

Current status of ocular drugsAnti-VEGF therapy. Over the past decade, considerable progress has been made in the treatment of ocular neo-vascular diseases owing to an increased understanding of the mechanisms of ocular angiogenesis32,33 (FIG. 4). Anti-angiogenic therapies targeting VEGFA have proven to be highly effective in treating neovascular AMD and have become the mainstay of treatment. Anti-VEGF treatment is the only therapy to date that improves vision in patients with neovascular ocular diseases, as dem-onstrated by several randomized, controlled Phase III studies49–51.

Ranibizumab, a recombinant humanized antibody fragment that binds all known isoforms of human VEGFA, is formulated for ocular administration (via an intravitreal injection) and approved in the United States for the treatment of neovascular AMD. Bevacizumab, which was initially approved for the treatment of cer-tain types of cancers, has also been used (off-label) to treat wet AMD52 but it is not formulated for intraocular administration. A recent study compared the safety and efficacy of these two drugs in a multicentre, rand-omized controlled trial51. The primary results of the 1-year follow-up study showed that monthly administration of bevacizumab is non-inferior to monthly administration of ranibizumab. However, there were some safety con-cerns in the trial; a statistically higher number of serious adverse events was observed in the bevacizumab group51 (mainly hospitalizations, perhaps owing to longer systemic retention of bevacizumab compared to ranibizumab).

Another therapeutic approach is the use of aflibercept, which is a fusion protein that incorporates portions of extracellular domains of the human VEGF receptor 1 (VEGFR1) and VEGFR2 that are fused to the constant region of human immunoglobulin G53,54. Aflibercept binds to all isoforms of VEGFA with a higher affinity than ranibizumab and bevacizumab. It also binds placen-tal growth factor 1 and placental growth factor 2 (REF. 55). Two Phase III trials using aflibercept (as an intravitreal injection) to treat wet AMD have been completed and showed equivalent efficacy to ranibizumab. The drug was well tolerated and approved by the US Food and Drug Administration (FDA) for the treatment of wet AMD in 2011. The application of anti-VEGF agents in the treatment of retinal vascular diseases, including diabetic retinopathy and retinal vein occlusions, has been very successful56–58 but ranibizumab has only been approved by the FDA for treating retinal vein-occlusive diseases.

IOP-lowering therapies. There are two major forms of glaucoma: open-angle glaucoma and closed-angle glau-coma. Both forms are initially treated by lowering IOP, typically with eye drops, but rapid advancement to surgi-cal treatment is required for some types of closed-angle glaucoma.

IOP is determined by the balance between the aqueous humour secreted into the eye by the ciliary body and the amount that is drained from the eye via the two major outflow pathways: the trabecular meshwork and the uveo scleral pathway (consisting of the iris root, longitu-dinal ciliary muscle, anterior choroid and sclera). IOP is in a steady state when the rate of aqueous inflow is the same as the rate of aqueous outflow.

Pharmacological treatment of glaucoma lowers IOP by decreasing the rate of aqueous inflow and/or increas-ing the rate of aqueous outflow (FIG. 5a). There are five major classes of drugs (administered as eye drops) that are approved for lowering IOP in glaucoma (TABLE 2). Cholinergic agents have been used for more than 100 years to lower IOP. They act on muscarinic recep-tors located on the ciliary muscle to increase outflow through the trabecular meshwork59. Although the most

R E V I E W S



Fovea

Fovea

Vitreous

Retina

Choroid


d

f

a

e

RPE

Bruch’s membrane

Choroid

Horizontal cell

Amacrine cell

Vitreous

Bipolar cell

Ganglion cell

RodCone

RetinaRPE

Choroid

RetinaRPE

Choroid

c

b

Fovea

ON

Macula

R E V I E W S



Complement systemA part of the innate immune system that consists of approximately 25 proteins. Three pathways activate the complement system: the classical complement pathway, the alternative complement pathway and the mannose-binding pathway.

widely used drug in this class — pilocarpine — is inex-pensive and has few systemic side effects, its widespread use is largely limited by ocular side effects (such as mio-sis, myopia, brow ache and dimming of vision) and the inconvenience of requiring dosing four times daily59,60.

Owing to their convenient once-daily dosing, pros-taglandin F receptor (PTGFR) analogues are generally administered as first-line agents and increase aqueous outflow by altering the composition of the extracell-ular matrix in the ciliary muscle and trabecular mesh-work60,61. These agents are the most effective of the IOP-lowering agents, and a single daily dose is effec-tive at reducing IOP throughout the day and during the night62. PTGFR analogues cause lengthening and thick-ening of eyelashes and can also cause hazel and blue irides with brown patches to become diffusely brown.

β-adrenergic receptor blockers have been widely used for more than 30 years and have various strengths, including their ocular tolerability and IOP-lowering effi-cacy (which is surpassed only during the day by PTGFR analogues). They reduce aqueous humour secretion by inhibiting the activity of the predominant β-adrenergic receptor in the ciliary epithelium63. However, they are only effective during the day and not during the night62, and can reduce ocular perfusion pressure. They have minimal efficacy in combination with PTGFR ana-logues, and for this reason the use of these agents in a combination product has not been approved in the United States.

Another approach is the use of carbonic anhydrase inhibitors to inhibit the activity of carbonic anhydrase 2 in the non-pigmented ciliary epithelium, thus reduc-ing aqueous humour formation. These drugs lower

IOP throughout the day and during the night, and are widely used as second-line agents64. α-adrenergic recep-tor agonists lower IOP primarily through stimulation of α2-adrenergic receptors in the eye. Like the beta block-ers, they effectively lower IOP during the day but not during the night65. They appear to decrease aqueous inflow and increase uveoscleral outflow66. In experimen-tal animal models they have a neuroprotective effect on RGCs67 but this has not been demonstrated convincingly in patients with glaucoma.

Ongoing clinical trialsComplement pathway inhibitors. Recent studies have shown that AMD is associated with certain genotypic variants of complement pathways, including comple-ment factor H (CFH)21,68–70, complement component 2 (C2)71, complement factor B71,72, complement compo-nent 3 (C3)72,73 and complement factor I74. Owing to multiple gene associations in the complement activa-tion pathway, several complement inhibitors are now in clinical trials. Various strategies have been exploited to treat AMD by modulating the complement system, such as the replacement of defective complement components as well as inhibition of the complement pathway (FIG. 6). Inhibition of the complement pathway disables down-stream complement cascade activation, thus hindering inflammatory responses. However, the complement system is an important component of the immune sys-tem, and local or systemic complement inhibition may increase the risk of infections such as endophthalmitis.

POT-4 (also known as AL-78898A) is a synthetic peptide C3 inhibitor that binds reversibly to C3. An appealing feature of this intravitreal drug is its gel-like preparation, which can be deposited in the vitreous at high concentrations and can last for 6 months as a sustained-delivery vesicle. A Phase I study of POT-4 in patients with wet AMD was completed successfully with-out any safety concerns74. A Phase II study of AL-78898A in conjunction with ranibizumab is currently underway.

ARC1905 is a complement component 5 (C5) inhibi-tor that is administered by intravitreal injection. As a small-molecule aptamer-based drug it exhibits several advantages, including low activation of the immune response and the ability to formulate it in a sustained-release form. A Phase I trial investigating the efficacy of ARC1905 in combination with ranibizumab for treating wet AMD was performed and the agent was shown to be safe and well tolerated75. Another Phase I study investi-gating the efficacy of ARC1905 in dry AMD is underway.

Eculizumab is a monoclonal antibody derived from a murine anti-human C5-specific antibody. Like ARC1905, it is also intravenously administered and inhibits the cleav-age of C5 to C5a and C5b. Eculizumab is already approved by the FDA for the treatment of paroxysmal nocturnal haemoglobinuria and is currently in Phase II trials known as COMPLETE (‘complement inhibition with eculizumab for the treatment of non-exudative age-related macular degeneration’) for the treatment of dry AMD.

FCFD4514S is a recombinant, humanized monoclo-nal antibody Fab fragment directed against complement factor D, currently in a Phase I/II study to evaluate its

Figure 2 | Retinal anatomy and structure in health and diseases. Imaging of the structures of an eye by optical coherence tomography. a | A schematic diagram of the retina is shown at the top of the panel. The inset shows the choroid and layers of the retina, including the retinal pigment epithelium (RPE). Optical coherence tomography images are also shown, of a normal retina (top) and a retina with pigmented epithelium detachment due to choroidal neovascularization (CNV) (indicated by the white arrows) in wet age-related macular degeneration (AMD) (bottom). b | The image shown is of a normal retina and macula. c | The image depicts a macula with confluent soft drusen (indicated by the white arrows) — a hallmark of early AMD. d | The image shows a macula of geographic atrophy (within the dashed circle). e | A macula of CNV with haemorrhage (indicated by the white arrows) is shown. f | The image illustrates an optic nerve with glaucomatous excavation and loss of neuroretinal rim between the disc margin (white dashed lines) and the border of the optic cup (black dashed lines). Images courtesy of K.Z. ON, optic nerve.

Table 1 | Leading causes of irreversible visual impairment

Disease Incidence Refs

Cataract 8.4–29.7% of patients over 43 years of age 183

Age-related macular degeneration

6–22% of patients over 70 years of age 184

Glaucoma 1–4% of patients over 45 years of age 6

Diabetic retinopathy 74.9–92.3% of diabetic patients over 30 years of age

185,186

Retinitis pigmentosa 1 in 4,000 187▶

R E V I E W S




Abnormal complement activity

Chronic inflammation

Photoreceptor–RPE–Bruch’schoriocapillaris complex

Chronic oxidative stress

++

+

+Lipofuscin

Antioxidants

Neuroprotectants

Anti-angiogenic agents

Accumulation of extracellular and intracellular debris

Drusen formation, damage to photoreceptors and RPE

Visual cycleinhibitors

Anti-inflammatoryagents

Geographic atrophy Neovascularization


Abnormal complement activity

Chronic inflammation

Photoreceptor–RPE–Bruch’schoriocapillaris complex

Chronic oxidative stress

++

+

+Lipofuscin

Antioxidants

Neuroprotectants

Anti-angiogenic agents

Accumulation of extracellular and intracellular debris

Drusen formation, damage to photoreceptors and RPE

Visual cycleinhibitors

Anti-inflammatoryagents

Geographic atrophy Neovascularization

Small interfering RNA(siRNA). Small, double-stranded RNA molecules that interfere with gene expression by binding to and promoting the degradation of mRNA.

safety, tolerability, pharmacokinetics and immuno-genicity when intravitreally injected in the treatment of geographic atrophy.

Visual cycle inhibitors. A prominent feature of AMD is the aberrant accumulation of cellular debris or lipofuscin within the RPE. Lipofuscin is a product of autofluores-cent material composed of lipids, proteins and vitamin A derivatives76–78. The rationale for the use of visual cycle modulators is to reduce the accumulation of fluoro-phores (for example, A2E and lipofuscin) in RPE cells (FIG. 3). Retinol is essential for vision, and its metabolism is mediated through the visual cycle (see the Webvision website). Retinol binding protein (RBP) binds to all-trans-retinol with a high affinity. The retinol–RBP complex serves as a substrate for the uptake of retinol by the RPE within the eye. Two visual cycle inhibitors are currently in clinical trials.

N-(4-hydroxyphenyl)-retinamide (Fenretinide; Sirion Therapeutics) is an oral synthetic retinoid derivative that binds to RBP in the circulation and blocks its association with retinol, thereby preventing the transport of retinol to the RPE and lowering its availability in the visual cycle. Fenretinide reduces lipofuscin and A2E accumu-lation in the RPE of mice in which ATP-binding cas-sette subfamily A member 4 (ABCA4) is knocked out (ABCA4-knockout mice) and causes modest delays in dark adaptation79. A Phase II clinical trial of this oral agent has been completed, and the drug has been shown to slow the progression of geographic atrophy and reduce the incidence of CNV. Potential side effects include night blindness and dry eye.

ACU-4429 is another orally administered visual cycle modulator. It inhibits the conversion of all-trans-retinyl ester to 11-cis-retinol via inhibition of the isomerase retinoid isomerohydrolase (RPE65). It was shown to slow retinal degeneration in preclinical models. However, side effects of this drug include nyctalopia, dyschroma-topsia and delayed dark adaptation. By modulating isomerization, ACU-4429 slows the visual cycle in rod photoreceptors and decreases the accumulation of A2E. A Phase II study of ACU-4429 is currently underway for the treatment of patients with AMD who have geographic atrophy.

Anti-angiogenic agents. Because ocular angiogenesis is a multistep process (FIG. 4), it can be intercepted or inhib-ited at many steps using various approaches, and several agents are now under investigation (TABLE 3). The various strategies and related agents are discussed below.

One strategy is to inhibit mammalian target of rapa-mycin (mTOR), which is a tyrosine kinase that has a key role in regulating cell growth and proliferation. Its activation leads to the production of hypoxia-inducible factors (for example, HIF1α), which in turn can induce the expression of the angiogenesis stimulator VEGF80. Examples of mTOR inhibitors include rapamycin (also known as sirolimus), everolimus and Palomid 529. Sirolimus is currently in Phase II clinical trials for the treatment of patients with diabetic macular oedema, and everolimus in combination with ranibizumab is in Phase II trials in patients with wet AMD.

Small interfering RNA (siRNA) is another approach that is used to inhibit VEGF expression. PF-655 is a synthetic siRNA that inhibits the expression of DNA damage-inducible transcript 4 protein (DDIT4; also known as RTP801), which has been shown to be involved in pathological retinal neovascularization in animal models81. It is currently in Phase II trials for diabetic macular oedema and wet AMD. Bevasiranib is a naked, 21-nucleotide-long siRNA that specifically targets VEGF. However, clinical trials of bevasiranib were discontinued because they did not meet their end point (stabiliza-tion of vision), which was defined as a loss of less than three lines of vision on the ETDRS (Early Treatment for Diabetic Retinopathy Study) vision chart. However, in hindsight, the failure of these trials is not surprising as siRNAs need to be formulated to achieve cell permea-tion in order to cause bona fide RNA interference. They

Figure 3 | Proposed pathophysiology of AMD, and locations in the pathway in which different therapeutic interventions might be effective. One or more of the following key processes is likely to have a role in age-related macular degeneration (AMD): oxidative damage; lipofuscin accumulation and impaired function of retinal pigment epithelium (RPE); increased cell death; abnormal immune system activation; senescent loss of homeostatic control; and abnormalities in Bruch’s membrane. VEGF, vascular endothelial growth factor. Potential strategies for the treatment of AMD are shown in orange boxes.

R E V I E W S



http://webvision.med.utah.edu/

http://webvision.med.utah.edu/


Smoothmuscle cell

Release

Binding toendothelialcell receptor

Angiogenicfactorproduction

PEDF

• JSM6427• Volociximab

Intracellularsignalling

Blood vessel

Endothelialcell

Endothelial cell proliferation

Directionalmigration

Tubeformation

α5β1 integrin

PazopanibVascularstabilizationE10030

Sirna-027

Fosbretabulin

Sonepcizumab

• Sirolimus• RAD001• Palomid 529• PF4523665

• Aflibercept• Ranibizumab• Bevacizumab

also need to be modified to avoid off-target effects and recognition of their nucleotide structure by the innate immune system82–84.

In contrast to VEGF, pigment epithelium-derived factor (PEDF) has an antagonistic effect on endothe-lial cell proliferation and migration, and its levels are decreased in AMD85–87. A Phase I clinical trial has been initiated using an adenoviral vector (AdGVPEDF.11D) to deliver PEDF via intravitreal injections. Many agents, such as ranibizumab or bevacizumab, have now been developed to inhibit the binding of VEGF to its receptor. Aflibercept is another example of an agent that prevents receptor binding by trapping VEGF in the extracellu-lar space. AAV2-sFLT01 is an adeno-associated virus (AAV)88 that produces a soluble VEGFR1 that can bind to free VEGF89,90 to interrupt VEGF signalling.

VEGFA binds primarily to VEGFR1 and VEGFR2, both of which are tyrosine kinases91. A siRNA (AGN211745) designed to inhibit VEGFR1 synthesis has been tested in a Phase II trial, but this was terminated presumably because AGN211745 was not formulated for cell permeation. Another approach that is currently undergoing testing involves the blockade of chemokine CC motif recep-tor 3, which mediates angiogenic signalling initiated by eotaxins92.

VEGF binds and activates its receptors VEGFR1, VEGFR2 and VEGFR3. Phosphorylation of the tyrosine

residue of the kinase domain of VEGFRs is crucial for receptor activation. Small-molecule tyrosine kinase inhib-itors therefore have considerable potential in targeting the kinase domain and blocking the action of VEGFR1, VEGFR2 and VEGFR3. Such inhibitors are usually also active against platelet-derived growth factor receptor (PDGFR), KIT and fibroblast growth factor receptor 1 (FGFR1). Some non-specific tyrosine kinase inhibition and cross-inhibition may be beneficial. As these growth factors are also involved in ocular angiogenesis, they can potentially provide synergistic effects on CNV inhibition.

Pazopanib is a tyrosine kinase inhibitor that inhibits VEGFR1, VEGFR2, VEGFR3, PDGFR, KIT and FGFR1. Pazopanib is effective in preclinical models of CNV93. It is administered topically and has shown promising results in a Phase II trial for CNV94. Vatalanib is an oral tyrosine kinase inhibitor that has been tested previously as a treatment for CNV in Phase I and Phase II clinical studies. AL39324 is an intravitreally administered tyro-sine kinase inhibitor that is in a Phase II clinical trial in combination with ranibizumab.

TG100801 is a tyrosine kinase inhibitor that binds and inhibits VEGFR and PDGFR. It is administered as an eye drop and has been found to suppress CNV and retinal oedema95. A Phase I trial was completed success-fully but a Phase II trial was discontinued because of corneal toxicity.

Figure 4 | Mechanism of action of ocular angiogenesis inhibitors in clinical care or development. Schematic diagram of new capillary blood vessel growth. Several classes of angiogenic factors, including vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF) and pigment epithelium-derived factor (PEDF), bind to specific receptors on vascular endothelial cells to induce specific signalling pathways that activate the cells. Angiogenesis can be inhibited by blocking these signalling pathways as well as through various other approaches. Tyrosine kinase inhibitors (sirolimus, RAD001, Palomid 529 and PF4523665) prevent the production of angiogenic factors. Bevacizumab, ranibizumab and aflibercept bind to and sequester VEGF, thereby inhibiting its interaction with VEGF receptors. Sirna-027 inhibits the synthesis of VEGF receptor 1. JSM6427 and volociximab both bind to α5β1 integrin and block the migration of endothelial cells. The active metabolite of fosbretabulin (combretastatin A4) inhibits microtubule polymerization in proliferating endothelial cells, inducing vessel regression.

R E V I E W S



Sclera

Cornea

Ciliary muscleCiliary processes

Iris

Anterior chamber

Lens

Trabecular meshwork outflow

• Cholinergic agents

• Latrunculins• ROCK inhibitors

Uveoscleral outflow

• Prostaglandin analogues• α-adrenergic receptor agonists

• Prostaglandin EP2 agonists• Nitric oxide-donating prostaglandin F2α analogue • Drug-eluting punctal plug with latanoprost

Inflow

• β-adrenergic receptor blockers• α-adrenergic receptor agonists• Carbonic anhydrase inhibitors

• siRNA β-adrenergic receptor antagonist


Current treatmentsInvestigational drugs

An alternative approach to inhibiting the action of VEGF involves preventing the activation of the nicotinic acetylcholine receptor (nAChR) pathway in the vascu-lature, which inhibits VEGF-induced angiogenesis in endothelial cells96. ATG-3 is an eye drop formulation of mecamylamine, an antagonist of the nAChR pathway97. It is the first anti-angiogenic eye drop that has been stud-ied in humans. ATG-3 has successfully completed two Phase II clinical trials, one in patients with diabetic mac-ular oedema98 and the other in patients with wet AMD who were receiving anti-VEGF treatments.

Sphingosine-1-phosphate (S1P) is the extracellu-lar ligand for S1P receptor 1 (also known as EDG1), a G protein-coupled lysophospholipid receptor, and it is a pro-angiogenic and profibrotic mediator99,100. Intra-vitreous injection of an anti-S1P monoclonal antibody inhibited CNV formation and subretinal collagen deposi-tion in a preclinical model of laser-induced CNV and in a preclinical model of diabetic retinopathy101,102. A human-ized S1P monoclonal antibody (iSONEP) has completed a Phase I clinical trial in patients with wet AMD. iSONEP was well tolerated, and no drug-related serious adverse events were observed in any of the patients. iSONEP also exhibited positive biological effects (including lesion regression, reduction of retinal thickness and resolution of pigment epithelium detachment). The mechanisms responsible for the positive effects exerted by S1P appear to be independent to those of anti-VEGF agents, indicat-ing the potential of S1P antibodies to serve as a mono-therapy (or an adjunct therapy to anti-VEGF agents) for the treatment of neovascular AMD.

Endothelial cell migration involves interactions between integrins (transmembrane heterodimeric pro-teins) and extracellular matrix ligands. For example, α5β1 integrin is expressed on the surface of vascular endothelial cells and mediates cell migration103. JSM6427 and volociximab both bind to α5β1 integrin and block the migration of endothelial cells. Both agents have been shown to inhibit CNV formation in preclinical models104,105.

Fosbretabulin is a novel vascular disrupting agent, and its active metabolite is combretastatin A4 (CA4)106. CA4 binds to tubulin and prevents microtubule poly-merization in proliferating endothelial cells, thus induc-ing vessel regression107. In animal models, CA4 has been demonstrated to inhibit retinal neovascularization and suppress the development of CNV108,109. Phase II trials are underway to evaluate the safety and efficacy of fos-bretabulin in patients with CNV and vasculopathy.

Pericyte recruitment is a crucial step in vascular maturation and stabilization. PDGF has a key role in this process110,111. With this in mind, E10030 — a pegylated aptamer that inhibits PDGF-β — was developed. Inhibition of PDGF results in the stripping of pericytes from endothe-lial cells, which increases the sensitivity of mature CNV to VEGF inhibition. PDGF inhibition may therefore syn-ergize with VEGF inhibition in CNV treatment. Indeed, in a Phase I dose-escalating study, some CNV regression was observed in over 90% of patients receiving the com-bination therapy of E10030 and ranibizumab, compared to about 10% of patients receiving ranibizumab alone112. A Phase II study is currently underway.

Pharmacologic vitreolysis. Pharmacological interven-tions for retinal diseases, such as macular oedema and macular holes, that are associated with the vitreoretinal interface (VRI) may soon include one for pharmacologic vitreolysis. The goals of an intravitreal pharmacologic vit-reolysis agent are to cleanly cleave the VRI at the internal limiting membrane, removing the potential for fibro-cellular traction, and to liquefy the vitreous by altering its molecular organization and structure203.

Ocriplasmin (ThromboGenics, Belgium) is currently being investigated as a pharmacologic vitreolysis agent. A stable, truncated form of human plasmin retaining pro-teolytic activity, ocriplasmin (molecular weight 27.2 kDa) is manufactured using recombinant DNA technology204. Preclinical studies suggest that ocriplasmin targets sub-strates that are important to both the vitreous structure and the VRI, including collagen fibrils, fibronectin and laminin204. Ocriplasmin was recently studied in two Phase III pivotal clinical trials as a single intravitreal injection for treatment of patients with symptomatic vitreomacular adhesion (VMA), including macular holes. Results from both trials indicated that treatment with ocriplasmin met the primary end point of VMA reso-lution as well as secondary end points, including visual acuity improvement and closure of full-thickness macu-lar holes in a subset of patients with VMA when com-pared to placebo. Most adverse events were considered mild and were either transient or temporary in nature, with a majority occurring within the first 7 days after

Figure 5 | Agents that lower intraocular pressure. Current and investigational drugs for lowering intraocular pressure, decreasing aqueous inflow and/or increasing aqueous outflow are indicated on the figure. Aqueous humour is secreted into the posterior chamber by the ciliary processes and eventually circulates into the anterior chamber angle, where it drains from the eye via one or both outflow pathways: the trabecular meshwork or the uveoscleral outflow route. ROCK, RHO-associated protein kinase.

R E V I E W S



treatment205. Ocriplasmin is currently under review by the FDA. Other indications may include: vitreous liquefaction and lysis to facilitate complex surgical cases such as paedi-atric vitrectomy, proliferative vitreoretinopathy and prolif-erative diabetic retinopathy; relief of vitreous traction on retinal breaks in pneumatic retinopexy; and treatment of macular oedema due to vitreomacular traction.

Novel IOP-lowering drugs. In addition to the five major drug classes currently being used to lower IOP (FIG. 5), several novel drugs are being evaluated in clinical trials (FIG. 5; TABLE 4) but results of these studies have not been published. Approved ocular hypotensive drugs with new delivery systems, including drug-eluting punctal plugs (for example, with a prostaglandin analogue), are also being investigated for sustained release and to improve patient adherence. Existing drug classes can be separated into those that lower habitual IOP throughout the day

and during the night when administered topically (such as prostaglandin analogues62,113 and carbonic anhydrase inhibitors64), and those that are effective only during the day and not during the nocturnal period (such as beta blockers64 and α-adrenergic receptor agonists65). The relationship of IOP lowering (throughout the day and during the night) to glaucoma progression is unknown and under investigation114.

The latrunculins are macrolides from marine sponges that inhibit actin polymerization115. Studies in non-human primates and post-mortem analyses of eyes from patients have demonstrated that latrunculins increase trabecular meshwork outflow via a novel mechanism that disrupts the actin cytoskeleton in the trabecular mesh-work116. However, their performance in clinical trials has so far been marginal and they have poor solubility. It has been suggested that their efficacy may be improved by changes in their delivery system117.

Table 2 | Available classes of IOP-lowering eye drops for glaucoma

Class Drugs Mechanisms of action Notes

Aqueous humour outflow (conventional pathway)

Cholinergic drugs Pilocarpine, carbachol

•Increase trabecular meshwork outflow through ciliary muscle contraction59

•Highly effective but their use is compromised by dim vision (from small pupil) and discomfort59,60

Aqueous humour outflow (uveoscleral pathway)

Prostaglandins PGF2α analogues : latanoprost, travoprost, bimatoprost and tafluprost

•Increase outflow by increasing matrix metalloproteinase expression and altering the extracellular matrix in the ciliary muscle and trabecular meshwork61,192

•Highly effective and well tolerated: generally used as first-line therapy62

Aqueous humour formation

β-adrenergic receptor blockers

Timolol, betaxol, carteolol and levobunolol

•Decrease inflow by regulating aqueous humour formation in the ciliary processes63

•Few ocular side effects but occasional systemic effects include fatigue and bradycardia62

α-adrenergic receptor agonists

Apraclonidine and brimonidine

•Decrease inflow by inactivating adenylyl cyclase in ciliary processes and mediating noradrenaline release via activation of pre-synaptic α2-adrenergic receptors; may also increase uveoscleral outflow65

•Sedation is minimized by occluding nasolacrimal tear duct for up to 2 minutes

•These drugs have also been suggested to have neuroprotective effects67

•Allergy is not uncommon, particularly with the 0.2% formulation

Carbonic anhydrase inhibitors

Dorzolamide, brinzolamide, acetazolamide* and methazolamide*

•Decrease aqueous humour formation by inhibiting carbonic anhydrase and HCO

3 production in the

ciliary epithelium64

•Eye drops are better tolerated than orally administered drugs193

Combination of existing classes of IOP-lowering drugs

β-adrenergic receptor blockers plus carbonic anhydrase inhibitors

Timolol and brinzolamide

Decrease inflow196 –

β-adrenergic receptor blockers plus α-adrenergic receptor agonists

Timolol and brimonidine

Decrease inflow and increase outflow197

•Combination product has less hyperaemia than brimonidine alone

β-adrenergic receptor blockers plus prostaglandins

Timolol and PGF2α analogues

Decrease inflow and increase outflow198–202

•Not available in the United States

IOP, intraocular pressure; PGF2α, prostaglandin F2α receptor. *Available as oral medications.

R E V I E W S




C3 convertase (C4b2a)

C5 convertase

C3b C3a

C3 (H2O)

CFB

C6, C7, C8 and C9

C5bC5a

C5

Terminal membrane attack complex

CFH CFI+

CFI+C2

C3+ C4

C1

C1 complex C3 (H2O)-B

IgM, IgG immunecomplexes or CRP

Classical pathway Lectin pathway Alternative pathway

Mannose-bindinglectin complex

Endogenous orforeign pathogens

RHO-associated protein kinase (ROCK) is a down-stream effector of RHO (a small guanine triphosphatase) in the RHO-dependent signal transduction pathway. ROCK inhibitors induce changes in the actin cytoskel-eton and the cellular motility of the outflow pathways, and they can also lower IOP118. Several drugs in this class have been tested in clinical trials. However, it is still unknown whether the efficacy of these agents will outweigh their side effects, so the use of this class of compounds may be limited by conjunctival hyperaemia and subconjunctival haemorrhages119,120. One strategy for addressing these side effects is to use a pro-drug that is converted into a more active compound once it passes through the cornea and is present in the anterior cham-ber121. Such an example is ATS907; the pro-drug form of ATS907 is less active and highly permeable across the cornea. After it passes through the cornea, ATS907 is then converted into a more active ROCK inhibitor in the aqueous humour of the anterior chamber. ATS907 is currently in Phase I/II clinical trials.

Other ROCK inhibitors that are currently in Phase II glaucoma trials include AR-12286 and K-115. AR-12286 is currently in a Phase II clinical trial to evaluate its 24-hour efficacy in patients with open-angle glaucoma or ocular hypertension. K-115 has completed two Phase II clinical trials, which demonstrated the ability of the compound to lower IOP in patients with primary open-angle glaucoma or hypertension122,123. Amakem

also has a ROCK inhibitor (AMA0076) in preclinical development. It is currently unknown whether ROCK inhibitors will be effective when used only once a day or whether they will require administration at least twice a day. In addition, it has not yet been determined whether ROCK inhibitors will be effective at lowering IOP dur-ing the night or whether they will be used as primary — rather than adjunctive — therapy.

Drugs in this class are also being experimentally evaluated for their neuroprotective potential124. The only ROCK inhibitor that is approved and clinically used is fasudil (Eril; Asahi Kasei). Fasudil is used in Japan for neuroprotection in subarachnoid haemorrhage. ROCK inhibitors can also be useful in trabeculectomy surgery to reduce fibrosis125. Another potential use of ROCK inhibitors is as anti-inflammatory agents, which may be beneficial at ameliorating ocular surface disease and inflammation.

Other classes of drugs are also being studied for their ability to lower IOP. Adenosine receptor agonists increase outflow by increasing trabecular meshwork outflow126. Several drug companies are pursuing strat-egies involving modulation of the adenosine receptor pathway by targeting different receptors. Examples of such drugs include ATL313 (an adenosine A2A receptor agonist), INO8875 (an adenosine A1 receptor agonist) and OPA-6566. However, these drugs may have limited use owing to desensitization of their receptors.

The angiotensin II type 1 receptor antagonist olmesar-tan is being tested in Japan for its ability to lower IOP by increasing outflow127. Cannabinoid receptor agonists128 are also being tested. The evidence for their use in glau-coma is based on the observation that smoking marijuana transiently lowers IOP129. In a non-human primate model of glaucoma, topical application of a cannabinoid receptor agonist that is selective for cannabinoid receptor 1 low-ered IOP by decreasing aqueous humour flow128. A can-nabinoid receptor-selective agonist that lowers IOP might be better tolerated and have a longer ocular hypotensive duration of action than marijuana.

SYL040012, a siRNA targeting the β2-adrenergic receptor, is currently in clinical trials to investigate its effect on IOP in patients with ocular hypertension130. PF-04217329 is a selective agonist of prostaglandin E receptor 2 that has completed Phase II clinical trials. The results showed that PF-04217329 significantly reduces IOP in patients with primary open-angle glau-coma and ocular hyper tension, but doses higher than 0.015% were associated with a higher incidence of photo-phobia and iritis131,132.

BOL-303259-X (latanoprostene bunod) is a nitric oxide-donating prostaglandin F2α analogue. A Phase II clinical trial is currently underway to compare the safety and efficacy of BOL-303259-X with latanoprost in patients with glaucoma or ocular hypertension. The L-PPDS (Latanoprost Punctal Plug Delivery System) study133 by QLT recently completed a Phase II clinical trial and latanoprost was found to be well tolerated; adverse events observed were similar to those reported for punctal plugs (for example, tearing). In addition, the L-PPDS was shown to significantly lower IOP133.

Figure 6 | Complement pathways, their association with AMD and drug targets. Schematic diagram of classical, alternative and lectin complement activation pathways. Circled proteins have corresponding gene variants that are associated with age-related macular degeneration (AMD). C1, complement component 1; CFB, complement factor B; CFH, complement factor H; CFI, complement factor I; CRP, C-reactive protein; IgG, immunoglobulin G.

R E V I E W S



Neuroprotective agents. Both retinal diseases and glaucoma are neurodegenerative disorders in which a common end point is the apoptosis of retinal neurons. Therefore, the use of agents that confer neuroprotection is a very appealing therapeutic approach. Stimulation of glutamate receptors, interleukin-1 receptors (IL-1Rs), JUN receptors and tumour necrosis factor receptors (TNFRs) triggers retinal neurons to undergo apoptosis through a cascade of cellular signalling events134. These signals include activation of the pro-apoptotic proteins BCL-2 antagonist of cell death (BAD), BH3-interacting domain death agonist (BID) and BCL-2-associated X protein (BAX), which then promote the release of cytochrome c and activate the caspase pathways.

The anti-apoptotic mechanisms can also be stimulated through neurotrophin signalling pathways, including

pathways involving brain-derived neurotrophic factor (BDNF), ciliary neurotrophic factor (CNTF) and glial cell-derived neurotrophic factor (GDNF), resulting in the inhibition of the pro-apoptotic cascade through signalling molecules such as AKT, B cell lymphoma 2 (BCL-2), BCL-XL as well as X-linked inhibitor of apop-tosis (XIAP)134.

To date, only a few clinical trials (TABLE 5) have been carried out to investigate the efficacy of neuroprotective agents in glaucoma (for example, memantine (Namenda; Forest/Lundbeck) and brimonidine (Alphagan; Allergan)) and in AMD (for example, brimonidine, AL-8309B, CNTF and RN6G). These are discussed in more detail below.

AL-8309B is a topical selective agonist of the 5-hydroxy -tryptamine 1A receptor (5-HT1A). 5‑HT1A agonists have been shown to have neuroprotective effects against

Table 3 | Anti-angiogenic drugs in clinical trials

Drug Company Description Target Clinical phase

ClinicalTrials.gov identifiers

Sirolimus (also known as rapamycin)

MacuSight/Santen

Tyrosine kinase inhibitor

mTOR Phase I/II NCT01445548

Phase I/II NCT00766649

Phase II NCT00656643

RAD-001 (everolimus)

Novartis Tyrosine kinase inhibitor

mTOR Phase II NCT00304954

Palomid 529 Paloma Tyrosine kinase inhibitor

mTOR Phase I NCT01033721

Phase I NCT01271270

PF-655 (also known as PF-04523655)

Pfizer Synthetic siRNA DDIT4 Phase II NCT01445899

AdGVPEDF.11D GenVec Adenovirus vector containing PEDF

Endothelial cells Phase I NCT00109499 (completed)

AAV2-sFLT01 Genzyme Adeno-associated virus (AAV) vector carrying VEGFR

VEGF Phase I NCT01024998

Pazopanib GlaxoSmithKline (United States)

Tyrosine kinase inhibitor

VEGFRs Phase II NCT01362348


PTK787 Novartis Oral tyrosine kinase inhibitor

VEGFRs Phase I NCT00138632 (completed)

AL-39324 Alcon Tyrosine kinase inhibitor

VEGFRs Phase II NCT00992563 (completed)

ATG-3 CoMentis Eye drop formulation of mecamylamine

Nicotinic acetylcholine receptor

Phase II NCT00536692 (completed)


iSONEP (sonepcizumab)

Lpath Monoclonal antibody

Sphingosine-1-phosphate


Phase I NCT01334255

JSM6427 Jerini Ophthalmic Monoclonal Antibody

α5β1 Integrin Phase I NCT00536016

Volociximab Ophthotech Corporation

Monoclonal Antibody

α5β1 Integrin Phase I NCT00782093

Fosbretabulin OXiGENE Combrestatin A4 phosphate

Microtubule assembly


E10030 Ophthotech Corporation

Pegylated aptamer Platelet-derived growth factor-β


DDIT4, DNA damage-inducible transcript 4 protein; mTOR, mammalian target of rapamycin; PEDF, pigment epithelium-derived factor; siRNA, small interfering RNA; VEGF, vascular endothelial growth factor; VEGFR, VEGF receptor.

R E V I E W S



excitotoxic neuronal damage in animal models of central nervous system injury135,136. However, the mechanisms under lying the neuroprotective properties of 5-HT1A ago-nists remain elusive. Several putative mechanisms that have been identified include inhibition of caspase 3 and activation of the mitogen-activated protein kinase (MAPK) signalling pathway137, which leads to increased expression of anti-apoptotic proteins (for example, XIAP, BCL-2 and BCL-XL)

138.

AL‑8309B exhibited neuroprotective effects in animal models of excitotoxic neuronal damage and light damage, in which it reduced the incidence of neuronal death139. A 2-year, multicentre, randomized, double-masked, placebo-controlled Phase III clinical trial (the Geographic Atrophy Treat ment Evaluation (GATE) trial) is currently underway to investigate the effects of topical AL-8309B administration (at doses of 1.0% and 1.75%) in AMD patients with geographic atrophy; the trial is expected

Table 4 | Other novel glaucoma drugs and possible mechanisms of action

Drugs Companies Mechanisms of action Clinical phase

Clinical trial identifiers

Refs

Latrunculins Wisconsin Alumni Research Fund (WARF)

Increase outflow by altering actin cytoskeleton and increasing trabecular meshwork outflow

Preclinical NA 116

SYL040012 Sylentis β-adrenergic receptor antagonist; small interfering RNA that reduces aqueous humour inflow

Phase II NCT01227291 130

PF-04217329 (also known as taprenepag)

Pfizer Prostaglandin E receptor 2 agonist; increases outflow

Phase II NCT00934089 (completed); NCT00572455 (completed)

194

BOL-303259-X NicOx (with Bausch & Lomb)

Nitric oxide-donating prostaglandin F2α analogue; increases outflow by increasing uveoscleral outflow


Latanoprost Punctal Plug Delivery System (L-PPDS)

QLT Sustained delivery of latanoprost by the plug increases outflow


INO-8875 Inotek Pharmaceuticals

Adenosine receptor agonist; increases outflow via trabecular meshwork

Phase I/II NCT01123785 126

OPA-6566 Acucela/Otsuka Adenosine receptor agonist; increases outflow via trabecular meshwork


ATL313 Santen Adenosine receptor agonist; increases outflow via trabecular meshwork

Preclinical NA 126

Angiotensin II type 1 receptor antagonists (olmesartin, candesartin and losartin)

Santen Increase outflow and may also have neuroprotective effects on retinal ganglion cells

Preclinical NA 127

Cannabinoid receptor agonist

Investigational Decrease aqueous humour outflow

Preclinical NA 128

ROCK inhibitors

AR-12286 Aerie ROCK inhibitor; alters actin cytoskeleton and cellular motility of outflow pathways



ATS907 Altheos ROCK inhibitor; alters actin cytoskeleton and cellular motility of outflow pathways


AMA0076 Amakem ROCK inhibitor; alters actin cytoskeleton and cellular motility of outflow pathways

Preclinical NA 118

K-115 Kowa ROCK inhibitor; alters actin cytoskeleton and cellular motility of outflow pathways

Phase II JapicCTI-101015 (completed)

118

Phase II JapicCTI-090708 (completed)

NA, not available; ROCK, RHO-associated protein kinase.

R E V I E W S



to be completed in 2012. The primary efficacy end point is the rate of progression to geographic atrophy.

Another promising approach is the use of brimon-idine tartrate (delivered through an intravitreal implant), which is an α2-adrenergic receptor agonist that was origi-nally designed to lower IOP in the treatment of glaucoma. Although the mechanism of action of the compound is largely unknown, brimonidine was shown to promote the production of neurotrophic factors (such as CNTF) and protect photoreceptors in animal models of retinal degeneration induced by light damage140. Brimonidine tartrate has been formulated as an intravitreal implant for delivering brimonidine to the retinal tissue over a period of 3 months. A Phase II study to evaluate the safety and efficacy of the compound is expected to be completed in 2012. The primary end point is the change from the base-line in the surface area of geographic atrophy after 1 year.

RN6G (also known as PF-4382923) is a humanized monoclonal antibody that is under development for the treatment of Alzheimer’s disease. It targets the carboxyl termini of amyloid-β40 and amyloid-β42. Intravenous treatment with RN6G is intended to prevent the accumu-lation of amyloid-β40 and amyloid-β42, and to prevent their cytotoxic effects141. Amyloid-β is present in the eyes of patients with AMD; systemic treatment with RN6G decreased the amount of amyloid-β in the eyes of a mouse model of AMD and prevented damage to the retina142. A Phase I clinical trial has been successfully completed.

CNTF, which protects photoreceptors from apop-tosis during retinal degeneration143, also has potential as a treatment for geographic atrophy. Although the intrinsic function of CNTF is not fully understood, exogenous CNTF affects the survival and differentia-tion of various cells in the nervous system, including retinal cells144,145. The CNTF study used an implanted

semipermeable capsule that encapsulates genetically modified RPE cells that overexpress CNTF. RPE cells are encapsulated within a semipermeable membrane with small pores, which allows the release of CNTF into the vitreous yet prevents the allogeneic rejection of RPE cells. CNTF effectively protected photoreceptors in 12 animal models of photoreceptor degeneration94,146–149. A multicentre, 1-year, double-masked, sham-controlled dose-ranging study found that CNTF delivered by the encapsulated cell technology (ECT) implant appears to slow the progression of vision loss in patients with geo-graphic atrophy, especially in patients with 20/63 vision or better vision on an eye chart at the baseline150.

Degeneration and functional loss of the glaucoma-tous optic nerve was once thought to be primarily due to diffuse RGC injury; however, it has recently been proposed that this form of neurodegeneration appears to be compartmentalized, thus affecting neuronal pro-cesses well before the RGC body in the inner retina150. This has important implications for the development of neuroprotective therapies for glaucoma, as functional changes or axonal dystrophy may be more amenable to treatment than the replacement of lost RGC bodies or long stretches of their axons in the optic nerve or their projections into the brainstem151.

Excessive activation of the NMDA (N-methyl-d-aspartate) receptor signalling cascade leads to excito-toxicity, wherein intracellular calcium overloads RGCs and other neurons, causing cell death through apoptosis. Oral administration of memantine to non-human pri-mates with experimentally induced glaucoma conferred protection of RGCs and their target — the relay neurons in the lateral geniculate body44. A Phase III clinical trial has been completed to investigate the efficacy of the NMDA glutamate receptor antagonist memantine in glaucoma

Table 5 | Neuroprotective agents in clinical trials

Drugs Company Description and mechanism of action

Indication (clinical phase)

ClinicalTrials.gov identifier

Refs

AL-8309B Alcon Selective 5-HT1A

agonist; neuroprotective against excitotoxic neuronal damage

Geographic atrophy (Phase III)

NCT00890097 (completed)

139

Brimonidine tartrate (intravitreal implant)

Allergan α-adrenergic receptor agonist; promotes the production of neurotrophic factors such as CNTF, and protects photoreceptors in animal models of retinal degeneration caused by light damage of photoreceptor cells

Geographic atrophy (Phase II)

NCT00658619 140

RN6G Pfizer Monoclonal antibody; prevents accumulation of amyloid-β40 and amyloid-β42, thus preventing damage to retina

AMD (Phase I) NCT01003691 141,142

CNTF Neurotech Neurotrophic factor; protects photoreceptors from degeneration, slowing progression of vision loss

Geographic atrophy (Phase II)

NCT01408472 94, 143–150

Memantine Allergan NMDA glutamate receptor antagonist; protects retinal ganglion cells and their target (the relay neurons) in the lateral geniculate body

Primary open-angle glaucoma (Phase III)

NCT00168350 (completed)

44

5-HT1A

, 5-hydroxytryptamine 1A receptor; AMD, age-related macular degeneration; CNTF, ciliary neurotrophic factor; NMDA, N-methyl-d-aspartate.

R E V I E W S



neuroprotection (protection of the structure and function of the optic nerve to halt glaucoma progression indepen-dently of lowering IOP). However, memantine failed to reach the primary efficacy end point of reducing visual field loss in patients at a high risk of developing glaucoma. Data from this study have not yet been reported.

The efficacy of topical brimonidine in preserving visual field loss in glaucoma has also been investigated. Brimonidine is approved by the FDA for lowering IOP in glaucoma, and systemic administration of the drug has been reported to protect RGCs in an experimental rat model of ocular hypertension152. In the low-tension glau-coma treatment study, patients who were randomized to receive topical brimonidine monotherapy had less visual field loss than those who were randomized to receive topical timolol (Timoptic; Merck & Co.), even though both drugs invoked similar IOP-lowering effects153.

Although it has been suggested that this result is consistent with the neuroprotective effects of topical brimonidine, there are several important limitations of the study, including the considerably small sample size, the large number of dropouts in the brimonidine group and the possibility that the patients who were treated with brimonidine were unmasked as they often had con-junctival hyperaemia. Therefore, although the results are intriguing, they do need corroboration. The brimonidine tartrate intravitreal implant that is being investigated for AMD might also be useful for treating glaucoma, par-ticularly if it is used for intracameral drug delivery to lower IOP and provide neuroprotection in glaucoma, but such studies have not yet been undertaken.

Potential future therapeutic approachesTarget discovery through genomic research. A crucial role of genetic variation in the occurrence and develop-ment of primary open-angle glaucoma has been affirmed by the varying prevalence of primary open-angle glau-coma observed in different populations, family aggrega-tion trends154,155 and in twin studies. Some populations have a higher prevalence of primary open-angle glau-coma than others (for example, the prevalence of the disease in individuals of African ancestry is several-fold higher than in those of European ancestry). This suggests that genetic background may be a primary cause of the disease. A growing number of studies, including genome-wide association studies, have identified glaucoma sus-ceptibility loci for monogenic and common primary open-angle glaucoma (reviewed in REFS 156,157). Genes that are involved in cell cycle control and transforming growth factor-β (TGFβ) pathways have emerged as the main risk loci for primary open-angle glaucoma158,159. However, it is not clear at present how these findings could be translated into therapies.

There is increasing evidence that genetic susceptibili-ties contribute to the development of diabetic retinopathy. For instance, the frequency of diabetic retinopathy varies among different ethnicities and its prevalence is higher in individuals with a positive family history160–162. Identifying genes (such as erythropoietin)163 that are associated with diabetic retinopathy could lead to the development of targeted treatment strategies in the near future.

AMD is an excellent example in which novel target discoveries have been achieved through genomic research. More than ten genes have been identified for AMD, which point to several new pathways that may be involved in the pathogenesis of AMD and could be used to predict the risk of developing the disease164. These genes encode pro-teins such as hepatic lipase and apolipoprotein E (which are involved in lipid metabolism)165,166, proteases such as HTRA1 (REFS 27,40) as well as components of the com-plement pathway. Owing to multiple gene associations in each of these pathways, various drugs targeting these pathways (for example, the alternative complement path-way) are now in clinical trials.

Gene therapy. Gene therapy has been an active area of clinical investigation. It can provide long-term, sustained effects with a single injection; however, reversibility needs to be addressed as long-term continuous blockade may not be desirable. Various ocular gene therapy trials have been highly successful in treating retinal degenera-tion167–169. An AAV2-mediated gene transfer of the MER receptor tyrosine kinase (MERTK) gene rescued retinal degeneration resulting from a MERTK mutation in RCS rats170, and a Phase I clinical trial is underway in patients with recessive MERTK mutations. TT30 is a recombi-nant CFH fusion protein that is under development for replacement therapy of defective CFH (see the Taligen Therapeutics website). Knockdown or silencing of CFH-related 1 (CFHR1) by preventing its mRNA expression might be useful for the treatment of AMD because dele-tion of CFHR1 protects against AMD independently of CFH modulation itself 171,172.

Stem cell biology and therapy. More recently, there has been increasing interest in the potential use of stem cell therapy to achieve ocular tissue repair and/or regenera-tion173,174. The eye is a particularly attractive target organ for stem cell therapy as it is in a contained intraocular space and is therefore associated with minimal systemic safety concerns and immune reactions. The effect of stem cell therapy can also be monitored through non-invasive diagnostic tools such as optical coherence tomography. These stem cell advancements include our considerably improved ability to manipulate and control stem cell fate and function in a more precise and defined fashion, as well as new approaches to reprogramme somatic cells towards various tissue-specific lineages173,175. For exam-ple, RPE can be derived from patients with geographic atrophy who have a high genetic risk genotype, and this RPE can be subjected to an in vitro high-throughput screen to identify small molecules that can prevent the death of RPE cells or enhance their survival173. Ultimately, it is possible that conventional small-molecule or biologi-cal therapeutics may be developed that could enable one’s own cells to regenerate in vivo to alleviate damage caused by diseases and injuries.

Advanced Cell Technology has produced human embryonic stem cell-derived RPE cells and has received FDA approval for their use in clinical trials. Two com-bined, multicentre, open-label Phase I/II trials are cur-rently underway, using RPE cell subretinal transplantation

R E V I E W S



http://www.taligentherapeutics.com/pipeline/index.html


to treat patients with Stargardt macular dystrophy and geographic atrophy. These trials are prospective open-label studies to determine the safety and tolerability of transplanted RPE cells. Preclinical studies in animals showed that transplanted RPE cells preserved overlying photoreceptors, slowed the progression of degeneration and improved visual functions176. StemCells has recently filed an investigational new drug application with the FDA to initiate a Phase I/II multicentre study using adult-derived neural stem cells to treat patients with geographic atrophy.

Ocular drug deliveryDespite advances made in the discovery of novel targets and an understanding of their mechanisms of action, drug delivery to these targets — especially when it is to the posterior eye — is continually challenged by anatomi-cal and physiological constraints, particularly the blood–ocular barriers and an array of efflux transporters. As a result, therapeutic agents that are systemically admin-istered are inevitably associated with adverse systemic exposure and unsatisfying efficacy177. Although using intraocular or periocular injections can substantially lower systemic exposure when delivering therapeutic entities to the retina, poor patient compliance and the high risk of complications (including retinal detach-ment, endophthalmitis and increased IOP) have limited the frequency of administration via these routes178. For example, anti-VEGF therapies are highly effective in treating wet AMD, diabetic macular oedema and reti-nal vein-occlusive diseases, but a monthly intraocular injection is not an ideal drug delivery platform and it is hard to maintain patient compliance.

Overall, these challenges have hindered the use of many novel therapeutic agents, thus emphasizing the urgency to develop novel drug delivery strategies for the eye. Among various approaches to optimize drug delivery, ECT and applications of nanomedicine can have essential roles.

Encapsulated cell technology. ECT is a cell-based delivery system that is used to treat chronic disorders of the eye179. In this technology, specific cells are first genetically engi-neered to overexpress desired therapeutic proteins, and are then encapsulated into semipermeable capsules that facilitate the diffusion of nutrients and proteins but pre-vent attack by the host immune system. For treatment, the capsules are surgically implanted into target sites, where the cells will survive and release proteins. As any gene encoding a therapeutic protein can now be engi-neered into cells, ECT has a broad range of applications. The implantation of cell-loaded capsules also allows for controlled, continuous and long-term administration of therapeutic proteins to the proximity of the eye in situ with no systemic exposure. The implant can also be retrieved, providing an added level of safety180.

ECT has advanced into the clinic as a result of an improved understanding of the mechanisms of protein functions related to visual impairments, coupled with advances in genetic engineering and implant technol-ogy181. In particular, encapsulated human cells that are

genetically modified to secrete CNTF (for the treatment of geographic atrophy associated with dry macular degeneration and retinitis pigmentosa) have advanced to Phase II clinical trials, with promising results150.

ECT can also be extended to other ophthalmic dis-eases, depending on the secreted protein. For example, the delivery of alginate-encapsulated cells producing glucagon-like peptide 1 (GLP1) resulted in the intraocular production of GLP1 without causing the obvious cyto-toxic effects in animal studies182. Meanwhile, a Phase I clinical trial that is currently being planned by Neurotech involves the implantation of encapsulated human cells that are genetically modified to deliver a structural antag-onist of VEGF for the treatment of wet AMD150.

Nanomedicine. Nanomedicine, which exploits nanotech-nology to solve medical challenges, has offered numer-ous exciting possibilities in health care. In particular, nanoparticle drugs have become established therapeutics by formulating nanometre-scale carriers composed of various therapeutic entities (including small-molecule drugs, peptides, proteins and nucleic acids), together with excipients such as lipids and polymers that assemble with these therapeutic entities183,184. Although nanopar-ticle therapeutics were initially developed for cancer treatment, their application has expanded into oph-thalmology owing largely to their unique capability of enhancing drug efficacy — through improved drug encapsulation, sustained or triggered drug release, and preferential targeting to disease sites185. Some nano-particle therapeutics have been clinically approved for treating ophthalmic diseases. For example, pegaptanib — an anti-VEGF aptamer conjugated with branched polyethylene glycol — has been approved for the treat-ment of AMD186.

In addition, nanomedicine has provided new oppor-tunities for overcoming obstacles in treating ophthalmic diseases. For example, by combining synthetic chem-istry with a basic understanding of protein–polymer interactions, nanotechnology has led to the efficient encapsulation and controlled delivery of proteins within biological fluid under mild conditions187. These technologies are particularly promising as the thera-peutic potential of proteins and monoclonal antibodies is increasingly becoming realized in ophthalmology, particularly for halting retinal angiogenesis188. In addi-tion, nanotechnology has resulted in the formation of sophisticated carriers that are particularly suitable for the delivery of therapeutic agents to the cytoplasm for bio-activity189. Technology for intracellular delivery, if availa-ble, can be used in siRNA-based therapeutics to selectively silence gene expression for the treatment of ophthalmic diseases.

Moreover, nanotechnology has provided new oppor-tunities for drug delivery, with the goal of attaining pro-longed ocular drug retention. In particular, approaches that are under active investigation include the incorpora-tion of bioadhesive components to formulate multifunc-tional nanoparticle drug reservoirs that can be attached to the corneal tissue with a high affinity190. Furthermore, the recent development of polymeric nanoparticles that

NanotechnologyThe understanding and control of matter at dimensions between approximately 1 nm and 100 nm. It involves imaging, measuring, modelling and manipulating matter at

this length scale.

Nanoparticle drugsNanometre-scale particles that are used to carry and transport pharmaceutical agents to improve therapeutic efficacy, drug safety and patient compliance.

R E V I E W S



are camouflaged by the red blood cell membrane allows man-made delivery vehicles to share functionalities that have been engineered and perfected by nature, hence taking another step towards bridging synthetic materials with natural components191. These biomimetic nano-particles — that have excellent biocompatibility and well-controlled drug release kinetics — will have many applications in the treatment of ophthalmic diseases.

ConclusionsCurrently, anti-VEGF therapies are highly effective in treating wet AMD, diabetic macular oedema and retinal vein-occlusive diseases, and have become the first-line treatment for these conditions. However, there are still several common blinding ophthalmic disorders, such as geographic atrophy, for which no effective therapies are currently available. Moreover, many individuals with

1. Rein, D. B. et al. The economic burden of major adult visual disorders in the United States. Arch. Ophthalmol. 124, 1754–1760 (2006).

2. Gaudana, R., Ananthula, H. K., Parenky, A. & Mitra, A. K. Ocular drug delivery. AAPS J. 12, 348–360 (2010).

3. Jager, R. D., Mieler, W. F. & Miller, J. W. Age-related macular degeneration. N. Engl. J. Med. 358, 2606–2617 (2008).This paper provides a general overview of AMD.

4. Zarbin, M. A. & Rosenfeld, P. J. Pathway-based therapies for age-related macular degeneration: an integrated survey of emerging treatment alternatives. Retina 30, 1350–1367 (2010).This is a review on the molecular pathways that are involved in AMD as well as targets for drug development.

5. Yehoshua, Z., Rosenfeld, P. J. & Albini, T. A. Current clinical trials in dry AMD and the definition of appropriate clinical outcome measures. Semin. Ophthalmol. 26, 167–180 (2011).

6. Weinreb, R. N. & Khaw, P. T. Primary open-angle glaucoma. Lancet 363, 1711–1720 (2004).This is a general overview of primary open-angle glaucoma.

7. Haddad, S., Chen, C. A., Santangelo, S. L. & Seddon, J. M. The genetics of age-related macular degeneration: a review of progress to date. Surv. Ophthalmol. 51, 316–363 (2006).

8. Rattner, A. & Nathans, J. Macular degeneration: recent advances and therapeutic opportunities. Nature Rev. Neurosci. 7, 860–872 (2006).

9. Penfold, P. L., Killingsworth, M. C. & Sarks, S. H. Senile macular degeneration: the involvement of immunocompetent cells. Graefes Arch. Clin. Exp. Ophthalmol. 223, 69–76 (1985).

10. Klein, R., Klein, B. E. & Linton, K. L. Prevalence of age-related maculopathy. The Beaver Dam Eye Study. Ophthalmology 99, 933–943 (1992).

11. Vingerling, J. R. et al. The prevalence of age-related maculopathy in the Rotterdam study. Ophthalmology 102, 205–210 (1995).

12. Hageman, G. S. et al. An integrated hypothesis that considers drusen as biomarkers of immune-mediated processes at the RPE-Bruch’s membrane interface in aging and age-related macular degeneration. Prog. Retin. Eye Res. 20, 705–732 (2001).

13. Kaplan, H. J., Leibole, M. A., Tezel, T. & Ferguson,T. A. Fas ligand (CD95 ligand) controls angiogenesis beneath the retina. Nature Med. 5, 292–297 (1999).

14. Krzystolik, M. G. et al. Prevention of experimental choroidal neovascularization with intravitreal anti-vascular endothelial growth factor antibody fragment. Arch. Ophthalmol. 120, 338–346 (2002).

15. Okamoto, N. et al. Transgenic mice with increased expression of vascular endothelial growth factor in the retina: a new model of intraretinal and subretinal neovascularization. Am. J. Pathol. 151, 281–291 (1997).

16. Friedman, D. S. et al. Prevalence of age-related macular degeneration in the United States. Arch. Ophthalmol. 122, 564–572 (2004).

17. Dithmar, S. et al. Murine high-fat diet and laser photo -chemical model of basal deposits in Bruch membrane. Arch. Ophthalmol. 119, 1643–1649 (2001).

18. Green, W. R. & Key, S. N. Senile macular degeneration: a histopathologic study. Trans. Am. Ophthalmol. Soc. 75, 180–254 (1977).

19. Hageman, G. S. & Mullins, R. F. Molecular composition of drusen as related to substructural phenotype. Mol. Vis. 5, 28 (1999).

20. Kvanta, A., Algvere, P. V., Berglin, L. & Seregard, S. Subfoveal fibrovascular membranes in age-related macular degeneration express vascular endothelial growth factor. Invest. Ophthalmol. Vis. Sci. 37, 1929–1934 (1996).

21. Hageman, G. S. et al. A common haplotype in the complement regulatory gene factor H (HF1/CFH) predisposes individuals to age-related macular degeneration. Proc. Natl Acad. Sci. USA 102, 7227–7232 (2005).

22. Mullins, R. F., Russell, S. R., Anderson, D. H. & Hageman, G. S. Drusen associated with aging and age-related macular degeneration contain proteins common to extracellular deposits associated with atherosclerosis, elastosis, amyloidosis, and dense deposit disease. FASEB J. 14, 835–846 (2000).

23. Mullins, R. F., Aptsiauri, N. & Hageman, G. S. Structure and composition of drusen associated with glomerulonephritis: implications for the role of complement activation in drusen biogenesis. Eye 15, 390–395 (2001).

24. Anderson, R. E. et al. Low docosahexaenoic acid levels in rod outer segments of rats with P23H and S334ter rhodopsin mutations. Mol. Vis. 8, 351–358 (2002).

25. Johnson, L. V., Leitner, W. P., Staples, M. K. & Anderson, D. H. Complement activation and inflammatory processes in Drusen formation and age related macular degeneration. Exp. Eye Res. 73, 887–896 (2001).

26. Crabb, J. W. et al. Drusen proteome analysis: an approach to the etiology of age-related macular degeneration. Proc. Natl Acad. Sci. USA 99, 14682–14687 (2002).

27. Yang, Z. et al. A variant of the HTRA1 gene increases susceptibility to age-related macular degeneration. Science 314, 992–993 (2006).

28. Gehrs, K. M., Anderson, D. H., Johnson, L. V. & Hageman, G. S. Age-related macular degeneration — emerging pathogenetic and therapeutic concepts. Ann. Med. 38, 450–471 (2006).

29. Kaneko, H. et al. DICER1 deficit induces Alu RNA toxicity in age-related macular degeneration. Nature 471, 325–330 (2011).

30. Weismann, D. et al. Complement factor H binds malondialdehyde epitopes and protects from oxidative stress. Nature 478, 76–81 (2011).

31. Nozaki, M. et al. Drusen complement components C3a and C5a promote choroidal neovascularization. Proc. Natl Acad. Sci. USA 103, 2328–2333 (2006).

32. Ferrara, N. Vascular endothelial growth factor and age-related macular degeneration: from basic science to therapy. Nature Med. 16, 1107–1111 (2010).This is a good review on the biology of VEGF and the use of anti-VEGF therapy in AMD.

33. Aiello, L. P. et al. Vascular endothelial growth factor in ocular fluid of patients with diabetic retinopathy and other retinal disorders. N. Engl. J. Med. 331, 1480–1487 (1994).

34. Adamis, A. P. et al. Increased vascular endothelial growth factor levels in the vitreous of eyes with proliferative diabetic retinopathy. Am. J. Ophthalmol. 118, 445–450 (1994).

35. Hollyfield, J. G. et al. Oxidative damage-induced inflammation initiates age-related macular degeneration. Nature Med. 14, 194–198 (2008).

36. Tuo, J. et al. Murine ccl2/cx3cr1 deficiency results in retinal lesions mimicking human age-related macular degeneration. Invest. Ophthalmol. Vis. Sci. 48, 3827–3836 (2007).

37. Karan, G. et al. Lipofuscin accumulation, abnormal electrophysiology, and photoreceptor degeneration in mutant ELOVL4 transgenic mice: a model for macular degeneration. Proc. Natl Acad. Sci. USA 102, 4164–4169 (2005).

38. Maeda, A., Maeda, T., Golczak, M. & Palczewski, K. Retinopathy in mice induced by disrupted all-trans-retinal clearance. J. Biol. Chem. 283, 26684–26693 (2008).

39. Yang, Z. et al. Mutant prominin 1 found in patients with macular degeneration disrupts photoreceptor disk morphogenesis in mice. J. Clin. Invest. 118, 2908–2916 (2008).

40. Jones, A. et al. Increased expression of multifunctional serine protease, HTRA1, in retinal pigment epithelium induces polypoidal choroidal vasculopathy in mice. Proc. Natl Acad. Sci. USA 108, 14578–14583 (2011).

41. Age-Related Eye Disease Study Research Group. A randomized, placebo-controlled, clinical trial of high-dose supplementation with vitamins C and E, beta carotene, and zinc for age-related macular degeneration and vision loss: AREDS report no. 8. Arch. Ophthalmol. 119, 1417–1436 (2001).

42. Varma, R. et al. Biologic risk factors associated with diabetic retinopathy: the Los Angeles Latino Eye Study. Ophthalmology 114, 1332–1340 (2007).

43. Patel, S., Chen, H., Tinkham, N. H. & Zhang, K. Genetic susceptibility of diabetic retinopathy. Curr. Diab. Rep. 8, 257–262 (2008).

44. Yucel, Y. H., Zhang, Q., Weinreb, R. N., Kaufman, P. L. & Gupta, N. Effects of retinal ganglion cell loss on magno-, parvo-, koniocellular pathways in the lateral geniculate nucleus and visual cortex in glaucoma. Prog. Retin. Eye Res. 22, 465–481 (2003).

45. Quigley, H. A. Glaucoma. Lancet 377, 1367–1377 (2011).

46. Resnikoff, S. et al. Global data on visual impairment in the year 2002. Bull. World Health Organ. 82, 844–851 (2004).

47. Weinreb, R. N. & Harris, A. (eds) Ocular Blood Flow in Glaucoma (Kugler Publications Amsterdam, 2009).

glaucoma continue to develop visual impairment despite receiving treatment with IOP-lowering drugs. Present research aims to improve our understanding of the molecular mechanisms underlying the development of ophthalmic disorders and identify potential therapeutic targets. Clinical trials are currently underway to evaluate new drugs directed against several novel targets, such as complement inhibitors for AMD and ROCK inhibi-tors for glaucoma. We need to develop animal models that more accurately recapitulate ophthalmic diseases in humans in order to validate new therapeutic agents before embarking on expensive clinical trials. Sustained ocular drug release platforms to deliver either large bio-logics or small-molecule drugs are an important focus. Developing new therapeutics and preventing the loss of vision will present both great opportunities and challenges for the next decades to come.

R E V I E W S



48. Weinreb, R. N. et al. Risk assessment in the management of patients with ocular hypertension. Am. J. Ophthalmol. 138, 458–467 (2004).

49. Brown, D. M. et al. Ranibizumab versus verteporfin for neovascular age-related macular degeneration. N. Engl. J. Med. 355, 1432–1444 (2006).This was one of the first reports to demonstrate the efficacy of anti-VEGF antibody therapy in exudative AMD.

50. Rosenfeld, P. J. et al. Ranibizumab for neovascular age-related macular degeneration. N. Engl. J. Med. 355, 1419–1431 (2006).This was one of the first reports to demonstrate the efficacy of anti-VEGF antibody therapy in exudative AMD.

51. Martin, D. F. et al. Ranibizumab and bevacizumab for neovascular age-related macular degeneration. N. Engl. J. Med. 364, 1897–1908 (2011).

52. Rich, R. M. et al. Short-term safety and efficacy of intravitreal bevacizumab (Avastin) for neovascular age-related macular degeneration. Retina 26, 495–511 (2006).

53. Shih, T. Y., Gaydos, C. A., Rothman, R. E. & Hsieh, Y. H. Poor provider adherence to the Centers for Disease Control and Prevention treatment guidelines in US emergency department visits with a diagnosis of pelvic inflammatory disease. Sex Transm. Dis. 38, 299–305 (2011).

54. Zhang, M. et al. A Phase 1 study of KH902, a vascular endothelial growth factor receptor decoy, for exudative age-related macular degeneration. Ophthalmology 118, 672–678 (2011).

55. Nguyen, Q. D. et al. A Phase I trial of an IV-administered vascular endothelial growth factor trap for treatment in patients with choroidal neovascularization due to age-related macular degeneration. Ophthalmology 113, 1522.e1–1522.e14 (2006).

56. Brown, D. M. et al. Ranibizumab for macular edema following central retinal vein occlusion: six-month primary end point results of a Phase III study. Ophthalmology 117, 1124–1133 (2010).

57. Elman, M. J. et al. Randomized trial evaluating ranibizumab plus prompt or deferred laser or triamcinolone plus prompt laser for diabetic macular edema. Ophthalmology 117, 1064–1077 (2010).

58. Elman, M. J. et al. Expanded 2-year follow-up of ranibizumab plus prompt or deferred laser or triamcinolone plus prompt laser for diabetic macular edema. Ophthalmology 118, 609–614 (2011).

59. Migdal, C. Glaucoma medical treatment: philosophy, principles and practice. Eye 14, 515–518 (2000).

60. Medeiros, F. A. & Weinreb, R. N. Medical back-grounders: glaucoma. Drugs Today 38, 563–570 (2002).

61. Sagara, T. et al. Topical prostaglandin F2alpha treatment reduces collagen types I, III, and IV in the monkey uveoscleral outflow pathway. Arch. Ophthalmol. 117, 794–801 (1999).

62. Liu, J. H., Kripke, D. F. & Weinreb, R. N. Comparison of the nocturnal effects of once-daily timolol and latanoprost on intraocular pressure. Am. J. Ophthalmol. 138, 389–395 (2004).

63. Nathanson, J. A. Human ciliary process adrenergic receptor: pharmacological characterization. Invest. Ophthalmol. Vis. Sci. 21, 798–804 (1981).

64. Liu, J. H., Medeiros, F. A., Slight, J. R. & Weinreb, R. N. Comparing diurnal and nocturnal effects of brinzolamide and timolol on intraocular pressure in patients receiving latanoprost monotherapy. Ophthalmology 116, 449–454 (2009).

65. Liu, J. H., Medeiros, F. A., Slight, J. R. & Weinreb, R. N. Diurnal and nocturnal effects of brimonidine monotherapy on intraocular pressure. Ophthalmology 117, 2075–2079 (2010).

66. Toris, C. B., Gleason, M. L., Camras, C. B. & Yablonski, M. E. Effects of brimonidine on aqueous humor dynamics in human eyes. Arch. Ophthalmol. 113, 1514–1517 (1995).

67. Hernandez, M., Urcola, J. H. & Vecino, E. Retinal ganglion cell neuroprotection in a rat model of glaucoma following brimonidine, latanoprost or combined treatments. Exp. Eye Res. 86, 798–806 (2008).

68. Edwards, A. O. et al. Complement factor H polymorphism and age-related macular degeneration. Science 308, 421–424 (2005).

69. Haines, J. L. et al. Complement factor H variant increases the risk of age-related macular degeneration. Science 308, 419–421 (2005).

70. Klein, R. J. et al. Complement factor H polymorphism in age-related macular degeneration. Science 308, 385–389 (2005).

71. Li, M. et al. CFH haplotypes without the Y402H coding variant show strong association with susceptibility to age-related macular degeneration. Nature Genet. 38, 1049–1054 (2006).

72. Maller, J. et al. Common variation in three genes, including a noncoding variant in CFH, strongly influences risk of age-related macular degeneration. Nature Genet. 38, 1055–1059 (2006).

73. Yates, J. R. et al. Complement C3 variant and the risk of age-related macular degeneration. N. Engl. J. Med. 357, 553–561 (2007).

74. Fagerness, J. A. et al. Variation near complement factor I is associated with risk of advanced AMD. Eur. J. Hum. Genet. 17, 100–104 (2009).

75. Cousins, S. W. & the Ophthotech Study Group. Targeting complement factor 5 in combination with vascular endothelial growth factor (VEGF) inhibition for neovascular age related macular degeneration (AMD): results of a Phase 1 study. Invest. Ophthalmol. Vis. Sci. 51, e-Abstract 1251 (2010).

76. Mata, N. L., Weng, J. & Travis, G. H. Biosynthesis of a major lipofuscin fluorophore in mice and humans with ABCR-mediated retinal and macular degeneration. Proc. Natl Acad. Sci. USA 97, 7154–7159 (2000).

77. Sparrow, J. R., Nakanishi, K. & Parish, C. A. The lipofuscin fluorophore A2E mediates blue light-induced damage to retinal pigmented epithelial cells. Invest. Ophthalmol. Vis. Sci. 41, 1981–1989 (2000).

78. Kubota, R. et al. Safety and effect on rod function of ACU-4429, a novel small-molecule visual cycle modulator. Retina 32, 183–188 (2012).

79. Weng, J. et al. Insights into the function of Rim protein in photoreceptors and etiology of Stargardt’s disease from the phenotype in abcr knockout mice. Cell 98, 13–23 (1999).

80. Land, S. C. & Tee, A. R. Hypoxia-inducible factor 1α is regulated by the mammalian target of rapamycin (mTOR) via an mTOR signaling motif. J. Biol. Chem. 282, 20534–20543 (2007).

81. Brafman, A. et al. Inhibition of oxygen-induced retinopathy in RTP801-deficient mice. Invest. Ophthalmol. Vis. Sci. 45, 3796–3805 (2004).

82. Kleinman, M. E. et al. Sequence- and target-independent angiogenesis suppression by siRNA via TLR3. Nature 452, 591–597 (2008).

83. Hornung, V. et al. Sequence-specific potent induction of IFN-α by short interfering RNA in plasmacytoid dendritic cells through TLR7. Nature Med. 11, 263–270 (2005).

84. Sledz, C. A., Holko, M., de Veer, M. J., Silverman, R. H. & Williams, B. R. Activation of the interferon system by short-interfering RNAs. Nature Cell Biol. 5, 834–839 (2003).

85. Dawson, D. W. et al. Pigment epithelium-derived factor: a potent inhibitor of angiogenesis. Science 285, 245–248 (1999).

86. Bhutto, I. A. et al. Reduction of endogenous angiogenesis inhibitors in Bruch’s membrane of the submacular region in eyes with age-related macular degeneration. Arch. Ophthalmol. 126, 670–678 (2008).

87. Holekamp, N. M., Bouck, N. & Volpert, O. Pigment epithelium-derived factor is deficient in the vitreous of patients with choroidal neovascularization due to age-related macular degeneration. Am. J. Ophthalmol. 134, 220–227 (2002).

88. Maclachlan, T. K. et al. Preclinical safety evaluation of AAV2-sFLT01— a gene therapy for age-related macular degeneration. Mol. Ther. 19, 326–334 (2011).

89. Kendall, R. L. & Thomas, K. A. Inhibition of vascular endothelial cell growth factor activity by an endogenously encoded soluble receptor. Proc. Natl Acad. Sci. USA 90, 10705–10709 (1993).

90. Ambati, B. K. et al. Corneal avascularity is due to soluble VEGF receptor-1. Nature 443, 993–997 (2006).

91. Ferrara, N., Gerber, H. P. & LeCouter, J. The biology of VEGF and its receptors. Nature Med. 9, 669–676 (2003).

92. Takeda, A. et al. CCR3 is a target for age-related macular degeneration diagnosis and therapy. Nature 460, 225–230 (2009).

93. Takahashi, K., Saishin, Y., King, A. G., Levin, R. & Campochiaro, P. A. Suppression and regression of choroidal neovascularization by the multitargeted kinase inhibitor pazopanib. Arch. Ophthalmol. 127, 494–499 (2009).

94. LaVail, M. M. et al. Protection of mouse photoreceptors by survival factors in retinal degenerations. Invest. Ophthalmol. Vis. Sci. 39, 592–602 (1998).

95. Palanki, M. S. et al. Development of prodrug 4-chloro-3-(5-methyl-3-{[4-(2-pyrrolidin-1-ylethoxy)phenyl]amino}-1,2,4-benzotria zin-7-yl)phenyl benzoate (TG100801): a topically administered therapeutic candidate in clinical trials for the treatment of age-related macular degeneration. J. Med. Chem. 51, 1546–1559 (2008).

96. Heeschen, C., Weis, M., Aicher, A., Dimmeler, S. & Cooke, J. P. A novel angiogenic pathway mediated by non-neuronal nicotinic acetylcholine receptors. J. Clin. Invest. 110, 527–536 (2002).

97. Kiuchi, K. et al. Mecamylamine suppresses basal and nicotine-stimulated choroidal neovascularization. Invest. Ophthalmol. Vis. Sci. 49, 1705–1711 (2008).

98. Campochiaro, P. A. et al. Topical mecamylamine for diabetic macular edema. Am. J. Ophthalmol. 149, 839–851 (2010).

99. Ozaki, H., Hla, T. & Lee, M. J. Sphingosine-1-phosphate signaling in endothelial activation. J. Atheroscler. Thromb. 10, 125–131 (2003).

100. Watterson, K. R., Lanning, D. A., Diegelmann, R. F. & Spiegel, S. Regulation of fibroblast functions by lysophospholipid mediators: potential roles in wound healing. Wound Repair Regen. 15, 607–616 (2007).

101. Caballero, S. et al. Anti-sphingosine-1-phosphate monoclonal antibodies inhibit angiogenesis and sub-retinal fibrosis in a murine model of laser-induced choroidal neovascularization. Exp. Eye Res. 88, 367–377 (2009).

102. Xie, B. et al. Blockade of sphingosine-1-phosphate reduces macrophage influx and retinal and choroidal neovascularization. J. Cell Physiol. 218, 192–198 (2009).

103. Orecchia, A. et al. Vascular endothelial growth factor receptor-1 is deposited in the extracellular matrix by endothelial cells and is a ligand for the α5β1 integrin. J. Cell Sci. 116, 3479–3489 (2003).

104. Zahn, G. et al. Preclinical evaluation of the novel small-molecule integrin α5β1 inhibitor JSM6427 in monkey and rabbit models of choroidal neovascularization. Arch. Ophthalmol. 127, 1329–1335 (2009).

105. Nirmalan, P. K. et al. Relationship between vision impairment and eye disease to vision-specific quality of life and function in rural India: the Aravind Comprehensive Eye Survey. Invest. Ophthalmol. Vis. Sci. 46, 2308–2312 (2005).

106. Pettit, G. R. et al. Antineoplastic agents 322. synthesis of combretastatin A-4 prodrugs. Anticancer Drug Des. 10, 299–309 (1995).

107. Dark, G. G. et al. Combretastatin A-4, an agent that displays potent and selective toxicity toward tumor vasculature. Cancer Res. 57, 1829–1834 (1997).

108. Nambu, H., Nambu, R., Melia, M. & Campochiaro, P. A. Combretastatin A-4 phosphate suppresses development and induces regression of choroidal neovascularization. Invest. Ophthalmol. Vis. Sci. 44, 3650–3655 (2003).

109. Griggs, J. et al. Inhibition of proliferative retinopathy by the anti-vascular agent combretastatin-A4. Am. J. Pathol. 160, 1097–1103 (2002).

110. Benjamin, L. E., Hemo, I. & Keshet, E. A plasticity window for blood vessel remodelling is defined by pericyte coverage of the preformed endothelial network and is regulated by PDGF-B and VEGF. Development 125, 1591–1598 (1998).

111. Jo, N. et al. Inhibition of platelet-derived growth factor B signaling enhances the efficacy of anti-vascular endothelial growth factor therapy in multiple models of ocular neovascularization. Am. J. Pathol. 168, 2036–2053 (2006).

112. Boyer, D. S. & the Ophthotech Anti-PDGF in AMD Study Group. Combined inhibition of platelet derived (PDGF) and vascular endothelial (VEGF) growth factors for the treatment of neovascular age-related macular degeneration (NV-AMD) — results of a Phase 1 study. Invest. Ophthalmol. Vis. Sci. 50, e-Abstract 1260 (2009).

113. Sit, A. J., Weinreb, R. N., Crowston, J. G., Kripke, D. F. & Liu, J. H. Sustained effect of travoprost on diurnal and nocturnal intraocular pressure. Am. J. Ophthalmol. 141, 1131–1133 (2006).

114. Liu, J. H. & Weinreb, R. N. Monitoring intraocular pressure for 24 h. Br. J. Ophthalmol. 95, 599–600 (2011).

115. Coue, M., Brenner, S. L., Spector, I. & Korn, E. D. Inhibition of actin polymerization by latrunculin A. FEBS Lett. 213, 316–318 (1987).

R E V I E W S



116. Peterson, J. A. et al. Latrunculins’ effects on intraocular pressure, aqueous humor flow, and corneal endothelium. Invest. Ophthalmol. Vis. Sci. 41, 1749–1758 (2000).

117. Chen, J., Runyan, S. A. & Robinson, M. R. Novel ocular antihypertensive compounds in clinical trials. Clin. Ophthalmol. 5, 667–677 (2011).

118. Rao, P. V., Deng, P. F., Kumar, J. & Epstein, D. L. Modulation of aqueous humor outflow facility by the Rho kinase-specific inhibitor Y-27632. Invest. Ophthalmol. Vis. Sci. 42, 1029–1037 (2001).

119. Honjo, M. The possibility of selective Rho-associated kinase (ROCK) inhibitors as a medical treatment for glaucoma. [Article in Japanese] Nihon Ganka Gakkai Zasshi 113, 1071–1081 (2009).

120. Williams, R. D., Novack, G. D., van Haarlem, T. & Kopczynski, C. Ocular hypotensive effect of the Rho kinase inhibitor AR-12286 in patients with glaucoma and ocular hypertension. Am. J. Ophthalmol. 152, 834–841 (2011).

121. Wirostko B. M., Umeno, H., Hsu, H. H. & Kengatharan, M. Safety and efficacy of a novel topical Rho kinase inhibitor ATS907 in normotensive cynomolgus monkeys. Invest. Ophthalmol. Vis. Sci. 52, e-Abstract 3096 (2011).

122. Yamamoto, T., Abe, H., Kuwayama, Y., Tanihara, H. & Araie, M. Efficacy and safety of the Rho kinase inhibitor, K-115, over 24 hours in patients with primary open-angle glaucoma and ocular hypertension. Invest. Ophthalmol. Vis. Sci. 52, e-Abstract 216 (2011).

123. Mizuno, K. et al. Ocular hypotensive mechanism of K-115, a Rho-kinase inhibitor, and Rho-kinase expression in the eye. Invest. Ophthalmol. Vis. Sci. 52, e-Abstract 237 (2011).

124. Sugiyama, T. et al. Effects of fasudil, a Rho-associated protein kinase inhibitor, on optic nerve head blood flow in rabbits. Invest. Ophthalmol. Vis. Sci. 52, 64–69 (2011).

125. Honjo, M. et al. Potential role of Rho-associated protein kinase inhibitor Y-27632 in glaucoma filtration surgery. Invest. Ophthalmol. Vis. Sci. 48, 5549–5557 (2007).

126. Tian, B., Gabelt, B. T., Crosson, C. E. & Kaufman, P. L. Effects of adenosine agonists on intraocular pressure and aqueous humor dynamics in cynomolgus monkeys. Exp. Eye Res. 64, 979–989 (1997).

127. Wang, R. F., Podos, S. M., Mittag, T. W. & Yokoyoma, T. Effect of CS-088, an angiotensin AT1 receptor antagonist, on intraocular pressure in glaucomatous monkey eyes. Exp. Eye Res. 80, 629–632 (2005).

128. Chien, F. Y., Wang, R. F., Mittag, T. W. & Podos, S. M. Effect of WIN 55212-2, a cannabinoid receptor agonist, on aqueous humor dynamics in monkeys. Arch. Ophthalmol. 121, 87–90 (2003).

129. Hepler, R. S. & Frank, I. R. Marihuana smoking and intraocular pressure. JAMA 217, 1392 (1971).

130. Ruz, V., Moreno-Montañés, J., Sadaba, B., González, V. & Jiménez, A. I. Phase I study with a new siRNA: SYL040012. Tolerance and effect on intraocular pressure. Invest. Ophthalmol. Vis. Sci. 52, e-Abstract 223 (2011).

131. Schachar, R. A., Raber, S., Courtney, R. & Zhang, M. A Phase 2, randomized, dose-response trial of taprenepag isopropyl (PF-04217329) versus latanoprost 0.005% in open-angle glaucoma and ocular hypertension. Curr. Eye Res. 36, 809–817 (2011).

132. Schachar, R. A., Raber, S., Courtney, R., Zhang, M. & Bosworth, C. Dose-escalating, double-masked, vehicle-controlled trial of the IOP-reducing effect of the EP2 agonist PF-04217329. Invest. Ophthalmol. Vis. Sci. 51, e-Abstract 175 (2010).

133. Goldberg, D. F. & Williams, R. A Phase 2 study evaluating safety and efficacy of the latanoprost punctal plug delivery system (L-PPDS) in subjects with ocular hypertension (OH) or open-angle glaucoma (OAG). Invest. Ophthalmol. Vis. Sci. 53, e-Abstract 5095 (2012).

134. Clark, A. F. & Yorio, T. Ophthalmic drug discovery. Nature Rev. Drug Discov. 2, 448–459 (2003).

135. Mauler, F. & Horvath, E. Neuroprotective efficacy of repinotan HCl, a 5-HT1A receptor agonist, in animal models of stroke and traumatic brain injury. J. Cereb. Blood Flow Metab. 25, 451–459 (2005).

136. Ramos, A. J. et al. The 5HT1A receptor agonist, 8-OH-DPAT, protects neurons and reduces astroglial reaction after ischemic damage caused by cortical devascularization. Brain Res. 1030, 201–220 (2004).

137. Adayev, T., Ray, I., Sondhi, R., Sobocki, T. & Banerjee, P. The G protein-coupled 5-HT1A receptor causes suppression of caspase-3 through MAPK and protein kinase Cα. Biochim. Biophys. Acta 1640, 85–96 (2003).

138. Hsiung, S. C., Tamir, H., Franke, T. F. & Liu, K. P. Roles of extracellular signal-regulated kinase and Akt signaling in coordinating nuclear transcription factor-κB- dependent cell survival after serotonin 1A receptor activation. J. Neurochem. 95, 1653–1666 (2005).

139. Collier, R. J. et al. Agonists at the serotonin receptor (5-HT1A) protect the retina from severe photo-oxidative stress. Invest. Ophthalmol. Vis. Sci. 52, 2118–2126 (2011).

140. Wen, R., Cheng, T., Li, Y., Cao, W. & Steinberg, R. H. α2-adrenergic agonists induce basic fibroblast growth factor expression in photoreceptors in vivo and ameliorate light damage. J. Neurosci. 16, 5986–5992 (1996).

141. Johnson, L. V. et al. The Alzheimer’s Aβ-peptide is deposited at sites of complement activation in pathologic deposits associated with aging and age-related macular degeneration. Proc. Natl Acad. Sci. USA 99, 11830–11835 (2002).

142. Ding, J. D. et al. Targeting age-related macular degeneration with Alzheimer’s disease based immunotherapies: anti-amyloid-β antibody attenuates pathologies in an age-related macular degeneration mouse model. Vision Res. 48, 339–345 (2008).

143. Faktorovich, E. G., Steinberg, R. H., Yasumura, D., Matthes, M. T. & LaVail, M. M. Photoreceptor degeneration in inherited retinal dystrophy delayed by basic fibroblast growth factor. Nature 347, 83–86 (1990).

144. Fuhrmann, S., Kirsch, M. & Hofmann, H. D. Ciliary neurotrophic factor promotes chick photoreceptor development in vitro. Development 121, 2695–2706 (1995).

145. Fuhrmann, S., Grabosch, K., Kirsch, M. & Hofmann, H. D. Distribution of CNTF receptor α protein in the central nervous system of the chick embryo. J. Comp. Neurol. 461, 111–122 (2003).

146. Tao, W. et al. Encapsulated cell-based delivery of CNTF reduces photoreceptor degeneration in animal models of retinitis pigmentosa. Invest. Ophthalmol. Vis. Sci. 43, 3292–3298 (2002).

147. LaVail, M. M. et al. Multiple growth factors, cytokines, and neurotrophins rescue photoreceptors from the damaging effects of constant light. Proc. Natl Acad. Sci. USA 89, 11249–11253 (1992).

148. Cayouette, M. & Gravel, C. Adenovirus-mediated gene transfer of ciliary neurotrophic factor can prevent photoreceptor degeneration in the retinal degeneration (rd) mouse. Hum. Gene Ther. 8, 423–430 (1997).

149. Cayouette, M., Behn, D., Sendtner, M., Lachapelle, P. & Gravel, C. Intraocular gene transfer of ciliary neurotrophic factor prevents death and increases responsiveness of rod photoreceptors in the retinal degeneration slow mouse. J. Neurosci. 18, 9282–9293 (1998).

150. Zhang, K. et al. Ciliary neurotrophic factor delivered by encapsulated cell intraocular implants for treatment of geographic atrophy in age-related macular degeneration. Proc. Natl Acad. Sci. USA 108, 6241–6245 (2011).This is the first study to demonstrate neuro protection by CNTF in a Phase II clinical trial.

151. Lambert, W. S., Ruiz, L., Crish, S. D., Wheeler, L. A. & Calkins, D. J. Brimonidine prevents axonal and somatic degeneration of retinal ganglion cell neurons. Mol. Neurodegener. 6, 4 (2011).

152. WoldeMussie, E., Ruiz, G., Wijono, M. & Wheeler, L. A. Neuroprotection of retinal ganglion cells by brimonidine in rats with laser-induced chronic ocular hypertension. Invest. Ophthalmol. Vis. Sci. 42, 2849–2855 (2001).

153. Krupin, T., Liebmann, J. M., Greenfield, D. S., Ritch, R. & Gardiner, S. A randomized trial of brimonidine versus timolol in preserving visual function: results from the low-pressure glaucoma treatment study. Am. J. Ophthalmol. 151, 671–681 (2011).

154. Drance, S. M., Schulzer, M., Thomas, B. & Douglas, G. R. Multivariate analysis in glaucoma. Use of discriminant analysis in predicting glaucomatous visual field damage. Arch. Ophthalmol. 99, 1019–1022 (1981).

155. Wolfs, R. C. et al. Genetic risk of primary open-angle glaucoma. Population-based familial aggregation study. Arch. Ophthalmol. 116, 1640–1645 (1998).

156. Wiggs, J. L. Genetic etiologies of glaucoma. Arch. Ophthalmol. 125, 30–37 (2007).

157. Stone, E. M. et al. Identification of a gene that causes primary open angle glaucoma. Science 275, 668–670 (1997).

158. Ramdas, W. D. et al. Common genetic variants associated with open-angle glaucoma. Hum. Mol. Genet. 20, 2464–2471 (2011).

159. Burdon, K. P. et al. Genome-wide association study identifies susceptibility loci for open angle glaucoma at TMCO1 and CDKN2B‑AS1. Nature Genet. 43, 574–578 (2011).

160. Hietala, K., Forsblom, C., Summanen, P. & Groop, P. H. Heritability of proliferative diabetic retinopathy. Diabetes 57, 2176–2180 (2008).

161. Arar, N. H. et al. Heritability of the severity of diabetic retinopathy: the FIND-Eye study. Invest. Ophthalmol. Vis. Sci. 49, 3839–3845 (2008).

162. Wong, T. Y. et al. Diabetic retinopathy in a multi-ethnic cohort in the United States. Am. J. Ophthalmol. 141, 446–455 (2006).

163. Tong, Z. et al. Promoter polymorphism of the erythropoietin gene in severe diabetic eye and kidney complications. Proc. Natl Acad. Sci. USA 105, 6998–7003 (2008).

164. Chen, Y. et al. Assessing susceptibility to age-related macular degeneration with genetic markers and environmental factors. Arch. Ophthalmol. 129, 344–351 (2011).This is a good survey on the genetic and environmental factors that influence AMD.

165. Neale, B. M. et al. Genome-wide association study of advanced age-related macular degeneration identifies a role of the hepatic lipase gene (LIPC). Proc. Natl Acad. Sci. USA 107, 7395–7400 (2010).

166. Klaver, C. C. et al. Genetic association of apolipo-protein E with age-related macular degeneration. Am. J. Hum. Genet. 63, 200–206 (1998).

167. Bainbridge, J. W. et al. Effect of gene therapy on visual function in Leber’s congenital amaurosis. N. Engl. J. Med. 358, 2231–2239 (2008).This is one of the first studies to demonstrate the efficacy of gene therapy in patients with limited visual function owing to retinal disease.

168. Maguire, A. M. et al. Safety and efficacy of gene transfer for Leber’s congenital amaurosis. N. Engl. J. Med. 358, 2240–2248 (2008).This is one of the first studies to demonstrate the efficacy of gene therapy in patients with limited visual function owing to retinal disease.

169. Cideciyan, A. V. et al. Human gene therapy for RPE65 isomerase deficiency activates the retinoid cycle of vision but with slow rod kinetics. Proc. Natl Acad. Sci. USA 105, 15112–15117 (2008).

170. Deng, W. T. et al. Tyrosine-mutant AAV8 delivery of human MERTK provides long-term retinal preservation in RCS rats. Invest. Ophthalmol. Vis. Sci. 53, 1895–1904 (2012).

171. Hughes, A. E. et al. A common CFH haplotype, with deletion of CFHR1 and CFHR3, is associated with lower risk of age-related macular degeneration. Nature Genet. 38, 1173–1177 (2006).

172. Hageman, G. S. et al. Extended haplotypes in the complement factor H (CFH) and CFH-related (CFHR) family of genes protect against age-related macular degeneration: characterization, ethnic distribution and evolutionary implications. Ann. Med. 38, 592–604 (2006).

173. Du, H., Lim, S. L., Grob, S. & Zhang, K. Induced pluripotent stem cell therapies for geographic atrophy of age-related macular degeneration. Semin. Ophthalmol. 26, 216–224 (2011).

174. Zhang, K. & Ding, S. Stem cells and eye development. N. Engl. J. Med. 365, 370–372 (2011).This is a summary and review of eye development and its relevance to stem cells.

175. Kim, J. et al. Direct reprogramming of mouse fibroblasts to neural progenitors. Proc. Natl Acad. Sci. USA 108, 7838–7843 (2011).

176. Gouras, P., Kong, J. & Tsang, S. H. Retinal degeneration and RPE transplantation in Rpe65–/– mice. Invest. Ophthalmol. Vis. Sci. 43, 3307–3311 (2002).

177. Urtti, A. Challenges and obstacles of ocular pharmacokinetics and drug delivery. Adv. Drug Deliv. Rev. 58, 1131–1135 (2006).

178. Thrimawithana, T. R., Young, S., Bunt, C. R., Green, C. & Alany, R. G. Drug delivery to the posterior segment of the eye. Drug Discov. Today 16, 270–277 (2011).

179. Burnham, C. M. Encapsulated cell technology could prevent blindness. Drug Discov. Today 8, 146–147 (2003).

R E V I E W S



180. Tao, W. Application of encapsulated cell technology for retinal degenerative diseases. Expert Opin. Biol. Ther. 6, 717–726 (2006).

181. Sieving, P. A. et al. Ciliary neurotrophic factor (CNTF) for human retinal degeneration: Phase I trial of CNTF delivered by encapsulated cell intraocular implants. Proc. Natl Acad. Sci. USA 103, 3896–3901 (2006).

182. Zhang, R. et al. Intravitreal cell-based production of glucagon-like peptide-1. Retina 31, 785–789 (2011).

183. Petros, R. A. & DeSimone, J. M. Strategies in the design of nanoparticles for therapeutic applications. Nature Rev. Drug Discov. 9, 615–627 (2010).This is an excellent review on the rational design of therapeutic nanoparticles.

184. Shi, J., Votruba, A. R., Farokhzad, O. C. & Langer, R. Nanotechnology in drug delivery and tissue engineering: from discovery to applications. Nano Lett. 10, 3223–3230 (2010).

185. Sultana, Y., Maurya, D. P., Iqbal, Z. & Aqil, M. Nanotechnology in ocular delivery: current and future directions. Drugs Today 47, 441–455 (2011).

186. Zhang, L. et al. Nanoparticles in medicine: therapeutic applications and developments. Clin. Pharmacol. Ther. 83, 761–769 (2008).This is a good overview of the development and clinical approval of nanomedicines.

187. Yan, M. et al. A novel intracellular protein delivery platform based on single-protein nanocapsules. Nature Nanotechnol. 5, 48–53 (2010).

188. Gariano, R. F. & Gardner, T. W. Retinal angiogenesis in development and disease. Nature 438, 960–966 (2005).

189. Whitehead, K. A., Langer, R. & Anderson, D. G. Knocking down barriers: advances in siRNA delivery. Nature Rev. Drug Discov. 8, 129–138 (2009).

190. du Toit, L. C., Pillay, V., Choonara, Y. E., Govender, T. & Carmichael, T. Ocular drug delivery — a look towards nanobioadhesives. Expert Opin. Drug Deliv. 8, 71–94 (2011).

191. Hu, C.-M. J. et al. Erythrocyte membrane-camouflaged polymeric nanoparticles as a biomimetic delivery platform. Proc. Natl Acad. Sci. USA 108, 10980–10985 (2011).

192. Weinreb, R. N. & Lindsey, J. D. Metalloproteinase gene transcription in human ciliary muscle cells with latanoprost. Invest. Ophthalmol. Vis. Sci. 43, 716–722 (2002).

193. Hiett, J. A. & Dockter, C. A. Topical carbonic anhydrase inhibitors: a new perspective in glaucoma therapy. Optom. Clin. 2, 97–112 (1992).

194. Nilsson, S. F. et al. The prostanoid EP2 receptor agonist butaprost increases uveoscleral outflow in the cynomolgus monkey. Invest. Ophthalmol. Vis. Sci. 47, 4042–4049 (2006).

195. Krauss, A. H. et al. Ocular hypotensive activity of BOL-303259-X, a nitric oxide donating prostaglandin F2α agonist, in preclinical models. Exp. Eye Res. 93, 250–255 (2011).

196. Choudhri S., Wand, M. & Shields, M. B. A comparison of dorzolamide–timolol combination versus the concomitant drugs. Am. J. Ophthalmol. 130, 832–833 (2000).

197. Craven, E. R. et al. Brimonidine and timolol fixed-combination therapy versus montherapy: a 3-month randomized trial in patients with glaucoma or ocular hypertension. J. Ocul. Pharmacol. Ther. 21, 337–338 (2005).

198. Pfeiffer, N. & the German Latanoprost Fixed Combination Study Group. A comparison of the fixed combination of latanoprost and timolol with its individual components. Graefe’s Arch. Clin. Exp. Ophthalmol. 240, 893–899 (2002).

199. Higginbotham, E. J. et al. Latanoprost and timolol combination therapy vs monotherapy: one-year randomized trial. Arch. Ophthalmol. 120, 915–922 (2002).

200. Barneby, H. S. et al. The safety and efficacy of travoprost 0.004%/timolol 0.5% fixed combination ophthalmic solution. Am. J. Ophthalmol. 140, 1–7 (2005).

201. Schuman, J. S. et al. Efficacy and safety of a fixed combination of travoprost 0.004%/timolol 0.5% ophthalmic solution once daily for open-angle glaucoma or ocular hypertension. Am. J. Ophthalmol. 140, 242–250 (2005).

202. Hommer A & the Ganfort Investigators Group I. A double-masked, randomized, parallel comparison of a fixed combination of bimatoprost 0.03%/timolol

0.5% with non-fixed combination use in patients with glaucoma or ocular hypertension. Eur. J. Ophthalmol. 17, 53–62 (2007).

203. Rhéaume, M. A. & Vavvas, D. Pharmacologic vitreolysis. Semin. Ophthalmol. 25, 295–302 (2010).

204. Gandorfer, A. et al. Posterior vitreous detachment induced by microplasmin. Invest. Ophthalmol. Vis. Sci. 45, 641–647 (2004).

205. Pieramici, D. J. & Boyer, D. S. The phase III MIVI-TRUST clinical trial data: subgroup analysis of a single intravitreal injection of ocriplasmin in patients with full-thickness macular hole (Poster). In The Annu. Meeting of The Assoc. for Research in Vision and Ophthalmology (ARVO) (6–10 May 2012; Ft. Lauderdale, Florida, USA).

AcknowledgementsWe thank J. Ambati, S. Ding, I. Kozak, J. Lee, L. Zhao and P. Shaw for their helpful comments. K.Z. is supported by grants from the Chinese National 985 Project to Sichuan University and West China Hospital, the National Eye Institute and the US National Institutes of Health, VA Merit Award, Research to Prevent Blindness, King Abdulaziz City for Science and Technology (through the University of California San Diego Center of Excellence in Nanomedicine centre grant) and the BWF (Burroughs Wellcome Fund) Clinical Scientist Award in Translational Research. L.Z. is supported by the National Science Foundation (NSF) grants CMMI1031239 and DMR1216461; R.N.W. is supported by grants from the National Eye Institute and the US National Institutes of Health (EY019692) and an unrestricted grant from Research to Prevent Blindness, New York, USA. We apologize for the omission of references owing to page limitations.

Competing interests statementThe authors declare competing financial interests: see Web version for details.

FURTHER INFORMATIONKang Zhang’s homepage: http://eyesite.ucsd.edu/retina/KZL/index.html Taligen Therapeutics website: http://www.taligentherapeutics.com/pipeline/index.htmlWebvision website: http://webvision.med.utah.edu

ALL LINKS ARE ACTIVE IN THE ONLINE PDF

R E V I E W S



http://www.nature.com/nrd/journal/v11/n7/box/nrd3745_audecl.html


http://webvision.med.utah.edu

ophthalmic drug discovery: novel targets and mechanisms … · test visual function, ......

Documents