Rho-associated kinase inhibitors: A novel glaucoma therapy

https://doi.org/10.1016/j.preteyeres.2013.05.002Get rights and content

Abstract

The rho-associated kinase (ROCK) signaling pathway is activated via secreted bioactive molecules or via integrin activation after extracellular matrix binding. These lead to polymerization of actin stress fibers and formation of focal adhesions. Accumulating evidence suggests that actin cytoskeleton-modulating signals are involved in aqueous outflow regulation. Aqueous humor contains various biologically active factors, some of which are elevated in glaucomatous eyes. These factors affect aqueous outflow, in part, through ROCK signaling modulation.

Various drugs acting on the cytoskeleton have also been shown to increase aqueous outflow by acting directly on outflow tissue. In vivo animal studies have shown that the trabecular meshwork (TM) actin cytoskeleton in glaucomatous eyes is more disorganized and more randomly oriented than in non-glaucomatous control eyes. In a previous study, we introduced ROCK inhibitors as a potential glaucoma therapy by showing that a selective ROCK inhibitor significantly lowered rabbit IOP. Rho-associated kinase inhibitors directly affect the TM and Schlemm's canal (SC), differing from the target sight of other glaucoma drugs. The TM is affected earlier and more strongly than ciliary muscle cells by ROCK inhibitors, largely because of pharmacological affinity differences stemming from regulatory mechanisms. Additionally, ROCK inhibitors disrupt tight junctions, result in F-actin depolymerization, and modulate intracellular calcium level, effectively increasing SC-cell monolayer permeability.

Perfusion of an enucleated eye with a ROCK inhibitor resulted in wider empty spaces in the juxtacanalicular (JCT) area and more giant vacuoles in the endothelial cells of SC, while the endothelial lining of SC was intact. Interestingly, ROCK inhibitors also increase retinal blood flow by relaxing vascular smooth muscle cells, directly protecting neurons against various stresses, while promoting wound healing. These additional effects may help slow progressing visual field loss in glaucoma patients, making ROCK inhibitors an even more desirable anti-glaucoma agent. All evidence indicates that aqueous humor outflow is affected by cytoskeleton physiology and this information may provide valuable insight into understanding glaucoma pathology and treatment.

Introduction

Glaucoma is estimated to be the second leading cause of vision loss in the world, with an estimated 60.5 million people affected worldwide (Quigley and Broman, 2006). Despite the high prevalence, relatively little progress has been made in glaucoma management over the past several decades. Lowering intraocular pressure (IOP) is the classic treatment approach for glaucoma and is the only therapy that has been shown to be effective in large-scale clinical studies. Medications, mainly topical eye drops, are generally used first to reduce IOP. When these are insufficient, more invasive therapies, such as laser iridotomy and/or filtration surgeries, are performed. Unfortunately, the lowering of IOP following successful filtration surgery can be temporary because of excess scarring at the surgical site. Severe vision loss often results in these cases.

Glaucoma is a multifactorial disease and elevated IOP is not the only risk factor for the development and progression of glaucoma. Therefore, novel treatment strategies, such as protecting retinal ganglion cells or increasing retinal blood flow, have been explored. Memantine, a neuroprotectant that has shown promise in animal glaucoma models, only resulted in insignificant delays in glaucoma progression in clinical trials (Osborne, 2009). Still, this drug established the need and desire for glaucoma therapeutics that do not affect IOP.

In the study that first introduced Rho-associated kinase (ROCK) inhibitors as potential glaucoma therapeutics, we showed that the selective ROCK inhibitor Y-27632 (Uehata et al., 1997) lowered rabbit IOP (Honjo et al., 2001b). These compounds directly affect trabecular meshwork (TM, Fig. 1) and Schlemm's canal (SC) cells in vitro, which are different therapeutic targets than commercially available glaucoma drugs. Because of this, ROCK inhibitors have recently gained interest and are now the focus of several ongoing clinical trials. Moreover, accumulating evidence shows that ROCK inhibitors have favorable effects because of their ability to lower IOP in a novel way, but also because of neuroprotection. Interestingly, ROCK inhibitors also reduce tissue scarring after filtration surgery.

Section snippets

Tissue composition and function of aqueous outflow structures

Intraocular pressure depends on the balance between the inflow and outflow of aqueous humor. In glaucoma patients, IOP is elevated because of an increase in outflow resistance. Two aqueous humor outflow pathways exist in the eye. Aqueous mainly flows through the TM and SC to the episcleral vein, but an auxiliary uveoscleral pathway through the iris root and ciliary muscle exists, with fluid leaving the eye through the choroidal circulation or orbital tissues (Fig. 2). An increased resistance to

Molecular mechanisms by which ROCK inhibitors increase outflow facility

Most IOP-lowering drugs currently in clinical use suppress aqueous humor production (sympathomimetics, β-adrenergic receptor antagonists, carbonic anhydrase inhibitors) or enhance uveoscleral outflow (prostaglandin analogs; Brubaker, 2003). Pilocarpine, a miotic agent, increases conventional outflow through the TM and SC and also causes ciliary muscle contraction. This widens the spaces of the TM, thereby decreasing flow resistance and increasing aqueous humor outflow (Kaufman and Barany, 1976

Side effects of ROCK inhibitors

Rho kinase inhibitors can increase blood flow by inhibiting calcium sensitization and relaxing vascular smooth muscles (Uehata et al., 1997). From a therapeutic point of view, vasodilating conjunctival vessels would manifest as conjunctival hyperemia (Tanihara et al., 2008, Williams et al., 2011). This harmless cosmetic side effect does not affect vision, but would likely reduce patient satisfaction and compliance. To reduce the cosmetic implications of conjunctival hyperemia, a once-daily

Future directions

In this paper, we review the background, molecular mechanisms, and side effects of ROCK inhibitors. As stated in the opening paragraph, several clinical trials with ROCK inhibitors are currently underway. We reported that the topical ROCK inhibitor, Y-39983, when used twice daily, dramatically lowered IOP in healthy volunteers (Fig. 7; Tanihara et al., 2008). In addition, another ROCK inhibitor, AR-12286, safely lowered IOP in a recent clinical study (Williams et al., 2011), but approval for

Acknowledgments

This work was supported in part by JSPS KAKENHI Grant Number 23390403 and 23791994.

References (145)

  • Y. Itoh et al.

    Inhibition of RhoA/Rho-kinase pathway suppresses the expression of type I collagen induced by TGF-beta2 in human retinal pigment epithelial cells

    Exp. Eye Res.

    (2007)
  • M.A. Johnstone et al.

    Pressure-dependent changes in structures of the aqueous outflow system of human and monkey eyes

    Am. J. Ophthalmol.

    (1973)
  • B. Junglas et al.

    Connective tissue growth factor causes glaucoma by modifying the actin cytoskeleton of the trabecular meshwork

    Am. J. Pathol.

    (2012)
  • J. Kayes

    Pressure gradient changes on the trabecular meshwork of monkeys

    Am. J. Ophthalmol.

    (1975)
  • R.N. Khurana et al.

    The role of protein kinase C in modulation of aqueous humor outflow facility

    Exp. Eye Res.

    (2003)
  • Y. Kitaoka et al.

    Involvement of RhoA and possible neuroprotective effect of fasudil, a rho kinase inhibitor, in NMDA-induced neurotoxicity in the rat retina

    Brain Res.

    (2004)
  • A. Lepple-Wienhues et al.

    Differential smooth muscle-like contractile properties of trabecular meshwork and ciliary muscle

    Exp. Eye Res.

    (1991)
  • Z. Lu et al.

    The mechanism of increasing outflow facility by rho-kinase inhibition with Y-27632 in bovine eyes

    Exp. Eye Res.

    (2008)
  • O. Maepea et al.

    Pressures in the juxtacanalicular tissue and Schlemm's canal in monkeys

    Exp. Eye Res.

    (1992)
  • P.G. McMenamin et al.

    Effects of prolonged intracameral perfusion with mock aqueous humour on the morphology of the primate outflow apparatus

    Exp. Eye Res.

    (1986)
  • M. Miura et al.

    Critical role of the rho-kinase pathway in TGF-beta2-dependent collagen gel contraction by retinal pigment epithelial cells

    Exp. Eye Res.

    (2006)
  • E. Nakajima et al.

    Contribution of ROCK in contraction of trabecular meshwork: proposed mechanism for regulating aqueous outflow in monkey and human eyes

    J. Pharm. Sci.

    (2005)
  • T. Nishikimi et al.

    Molecular mechanisms and therapeutic strategies of chronic renal injury: renoprotective effect of rho-kinase inhibitor in hypertensive glomerulosclerosis

    J. Pharmacol. Sci.

    (2006)
  • J.P. Alexander et al.

    Involvement of the Erk-MAP kinase pathway in TNF-alpha regulation of trabecular matrix metalloproteinases and TIMPs

    Invest. Ophthalmol. Vis. Sci.

    (2003)
  • J.A. Alvarado et al.

    Endothelia of Schlemm's canal and trabecular meshwork: distinct molecular, functional, and anatomic features

    Am. J. Physiol. Cell Physiol.

    (2004)
  • J.A. Alvarado et al.

    Effect of beta-adrenergic agonists on paracellular width and fluid flow across outflow pathway cells

    Invest. Ophthalmol. Vis. Sci.

    (1998)
  • J.A. Alvarado et al.

    Interactions between endothelia of the trabecular meshwork and of Schlemm's canal: a new insight into the regulation of aqueous outflow in the eye

    Trans. Am. Ophthalmol. Soc.

    (2005)
  • J.J. Arnold et al.

    The effect of rho-associated kinase (ROCK) inhibition on the ocular penetration of timolol maleate

    Invest. Ophthalmol. Vis. Sci.

    (2013)
  • A. Bill et al.

    Scanning electron microscopic studies of the trabecular meshwork and the canal of Schlemm – an attempt to localize the main resistance to outflow of aqueous humor in man

    Acta Ophthalmol. (Copenh)

    (1972)
  • A.A. Birukova et al.

    Involvement of microtubules and Rho pathway in TGF-beta1-induced lung vascular barrier dysfunction

    J. Cell Physiol.

    (2005)
  • T. Borras et al.

    Gene transfer to the human trabecular meshwork by anterior segment perfusion

    Invest. Ophthalmol. Vis. Sci.

    (1998)
  • T. Borras et al.

    Adenoviral reporter gene transfer to the human trabecular meshwork does not alter aqueous humor outflow. Relevance for potential gene therapy of glaucoma

    Gene Ther.

    (1999)
  • J.M. Bradley et al.

    Effect of matrix metalloproteinases activity on outflow in perfused human organ culture

    Invest. Ophthalmol. Vis. Sci.

    (1998)
  • H.S. Brilakis et al.

    Giant vacuole survival time and implications for aqueous humor outflow

    J. Glaucoma

    (2001)
  • S.B. Campos et al.

    Cytokine-induced F-actin reorganization in endothelial cells involves RhoA activation

    Am. J. Physiol. Renal Physiol.

    (2009)
  • S. Cellek et al.

    A Rho-kinase inhibitor, soluble guanylate cyclase activator and nitric oxide-releasing PDE5 inhibitor: novel approaches to erectile dysfunction

    Expert Opin. Inves. Drugs

    (2002)
  • L. Choritz et al.

    Endothelin-1 concentration in aqueous humor correlates with intraocular pressure in primary open angle and pseudoexfoliation glaucoma

    Invest. Ophthalmol. Vis. Sci.

    (2012)
  • S.M. Chudgar et al.

    Regulation of connective tissue growth factor expression in the aqueous humor outflow pathway

    Mol. Vis.

    (2006)
  • A.F. Clark et al.

    Dexamethasone alters F-actin architecture and promotes cross-linked actin network formation in human trabecular meshwork tissue

    Cell Motil. Cytoskeleton

    (2005)
  • A.F. Clark et al.

    Cytoskeletal changes in cultured human glaucoma trabecular meshwork cells

    J. Glaucoma

    (1995)
  • T. Cumurcu et al.

    Aqueous humor erythropoietin levels in patients with primary open-angle glaucoma

    J. Glaucoma

    (2007)
  • A.W. de Kater et al.

    Localization of smooth muscle and nonmuscle actin isoforms in the human aqueous outflow pathway

    Invest. Ophthalmol. Vis. Sci.

    (1992)
  • A.W. de Kater et al.

    Localization of smooth muscle myosin-containing cells in the aqueous outflow pathway

    Invest. Ophthalmol. Vis. Sci.

    (1990)
  • D.L. Epstein et al.

    Influence of ethacrynic acid on outflow facility in the monkey and calf eye

    Invest. Ophthalmol. Vis. Sci.

    (1987)
  • D.L. Epstein et al.

    Morphology of the trabecular meshwork and inner-wall endothelium after cationized ferritin perfusion in the monkey eye

    Invest. Ophthalmol. Vis. Sci.

    (1991)
  • D.L. Epstein et al.

    Acto-myosin drug effects and aqueous outflow function

    Invest. Ophthalmol. Vis. Sci.

    (1999)
  • C.R. Ethier et al.

    Effects of latrunculin-B on outflow facility and trabecular meshwork structure in human eyes

    Invest. Ophthalmol. Vis. Sci.

    (2006)
  • Y. Feng et al.

    Discovery of substituted 4-(pyrazol-4-yl)-phenylbenzodioxane-2- carboxamides as potent and highly selective Rho kinase (ROCK-II) inhibitors

    J. Med. Chem.

    (2008)
  • M.S. Filla et al.

    Regulation of cross-linked actin network (CLAN) formation in human trabecular meshwork (HTM) cells by convergence of distinct beta1 and beta3 integrin pathways

    Invest. Ophthalmol. Vis. Sci.

    (2009)
  • M.S. Filla et al.

    Beta1 and beta3 integrins cooperate to induce syndecan-4-containing cross-linked actin networks in human trabecular meshwork cells

    Invest. Ophthalmol. Vis. Sci.

    (2006)
  • Cited by (127)

    • Induced pluripotent stem cells for modeling open-angle glaucoma

      2022, Novel Concepts in iPSC Disease Modeling
    View all citing articles on Scopus
    1

    Percentage of work contributed by each author in the production of the manuscript is as follows: Toshihiro Inoue, 80%; Hidenobu Tanihara, 20%.

    View full text