Update on the Mechanism of Action of Topical Prostaglandins for Intraocular Pressure Reduction Carol B

Home , Bimatoprost, Latanoprost, Prostaglandin E, Sulprostone, Tafluprost, Travoprost

SURVEY OF OPHTHALMOLOGY VOLUME 53 SUPPLEMENT 1 NOVEMBER 2008

Update on the Mechanism of Action of Topical Prostaglandins for Intraocular Pressure Reduction Carol B. Toris, PhD,1 B’Ann T. Gabelt, MS,2 and Paul L. Kaufman, MD2

1Department of Ophthalmology and Visual Sciences, University of Nebraska Medical Center, Omaha, Nebraska, USA; and 2Department of Ophthalmology and Visual Sciences, University of Wisconsin, Madison, Wisconsin, USA

Abstract. A decade has passed since the first topical prostaglandin analog was prescribed to reduce intraocular pressure (IOP) for the treatment of glaucoma. Now four prostaglandin analogs are available for clinical use around the world and more are in development. The three most efficacious of these drugs are latanoprost, travoprost, and bimatoprost, and their effects on IOP and aqueous humor dynamics are similar. A consistent finding is a substantial increase in uveoscleral outflow and a less consistent finding is an increase in trabecular outflow facility. Aqueous flow appears to be slightly stimulated as well. Prostaglandin receptors and their associated mRNAs have been located in the trabecular meshwork, ciliary muscle, and sclera, providing evidence that endogenous prostaglandins have a functional role in aqueous humor drainage. Earlier evidence found that topical PG analogs release endogenous prostaglandins. One well-studied mechanism for the enhancement of outflow by prostaglandins is the regulation of matrix metalloproteinases and remodeling of extracellular matrix. Other proposed mechanisms include widening of the connective tissue-filled spaces and changes in the shape of cells. All of these mechanisms alter the permeability of tissues of the outflow pathways leading to changes in outflow resistance and/or outflow rates. This review summarizes recent (since 2000) animal and clinical studies of the effects of topical prostaglandin analogs on aqueous humor dynamics and recent cellular and molecular studies designed to clarify the outflow effects. (Surv Ophthalmol 53:S107--S120, 2008. Ó 2008 Elsevier Inc. All rights reserved.)

Key words. aqueous humor intraocular pressure matrix metalloproteinases prostaglandin trabecular outﬂow uveoscleral outﬂow

All of the clinically available drugs for the treatment through the uveoscleral outflow pathway but signif- of elevated intraocular pressure (IOP) have direct icant effects on trabecular outflow facility also have 67,85 effects on one or more parameters of aqueous been reported. Three PGF2a analogs (bimato- humor dynamics. IOP usually is reduced by slowing prost, latanoprost, and travoprost) are approved for the production rate of aqueous humor, by de- glaucoma therapy in the United States, one addi- creasing the resistance to flow through the trabec- tional analog (unoprostone) is prescribed in Japan ular meshwork, by increasing drainage through the and a new analog (tafluprost) is in clinical trials in uveoscleral outflow pathway, or by a combination of Japan. Travoprost and latanoprost are ester pro- these mechanisms. Currently, the most effective drugs of PGF2a. Bimatoprost is the amide prodrug 86 outflow drugs approved for clinical use are prosta- of 17-phenyl-PGF2a and has been classified 86 glandin (PG) F2a analogs. These drugs reduce IOP by some as a prostamide, although this classifica- by stimulation of aqueous humor drainage primarily tion has been somewhat controversial.7,37,56--58,86,87

44 23 Unoprostone is an analog of a pulmonary metabo- agonists butaprost and 8-iso PGE2 (Table 1). 26 lite of PGF2a and sometimes labeled a docosanoid. Exceptions do exist. One drop of 17-phenyl trinor 8- Tafluprost is a difluoroprostaglandin derivative of iso PGE2 and the selective EP4 receptor agonist 3,7- 62 PGF2a. Agonists of DP, EP, and TP receptors have dithia PGE1 increased outflow facility sufficiently in been or are being investigated in animal models for normotensive monkey eyes to account for most, if their IOP efficacy and potential as new glaucoma not all of the IOP reduction (Kharlamb, ARVO 2004 therapeutic drugs. None of these agonists has yet to abstract 1035, Table 1). An older study found an be approved for clinical use. increase in outflow facility at 4 hours after one 35 This review summarizes recent (since 2000) topical drop of PGF2a. animal and clinical English-language studies of the One potential reason for the apparent differences effects of topical PG agonists on aqueous humor in trabecular outflow facility effect among the dynamics and recent cellular and molecular studies various PG agonists is the method of measurement. designed to clarify the outflow effects. Two noninvasive methods, tonography and fluorophotometry, and two invasive methods, two-level constant pressure perfusion and isotope accumula- Aqueous Humor Dynamics tion, are used to make the assessment. All methods measure trabecular outflow facility, but confound- Latanoprost, travoprost, and bimatoprost pro- ing factors are known to exist, including ocular duce similar increases in uveoscleral outflow of rigidity (a measure of eye stiffness), pseudofacility several-fold. Increases in trabecular outflow facility (the facility of flow of aqueous humor from the also have been reported but this finding has not posterior to anterior chamber resulting from the been found consistently (Table 1). Unoprostone, probe-induced increase in IOP), and uveoscleral the least efficacious of the four compounds, appears outflow facility. The name of the measured variable to have little effect on uveoscleral outflow in trabecular outflow facility is not entirely accurate humans. Rather it works mainly by increasing because of the inclusion of these other factors in trabecular outflow facility.69 The new fluoroprosta- the various measurements. All of the measurement glandin F , tafluprost, increases uveoscleral outflow 2a techniques assume that uveoscleral outflow facility is and aqueous flow in monkeys62 but has yet to be very small and affected little by the measurement studied in humans. Earlier studies13,89 have re- itself. This assumption is based on monkey stud- ported that the topical PG analogs release endoge- ies8,68 reporting that uveoscleral outflow is relatively nous prostaglandins that may contribute to the pressure-independent. However, if an experimental observed ocular hypotensive effects. Tafluprost in manipulation were to increase uveoscleral outflow mice reportedly works in part through FP receptor- facility, this could be interpreted erroneously, as an mediated prostaglandin production acting through increase in trabecular outflow facility. It is thought the prostanoid EP receptor.49 Studies published 3 by some that PGs increase uveoscleral outflow between 2000 and 2008 of FP, DP, and EP receptor facility but this has yet to be proven experimentally. agonists and their effects on aqueous humor These methods may not detect changes in trabecu- dynamics in humans and nonhuman primates are lar outflow facility if the changes are overshadowed summarized in Table 1. Studies predating 2000 are by strong effects on uveoscleral outflow and/or found in an earlier review.67 uveoscleral outflow facility. The length of time of treatment could be another TRABECULAR OUTFLOW FACILITY factor contributing to the differing effects of PGs on Trabecular outflow facility is not always increased trabecular outflow facility. The immediate IOP following topical treatment with PG analogs but effects that occur from a single dose of a PG analog evidence is building that the effect is real and not may be mediated by cellular mechanisms different unique to any one drug of this class. Latanoprost, from those that occur after repeated applications or travoprost, bimatoprost, and unoprostone all have continuous exposure.9,83,90 Therefore, findings been found to significantly increase trabecular from multiple doses of each PG should be com- outflow facility in at least one clinical study pared before concluding that one PG analog acts (Table 1). through mechanisms different from all others. Compared to humans, the trabecular meshwork Mice are being used with increasing frequency to of monkeys seems to be less affected by PG analogs. evaluate aqueous humor dynamics because of their Trabecular outflow facility was unchanged following potential for genetic manipulation in addition to multiple topical treatments of monkeys with PGF2a- their ocular similarities to humans. These animals isopropyl ester,22 tafluprost,62 and travoprost,71 the exhibit increases in outflow facility 2 hours after one DP receptor agonist AL-6598,72 and EP receptor 4-ml drop of latanoprost.18 Several limitations to OIA RSALNISADAUOSHMRDYNAMICS HUMOR AQUEOUS AND PROSTAGLANDINS TOPICAL

TABLE 1 Studies of Aqueous Humor Dynamics in Humans and Nonhuman Primates Treated with Prostaglandin Analogs Published from 2000 to 2008 Type and Number Duration of Analog of Subjects Treatment IOP Fa CPev Fu Reference FP receptor analogs, prostamides Bimatoprost ONT volunteers QD 2 days Y(day) [(day) [day [day Brubaker et al, 200111 (n 5 25) QD 3 days Y(night) [(night) OHT or POAG patients QD 7 days Y4 [ [Christiansen et al, (n 5 29) 200416 ONT volunteers QD 7 days Y4 [ [Lim et al, 200838 (n 5 30) Latanoprost OHT patients (n 5 30) QD 7 days Y4 44[Toris et al, 200173 OHT or POAG patients QD 14 days Y4 [ Dinslage et al, 200420 (n 5 30) ONT volunteers QD 7 days Y4 [ [Lim et al , 200838 (n 5 30) 15-keto latanoprost Cynomolgus, ONT One drop Y4 4 Wang et al, 200777 monkeys (n 5 30) Travoprost OHT & POAG patients QD 17 days Y(day and night) 4[(day) 4[(marginally Toris et al, 200770 (n 5 26) insigniﬁcant) ONT volunteers QD 7 days Y4 [ [Lim et al, 200838 (n 5 30) cynomolgus BID 3 days Y4 4 [a Toris et al, 200571 monkeys unilateral OHT (n 5 12) Unoprostone OHT patients (n 5 30) BID 5 days Y4 [44Toris et al, 200469 and 28 days Taﬂuprost cynomolgus monkeys QD 4 to 5 days Y[ 4 Takagi et al, 200462 (n 5 8--12) DP receptor agonist AL-6598 cynomolgus monkeys, BID 3 days YONT eye only [ONT eye only 4[ONT eye only Toris et al, 200672 unilateral OHT (n 5 11) EP2 receptor agonists Butaprost Cynomolgus monkeys, One drop YOHT and ONT 4One drop, 4One drop, ONT [QD 5 Nilsson et al, 200644 ONT and OHT or QD 5 days ONT days, ONT (n 5 6--8) 23 8-iso PGE2 Cynomolgus monkeys, BID, 9--29 doses Y44Two methods [ Gabelt et al, 2004

ONT (n 5 7--10) S109 (continued on next page) S110 Surv Ophthalmol 53 (Suppl 1) November 2008 TORIS ET AL

using this animal model should be mentioned. The no

5 size of the eye makes accurate measurement ocular difficult. Inflow and outflow are very slow (many- 4 5 fold slower than in humans),1 and changes in flow can be near the limit of detection. Additionally, the

Reference anesthesia needed for most measurements quickly and profoundly affects IOP.31 IOPs vary among abstract 4803) abstract 1035) strains of mice,30 and aqueous humor outﬂow rates Wang (ARVO 2007 Kharlamb (ARVO 2004

decreased effect; may vary among strains as well. Further research is 5

5 needed to characterize differences in aqueous Y

humor dynamics among the murine strains. u F QD Intraocular pressure; OHT days, ONT 4 5

once daily; UVEOSCLERAL OUTFLOW 5 ev Increases in uveoscleral outflow have been reported with topical treatment of bimatoprost and latanoprost in ocular normotensive and hypertensive subjects (Table 1).11,16,20,73 Travoprost increased CP uveoscleral outflow in monkeys71 and marginally increased it in ocular hypertensive patients as well.70 One drop, ONT uveoscleral outflow IOP

[ Unoprostone, the weakest of the four prescribed PG 5

u analogs, is the only one that did not affect uveoscleral outﬂow in humans despite 5 days of twice-daily dosing.69 a F Multiple topical drops of agonists for FP receptors

primary open angle glaucoma; QD 62 72

One (tafluprost ), DP receptors (AL-6598 ), and EP drop, ONT 2 5 44 78 4 receptors (butaprost and 8-iso PGE2 ) increased aqueous flow; F uveoscleral outflow in monkeys (Table 1). In 5

a contrast, one drop of the EP4 agonist 3,7-dithia PGE1 had no effect on uveoscleral outﬂow in

IOP monkeys (Kharlamb et al, unpublished data, abstract 1035 presented at the 2004 ARVO annual OHT and ONT,

Y4Y [ meeting). A multiple dose study is needed to clarify whether the effect persists with repeat dosing. 5 days

episcleral venous pressure; POAG AQUEOUS FLOW Treatment 5 Duration of or QD

ev Most studies investigating aqueous ﬂow have One drop One drop found that PG analogs produce a small (10--15%) increase that may or may not be statistically

6--8) signiﬁcant and is not clinically important. A

5 significant increase in aqueous flow was found at 6) night in young healthy Japanese volunteers treated 5 with latanoprost,41 and during the day and at night outflow facility determined by fluorophotometry; F of Subjects in healthy predominantly white volunteers treated

5 11 ocular normotension; P

Type and Number with bimatoprost. Additionally, aqueous ﬂow in- ﬂ ONT (n ONT and OHT (n 5

Cynomolgus monkeys, Cynomolgus monkeys, creased during the day in monkeys treated with a DP 72 ) agonist. This effect does not contribute to any 1 ONT reduction in IOP but an increase in aqueous ﬂow 5 increased effect. Blank cell indicates no data reported. 4 2 could be considered a healthy side effect of topical twice daily; C 5 continued PG analogs because aqueous humor carries essential 5 [ nutrients and removes waste products, crucial for keeping the avascular tissues of the anterior BID receptor agonist 3,7-dithia PGE 8-iso PGE hypertension Table 1 ( 17-phenyl trinor selective EP Analog effect; segment healthy. TOPICAL PROSTAGLANDINS AND AQUEOUS HUMOR DYNAMICS S111

EPISCLERAL VENOUS PRESSURE pig organ--cultured anterior segments perfused at Three studies have reported no change in a constant pressure of 15 mm Hg, showed increased episcleral venous pressure in patients with ocular outflow by up to 62% at 8 hours after topical hypertension treated for one week with latanoprost, administration of bimatoprost and by 30% at 5 74 travoprost, or unoprostone (Table 1). The measure- hours after intracameral administration of PGF2a. 6 ments (made with a commercially available epis- Human trabecular meshwork monolayer cultures cleral venomanometer, EyeTech, Morton Grove, IL), treated with bimatoprost had a 78% increase in hydraulic conductivity which also was blocked by are difficult to make with accuracy and consistency. 76 Nevertheless, because no study found any trend to AGN211334. PGE1 increases outflow facility by 26% in human suggest a change, it seems reasonable to conclude 19 that the PG effect on episcleral venous pressure is organ-cultured anterior segments. This effect minimal. probably occurs by an adenylyl cyclase-dependent pathway activated primarily by the predominant EP2 receptors in the trabecular meshwork.32 Stimulation Cellular and Molecular Studies of EP2 receptors in trabecular meshwork is coupled to the activation of high-conductance Caþ2-activated TRABECULAR MESHWORK Kþ channels (BK) which may contribute to the Organ-cultured anterior segments and trabecular relaxant activity of EP2 agonists in isolated trabec- meshwork cell cultures provide strong evidence that ular meshwork strips.61 However, outflow facility was PG analogs can alter the resistance in trabecular not stimulated and cAMP production was not outflow (Table 2). PGs have direct effects on matrix altered after short exposure periods with PGF2a or metalloproteinases (MMPs), neutral proteases that placebo in human organ-cultured anterior initiate degradation of the extracellular matrix and segments.19 play a major role in regulating resistance to flow Prostaglandin receptors have been identified in through tissues. MMPs are expressed by human human trabecular meshwork tissue but a prostamide trabecular meshwork2 and directly control outflow receptor has not yet been cloned. The genes for all resistance in human organ-cultured anterior seg- prostanoid receptors are expressed in human 9 ments. The activity of MMPs is regulated by tissue trabecular meshwork. Gene expression for the EP2 inhibitors of metalloproteinases (TIMPs).10 Cur- receptor is most abundant, followed by FP, TP, IP, 32 rently four TIMPS (TIMP-1, -2, -3, -4) have been and EP4, with DP and EP3 at the lowest levels. identified in mammals and each TIMP targets Immunofluorescent labeling of FP and EP prosta- specific MMPs.42 In one study,47 cultures of human noid receptor subtypes in normal human trabecular trabecular meshwork cells treated with latanoprost meshwork tissue showed positive staining for EP1 on acid for 24 hours had increased expression of MMPs trabecular cells of the outer portion of the 6 -1, -3, -17, and -24 and TIMPs -2, -3, and -4. A study meshwork and cells lining Schlemm’s canal. EP2 of human organ-cultured anterior segments infused was localized to the outer wall and periphery of with latanoprost acid found increased outflow Schlemm’s canal. EP3 and EP4 labeling was present facility at 24 hours after administration when on trabecular cells along the entire meshwork. compared to control anterior segments (67% vs Moderate expression of FP receptor protein was 6%) but no changes in the amount of MMPs or present in the outer portion of the trabecular scleral hydraulic conductivity.6 Histological exami- meshwork and endothelial cells of Schlemm’s canal, nation found regions of focal detachment and loss collector channels and aqueous veins.55 of Schlemm’s canal endothelial cells and extracel- Human trabecular meshwork cells in culture lular matrix in some areas of the trabecular express many of the same PG receptors as are found meshwork.6 The focal cell loss was reasoned to be in intact tissue. Cultured trabecular meshwork cells 6 83,84 due to cytoskeletal and focal adhesion changes can produce PGE2 and low levels of PGF2a. because PGs in aortic smooth muscle cells cause Additionally, FP receptors have been identified in disassembly of actin stress fibers and inhibition of human trabecular meshwork cells as determined by phosphorylation of paxillin and other focal adhe- the presence of mRNA, protein, and a functional sion proteins.12 response (increased inositol phosphate accumula- 5 Bimatoprost increases outflow facility by 40% in tion and intracellular calcium release) to PGF2a human organ-cultured anterior segments within 48 and numerous synthetic FP-selective PG agonists.58 hours of treatment. This effect was blocked by PGE2 elicits its biological effects via four G-protein- preincubation with AGN211334, thought to be coupled receptor subtypes which stimulate a prostamide-selective antagonist,76 or , alternatively, phosphoinositide turnover with elevation in intra- 7 an inhibitor of bimatoprost hydrolysis. Similarly, cellular-free calcium (EP1 and some EP3), activation TABLE 2 2008 November 1) (Suppl 53 Ophthalmol Surv S112 Molecular and Cellular Studies of the Effects of Prostaglandin-Related Compounds on the Aqueous Humor Outflow Pathways Cellular/Molecular Analog Animal/ Tissue Duration of Treatment Pathway/Effect Events Reference Trabecular meshwork 64 PGF2a, fluprostenol trabecular meshwork Up to 2 hours FP receptor mediated Thieme 2006 strips (bovine) inhibition of endothelin-1 contractility; no effect on baseline tension or carbachol-induced contraction 5 PGF2a human trabecular 1 hour FP mediated inositol Anthony 1998 meshwork cells phosphate accumulation; intracellular calcium release 19 PGF2a human anterior segments 60 minutes per dose outflow facility no change in cAMP Dijkstra 1999 unchanged production 36,37 PGF2a, butaprost human trabecular 6 hours upregulate mRNA for Liang 2003,2004 meshwork cells Nur77 and connective tissue growth factor Bimatoprost human anterior segments 48--72 hour continuous outflow facility increase Wan 200776 infusion Bimatoprost human trabecular 48--72 hour continuous hydraulic conductivity Wan 200776 meshwork monolayer infusion increase Bimatoprost human trabecular 6 hours no change in Nur77 Liang 2003, 200436,37 meshwork cells mRNA and connective tissue growth factor expression bimatoprost acid (B), human trabecular 1 hr (PI turnover); 3 min FP receptor activation: Sharif 200358 latanoprost acid (L), meshwork cells (Ca 2þ mobilization) stimulate travoprost acid (T), phosphoinositide unoprostone (U), turnover PGF2a (P) (B,L,T,U,P)stimulate intracellular Ca 2þ mobilization (T,U,P) 46 latanoprost acid, PGE1 human trabecular 24 hours increase mRNA for MMP- Oh 2006 meshwork cells 1,-3,-17,-24; TIMP-2,-3,-4 6

latanoprost acid; PGE1 human anterior segments 24 hours outflow facility increase; focal detachment and loss Bahler 2008 AL ET TORIS scleral hydraulic of Schlemm’s canal conductivity cells; extracellular unchanged matrix loss; no change in MMPs latanoprost acid human trabecular 9 days in vivo increase insulin growth Zhao 200390 meshwork cells factor-1 gene and fibroleukin gene expression OIA RSALNISADAUOSHMRDYNAMICS HUMOR AQUEOUS AND PROSTAGLANDINS TOPICAL Unoprostone human trabecular Up to 2 hours decrease activity of L-type Thieme 200565 meshwork cells Ca 2þ channels via tyrosine kinase pathways 19 PGE1 human anterior segments 60 minutes increase outflow facility increase cAMP Dijkstra 1999 production 61 AH13205 human trabecular Less than 10 minutes EP2 agonist activation of Stumpff 2005 meshwork cells BK calcium channels Uveoscleral tissues 53 PGF2a monkeys multiple topical decrease collagen types I, Sagara 1999 treatments III, IV in ciliary muscle 25 PGF2a monkeys multiple topical increase MMP-1,-2,-3 in Gaton 2001 treatments ciliary muscle 79 PGF2a monkeys multiple topical scleral permeability increase MMP-1,-2,-3 in Weinreb 2001 treatments sclera PGF2a human sclera explants collagen gelatinization Molik 2006 (ARVO 2006 abstract) 33 PGF2a, latanoprost human sclera 1--3 days increase scleral increase MMP-1,-2,-3 Kim 2001 permeability 82 PGF2a, PhXA85 scleral organ cultures 24 hr increase mRNA for MMPs Weinreb 2004 and TIMPx 28 PGF2a, latanoprost acid human ciliary muscle 5 min for ERK1/2, PKC; increase MMP-2 via Husain 2005 cells 4hr for MMP-2 analysis protein kinase C- and extracellular signal regulated protein kinase 1/2- dependent pathways 56 PGF2a, latanoprost acid, human ciliary muscle 1hr for PI turnover; 3 min increase mitogen Sharif 2003 bimatoprost acid, cells for Ca2þ mobilization; activated protein kinase travoprost acid 5 min for p42/p44 MAP activity; kinase phosphoinositide hydrolysis; intracellular calcium mobilization; inhibited by FP antagonist AL-8810 89 PGF2a, latanoprost, ciliary muscle tissue 5--10 min increase phospholipase Yousufzai 1996 PhXA85 (several species) A2 and release of arachidonic acid leading to formation of PGE2, PGD2 and PGF2a 36,37 PGF2a, butaprost human ciliary muscle 6 hours upregulate Nur77 and Liang 2003, 2004 cells connective tissue growth factor

(Continued on next page) S113 Table 2 (continued) 2008 November 1) (Suppl 53 Ophthalmol Surv S114 Cellular/Molecular Analog Animal/ Tissue Duration of Treatment Pathway/Effect Events Reference latanoprost acid human ciliary muscle 24 hours increase mRNA for MMP- Oh 200647 cells 3,-9,-17; increase mRNA for TIMP-3; decrease mRNA for MMP-1,-2,- 12,-14,-15,-16, TIMP-4 latanoprost acid human ciliary muscle 24 hours increase MMP-1,-3,-9 Weinreb 200281 cells latanoprost acid human ciliary muscle Up to 24 hours increase TIMP-1 Anthony 20024 cells latanoprost acid nonpigmented ciliary Up to 48 hr Cyclooxygenase-2 Hinz 200527 epithelial cells induction leading to increase MMP-1 latanoprost acid rats Single topical dose Initial IOP elevation Husain 200629 followed by prolonged IOP reduction latanoprost acid human ciliary muscle 9 days in vivo decrease aquaporin-1 and Zhao 200390 cells versican gene expression; decrease in mRNA for FP receptor latanoprost, bimatoprost, monkeys topical treatments for one tissue spaces of ciliary Richter 200352 sulprostone, AH13205 year muscle enlarged and organized; myelinated nerve fiber bundles present Bimatoprost human ciliary muscle 6 hours no change in Nur77 and Liang 2003, 200436,37 cells connective tissue growth factor expression; upregulation of Cyr61 3,7-dithia PGE1 ciliary muscle tissue No details -- probably less no EP4 mediated Kharlamb, 2006 (ARVO (human and monkey) than 2 hours relaxation 2006 abstract) Genetic studies latanoprost, travoprost, FP knockout mice single topical treatment IOP decrease FP receptor needed for Ota 200548 bimatoprost, IOP decrease unoprostone Latanoprost FP knockout mice 7 days topical sclera intact FP receptor gene Crowston 2007 (ARVO needed for 2007 abstract) upregulation of MMP- AL ET TORIS 2,-3,-9 48 latanoprost, travoprost, EP1,EP2,EP3 knockout single topical treatment IOP decrease EP1 and EP2 not involved Ota 2005 bimatoprost, mice in IOP decrease; EP3 unoprostone may have a role IOP 5 intraocular pressure; MMP 5 matrix metalloproteinase; PG 5 prostaglandin; PI 5 phospholipase C-mediated phosphoinositide; TIMP 5 tissue inhibitor of metalloproteinase. TOPICAL PROSTAGLANDINS AND AQUEOUS HUMOR DYNAMICS S115 of adenylyl cyclase and elevation of intracellular 2006 ARVO annual meeting), PGF2a and latano- 88 cAMP (EP2 and EP4) or inhibition of adenylyl prost did not. 17,43 cyclase (EP3). Prolonged treatment of human Remodeling of the extracellular matrix within the trabecular meshwork cells with latanoprost or PGF2a ciliary muscle and sclera is the most thoroughly ethanolamide causes an increase in expression of understood effect of PG treatment. Dissolution of genes for insulin-like growth factor-1 (IGF-1) and collagen types I and III within the connective tissue- fibroleukin that could act to increase outflow filled spaces between the outer longitudinally facility. IGF-1 is reported to increase the level of oriented muscle bundles39,63 results from PG-stim- MMPs, stromelysin, and gelatinase in trabecular ulated induction of enzymes MMP-1, -2, and -3 in meshwork cells. The protease activity of fibroleukin the ciliary muscle and surrounding sclera. 25 Long- may be active against a component in the extracel- term (1 year) unilateral treatment of monkey eyes lular matrix.90 with topical bimatoprost, latanoprost, sulprostone Unoprostone free acid (UF-021) shows low (EP3/EP1 agonist) or AH13205 and butaprost (EP2 affinity for all prostanoid receptors and weak agonists) found that in all cases the tissue spaces of functional responses via FP receptor activation.57 the ciliary muscle were enlarged and organized into Several molecular events have been associated with elongated tube-like spaces, covered by endothelial- unoprostone exposure. A reduction in the activity of like cells often in contact with the basement L-type Ca2þ channels via a signal transduction membrane, and contained myelinated nerve fiber pathway was mediated by tyrosine kinases.65 Activa- bundles.44,52 These fluid pathways resembled a kind tion of BK channels by unoprostone free acid of lymphatic system described in the choroid.34 inhibited trabecular meshwork contraction leading Changes in the trabecular meshwork also were to an increase in outflow.66 present. MMP expression in human ciliary body Endothelin-1 is involved in regulating the con- tissue and ciliary muscle cells was determined after tractility of the trabecular meshwork. FP receptor latanoprost acid treatment for 24 hours. The mRNA agonists can block endothelin-1 induced contractil- of MMP-1, -2, -3, -11, -12, -14, -15, -16, -17, -19, and ity of the trabecular meshwork. Evidence indicates -24 as well as TIMP-1 to -4 were found in ciliary body this inhibition is mediated by the FP receptor.64 tissue and ciliary muscle cells. Latanoprost increased MMP-3, -9, -17, and TIMP-3 and down-regulated MMP-1, -2, -12, -14, -15, and -16 and TIMP-4.46 UVEOSCLERAL TISSUES Latanoprost acid induced a concentration-depen- Immunohistochemistry studies of EP and FP dent increase in MMP-1, -3, and -9 gene transcrip- receptor localization in the uveoscleral tissue in tion81 and a concentration- and time-dependent normal human donor eyes indicate the presence of increase in TIMP-1 but not TIMP-2 mRNA and EP-1, -2, -3, -4 and FP receptors in the ciliary body protein.4 and sclera. FP receptors are most abundant in the Loss of cyclooxygenase (COX)-2 expression in the circular portion of the ciliary muscle.55 ciliary body of humans has been associated with Several mechanisms have been proposed to glaucoma.40 This association has led to studies explain the PG-induced increase in uveoscleral investigating the connection between PGs, COX-2 outflow: remodeling of the extracellular matrix of expression and MMPs. Indeed, the IOP-lowering the ciliary muscle45,53,80 and sclera33,82 (Molik et al, action of latanoprost appears to be associated with unpublished data, abstract 406 presented at the induction of COX-2 and subsequent MMP-1 expres- 2006 ARVO annual meeting) causing changes in the sion in human nonpigmented epithelial cells.27 permeability of these tissues; widening of the MMP-1 released into the aqueous humor would be connective tissue-filled spaces among the ciliary expected to flow into the ciliary muscle and through muscle bundles,39,63 which may be caused in part the trabecular meshwork and Schlemm’s canal to by relaxation of the ciliary muscle51,75 and changes potentially increase outflow via multiple routes. in the shape of ciliary muscle cells as a result of Studies to elucidate additional cellular mecha- alterations in actin and vinculin localization within nisms associated with PG-induced MMP secretion are 60 the cells. ongoing. PGF2a- and latanoprost-induced secretion Ciliary muscle relaxation has been suggested as and activation of MMP-2 in ciliary muscle cells were responsible for the initial reduction in IOP from shown to occur via protein kinase C and extracellular topical PGs. This does not appear to be the case for signal regulated protein kinase 1/2-dependent 28 all PGs and their agonists. PGE1 and PGE2 relaxed pathways. Mitogen-activated protein kinase activity isolated monkey ciliary muscle strips precontracted was stimulated in human ciliary muscle cells with 88 O O O with carbachol but 3,7-dithia PGE1 (Kharlamb et travoprost acid PGF2a latanoprost acid al, unpublished data, abstract 413 presented at the bimatoprost O latanoprost 5 bimatoprost acid. The S116 Surv Ophthalmol 53 (Suppl 1) November 2008 TORIS ET AL

FP antagonist AL-8810 completely inhibited the subtypes in mediating the IOP-lowering response to mitogen-activated protein kinase activity induced by clinical PG analogs. Studies in FP receptor--deficient bimatoprost and bimatoprost acid, indicating that mice have shown that the FP receptor is essential for both agonists were activating the FP receptor.57 the early IOP-lowering response to topical latano- Inhibition of the latanoprost-induced reduction prost, travoprost, bimatoprost, and unoprostone.48 of IOP in rats by thalidomide suggested that the The involvement of the FP receptor in the IOP IOP-lowering response is mediated, in part, through reduction with long-term dosing is unknown. tumor necrosis factor-a-dependent signaling path- Upregulation of MMP-2, -3, -9 and FP mRNA in ways. Treatment of human ciliary muscle cells with the sclera following 7 days of topical treatment with tumor necrosis factor-a increased the secretion of latanoprost also was dependent on an intact FP MMP-1, and -2 (Husain et al, unpublished data, receptor gene (Crowston et al, unpublished data, abstract 219 presented at the 2006 ARVO annual abstract 1551 presented at the 2007 ARVO annual meeting). meeting). EP receptor-deficient mice have been PGF2a can stimulate the formation of endogenous studied in similar ways. When EP1,EP2, and EP3 PGs by stimulation of phospholipase A2 and release receptor--deficient mice and their wild-type back- of arachidonic acid for PG synthesis.89 Human ground strain were treated topically with latano- ciliary muscle cells exposed to PGF2a ethanolamide prost, travoprost, bimatoprost, or unoprostone, it or latanoprost for 9 days show a downregulation of was found that EP3 receptors were involved in the the FP receptor. In the same study, downregulation IOP-lowering response to latanoprost, travoprost, of the aquaporin-1 and versican genes are proposed and bimatoprost at 3 hours after drug administra- 50 to increase flow through the ciliary muscle and tion but EP1 and EP2 receptors were not. decrease IOP.90 PG-induced changes in the sclera also are important in the regulation of uveoscleral outflow and may be used to enhance transscleral delivery of PHARMACOLOGIC DIFFERENCES AMONG THE peptides and other high-molecular-weight sub- PROSTAGLANDINS stances to the posterior segment of the eye. Five Natural prostaglandins (PGF2a, PGE2, PGD2, days of topical treatment with PGF2a-isopropyl ester PGI2) have the highest affinity for their respective increased MMP-1, -2, and -3 in the sclera of receptors (FP, EP, DP, IP) but are relatively non- monkeys.79 Immunocytochemistry studies and selective for these and other PG receptors (TP) and mRNA analysis of human sclera and cultured their subtypes (DP1,DP2,EP1--4) in receptor-binding human scleral fibroblasts showed the presence of studies.57 Prostamides are also naturally occurring EP1,EP2 and FP receptor subtypes but not EP3 and neutral lipids which have very little activity at 3 EP4 subtypes. Human scleral permeability to prostaglandin receptors but, thus far, only pharma- dextrans was measured in an Ussing chamber cologic evidences exists for a prostamide receptor.87 following exposure to PGF2a and latanoprost acid The therapeutic derivatives of PGF2a, either amide for 1--3 days. Scleral permeability increased in a dose- or ester prodrugs, are powerful ocular hypotensive and time-dependent manner. This was accompanied drugs and either mediate their effects primarily via by an increase in MMP concentration in the media FP receptor activation or potentially via some yet with the greatest increases in MMP-2 and -3 unidentified receptor activation.57 33 compared to MMP-1. PGF2a and latanoprost acid It has been reported that the prostamide bimato- also induced increases in mRNA for MMPs and prost stimulates neither FP nor EP2 receptors and its TIMPs in human scleral organ cultures.82 X-ray effects on aqueous humor outflow, although similar diffraction analysis of human scleral explants to latanoprost and travoprost, are accomplished via showed that incubation in PGF2a-containing media a receptor distinct from these pure FP receptor caused the collagen helix to undergo gelatinization agonists. However, bimatoprost acid (17-phenyl similar to what was found after incubation with PGF2a), which is found in the aqueous humor of MMP-enriched media (Molik et al, unpublished humans after topical application of bimatoprost data, abstract 406 presented at the 2006 ARVO (Dahlin et al, ARVO 2004 abstract 2096),14,15 annual meeting). exhibits a relatively high affinity for three different PG receptors (i.e., FP [Ki 5 83 nM], EP1 [Ki 5 95 nM], EP3 [Ki 5 387 nM]). Bimatoprost acid also exhibits functional activity (phosphoinositide turn- GENETIC STUDIES over) at the EP1 (EC50 5 2.7 nM) and FP (EC50 5 Mice deficient in various PG receptors have been 2.8--3.8 nM) receptors in most cell types.57 Bimato- used to determine the role of prostanoid receptor prost acid is a potent, non-selective PGF2a analog. TOPICAL PROSTAGLANDINS AND AQUEOUS HUMOR DYNAMICS S117

Unlike PGF2a or the EP2 agonist butaprost, including activation of multiple signaling pathways, bimatoprost did not upregulate orphan nuclear and increased expression of some factors and receptor (Nur77) or connective tissue growth factor downregulation of others. Genetic studies of knock- (CTNF) expression in human ciliary muscle cells or out mice treated with PGs have found that a re- trabecular meshwork. In addition, the FP antagonist duction in IOP requires intact FP and EP3 receptors. AL-8810 blocked the PGF2a-induced Nur77 mRNA Many questions remain unanswered but progress expression in human ciliary muscle cells and continues to be made. Prostamide antagonists have 76,87 trabecular meshwork indicating PGF2a-induced been described but this has raised new ques- Nur77 mRNA expression is via the activation of FP tions.7 A prostamide receptor needs to be cloned receptors.36,37 Bimatoprost induced the upregula- and its biosynthesis enzyme identified to conclude tion of Cyr61 (cysteine-rich angiogenic protein 61) that a unique prostamide-sensitive receptor exists. gene expression in cat iris and human ciliary muscle Further work is required to confirm that bimato- cells. Cry61 is involved in regulating extracellular prost acts through this receptor. Multiple-dosing matrix-associated signaling proteins and may be studies of each PG should be compared before a unique mechanism by which bimatoprost exerts its concluding that one PG analog acts through pharmacological action to lower IOP independent mechanisms different from all others. Live animal of Nur77 or CTNF.37 The importance of the and clinical studies are needed to support claims induction or lack of induction of these various made by in vitro experiments. Receptor subtype genes for IOP reduction remains to be determined. selectivity of topical PGs should be identified. The The production of transgenic mice lacking these importance of induction or lack of induction of genes and their IOP responses to PGF2a, bimato- various genes for IOP reduction needs to be prost, and butaprost is needed. clarified. One day, the current research may lead Bimatoprost appears to reduce the IOP of to future new drugs that exceed the utility of the 24 patients who are unresponsive to latanoprost, PGF2a analogs. suggesting that the prostamide bimatoprost and the FP receptor agonist latanoprost stimulate different receptor populations. This is consistent Method of Literature Search with studies on isolated iridial cells where bimato- These studies, dating between 2000 and 2008 prost stimulated an entirely different cell popula- were found from a series of literature searches of tion to those sensitive to PGF2a and bimatoprost 59 PubMed and from the reference lists of these acid (17-phenyl PGF2a). An equally plausible articles. The searches included the following terms explanation is that some eyes may be deficient in in various combinations: anterior segment organ corneal esterase and thus are not able to adequately culture, aqueous flow, aqueous humor dynamics, bimato- convert the prodrug latanoprost into its free acid 21 prost, ciliary muscle, DP receptor, EP receptor, fluoropho- active form. Also splice variants of the FP receptor tometry, FP receptor, latanoprost, matrix metalloproteinase, may exist that have not yet been discovered. Single monkey, ocular, outflow facility, prostaglandins, prosta- nucleotide polymorphisms in the promoter and mide, prostanoid, tafluprost, tonography, travoprost, intron 1 regions of the FP receptor gene are unoprostone, uveoscleral outflow. Original research correlated with the variability in the IOP-lowering 54 articles, review articles, and meeting abstracts are response to latanoprost in normal human eyes. included in this review.

Summary References Clinical and animal studies of aqueous humor 1. Aihara M, Lindsey JD, Weinreb RN: Aqueous humor dynamics in mice. Invest Ophthalmol Vis Sci 44:5168--73, dynamics have reported that PG analogs effectively 2003 reduce IOP by enhancing aqueous humor outflow. 2. Alexander JP, Samples JR, Van Buskirk EM, et al: Expression The relative effects of PG analogs on each of the two of matrix metalloproteinases and inhibitor by human trabecular meshwork. Invest Ophthalmol Vis Sci 32:172-- outflow pathways may vary, but it appears that they 80, 1991 reduce IOP predominantly by increasing uveoscleral 3. Anthony TL, Lindsey JD, Aihara M, et al: Detection of outflow and to a lesser extent trabecular outflow prostaglandin EP(1), EP(2), and FP receptor subtypes in human sclera. Invest Ophthalmol Vis Sci 42:3182--6, 2001 facility. Morphological studies have identified PG 4. Anthony TL, Lindsey JD, Weinreb RN: Latanoprost’s effects receptors and described significant cellular changes on TIMP-1 and TIMP-2 expression in human ciliary muscle in PG-treated tissues of both outflow pathways. cells. Invest Ophthalmol Vis Sci 43:3705--11, 2002 5. Anthony TL, Pierce KL, Stamer WD, et al: Prostaglandin F2 Biochemical studies have reported many complex alpha receptors in the human trabecular meshwork. Invest cellular events in PG-treated outflow tissues, Ophthalmol Vis Sci 39:315--21, 1998 S118 Surv Ophthalmol 53 (Suppl 1) November 2008 TORIS ET AL

6. Bahler CK, Howell KG, Hann CR, et al: Prostaglandins 27. Hinz B, Ro¨sch S, Ramer R, et al: Latanoprost induces matrix increase trabecular meshwork outflow facility in cultured metalloproteinase-1 expression in human nonpigmented human anterior segments. Am J Ophthalmol 145:114--9, ciliary epithelial cells through a cyclooxygenase-2-dependent 2008 mechanism. FASEB J 19:1929--31, 2005 7. Bean GW, Camras CB, et al: Commercially Available 28. Husain S, Jafri F, Crosson CE: Acute effects of PGF2alpha on Prostaglandin Analogs for the Reduction of Intraocular MMP-2 secretion from human ciliary muscle cells: a PKC- Pressure -- Similarities and Differences. Surv Ophthalmol and ERK-dependent process. Invest Ophthalmol Vis Sci 46: 53(Suppl 1):S69--S84, 2008 1706--13, 2005 8. Bill A: Further studies on the influence of the intraocular 29. Husain S, Whitlock NA, Rice DS, et al: Effects of latanoprost pressure on aqueous humor dynamics in cynomolgus on rodent intraocular pressure. Exp Eye Res 83:1453--8, monkeys. Invest Ophthalmol 6:364--72, 1967 2006 9. Bradley JM, Vranka J, Colvis CM, et al: Effect of matrix 30. John SW, Hagaman JR, MacTaggart TE, et al: Intraocular metalloproteinases activity on outflow in perfused human pressure in inbred mouse strains. Invest Ophthalmol Vis Sci organ culture. Invest Ophthalmol Vis Sci 39:2649--58, 1998 38:249--53, 1997 10. Brew K, Dinakarpandian D, Nagase H: Tissue inhibitors of 31. Johnson TV, Fan S, Toris CB: Rebound tonometry in metalloproteinases: evolution, structure and function. Bio- conscious, conditioned mice avoids the acute and profound chim Biophys Acta 1477:267--83, 2000 effects of anesthesia on intraocular pressure. J Ocul 11. Brubaker RF, Schoff EO, Nau CB, et al: Effects of AGN Pharmacol Ther 24:175--85, 2008 192024, a new ocular hypotensive agent, on aqueous 32. Kamphuis W, Schneemann A, van Beek LM, et al: Prosta- dynamics. Am J Ophthalmol 131:19--24, 2001 noid receptor gene expression profile in human trabecular 12. Bulin C, Albrecht U, Bode JG, et al: Differential effects of meshwork: a quantitative real-time PCR approach. Invest vasodilatory prostaglandins on focal adhesions, cytoskeletal Ophthalmol Vis Sci 42:3209--15, 2001 architecture, and migration in human aortic smooth muscle 33. Kim JW, Lindsey JD, Wang N, et al: Increased human scleral cells. Arterioscler Thromb Vasc Biol 25:84--9, 2005 permeability with prostaglandin exposure. Invest Ophthal- 13. Camras CB, Podos SM: The role of endogenous prostaglan- mol Vis Sci 42:1514--21, 2001 din in clinically-used and investigational glaucoma therapy, 34. Krebs W, Krebs IP: Ultrastructural evidence for lymphatic in Bito LZ, Stjernschantz J: The Ocular Effects of Prosta- capillaries in the primate choroid. Arch Ophthalmol 106: glandins and Other Eicosanoids. New York, NY, Alan R. Liss, 1615--6, 1988 Inc, 1989. pp 459--75 35. Lee PY, Podos SM, Severin C: Effect of prostaglandin F2 14. Camras CB, Toris CB, Sjoquist B, et al: Detection of the free alpha on aqueous humor dynamics of rabbit, cat, and acid of bimatoprost in aqueous humor samples from human monkey. Invest Ophthalmol Vis Sci 25:1087--93, 1984 eyes treated with bimatoprost before cataract surgery. 36. Liang Y, Li C, Guzman VM, et al: Upregulation of orphan Ophthalmology 111:2193--8, 2004 nuclear receptor Nur77 following PGF(2alpha), Bimato- 15. Cantor LB, Hoop J, Wudunn D, et al: Levels of bimatoprost prost, and Butaprost treatments. Essential role of a protein acid in the aqueous humour after bimatoprost treatment of kinase C pathway involved in EP(2) receptor activated patients with cataract. Br J Ophthalmol 91:629--32, 2007 Nur77 gene transcription. Br J Pharmacol 142:737--48, 2004 16. Christiansen GA, Nau CB, McLaren JW, et al: Mechanism of 37. Liang Y, Li C, Guzman VM, et al: Comparison of ocular hypotensive action of bimatoprost (Lumigan) in prostaglandin F2alpha, bimatoprost (prostamide), and patients with ocular hypertension or glaucoma. Ophthal- butaprost (EP2 agonist) on Cyr61 and connective tissue mology 111:1658--62, 2004 growth factor gene expression. J Biol Chem 278:27267--77, 17. Coleman RA, Grix SP, Head SA, et al: A novel inhibitory 2003 prostanoid receptor in piglet saphenous vein. Prostaglan- 38. Lim KS, Nau CB, O’Byrne MM, et al: Mechanism of action of dins 47:151--68, 1994 bimatoprost, latanoprost, and travoprost in healthy subjects. 18. Crowston JG, Aihara M, Lindsey JD, et al: Effect of A crossover study. Ophthalmology 115:790--5. e4, 2008 latanoprost on outflow facility in the mouse. Invest 39. Lu¨tjen-Drecoll E, Tamm E: Morphological study of the Ophthalmol Vis Sci 45:2240--5, 2004 anterior segment of cynomolgus monkey eyes following 19. Dijkstra BG, Schneemann A, Hoyng PF: Flow after prosta- treatment with prostaglandin F2 alpha. Exp Eye Res 47:761-- glandin E1 is mediated by receptor-coupled adenylyl cyclase 9, 1988 in human anterior segments. Invest Ophthalmol Vis Sci 40: 40. Maiho¨fner C, Schlo¨tzer-Schrehardt U, Gu¨hring H, et al: 2622--6, 1999 Expression of cyclooxygenase-1 and -2 in normal and 20. Dinslage S, Hueber A, Diestelhorst M, et al: The influence of glaucomatous human eyes. Invest Ophthalmol Vis Sci 42: Latanoprost 0.005% on aqueous humor flow and outflow 2616--24, 2001 facility in glaucoma patients: a double-masked placebo- 41. Mishima HK, Kiuchi Y, Takamatsu M, et al: Circadian controlled clinical study. Graefes Arch Clin Exp Ophthalmol intraocular pressure management with latanoprost: diurnal 242:654--60, 2004 and nocturnal intraocular pressure reduction and increased 21. Eisenberg D: Latanoprost versus bimatoprost. Ophthalmol- uveoscleral outflow. Surv Ophthalmol 41(Suppl 2):S139--44, ogy 110:2003, 1861--2; author reply, 1862 1997 22. Gabelt BT, Kaufman PL: The effect of prostaglandin F2 42. Murphy G, Docherty AJ: The matrix metalloproteinases and alpha on trabecular outflow facility in cynomolgus monkeys. their inhibitors. Am J Respir Cell Mol Biol 7:120--5, 1992 Exp Eye Res 51:87--91, 1990 43. Negishi M, Sugimoto Y, Irie A, et al: Two isoforms of 23. Gabelt BT, Seeman JL, Podos SM, et al: Aqueous humor prostaglandin E receptor EP3 subtype. Different COOH- dynamics in monkeys after topical 8-iso PGE(2). Invest terminal domains determine sensitivity to agonist-induced Ophthalmol Vis Sci 45:892--9, 2004 desensitization. J Biol Chem 268:9517--21, 1993 24. Gandolfi SA, Cimino L: Effect of bimatoprost on patients 44. Nilsson SF, Drecoll E, Lu¨tjen-Drecoll E, et al: The prostanoid with primary open-angle glaucoma or ocular hypertension EP2 receptor agonist butaprost increases uveoscleral outflow who are nonresponders to latanoprost. Ophthalmology 110: in the cynomolgus monkey. Invest Ophthalmol Vis Sci 47: 609--14, 2003 4042--9, 2006 25. Gaton DD, Sagara T, Lindsey JD, et al: Increased matrix 45. Ocklind A: Effect of latanoprost on the extracellular matrix metalloproteinases 1, 2, and 3 in the monkey uveoscleral of the ciliary muscle. A study on cultured cells and tissue outflow pathway after topical prostaglandin F(2 alpha)- sections. Exp Eye Res 67:179--91, 1998 isopropyl ester treatment. Arch Ophthalmol 119:1165--70, 46. Oh DJ, Martin JL, Williams AJ, et al: Analysis of expression of 2001 matrix metalloproteinases and tissue inhibitors of metal- 26. Haria M, Spencer CM: Unoprostone (isopropyl unopro- loproteinases in human ciliary body after latanoprost. Invest stone). Drugs Aging 9:213--8, discussion 219--20, 1996 Ophthalmol Vis Sci 47:953--63, 2006 TOPICAL PROSTAGLANDINS AND AQUEOUS HUMOR DYNAMICS S119

47. Oh DJ, Martin JL, Williams AJ, et al: Effect of latanoprost on 65. Thieme H, Steinhausen K, Ottlecz A, et al: Effects of the expression of matrix metalloproteinases and their tissue unoprostone and endothelin 1 on L-type channel currents inhibitors in human trabecular meshwork cells. Invest in human trabecular meshwork cells. Ophthalmic Res 37: Ophthalmol Vis Sci 47:3887--95, 2006 293--300, 2005 48. Ota T, Aihara M, Narumiya S, et al: The effects of 66. Thieme H, Stumpff F, Ottlecz A, et al: Mechanisms of action prostaglandin analogues on IOP in prostanoid FP-recep- of unoprostone on trabecular meshwork contractility. Invest tor-deficient mice. Invest Ophthalmol Vis Sci 46:4159--63, Ophthalmol Vis Sci 42:3193--201, 2001 2005 67. Toris CB, Camras CB, Yablonski ME, et al: Effects of 49. Ota T, Aihara M, Saeki T, et al: The IOP-lowering effects and exogenous prostaglandins on aqueous humor dynamics mechanism of action of tafluprost in prostanoid receptor- and blood-aqueous barrier function. Surv Ophthalmol deficient mice. Br J Ophthalmol 91:673--6, 2007 41(Suppl 2):S69--75, 1997 50. Ota T, Aihara M, Saeki T, et al: The effects of 68. Toris CB, Pederson JE: Effect of intraocular pressure on prostaglandin analogues on prostanoid EP1, EP2, and uveoscleral outflow following cyclodialysis in the monkey EP3 receptor-deficient mice. Invest Ophthalmol Vis Sci 47: eye. Invest Ophthalmol Vis Sci 26:1745--9, 1985 3395--9, 2006 69. Toris CB, Zhan G, Camras CB: Increase in outflow facility 51. Poyer JF, Millar C, Kaufman PL: Prostaglandin F2 alpha with unoprostone treatment in ocular hypertensive patients. effects on isolated rhesus monkey ciliary muscle. Invest Arch Ophthalmol 122:1782--7, 2004 Ophthalmol Vis Sci 36:2461--5, 1995 70. Toris CB, Zhan G, Fan S, et al: Effects of travoprost on 52. Richter M, Krauss AH, Woodward DF, et al: Morphological aqueous humor dynamics in patients with elevated in- changes in the anterior eye segment after long-term traocular pressure. J Glaucoma 16:189--95, 2007 treatment with different receptor selective prostaglandin 71. Toris CB, Zhan GL, Camras CB, et al: Effects of travoprost agonists and a prostamide. Invest Ophthalmol Vis Sci 44: on aqueous humor dynamics in monkeys. J Glaucoma 14: 4419--26, 2003 70--3, 2005 53. Sagara T, Gaton DD, Lindsey JD, et al: Topical prostaglandin 72. Toris CB, Zhan GL, Feilmeier MR, et al: Effects of F2alpha treatment reduces collagen types I, III, and IV in a prostaglandin DP receptor agonist, AL-6598, on aqueous the monkey uveoscleral outflow pathway. Arch Ophthalmol humor dynamics in a nonhuman primate model of 117:794--801, 1999 glaucoma. J Ocul Pharmacol Ther 22:86--92, 2006 54. Sakurai M, Higashide T, Takahashi M, et al: Association 73. Toris CB, Zhan GL, Zhao J, et al: Potential mechanism for between genetic polymorphisms of the prostaglandin the additivity of pilocarpine and latanoprost. Am J Oph- F2alpha receptor gene and response to latanoprost. thalmol 131:722--8, 2001 Ophthalmology 114:1039--45, 2007 74. Vaajanen A, Vapaatalo H, Oksala O: A modified in vitro 55. Schlotzer-Schrehardt U, Zenkel M: Nu¨sing RM: Expression method for aqueous humor outflow studies in enucleated and localization of FP and EP prostanoid receptor subtypes porcine eyes. J Ocul Pharmacol Ther 23:124--31, 2007 in human ocular tissues. Invest Ophthalmol Vis Sci 43:1475-- 75. Van Alphen GWHM, Wilhelm PB, Elsenfeld PW: The effect 87, 2002 of prostaglandins on the isolated internal muscles of the 56. Sharif NA, Crider JY, Husain S, et al: Human ciliary muscle mammalian eye, including man. Doc Ophthalmol 42:397-- cell responses to FP-class prostaglandin analogs: phosphoi- 415, 1977 nositide hydrolysis, intracellular Ca2þ mobilization and 76. Wan Z, Woodward DF, Cornell CL, et al: Bimatoprost, MAP kinase activation. J Ocul Pharmacol Ther 19:437--55, prostamide activity, and conventional drainage. Invest 2003 Ophthalmol Vis Sci 48:4107--15, 2007 57. Sharif NA, Kelly CR, Crider JY, et al: Ocular hypotensive FP 77. Wang RF, Gagliuso DJ, Mittag TW, et al: Effect of 15-keto prostaglandin (PG) analogs: PG receptor subtype binding latanoprost on intraocular pressure and aqueous humor affinities and selectivities, and agonist potencies at FP and dynamics in monkey eyes. Invest Ophthalmol Vis Sci 48: other PG receptors in cultured cells. J Ocul Pharmacol Ther 4143--7, 2007 19:501--15, 2003 78. Wang RF, Lee PY, Mittag TW, et al: Effect of 8-iso 58. Sharif NA, Kelly CR, Crider JY: Human trabecular meshwork prostaglandin E2 on aqueous humor dynamics in monkeys. cell responses induced by bimatoprost, travoprost, unopro- Arch Ophthalmol 116:1213--6, 1998 stone, and other FP prostaglandin receptor agonist ana- 79. Weinreb RN: Enhancement of scleral macromolecular logues. Invest Ophthalmol Vis Sci 44:715--21, 2003 permeability with prostaglandins. Trans Am Ophthalmol 59. Spada CS, Krauss AH, Woodward DF, et al: Bimatoprost and Soc 99:319--43, 2001 prostaglandin F(2 alpha) selectively stimulate intracellular 80. Weinreb RN, Kashiwagi K, Kashiwagi F, et al: Prostaglandins calcium signaling in different cat iris sphincter cells. Exp increase matrix metalloproteinase release from human Eye Res 80:135--45, 2005 ciliary smooth muscle cells. Invest Ophthalmol Vis Sci 38: 60. Stjernschantz J, Seleń G, Ocklind A, Resul B: Weinreb RN: 2772--80, 1997 Uveoscleral Outflow. Biology and Clinical Aspects: Effects of 81. Weinreb RN, Lindsey JD: Metalloproteinase gene transcrip- latanoprost and related prostaglandin analogues, in Alm A. tion in human ciliary muscle cells with latanoprost. Invest London, UK, Mosby International Limited, 1998. pp Ophthalmol Vis Sci 43:716--22, 2002 57--72 82. Weinreb RN, Lindsey JD, Marchenko G, et al: Prostaglandin 61. Stumpff F, Boxberger M, Krauss A, et al: Stimulation of FP agonists alter metalloproteinase gene expression in cannabinoid (CB1) and prostanoid (EP2) receptors opens sclera. Invest Ophthalmol Vis Sci 45:4368--77, 2004 BKCa channels and relaxes ocular trabecular meshwork. 83. Weinreb RN, Mitchell MD, Polansky JR: Prostaglandin Exp Eye Res 80:697--708, 2005 production by human trabecular cells: in vitro inhibition 62. Takagi Y, Nakajima T, Shimazaki A, et al: Pharmacological by dexamethasone. Invest Ophthalmol Vis Sci 24:1541--5, characteristics of AFP-168 (tafluprost), a new prostanoid FP 1983 receptor agonist, as an ocular hypotensive drug. Exp Eye Res 84. Weinreb RN, Polansky JR, Alvarado JA, et al: Arachidonic 78:767--76, 2004 acid metabolism in human trabecular meshwork cells. Invest 63. Tamm E, Rittig M: Lu¨tjen-Drecoll E: [Electron microscopy Ophthalmol Vis Sci 29:1708--12, 1988 and immunohistochemical studies of the intraocular pres- 85. Weinreb RN, Toris CB, Gabelt BT, et al: Effects of sure lowering effect of prostaglandin F2 alpha]. Fortschr prostaglandins on the aqueous humor outflow pathways. Ophthalmol 87:623--9, 1990 Surv Ophthalmol 47(Suppl 1):S53--64, 2002 64. Thieme H, Schimmat C, Mu¨nzer G, et al: Endothelin 86. Woodward DF, Krauss AH, Chen J, et al: Pharmacological antagonism: effects of FP receptor agonists prostaglandin characterization of a novel antiglaucoma agent, Bimato- F2alpha and fluprostenol on trabecular meshwork contrac- prost (AGN 192024). J Pharmacol Exp Ther 305:772--85, tility. Invest Ophthalmol Vis Sci 47:938--45, 2006 2003 S120 Surv Ophthalmol 53 (Suppl 1) November 2008 TORIS ET AL

87. Woodward DF, Liang Y, Krauss AH: Prostamides (prosta- The supplement in which this article is published was glandin-ethanolamides) and their pharmacology. Br J funded by Pﬁzer. The authors reported no proprietary or Pharmacol 153:410--9, 2008 commercial interest in any product mentioned or concept 88. Yamaji K, Yoshitomi T, Ishikawa H, et al: Prostaglandins E1 discussed in this article. The article was supported in part by and E2, but not F2alpha or latanoprost, inhibit monkey unrestricted grants from Research to Prevent Blindness, Inc., ciliary muscle contraction. Curr Eye Res 30:661--5, 2005 New York, NY (C.T., P.K.), NIH/NEI EY002698 (P.K.); NEI P30 89. Yousufzai SY, Ye Z: Abdel-Latif AA: Prostaglandin F2 alpha EY016665 (Core Grant for Vision Research) (P.K.); unre- and its analogs induce release of endogenous prostaglan- stricted departmental and Physician-Scientist awards (P.K.); dins in iris and ciliary muscles isolated from cat and other Ocular Physiology Research and Education Foundation (P.K.); mammalian species. Exp Eye Res 63:305--10, 1996 and the Walter Helmerich Chair from the Retina Research 90. Zhao X, Pearson KE, Stephan DA, et al: Effects of Foundation (P.K.). prostaglandin analogues on human ciliary muscle and Reprint address: Carol B. Toris, PhD, Department of trabecular meshwork cells. Invest Ophthalmol Vis Sci 44: Ophthalmology, 985840 Nebraska Medical Center, Omaha, NE 1945--52, 2003 68198-5840. e-mail: [email protected].