DRUG DELIVERY BY COLLAGEN SHIELDS: EXPERIMENTAL STUDIES

III. DRUG DELIVERY BY COLLAGEN SHIELDS: EXPERIMENTAL STUDIES

A variety of studies have described the pharmacokinetics of ocular delivery of dyes and drugs by collagen shields as well as the use of the shields in the chemotherapy of various disorders. These studies are reviewed below and summarized in Tables 2 and 3 (13,14).

A. Fluorescein, a Water-Soluble Dye To determine the ocular penetration of water-soluble compounds delivered

by collagen shields, Reidy et al. (15) applied shields hydrated in a solution of

Corneal Collagen Shields for Ocular Drug Delivery 313 sodium fluorescein to normal eyes of volunteers and measured the fluores-

cence in the anterior chamber by fluorophotometry. The shields delivered significantly larger amounts of dye to the aqueous humor at 2 and 4 hours compared with drops of the same concentration instilled every 30 minutes over 4 hours, as well as in comparison with daily wear soft contact lenses presoaked in 0.01% fluorescein. The collagen shields did not induce any damage to the corneal epithelium over a 2-hour period. These results demonstrate that the collagen shield is superior to topical drops and some soft contact lenses in delivering fluorescein to the cornea and aqueous humor. The collagen shields might also successfully deliver other water- soluble compounds, such as antibiotics, to the eye in amounts comparable to or greater than the amounts delivered by drops over the same period of time.

B. Antibacterial Agents Ideally, chemotherapy for bacterial keratitis would delivery antibiotics

rapidly to both the cornea and aqueous humor, produce concentrations of antibiotic significantly above the minimum inhibitory concentration (MIC) or minimum bactericidal concentration (MBC) of ocular pathogens, and sustain this high concentration for many hours. However, there are numer- ous problems associated with achieving this ideal, and numerous approaches have been taken to solve these problems (13,16,17).

Various investigators have examined the utility of the collagen shield for the delivery of antibiotics to the cornea and aqueous humor. In one of the earliest pharmacokinetic studies, Unterman et al. (18) assessed the phar- macokinetics of tobramycin delivered to rabbit eyes by means of collagen shields hydrated in solutions of either 40 or 200 mg/mL of tobramycin. Tobramycin concentrations in the cornea and aqueous humor were deter- mined at 2, 4, and 8 hours after application. No toxicity was observed with shields hydrated in the 40 mg/mL solution at any time. Eight hours after application, the corneas with shields hydrated in the 200 mg/mL solution of tobramycin had some epithelial defects. At all times and with either hydra- tion solution, the concentration of tobramycin in the cornea and aqueous humor exceeded the MIC for most aminoglycoside-sensitive strains of Pseudomonas .

O’Brien et al. (19) compared collagen shields with soft contact lenses in pharmacokinetic studies of the ocular penetration of tobramycin. Three groups were compared: (a) eyes with collagen shields rehydrated in 3 mg/ mL of tobramycin, (b) eyes with therapeutic soft contact lenses, and (c) eyes with neither lenses nor shields. Topical tobramycin (3 mg/mL) was applied to all eyes every 5 minutes for a total of six doses. Aqueous humor samples

Table 2 Studies of Collagen Shield Drug Delivery

Overall result with collagen Ref.

Compared with Collagen

shield (CS) Phinney et al., 1988 (30)

Drug

shield (CS)

Assay site

Gentamicin

Loading dose + frequent

Tears

CS comparable at all sites

drops

Cornea Aqueous

Phinney et al., 1988 (30)

Vancomycin

Loading dose + frequent

Tears

CS comparable at all sites

drops

Cornea Aqueous

O’Brien et al., 1988 (18)

CS superior Unterman et al., 1988

Tobramycin

Soft contact lens

Aqueous

CS comparable at both sites (17)

Tobramycin

Subconjunctival injection

Cornea

Aqueous

Hwang et al., 1989 (32)

Dexamethasone

Single drop

Cornea

CS superior at all sites

Aqueous Iris Vitreous

Hwang et al., 1989 (32)

Dexamethasone

Frequent drops

Cornea

CS superior at all sites

Aqueous Iris Vitreous

Hwang et al., 1989 (32)

Dexamethasone

CS + frequent drops

Cornea

CS superior at all sites

versus frequent drops

Aqueous

Higaki

Iris Vitreous

et Sawusch et al., 1989 (33)

Prednisolone

Single drop

Cornea

CS superior at both sites al.

Aqueous

Corneal Reidy et al., 1990 (13)

Fluorescein

Frequent drops

Anterior

CS superior to both

Soft contact lens

chamber

Schwartz et al., 1990 (40)

Amphotericin B

Frequent drops

Cornea

CS comparable at both

sites Collagen Reidy et al., 1990 (44)

Aqueous

Cyclosporin A

Frequent drops

Cornea

CS superior

CS comparable Murray et al., 1990 (42)

Aqueous

CS Superior Shields Gussler et al., 1990 (47)

Heparin

Subconjunctival injection

Aqueous

Trifluorothymidine

Drops

Aqueous

Cornea with epithelial

CS + drops

defect: CS + drops superior CS superior from 0–2 h

for Drops superior from 4–8 h

Ocular Chen et al., 1990 (46)

CS superior at all sites

Aqueous Conjunctiva

Drug Taravella et al., 1998 (65)

No difference Kuster et al., 1998 (42)

Tobramycin

Three drops

Aqueous

Trifluorothymidine

Drops

Cornea

CS higher Delivery

Aqueous

Taravella et al., 1999 (64)

Ofloxacin

Three drops

Aqueous

CS superior

316 Table 3 Studies of Collagen Shield Drug Delivery in Rabbit Models of Disease

Compared with collagen

Ref.

Result Sawusch et al., 1988

Experimental model

Drug

shield (CS)

Enhanced antimicrobial (21)

Pseudomonas keratitis

Tobramycin

CS + frequent drops

effect with CS Hobden et al., 1988 (20)

versus frequent drops

Pseudomonas keratitis

Tobramycin

Frequent drops

CS comparable

CS + delayed drops

antimicrobial effect

CS comparable antimicrobial effect Hobden et al., 1990 (29)

versus second CS

Aminoglycoside-

Ciprofloxacin

CS with vehicle

Ciprofloxacin >

resistant Pseudomonas

Norfloxacin

CS with water

norfloxacin for

antimicrobial effect Tobramycin, vehicle, water—no effect Chen et al., 1990 (19)

keratitis

Tobramycin

High-risk keratoplasty

Cyclosporine

Drops

CS superior preventive

A Drops

effect on graft rejection

CS superior therapeutic effect on graft rejection

Hagenah et al., 1990

aFGF and CS alone (52)

Epithelial wound

EGF, aFGF

CS alone

healing

Untreated corneas

superior to untreated EGF and CS

Higaki comparable to

untreated Clinch et al., 1992 (22)

Enhanced antimicrobial et

Pseudomonas keratitis

Tobramycin

CS + frequent drops

effect with CS al.

versus frequent drops

Corneal Silbiger et al., 1992 (24)

Pseudomonas keratitis

Gentamicin

CS + frequent drops

CS with topical drops

versus frequent drops

every 3 hours was effective, but less effective than the

Collagen topical drops every

half hour Baziuk et al., 1992 (28)

Lensectomy and

Gentamicin

Frequent eye drops (every

CS: gentamicin Shields

vitrectomy

30 minutes)

concentrations were lower than frequent eye drops in aqueous humor

for Murray et al., 1992 (43)

Anterior chamber fibrin

Tissue

tPA-hydrated CS versus

CS + tPA significantly Ocular

clot

plasminogen

control CS

shortened the time to

fibrin clot lysis Assil et al., 1992 (23)

activator

No significant difference Drug in efficacy Pleyer et al., 1992 (41)

Pseudomonas keratitis

Tobramycin

Eye drops

Candida albicans

Amphotericin

Eye drops (hourly)

CS group had Delivery

keratitis

B significantly lower fungal counts

Callegan et al., 1994

Enhanced antimicrobial (31)

Staphylococcus aureus

Tobramycin

CS + frequent drops

keratitis

versus frequent drops

effect with CS

318 Higaki et al. were taken 15 and 60 minutes following the last dose. At both times, the eyes

with the collagen shields had a significantly greater concentration of tobra- mycin than the eyes with soft contact lenses or the eyes that received topical drops only.

Chen et al. (20) compared the ocular bioavailability in rabbits of 0.3% tobramycin applied with a collagen shield with topical drop application of tobramycin. Groups of rabbits received either a collagen shield presoaked in tobramycin with a tobramycin drop before and after shield application or three drops of tobramycin. The collagen shield group had higher tobramy- cin levels in the cornea, aqueous humor, and conjunctiva than the second group. They concluded that the use of collagen shields together with stan- dard ophthalmic concentrations of tobramycin is useful in achieving higher concentrations of topically delivered drugs into the anterior segment of the eye.

Hobden et al. (21), Sawusch et al. (22), and Clinch et al. (23) reported the efficacy of collagen shields rehydrated with tobramycin in the therapy of experimental Pseudomonas keratitis in rabbit eyes. Hobden et al. (21) demonstrated that collagen shields hydrated in 4% tobramycin were as efficacious as 4% topical drops given every 30 minutes over a 4-hour period; the number of colony-forming units in both the shield-treated and drop- treated corneas were reduced 4–5 log. Also, eyes with antibiotic-hydrated collagen shields plus one topical application of tobramycin drops over the shield halfway through the 9-hour experimental period were compared to eyes with shields in which the shield was replaced half way through the experimental period. No difference in the number of bacteria was seen. Additionally, these studies showed that the shield alone does not enhance bacterial growth; the number of bacteria was no greater in infected corneas treated with collagen shields hydrated in distilled water (or balanced saline solution) than in untreated control corneas. The overall results provided support for the efficacy and convenience of collagen shields rehydrated in

a water-soluble antibiotic such as tobramycin for the treatment of Pseudomonas keratitis. Assil et al. (24) compared the efficacy of a fortified (14 mg/mL) tobra- mycin-soaked collagen shield to the efficacy of a single loading dose (four 50 m L drops) of fortified tobramycin eyedrops in the treatment of rabbits with Pseudomonas aeruginosa –induced keratitis. Six hours after a single treat- ment, significantly fewer colony-forming units of Pseudomonas were present in the corneas of all three drug-treated groups as compared to the number of colonies in the corneas of balanced salt solution–treated control rabbits. However, no significant difference was found between a collagen shield presoaked in tobramycin and a single loading dose of tobramycin eyedrops in terms of the ability to reduce Pseudomonas.

Corneal Collagen Shields for Ocular Drug Delivery 319 Silbiger and Stern (25) studied the effectiveness of topical gentamicin

treatment, with and without the use of corneal collagen shields, in a rabbit model of Pseudomonas keratitis. A 13.6 mg/mL solution of gentamicin was topically administered, and collagen shields were soaked in 13.6 mg/mL gentamicin for 5 minutes before being placed on the cornea. One hour after the end of the treatment, the corneas were obtained and cultured. The use of an antibiotic-impregnated collagen shield supplemented with topical therapy was more effective than the use of topical treatment alone. However, the collagen shield augmented with topical treatment every 3 hours was significantly less effective than the topical treatment every half hour. They concluded that the use of antibiotic-impregnated collagen shields should not replace the use of topical treatment every half hour with fortified antibiotics as a mainstay of initial drug treatment. However, Liang et al. (26) reported that shield therapy provided significantly higher gentamicin levels in the cornea and aqueous humor than the hourly drop treatment at

0.5 and 2 hours after the end of the treatment in uninfected rabbit eyes.

Callegan et al. (27) compared the efficacy of topical fortified tobramy- cin (1.36%) administered by collagen shields or in topical drop form to rabbit corneas infected with Staphylococcus aureus. Eyes were treated with shields hydrated in and supplemented with fortified tobramycin drops applied every 1, 2, 5 or 10 hours after infection. For topical treatment alone, tobramycin was applied following the identical regimen. Shields sup- plemented with tobramycin drops applied every 1, 2, or 5 hours and topical delivery of tobramycin ever hour sterilized all corneas. Collagen shield deliv- ery of tobramycin with supplemental topical drops can eradicate staphylo- cocci in rabbits with less frequent dosing intervals than required with topical therapy alone.

Dorigo et al. (28) studied collagen shield delivery of netilmicin, an aminoglycoside antibiotic, in rabbits. Collagen shields were immersed for

10 minutes in a commercially available solution of netilmicin, at the stan- dard concentration of 3 mg/mL. The drug levels remained above the MIC for the usual pathogens for 18 hours in the cornea and for 6 hours in the aqueous humor. The study showed that a very concentrated drug solution is not required to obtain high and persistent levels of netilmicin in the cornea.

Baziuk et al. (29) investigated the intraocular drug delivery of collagen shield and fortified eye drops in rabbits that had undergone bilateral len- sectomy and vitrectomy. The left eyes were fitted with collagen shields that had been soaked for 5 minutes in 2.0 mL of gentamicin solution (40 mg/mL) and compared with the right eyes treated with fortified gentamicin drops (13.6 mg/mL) every 30 minutes for 12 hours. The gentamicin concentration was higher in the aqueous humor of all eyes treated with fortified gentamicin drops.

320 Higaki et al. Hobden et al. (30) reported the use of collagen shields hydrated with

various fluoroquinolones for chemotherapy of aminoglycoside-resistant Pseudomonas . The fluoroquinolones used were norfloxacin (40 mg/mL) and ciprofloxacin (25 mg/mL), and the aminoglycoside control was tobra- mycin (40 mg/mL). In these experiments, Pseudomonas was made amino- glycoside-resistant by conjugal transfer of a plasmid. The MICs were 31.25 m g/mL for tobramycin, 0.25 mg/mL for ciprofloxacin, and 0.48 mg/mL for norfloxacin. The colony-forming units from rabbit corneas treated with ciprofloxacin were reduced by 4 log compared to corneas treated with col- lagen shields containing tobramycin or untreated corneas. Norfloxacin, which decreased the colony-forming units approximately 2 log, was not as effective as ciprofloxacin.

Phinney et al. (31) were the first to report the delivery of two anti- biotics in combination (gentamicin and vancomycin) to uninfected rabbit eyes using the collagen shield. Tear, corneal, and aqueous humor concen- trations of each of the two antibiotics were generally higher than, or at least similar to, those achieved by frequent topical application. Combinations of antibiotics have the potential to cover a broad spectrum of infectious agents, but care must be taken to test for pharmacological compatibility to avoid potential therapeutic interference and/or toxicity.

In conclusion, these results suggested that collagen shields containing an antibiotic could serve as a vehicle for drug delivery and could be used for preoperative and postoperative antibiotic prophylaxis and initial treatment of bacterial keratitis (32).

C. Anti-Inflammatory Agents Hwang et al. (33) and Sawusch et al. (34) used collagen shields to enhance

the penetration of anti-inflammatory agents. Hwang et al. (33) compared the deliver of dexamethasone to the cornea and aqueous humor in normal rabbit eyes by four methods: single 0.1% dexamethasone drop, hourly drops, collagen shields hydrated in 0.1% dexamethasone, and collagen shields hydrated in 0.1% dexamethasone followed by hourly topical 0.1% drops. Treatment with the drug-hydrated collagen shields plus hourly drops resulted in both peak and cumulative drug concentrations in the cornea and aqueous humor that were two- to fourfold greater than the concentration achieved by hourly drops alone. Collagen shields without accompanying drops yielded drug concentrations either equal to or greater than the peak and cumulative drug concentrations produced by hourly drops. The authors concluded that collagen shields significantly enhance dexamethasone pene- tration and would be useful for maximizing the delivery of this anti-inflam-

Corneal Collagen Shields for Ocular Drug Delivery 321 matory agent. They also suggested that the use of collagen shields would

decrease the requirement for frequent topical drops. Sawusch et al. (34) compared (a) collagen shields hydrated in 1% prednisolone, (b) collagen shields receiving topical drops in situ, and (c) topical application of 1% prednisolone drops alone. Cornea and aqueous humor were assessed for prednisolone acetate at 30 and 120 minutes after drug application. Both collagen shield delivery systems produced signifi- cantly greater drug levels than topical drops alone at both times. Thus, both these reports support the potential for collagen shield delivery of cor- ticosteroid anti-inflammatory agents (33,34).

D. Combination Therapy In clinical cases, combinations of drugs are often used. Milani et al. (35)

investigated the ability of collagen shields impregnated with gentamicin sulfate and dexamethasone to deliver medication to rabbit eyes. They com- pared collagen shields with subconjunctival injection therapy. The collagen shields produced aqueous humor levels of gentamicin and dexamethasone that were lower than those produced by subconjunctival injection therapy at

30 and 60 minutes, respectively, but that were comparable to subconjuncti- val injection at 3, 6, and 10 hours. They concluded that collagen shield deliver of gentamicin-dexamethasone might be comparable to subconjunc- tival injections and provide an alternative therapy after intraocular surgery. Renard et al. (36) used collagen shields soaked in gentamicin and dexa- methasone for patients after cataract surgery. No adverse effect was reported.

Mahlberg et al. (37) studied the aggregate formation of tobramycin sulfate in combination with methylprednisolone acetate or dexamethasone sodium phosphate on collagen shields. Aggregates were formed on the sur- face of the shields when they were immersed in methylprednisolone acetate and tobramycin. Dexamethasone sodium phosphate and tobramycin resulted in a completely transparent shield. To avoid undesired side effects, such as epithelial sloughing and corneal edema after collagen shield applica- tion, antibiotics and steroids must be carefully selected. Combinations of gentamicin and methylprednisolone sodium succinate or gentamicin and cefazolin on collagen shield also result in precipitates (38). Additionally, care must be taken when mixing drugs to prevent adverse reactions. An aminoglycoside such as gentamicin and a penicillin such as mezlocillin should not be combined because of inactivation of the aminoglycoside by the penicillin (39).

322 Higaki et al.

E. Antifungal Agents Schwartz et al. (40) compared the delivery of amphotericin B in collagen

shields hydrated in a 0.5% drug solution with frequent topical drops (0.15%) in uninfected rabbit eyes. Drops were applied every 5 minutes for the first half hour and at hourly intervals thereafter. The corneas and aqeous humor were assessed at 1, 2, 3, and 6 hours following the initiation of drug delivery. Drug levels in the shield-treated corneas were significantly higher than levels in the drop-treated corneas at 1 and 2 hours after therapy began. At 3 hours, the concentrations of the antifungal drug in the corneal tissues were similar for both delivery methods. At 6 hours, both groups had sig- nificant concentrations of the antifungal agent, but the amount of the drug was greater in the drop-treated corneas than in the shield-treated corneas. Drug levels in the aqueous humor did not differ between the two groups at any time. The results suggest that amphotericin B can be delivered to the cornea via collagen shields at a rate that is comparable with frequent drop deliver.

Pleyer et al. (41) evaluated the effect of collagen shields presoaked with amphotericin B on the treatment of experimental Candida albicans–induced keratitis. Treatment results were compared to those of amphotericin B eye- drops instilled hourly in rabbits. Treatment groups were (a) hourly instilla- tion of 0.15% amphotericin B drops, (b) application of a collagen shield presoaked in 0.5% amphotericin B for one hour, and (c) hourly instillation of saline drops. Rabbit eyes treated with amphotericin B–soaked collagen shields had significantly lower fungal counts compared with eyes receiving hourly amphotericin B drops at days 1 and 3 after the beginning of treat- ment. They concluded that collagen shields soaked in amphotericin B could

be a useful and convenient treatment device in keratomycosis such as that caused by Candida albicans.

F. Anticoagulant Therapy Heparin, a large molecule with a molecular weight between 6000 and 20,000

daltons, has been studied experimentally as a possible agent for the reduc- tion of postoperative fibrin formation after vitrectomy. The intravenous route of administration, however, can be associated with increased post- operative hemorrhage. In an attempt to discover a vehicle that would permit more localized drug delivery, Murray et al. (42) examined the pharmacoki- netics and anticoagulation efficacy of heparin delivered by collagen shields. Collagen shields hydrated with radiolabeled heparin were applied to rabbit eyes, and the amount of heparin in aqueous humor, cornea, and iris was measured at intervals from 15 minutes to 6 hours. The peak of radioactivity

Corneal Collagen Shields for Ocular Drug Delivery 323 was detected in the cornea and aqueous humor 1 hour after application.

Also in this study, 12-hour collagen shields hydrated with heparin were compared to subconjunctival heparin injection. The highest biological activ- ity was seen 30 minutes after application of the collagen shield, and there was a significant amount of anticoagulant activity in the aqueous humor 6 hours after application. At no time was any anticoagulant activity seen in the aqueous humor following subconjunctival injection. The results of this study demonstrate that a high molecular weight compound such as heparin can be delivered by 12- or 24-hour collagen shields, producing significant levels in the aqueous humor. This suggests that collagen shields hydrated with heparin might be effective in the prevention of treatment of fibrin formation in the aqueous humor.

Murray et al. (43) also studied collagen shield delivery of tissue plas- minogen activator (tPA) to the anterior segment and vitreous of rabbit eyes. Time to complete lysis of anterior chamber fibrin clots in the eyes treated

Treatment with tPA-hydrated collagen shields shortened the mean time to Elevated aqueous tPA levels were first measured 18 hours after application

of tPA-hydrated collagen shields. Aqueous tPA levels peaked at 36 hours and remained elevated throughout the 48-hour study period. Vitreous tPA levels were elevated by 2 hours, peaked at 24 hours, and also remained elevated throughout the study period. These results document the efficacy and safety of tPA delivery to the aqueous and vitreous humor via a hydrated collagen shield in the rabbit.

G. Immunosuppressive Agents Cyclosporine has been used successfully to prevent graft rejection in many

types of transplantation. The side effects of systemic administration, how- ever, are considerable, and the systemic dose needed to provide sufficient drug in the cornea makes this route less than useful to prevent rejection of corneal transplants. Also, the drug penetrates the cornea poorly, frustrating the efforts at topical administration. Reidy et al. (44) demonstrated that collagen shields with 4 mg of cyclosporin A incorporated during manufac- ture delivered significantly higher concentrations of drug to the cornea and aqueous humor than an equivalent amount of cyclosporin A prepared in olive oil and given as drops at 15-minute intervals. Four hours after appli- cation of the collagen shield, the corneas contained almost 2500 ng of the drug and the aqueous humor contained about 250 ng. At all times, the concentration of the drug in the aqueous humor was higher in rabbits receiving cyclosporin A via collagen shields compared to drops.

Kanpolat et al. (45) also investigated the penetration of cyclosporin A into the rabbit cornea and aqueous humor after topical drop and collagen shield administration. The rabbits were divided into three groups. The first group received 6 mg of cyclosporin A in castor oil and the second group received 6 mg of cyclosporin A in olive oil applied as topical drops to rabbit eyes within 12 hours. In the third group 12-hour collagen shields soaked in 6 mg of cyclosporin A in olive oil were applied. The cyclosporin A of castor oil drops were higher than those obtained with olive oil drops. In eyes with collagen shields, cyclosporin A levels were higher than olive oil drops but nearly equal to the castor oil drops. In an extension of the studies, Chen et al. (46) showed that cyclosporin A–containing collagen shields suppress corneal allograft rejection. Rabbit eyes with penetrating keratoplasty grafts were placed in vascularized beds to enhance the possibility of graft rejection. The grafts were treated with equivalent amounts of cyclosporin A in the collagen shields or in olive oil drops. The mean survival time of shield- treated grafts was significantly longer than that of drop-treated grafts. Grafts showing early signs of graft reaction treated with cyclosporin–con- taining shields showed reversal of the rejection process. The results of these two studies indicate that the collagen shield is an effective delivery system for cyclosporin A and that the drug delivered in this manner can both suppress the initiation of graft rejection and reverse a graft reaction in progress.

H. Antiviral Agent Gussler et al. (47) investigated the delivery of trifluorothymidine (TFT) in

collagen shields and topical drops in normal rabbit corneas and corneas with experimental epithelial defects. Collagen shields hydrated in 1% TFT and 1% topical drops were used. The eyes were treated with either collagen shields hydrated with TFT, TFT drops, or a combination of collagen shields and drops. Rabbits with normal corneas showed no difference among the treatment groups in terms of TFT levels in the cornea or aqueous humor 30 minutes and 2, 4, and 8 hours after application of the antiviral. Among the rabbits with experimental epithelial defects, the highest drug concentrations were found in the eyes treated with the combination of shields and drops, and the second highest tissue concentrations were seen in the eyes treated with collagen shields hydrated in TFT. Treatment with drops alone pro- duced lower concentrations of TFT than either treatment involving the collagen shields. The authors suggested that this method of drug delivery may be useful to enhance the eradication of herpes simplex virus in eyes with epithelial defects. The authors noted, however, the need for studies of cor-

324 Higaki et al.

Corneal Collagen Shields for Ocular Drug Delivery 325 neal toxicity and efficacy in herpes-infected eyes before a definitive thera-

peutic regimen can be established.

I. Wound Healing After Fyodorov et al. (5) first described the clinical use of collagen shields

for the protection and enhancement of epithelial healing, a number of experimental studies were published confirming their findings. Frantz et al. (48) showed that rabbit eyes with 6 mm superficial keratectomies treated with collagen shields healed significantly faster than untreated eyes. Simsek et al. (49) studied the effects of collagen shields and therapeutic contact lenses on corneal wound healing in rabbits. A corneal wound was created by mechanical removal of the central 6 mm zone of the corneal epithelium

mm 2

2 /hour with the thera-

2 /hour in the control group. Comparison of the study groups revealed no statistically significant difference between the

collagen shield and the therapeutic lens group at any time, whereas a sig- nificantly larger wound size was observed in the control group compared with the treatment groups. In conclusion, these results indicated that both collagen shields and therapeutic lenses enhance wound healing in rabbit eyes.

On the other hand, the results of some studies suggested that more evidence would be needed to establish effectiveness of collagen shields in promoting wound healing (50,51). Shaker et al. (50) reported that cat eyes treated with non–cross-linked porcine collagen shields had a significantly greater healing response than untreated eyes. However, there was no differ- ence in the slope of the healing curve, suggesting that the shield did not increase the speed of epithelial cell migration. Callizo et al. (51) suggested that collagen shields might be ineffective or have adverse effects in deeply injured rabbit corneas. They performed bilateral keratectomies, then treated only the left eye with a collagen shield. No differences were seen in the time course of the healing process between control (untreated) and treated eyes. More polymorphonuclear infiltration, mainly composed of eosinophils, was shown in treated eyes. They concluded that the usefulness of collagen shields should be reappraised, especially in injured corneas.

Additional studies combined the healing properties of collagen shields with delivery of growth factors to influence the rate of reepithelialization. Hagenah et al. (52) used rabbit eyes with superficial keratectomies to exam- ine the effect of collagen shields alone or in combination with epidermal growth factor (EGF) or acidic fibroblastic growth factor on epithelial wound healing. Eyes treated with collagen shields alone or collagen shields

326 Higaki et al. hydrated with a fibroblastic growth factor healed significantly faster than

untreated controls. Shields containing EGF had no enhanced effect. Although it is apparent from this and other studies that the shields alone promote healing, the utility of growth factors in and of themselves, includ- ing optimal dosage, timing, and duration of application, is still uncertain. Therefore, even if the shields enhance the delivery of growth factors, it is not clear at this time how such enhanced delivery can be used to improve epithelial healing.

J. Other Uses Wentworth et al. (53) determined the impact of collagen shields on ulcera-

tion of rabbit corneas after alkali burn. After a 60-second sodium hydroxide burn to rabbit corneas, 24-hour collagen shields were replaced once daily for

21 days. They showed daily use of 24-hour collagen shields after a severe alkali burn to the rabbit cornea exacerbates the progression of corneal ulceration and perforation. They thought that the harmful effects of col- lagen shields might have resulted from the repeated removal and insertion of the shields. The trapping of activated polymorphonuclear leukocytes in the collagen shields that were in contact with the corneal surface caused a locally high concentration of metalloproteinases.

IV. CLINICAL STUDIES The effect of collagen shields on healing and antibacterial or other che-

motherapeutic effects in human eyes has been reported.

A. Therapy to Assist Wound Healing Aquavella et al. (54) reported the use of porcine collagen shields hydrated

with ophthalmic drugs to treat patients following penetrating keratoplasty and cataract extraction. The drugs included tobramycin, gentamicin, pilo- carpine, dexamethasone, and flurbiprofen. No adverse effects were noted. In another study by this group (55), collagen shields used as bandage lenses appeared to accelerate corneal reepithelialization after keratoplasty or other types of anterior segment surgery.

Poland and Kaufman (56) reported the use of collagen shields hydrated with tobramycin in patients who had cataract extraction, penetrat- ing keratoplasty, epikeratophakia, or nonsurgical epithelial healing problems. All surgical patients showed more rapid healing of epithelial defects. Acute nonsurgical epithelial problems with impaired healing also

Corneal Collagen Shields for Ocular Drug Delivery 327 benefited from the use of collagen shields. In contrast, chronic epithelial

defects responded poorly. No infections occurred in any of the patients. Similarly, Groden and White (57) found that 24-hour porcine collagen shields did not contribute to healing of persistent (chronic) epithelial defects following penetrating keratoplasty. They defined a persistent defect as an epithelial erosion (noninfectious) that did not heal in 2 weeks with patching, frequent lubrication, and/or temporary tape tarsorrhaphy. Patients with such defects were assigned to either collagen shield treatment or treatment with a hydrophilic bandage soft contact lens. When none of the collagen shield–treated defects healed, this approach was abandoned and bandage contact lens treatment instituted for all patients. The authors suggested that

a longer-lasting corneal shield (72 hours dissolution time) might be more effective. Marmer (58) described postsurgical healing in human eyes with por- cine collagen shields after radial keratotomy. The patients who were treated with the collagen shields reported less glare and discomfort than patients who did not receive shields. Also, the eyes with shields showed less inflam- matory reaction and edema.

Palmer and McDonald (59) used a disposable soft contact lens piggy- backed onto a medicated, 12-hour corneal collagen shield after corneal surgery in three patients known to have poor corneal epithelial wound healing characteristics. This piggyback lens/shield system delivered sus- tained high levels of medication and promoted epithelial healing in the acute postoperative period without unnecessary postoperative orbital manipulation.

In a different kind of healing study, Fourman and Wiley (60) reported the results of treating a glaucoma filter bleb with a 24-hour collagen shield hydrated in 4 mg/mL of gentamicin. The shield was placed over the leaking site; the bleb leak was sealed within 2 days and remained sealed 2 months later. The collagen shield was helpful in the management of filter bleb leak.

B. Antibacterial or Antiviral Efficacy Lois and Molino (61) treated a case of Mycobacterium chelonae keratitis

with topical fortified amikacin, with no response. They then debrided the epithelial and stromal lesions, and applied an amikacin-soaked corneal col- lagen shield, supplemented every 4 hours with topical fortified amikacin drops. After this treatment, clinical and laboratory examinations showed that no infectious organisms could be detected.

Kuster et al. (62) investigated the ability of collagen shields to deliver TFT to human cornea and aqueous humor. Collagen shields were soaked

328 Higaki et al. in commercially prepared TFT. Patients undergoing penetrating kerato-

plasty wore a presoaked collagen shield for at least 30 minutes preopera- tively. Control patients received drops of TFT only. Cornea and aqueous samples were obtained during surgery. Collagen shields did not enhance delivery of TFT to the cornea with an intact epithelium. In corneas with poor epithelium, drug penetration was higher but variable. They concluded that the role of collagen shields as a drug delivery system for the treatment of herpes simplex keratitis remains to be determined.

C. Postoperative Application Renard et al. (36) compared the effectiveness of subconjunctival injections

and collagen shields in delivering anti-inflammatory agents and antibiotics after cataract surgery. The occurrence of folds in Descemet’s membrane was less frequent and aqueous flare less severe in the collagen shield–treated group than in those treated with subconjunctival injections.

Haaskjold et al. (63) compared the efficacy of collagen shields with that of peribulbar/retrobulbar injection after cataract surgery. Collagen shields were saturated with an antibiotic and a steroid and placed over the cornea postoperatively. The second group received the same drugs through a peribulbar/retrobulbar injection. One day after surgery, the shield group had significantly less corneal edema, conjunctival hemor- rhage, postoperative pain, and fewer corneal opacities. They suggested that using collagen shields for drug delivery after cataract surgery decreases tissue damage and increases patient comfort without adverse side effects.

Taravella et al. (64) investigated whether collagen shields are more effective than topical eye drops for infection prophylaxis after cataract surgery. In their studies, the patients were divided into two groups: the first received three postoperative drops of commercially available topical ofloxacin (0.3%) given 10 minutes apart; the second had a collagen shield soaked in the same medication applied to the eye before surgery. Aqueous humor was extracted immediately before surgery for analysis. Aqueous concentration of ofloxacin in the shield group was significantly higher than that in the drop group. The MICs of ofloxacin for many common ocular pathogens were reached or exceeded in the shield group. However, in a similar study using tobramycin, the ocular penetration of the anti- biotic into the anterior chamber of the human eye in the shield group was not different from that in the drop group (65). The aqueous concentration did not approach the MIC of tobramycin for many common ocular patho- gens.

Corneal Collagen Shields for Ocular Drug Delivery 329

D. Dry Eye Though collagen shields have a lubricating effect, they are not useful as a

treatment for keratoconjunctivitis sicca because they must be applied in the physician’s office and they are not transparent. Kaufman et al. (66) tested a similar system, with the addition of therapeutic agents during manufactur- ing (Collasomes), for drug delivery to the ocular surface. The size of

interference with vision were the advantages of this system.

E. Clinical Uses for Corneal Collagen Shields At the LSU Eye Center, our ocular ophthalmologists performing cataract

surgeries, corneal transplants, and other invasive corneal procedures use the corneal collagen shields as a bandage. Most often they will use antibiotics and anti-inflammatories and hydrate the shields in the solution for 3 min- utes. This is the most common use of the corneal collagen shields following many surgical procedures. Since the eye is bandaged, there is no problem involving visual acuity and often only one shield is used until the bandage is removed (24–48 hours) and the patient’s eye is evaluated.

Another use for the collagen shield is when an ophthalmologist has decided to hospitalize a patient with suspected ocular infection and there is concern about compliance for administering antibiotic drops. In the office, the ophthalmologist can hydrate the shield in a solution of fortified tobra- mycin or commercially available fluoroquinolone and then apply the corneal collagen shield. Patients are instructed to apply drops every 5 or 10 minutes until they are admitted and are being administered antibiotics as a single entity or in combinations in a hospital setting. The placing of the collagen shield hydrated with fortified antibiotics assures that a reliable amount of antibiotic will be in contact with the corneal surface over the next 2–4 hours. The shield should be removed after 3–5 hours if antibiotics are not adminis- tered. It is important not to have any oxygen deprivation of a cornea sus- pected to have an infection.

V. CONCLUSIONS Collagen is a protein that can be safely applied to the body for a variety of

medical and cosmetic purposes. The creation of the corneal collagen shield has provided a means to promote wound healing and, perhaps more impor- tantly, to deliver a variety of medications to the cornea and other ocular tissues. There are many indications that the shields deliver drugs as well as,

330 Higaki et al. if not better than, topical drops. The simplicity of use and convenience

afforded by shields make them an attractive delivery device. Although col- lagen shields produce some discomfort and interfere with vision, corneal collagen shields could become a commonly employed technological improvement in ophthalmic drug delivery.

ACKNOWLEDGMENTS The authors wish to acknowledge the editorial assistance and research col-

laboration of A. K. Mitra, Ph.D. Specific research from the laboratory of Dr. Hill was supported in part by U.S. Public Health Service grant EY06311 and EY08871 (JMH); EY02377 (Eye Center Core Grant); an unrestricted grant from Research to Prevent Blindness (RPB); Dr. Hill is an RPB Scientific Investigator Award recipient. The authors wish to acknowledge the secretarial assistance of Mrs. Carole Hoth. The authors also wish to acknowledge special thanks to Ms. Kristina Braud and Ms. Kathy Vu. None of the authors have any financial or proprietary interest in any agents or devices mentioned in this review.

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