The anterior segment of a human glaucomatous eye after subconjunctival injection of 80 µg of 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester solution displaying a subconjunctival bleb filled with the yellow solution.
Preoperative (n = 10 patients) and postoperative (n = 8 patients) intraocular pressure (IOP) of study eyes after trabeculectomy with photodynamic therapy. There was a statistically significant reduction (P<.001) of mean IOP up to 400 days. One eye with pseudoexfoliation glaucoma had an IOP of 17 mm Hg at the last examination, but a topical antiglaucomatous medication (qualified success) was needed.
The treated eye of patient 5 nineteen days after surgery. The anterior segment shows no inflammation. There is new filtration area and basal iridectomy at the 2-o'clock position. A functional filtration bleb is visible. The intraocular pressure in this eye had decreased to normal levels.
The treated eye of patient 5 (see Figure 3) 4 months after surgery. The anterior segment displays no inflammatory response. The intraocular pressure in this eye was significantly reduced from 26 to 32 mm Hg to less than 20 mm Hg without the patient receiving any antiglaucomatous therapy.
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Diestelhorst M, Grisanti S. Photodynamic Therapy to Control Fibrosis in Human Glaucomatous Eyes After Trabeculectomy: A Clinical Pilot Study. Arch Ophthalmol. 2002;120(2):130–134. doi:10.1001/archopht.120.2.130
To evaluate the safety, tolerability, and clinical effect on intraocular pressure (IOP) of a carboxyfluorescein ester as an adjunctive antifibrotic therapy in human glaucomatous eyes to control postoperative wound healing after trabeculectomy.
In 10 human glaucomatous eyes with high IOPs that underwent 1 to 3 previous surgical procedures for glaucoma, 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester was applied 15 minutes prior to trabeculectomy via a subconjunctival injection followed by intraoperative illumination with diffuse blue light(450-490 nm; approximately 51.9 × 103 candelas/m2)for 8 minutes. Antifibrotic efficacy was established by clinical response, postoperative IOP reduction, slitlamp examination, and filtering bleb site photographs. Success was defined as an IOP less than 21 mm Hg from baseline without the eye receiving any antiglaucomatous medication or an application of antimetabolites.
The mean (SD) IOP on the day before surgery was 37.2 (11.2) mm Hg. The IOP was 16.6 (3.8) mm Hg in 8 successful eyes after a mean (SD) follow-up of 400.1 (38.1) days (P<.001). Two eyes showed scarring at the site of the filtering bleb within 1 month. In 1 eye with pseudoexfoliation glaucoma IOP decreased to 17 mm Hg but needed topical antiglaucomatous medication. Clinical examination revealed none of the following: blebitis, uveitis, endophthalmitis, or toxic damage of the adjacent tissues.
We report our findings about the first 10 consecutive human glaucomatous eyes treated with a single dose of 80 µg of 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester during trabeculectomy. In contrast to chemotherapeutic agents, cellular photoablation acts only on cells having incorporated 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester and having been exposed to light of an appropriate wavelength. Though safety and efficacy demand a controlled randomized study, our method seems to be an effective therapeutic approach to control postoperative fibrosis in human glaucomatous eyes with a poor surgical prognosis. Multiple factors such as dose of light, means of application, wavelength, irradiation area, total dose of the dye, and multiple dosing may be altered in the future to improve the antifibrotic effect of photodynamic therapy during surgery for glaucoma.
TRABECULECTOMY is the most frequently applied surgical method to reduce intraocular pressure (IOP) in patients with glaucoma. The wound healing process at the site of the filtration area, however, causes failure in about 30% of these cases within 6 to 8 weeks after surgery.1 Characteristically fibroblasts from Tenon capsule and episclera lead to a fibroproliferative response that involves and closes the created fistula.2-15 To increase the success rates of filtering surgery, agents such as mitomycin C and fluorouracil4,16-34 have been used perioperatively as an antifibrotic therapy. Despite their positive effect on the success rates of filtering surgery in patients with a poor surgical prognosis, diffusion into adjacent ocular tissues causes toxic effects.35,36 New surgical complications, such as blebitis, endophthalmitis, and hypotonic maculopathy, and an increased incidence of already known postoperative complications have limited its clinical use and stimulated the search for clinically less harmful alternatives.37,38
Photodynamic therapy (PDT) is a selective and localized treatment based on the photosensitized oxidation of biological matter.39-42 A photosensitizer can be used as a mediator of controlled light-induced cell toxicity. The mechanism of action is reported to be due to type I and II photooxidative reactions. Oxygen is transformed into singlet oxygen that oxidizes amino acids, nucleic acids, and unsaturated fatty acids. Within hours, the cytotoxic effect is seen at cell membranes (blebs), plasma membranes, mitochondria, lysosomes, and nuclei. Selective activation of the photosensitizer by the application of light at an appropriate wavelength limits the drug effect to a selected area.40,41
This carboxyfluorescein ester has been shown to inhibit the proliferation of human Tenon fibroblasts in vitro after using the combined treatment of 10 µg of 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester and irradiation for 10 minutes.39 In living eyes of pigmented rabbits 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester was shown to effectively inhibit scarring after filtration surgery and demonstrated a dose-response curve from 40 µg to 100 µg after irradiation for 10 minutes.40 The objective of this open-label phase 2 study was to investigate the effect of cellular photoablation mediated by 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester therapy on the postoperative fibrosis after filtration surgery in human glaucomatous eyes with a poor prognosis.
All patients had uncontrolled glaucoma due to high IOPs despite maximum tolerable medical therapy. They were referred to the Department of Ophthalmology, Center of Ophthalmology, University of Cologne, Cologne, Germany, to undergo glaucoma surgery. We included only eyes with advanced glaucomatous cupping of the optic nerve head (glaucoma fere absolutum and glaucoma absolutum; cup-disc ratio, >0.9).
The study was approved by the ethics review board of the University of Cologne and followed the guidelines of the Declaration of Helsinki as revised in Tokyo Japan, and Venice, Italy. Each patient gave written informed consent after receiving a detailed explanation of the purpose of the study and the method of surgery.
The fluorescent probe, 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester, is a cell membrane permeable compound rendered membrane-impermeable and fluorescent on cleavage by intracellular esterases. The clinical samples of the photosensitizer were provided in tuberculin syringes by the pharmacy of the University of Cologne. A dose of 80 µg was diluted in 300 µL of balanced salt solution and stored at −70°C.
Surgery and clinical examinations were performed in 10 consecutive eyes of 5 men and 5 women with glaucoma. All patients were hospitalized before surgery and remained indoors until the fifth postoperative day. Patients were examined for safety, tolerability, and efficacy of therapy after 14 days, and after 1, 3, 6, 9, 12, and 14 months.
The surgical procedure was performed under topical or general anesthesia by one surgeon (M.D.). Fifteen minutes prior to surgery the eye received a single subconjunctival injection of 80 µg of 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester solution in one superior quadrant where the trabeculectomy was to be performed (Figure 1).
Following a limbus-based conjunctival flap, the episcleral Tenon and the subconjunctival tissue were irradiated. Illumination with diffuse blue light (450-490 nm; approximately 51.9 × 103 candelas/m2) was performed for 8 minutes. Photoactivating light was delivered by a portable lamp (Zeiss GmbH, Oberkochen, Germany) equipped with a blue filter fixed at a distance of about 2 cm from the episclera. Light was focused to an area of about 20 mm in diameter, encompassing the subconjunctival and episcleral tissues and sparing the cornea, which was covered by tape. The illuminated area produced a 3.5 × 3.5-mm scleral flap. Since the illuminated area was well focused, no further mask was needed to minimize light absorption to adjacent tissues.
A standard trabeculectomy and a full-thickness iridectomy followed. The scleral flap was closed with two 10/0 nylon sutures. The Tenon capsule and conjunctiva were closed with a single 8/0 polyglactin (Vicryl) running suture. At the end of surgery all eyes were treated with a subconjunctival injection of a water solution of betamethasone and gentamicin sulfate.
Postoperatively all eyes received prednisolone acetate and tobramycin eye drops for 14 consecutive days. No antiglaucomatous medication or antimetabolites were applied. The postoperative treatment protocol allowed for subconjunctival injection of the betamethasone water solution and laser suture lysis of the 2 scleral nylon sutures. Postoperative success was defined as an IOP of less than 21 mm Hg without the eye receiving any antiglaucomatous medication or an application of antimetabolites.
Clinical examination at the slitlamp was performed on designated days to evaluate both the general appearance, tolerability, and safety of the therapy in the treated eyes and to measure the IOP and the visual acuity. The IOP was measured using Goldmann applanation tonometry before and after surgery. The anterior segment and the site of the filtering bleb was documented by taking photographs with a 35-mm camera (Nikon Corp, Tokyo, Japan).
A 2-tailed t test was performed. Statistical significance was set at P<.05. All values are given as mean (SD).
We included 5 men and 5 women with a mean age of 63.1 years (age range, 18-85 years); with primary open-angle glaucoma (n = 2), pseudoexfoliation glaucoma (n = 3), secondary glaucoma (n = 4), or Axenfeld-Rieger syndrome(n = 1) (Table 1). Most of the eyes had a history of at least 1 glaucoma surgical intervention that had failed. The study eyes were evaluated daily within the first 5 postoperative days and then at designated time points (+14 days, and 1, 3, 6, 9, 12, and 14 months) thereafter. A mean follow-up was 400.1 (38.1) days. The demographic and clinical data of the 5 men and 5 women treated with PDT during trabeculectomy are listed in Table 1.
The preoperative IOP in 10 eyes ranged from 32 to 60 mm Hg with a mean of 37.2 (11.2) mm Hg. On the first postoperative day IOP ranged from 2 to 47 mm Hg with a mean of 9 (16) mm Hg. After surgery (duration, 400.1 [38.1] days), the mean IOP of 8 successful eyes was 16.6 (3.8) mm Hg (P<.001) (Figure 2). One eye with pseudoexfoliation glaucoma had an IOP of 17 mm Hg at the last examination, but a topical antiglaucomatous medication (qualified success) was needed.
Within the first 5 postoperative days, we saw a shallow anterior chamber, hypotony, and choroidal detachment due to an IOP of less than 6 mm Hg in 1 eye that resolved without surgical intervention. None of the eyes demonstrated hypotonic maculopathy. Patients did not report any symptoms of intraocular pain or other unexpected adverse effects.
In some eyes, the postoperative inflammatory reaction observed at the slitlamp examination included conjunctival hyperemia and mild anterior chamber flare (Figure 3). The intraoperative application of 80 µg of 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester did not cause clinically detectable tissue damage at the cornea or conjunctiva (Figure 4). No blebitis or endophthalmitis was observed in any eye. The bleb appearance was variable but within normal ranges. The blebs did not have avascularity as seen with mitomycin C or fluorouracil therapy.
Wound healing was clinically evident as a rise of the IOP and as closure of the filtration area. This process was prominent in 2 young patients (1 eye with secondary glaucoma due to trauma and 1 eye with Axenfeld-Rieger syndrome). Both eyes showed scarring within 1 month after surgery despite the application of PDT.
Photodynamic therapy is an alternative method for the treatment of localized pathologic conditions, such as skin cancer.42 Specific activation of pertinent drugs at the targeted area should avoid adverse effects. Photodynamic therapy has also been evaluated for some distinct ophthalmic diseases, such as ocular tumors, choroidal and corneal neovascularization, proliferative vitreoretinal disorders, and postoperative fibrosis in glaucoma surgery.41,43-47 Hill et al41 investigated the feasibility of PDT in a rabbit model of filtration surgery. Using ethyl etiopurin, a photosensitizer traditionally provided by intravenous injection, they showed that subconjunctival provision could have an effect on filtering bleb survival.41
2′7′-Bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester is a cell membrane–permeable compound rendered membrane impermeable and fluorescent on cleavage by intracellular esterases.48-50 Exposure of cells having incorporated 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester to light at an appropriate wavelength leads to cellular photoablation. The light-induced cytotoxicity of 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester has been demonstrated in in vitro and in vivo studies.39,40
In this study we investigated the antifibrotic effect of a single dose of 80 µg of 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester during glaucoma filtration surgery in human eyes with a poor surgical prognosis. This treatment model may be an alternative approach to control the more aggressive wound healing response of human eyes where filtration surgery had already failed. The use of antimetabolites, such as fluorouracil and mitomycin C, have prolonged bleb survival. The effect of the agents depends on drug concentration, total dose, application time, and method.3,21 Until now no dose-response curve of mitomycin C has been demonstrated to be both clinically safe and effective in human glaucomatous eyes.
Our data reveal that the success of trabeculectomy in human glaucomatous eyes with a poor surgical prognosis can be prolonged when cellular photoablation is applied during filtration surgery (Figure 2). Using a single dose of 80 µg of 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester, the success of filtration surgery was prolonged for a mean follow-up of 400 days. This was expressed both in lower IOP levels and prolonged filtration(Figure 3). The creation of filtration blebs as evidence of successful surgery was associated with a reduced IOP. Slitlamp microscopy revealed no detectable damage of the cornea or conjunctiva(Figure 4).
As 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester is a lipophilic drug, it may diffuse into other tissues adjacent to the area of subconjunctival injection. However, subconjunctival injection of 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester followed by illumination and surgery did not cause a conjunctival defect. Since activation of the dye by illumination is achieved only at the surface of the tissue, it is inconceivable that the photoablative effect affects the ciliary body. In preevaluating in vitro studies, cellular photoablation could only be detected in cells that had incorporated 2′7′-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein, acetoxymethyl ester and were exposed to light at an appropriate wavelength.39 In the rabbit model, histologic and morphologic studies did not reveal any damage to adjacent tissues.40
Cellular photoablation seems to be an effective therapeutic approach to control postoperative fibrosis in human glaucomatous eyes with a poor surgical prognosis. Multiple factors such as light dose, light application, wavelength, irradiation area, total dose of the dye, or multiple dosing may be altered in the future to improve the antifibrotic effect of PDT during glaucoma surgery. The safety and efficacy will be tested in a multicenter, randomized clinical study in 2002.
Accepted for publication October 17, 2001.
This study received a research award from the European Glaucoma Society, London, England, June 8, 2000.
The study was presented in part at the annual meeting of the Association of Research in Vision and Ophthalmology, Fort Lauderdale, Fla, May 4, 2000.
Corresponding author: Salvatore Grisanti, MD, Center of Ophthalmology, Eberhard-Karls-University Tübingen, Schleichstrasse 12-15, 72070 Tübingen, Germany (e-mail: firstname.lastname@example.org).
Reprints: Michael Diestelhorst, MD, Center of Ophthalmology, University of Cologne, Joseph-Stelzmann-Strasse 9, 50931 Cologne, Germany (e-mail: email@example.com).