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Vitreous concentration of topically applied brimonidine tartrate 0.2%
Vitreous Concentration of Topically Applied Brimonidine Tartrate 0.2% Alexander R. Kent, MD,1 Jonathan D. Nussdorf, MD,2 Robert David, MD,3 Farrell Tyson, MD,1 David Small, PhD,3 Dan Fellows, BS4 Objective: To determine the vitreous concentration of brimonidine after topical administration of Alphagan. Design: Prospective observational case series. Participants: Eighteen patients scheduled for elective pars plana vitrectomy. Methods: Brimonidine tartrate, 0.2%, was topically administered twice or three times daily for 4 to 14 days preoperatively in 13 patients. Four patients served as controls, without application of brimonidine. A dry, undiluted vitrectomy specimen obtained intraoperatively was collected, frozen, and sent to an independent bioanalytical facility for quantitative determination of vitreous concentration of brimonidine using gas chromatography/mass spectrometry. Main Outcome Measures: The concentration of brimonidine in human vitreous. Results: All patients treated with brimonidine measured above the lower limit of quantitation with a mean vitreous concentration of 185 ⫾ 500 nM. All patients not treated with brimonidine measured at or below the lower limit of quantitation of 0.05 nM. There was a trend toward higher concentration in patients who were either aphakic or pseudophakic compared with those that were phakic. Conclusions: Topically applied brimonidine results in vitreous levels at or above 2 nM, the concentration shown to activate ␣2-receptors. Ophthalmology 2001;108:784 –787 © 2001 by the American Academy of Ophthalmology. Glaucoma is a progressive optic neuropathy associated with elevated intraocular pressure, as well as characteristic cupping of the optic nerve and visual field defects. Currently, the mainstay of treatment is to lower intraocular pressure by means of topical medication, laser trabeculoplasty, incisional surgery, or a combination of these. Despite optimal pressure control, some patients continue to lose peripheral visual field. The mechanism of this nonpressure-related progression is controversial and may include a deficiency in optic nerve blood flow, an increase in toxic neurotransmitters, or altered collagen in the lamina cribosa, in isolation or in combination.1–3 Research in animal models has identified several pharmacologic agents that modulate neurodegenerative pathways. In a rat optic nerve crush model, intraperitoneal Originally received: May 9, 2000. Accepted: November 22, 2000. Manuscript no. 200281. 1 Storm Eye Institute, Medical University of South Carolina, Charleston, South Carolina. 2 Oschner Clinic, Department of Ophthalmology, New Orleans, Louisiana. 3 Allergan, Inc., Irvine, California. 4 Oneida Research Services, Whitesboro, New York. Presented in part at the American Glaucoma Society, San Antonio, Texas, March 2000. Supported in part by an unrestricted grant from Allergan, Inc., and Research to Prevent Blindness, New York, New York. The authors have no proprietary or commercial interest with the products mentioned. Drs. David and Small are employees of Allergan, Inc. Reprint requests to Alexander R. Kent, MD, Storm Eye Institute, 167 Ashley Avenue, Charleston, SC 29425-2232.
784
© 2001 by the American Academy of Ophthalmology Published by Elsevier Science Inc.
pretreatment with brimonidine tartrate, an ␣2-adrenergic agonist, has been shown to significantly reduce the secondary optic nerve degeneration; a similar effect was seen in an acute ischemia model.4,5 Brimonidine tartrate 0.2% (Alphagan), is currently used clinically as a topical ocular hypotensive agent, and studies are in progress to assess optic nerve or retinal ganglion cell protective activity in humans.6 To have a neuroprotective effect, a drug must penetrate through the ocular barriers of the corneal epithelium, the lens-iris-capsular complex, and the vitreous to reach the target and exert an effect on the retinal ganglion cell layer and optic nerve head. Animal research has found the penetration of brimonidine into the vitreous after topical administration to be significantly less than into the aqueous but still well above the concentration needed to activate ␣2-receptors. Before this study, the concentration of brimonidine in human vitreous after topical administration was unknown.7 We present here quantitative measurement of the vitreous concentration after topically applied brimonidine tartrate 0.2% in humans.
Materials and Methods Patients scheduled for elective pars plana vitrectomy were offered participation in the study. The indications for surgery were macular pucker, macular hole, seton placement in the pars plana, and retinal detachment. These samples were obtained from patients at the Medical University of South Carolina, Charleston, SC (AK) and a private practice location in Louisville, KY (JN) between July 1998 and January 1999. The treatment protocol was approved by the investigational review board at each center, and informed ISSN 0161-6420/00/$–see front matter PII S0161-6420(00)00654-0
Kent et al 䡠 Vitreous Brimonidine Concentration Table 1. Patient Demographics
Age (yrs) Race Caucasian Black Sex Male Female
Table 2. Mean, Median, and Range of Vitreous Concentration of Brimonidine in Aphakic, Pseudophakic, and Aphakic Eyes
Mean
Standard Deviation
67.5
11.5
12 1 4 9
consent was obtained from all patients. Patients were excluded from the study if they had active or advanced diabetic retinopathy, active ocular inflammation, vitreous hemorrhage, prior vitrectomy, or an inability to adhere to the study protocol. Patients were instructed to instill brimonidine tartrate 0.2% in the eye to be operated for 4 to 14 days before surgery (mean 9.5 ⫾ 4.0 days). Eight patients were instructed to instill the medication three times daily with the last drop of brimonidine 2, 4, 6, or 8 hours before surgery. Five patients received brimonidine with twice daily dosing for 14 days, placing the last drop at bedtime the night before surgery, 9 to 12 hours before the collection of the vitreous sample. Four additional patients served as controls, with no topical administration of brimonidine. Vitrectomy by means of the pars plana was performed under retrobulbar anesthesia with a standard three-port technique. The infusion line was closed before insertion to eliminate dilution of the vitreous sample. Approximately 0.5 to 1.0 ml was removed by dry vitrectomy from the center of the vitreous cavity into a syringe. The vitrector was removed from the eye, and the infusion line to the eye was opened. The collected vitreous was immediately coded, frozen, and later forwarded to an independent facility (Oneida Research Services, Whitesboro, NY) for bioanalytical quantitation in a masked fashion.
Method of Quantitation Deuterated brimonidine was added to samples as an internal standard before processing. Brimonidine was extracted from vitreous humor into ethyl acetate under basic conditions, then back-extracted from the ethyl acetate into 0.1 M HCl. The aqueous layer was then made basic by addition of NaOH, and brimonidine was extracted into methylene chloride. The organic layer was removed and evaporated with nitrogen, and the dried residue was derivatized with 3,5 bis (tirfluoromethyl)benzoyl chloride and again evaporated with nitrogen. Samples were reconstituted in 25 l of ethyl acetate, and 1 to 2 l was injected into a Hewlett-Packard 5890 gas chromatograph (Hewlett Packard, Avondale, PA) interfaced with a Finnigan 4600 mass spectrometer (Finnigan Thermo Quest, San Jose, CA). The quantitation range was 50 to 10,000 pg/ml (0.17– 34.4 M). The molecular weight of brimonidine free base is 291.1.
Vitreous Concentration of Brimonidine (nM)
Aphakic Pseudophakic Phakic Overall
Mean
Median
Range
164 256 9.3 185
164 14.7 9.0 15.1
115–213 4.4–1836 1.4–17.4 1.4–1836
trend toward higher vitreous brimonidine concentration in patients after cataract surgery: 256 ⫾ 639 nM (median, 14.7 nM), 164 ⫾ 69 nM (median, 164 nM), and 9.3 ⫾ 8.0 nM (median, 9.0 nM) in pseudophakic, aphakic/anterior chamber lens, and phakic patients, respectively. The sample size in each category is too small for meaningful statistical analysis. Nevertheless, it was noted that the brimonidine concentrations in the vitreous depended more on the status of the lens than on the time since the last instillation or the duration of treatment: the mean and median vitreous concentration of brimonidine in the five pseudophakic eyes, 12 hours after the last dose of brimonidine was 14.9 ⫾ 8.1 nM and 14.3 nM (range, 4.4 –27.2 nM), respectively (Fig 1).
Discussion Brimonidine has been used as an effective ocular hypotensive agent in patients with ocular hypertension and glaucoma.6,8 Recent experimental studies have suggested that brimonidine may have benefits extending beyond the traditional therapy of lowering intraocular pressure. Brimonidine has been shown to exert a neuroprotective activity by means of the ␣2-receptor in a calibrated rat optic nerve crush model, as well as in studies involving ischemia/ reperfusion.4,5 To date, there have been no long-term studies confirming this neuroprotective activity specifically in humans or animal models of glaucoma. Mastropasqua et al9 observed a slight improvement over baseline in one visual field parameter (corrected pattern standard deviation) in patients 2 hours after topical administration of brimonidine. For a drug to have neuroprotective capabilities from a
Results The demographics can be found in Table 1. Nearly all patients were white. Samples from patients who did not receive brimonidine were at or below the lower limits of quantitation, and all samples from patients who received brimonidine were above the lower limit of quantitation. Four brimonidine-treated patients had levels above the upper limit of quantitation, but still within what was considered to be the linear portion of the standard curve. Mean and median brimonidine concentrations in all treated patients were 185 ⫾ 500 nM and 15.1 nM, respectively (Table 2). There was a
Figure 1. Vitreous concentration of brimonidine in phakic (E), pseudophakic (■), and aphakic (Œ) subject as a function of time between last dose and vitrectomy procedure. All patients at 12 hours were dosed twice daily.
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Ophthalmology Volume 108, Number 4, April 2001 topically applied ocular preparation, the drug must overcome several hurdles to stimulate its target. Initially, the drug must penetrate the corneal epithelial and lens–iris barriers into the vitreous, where it is diluted, and must then bind to and activate receptors in the retina (nerve fiber layer) and optic nerve head. Only then can subsequent intracellular changes interrupt the programmed cell death (apoptosis) mechanisms, slowing the neurodegenerative glaucomatous process. This study attempts to determine the amount of brimonidine present in the human vitreous during typical brimonidine treatment. Previous animal studies have shown that the aqueous concentrations of brimonidine in albino and pigmented rabbits range between 0.03 and 2.2 g/ml over a 6-hour period after instillation.10 Brimonidine binds to melanin and maintains its peak concentration in the ciliary body over 6 hours after a single dose.10 This may help explain the significant vitreous concentration detected 10 to 14 hours after the last brimonidine dose in several of our patients. Similarly, monkey studies have demonstrated that a concentration of 100 to 170 nM is attained in the vitreous after 14 days of treatment with brimonidine twice daily (Chien DS, et al. Pharm Res 1992;9:S336).7 Activation of the ␣2-receptor by brimonidine has been shown to up-regulate an intrinsic signaling cascade and antiapoptotic genes such as bcl-2 and bcl-xl.5 Wen et al11 have shown clonidine to induce a threefold increase in bFGF mRNA expression in photoreceptors in vivo within 12 hours after intraperitoneal injection in Sprague-Dawley rats; this response was completely inhibited by yohimbine, an ␣2-receptor antagonist. Alpha2-adrenoreceptors may mediate in neuroprotection through other mechanisms: phosphorylation of mitogen-activated retinal protein kinase or a decrease in glutamate release from neurons (Peng, Lai, et al. Activation of cell survival signaling pathway in the retina by selective alpha-2-adrenoceptor agonist— brimonidine. IOVS 40,4(S);S763, ARVO, 1999). Preclinical studies have shown that brimonidine activates the ␣2-receptor at 2 nM while requiring at least 2000 nM for the ␣2-receptor.7 In this study, we demonstrate that brimonidine concentrations obtained after topical administration exceed the concentration required for ␣2-receptor activation. Although few of these patients had glaucoma, their age distribution was similar to that of patients with ocular hypertension and glaucoma. Most patients had mild-to-moderate retinal pathology such as macular holes and epiretinal membranes requiring pars plana vitrectomies; it is unlikely that the retinal pathology present would have altered the ocular absorption, distribution, or metabolism of brimonidine. Topical brimonidine was placed preoperatively for at least 4 days (mean, 9.5 ⫾ 4.0 days) in an attempt to reach steadystate concentrations. The affinity of brimonidine for melanin may have attenuated vitreous brimonidine concentrations in patients administering the medication for a shorter period of time.10 However, in our study, there was no significant effect of longer treatment with higher concentrations of detected brimonidine (logistic regression). The absence of a crystalline lens appears to increase the vitreal concentration (Table 2), although this was not statistically significant because of the few numbers in this study and
786
wide variation in concentration. Twelve hours after last dosing, the brimonidine concentration was a mean of 14.9 ⫾ 8.1 nM (median, 14.3 nM) in a group of five pseudophakic patients that had instilled brimonidine on a twice daily basis for 2 weeks before surgery. This would suggest trough effect concentration in pseudophakic patients using brimonidine on a three times daily basis to be well above receptor activation levels. All but one subject succeeded in having a vitreous concentration of brimonidine higher than that previously known needed to activate ␣2-receptors. To activate such receptors, the drug present in the vitreous must not be bound to protein and must be in equilibrium with the retinal concentration that may or may not be at the same level. Unfortunately, in vivo determination of the retinal concentration is very difficult because of the absence of retinal tissue obtained during such operations. The retinal concentration needs further study for precise quantitation. We were unable to detect a correlation with dosing at various times less than 8 hours before vitreous sampling. Although there seemed to be a decay curve of brimonidine concentration over the 12 hours after last dose, it was not statistically significant given the small data set with high variability. Our study demonstrates a significant presence of brimonidine in the vitreous at concentrations that are well above the level to activate ␣2-receptors. Whether this is sufficient to impart an effective, detectable neuroprotective effect has yet to be determined. We look forward to the results of other studies currently underway that are addressing the potential long-term neuroprotective effect of brimonidine and other medications in patients with open-angle and low-tension glaucoma.
References 1. Flammer J, Haefliger IO, Orgul S, Resink T. Vascular dysregulation: a principal risk factor for glaucomatous damage? J Glaucoma 1999;8:212–9. 2. Dreyer EB, Zurakowski D, Schumer RA, et al. Elevated glutamate levels in the vitreous body of humans and monkeys with glaucoma. Arch Ophthalmol 1996;114:299 –305. 3. Quigley HA, Dorman-Pease ME, Brown AE. Quantitative study of collagen and elastin of the optic nerve head and sclera in human and experimental monkey glaucoma. Curr Eye Res 1991;10:877– 88. 4. Yoles E, Wheeler LA, Schwartz M. Alpha2-adrenoreceptor agonists are neuroprotective in a rat model of optic nerve degeneration [published erratum appears in Invest Ophthalmol Vis Sci 1999;40:2470]. Invest Ophthalmol Vis Sci 1999;40: 65–73. 5. Wheeler LA, Lai R, Woldemussie E. From the lab to the clinic: activation of an ␣2 agonist pathway is neuroprotective in models of retinal and optic nerve injury. Eur J Ophthalmol 1999;9(Suppl 1):S17–21. 6. Schuman JS. Clinical experience with brimonidine 0.2% and timolol 0.5% in glaucoma and ocular hypertension. Surv Ophthalmol 1996;41(Suppl 1):S27–37. 7. Burke J, Schwartz M. Preclinical evaluation of brimonidine. Surv Ophthalmol 1996;41(Suppl 1):S9 –18. 8. Greenfield DS, Liebmann JM, Ritch R. Brimonidine: a new
Kent et al 䡠 Vitreous Brimonidine Concentration alpha2-adrenoreceptor agonist for glaucoma treatment. J Glaucoma 1997;6:250 – 8. 9. Mastropasqua L, Ciancaglini M, Carpineto P, et al. Effects of brimonidine 0.2% on blue-yellow perimetry of glaucomatous patients. Acta Ophthalmol Scand Suppl 1998;76 Suppl 227: 36. 10. Acheampong AA, Shackleton M, Tang-Liu DD. Comparative
ocular pharmacokinetics of brimonidine after a single dose application to the eyes of albino and pigmented rabbits. Drug Metab Dispos 1995;23:708 –12. 11. Wen R, Cheng T, Li Y, et al. Alpha 2-adrenergic agonists induce basic fibroblast growth factor expression in photoreceptors in vivo and ameliorate light damage. J Neurosci 1996; 16:5986 –92.
787
784
© 2001 by the American Academy of Ophthalmology Published by Elsevier Science Inc.
pretreatment with brimonidine tartrate, an ␣2-adrenergic agonist, has been shown to significantly reduce the secondary optic nerve degeneration; a similar effect was seen in an acute ischemia model.4,5 Brimonidine tartrate 0.2% (Alphagan), is currently used clinically as a topical ocular hypotensive agent, and studies are in progress to assess optic nerve or retinal ganglion cell protective activity in humans.6 To have a neuroprotective effect, a drug must penetrate through the ocular barriers of the corneal epithelium, the lens-iris-capsular complex, and the vitreous to reach the target and exert an effect on the retinal ganglion cell layer and optic nerve head. Animal research has found the penetration of brimonidine into the vitreous after topical administration to be significantly less than into the aqueous but still well above the concentration needed to activate ␣2-receptors. Before this study, the concentration of brimonidine in human vitreous after topical administration was unknown.7 We present here quantitative measurement of the vitreous concentration after topically applied brimonidine tartrate 0.2% in humans.
Materials and Methods Patients scheduled for elective pars plana vitrectomy were offered participation in the study. The indications for surgery were macular pucker, macular hole, seton placement in the pars plana, and retinal detachment. These samples were obtained from patients at the Medical University of South Carolina, Charleston, SC (AK) and a private practice location in Louisville, KY (JN) between July 1998 and January 1999. The treatment protocol was approved by the investigational review board at each center, and informed ISSN 0161-6420/00/$–see front matter PII S0161-6420(00)00654-0
Kent et al 䡠 Vitreous Brimonidine Concentration Table 1. Patient Demographics
Age (yrs) Race Caucasian Black Sex Male Female
Table 2. Mean, Median, and Range of Vitreous Concentration of Brimonidine in Aphakic, Pseudophakic, and Aphakic Eyes
Mean
Standard Deviation
67.5
11.5
12 1 4 9
consent was obtained from all patients. Patients were excluded from the study if they had active or advanced diabetic retinopathy, active ocular inflammation, vitreous hemorrhage, prior vitrectomy, or an inability to adhere to the study protocol. Patients were instructed to instill brimonidine tartrate 0.2% in the eye to be operated for 4 to 14 days before surgery (mean 9.5 ⫾ 4.0 days). Eight patients were instructed to instill the medication three times daily with the last drop of brimonidine 2, 4, 6, or 8 hours before surgery. Five patients received brimonidine with twice daily dosing for 14 days, placing the last drop at bedtime the night before surgery, 9 to 12 hours before the collection of the vitreous sample. Four additional patients served as controls, with no topical administration of brimonidine. Vitrectomy by means of the pars plana was performed under retrobulbar anesthesia with a standard three-port technique. The infusion line was closed before insertion to eliminate dilution of the vitreous sample. Approximately 0.5 to 1.0 ml was removed by dry vitrectomy from the center of the vitreous cavity into a syringe. The vitrector was removed from the eye, and the infusion line to the eye was opened. The collected vitreous was immediately coded, frozen, and later forwarded to an independent facility (Oneida Research Services, Whitesboro, NY) for bioanalytical quantitation in a masked fashion.
Method of Quantitation Deuterated brimonidine was added to samples as an internal standard before processing. Brimonidine was extracted from vitreous humor into ethyl acetate under basic conditions, then back-extracted from the ethyl acetate into 0.1 M HCl. The aqueous layer was then made basic by addition of NaOH, and brimonidine was extracted into methylene chloride. The organic layer was removed and evaporated with nitrogen, and the dried residue was derivatized with 3,5 bis (tirfluoromethyl)benzoyl chloride and again evaporated with nitrogen. Samples were reconstituted in 25 l of ethyl acetate, and 1 to 2 l was injected into a Hewlett-Packard 5890 gas chromatograph (Hewlett Packard, Avondale, PA) interfaced with a Finnigan 4600 mass spectrometer (Finnigan Thermo Quest, San Jose, CA). The quantitation range was 50 to 10,000 pg/ml (0.17– 34.4 M). The molecular weight of brimonidine free base is 291.1.
Vitreous Concentration of Brimonidine (nM)
Aphakic Pseudophakic Phakic Overall
Mean
Median
Range
164 256 9.3 185
164 14.7 9.0 15.1
115–213 4.4–1836 1.4–17.4 1.4–1836
trend toward higher vitreous brimonidine concentration in patients after cataract surgery: 256 ⫾ 639 nM (median, 14.7 nM), 164 ⫾ 69 nM (median, 164 nM), and 9.3 ⫾ 8.0 nM (median, 9.0 nM) in pseudophakic, aphakic/anterior chamber lens, and phakic patients, respectively. The sample size in each category is too small for meaningful statistical analysis. Nevertheless, it was noted that the brimonidine concentrations in the vitreous depended more on the status of the lens than on the time since the last instillation or the duration of treatment: the mean and median vitreous concentration of brimonidine in the five pseudophakic eyes, 12 hours after the last dose of brimonidine was 14.9 ⫾ 8.1 nM and 14.3 nM (range, 4.4 –27.2 nM), respectively (Fig 1).
Discussion Brimonidine has been used as an effective ocular hypotensive agent in patients with ocular hypertension and glaucoma.6,8 Recent experimental studies have suggested that brimonidine may have benefits extending beyond the traditional therapy of lowering intraocular pressure. Brimonidine has been shown to exert a neuroprotective activity by means of the ␣2-receptor in a calibrated rat optic nerve crush model, as well as in studies involving ischemia/ reperfusion.4,5 To date, there have been no long-term studies confirming this neuroprotective activity specifically in humans or animal models of glaucoma. Mastropasqua et al9 observed a slight improvement over baseline in one visual field parameter (corrected pattern standard deviation) in patients 2 hours after topical administration of brimonidine. For a drug to have neuroprotective capabilities from a
Results The demographics can be found in Table 1. Nearly all patients were white. Samples from patients who did not receive brimonidine were at or below the lower limits of quantitation, and all samples from patients who received brimonidine were above the lower limit of quantitation. Four brimonidine-treated patients had levels above the upper limit of quantitation, but still within what was considered to be the linear portion of the standard curve. Mean and median brimonidine concentrations in all treated patients were 185 ⫾ 500 nM and 15.1 nM, respectively (Table 2). There was a
Figure 1. Vitreous concentration of brimonidine in phakic (E), pseudophakic (■), and aphakic (Œ) subject as a function of time between last dose and vitrectomy procedure. All patients at 12 hours were dosed twice daily.
785
Ophthalmology Volume 108, Number 4, April 2001 topically applied ocular preparation, the drug must overcome several hurdles to stimulate its target. Initially, the drug must penetrate the corneal epithelial and lens–iris barriers into the vitreous, where it is diluted, and must then bind to and activate receptors in the retina (nerve fiber layer) and optic nerve head. Only then can subsequent intracellular changes interrupt the programmed cell death (apoptosis) mechanisms, slowing the neurodegenerative glaucomatous process. This study attempts to determine the amount of brimonidine present in the human vitreous during typical brimonidine treatment. Previous animal studies have shown that the aqueous concentrations of brimonidine in albino and pigmented rabbits range between 0.03 and 2.2 g/ml over a 6-hour period after instillation.10 Brimonidine binds to melanin and maintains its peak concentration in the ciliary body over 6 hours after a single dose.10 This may help explain the significant vitreous concentration detected 10 to 14 hours after the last brimonidine dose in several of our patients. Similarly, monkey studies have demonstrated that a concentration of 100 to 170 nM is attained in the vitreous after 14 days of treatment with brimonidine twice daily (Chien DS, et al. Pharm Res 1992;9:S336).7 Activation of the ␣2-receptor by brimonidine has been shown to up-regulate an intrinsic signaling cascade and antiapoptotic genes such as bcl-2 and bcl-xl.5 Wen et al11 have shown clonidine to induce a threefold increase in bFGF mRNA expression in photoreceptors in vivo within 12 hours after intraperitoneal injection in Sprague-Dawley rats; this response was completely inhibited by yohimbine, an ␣2-receptor antagonist. Alpha2-adrenoreceptors may mediate in neuroprotection through other mechanisms: phosphorylation of mitogen-activated retinal protein kinase or a decrease in glutamate release from neurons (Peng, Lai, et al. Activation of cell survival signaling pathway in the retina by selective alpha-2-adrenoceptor agonist— brimonidine. IOVS 40,4(S);S763, ARVO, 1999). Preclinical studies have shown that brimonidine activates the ␣2-receptor at 2 nM while requiring at least 2000 nM for the ␣2-receptor.7 In this study, we demonstrate that brimonidine concentrations obtained after topical administration exceed the concentration required for ␣2-receptor activation. Although few of these patients had glaucoma, their age distribution was similar to that of patients with ocular hypertension and glaucoma. Most patients had mild-to-moderate retinal pathology such as macular holes and epiretinal membranes requiring pars plana vitrectomies; it is unlikely that the retinal pathology present would have altered the ocular absorption, distribution, or metabolism of brimonidine. Topical brimonidine was placed preoperatively for at least 4 days (mean, 9.5 ⫾ 4.0 days) in an attempt to reach steadystate concentrations. The affinity of brimonidine for melanin may have attenuated vitreous brimonidine concentrations in patients administering the medication for a shorter period of time.10 However, in our study, there was no significant effect of longer treatment with higher concentrations of detected brimonidine (logistic regression). The absence of a crystalline lens appears to increase the vitreal concentration (Table 2), although this was not statistically significant because of the few numbers in this study and
786
wide variation in concentration. Twelve hours after last dosing, the brimonidine concentration was a mean of 14.9 ⫾ 8.1 nM (median, 14.3 nM) in a group of five pseudophakic patients that had instilled brimonidine on a twice daily basis for 2 weeks before surgery. This would suggest trough effect concentration in pseudophakic patients using brimonidine on a three times daily basis to be well above receptor activation levels. All but one subject succeeded in having a vitreous concentration of brimonidine higher than that previously known needed to activate ␣2-receptors. To activate such receptors, the drug present in the vitreous must not be bound to protein and must be in equilibrium with the retinal concentration that may or may not be at the same level. Unfortunately, in vivo determination of the retinal concentration is very difficult because of the absence of retinal tissue obtained during such operations. The retinal concentration needs further study for precise quantitation. We were unable to detect a correlation with dosing at various times less than 8 hours before vitreous sampling. Although there seemed to be a decay curve of brimonidine concentration over the 12 hours after last dose, it was not statistically significant given the small data set with high variability. Our study demonstrates a significant presence of brimonidine in the vitreous at concentrations that are well above the level to activate ␣2-receptors. Whether this is sufficient to impart an effective, detectable neuroprotective effect has yet to be determined. We look forward to the results of other studies currently underway that are addressing the potential long-term neuroprotective effect of brimonidine and other medications in patients with open-angle and low-tension glaucoma.
References 1. Flammer J, Haefliger IO, Orgul S, Resink T. Vascular dysregulation: a principal risk factor for glaucomatous damage? J Glaucoma 1999;8:212–9. 2. Dreyer EB, Zurakowski D, Schumer RA, et al. Elevated glutamate levels in the vitreous body of humans and monkeys with glaucoma. Arch Ophthalmol 1996;114:299 –305. 3. Quigley HA, Dorman-Pease ME, Brown AE. Quantitative study of collagen and elastin of the optic nerve head and sclera in human and experimental monkey glaucoma. Curr Eye Res 1991;10:877– 88. 4. Yoles E, Wheeler LA, Schwartz M. Alpha2-adrenoreceptor agonists are neuroprotective in a rat model of optic nerve degeneration [published erratum appears in Invest Ophthalmol Vis Sci 1999;40:2470]. Invest Ophthalmol Vis Sci 1999;40: 65–73. 5. Wheeler LA, Lai R, Woldemussie E. From the lab to the clinic: activation of an ␣2 agonist pathway is neuroprotective in models of retinal and optic nerve injury. Eur J Ophthalmol 1999;9(Suppl 1):S17–21. 6. Schuman JS. Clinical experience with brimonidine 0.2% and timolol 0.5% in glaucoma and ocular hypertension. Surv Ophthalmol 1996;41(Suppl 1):S27–37. 7. Burke J, Schwartz M. Preclinical evaluation of brimonidine. Surv Ophthalmol 1996;41(Suppl 1):S9 –18. 8. Greenfield DS, Liebmann JM, Ritch R. Brimonidine: a new
Kent et al 䡠 Vitreous Brimonidine Concentration alpha2-adrenoreceptor agonist for glaucoma treatment. J Glaucoma 1997;6:250 – 8. 9. Mastropasqua L, Ciancaglini M, Carpineto P, et al. Effects of brimonidine 0.2% on blue-yellow perimetry of glaucomatous patients. Acta Ophthalmol Scand Suppl 1998;76 Suppl 227: 36. 10. Acheampong AA, Shackleton M, Tang-Liu DD. Comparative
ocular pharmacokinetics of brimonidine after a single dose application to the eyes of albino and pigmented rabbits. Drug Metab Dispos 1995;23:708 –12. 11. Wen R, Cheng T, Li Y, et al. Alpha 2-adrenergic agonists induce basic fibroblast growth factor expression in photoreceptors in vivo and ameliorate light damage. J Neurosci 1996; 16:5986 –92.
787