Clinical Ophthalmology
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`Open access Full Text article
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`O r i g i n a l re s e a rCh
`
`The ocular distribution of 14C-labeled bromfenac
`ophthalmic solution 0.07% in a rabbit model
`
`Journal name: Clinical Ophthalmology
`Journal Designation: Original Research
`Year: 2014
`Volume: 8
`Running head verso: Baklayan and Muñoz
`Running head recto: Ocular distribution of 14C-bromfenac solution
`DOI: http://dx.doi.org/10.2147/OPTH.S66638
`
`george a Baklayan
`Mauricio Muñoz
`Bausch + lomb, irvine, Ca, Usa
`
`Correspondence: george a Baklayan
`Bausch + lomb, 50 Technology Drive,
`irvine, Ca, Usa 92618
`Tel +1 949 788 5314
`Fax +1 949 788 6020
`email george.baklayan@bausch.com
`
`Purpose: To evaluate the ocular distribution of an advanced formulation of bromfenac
`ophthalmic solution. Two studies were conducted in rabbits: 1) a 12-hour parallel-group study
`comparing the ocular distribution of 14C-bromfenac ophthalmic solution 0.07%, pH 7.8 with
`that of 14C-bromfenac ophthalmic solution 0.09%, pH 8.3, and 2) a 24-hour study evaluating
`the ocular distribution of 14C-bromfenac ophthalmic solution 0.07%, pH 7.8.
`Methods: In the 12-hour study, rabbits were randomized to receive 50 µL of 14C-bromfenac
`0.07%, pH 7.8 or 50 µL 14C-bromfenac 0.09%, pH 8.3 in one eye, whereas, in the 24-hour,
`study both eyes received 50 µL of 14C-bromfenac 0.07%, pH 7.8. Ocular tissues were collected
`at 1, 2, 4, 8, 12 (both studies) and 24 hours (second study only) following drug instillation, and
`tissue radioactivity was determined using liquid scintillation chromatography.
`Results: Measureable levels of bromfenac were observed in all ocular tissues, with the excep-
`tion of vitreous humor, regardless of formulation. In the 12-hour study, high concentrations of
`14C-bromfenac were found in the sclera, followed by the iris/ciliary body, aqueous humor, choroid,
`retina, and lens. There was no significant difference between the bromfenac 0.07%, pH 7.8 and
`bromfenac 0.09%, pH 8.3 formulations in any 14C-bromfenac tissue levels at any time point,
`with the exception of in sclera at 2 hours post-instillation (0.451 µg eq/g versus 0.302 µg eq/g,
`respectively, P0.001). There was also no significant difference in the total amount of
`14C- bromfenac in the tissues evaluated following instillation of the two formulations. In the
`24-hour study evaluating bromfenac 0.07%, pH 7.8 only, high concentrations of 14C-bromfenac
`were found 1 hour post-instillation in the cornea (2.402 µg eq/g) and conjunctiva (1.049 µg eq/g),
`two tissues not evaluated in the 12-hour study. The rank order of 14C-bromfenac levels in the other
`ocular tissues was the same as that observed in the 12-hour study, with measureable amounts of
`14C-bromfenac detected through 24 hours in all tissues with the exception of vitreous humor.
`Conclusion: Bromfenac ophthalmic solution 0.07%, pH 7.8 readily penetrated ocular tissues
`with levels similar to those of bromfenac ophthalmic solution 0.09%, pH 8.3.
`Keywords: penetration, topical NSAIDs, pharmacokinetics, eye, pH
`
`Introduction
`Topical ophthalmic drugs are often reformulated for improved bioavailability and/
`or reduced dosing frequency. Most often this is accomplished by increasing the con-
`centration of the active ingredient in the formulation. Another approach is to modify
`the vehicle without increasing the concentration.1 Recently, bromfenac ophthalmic
`solution was reformulated with a lower pH to allow for a decreased concentration of
`the active ingredient while maintaining high intraocular tissue exposure. In order to
`provide clinicians insight into the applicability of the modified bromfenac formulation
`in the management of various ocular disorders, studies were conducted to evaluate
`the ocular penetration of bromfenac in the modified formulation and compare it to a
`previously marketed reference formulation.
`
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`1717
`Clinical Ophthalmology 2014:8 1717–1724
`© 2014 Baklayan and Muñoz. This work is published by Dove Medical Press Limited, and licensed under Creative Commons Attribution – Non Commercial (unported, v3.0)
`License. The full terms of the License are available at http://creativecommons.org/licenses/by-nc/3.0/. Non-commercial uses of the work are permitted without any further
`permission from Dove Medical Press Limited, provided the work is properly attributed. Permissions beyond the scope of the License are administered by Dove Medical Press Limited. Information on
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`Topical nonsteroidal anti-inflammatory drugs (NSAIDs)
`are routinely used for treatment of inflammation and pain sec-
`ondary to cataract surgery, and are sometimes preferred over
`topical corticosteroids to avoid steroid-associated adverse
`events such as elevated intraocular pressure and cataract
`formation.2–4 In the United States, bromfenac ophthalmic
`0.09% solution is a topical NSAID indicated for the treat-
`ment of postoperative inflammation and reduction of ocular
`pain in patients who have undergone cataract extraction. The
`0.09% formulation (or 0.1% when labeled as the salt form)
`has been shown to be as effective as betamethasone in mini-
`mizing inflammation following cataract surgery.5 In April
`2013, Bausch + Lomb gained US Food and Drug Admin-
`istration (FDA) approval for bromfenac 0.07% (Prolensa®;
`Bausch + Lomb, Irvine, CA, USA), a once-daily, advanced
`formulation of bromfenac ophthalmic solution.6 Prolensa®
`was reformulated from bromfenac 0.09% (Bromday®;
`Bausch + Lomb) to achieve similar ocular bioavailability
`with a lower concentration of active drug, thereby ensuring
`similar clinical efficacy to Bromday® but with reduced expo-
`sure of the surgically compromised ocular surface to the drug.
`In order to lower the concentration yet maintain the same
`degree of ocular penetration, the pH of the formulation was
`reduced from 8.3 (Bromday®) to 7.8 (Prolensa®). Bromfenac,
`like most NSAIDs, is a weakly acidic drug.7 Decreasing the
`pH of the formulation increases the unionized fraction of the
`drug, which, in turn, enhances ocular penetration.7
`NSAIDs have anti-inflammatory properties mediated by
`the inhibition of the cyclooxygenase (COX) pathway and
`prostaglandin (PG) synthesis. PGs, which increase after cata-
`ract surgery, disrupt the blood–aqueous humor barrier and
`increase vascular permeability, vasodilation, and intraocular
`pressure, leading to ocular inflammation.8,9 Studies have
`shown that the COX-2 enzyme, considered the primary
`mediator of ocular inflammation,10 is upregulated in animal
`models of ocular inflammation and ocular injury.11,12
`In order to reduce inflammation effectively, ophthalmic
`NSAIDs need to both penetrate the affected tissues and
`bind to the COX-2 enzyme. A study of the physiochemi-
`cal properties of amfenac and its derivatives found that
`the addition of a substituent group to the aromatic ring of
`the molecule decreased, while the addition of a halogen
`group (Br~IClF) to the 4′-position of the aromatic ring
`increased anti-inflammatory potency.13 The presence of a
`bromine in the 4′-position, as in bromfenac, was found to
`improve in vitro and in vivo potency, absorption across the
`cornea, and penetration into ocular tissues.13–15 Bromfenac
`is a selective COX-2 inhibitor with an IC50 approximately
`
`32 times more active against COX-2 than COX-1 (IC50
`of 0.0066 µM and 0.210 µM, respectively),16 and it has
`been shown in an animal model to significantly prevent
`PGE2 production compared with nepafenac (P0.05).17
`Bromfenac is also highly lipophilic, with a ClogP (octanol/
`water partition coefficient) of 2.23, which is higher than
`that of amfenac (1.23), suggesting that bromfenac may
`have increased pene tration into ocular tissues and poten-
`tially a more rapid onset of action.15 A pharmacokinetic
`study in patients undergoing cataract surgery showed that
`bromfenac was absorbed within 30 minutes following
`instillation of bromfenac ophthalmic solution 0.1%, with
`a peak aqueous humor concentration at 2.5 to 3 hours after
`instillation.18 In addition, a previous animal study demon-
`strated that bromfenac 0.09%, pH 8.3 readily penetrates
`ocular tissues.19
`The objective of the current two studies was to evaluate
`the ocular distribution of the new modified formulation of
`bromfenac ophthalmic solution. The first study evaluated the
`12-hour ocular distribution profile of 14C-bromfenac ophthal-
`mic solution 0.07%, pH 7.8 compared with 14C-bromfenac
`ophthalmic solution 0.09%, pH 8.3, while the second study
`evaluated the 24-hour ocular distribution of 14C-bromfenac
`ophthalmic solution 0.07%, pH 7.8 to confirm sustained
`ocular penetration with once-daily dosing.
`
`Methods
`Two separate preclinical studies were performed at the
` Biological Test Center ([BTC] Irvine, CA, USA) using
`similar protocols. Each study complied with the animal wel-
`fare policies of the BTC, was approved by the Institutional
`Animal Care and Use Committee (IACUC), and conformed
`to the ARVO (Association for Research in Vision and
` Ophthalmology) “Statement on the Use of Animals in
` Ophthalmic and Visual Research.”
`
`Test article
`The 14C-bromfenac was manufactured by Daiichi Pure
`Chemicals Co., Ltd., and provided by Senju Pharmaceuticals
`Co., Ltd., (Osaka, Japan). It was determined to be 98.3% pure
`by high-performance liquid chromatography (HPLC). The
`radiolabeled carbon atom was that of the carbonyl within
`the bromobenzoyl group of bromfenac (Figure 1), a stable
`position within the molecule.
`Dosing solutions were prepared by adding 14C-bromfenac
`to diluent to reach a concentration of 0.07%, pH 7.8 or
`0.09%, pH 8.3. The resulting 14C-bromfenac 0.07%, pH 7.8
`and the 0.09%, pH 8.3 formulations used in the studies
`
`1718
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`Ocular distribution of 14C-bromfenac solution
`
`NH2
`
`O
`
`*
`
`COONa 1.5 H2O
`
`•
`
`Br
`
`Figure 1 Molecular structure of bromfenac showing the position of the 14C label.
`Note: *14C.
`
`were identical to the marketed Prolensa® and Bromday®
`formulations, respectively, with the exception of the inclusion
`of 14C-bromfenac. Following dosing, dosing solutions were
`reanalyzed by HPLC to confirm radioactive purity; all
`samples were determined to be 95% to 99% pure.
`
`animals
`In both studies, female New Zealand White rabbits were
`obtained from The Rabbit Source (Ramona, CA, USA). Ani-
`mals were at least 12 weeks of age and weighed 1.9 to 3.6 kg
`at the time of dosing. Animals were housed in individual
`cages and identified with ear tags and cage cards.
`
`Pretreatment examination and dosing
`procedures
`Prior to placement in the study, each animal underwent a
`pretreatment ophthalmic examination as well as a slit-lamp
`examination. Criteria for acceptance into the study were
`severity scores of 1 (on 0–3 or 0–4 scales) for conjuncti-
`val congestion and swelling, and the absence of any other
`slit-lamp signs. Prior to dosing, animals were weighed
`and randomly assigned to a study group. For the 12-hour
`study, animals were divided into groups receiving either
`14C-bromfenac 0.07%, pH 7.8 or 14C-bromfenac 0.09%,
`pH 8.3 for collection of ocular tissues at one of five different
`time points (1, 2, 4, 8, and 12 hours). For the 24-hour study,
`animals were divided into groups receiving 14C-bromfenac
`0.07%, pH 7.8 for collection of ocular tissue at one of six
`different time points (1, 2, 4, 8, 12, and 24 hours). There was
`no comparator arm in the 24-hour study.
`On day 1 of each study, 50 µL of 14C-bromfenac dosing
`solution was administered into the conjunctival sac of each
`animal using a calibrated digital pipette, and the time of dose
`administration was noted. The eyelids were held closed post-
`dose for 5–10 seconds. Actual dose in mg and µCi were used
`in all subsequent calculations.
`In the 12-hour parallel-group study, animals received
`either 14C-bromfenac 0.07%, pH 7.8 or 14C-bromfenac 0.09%,
`pH 8.3 in one eye. The specific activity of bromfenac in these
`
`formulations was 528,494 dpm/µg (or 0.23 µCi/µg) and
`359,952 dpm/µg (or 0.16 µCi/µg), respectively. Three rabbit
`eyes were dosed per treatment per time point for a total of
`15 eyes dosed for each test agent. Animals were euthanized by
`intravenous injection of a commercial euthanasia solution per
`the standard operating procedure of the animal facility (BTC)
`at predetermined time points (1 hour [±5 minutes], 2 hours
`[±15 minutes], 4 hours [±15 minutes], 8 hours [±15 minutes],
`or 12 hours [±15 minutes]) after instillation of test agent, and
`ocular tissues were immediately harvested. In the 24-hour
`study, animals were administered 14C-bromfenac 0.07%, pH
`7.8 only (specific activity of 411,032 dpm/µg or 0.18 µCi/µg).
`Both eyes from three rabbits were dosed per time point
`over a 24-hour period for a total of six eyes dosed per time
`point. Animals were euthanized as described above at 1 hour
`(±5 minutes), 2 hours (±15 minutes), 4 hours (±15 minutes),
`8 hours (±15 minutes), 12 hours (±15 minutes), or 24 hours
`(±15 minutes) after administration of test agent, and ocular
`tissues harvested.
`In both studies, animals were observed for morbidity and
`mortality throughout the study.
`
`sample processing and radioactivity
`measurements
`Tissues harvested included the aqueous humor, vitreous
`humor, iris/ciliary body, lens, retina, choroid, sclera (both
`studies), and cornea and conjunctiva (24-hour study only).
`While the 12-hour study focused on the tissues most relevant
`to treating postoperative inflammation, the 24-hour study,
`conducted later, was patterned from a previously developed
`24-hour drug exposure protocol19 and focused on all ocular
`tissues and therefore also included the cornea and conjunc-
`tiva. Immediately following euthanasia, the aqueous humor
`was collected from the eye and frozen at -20°C if not imme-
`diately analyzed. The globe was then enucleated, snap-frozen
`in liquid nitrogen, and stored at -70°C until dissection.
`Iris/ciliary body, lens, retina, choroid, sclera (both
`studies), and cornea and conjunctiva (24-hour study only)
`were dissected from each eye, weighed into combustion
`cones, and combusted. Combusted samples were trapped
`in 14C-Cocktail (RJ Harvey, Hillsdale, NJ, USA) in liquid
`scintillation chromatography (LSC) vials and the amount
`of radioactivity was determined by LSC. To determine the
`amount of radioactivity in aqueous humor and vitreous
`humor, duplicate aliquots of each aqueous humor sample
`(25 µL) and homogenized vitreous humor sample (100 µL)
`were transferred to LSC vials and the amount of radioactiv-
`ity determined following the addition of 10 mL of Insta-Gel
`
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`Baklayan and Muñoz
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`(Perkin-Elmer Life and Analytical Sciences, Inc., Waltham,
`MA, USA).
`Radioactivity measurements were made using a Beckman
`Liquid Scintillation Spectrometer (Beckman Coulter Inc.,
`Fullerton, CA, USA). Counting time was to a statistical accu-
`racy of ±2% or a maximum of 10 minutes, whichever came
`first. Background noise was automatically subtracted from
`each measurement and counts per minute were automatically
`converted to disintegrations per minute.
`
`statistical analysis
`To determine the specific activity (dpm/µg) of 14C-bromfenac,
`the following formula was used:
`
`Specific
`activity
`(dpm/µg)
`
`=
`
`Dosing solution
`radioactivity (dpm/g)
`Bromfenac concentration
`of dosing solution (µg/mL)
`
` ×
`
`Dosing
` (1)
`solution
`density (g/mL)
`
`To determine bromfenac-equivalent concentration of
`dosed 14C-bromfenac, the following formula was used:
`
`
`
`Bromfenac equivalent
`concentration (µg eq/g)
`
`=
`
`Sample radioactivity
`(dpm/g)
`Specific activity of
`bromfenac (dpm/µg)
`
`
`
`(2)
`
`Mean and standard deviation of each set of samples were
`used to characterize the data. Although the sample size (N=3
`eyes per treatment per time period) in the 12-hour study was
`not powered to find a specific difference, Student’s t-tests
`were used to compare the groups.
`
`Results
`12-hour pharmacokinetic study
`Following the administration of either 14C-bromfenac 0.07%,
`pH 7.8 or 14C-bromfenac 0.09%, pH 8.3, concentrations of
`bromfenac ranged from 0.000 to 0.595 µg eq/g depending on
`the tissue. The highest 14C-bromfenac levels were found in
`the sclera, followed by the iris/ciliary body, aqueous humor,
`choroid, retina, and lens (Table 1, Figures 1 and 2). The tim-
`ing of peak concentrations of radiolabeled bromfenac varied
`among the tissues, with peak levels being reached at 1 hour
`in the sclera and retina and at 2 hours in the aqueous humor,
`iris/ciliary body, lens, and choroid. There was a significant
`difference between the formulations at the 2-hour time
`point for scleral 14C-bromfenac levels, with higher levels
`in the animals randomized to the bromfenac 0.07%, pH 7.8
`treatment group (P0.001). However, there were no other
`
`between-treatment differences at any other time points in
`any tissue. The total amount (ie, sum) of 14C-bromfenac in
`all ocular tissues evaluated was directionally higher in the
`14C-bromfenac 0.07%, pH 7.8 group compared with the
`14C-bromfenac 0.09%, pH 8.3 group in four out of five time
`points, but the differences were not statistically significant.
`
`24-hour pharmacokinetic study
`After administration of 14C-bromfenac 0.07%, pH 7.8,
`14C-bromfenac was detected in all tissues of the eye over a
`24-hour period, with the exception of vitreous humor. As
`expected, the highest concentrations were seen in the cornea,
`followed by conjunctiva and sclera (Table 2 and Figure 3).
`The concentrations in all ocular tissues decreased to varying
`degrees over the 24-hour study period, with the exception
`of the lens, which exhibited little change from the 1-hour
`time point.
`No adverse events were observed in either study.
`
`Discussion
`Ocular inflammation following surgery arises from a
`complex series of intracellular signaling events involving
`COX-1 and COX-2 enzymes. In particular, activation of the
`COX-2 signaling pathway leads to the production of PGs and
` thromboxane A2, which ultimately alters vascular permeabil-
`ity by changing the function of vascular endothelial cells.8
`Topical NSAIDs are a class of anti-inflammatory drugs that
`have been used following cataract and other ocular surgeries,
`as well as in ocular disorders affecting the posterior chamber
`of the eye, due to their ability to inhibit the COX signaling
`pathways, thereby decreasing inflammation. In order for any
`topical NSAID to achieve its desired effects, the drug must
`penetrate the tissues of the eye, target the induced COX
`enzymes, and remain sustained within the tissue long enough
`to prevent or reduce PG effects. Once-daily dosing reduces
`the need for repeated applications and potentially improves
`patient adherence.
`Bromfenac topical ophthalmic solution has been shown to
`quickly permeate the tissues of the eye, maintain measurable
`concentrations for up to 24 hours, and effectively inhibit the
`COX-2 signaling pathway, making it an effective NSAID in
`the treatment of ocular inflammation.9 Several studies have
`compared bromfenac with other topical NSAIDs for the
`ability to inhibit COX-2 activity and hence inhibit inflam-
`mation. Yanni et al compared the IC50 values of bromfenac,
`diclofenac, and amfenac for purified rabbit COX-2 and
`found that the inhibitory effect of bromfenac on COX-2 was
`3.7 times greater than that of diclofenac and 6.5 times greater
`
`1720
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`Ocular distribution of 14C-bromfenac solution
`
`Table 1 Concentration of 14C-bromfenac in ocular tissues over 12 hours following topical administration of 14C-bromfenac 0.07%, ph
`7.8 or 14C-bromfenac 0.09%, ph 8.3
`
`Sample,
`mean (SD)
`
`Mean (± SD) bromfenac equivalents in µg eq/g
`12 hours
`8 hours
`4 hours
`1 hour
`2 hours
`0.09%,
`0.07%,
`0.09%,
`0.07%,
`0.09%,
`0.07%,
`0.09%,
`0.09%,
`0.07%,
`0.07%,
`pH 8.3
`pH 7.8
`pH 8.3
`pH 7.8
`pH 8.3
`pH 7.8
`pH 8.3
`pH 8.3
`pH 7.8
`pH 7.8
`0.010
`0.011
`0.018
`0.018
`0.052
`0.038
`0.137
`0.051
`0.048
`0.090
`(0.005)
`(0.003)
`(0.015)
`(0.004)
`(0.021)
`(0.011)
`(0.063)
`(0.017)
`(0.024)
`(0.024)
`0.001
`0.000
`0.000
`0.000
`0.000
`0.000
`0.000
`0.000
`0.000
`0.000
`(0.000)
`(0.000)
`(0.000)
`(0.000)
`(0.000)
`(0.000)
`(0.000)
`(0.000)
`(0.000)
`(0.000)
`0.022
`0.018
`0.034
`0.029
`0.061
`0.048
`0.083
`0.078
`0.081
`0.110
`(0.009)
`(0.001)
`(0.017)
`(0.013)
`(0.016)
`(0.005)
`(0.013)
`(0.027)
`(0.037)
`(0.044)
`0.003
`0.002
`0.002
`0.002
`0.002
`0.002
`0.002
`0.002
`0.001
`0.003
`(0.003)
`(0.000)
`(0.002)
`(0.001)
`(0.000)
`(0.000)
`(0.001)
`(0.001)
`(0.000)
`(0.001)
`0.002
`0.003
`0.002
`0.002
`0.010
`0.006
`0.015
`0.018
`0.013
`0.010
`(0.001)
`(0.004)
`(0.001)
`(0.000)
`(0.004)
`(0.001)
`(0.003)
`(0.005)
`(0.005)
`(0.002)
`0.008
`0.008
`0.010
`0.011
`0.043
`0.035
`0.083
`0.058
`0.045
`0.062
`(0.002)
`(0.002)
`(0.004)
`(0.004)
`(0.022)
`(0.007)
`(0.030)
`(0.010)
`(0.012)
`(0.011)
`0.039
`0.067
`0.026
`0.058
`0.220
`0.170
`0.302
`0.346
`0.595
`0.451
`(0.015)
`(0.045)
`(0.020)
`(0.019)
`(0.160)
`(0.026)
`(0.006)
`(0.134)
`(0.156)
`(0.028)
`0.388
`0.300
`0.623
`0.553
`0.783
`0.726
`0.085
`0.109
`0.092
`0.121
`(0.218)
`(0.048)
`(0.080)
`(0.186)
`(0.231)
`(0.088)
`(0.031)
`(0.043)
`(0.058)
`(0.005)
`Notes: n=15 for each treatment, three rabbits per time point. All differences between formulations were statistically insignificant at all time points with the exception of
`the 2-hour scleral drug level (P0.001). All differences between formulations in total drug levels were statistically insignificant (P0.20) using a student’s t-test, without
`correction for multiple comparisons.
`Abbreviation: sD, standard deviation.
`
`aqueous humor
`
`Vitreous humor
`
`iris/ciliary body
`
`lens
`
`retina
`
`Choroid
`
`sclera
`
`Total
`
`than that of amfenac.14 An additional study comparing the
`IC50 of bromfenac to amfenac, diclofenac, and ketorolac using
`recombinant human COX-2 demonstrated that bromfenac
`had the greatest activity against human COX-2, followed by
`amfenac, ketorolac, and diclofenac.20
`
`An advanced formulation of bromfenac ophthalmic
`solution was recently developed which contains a lower
`concentration of bromfenac at a lower pH than previous
`formulations. It was hypothesized that reducing the pH
`from 8.3 from 7.8 would allow for a lower-concentration
`
`Table 2 Concentration of 14C-bromfenac in ocular tissues over 24 hours following topical administration of 14C-bromfenac 0.07%,
`ph 7.8
`
`Sample,
`mean (SD)
`
`aqueous humor
`
`Vitreous humor
`
`iris/ciliary body
`
`lens
`
`retina
`
`Choroid
`
`sclera
`
`Cornea
`
`Conjunctiva
`
`Mean (± SD) bromfenac equivalents in µg eq/g
`1 hour
`2 hours
`4 hours
`0.057
`0.064
`0.049
`(0.014)
`(0.008)
`(0.019)
`0.000
`0.000
`0.000
`(0.000)
`(0.000)
`(0.000)
`0.077
`0.082
`0.058
`(0.016)
`(0.015)
`(0.020)
`0.001
`0.001
`0.001
`(0.000)
`(0.000)
`(0.001)
`0.012
`0.010
`0.007
`(0.006)
`(0.004)
`(0.001)
`0.054
`0.036
`0.032
`(0.012)
`(0.012)
`(0.010)
`0.477
`0.265
`0.149
`(0.085)
`(0.104)
`(0.062)
`2.402
`1.998
`1.219
`(0.534)
`(0.407)
`(0.529)
`1.049
`0.485
`0.355
`(0.138)
`(0.213)
`(0.181)
`
`Note: n=3 per time point.
`Abbreviation: sD, standard deviation.
`
`Clinical Ophthalmology 2014:8
`
`8 hours
`0.017
`(0.006)
`0.000
`(0.000)
`0.026
`(0.007)
`0.001
`(0.000)
`0.002
`(0.001)
`0.010
`(0.005)
`0.075
`(0.058)
`0.471
`(0.230)
`0.137
`(0.067)
`
`12 hours
`0.006
`(0.002)
`0.000
`(0.000)
`0.017
`(0.002)
`0.001
`(0.000)
`0.001
`(0.000)
`0.005
`(0.001)
`0.018
`(0.005)
`0.228
`(0.092)
`0.079
`(0.056)
`
`24 hours
`0.00
`(0.001)
`0.000
`(0.000)
`0.010
`(0.005)
`0.002
`(0.001)
`0.001
`(0.000)
`0.002
`(0.001)
`0.009
`(0.013)
`0.080
`(0.050)
`0.031
`(0.040)
`
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`bromfenac 0.07%, pH 7.8 was able to penetrate various
`ocular tissues and remain in those tissues at measurable
`levels for up to 24 hours. Even at a lower concentration, the
`bromfenac 0.07%, pH 7.8 solution exhibited similar, and in
`the case of scleral tissue, increased, penetration of ocular
`tissues studied, when compared with bromfenac 0.09%,
`pH 8.3 solution. This is likely due to the difference in pH
`between the solutions; lowering the pH increases the union-
`ized fraction of drug, which can lead to enhanced corneal
`permeability.7 Additionally, the reduction in pH to a more
`physiologic level could reduce any potential for discomfort
`and irritation upon instillation. Although we used a healthy
`eye model in these studies, ocular penetration is likely to be
`higher in inflamed eyes due to weakened barriers from the
`underlying pathological condition.21 However, any differ-
`ences between formulations is expected to be the same in
`an inflamed eye model.
`Of note, if one uses an estimate of 1 mL for the volume
`of a gram of tissue, it follows that ocular tissue levels of
`bromfenac following topical administration of bromfenac
`ophthalmic solution 0.07%, pH 7.8 were sustained at con-
`centrations above the reported IC50 of bromfenac for the
`COX-2 enzyme (0.0066 µM or 2.5 pg/mL) in all tissues
`evaluated for the duration of each study, with the exception of
`vitreous humor. That negligible levels of radiolabeled brom-
`fenac were found in the vitreous humor following dosing
`with bromfenac in the 12-hour study as well as the 24-hour
`study independent of formulation was not surprising and was
`consistent with previous findings.19 As a highly lipophilic
`drug, bromfenac has a more rapid drug-transit time through
`tissues and/or choroidal blood flow to posterior-segment
`
`Sclera
`Iris/ciliary
`body
`Aqueous
`humor
`Choroid
`Retina
`Lens
`
`Sclera
`Iris/ciliary
`body
`Aqueous
`humor
`Choroid
`Retina
`Lens
`
`1
`
`2
`
`4
`Time (h)
`
`8
`
`12
`
`1
`
`2
`
`4
`Time (h)
`
`8
`
`12
`
`10
`
`1
`
`0.1
`
`0.01
`
`0.001
`
`0.0001
`
`10
`
`1
`
`0.1
`
`0.01
`
`0.001
`
`0.0001
`
`A
`
`Concentration (μg eq/g)
`
`Concentration (μg eq/g)B
`
`Figure 2 Ocular distribution of 14C-bromfenac in rabbits over 12 hours following
`topical administration of 14C-bromfenac 0.07%, ph 7.8 (A) or 14C-bromfenac 0.09%,
`ph 8.3 (B).
`
`formulation to provide comparable penetration into ocular
`tissues as the reference product, bromfenac 0.09%, which has
`had considerable marketplace success. The 12- and 24-hour
`ocular distribution studies presented here demonstrate that
`
`Cornea
`Conjunctiva
`Sclera
`Iris/ciliary
`body
`Aqueous
`humor
`Choroid
`Retina
`Lens
`
`1
`
`2
`
`4
`
`8
`
`12
`
`24
`
`Time (h)
`
`10
`
`1
`
`0.1
`
`0.01
`
`0.001
`
`0.0001
`
`Concentration (μg eq/g)
`
`Figure 3 Ocular distribution of 14C-bromfenac in rabbits over 24 hours following topical administration of 14C-bromfenac 0.07%, ph 7.8.
`
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`
`Ocular distribution of 14C-bromfenac solution
`
`tissues, and is not expected to concentrate appreciably in
`the vitreous humor.
`Previous studies of 14C-bromfenac ophthalmic solution
`0.09%, pH 8.3 in rabbits have demonstrated that this solu-
`tion rapidly achieved measurable levels in all major ocular
`tissues at or before 2 hours and detectable levels of brom-
`fenac were sustained for up to 24 hours following a single
`ophthalmic dose.19 Similar to the results of the 24-hour study
`with bromfenac 0.07%, pH 7.8, the highest concentrations
`of radiolabeled bromfenac were detected in the cornea, con-
`junctiva, and sclera.19 Additional studies in human subjects
`undergoing cataract surgery have shown that a single dose of
`bromfenac ophthalmic solution 0.1% led to rapid absorption
`of the drug within 30 minutes of treatment and therapeutic
`levels (ie, above IC50 for bromfenac) were maintained within
`the ocular tissue for up to 12 hours.18
`The ability of ophthalmic NSAIDs to inhibit PG synthe-
`sis suggests that they may be effective for ocular disorders
`affecting posterior tissues, such as cystoid macular edema
`(CME).22–25 While the use of NSAIDs in the prevention and
`treatment of CME is not approved by the US FDA, various
`studies evaluating the efficacy of bromfenac 0.9% ophthalmic
`solution in the management of CME26–30 and choroidal neo-
`vascularization associated with age-related macular degener-
`ation31–33 have shown a potential benefit of bromfenac in the
`management of these disorders. The results of these 12- and
`24-hour ocular distribution studies suggest that the ability of
`bromfenac to quickly and effectively penetrate ocular tissues,
`including posterior tissues, may support its use in treating
`other ocular inflammatory diseases, including CME.
`
`Conclusion
`As a new, advanced formulation of bromfenac ophthalmic
`solution, developed for once-daily dosing with a lower
`concentration and lower pH than previous formulations,
`it was important to evaluate the ocular distribution of
`14C-bromfenac ophthalmic solution 0.07%, pH 7.8 up to
`24 hours post-administration and to compare its ocular
`distribution with that of 14C-bromfenac ophthalmic solution
`0.09%, pH 8.3. The pH of bromfenac ophthalmic solution
`0.07% (Prolensa®) was reduced to 7.8, thereby increasing the
`ratio of non-ionized to ionized fraction and allowing for a
`lower concentration of active ingredient, while maintaining
`the ability to rapidly penetrate ocular tissues and remain in
`relevant ocular tissues for up to 24 hours. While these studies
`on the ocular distribution of bromfenac ophthalmic solution
`0.07% were conducted in rabbits, the results suggest that the
`new formulation has the potential to be an effective agent
`
`for the treatment of inflammation in a number of ocular
`inflammatory diseases. Further clinical explorations into
`the use of bromfenac solution 0.07% in these settings will
`provide insight into the full range of therapeutic possibilities
`for bromfenac ophthalmic solution.
`
`Acknowledgments
`The authors wish to express our appreciation for the assis-
`tance of Chantelle Rein-Smith, PhD and Natalie Her, PhD
`of Whitsell Innovations Inc., for manuscript preparation
`and for the assistance of Linda Wang, PharmD, University
`of Southern California fellow training at Bausch + Lomb,
`for editorial assistance and data review/verification. This
`study was presented in part at the 85th Annual Meeting of
`the Association for Research in Vision and Ophthalmology,
`Inc.; May 5–9, 2013; Seattle, WA, USA.
`
`Disclosure
`George A Baklayan is an employee of Bausch + Lomb.
`Mauricio Muñoz was an employee of Bausch + Lomb at
`the time the manuscript was developed. Study support was
`provided by ISTA Pharmaceuticals, Inc., (Irvine, CA, USA).
`The sponsor participated in the design of the study, data col-
`lection, data management, data analysis, interpretation of the
`data, and preparation, review, and approval of the manuscript.
`Bausch + Lomb acquired ISTA Pharmaceuticals in June 2012.
`All bromfenac products are now marketed under the Bausch +
`Lomb trademark, unless otherwise specified. The preclinical
`trial discussed in this manuscript was conducted by ISTA
`Pharmaceuticals, Inc., and all rights were transferred to
`Bausch + Lomb upon completion of the acquisition. The
`authors report no other conflicts of interest in this work.
`
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