Rabbit Ocular Tissues Research Articles

Biocompatibility of Nb2C MXene film for ophthalmic implants

The Nb2C MXene film was prepared by tetrafluoroboric acid (HBF4) and hydrofluoric acid (HF) etching and vacuum extraction. The surface structure, properties, and composition of Nb2C MXene were characterized by scanning electron microscopy (SEM), atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS), and other methods. The biocompatibility of Nb2C MXene film was evaluated by in vivo and in vitro experiments. The results show the layered, micro–nano surface structure of Nb2C MXene with abundant hydroxyl groups (—OH) on the surface. Compared with the control group, the proliferation rate of conjunctival fibroblasts cultured with 1 mg/mL Nb2C MXene is 72.15% (p<0.05). The cytotoxicity of Nb2C MXene is Grade 0 (p>0.05), and the hemolysis rate is 0.38% (<5%). The oxidative stress and inflammatory responses in rabbit ocular tissues are inhibited by Nb2C MXene film, which promotes wound healing without inducing fibrosis, and its micro–nano structure promotes cell adhesion and proliferation, demonstrating good biocompatibility.

Phospholipid analyses of rabbit ocular surface tissues

The tissues of the integument covering the ocular surface comprise a mucus membrane functioning as a protective physical barrier and has the ability to mount a defensive inflammatory response. Since lipid metabolism has a role in both of these functions, we studied normal membrane phospholipids (PL) of the cornea and bulbar conjunctiva to (1) determine baseline PL profiles of these tissues, (2) compare and contrast these individual PL metabolite profiles as well as groups of metabolites, and (3) describe pathway-specific metabolic interrelations among these tissues. Corneal and conjunctival tissue samples were isolated from rabbit eyes (n = 30) and extracted with chloroform-methanol using a modified Folch procedure. 31P nuclear magnetic resonance spectroscopy was used to qualitatively and quantitatively measure tissue PL profiles. The cornea and conjunctiva, respectively, have the following PL composition (mole % of total detected phospholipid): phosphatidylglycerol (PG) –, 0.4; lysophosphatidylethanolamine 1.2, –; phosphatidic acid –, 0.4; diPG (cardiolipin) 2.1, 3.5; unknown PL at the chemical shift of 0.13 δ 1.5, 0.9; ethanolamine plasmalogen 11.2, 13.0; phosphatidylethanolamine 11.5, 12.8; phosphatidylserine 8.9, 10.1; sphingomyelin 10.2, 10.7; lysophosphatidylcholine 0.9, 1.4; phosphatidylinositol 5.3, 5.3; phosphatidylcholine (PC) plasmalogen or alkylacylPC 2.2, 1.9; PC 45.1, 40.0. In addition, 28 PL metabolic indices were calculated from these data, which permitted pathway-specific lipid analyses. This study (1) establishes PL profiles of the two ocular tissues of the integument that cover the surface of the eye, (2) compares and contrasts indices comprised of ratios and combinations of PL, and (3) describes pathway-specific metabolic interrelations among these tissues to serve as baselines for studies involving the distribution of tissue phospholipids.

Simultaneous determination of amphotericin B, tobramycin and vancomycin in rabbit ocular biofluids and tissues by LC-MS/MS: An antimicrobial therapy for keratitis and its PK-PD application

A rationale poly-microbial keratitis (PMK) therapy requires quick identification of pathogen (bacteria and fungi) and their efficient treatment. However, majority of healthcare providers are still having trouble finding an effective medicine to treat PMK due to constraints such as antimicrobial resistance, dose and dosing schedule. Thus, a broad spectrum anti-fungal and antibacterial having less resistance in community involving combination therapy such as amphotericin B (AmB), tobramycin (TBR) and vancomycin (VCM) is required. Hence, to characterize the pharmacokinetic (PK) and PK-pharmacodynamic (PD) indices, a rapid and sensitive simultaneous LC-MS/MS bioanalytical method was developed and validated for the quantification of AmB, TBR and VCM in rabbit ocular biofluids and tissues. Chromatographic resolution was achieved on a Zorbax C18 column with a mobile phase composed of acetonitrile and 0.4 % formic acid in deionized water using a gradient mode of elution. The calibration curves showed good linearity over the concentration range of 1.95–500 ng/mL for AmB and TBR, 3.9–800 ng/mL for VCM, respectively. The lower limit of quantification (LLOQ) was found to be 1.95 ng/mL for AmB and TBR, and 4.5 ng/mL for VCM. Analyte extraction was performed by simple protein precipitation method with minimal sample volume of 10 µL. Finally, the developed method was validated for selectivity, linearity (r2 > 0.99), precision, accuracy, matrix effects, and stability. The ocular pharmacokinetic profile of commercial AmB, TBR, and VCM formulations was further assessed using the validated method and the PK-PD indices along with dosing frequency was predicted by PK-PD modelling using Phoenix WinNonlin Software.

Aldehyde oxidase 1 activity and protein expression in human, rabbit, and pig ocular tissues

Aldehyde oxidase (AOX) is a cytosolic drug-metabolizing enzyme which has attracted increasing attention in drug development due to its high hepatic expression, broad substrate profile and species differences. In contrast, there is limited information on the presence and activity of AOX in extrahepatic tissues including ocular tissues. Because several ocular drugs are potential substrates for AOX, we performed a comprehensive analysis of the AOX1 expression and activity profile in seven ocular tissues from humans, rabbits, and pigs. AOX activities were determined using optimized assays for the established human AOX1 probe substrates 4-dimethylamino-cinnamaldehyde (DMAC) and phthalazine. Inhibition studies were undertaken in conjunctival and retinal homogenates using well-established human AOX1 inhibitors menadione and chlorpromazine. AOX1 protein contents were quantitated with targeted proteomics and confirmed by immunoblotting. Overall, DMAC oxidation rates varied over 10-fold between species (human ˃˃ rabbit ˃ pig) and showed 2- to 6-fold differences between tissues from the same species. Menadione seemed a more potent inhibitor of DMAC oxidation across species than chlorpromazine. Human AOX1 protein levels were highest in the conjunctiva, followed by most posterior tissues, whereas anterior tissues showed low levels. The rabbit AOX1 expression was high in the conjunctiva, retinal pigment epithelial (RPE), and choroid while lower in the anterior tissues. Quantification of pig AOX1 was not successful but immunoblotting confirmed the presence of AOX1 in all species. DMAC oxidation rates and AOX1 contents correlated quite well in humans and rabbits. This study provides, for the first time, insights into the ocular expression and activity of AOX1 among multiple species.

Establishment of an LC-MS/MS method for quantification of lifitegrast in rabbit plasma and ocular tissues and its application to pharmacokinetic study

Lifitegrast, a lymphocyte function-associated antigen 1 antagonist, was approved by the FDA for the treatment of dry eye disease. Cornea and conjunctiva have been reported to be the sites of action of lifitegrast. To investigate the pharmacokinetics of lifitegrast, a sensitive analytical method for the determination of lifitegrast in various biological matrices such as plasma and ocular tissues is required. However, only limited information about the analytical method for lifitegrast in biological samples is available. In the present study, we aimed to develop a new liquid chromatography-tandem mass spectrometry method for the determination of lifitegrast in rabbit plasma, cornea, conjunctiva, and sclera. Lifitegrast-d6 was used as an internal standard (IS). To prepare the biological samples, protein precipitation using acetonitrile was utilized. Analytes were separated from endogenous interferences on an Atlantis dC18 (5 µm, 2.1 × 150 mm), and a mixture of 0.1 % formic acid and acetonitrile was used as the mobile phase. The mass transition of precursor to product ion was monitored at 615.2 → 145.0 for lifitegrast and 621.2 → 145.1 for IS. The calibration curves were linear over the concentration range from 2 to 500 ng/mL for plasma and 5 to 500 ng/mL in ocular tissue homogenates. Intra- and inter-day accuracy ranged from 95.76 to 106.80 % in the plasma and 94.42 to 112.80 % in the ocular tissues. Precision was within 8.56 % in the plasma and 9.72 % in the ocular tissues. The short-term, long-term, auto-sampler, and freeze–thaw stabilities of lifitegrast were validated. The developed method was applied to a pharmacokinetic study of lifitegrast in rabbits. Following ophthalmic administration, only 3.26 % of administered lifitegrast was absorbed into the systemic circulation. Peak tissue concentrations were observed at 0.5 h after dosing, and topically administered lifitegrast was mainly distributed in the cornea and conjunctiva. The finding of this study is expected to be used in further pharmacokinetic studies and formulation development.

LC/MS Assessment of Glycoform Clearance of A Biotherapeutic MAb in Rabbit Ocular Tissues

Many biotherapeutics such as monoclonal antibodies (mAbs) consist of various glycoforms, which can have different PK properties upon administration to animals and human. As a result, it is necessary to monitor the abundance of glycoforms and limit lot-to-lot variability during the manufacturing process. However, limited information is known about the clearance of mAb glycoforms from ocular space upon intravitreal injection. We present here an assessment of glycoform clearance of a biotherapeutic mAb (IgG1) from rabbit vitreous humor, aqueous humor and retina tissue using LC/MS. The results show that G0, G0F and G1F have similar T1/2, while mannose-5 has a longer T1/2 and is cleared slower in rabbit ocular space, which contradicted with what has been reported in the literature in which Mann5 was cleared faster systematically.

Simultaneous Determination of Moxifloxacin Hydrochloride and Dexamethasone Sodium Phosphate in Rabbit Ocular Tissues and Plasma by LC-MS/MS: Application for Pharmacokinetics Studies.

Treatment of ocular infection involves pharmacotherapy with steroids and antibiotic drops, such as moxifloxacin hydrochloride (MFH) and dexamethasone sodium phosphate (DSP). To characterize the pharmacokinetics of these two compounds, we performed and validated a liquid chromatography-mass spectrometry (LC-MS/MS) method to quantify them in rabbit ocular tissues and plasma. We used protein precipitation to extract the compounds. The analyte and internal standard (IS) were separated using a Shim-pack Scepter C18 column. The mobile phase was composed of 0.1% formic acid water (A) and methanol (B). MFH and DSP were detected using positive ion electrostatic ionization (ESI) in multiple reaction monitoring mode (MRM). The calibration curves for both compounds showed good linearity over concentrations ranging from 0.5 to 200 ng/mL in rabbit ocular tissues and plasma. The lower limit of quantification for both MFH and DSP was 0.5 ng/mL. We validated this method for selectivity, linearity (r2 &gt; 0.99), precision, accuracy, matrix effects, and stability. Thus, we used this method to assess the pharmacokinetic (PK) characteristics of MFH and DSP in rabbit ocular tissues and plasma after single doses. Our results indicate that this method can be used for the simultaneous analysis of moxifloxacin hydrochloride and dexamethasone sodium phosphate in clinical samples.

A sensitive and rapid bioanalytical method for the quantitative determination of luliconazole in rabbit eye tissues using UPLC-MS/MS assay

Luliconazole (LCZ) is a novel antifungal imidazole with broad-spectrum and high susceptibility of Aspergillus and Fusarium are the dominant species of fungal keratitis, may potentially be a new medical treatment option for ocular fungal infection. To evaluate LCZ distribution in ocular tissues after topical application for the development of ophthalmic delivery system, it is important to have a bioanalytical method for measuring the drug concentrations in different ocular tissues and aqueous humor (AH). A selective and sensitive ultrahigh performance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS) method was developed for the quantification of LCZ in rabbit ocular tissues, including conjunctiva, cornea, AH, iris, lens, vitreous humor (VH), retinal choroid and sclera, using lanoconazole as internal standard (IS). Chromatographic separation was achieved on a Xterra MS, C18 column (2.1 × 50 mm, 3.5 μm) using mobile phase with formic acid solution (0.2%, v/v): acetonitrile (50:50, v/v) at a flow rate of 0.2 ml/min, and the run time was 2.5 min. Detection was performed using the transitions 354.1 → 150.3 m/z for LCZ and 320.1 → 150.3 m/z for IS by positive ion electrospray ionization in multiple reaction monitoring (MRM) mode. Method validation was conducted in accordance with U.S. Food and Drug Administration’s regulatory guidelines for bioanalytical method validation. The calibration curves were linear over the concentration range from 2.80 ng/ml to 2038 ng/ml for conjunctiva, cornea and sclera, 2.09 ng/ml to 1019 ng/ml for AH, 2.09 ng/ml to 509.5 ng/ml for iris, 2.09 ng/ml to 203.8 ng/ml for retinal choroid and VH, 2.04 ng/ml to 101.9 ng/ml for lens, with all the squared correlation coefficients (r2) more than 0.99. The accuracy of the method was within the acceptable limit of 89.34%∼112.78% at the lower limit of quantification and other concentrations, Inter-day and intra-day precision values, expressed in terms of RSD (%), in all tissues were within 15% at all concentrations. The mean recoveries of LCZ in rabbit ocular tissues was 84.85%∼100.52%. No interference was found due to matrix components. Luliconazole was stable during the stability studies, including autosampler stability, benchtop stability, freeze/thaw stability and long-term stability. The method was successfully applied to the ocular pharmacokinetic and tissues distribution studies of LCZ in rabbit after topical administration of LCZ ophthalmic drug delivery system.

Carboxylesterase Activities and Protein Expression in Rabbit and Pig Ocular Tissues.

Hydrolytic reactions constitute an important pathway of drug metabolism and a significant route of prodrug activation. Many ophthalmic drugs and prodrugs contain ester groups that greatly enhance their permeation across several hydrophobic barriers in the eye before the drugs are either metabolized or released, respectively, via hydrolysis. Thus, the development of ophthalmic drug therapy requires the thorough profiling of substrate specificities, activities, and expression levels of ocular esterases. However, such information is scant in the literature, especially for preclinical species often used in ophthalmology such as rabbits and pigs. Therefore, our aim was to generate systematic information on the activity and expression of carboxylesterases (CESs) and arylacetamide deacetylase (AADAC) in seven ocular tissue homogenates from these two species. The hydrolytic activities were measured using a generic esterase substrate (4-nitrophenyl acetate) and, in the absence of validated substrates for rabbit and pig enzymes, with selective substrates established for human CES1, CES2, and AADAC (d-luciferin methyl ester, fluorescein diacetate, procaine, and phenacetin). Kinetics and inhibition studies were conducted using these substrates and, again due to a lack of validated rabbit and pig CES inhibitors, with known inhibitors for the human enzymes. Protein expression levels were measured using quantitative targeted proteomics. Rabbit ocular tissues showed significant variability in the expression of CES1 (higher in cornea, lower in conjunctiva) and CES2 (higher in conjunctiva, lower in cornea) and a poor correlation of CES expression with hydrolytic activities. In contrast, pig tissues appear to express only CES1, and CES3 and AADAC seem to be either low or absent, respectively, in both species. The current study revealed remarkable species and tissue differences in ocular hydrolytic enzymes that can be taken into account in the design of esterase-dependent prodrugs and drug conjugates, the evaluation of ocular effects of systemic drugs, and in translational and toxicity studies.

Comparative study on pharmacokinetics and toxicity of intravitreal and sub-Tenon injection of triamcinolone acetonide in ocular tissues.

To compare the differences in kinetics, distribution, and toxicity of triamcinolone acetonide (TA) between the injection methods, sub-Tenon and intravitreal injections in rabbit ocular tissues. TA was injected into the vitreous or the sub-Tenon in rabbits. For pharmacokinetic study, rabbits were sacrificed periodically and then TA in blood and ocular tissues (retina/choroids, vitreous, and aqueous humor) were measured over 91d. For toxicological study, clinical signs, slit-lamp microscopic examination, ophthalmological test were performed. The eyeballs and surrounding tissues were collected and fixed with glutaraldehyde-formalin solution, and then paraffin embedded for histological investigation. Higher levels of TA were distributed in the intraocular tissues when injected into the vitreous compared to the sub-Tenon. Conversely, TA level was remarkably lower in the rabbits which received intravitreal TA injections than those treated with sub-Tenon injection throughout the study period in plasma. Optical discharge probably caused by systemic circulation of TA was observed by receiving sub-Tenon TA injection. Meanwhile, technic-associated toxicological ocular symptoms and findings were more frequently observed in intravitreal injection than in sub-Tenon injection. There are significant differences in kinetics and distribution of TA in vitreous body, aqueous humor and plasma, between the two injection methods. Although further study is needed to explain the species difference between human and rabbit, it is assumed that the difference in the frequency of intraocular pressure elevation and cataract formation by TA between the two injection methods are directly related to the TA concentrations in aqueous humor and vitreous body in each injection methods. Systemic toxicity and technic-associated toxicity are also closely related to kinetics of TA in plasma and each injection method itself, respectively.

A novel suprachoroidal microinvasive glaucoma implant: in vivo biocompatibility and biointegration

BackgroundA major challenge for any glaucoma implant is their ability to provide long-term intraocular pressure lowering efficacy. The formation of a low-permeability fibrous capsule around the device often leads to obstructed drainage channels, which may impair the drainage function of devices. These foreign body-related limitations point to the need to develop biologically inert biomaterials to improve performance in reaching long-term intraocular pressure reduction. The aim of this study was to evaluate in vivo (in rabbits) the ocular biocompatibility and tissue integration of a novel suprachoroidal microinvasive glaucoma implant, MINIject™ (iSTAR Medical, Wavre, Belgium).ResultsIn two rabbit studies, no biocompatibility issue was induced by the suprachoroidal, ab-externo implantation of the MINIject™ device. Clinical evaluation throughout the 6 post-operative months between the sham and test groups were similar, suggesting most reactions were related to the ab-externo surgical technique used for rabbits, rather than the implant material itself. Histological analysis of ocular tissues at post-operative months 1, 3 and 6 revealed that the implant was well-tolerated and induced only minimal fibroplasia and thus minimal encapsulation around the implant. The microporous structure of the device became rapidly colonized by cells, mostly by macrophages through cell migration, which do not, by their nature, impede the flow of aqueous humor through the device. Time-course analysis showed that once established, pore colonization was stable over time. No fibrosis nor dense connective tissue development were observed within any implant at any time point. The presence of pore colonization may be the process by which encapsulation around the implant is minimized, thus preserving the permeability of the surrounding tissues. No degradation nor structural changes of the implant occurred during the course of both studies.ConclusionsThe novel MINIject™ microinvasive glaucoma implant was well-tolerated in ocular tissues of rabbits, with observance of biointegration, and no biocompatibility issues. Minimal fibrous encapsulation and stable cellular pore colonization provided evidence of preserved drainage properties over time, suggesting that the implant may produce a long-term ability to enhance aqueous outflow.

Incurred sample reanalysis of cefuroxime in rabbit ocular tissues-A case study.

In this paper, we present the incurred sample reanalysis (ISR) data for cefuroxime in various ocular tissues of rabbits. Based on the cefuroxime concentration vs. time profile in various ocular tissues, three chosen time points enabled ISR assessment. Cefuroxime was quantitated in the ocular tissues using a published liquid chromatography-electrospray ionization/tandem mass spectrometry method operated under the multiple reaction-monitoring mode in positive ion mode. Regardless of the ocular tissue, the linearity range was 12.7-2760 ng/ml with a correlation coefficient (r2 ) of ≥0.996. All of the ISR samples representing various ocular tissues met the acceptance criteria. To the best of our knowledge, this is the first report showing the ISR of ocular tissues in any species.

Characterization and tissue localization of ocular carboxylesterases in multiple species

AbstractIntroductionOcular ADME (absorption, distribution, metabolism &amp; elimination) is gaining interest due to rapidly growing incidence of eye diseases and the need of new therapeutic applications. The information regarding drug‐metabolising enzymes (DME) in different ocular tissues is scattered. In addition, animal models may not be predictive of the human situation due to species differences.Aimof the studyWe sought to obtain quantitative and functional data of ocular carboxylesterases in multiple species to help comparisons between preclinical animal models and humans.Methods Esterase activity assays were optimized using established human‐specific substrates. Quantitative proteomics was done using specific marker peptides to quantify the selected esterases in ocular tissues of different species (porcine, rabbit and human). Finally, the functional and quantification data was compared between tissues and species. Main findingsThe rabbit ocular tissues had about 3‐fold higher hydrolysis rates for the generic esterase substrate, p‐nitrophenyl acetate, than the pig tissues. Carboxylesterase 1‐mediated activity (D‐luciferin methyl ester as substrate) in pig ocular tissues were low as compared to rabbit. Moreover, carboxylesterase 2‐mediated activities (fluorescein diacetate as substrate) varied over a 5‐ to 20‐fold range among seven ocular tissues (conjunctiva, cornea, aqueous humor, vitreous, retina, RPE and choroid) in the pig and rabbit, respectively. On average, corneal and choroidal tissues showed high expression and functional activity of carboxylesterase isoforms while retinal pigment epithelium and aqueous humor showed the least activity. More detailed enzymatic and proteomic data will be presented at the poster.ConclusionThis study provides new insight in ocular esterase expression and esterase‐mediated drug metabolism in multiple species.

Ophthalmic Drug Delivery Using Iontophoresis: Recent Clinical Applications.

Background: Iontophoresis is a noninvasive delivery system designed to overcome barriers to ocular penetration of topical ophthalmic medications by employing a low-amplitude electrical current to promote the migration of a charged drug substance across biological membranes. Trans-scleral iontophoresis of dexamethasone phosphate has demonstrated dramatically increased intraocular concentrations of dexamethasone in rabbit ocular tissues compared with topical instillation, including 50- to 100-fold greater aqueous humor concentrations. Methods: This article reviews available data on recent clinical applications of iontophoretic ophthalmic drug delivery. Results: The EyeGate II delivery system (EGDS) is a trans-scleral iontophoresis system that has been used in conjunction with EGP-437, a proprietary-charged formulation of dexamethasone phosphate for iontophoretic delivery. In patients with noninfectious anterior uveitis, EGP-437, delivered through 2 iontophoretic treatments using the EGDS, demonstrated similar efficacy to topical prednisolone acetate 1% eye drops instilled 8 times daily over 28 days, suggesting the potential to decrease or eliminate the need for daily dosing of topical steroids in this patient population. Other applications for EGP-437 delivered through the EGDS that have been explored in clinical trials include treatment of dry eye, postsurgical inflammation and pain, and scleritis. In addition, transcorneal iontophoresis has been used outside of the United States to enhance riboflavin penetration in patients undergoing corneal cross-linking as therapy for progressive keratoconus. Conclusions: The reviewed studies demonstrate the feasibility of using iontophoresis to enhance drug delivery to ocular tissues and support the potential of this noninvasive technique across a range of ophthalmic indications.

Historical Data: Histopathology Lesions Observed in the Eyes of Control Rabbits in Topical Ocular Administration and Contact Lens Studies.

Information on background changes in the ocular tissues of rabbits ( Oryctolagus cuniculus), a common species employed in ophthalmic toxicology studies, is sparse. This complicates interpretation of changes in light of small sample sizes on any single study. The purpose of this publication is to document the interstudy incidence of spontaneous or iatrogenic changes occurring in eyes of control rabbits. Photomicrographs of select lesions are provided. The data set was derived from a total of 54 studies conducted over an eleven-year period at Alcon Research Ltd., a Novartis Division, which featured topical ocular and contact lens routes of administration. It includes a total of 1,222 pigmented and albino New Zealand rabbits and a total of 2,084 eyes which were either untreated or treated with innocuous control articles. There were no noteworthy differences across routes of administration. Changes in anterior segment ocular and adnexal tissues were more common than in posterior segment ocular tissues. Overall, mononuclear cell infiltration was the most common finding. The retina was the posterior tissue most commonly observed with spontaneous changes, with folds and rosettes being the most common retinal finding. Retinal changes were more common in albino as compared to pigmented rabbits. Understanding the incidence and characteristics of spontaneous ocular lesions facilitates accurate and consistent diagnosis and data interpretation.

Esterase activity in porcine and albino rabbit ocular tissues

Corneal esterases are utilized in the activation of topically applied ester prodrugs. Esterases may also be involved in the metabolism of drugs in posterior eye tissues, but their physiological activity is unknown. Furthermore, extrapolation of the esterase activity from protein level to the tissues is missing.The aims of the current study were to determine esterase activities in porcine and albino rabbit ocular tissues, calculate the activities for whole tissues and compare esterase activity between the species. We conducted a hydrolysis study with ocular tissue homogenates using an esterase probe substrate 4-nitrophenyl acetate. The hydrolysis rates were first normalized to protein content and then scaled to whole tissues.The hydrolytic rate normalized to protein content was high in the cornea and iris-ciliary body and low in the lens and aqueous humor, and in general, the rabbit tissues had higher hydrolytic rates than the porcine ones. Esterase activity scaled to whole tissue was high in cornea and iris-ciliary body and low in aqueous humor and retinal pigment epithelium in both species.The current study revealed differences in esterase activities among the ocular tissues and the species. This basic knowledge on ocular esterases provides background information particularly for posterior segment drug development.

Ocular Distribution and Pharmacokinetics of Lifitegrast in Pigmented Rabbits and Mass Balance in Beagle Dogs.

Purpose: Lifitegrast is approved in the United States for the treatment of dry eye disease (DED). We assessed lifitegrast's ocular distribution/pharmacokinetic profile in rabbits, and 14C-lifitegrast mass balance/excretion in dogs.Methods: Female pigmented rabbits received a single topical ocular dose of lifitegrast (Formulation No. 1, n = 25; No. 2, n = 25) per eye twice daily (target, 1.75 mg/eye/dose). Blood/ocular tissues were collected on day 5. Beagle dogs received single intravenous (n = 10; target, 3 mg, 262 μCi/animal) and ocular (n = 8, target, 3 mg, 30 μCi/eye) doses of 14C-lifitegrast (∼8 weeks between doses). Blood, excreta, and cage rinse/wipes were collected. Concentrations were measured by mass spectrometry/liquid scintillation counting. Pharmacokinetic analyses (noncompartmental) included maximum concentration (Cmax), time to Cmax (tmax), and area under the concentration-time curve from 0 to 8 h (AUC0–8).Results: In rabbits, lifitegrast Cmax and AUC0–8 were similar between formulations. Cmax was highest in ocular anterior segment tissues: 5,190–14,200 ng/g [conjunctiva (palpebral/bulbar), cornea, anterior sclera]. Posterior segment tissues had lower concentrations (0–826 ng/g). AUC0–8 followed a similar trend. Plasma concentrations were low (Cmax <18 ng/mL). Tissue/plasma tmax was ∼0.25–1 h. In dogs, after intravenous/ocular doses, 14C-lifitegrast was eliminated primarily through feces. Excreted radioactivity was mainly unchanged lifitegrast.Conclusions: High exposure of lifitegrast in rabbit ocular anterior segment tissues and low exposure in posterior segment tissues/plasma suggests that lifitegrast reaches target tissues for DED treatment, with low potential for off-target systemic/ocular effects. Excretion of unchanged 14C-lifitegrast suggests minimal drug metabolism in vivo. This is consistent with lifitegrast clinical trial efficacy/safety data.

A sensitive UPLC-APCI-MS/MS method for the determination of dexamethasone and its application in an ocular tissue distribution study in rabbits following topical administration

A sensitive, specific and rapid ultra-performance liquid chromatography tandem mass spectrometry method has been developed and validated for the determination of dexamethasone (DEX) in ocular tissues of rabbits.

Targeted delivery of hyaluronic acid to the ocular surface by a polymer-peptide conjugate system for dry eye disease

Eye drop formulations containing HA are widely used to lubricate the ocular surface and relieve dry eye related symptoms, however its low residence time remains a challenge. We designed a polymer-peptide system for the targeted delivery of HA to the ocular surface using sialic acid or type I collagen as anchors for HA immobilization. The addition of the polymer-peptide system to HA eye drop exhibited a reduced friction coefficient, and it can keep the ocular surface enriched with HA for prolonged time. This system can be further developed into a more effective treatment for dry eye than a standard HA eye drop.

Topical delivery of aqueous micellar resolvin E1 analog (RX-10045)

PurposeThe primary objective of this study were to optimize aqueous micellar solution of isopropyl ester prodrug of resolvin (RX-10045), study in vivo ocular compatibility and tissue distribution following topical administration. MethodsAn optimized ratio of hydrogenated castor-oil and octoxynol-40 (1.0:0.05wt%) was prepared to entrap RX-10045 in the hydrophobic core of micelles. RX-10045 aqueous micelles were subjected to characterization. In vitro stability studies were performed at 4°C, 25°C and 40°C. In vivo studies were conducted in New Zealand albino rabbits following topical drop administration. ResultsAqueous RX-10045 micellar solutions were successfully prepared. Micelles had a mean diameter of ∼12nm with low negative surface charge. RX-10045 demonstrated high stability in citrate buffer (0.01M) at 40°C. Hackett–McDonald ocular irritation scores were extremely low comparable to negative control. No significant difference in intraocular pressure was noted. Electroretinography studies did not reveal any retinal damage after multiple dosing of RX-10045 micellar solution. Ocular tissue distribution studies demonstrated appreciable drug concentrations in anterior ocular tissues. Moreover, RX-10008 (active metabolite of RX-10045) was detected in retina/choroid upon topical drop instillation. ConclusionsA clear, stable, aqueous 0.1% RX-10045 micellar formulation was successfully prepared. Micellar solution was well-tolerated and did not have any measurable tissue damage in rabbit ocular tissues. Micelles appear to follow conjunctival/scleral pathway to reach back-of-the-eye tissue (retina). Topical aqueous formulations may be employed to treat posterior ocular diseases. Such micellar topical formulations may be more patient acceptable over invasive routes of administrations such as intravitreal injection/implants.