A triple crosslinked micelle-hydrogel lacrimal implant for localized and prolonged therapy of glaucoma.

Glaucoma is a chronic disease that requires lifelong treatment, whereas, discomfort caused by frequent medication may affect the quality of life. Moreover, the therapeutic efficacy of traditional local administration was unsatisfactory due to the rapid ocular clearance mechanism and the ocular barrier. Herein, a triple crosslinked micelle-hydrogel lacrimal implant with low polymer content was fabricated for localized and prolonged therapy of glaucoma. Latanoprost and timolol were simultaneously entrapped in the PEG-PLA micelles with high encapsulation efficiency and further loaded into the triple crosslinked hydrogel, facilitating a double sustained release of drugs. Subsequently, the implant was constructed by a unique molecular orientation fixation technology, which enables the implant to be fixed in the lacrimal duct. The triple crosslinked micelle-hydrogel lacrimal implant manifested a distinguished physicochemical characterization to sustain the release of latanoprost and timolol. In vitro release experiment demonstrated the duration of two drugs was extended for up to 28 days. The in vivo test of elevated intraocular pressure (IOP) in a rabbit model revealed that the IOP-lowering effects were sustained longer than 28 days as expected. The relative pharmacological availability (PA) of lacrimal implants was 5.7 times greater than that of the eye drops. The results of the studies on ocular irritation and histological examination demonstrated the good safety of the lacrimal implant. In conclusion, the triple crosslinked micelle-hydrogel lacrimal implant could effectively lower the IOP with splendid compatibility, demonstrating the promising prospect in the long-term noninvasive treatment of glaucoma.

Technical brief: Direct, real-time electrochemical measurement of nitric oxide in ex vivo cultured human corneoscleral segments.

Chronic elevation of intraocular pressure (IOP) is a major risk factor associated with primary open angle glaucoma (POAG), a common form of progressive optic neuropathy that can lead to debilitating loss of vision. Recent studies have identified the role of nitric oxide (NO) in the regulation of IOP, and as a result, several therapeutic ventures are currently targeting enhancement of NO signaling in the eye. Although a low level of NO is important for ocular physiology, excess exogenous NO can be detrimental. Therefore, the ability to directly measure NO in real time is essential for determining the role of NO signaling in glaucomatous pathophysiology. Historically, NO activity in human tissues has been determined by indirect methods that measure levels of NO metabolites (nitrate/nitrite) or downstream components of the NO signaling pathway (cGMP). In this proof-of-concept work, we assess the feasibility of direct, real-time measurement of NO in ex vivo cultured human corneoscleral segments using electrochemistry. A NO-selective electrode (ISO-NOPF200) paired to a free radical analyzer (TBR1025) was placed on the trabecular meshwork (TM) rim for real-time measurement of NO released from cells. Exogenous NO produced within cells was measured after treatment of corneoscleral segments with esterase-dependent NO-donor O2-acetoxymethylated diazeniumdiolate (DETA-NONOate/AM; 20 µM) and latanoprostene bunod (5-20 µM). A fluorescent NO-binding dye DAF-FM (4-Amino-5-methylamino- 2',7'-difluorofluorescein diacetate) was used for validation. A linear relationship was observed between the electric currents measured by the NO-sensing electrode and the NO standard concentrations, establishing a robust calibration curve. Treatment of ex vivo cultured human donor corneoscleral segments with DETA-NONOate/AM and latanoprostene bunod led to a significant increase in NO production compared with vehicle-treated controls, as detected electrochemically. Furthermore, the DAF-FM fluorescence intensity was higher in outflow pathway tissues of corneoscleral segments treated with DETA-NONOate/AM and latanoprostene bunod compared with vehicle-treated controls. In conclusion, these results demonstrate that NO-sensing electrodes can be used to directly measure NO levels in real time from the tissues of the outflow pathway.

Prostaglandin FP receptor antagonists: discovery, pharmacological characterization and therapeutic utility.

In contrast to the availability of potent and selective antagonists of several prostaglandin receptor types (including DP1 , DP2 , EP and TP receptors), there has been a paucity of well-characterized, selective FP receptor antagonists. The earliest ones included dimethyl amide and dimethyl amine derivatives of PGF2α , but these have failed to gain prominence. The fluorinated PGF2α analogues, AL-8810 and AL-3138, were subsequently discovered as competitive and non-competitive FP receptor antagonists respectively. Non-prostanoid structures, such as the thiazolidinone AS604872, the D-amino acid-based oligopeptide PDC31 and its peptidomimic analogue PDC113.824 came next, but the latter two are allosteric inhibitors of FP receptor signalling. AL-8810 has a sub-micromolar in vitro potency and ≥2 log unit selectivity against most other PG receptors when tested in several cell- and tissue-based functional assays. Additionally, AL-8810 has demonstrated therapeutic efficacy as an FP receptor antagonist in animal models of stroke, traumatic brain injury, multiple sclerosis, allodynia and endometriosis. Consequently, it appears that AL-8810 has become the FP receptor antagonist of choice. LINKED ARTICLES: This article is part of a themed section on Eicosanoids 35 years from the 1982 Nobel: where are we now? To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.8/issuetoc.

Treatment merits of Latanoprost/Thymoquinone - Encapsulated liposome for glaucomatus rabbits.

Elevation of the intraocular pressure (IOP) is recognized as a risk factor for glaucoma development. Latanoprost (LAT) is a prostaglandin analog used to reduce the (IOP). Thymoquinone (TQ) is a major bioactive ingredient of Nigella sativa. The aim of this study was to develop novel liposomal drug carriers for ocular delivery of LAT, TQ and a mixture of them to investigate their IOP lowering efficacy upon subconjunctival injection in glaucoma-induced rabbit's eye. The aim of the present work extends also to study the effect of the different liposome formulations on the aqueous humor oxidative stress. Liposome samples were prepared using thin film hydration method. The physiochemical properties of the prepared drugs were characterized. The IOP was recorded for 70 rabbits using Schiotz-tonometer. Malondialdehyde (MDA), reduced glutathione (GSH), catalase (CAT) activities and total antioxidant activity of the aqueous humor were estimated. Fourier transform infrared and differential scanning calorimetric studies confirmed the interaction between the drug and the vesicles, which resulted in high drug encapsulation efficiency ≥88%. The size of the prepared liposomes was less than 10 µm which make them suitable in ophthalmic applications. The sustained effect was achieved by liposome samples of Lip (LAT) and Lip (LAT + TQ) which were able to reduce the IOP significantly up to 84 h. Morever, the treatment of glaucomatous rabbits with liposome formulations containing TQ in their preparation [Lip (TQ) and Lip (LAT + TQ)] greatly improved the ocular tissue-induced histopathological lesions. None of the prepared liposome formulations succeeded to improve the glaucoma-induced oxidative stress damage.

The physical properties of generic latanoprost ophthalmic solutions are not identical.

PURPOSE: To compare various characteristics of Xalatan® and five generic latanoprost ophthalmic solutions. METHODS: Drop size, volume, pH values, buffer capacity, viscosity, hardness of bottles and costs were determined. Drop sizes were measured in triplicates by micropipettes, and the number of drops counted in three separate bottles of each generic product was determined. pH values were measured in triplicates by a calibrated pH meter. Buffer capacity was exploited by titrating known quantities of strong base into 2.5 ml of each brand and interpolated to neutral pH. Kinematic viscosity was determined by linear regression of timed gravity flow from a vertical syringe through a 21-G cannula. The hardness of the bottles was evaluated by gradually increasing tension on a hook placed around each bottle until a drop was expelled reading the tension on an attached spring scale. RESULTS: Drop sizes and the number of drops in the bottles varied significantly between the generic drugs. The control value of pH in the brand version (Xalatan® ) was markedly lower compared to the generic latanoprost products. Titration of Xalatan® to neutrality required substantially more NaOH compared to the generic latanoprost products. Finally, the viscosity revealed a significant variability between brands. Remarkable differences were found in bottle shapes, bottle hardness and costs of the latanoprost generics. CONCLUSION: Generic latanoprost eye drops should not be considered identical to the original brand version as regards to drop size, volumes, pH values, buffer capacity, viscosity, hardness of bottles and costs. It is likely that these issues affect compliance and intraocular pressure (IOP)-lowering effect. Therefore, re-evaluation of the requirements for introducing generic eye drops seems reasonable.

Evaluation of physical properties and dose equivalency of generic versus branded latanoprost formulations.

The purpose of this study was to comparatively evaluate the pharmaceutical characteristics of various marketed generic formulations of prostaglandin analogue latanoprost in the Indian market. Three generics of latanoprost and one branded (Xalatan) formulation (five vials each) were obtained from authorized agents from the respective commercial sourcing having the same batch number. These formulations were coded, and the labels were removed. At a standardized room temperature of 25 °C, the concentration, osmolarity, drop size, pH, and total drops per vial were determined for Xalatan and all the generics of latanoprost. The concentration of various brands varied between 50.49 ± 0.36 and 58.90 ± 0.52 µg/ml as compared to the standard labeled concentration of 50 µg/ml on the latanoprost vials. The concentration of drugs in individual drop varied from 1.30 ± 0.05 to 1.78 ± 0.04 µg/drop. The volume of drug formulation per bottle varied from 2.4 ± 0.12 to 2.6 ± 0.09 ml/bottle. The number of drops per bottle varied from minimum of 88.60 ± 0.10 drops to maximum of 102.0 ± 4.3 drops across all the formulations, while the drop size varied from 25.72 ± 2.70 to 29.97 ± 1.38 µl. The osmolarity of 2/4 drugs was within 300 mOs M (±10 %). The specific gravity varied between 0.98 ± 0.01 and 1.007 ± 0.01, while pH was between 7.05 ± 0.004 and 7.13 ± 0.005. Two of the generic brands were outside the United States pharmacopoeia limits (±10%) for ophthalmic formulation, with concentration exceeding the limits by 3 % (p = 0.151) and 8 % (p = 0.008), respectively. This pilot study highlights that there are significant variations in the drug concentrations and physical properties of generic latanoprost formulations. Although none of the brands had concentrations below the recommended level, two of the brands had concentrations exceeding the limits by 3 and 8 %, respectively.

Impact of prostaglandin glaucoma drops on platelet-activating factor action: an in vitro study.

AIM: The aim of this study was to investigate the effect of different prostaglandin analogs on platelet-activating factor (PAF) levels. METHODS: Three prostaglandin analogs were selected: bimatoprost 0.3 mg/mL, latanoprost 50 µg/mL, and tafluprost 15 µg/mL. Each drug sample was tested for its ability to cause platelet aggregation, which was measured as PAF-induced aggregation, before and after the addition of various concentrations of the examined sample, creating a linear curve of percentage inhibition (ranging from 0% to 100%) versus different concentrations of the sample. The concentration of the sample that inhibited 50% PAF-induced aggregation was calculated based on this curve, and this value was defined as IC50. In addition, the effect of eye drops on PAF metabolism was examined, through an in vitro analysis on PAF basic metabolic enzymes (PAF-cholinephosphotransferase, PAF-acetyl-CoA:1-O-alkyl-sn-glycero-3-phosphocholine acetyltransferase, and PAF-acetylhydrolase). RESULTS: The IC50 values for Lumigan UD® (bimatoprost 0.3 mg/mL), Monoprost® (latanoprost 50 µg/mL), and Saflutan (tafluprost 15 µg/mL) were 8.7, 0.28, and 1.4 µg/mL, respectively. DISCUSSION: All three prostaglandin analogs suspended PAF, but bimatoprost induced the most potent inhibition, compared to tafluprost and to the weak effect of latanoprost.

New prostaglandin analog formulation for glaucoma treatment containing cyclodextrins for improved stability, solubility and ocular tolerance.

Latanoprost is a practically insoluble prostaglandin F2α analog considered a first-line agent for glaucoma treatment. From a pharmaceutical point of view, latanoprost is challenging to be formulated as an eye drop due to its poor water solubility and the presence of an ester bond that needs to be cleaved in vivo but maintained unchanged during storage. Cyclodextrins (CDs) are known to form complexes with hydrophobic drugs, influencing their stability, availability, solubility, and tolerance in a non-predictable manner. A variety of CDs including native α, ß, and γCDs as well as substituted hydroxypropylßCD, hydroxypropylγCD, dimethylßCD, sulphatedßCD, and propylaminoßCD were screened and the most appropriate CD for the formulation of latanoprost for an ocular topical application was selected. Among the tested CDs, propylaminoßCD had the best trade-off between latanoprost stability and availability, which was confirmed by its complex constant value of 3129M(-1). Phase-solubility and NMR investigations demonstrated that the propylaminoßCD effectively formed a complex involving the ester group of latanoprost providing protection to its ester bond, while ensuring proper latanoprost solubilization. Furthermore, in vivo experiments demonstrated that the latanoprost-propylaminoßCD formulation led to lower ocular irritation than the commercial latanoprost formulation used as a reference. The latanoprost-propylaminoßCD formulation was demonstrated to successfully address the main stability, solubility, and tolerance limitations of topical ocular latanoprost therapy for glaucoma.

Synthesis of prostaglandin analogues, latanoprost and bimatoprost, using organocatalysis via a key bicyclic enal intermediate.

Two antiglaucoma drugs, bimatoprost and latanoprost, which are analogues of the prostaglandin, PGF2α, have been synthesized in just 7 and 8 steps, respectively. The syntheses employ an organocatalytic aldol reaction that converts succinaldehyde into a key bicyclic enal intermediate, which is primed for attachment of the required lower and upper side chains. By utilizing the crystalline lactone, the drug molecules were prepared in >99% ee.

[Original preparations versus generics--latanoprost: how similar is different?]. / Originalpräparat versus Generika--Latanoprost : Wie gleich ist unterschiedlich?

BACKGROUND: To test the interchangeability of the commercially available (in Germany) latanoprost drugs and their generics respectively, the concentration of the active substance was tested. Guidelines of the European Medicines Agency postulate a sufficient bioequivalence, if the range of the agent is within 80-125% of the original drug. METHODS: All compounds of latanoprost were procured registered. The concentration of latanoprost and benzalkoniumchloride was measured by high-performance liquid chromatography (HPLC) in a validated reference labroratory for 23 generics. In addition, the mean volume of drops and the pH of the formulation were measured. The packaging label and the readability of the enclosed information leaflet were checked. RESULTS: All products contained less than 50 µg/ml latanoprost. The deviating reduction of the active substance (mean: - 7.39%, ± 2.8%) was accompanied by fluctions of the eyedrops' mass (mean: 0.03 g, ± 0.002 g). The concentration of benzalkonium chloride was mostly increased (median: 5.45%, min: - 2.5%, max: 11.5%). The pH of the original drug and the generics (median 6.78, min: 6.62, max: 6.81) was similar to the original drug, but was significantly different from an unpreserved formulation (pH 7.18). Due to type size, the packaging leaflet was illegible for humans with impaired vision. CONCLUSIONS: Before prescribing generics in ophthalmology, different factors have to be considered, which might influence the amount of IOP lowering in effect. In the absence of healthcare research it is still unclear, how different bottle forms of eyedrops--such as appearance (e.g. Cyrillic characters) or pressure point (administration)--reduce the adherence of glaucoma patients.

Stability of latanoprost in generic formulations using controlled degradation and patient usage simulation studies.

PURPOSE: To evaluate the stability of latanoprost in generic formulations by using controlled degradation and patient usage simulation studies METHODS: Standard latanoprost was subjected to controlled degradation studies. Latanoprost content was assessed by using MRM, and generated Degradation Products (DP) were analysed by using the Information Dependent Acquisition (IDA) protocol of positive ESI-LC-MS/MS. Latanoprost content and formation of DP were assessed in generic formulations and were compared with Xalatan(®) in a controlled patient usage simulation studies. The last few drops of latanoprost, present in containers used by patients were also evaluated. RESULTS: Extreme pH conditions, oxidation, light and heat were found to be the significant factors for high degree of latanoprost degradation. Systematic analysis of 7 selected generics revealed that the latanoprost content varied from 90-330%. Concentration of the latanoprost in Xalatan was found to be 97% of the label claim. Degradation studies showed the formation of 3 novel and 3 already known impurities. Upon simulated patient usage, 2 of the generic formulations showed significant degradation of latanoprost. Generic formulations having thermally sealed gas tight packing showed good stability during patient usage. Overage of latanoprost was observed in generics with other than thermal sealing. Latanoprost bottles used by patients showed concentrations ranging from 20 to 250% of label claim (144% median). CONCLUSION: This study revealed the presence of overage of latanoprost in some generic formulations and formation of degradation products. Packaging with gas tight containers may be one of the important factors for latanoprost stability, along with its storage at low temperature during patient usage.

Extended latanoprost release from commercial contact lenses: in vitro studies using corneal models.

In this study, we compared, for the first time, the release of a 432 kDa prostaglandin F2a analogue drug, Latanoprost, from commercially available contact lenses using in vitro models with corneal epithelial cells. Conventional polyHEMA-based and silicone hydrogel soft contact lenses were soaked in drug solution (131 µg = ml solution in phosphate buffered saline). The drug release from the contact lens material and its diffusion through three in vitro models was studied. The three in vitro models consisted of a polyethylene terephthalate (PET) membrane without corneal epithelial cells, a PET membrane with a monolayer of human corneal epithelial cells (HCEC), and a PET membrane with stratified HCEC. In the cell-based in vitro corneal epithelium models, a zero order release was obtained with the silicone hydrogel materials (linear for the duration of the experiment) whereby, after 48 hours, between 4 to 6 µg of latanoprost (an amount well within the range of the prescribed daily dose for glaucoma patients) was released. In the absence of cells, a significantly lower amount of drug, between 0.3 to 0.5 µg, was released, (p <0:001). The difference observed in release from the hydrogel lens materials in the presence and absence of cells emphasizes the importance of using an in vitro corneal model that is more representative of the physiological conditions in the eye to more adequately characterize ophthalmic drug delivery materials. Our results demonstrate how in vitro models with corneal epithelial cells may allow better prediction of in vivo release. It also highlights the potential of drug-soaked silicone hydrogel contact lens materials for drug delivery purposes.

Sustained delivery of latanoprost by thermosensitive chitosan-gelatin-based hydrogel for controlling ocular hypertension.

Glaucoma is an irreversible ocular disease that may lead to progressive visual field loss and eventually to blindness with inadequately controlled intraocular pressure (IOP). Latanoprost is one of the most potent ocular hypotensive compounds, the current first-line therapy in glaucoma. However, the daily instillation required for efficacy and undesirable side-effects are major causes of treatment adherence failure and persistence in glaucoma therapy. In the present study, we developed an injectable thermosensitive chitosan/gelatin/glycerol phosphate (C/G/GP) hydrogel as a sustained-release system of latanoprost for glaucoma treatment. The latanoprost-loaded C/G/GP hydrogel can gel within 1min at 37°C. The results show a sustained release of latanoprost from C/G/GP hydrogel in vitro and in vivo. The latanoprost-loaded C/G/GP hydrogel showed a good in vitro and in vivo biocompatibility. A rabbit model of glaucoma was established by intravitreal injection of triamcinolone acetonide. After a single subconjunctival injection of latanoprost-loaded C/G/GP hydrogel, IOP was significantly decreased within 8days and then remained at a normal level. The results of the study suggest that latanoprost-loaded C/G/GP hydrogel may have a potential application in glaucoma therapy.

Biodegradable nanoparticles for controlled subconjunctival delivery of latanoprost acid: in vitro and in vivo evaluation. Preliminary results.

Purpose of the study was to develop and assess a novel controlled drug delivery system of latanoprost acid (LA). Poly(lactide)/Monomethoxy-poly(ethyleneglycol) (PLA-PEG) nanoparticles (NPs) were prepared using an emulsification-solvent evaporation technique. NPs were characterized in vitro according to their size, ζ-potential, drug entrapment efficiency and LA release. LA-loaded NPs (equivalent to 8.5 µg LA) were administered into the subconjunctival space of normotensive rabbits (group A). A free LA solution of the same drug content was subconjunctivally injected in a second rabbit group (group B), while blank NPs were administered in a third group (group C). A group of untreated animals (group D) served as the control. Intraocular pressure (IOP) was monitored for 8 consecutive days, using the Tono-pen XL. Aqueous humor (AH) levels of LA were evaluated for 6 days post-administration, by means of HPLC. Mean nanoparticle size was 80 nm. The drug entrapment efficiency was 18.3%. NPs sustained the release of LA over several days in vitro. Non-significant differences in baseline IOP were found between groups (p = 0.22). LA-loaded NPs exerted a significant hypotensive effect on group A, while IOP values remained significantly lower compared to the rest groups, throughout the study (p = 0.04). LA AH concentrations in group B continuously decreased with time, while LA levels in group A steadily increased. On day 6, LA levels were higher in group A compared to group B (344 ± 73.5 ng/ml and 228 ± 41.01 ng/ml, respectively). No adverse effects were observed. In conclusion, after subconjunctival administration, the LA-loaded NPs provided sustained LA delivery in vivo. They appear to be a promising system for the controlled subconjunctival delivery of LA.

A multivalent approach towards linked dual-pharmacology prostaglandin F receptor agonist/carbonic anhydrase-II inhibitors for the treatment of glaucoma.

Lowering of intra-ocular pressure is the primary pharmacologic approach for the treatment of glaucoma and a number of distinct mechanisms of action have been clinically validated. Targeting of multiple mechanisms in combination therapies has proven effective both clinically and commercially although potential improvements with regards to efficacy, tolerability and dosing frequency remain. Application of Theravance's multivalent approach to drug discovery towards linked dual-pharmacology prostaglandin F receptor (FP) agonist/carbonic anhydrase (CA)-II inhibitor compounds is described. Compound 29 exhibits weak potency (pEC(50)=5.7, IA>1.0) as an FP agonist with high binding affinity (pK(i)=8.1) to the CA-II enzyme, and has comparable corneal permeability to the CA-II inhibitor dorzolamide.

Stereocontrolled organocatalytic synthesis of prostaglandin PGF2α in seven steps.

Prostaglandins are hormone-like chemical messengers that regulate a broad range of physiological activities, including blood circulation, digestion and reproduction. Their biological activities and their complex molecular architectures have made prostaglandins popular targets for synthetic organic chemists for over 40 years. Prostaglandin analogues are widely used as pharmaceuticals and some, such as latanoprost, which is used to treat glaucoma, have become billion-dollar drugs. Previously reported syntheses of these compounds are quite lengthy, and every chemical step costs time and energy, generates waste and is accompanied by material losses. Using a new bond disconnection, here we report a concise synthesis of the most complex prostaglandin, PGF2α, with high levels of control of relative and absolute stereochemistry, and fewer steps. The key step is an aldol cascade reaction of succinaldehyde using proline organocatalysis to create a bicyclic enal in one step and an enantiomeric excess of 98%. This intermediate bicyclic enal is fully primed with the appropriate functionality for attachment of the remaining groups. Access to this bicyclic enal will not only render existing prostaglandin-based drugs more affordable, but will also facilitate the rapid exploration of related chemical structures around the ubiquitous five-membered ring motif, such as potentially therapeutic prostaglandin analogues.

Investigation of surfactants suitable for stabilizing of latanoprost.

The content of latanoprost, a therapeutic drug for glaucoma, is likely to decrease in solution. In a previous study, we confirmed that this was associated with latanoprost adsorption to the container and hydrolysis of latanoprost, and established a formulation of latanoprost eye drop solution that can be stored at room temperature. In addition, we clarified that a surfactant added to the formulation stabilized latanoprost by forming complex micelle. In this study, we investigated the influence of structure of surfactants in the stabilization of latanoprost. Non-ionic surfactants involving the polyoxyethylene chain (additive mol number: 20 or more) and the straight-chain alkyl group, with an HLB of 15.0 or more, markedly inhibited a decrease in the latanoprost content and its degradation. These surfactants are soluble in the formulation of eye drop solution, leading to micelle formation even at a low concentration, and they are not influenced by the temperature. Therefore, they may exhibit marked stabilization effects. In addition, there was no influence on the stability of latanoprost when adding benzalkonium chloride, as a preservative, to a formulation involving these surfactants.