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Introduction: Glaucoma is a chronic disease that requires long-term adherence to treatment. Topical application of conventional eye drops results in substantial drug loss due to rapid tear turnover, with poor drug bioavailability being a major challenge for efficient glaucoma treatment. We aimed to prepare the anti-glaucoma drug betaxolol hydrochloride (BH) as a novel nano-delivery system that prolonged the retention time at the ocular surface and improved bioavailability., Methods: We constructed multifunctional nanoparticles (MMt-BH-HA/CS-ED NPs) by ion cross-linking-solvent evaporation method. The particle size, zeta potential, encapsulation efficiency and drug loading of MMt-BH-HA/CS-ED NPs were physicochemically characterized. The structure of the preparations was characterized by microscopic techniques of SEM, TEM, XPS, XRD, FTIR and TGA, and evaluated for their in vitro release performance as well as adhesion properties. Its safety was investigated using irritation assays of hemolysis experiment, Draize test and histopathology examination. Precorneal retention was examined by in vivo fluorescence tracer method and pharmacokinetics in tear fluid was studied. A model of high IOP successfully induced by injection of compound carbomer solution was used to assess the IOP-lowering efficacy of the formulation, and it was proposed that micro-interactions between the formulation and the tear film would be used to analyze the behavior at the ocular surface., Results: The positively charged MMt-BH-HA/CS-ED NPs were successfully prepared with good two-step release properties, higher viscosity, and slower pre-corneal diffusion rate along with longer precorneal retention time compared to BH solution. The micro-interactions between nanoparticles and tear film converted the drug clearance from being controlled by fast aqueous layer turnover to slow mucin layer turnover, resulting in higher drug concentration on the ocular surface, providing more durable and stable IOP-lowering efficacy., Conclusion: The novel multifunctional MMt-BH-HA/CS-ED NPs can effectively reduce IOP and are suitable for the treatment of chronic disease glaucoma., Competing Interests: The authors report no conflicts of interest in this work., (© 2022 Hu et al.)

Physiological reflexes and anatomical barriers render traditional eye drop delivery inefficient. We previously reported that drug-loaded nanoparticles and microspheres prepared from montmorillonite and Eudragit polymers exhibited good sustained-release and lowered intraocular pressure. Here, we compared the performance of optimized formulations to select the most suitable formulation for glaucoma therapy. We found that the microspheres had much higher encapsulation efficiency and drug loading than nanoparticles. Moreover, cytocompatibility experiments demonstrated that nanoparticles showed more severe cytotoxicity than microspheres, probably due to their smaller particles, enhanced cell uptake, and intracellular solubility. Interestingly, the pre-corneal retention time of nanoparticles reflected a clear advantage over microspheres, while the duration of the pharmacological effect of nanoparticles was not as good as that of microspheres: compared with the nanoparticle depressurization duration of only 8 h, the microspheres continuously depressurized for 12 h. The slower release of the microspheres and its micro-interaction mechanism with the discontinuous mucin layer of the tear film led to the inconsistency between duration of pharmacodynamics and fluorescence ocular retention time. In summary, the lower cytotoxicity and longer pharmacological effect of microspheres indicate their potential advantages for glaucoma applications., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Artificial vitreous humor holds immense potential for use in in vitro intravitreal drug delivery assays. In this study, we investigated rheological properties and drug or nanoparticle migration in hyaluronic acid (HA) - agar based hydrogels and compared these characteristics with bovine vitreous humor. Gel compositions identified in literature containing HA (0.7-5.0 mg/ml) and agar (0.95-4.0 mg/ml) were classified as either high (VH), medium (VM) or low (VL) polymer load. Viscoelastic behavior was evaluated using oscillatory rheology, and migration of differently sized and charged polystyrene nanoparticles (NPs) through the different gels was determined via multiple particle tracking. Comparable rheological behaviour was observed between VL and bovine vitreous. Tracking evaluations revealed that increasing particle size and gel viscosity slowed NP migration. Additionally, 100 nm anionic NPs migrated slower than neutral NPs in VL and VM, while cationic NPs were immobile in all gels. Finally, distribution and clearance of sodium fluorescein was used to model drug mobility through the gels using a custom-built eye model. Flow and angular movement only influenced drug migration in VL and VM, but not VH. Finally, VL and VM demonstrated to have the most similar sodium fluorescein clearance to that of bovine vitreous humor. Together, these evaluations demonstrate that low viscosity HA-agar gels can be used to approximate nanoparticle and drug migration through biological vitreous humor., Competing Interests: Declaration of Competing Interest The near neutral particle technology described in this publication is being developed by Kala Pharmaceuticals. J.H. declares a financial, a management/advisor, and a paid consulting relationship with Kala Pharmaceuticals. J.H. is a cofounder of Kala Pharmaceuticals and owns company stock, which is subject to certain restrictions under Johns Hopkins University policy. The terms of this arrangement are being managed by Johns Hopkins University in accordance with its conflict-of-interest policies., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Purpose: To evaluate the long-term effect on inflammation and inflammasome activation of intravitreally delivered connexin43 mimetic peptide (Cx43MP) in saline or incorporated within nanoparticles (NPs) for the treatment of the light-damaged rat eye., Methods: Light-induced damage to the retina was created by exposure of adult albino Sprague-Dawley rats to intense light for 24 hours. A single dose of Cx43MP, Cx43MP-NPs, or saline was injected intravitreally at 2 hours after onset of light damage. Fluorescein isothiocyanate (FITC)-labelled Cx43MP-NPs were intravitreally injected to confirm delivery into the retina. Electroretinogram (ERG) recordings were performed at 24 hours, 1 week, and 2 weeks post cessation of light damage. The retinal and choroidal layers were analyzed in vivo using optical coherence tomography (OCT) and immunohistochemistry was performed on harvested tissues using glial fibrillary acidic protein (GFAP), leukocyte common antigen (CD45), and Cx43 antibodies., Results: FITC was visualized 30 minutes after injection in the ganglion cell layer and in the choroid. Cx43MP and Cx43MP-NP treatments improved a-wave and b-wave function of the ERG compared with saline-injected eyes at 1 week and 2 weeks post treatment, and prevented photoreceptor loss by 2 weeks post treatment. Inflammation was also reduced and this was in parallel with downregulation of Cx43 expression., Conclusions: The slow release of Cx43MP incorporated into NPs is more effective at treating retinal injury than a single dose of native Cx43MP in solution by reducing inflammation and maintaining both retinal structure and function. This NP preparation has clinical relevance as it reduces possible ocular complications associated with repeated intravitreal injections.

Intravitreal mobility of nanocarriers may have implications on the efficacy of the encapsulated drug in the treatment of vitreo-retinal diseases, with any changes in the integrity of the vitreous microstructure influencing nanoparticle biodistribution. This study investigated enzymatically digested vitreous models to mimic the aging eye. Collagenase, hyaluronidase, or trypsin was employed to selectively digest the structural components of the vitreous. Physical properties of digested bovine vitreous were initially assessed via texture analysis and oscillatory shear testing. Morphological changes in bovine vitreous microstructure were visualized by scanning electron microscopy and diffusion dynamics of hyaluronic acid coated nanoparticles through degraded porcine and bovine vitreous were examined using fluorescence spectroscopy and multiple particle tracking microscopy, respectively. After enzymatic treatment, the vitreous liquefied and its dynamic mechanical properties significantly changed with a decrease in stiffness and an increase in damping capacity. Micrographs confirmed specific digestion of each of the structural vitreous components. Furthermore, enzymatic degradation reduced steric hindrance and enhanced convective flow within the vitreous, resulting in increased intravitreal nanoparticle mobility which could alter the drug pharmacokinetics., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Recent studies have shown that Connexin43 mimetic peptide (Cx43 MP) can prevent secondary damage following retinal ischaemic and inflammatory disorders by blocking uncontrolled Cx43 hemichannel opening. However, limitations in peptide stability and the presence of various intraocular barriers limit efficient retinal delivery in the clinical setting. The present study aimed to achieve targeted and sustained peptide delivery to the retina by encapsulating Cx43 MP into hyaluronic acid (HA) coated albumin nanoparticles (NPs). Intraocular biodistribution, particle retention, retinal targeting, and therapeutic efficacy of intravitreally injected NPs encapsulating Cx43 MP were evaluated in a rat model of retinal ischaemia-reperfusion injury. NPs rapidly diffused through the vitreous and specifically targeted CD44-expressing retinal cells. NPs remained at the target site for extended periods enabling sustained peptide release and thus prolonged therapeutic action. Compared to free Cx43 MP, Cx43 MP loaded NPs enabled enhanced therapeutic efficacy preventing thinning of retinal layers and disruption of retinal blood vessels. Immunohistochemical results confirm that Cx43 MP loaded NPs efficiently reduced Cx43 expression, thereby suppressing ongoing inflammation and preventing the loss of retinal ganglion cells. Overall, HA coated NPs could have great potential as a peptide delivery platform in the treatment of chronic retinal degenerative and inflammatory disorders., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Frequent intravitreal injections are currently used to overcome the ocular barriers and provide sufficient drug to the posterior eye segment. However, intravitreal injections have been associated with a number of complications and high treatment costs. To overcome these limitations, peptide-loaded poly(d,l-lactic-co-glycolic acid) nanoparticles (PLGA NPs) were developed using the nanoprecipitation technique and were optimized via Box-Behnken Design (BBD) and Response Surface Methodology (RSM). Developed NPs were evaluated for potential toxicity and cell apoptosis using the zebrafish embryo toxicity (ZET) model with titanium dioxide NPs and ethanol (1% v/v) serving as positive controls. Developed NPs had a size of 75.6-153.8 nm, a polydispersity index between 0.11 and 0.25 and a zeta potential of -9.4 to -46.0 mV. Loaded peptide was found to be stable under various experimental conditions tested. BBD and RSM were validated through the characterization of optimized formulations. Survival and hatching rates of NP-treated zebrafish 0-144 h post-fertilization were found to be normal with no significant malformations. Cellular apoptosis studies also endorsed the non-cytotoxic nature of the NPs. The overall results indicate that optimized PLGA nanoparticles could be a promising platform for efficient peptide delivery to the posterior segment of the eye.

Effective drug delivery to the retina still remains a challenge due to ocular elimination mechanisms and complex barriers that selectively limit the entry of drugs into the eye. To overcome these barriers, frequent intravitreal injections are currently used to achieve high drug concentrations in vitreous and retina. However, these repetitive injections may result in several side effects. Recent advancements in the field of nanoparticle-based drug delivery could overcome some of these unmet needs and various preclinical studies conducted to date have demonstrated promising results of nanotherapies in the treatment of retinal diseases. Compared to the majority of commercially available ocular implants, the biodegradable nature of most nanoparticles (NPs) avoids the need for surgical implantation and removal after the release of the payload. In addition, the sustained drug release from NPs over an extended period of time reduces the need for frequent intravitreal injections and the risk of associated side effects. The nanometer size and highly modifiable surface properties make NPs excellent candidates for targeted ocular drug delivery. Studies have shown that nanocarriers enhance the intravitreal half-life and thus bioavailability of a number of drugs including proteins and peptides. In addition, they have shown promising results in delivering genetic material to the retinal tissues by protecting it from possible intravitreal degradation. This review covers the various challenges associated with drug delivery to the posterior segment of the eye, particularly the retina, and highlights the application of nanocarriers to overcome these challenges in context with recent advances in preclinical studies. WIREs Nanomed Nanobiotechnol 2018, 10:e1473. doi: 10.1002/wnan.1473 This article is categorized under: Therapeutic Approaches and Drug Discovery > Emerging Technologies Implantable Materials and Surgical Technologies > Nanomaterials and Implants., (© 2017 Wiley Periodicals, Inc.)

Intravitreal injection is the most common administration route for the treatment of retinal diseases. However, the vitreous and some of the retinal layers themselves act as significant barriers to efficient delivery of drugs administered intravitreally. This study aimed to improve the diffusive mobility of nanoparticles (NPs) in the vitreous and enhance their permeation across the retina after intravitreal injection by application of ultrasound (US). Ex vivo posterior bovine eye cups were used and the vitreous was either left intact or removed gently from the neural retina. Hyaluronic acid coated human serum albumin NPs were administered into the eye cups and continuous US with a frequency of 1MHz, an intensity of 0.5W/cm 2 , and a duration of 30s was applied once or repeatedly via the transscleral route. After pre-determined time points, fluorescence intensities in the vitreous and the retina were analyzed. Short pulses of US significantly improved the diffusive mobility of NPs through the vitreous as well as their penetration across the neural retina into the retinal pigment epithelium and choroid without causing any detectable damage to the ocular tissues. Therefore, transscleral US could be a powerful and safe tool to enhance retinal delivery of intravitreally injected NPs., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Diseases affecting the posterior segment the eye, such as age-related macular degeneration (AMD), are the leading cause of blindness worldwide. Conventional dosage forms, such as eye drops, have to surmount several elimination mechanisms and complex barriers to achieve therapeutic concentrations at the target site often resulting in low anterior segment bioavailability (ca. 2-5%) with generally none of the drug reaching posterior segment tissues. Thus, frequent intravitreal injections are currently required to treat retinal conditions which have been associated with poor patient compliance due to pain, risk of infection, hemorrhages, retinal detachment and high treatment related costs. To partially overcome these issues, ocular implants have been developed for some posterior segment indications; however, the majority require surgical implantation and removal at the end of the intended treatment period. The transparent nature of the cornea and lens render light-responsive systems an attractive strategy for the management of diseases affecting the back of the eye. Light-responsive in situ forming injectable implants (ISFIs) offer various benefits such as ease of application in a minimally invasive manner and more site specific control over drug release. Moreover, the biodegradable nature of such implants avoids the need for surgical removal after release of the payload. Incorporating drug-loaded polymeric nanoparticles (NPs) into these implants may reduce the high initial burst release from the polymeric matrix and further sustain drug release thus avoiding the need for frequent injections as well as minimizing associated side effects. However, light-responsive systems for ophthalmic application are still in their early stages of development with limited reports on their safety and effectiveness. We hypothesize that the innovative design and properties of NP-containing light-responsive ISFIs can serve as a platform for effective management of ocular diseases requiring long term treatment., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Recent studies have shown that Connexin43 mimetic peptide (Cx43 MP) can prevent secondary damage following several retinal ischemic and inflammatory disorders by blocking the pathological opening of gap junction hemichannels. However, the poor stability of peptides and the presence of various intraocular barriers limit efficient retinal delivery in the clinical setting. The present study aimed to prolong the bioactivity of Cx43 MP and achieve targeted delivery to the retina by loading the peptide into hyaluronic acid (HA) coated human serum albumin nanoparticles (HSA NPs). Two different loading methods, adsorption and incorporation, were used with the peptide released slowly over a period of up to four months. Compared to uncoated particles, HA coated HSA NPs exhibited enhanced in vitro cellular uptake and ex vivo retinal penetration via HA-CD44 receptor mediated interactions. Furthermore, cell viability and Cx43 MP functionality assays showed that NPs protected Cx43 MP from degradation, sustained its release, and thus prolonged its bioactivity without reducing cell viability at concentrations used for Cx43 hemichannel blocking. Therefore, HA coated HSA NPs could have great potential for sustained and targeted delivery of Cx43 MP to treat various retinal inflammatory conditions.

The efficacy of antisense oligodeoxynucleotides (AsODNs) is compromised by their poor stability in biological fluids and the inefficient cellular uptake due to their size and negative charge. Since chemical modifications of these molecules have resulted in a number of non-antisense activities, incorporation into particulate delivery systems has offered a promising alternative. The aim of this study was to evaluate various poly(D,L-lactic- co-glycolic acid) (PLGA) nanoparticles for AsODN entrapment and delivery. PLGA nanoparticles were prepared using the double emulsion solvent evaporation method. The influence of formulation parameters such as PLGA concentration and volume ratio of internal aqueous phase volume (Va1) to organic phase volume (Vo) to external aqueous phase volume (Va2) on particle size, polydispersity index (PDI) and zeta potential (ZP) was investigated using a full factorial study. The particle size increased with increasing PLGA concentrations and volume ratios, with an interaction detectable between the two factors. AsODN entrapment efficiencies ranged between 49.97% and 54.95% with no significant difference between various formulations. By fitting the in vitro release profiles to a dual first order release model it was shown that the AsODN release occurred via two processes: a diffusion controlled process in the early phase (25 to 32% within one day) and a PLGA degradation process in the latter (39 to 70% after 14 days). Cellular uptake studies using primary corneal epithelial cells suggested active transport of nanoparticles via endocytosis. PLGA nanoparticles therefore show potential to successfully entrap AsODNs, transport them into cells and release them over time due to polymer erosion. [ABSTRACT FROM AUTHOR]

Glaucoma is a common progressive eye disorder which remains the second leading cause of blindness worldwide. Current therapy involves frequent administration of eye drops which often results in poor patient adherence and therapeutic outcomes. The aim of this study was to overcome these limitations by developing a novel nanoparticle cross-linked collagen shield for sustained delivery of pilocarpine hydrochloride (PHCl). Three metal oxide nanoparticles (NPs); titanium dioxide (TiO 2 ), zinc oxide (ZnO) and polyvinylpyrrolidone (PVP) capped zinc oxide (ZnO/PVP), were evaluated for their cytotoxicity as well as shield transparency before selecting ZnO/PVP NPs as the ideal candidate. Cross-linked collagen shields were then characterized for their mechanical strength, swelling capacity and bioadhesive properties, with ZnO/PVP NP cross-linked shields showing the most favorable characteristics compared to plain films. The shield with the best properties was then loaded with PHCl and in vitro release of zinc ions as well as PHCl was measured without and with further cross-linking by ultraviolet irradiation. The concentration of zinc ions released was well below the IC 50 rendering them safe for ocular use. Moreover, collagen shields cross-linked with ZnO/PVP NPs released PHCl over a period of 14 days offering a promising sustained release treatment option for glaucoma. [ABSTRACT FROM AUTHOR]