Self-Assembling Topical Nanomicellar Formulation to Improve Curcumin Absorption Across Ocular Tissues.

The pathophysiological mechanisms for dry and wet age-related macular degeneration (AMD) involve oxidative stress and increased VEGF release and expression. An ideal drug candidate for both types of AMD is the one which offers significant protection to the retinal cells from oxidative stress and inhibit VEGF release. Curcumin is one such natural product which provides numerous beneficial effects including antioxidant, anti-inflammatory, and anti-VEGF activities and has the potential for the treatment of both types of AMD. The bioavailability of curcumin is negligible due to its poor aqueous solubility. The purpose of this work is to develop an aqueous nanomicellar drop formulation of curcumin (CUR-NMF) for back of the eye delivery utilizing hydrogenated castor oil (HCO-40) and octoxynol-40 (OC-40) to treat AMD. A full factorial design was performed with JMP software analysis to optimize the formulation size, polydispersity index (PDI), entrapment efficiency, loading, and precipitation. MTT and LDH assays on human retinal pigmented epithelial (D407) cells revealed that 5-10 µM CUR-NMF dose is safe for ophthalmic use. Furthermore, CUR-NMF exhibited significant protection of retinal (D407) cells against H2O2-induced oxidative stress. In vitro drug release kinetics suggested a sustained drug release profile indicating a long-term protection ability of CUR-NMF against oxidative stress to retinal cells. In addition, an ELISA suggested that CUR-NMF significantly reduces vascular endothelial growth factor (VEGF) release in D407 cell line, hence diminishes the risk of angiogenesis. Collectively, these results suggest that the proposed CUR-NMF can be tremendously effective in treating both types of AMD.

In vivo measurement of trabecular meshwork stiffness in a corticosteroid-induced ocular hypertensive mouse model.

Ocular corticosteroids are commonly used clinically. Unfortunately, their administration frequently leads to ocular hypertension, i.e., elevated intraocular pressure (IOP), which, in turn, can progress to a form of glaucoma known as steroid-induced glaucoma. The pathophysiology of this condition is poorly understood yet shares similarities with the most common form of glaucoma. Using nanotechnology, we created a mouse model of corticosteroid-induced ocular hypertension. This model functionally and morphologically resembles human ocular hypertension, having titratable, robust, and sustained IOPs caused by increased resistance to aqueous humor outflow. Using this model, we then interrogated the biomechanical properties of the trabecular meshwork (TM), including the inner wall of Schlemm's canal (SC), tissues known to strongly influence IOP and to be altered in other forms of glaucoma. Specifically, using spectral domain optical coherence tomography, we observed that SC in corticosteroid-treated mice was more resistant to collapse at elevated IOPs, reflecting increased TM stiffness determined by inverse finite element modeling. Our noninvasive approach to monitoring TM stiffness in vivo is applicable to other forms of glaucoma and has significant potential to monitor TM function and thus positively affect the clinical care of glaucoma, the leading cause of irreversible blindness worldwide.

Ocular Pharmacokinetics of a Topical Ophthalmic Nanomicellar Solution of Cyclosporine (Cequa®) for Dry Eye Disease.

Cequa®, a unique and first-in-class preservative free cyclosporine-A (CsA) nanomicellar topical formulation was recently approved by US FDA for treatment of dry eye disease or keratoconjuntivitis sicca (KCS). Being highly hydrophobic, CsA is currently available as an oil based emulsion, which has its own shortcomings. Developing an aqueous and clear formulation of CsA is imperative yet a challenging need in the quest for a safe and better drug product. In this regard, a novel, clear, aqueous nanomicellar solution of CsA was developed which has the potential to deliver therapeutic concentrations of CsA with minimal discomfort to patients. Highly promising pre-clinical results of Cequa® (OTX-101), has led to its advancement to the clinical trials. Phase III clinical trials have demonstrated that OTX-101 is highly effective, safe, and has a rapid onset of action in treating KCS. This review presents a comprehensive insight on formulation development, preclinical and clinical pharmacokinetic results of Cequa®. Additionally, the translational development of Cequa® from the laboratory benchtop to patient bedside has been discussed.

Multi-Layered Nanomicelles as Self-Assembled Nanocarrier Systems for Ocular Peptide Delivery.

Despite the great potential of peptides as therapeutics, there is an unmet challenge in sustaining delivery of sufficient amounts in their native forms. This manuscript describes a novel nanocarrier capable of delivering functional small peptides in its native form. Self-assembling multi-layered nanomicelles composed of two polymers, polyoxyethylene hydrogenated castor oil 40 (HCO-40) and octoxynol 40 (OC-40), were designed to combine hydrophilic interaction and solvent-induced encapsulation of peptides and proteins. The polymers are employed to encapsulate peptide or protein in the core of the organo-nanomicelles which are further encapsulated with another layer of the same polymers to form an aqueous stable nanomicellar solution. The size of the multi-layered nanomicelles ranges from ~ 16 to 20 nm with zeta potential close to neutral (~ - 2.44 to 0.39 mV). In vitro release studies revealed that octreotide-loaded multi-layered nanomicelles released octreotide at much slower rate in simulated tear fluid (STF) (~ 27 days) compared to PBST (~ 11 days) in its native form. MTT assay demonstrated negligible toxicity of the multi-layered nanomicelles at lower concentrations in human retinal pigment epithelial (HRPE, D407), human conjunctival epithelial (CCL 20.2), and rhesus choroid-retinal endothelial (RF/6A) cells. This work demonstrates an efficient small peptide delivery platform with significant advantages over existing approaches, as it does not require modification of the peptide, is biodegradable, and has a small size and high loading capacity.

Novel Random Triblock Copolymers for Sustained Delivery of Macromolecules for the Treatment of Ocular Diseases.

The objective of this study is to design, develop, and synthesize novel random triblock (RTB) copolymers for sustained delivery of macromolecules. RTB copolymers have not been utilized for the delivery of macromolecules for ocular diseases. RTB copolymers comprising of polyethylene glycol, glycolide, and ɛ-caprolactone blocks were synthesized and assessed for their molecular weights and purity using 1H-NMR spectroscopy, gel permeation chromatography, FTIR (functionality), and XRD (crystallinity). No toxicity was observed when ocular cell lines were treated with RTB copolymers. These materials were applied for encapsulation of peptides and proteins (catalase, IgG, BSA, IgG Fab fragment, lysozyme, insulin, and octreotide) in nanoparticles. Particle size ranged from 202.41 ± 2.45 to 300.1 ± 3.11 nm depending on the molecular size and geometry of proteins/peptides. Polydispersity indices were between 0.26 ± 0.02 and 0.46 ± 0.07 respectively. Percentage entrapment efficiency and drug loading ranged from 83.44 ± 2.24 to 45.35 ± 5.53 and 21.56 ± 0.46 to 13.08 ± 1.35 respectively depending on molecular weights of peptides or proteins. A sustained in vitro release of macromolecule was observed over 3-month period. These results suggest that RTB copolymers may be suitable for sustained delivery systems for various macromolecules for different diseases including ocular diseases.

The relationship between outflow resistance and trabecular meshwork stiffness in mice.

It has been suggested that common mechanisms may underlie the pathogenesis of primary open-angle glaucoma (POAG) and steroid-induced glaucoma (SIG). The biomechanical properties (stiffness) of the trabecular meshwork (TM) have been shown to differ between POAG patients and unaffected individuals. While features such as ocular hypertension and increased outflow resistance in POAG and SIG have been replicated in mouse models, whether changes of TM stiffness contributes to altered IOP homeostasis remains unknown. We found that outer TM was stiffer than the inner TM and, there was a significant positive correlation between outflow resistance and TM stiffness in mice where conditions are well controlled. This suggests that TM stiffness is intimately involved in establishing outflow resistance, motivating further studies to investigate factors underlying TM biomechanical property regulation. Such factors may play a role in the pathophysiology of ocular hypertension. Additionally, this finding may imply that manipulating TM may be a promising approach to restore normal outflow dynamics in glaucoma. Further, novel technologies are being developed to measure ocular tissue stiffness in situ. Thus, the changes of TM stiffness might be a surrogate marker to help in diagnosing altered conventional outflow pathway function if those technologies could be adapted to TM.

Ocular delivery of proteins and peptides: Challenges and novel formulation approaches.

The impact of proteins and peptides on the treatment of various conditions including ocular diseases over the past few decades has been advanced by substantial breakthroughs in structural biochemistry, genetic engineering, formulation and delivery approaches. Formulation and delivery of proteins and peptides, such as monoclonal antibodies, aptamers, recombinant proteins and peptides to ocular tissues poses significant challenges owing to their large size, poor permeation and susceptibility to degradation. A wide range of advanced drug delivery systems including polymeric controlled release systems, cell-based delivery and nanowafers are being exploited to overcome the challenges of frequent administration to ocular tissues. The next generation systems integrated with new delivery technologies are anticipated to generate improved efficacy and safety through the expansion of the therapeutic target space. This review will highlight recent advances in formulation and delivery strategies of protein and peptide based biopharmaceuticals. We will also describe the current state of proteins and peptides based ocular therapy and future therapeutic opportunities.

Nanoparticles in thermosensitive gel based composite nanosystem for ocular diseases.

The pentablock (PB) copolymers based composite nanosystems were designed to provide a long-term delivery of macromolecules to the back of the eye. A unique arrangement of each block (polyethylene glycol, polylactic acid, and polycaprolactone) with various molecular weights (PB-A and PB-B) was selected for the synthesis of nanoparticles (NPs) and thermosensitive gel (PB-C) by sequential ring-opening bulk copolymerization reaction. PB copolymers were characterized for their molecular weight and purity by 1H-NMR spectroscopy and crystallinity by PXRD. The macromolecule model drugs [lysozyme (Lyz ~ 14.5 kDa), IgG-Fab (~ 50 kDa), and IgG (~ 150 kDa)] were selected to delineate the effect of molecular weights on in vitro release profile of nanoformulations. Lyz-, Fab-, and IgG-encapsulated NPs were prepared by double emulsion solvent evaporation method. The entrapment efficiency (EE%) and drug loading (DL%) of macromolecules was higher for PB-B copolymers due to its higher molecular weight and hydrophobicity compare to PB-A. The particle size range of NPs was ~ 200-270 nm. In vitro release profiles of Lyz-, Fab-, and IgG-encapsulated in NPs alone and NPs suspended in gel (composite nanosystem) demonstrated a minimal burst release and drug release over a long period. The effect of hydrodynamic diameter of macromolecules and hydrophobicity of PB copolymers was investigated on the release profile of nanosystems. In vitro biocompatibility study showed negligible cytokine (IL-1, IL-6, and TNF-α) release, which confirmed the safety of the PB copolymers. Based on the results, it is anticipated that long-term ocular delivery of macromolecules can be achieved through composite nanosystems.

Recent Patents and Emerging Therapeutics on Ocular Inflammation and Allergy.

BACKGROUND: Ocular inflammation and allergic eye diseases range from mild to severe may disturb visual function and affect` quality of life. Since these diseases require intensive therapies, the pathophysiology and treatments of these conditions are highlighted. OBJECTIVE: The ocular diseases caused by inflammation and allergy are extensively studied in this review to provide an overview of the newer compounds, novel delivery approaches, preclinical and clinical trials for the treatment of allergic conjunctivitis, dry eye syndrome, and uveitis. METHOD: The eye is divided into two segments; anterior and posterior. Both segments provide barriers to the drug delivery to the eye. Despite many efforts by scientists, several potential drug candidates are often dropped from the initial screening portfolio due to failure in overcoming these barriers. Thus to overcome unmet challenges, remarkable progresses have been made towards the design of novel ocular therapeutics with enhanced activity and minimal toxicity to the ocular tissue. A comprehensible understanding of the diseased conditions, physiological barriers and pharmacokinetics of the eye would significantly accelerate the development of new therapeutics. Moreover, identification of new targets drives the discovery of novel drug molecules for the ocular disease treatment. RESULTS: The advancement in the drug discovery and dosage from design showcases the increasing number of patent applications being filed and issued for allergic conjunctivitis, dry eye syndrome, and uveitis. In addition, preclinical and clinical trials are now becoming available showing the newer generation of ocular drugs. CONCLUSION: This review presented a brief background on the disease condition, types, treatment, advancement in the delivery approaches, focus on emerging therapeutics, related patents and clinical trials for the treatment of allergic conjunctivitis, dry eye syndrome, and uveitis.

Pentablock copolymer dexamethasone nanoformulations elevate MYOC: in vitro liberation, activity and safety in human trabecular meshwork cells.

AIM: The aim of this study is to examine the elevation of MYOC in long-term treatment of human trabecular meshwork (HTM) cells using dexamethasone (DEX) encapsulated pentablock (PB) copolymer-based nanoparticles (NPs) (DEX-PB-NPs). MATERIALS & METHODS: PB copolymers and DEX-PB-NPs were synthesized and characterized using nuclear magnetic resonance, gel permeation chromatography, and X-ray diffraction analyses. MYOC levels secreted from HTM cells were measured by western blot (WB) analysis. RESULTS: DEX-PB-NPs were formulated in the size range of 109 ± 3.77 nm (n = 3). A long term DEX release from the NPs was observed over three months. Cell viability and cytotoxicity were not affected up to 12 weeks of treatment with PB-copolymer or DEX-PB-NPs. WB data from five HTM cell strains showed that MYOC levels increased by 5.2 ± 1.3, 7.4 ± 4.3, and 2.8 ± 1.1-fold in the presence of DEX-PB-NPs compared with 9.2 ± 3.8, 2.2 ± 0.5, and 1.5 ± 0.3-fold at 4, 8 and 12 weeks in control-DEX treatment group, respectively (n = 5). Based on the decline in MYOC levels after withdrawal of DEX from control wells, DEX-PB-NPs released the DEX for at least 10 weeks. CONCLUSION: The treatment of HTM cells using DEX-PB-NPs were analyzed in this study. The in vitro cell-based system developed here is a valuable tool for determining the safety and effects of steroids released from polymeric NPs.

Topical Formulation of Self-Assembled Antiviral Prodrug Nanomicelles for Targeted Retinal Delivery.

Topical drug administration for back of the eye delivery is extremely challenging due to the presence of protection mechanisms and physiological barriers. Self-assembled polymeric nanomicelles have emerged as promising vehicles for drug delivery. Apart from serving as an inert nanocarrier for therapeutic agents, polymeric nanomicelles are known to bypass mononuclear phagocytic system (MPS) and efflux transporters thereby improving drug bioavailability. In this investigation, a highly efficacious biotinylated lipid prodrug of cyclic cidofovir (B-C12-cCDF) was formulated within polymeric nanomicelles as a carrier for targeted retinal delivery. Polymeric nanomicelles were prepared from polyoxyethylene hydrogenated castor oil 40 (HCO-40) and octoxynol 40 (OC-40). In vitro release studies revealed that B-C12-cCDF-loaded nanomicelles released B-C12-cCDF at a faster rate in stimulated tear fluid (STF) in comparison to PBST. MTT and LDH assays demonstrated negligible cytotoxicity of B-C12-cCDF-loaded nanomicelles relative to CDF and B-C12-cCDF in HRPE (human retinal pigment epithelial, D407), HCE-T (human corneal epithelial), and CCL 20.2 (human conjunctival epithelial) cells. Confocal laser scanning microscopy and flow cytometry analyses indicated that B-C12-cCDF-loaded nanomicelles were efficiently internalized into D407 and HCE-T cells in contrast to CDF and B-C12-cCDF. Moreover, little B-C12-cCDF was also observed in the nuclei after 24 h of incubation. Polymeric nanomicelles carrying the transporter targeted prodrug did not produce any cytotoxic effects and were internalized into the cells effectively. Permeability experiments across HCE-T cells further confirmed significant transport of prodrug loaded nanomicelles and their subsequent uptake into D407 cells. These findings indicate that HCO-40/OC-40 based polymeric nanomicelles could become a promising topical delivery system for ocular administration of antiviral agents.

Clear, Aqueous Topical Drop of Triamcinolone Acetonide.

The objective of this study was to develop a clear aqueous mixed nanomicellar formulation (NMF) of triamcinolone acetonide (TA) with a combination of nonionic surfactant hydrogenated castor oil 60 (HCO-60) and octoxynol-40 (Oc-40). In order to delineate the effects of drug-polymer interactions on entrapment efficiency (EE), loading efficiency (LE), and critical micellar concentration (CMC), a design of experiment (DOE) was performed to optimize the formulation. In this study, full-factorial design has been used with HCO-60 and OC-40 as independent variables. All formulations were prepared following solvent evaporation and film rehydration method, characterized with size, polydispersity, shape, morphology, EE, LE, and CMC. A specific blend of HCO-60 and Oc-40 at a particular wt% ratio (5:1.5) produced highest drug EE, LE, and smallest CMC (0.0216 wt%). Solubility of TA in NMF improved 20 times relative to normal aqueous solubility. Qualitative 1H NMR studies confirmed the absence of free drug in the outer aqueous NMF medium. Moreover, TA-loaded NMF appeared to be highly stable and well tolerated on human corneal epithelial cells (HCEC) and human retinal pigment epithelial cells (D407 cells). Overall, these studies suggest that TA in NMF is safe and suitable for human topical ocular drop application.

Polymeric micelles for ocular drug delivery: From structural frameworks to recent preclinical studies.

Effective intraocular drug delivery poses a major challenge due to the presence of various elimination mechanisms and physiological barriers that result in low ocular bioavailability after topical application. Over the past decades, polymeric micelles have emerged as one of the most promising drug delivery platforms for the management of ocular diseases affecting the anterior (dry eye syndrome) and posterior (age-related macular degeneration, diabetic retinopathy and glaucoma) segments of the eye. Promising preclinical efficacy results from both in-vitro and in-vivo animal studies have led to their steady progression through clinical trials. The mucoadhesive nature of these polymeric micelles results in enhanced contact with the ocular surface while their small size allows better tissue penetration. Most importantly, being highly water soluble, these polymeric micelles generate clear aqueous solutions which allows easy application in the form of eye drops without any vision interference. Enhanced stability, larger cargo capacity, non-toxicity, ease of surface modification and controlled drug release are additional advantages with polymeric micelles. Finally, simple and cost effective fabrication techniques render their industrial acceptance relatively high. This review summarizes structural frameworks, methods of preparation, physicochemical properties, patented inventions and recent advances of these micelles as effective carriers for ocular drug delivery highlighting their performance in preclinical studies.

Recent perspectives on the delivery of biologics to back of the eye.

INTRODUCTION: Biologics are generally macromolecules, large in size with poor stability in biological environments. Delivery of biologics to tissues at the back of the eye remains a challenge. To overcome these challenges and treat posterior ocular diseases, several novel approaches have been developed. Nanotechnology-based delivery systems, like drug encapsulation technology, macromolecule implants and gene delivery are under investigation. We provide an overview of emerging technologies for biologics delivery to back of the eye tissues. Moreover, new biologic drugs currently in clinical trials for ocular neovascular diseases have been discussed. Areas covered: Anatomy of the eye, posterior segment disease and diagnosis, barriers to biologic delivery, ocular pharmacokinetic, novel biologic delivery system Expert opinion: Anti-VEGF therapy represents a significant advance in developing biologics for the treatment of ocular neovascular diseases. Various strategies for biologic delivery to posterior ocular tissues are under development with some in early or late stages of clinical trials. Despite significant progress in the delivery of biologics, there is unmet need to develop sustained delivery of biologics with nearly zero-order release kinetics to the back of the eye tissues. In addition, elevated intraocular pressure associated with frequent intravitreal injections of macromolecules is another concern that needs to be addressed.

Transporter effects on cell permeability in drug delivery.

INTRODUCTION: The role of drug transporters as one of the determinants of cellular drug permeability has become increasingly evident. Despite the lipophilicity of a drug molecule as rate-limiting factor for passive diffusion across biological membranes, carrier-mediated and active transport have gained attention over the years. A better understanding of the effects and roles of these influx transporters towards transmembrane permeability of a drug molecule need to be delineated for drug development and delivery. Areas covered: This review focuses on findings relative to role of transporters in drug absorption and bioavailability. Particularly the areas demanding further research have been emphasized. This review will also highlight various transporters expressed on vital organs and their effects on drug pharmacokinetics. Expert opinion: Significant efforts have been devoted to understand the role of transporters, their iterative interplay with metabolizing enzymes through molecular enzymology, binding and structure-activity relationship studies. A few assays such as parallel artificial membrane permeation assay (PAMPA) have been developed to analyze drug transport across phospholipid membranes. Although large web-accessible databases on tissue selective expression profiles at transcriptomic as well as proteomic are available, there is a need to collocate the scattered literature on the role of transporters in drug development and delivery.

How are we improving the delivery to back of the eye? Advances and challenges of novel therapeutic approaches.

INTRODUCTION: Drug delivery to the back of the eye requires strategic approaches that guarantee the long-term therapeutic effect with patient compliance. Current treatments for posterior eye diseases suffer from significant challenges including frequent intraocular injections of anti-VEGF agents and related adverse effects in addition to the high cost of the therapy. Areas covered: Treatment challenges and promising drug delivery approaches for posterior segment eye diseases, such as age-related macular degeneration (AMD) are summarized. Advances in the development of several nanotechnology-based systems, including stimuli-responsive approaches to enhance drug bioavailability and overcome existing barriers for effective ocular delivery are discussed. Stem cell transplantation and encapsulated cell technology (ECT) approaches to treat posterior eye diseases are elaborated. Expert opinion: There are several drug delivery systems demonstrating promising results. However, a better understanding of ocular barriers, disease pathophysiology, and drug clearance mechanisms is required for better therapeutic outcomes. The stem cell transplantation strategy and ECT approach provide positive results in AMD therapy, but there are a number of challenges that must be overcome for long-term efficiency. Ultimately, there are numerous multidimensional challenges to cure vision problems and a collaborative approach among scientists is required.

A comprehensive insight on ocular pharmacokinetics.

The eye is a distinctive organ with protective anatomy and physiology. Several pharmacokinetics compartment models of ocular drug delivery have been developed for describing the absorption, distribution, and elimination of ocular drugs in the eye. Determining pharmacokinetics parameters in ocular tissues is a major challenge because of the complex anatomy and dynamic physiological barrier of the eye. In this review, pharmacokinetics of these compartments exploring different drugs, delivery systems, and routes of administration is discussed including factors affecting intraocular bioavailability. Factors such as precorneal fluid drainage, drug binding to tear proteins, systemic drug absorption, corneal factors, melanin binding, and drug metabolism render ocular delivery challenging and are elaborated in this manuscript. Several compartment models are discussed; these are developed in ocular drug delivery to study the pharmacokinetics parameters. There are several transporters present in both anterior and posterior segments of the eye which play a significant role in ocular pharmacokinetics and are summarized briefly. Moreover, several ocular pharmacokinetics animal models and relevant studies are reviewed and discussed in addition to the pharmacokinetics of various ocular formulations.

Circumvention of P-gp and MRP2 mediated efflux of lopinavir by a histidine based dipeptide prodrug.

PURPOSE: This study was aimed to develop a novel Histidine-Leucine-Lopinavir (His-Leu-LPV) dipeptide prodrug and evaluate its potential for circumvention of P-gp and MRP2-mediated efflux of lopinavir (LPV) indicated for HIV-1 infection. METHODS: His-Leu-LPV was synthesized following esterification of hydroxyl group of LPV and was identified by (1)H NMR and LCMS/MS techniques. Aqueous solubility, stability and cell cytotoxicity of prodrug was determined. Uptake and permeability studies were carried out using P-gp (MDCK-MDR1) and MRP2 (MDCK-MRP2) transfected cell lines. To further delineate prodrug uptake, prodrug interaction with influx transporters (PepT1 and PHT1) was determined. Enzymatic hydrolysis and reconversion of His-Leu-LPV to LPV was examined using Caco-2 cell homogenates. RESULTS: Aqueous solubility generated by the prodrug was markedly higher relative to unmodified LPV. Importantly, His-Leu-LPV displayed significantly lower affinity towards P-gp and MRP2 as evident from higher uptake and transport rates. [3H]-GlySar and [3H]-l-His uptake receded to approximately 30% in the presence of His-Leu-LPV supporting the PepT1/PHT1 mediated uptake process. A steady regeneration of LPV and Leu-LPV in Caco-2 cell homogenates indicated His-Leu-LPV undergoes both esterase and peptidase-mediated hydrolysis. CONCLUSION: Histidine based dipeptide prodrug approach can be an alternative strategy to improve LPV absorption across poorly permeable intestinal barrier.