Nanofiber-coated implants: Development and safety after intravitreal application in rabbits.

Intravitreal injections are the preferred choice for drug administration to the posterior segment of the eye. However, the required frequent injections may cause complications to the patient and low adherence to the treatment. Intravitreal implants are able to maintain therapeutic levels for a long period. Biodegradable nanofibers can modulate drug release and allow the incorporation of fragile bioactive drugs. Age-related macular degeneration is one of the world major causes of blindness and irreversible vision loss. It involves the interaction between VEGF and inflammatory cells. In this work we developed nanofiber-coated intravitreal implants containing dexamethasone and bevacizumab for simultaneously delivery of these drugs. The implant was successfully prepared and the efficiency of the coating process was confirmed by scanning electron microscopy. Around 68% of dexamethasone was released in 35 days and 88% of bevacizumab in 48hs. The formulation presented activity in the reduction of vessels and was safe to the retina. It was not observed any clinical or histopathological change, neither alteration in retina function or thickness by electroretinogram and optical coherence tomography during 28 days. The nanofiber-coated implants of dexamethasone and bevacizumab may be considered as a new delivery system that can be effective for the treatment of AMD.

Intravitreal thalidomide ameliorates inflammation in a model of experimental uveitis induced by BCG.

Uveitis encompasses a heterogeneous and complex group of conditions characterized by intraocular inflammation, frequently affecting young individuals and representing an important cause of irreversible blindness worldwide. Animal models have been critical to understand etiology and pathogenesis of uveitis, being also employed to assess new therapeutic strategies, preceding human studies. However, there is still a need of developing and studying different models, due to the difficulties in recapitulating all forms of human uveitis effectively. Although corticosteroids are usually the first-line therapy for non-infectious uveitis, their long-term use is limited by potentially serious side effects in all possible delivery routes. Thus, thalidomide, a drug with anti-inflammatory and antiangiogenic properties, was investigated in a novel experimental model of uveitis, induced by Mycobacterium bovis Calmette-Guérin Bacillus (BCG), in rabbits. The experimental protocol consisted of two subcutaneous injections of BCG, followed by two intravitreal injections of the same antigen, inducing panuveitis. Animals were treated with a single intravitreal injection of thalidomide suspension or PBS. Clinical manifestations of uveitis improved after intravitreal thalidomide, involving both anterior and posterior segments. Protein content, N-acetyl-b-glucosaminidase (NAG) and myeloperoxidase (MPO) activities were elevated in ocular tissues after disease induction, further decreasing post-treatment with intravitreal thalidomide. This therapeutic response was also confirmed on ocular electrophysiology, as well as histopathology. This experimental model induced panuveitis in rabbits using a low-cost mycobacterial antigen, with intraocular inflammation subsequently improving after treatment. Intravitreal thalidomide may be a potential alternative to treat intraocular inflammation in corticosteroid-sparing therapies.

Toxicity and in vivo release profile of sirolimus from implants into the vitreous of rabbits' eyes.

PURPOSE: To assess the in vivo release profile and the retinal toxicity of a poly (lactic-co-glycolic acid) (PLGA) sustained-release sirolimus (SRL) intravitreal implant in normal rabbit eyes. METHODS: PLGA intravitreal implants containing or not SRL were prepared, and the viability of ARPE-19 and hES-RPE human retinal cell lines was examined after 24 and 72 h of exposure to implants. New Zealand rabbits were randomly divided into two groups that received intravitreal implants containing or not SRL. At each time point (1-8 weeks), four animals from the SRL group were euthanized, the vitreous was collected, and drug concentration was calculated. Clinical evaluation of the eyes was performed weekly for 8 weeks after administration. Electroretinography (ERG) was recorded in other eight animals, four for each group, at baseline and at 24 h, 1, 4, 6, and 8 weeks after the injection. ERG was carried out using scotopic and photopic protocols. The safety of the implants was assessed using statistical analysis of the ERG parameters (a and b waves, a and b implicit time, B/A ratio, oscillatory potential, and Naka-Rushton analysis) comparing the functional integrity of the retina between the PLGA and SRL-PLGA groups. After the last electrophysiological assessment, the rabbits were euthanized and retinal histopathology was realized. RESULTS: After 24 and 72 h of incubation with PLGA or SRL-PLGA implants, ARPE-19 and hES-RPE cells showed viability over 70%. The maximum concentration of SRL (199.8 ng/mL) released from the device occurred within 4 weeks. No toxic effects of the implants or increase in the intraocular pressure was observed through clinical evaluation of the eye. ERG responses showed no significant difference between the eyes that received PLGA or SRL-PLGA implants at baseline and throughout the 8 weeks of follow-up. No remarkable difference in retinal histopathology was detected in rabbit eyes treated with PLGA or SRL-PLGA implants. CONCLUSIONS: Intravitreal PLGA or SRL-PLGA implants caused no significant reduction in cell viability and showed no evident toxic effect on the function or structure of the retina of the animals. SRL was released from PLGA implant after application in the vitreous of rabbits during 8 weeks.

Biodegradable core-shell electrospun nanofibers containing bevacizumab to treat age-related macular degeneration.

Age-related macular degeneration (AMD) is a degenerative ocular disease that affects the central retina. It is considered the main cause of blindness and loss of vision worldwide. Angiogenic factors are associated with AMD, which has led to the use of antiangiogenic drugs, such as bevacizumab, to treat the disease using frequent intravitreal injections. In the present study, biodegradable core shell nanofibers containing bevacizumab were prepared by the coaxial electrospinning technique. It is thought that the shell could control the release of the drug, while the core would protect and store the drug. Poly(caprolactone) (PCL) and gelatin were used to form the shell of the nanofibers, while poly(vinyl alcohol) (PVA) and bevacizumab comprised the core. The nanofibers were characterized using microscopy techniques, thermal analysis, and FTIR. The results showed that core-shell nanofibers were produced as designed. Bevacizumab activity was evaluated using a chicken embryo chorioallantoic membrane (CAM) assay. An enzyme-linked immunosorbent assay was used to quantify the amount of the drug released from the different nanofibers in vitro. The toxicity of the nanofibers was evaluated in human retinal pigment epithelial (ARPE) cells. The CAM results demonstrated that bevacizumab maintained its antiangiogenic activity when incorporated into the nanofibers. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) tests revealed that the nanofibers showed no cellular toxicity, even in the presence of bevacizumab. The core-shell structure of the nanofibers reduced the release rate of bevacizumab compared with PVA nanofibers. The bevacizumab-loaded biodegradable nanofibers presented interesting properties that would potentially constitute an alternative therapy to intravitreal injections to treat AMD.

In vivo evaluation of antitumoral and antiangiogenic effect of imiquimod-loaded polymeric nanoparticles.

The chemotherapeutic agent imiquimod (Imq) is used to treat skin cancers, the most common type of human cancer. However, the high incidence of local and systemic side effects associated with its use as well as its low skin permeation impair patient compliance and therapeutic effectiveness To overcome these limitations, nanostructured systems such as nanoparticles can be a promising alternative. Nanoparticles are submicron particles (size less than 1000 nm) with high surface area that facilitates the interaction and cellular uptake by biological membranes. Therefore, the aim of the present work is to evaluate antiangiogenic effect and antitumoral activity of imiquimod-loaded nanoparticles compared to market Imq formulation. Polymeric nanoparticles containing Imq were obtained by the technique of precipitation of preformed polymer. Antiangiogenic activity of the formulations was determined in chicken embryo chorioallantoic membrane (CAM) and its chemopreventive potential was evaluate during multistage DMBA and croton oil model of skin carcinogenesis in mice. Nanoparticles containing Imq presented antiangiogenic activity superior than negative control, placebo dispersion and market Imq (p < 0.05) in the CAM model and also significantly reduced the number and size of papillomas compared to all other groups. These results suggest, therefore, that the obtained delivery system can be an alternative to treat diseases related to vessels formation and also potentially increase cutaneous permeation and efficacy of poor soluble drugs normally used to treat cutaneous diseases.