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Pathological neovascularization is a hallmark of many vision-threatening diseases. However, some patients exhibit poor responses to current anti-VEGF therapies due to resistance and limited efficacy. Recent studies have highlighted the roles of noncoding RNAs in various biological processes, paving the way for RNA-based therapeutics. In this study, we report a marked down-regulation of miR-205 under pathological conditions. miR-205 potently inhibits endothelial cell functions critical for pathological neovascularization, including proliferation, migration, and tube formation. Furthermore, miR-205 strengthens the endothelial barrier, thereby reducing vascular leakage. In mouse models of retinal and choroidal neovascularization, miR-205 administration effectively suppresses abnormal blood vessel formation and leakage. Mechanistically, miR-205 directly targets VEGFA and ANGPT2, which are key drivers of pathological neovascularization. To improve delivery, we successfully loaded miR-205 into exosomes derived from mesenchymal stem cells. This innovative approach avoids cytotoxicity while preserving therapeutic efficacy in both cellular and animal models. Collectively, our findings highlight miR-205 as a promising therapeutic for ocular neovascularization, with exosome delivery offering a novel and efficient strategy for treating vision-threatening vascular diseases. [ABSTRACT FROM AUTHOR]

Angiogenesis is vital to developmental, regenerative and repair processes. It is normally regulated by a balanced production of pro- and anti-angiogenic factors. Alterations in this balance under pathological conditions are generally mediated through up-regulation of pro-angiogenic and/or downregulation of anti-angiogenic factors, leading to growth of new and abnormal blood vessels. The pathological manifestation of many diseases including cancer, ocular and vascular diseases are dependent on the growth of these new and abnormal blood vessels. Thrompospondin-1 (TSP1) was the first endogenous angiogenesis inhibitor identified and its anti-angiogenic and anti-inflammatory activities have been the subject of many studies. Studies examining the role TSP1 plays in pathogenesis of various ocular diseases and vascular dysfunctions are limited. Here we will discuss the recent studies focused on delineating the role TSP1 plays in ocular vascular development and homeostasis, and pathophysiology of various ocular and vascular diseases with a significant clinical relevance to human health. [ABSTRACT FROM AUTHOR]

Purpose: Recent studies have reported the promising effect of intravitreal propranolol on retinal neovascularization. However, rapid clearance and short half-life of the drug in the vitreous are the main drawbacks of this therapeutic approach. This study investigates the extension of the residence time of propranolol in the vitreous by polymeric nanoparticles (NPs) with the prospect of improving choroidal neovascularization treatment. Methods: The poly (lactic-co-glycolic) acid (PLGA) NPs were fabricated by a modified double emulsion solvent evaporation method and the obtained NPs were characterized for their size, poly dispersity index (PDI), and surface image. The in vitro release, cell cytotoxicity, and uptake of NPs were also evaluated. To investigate the effect of the vitreous pharmacokinetic drug loaded NPs versus that of the free propranolol, they were intravitreally injected into the rabbits' eyes and the drug vitreous concentrations in defined intervals were analyzed by high performance liquid chromatography (HPLC). Results: The spherical NPs with about 230 nm size, and almost 10% drug loading were obtained. Based on the 3-(4, 5-Dimethylthiazol-2-Yl)-2, 5-Diphenyltetrazolium Bromide (MTT) outcomes, 30 µg/ml of propranolol was considered as the guide dosage in the intravitreal injection. Confocal microscopy images verified the presence of labeled NPs in the posterior segment after five days of receiving the injection. In vivo assay revealed that the vanishing rate of propranolol in rabbits treated with propranolol NPs was reduced at twice the rate as compared to that of the vanishing rate experienced with only the free drug. Conclusion: PLGA NPs can prolong the existence of propranolol in both vitreous and posterior ocular tissues, and thus, may provide an effective approach in treatment of posterior segment neovascularization. [ABSTRACT FROM AUTHOR]

Abstract Inflammatory ocular diseases are characterized by the presence of a persistent inflammatory response which cause tissue injury, decrease visual acuity and in severe cases, blindness. Several cytokines represent a therapeutic opportunity since they are key amplifiers of these pathologies, and thus neutralizing agents against them have been developed. Amongst others, macrophage migration inhibitory factor (MIF), an early produced inflammatory cytokine, has consistently been found elevated in patients with distinct ocular diseases (inflammatory and autoimmune). Here, we present and discuss evidence showing that preclinical trials using diverse strategies to neutralize MIF resulted in significant attenuation of disease signs and therefore MIF blockage might be a promising therapy for ocular diseases.

Abnormal ocular neovascularization, a major pathology of eye diseases, leads to severe visual loss. The role of lens epithelial cell (LEC)‐derived exosomes (Lec‐exo) is largely unknown. Thus, we aimed to investigate whether Lec‐exo can inhibit abnormal ocular neovascularization and explore the possible mechanisms. In our study, we proved the first evidence that exosomes derived from LECs attenuated angiogenesis in both oxygen‐induced retinopathy and laser‐induced choroidal neovascularization mice models. Further in vitro experiments proved that Lec‐exo inhibited proliferation, migration, and tube formation capability of human umbilical vein endothelial cells in high glucose condition. Further high‐throughput miRNAs sequencing analysis detected that miR‐146a‐5p was enriched in Lec‐exo. Mechanistically, exosomal miR‐146a‐5p was delivered to endothelial cells and bound to the NRAS coding sequence, which subsequently inactivated AKT/ERK signaling pathway. We successfully elucidated the function of Lec‐exo in inhibiting abnormal ocular neovascularization, which may offer a promising strategy for treatment of abnormal ocular neovascularization. [ABSTRACT FROM AUTHOR]

Inflammatory ocular diseases are characterized by the presence of a persistent inflammatory response which cause tissue injury, decrease visual acuity and in severe cases, blindness. Several cytokines represent a therapeutic opportunity since they are key amplifiers of these pathologies, and thus neutralizing agents against them have been developed. Amongst others, macrophage migration inhibitory factor (MIF), an early produced inflammatory cytokine, has consistently been found elevated in patients with distinct ocular diseases (inflammatory and autoimmune). Here, we present and discuss evidence showing that preclinical trials using diverse strategies to neutralize MIF resulted in significant attenuation of disease signs and therefore MIF blockage might be a promising therapy for ocular diseases. [ABSTRACT FROM AUTHOR]

Abnormal neovascularization is an important cause of blindness in many ocular diseases, for which the etiology and pathogenic mechanisms remain incompletely understood. Recent studies have revealed the diverse roles of noncoding RNAs in various biological processes and facilitated the research and development of the clinical application of numerous RNA drugs, including microRNAs. Here, we report the antiangiogenic activity of microRNA-29a (miR-29a) in three animal models of ocular neovascularization. The miR-29a knockout (KO) mice displayed enhanced vessel pruning, resulting in a decreased vascularized area during retinal development. In contrast, miR-29a deletion in adult mice accelerated angiogenesis in preclinical disease models, including corneal neovascularization, oxygen-induced retinopathy, and choroidal neovascularization, while the administration of agomir-29a ameliorated pathological neovascularization. Furthermore, miR-29a exerted inhibitory effects on endothelial cell proliferation, migration, and tube formation capacities. RNA sequencing analysis of retinas from miR-29a KO mice and RNA interference experiments identified platelet derived growth factor C and several extracellular matrix genes as downstream targets of miR-29a involved in regulating ocular angiogenesis. Our data suggest that miR-29a may be a promising clinical candidate for the treatment of neovascular diseases. [ABSTRACT FROM AUTHOR]

Background: To report the incidence and risk factors associated with ocular neovascularization (NV) in patients with central retinal artery occlusion (CRAO). Methods: This retrospective study included patients diagnosed with acute CRAO in a single tertiary center. Medical charts were reviewed for ocular NV occurrences. We analyzed systemic and ocular conditions on first visit and demographic data. Results: Eighty-seven eyes were eligible for this study. Among these, 13 eyes had ocular NV after CRAO, with an incidence of 15%. The prevalences of hypertension, diabetes mellitus, history of stroke, chronic kidney disease (CKD), and age at first visit were higher among patients with ocular NV than among patients without ocular NV after CRAO. Moreover, most patients with CKD in the ocular NV group had undergone dialysis. A multivariate regression analysis revealed that CKD (hazard ratio [HR]: 9.27, 95% CI, 1.87-46.05, p = 0.006) and glaucoma history (HR: 7.52, 95% CI, 1.14-49.46, p = 0.036) were significant risk factors for developing ocular NV among patients with CRAO. Conclusion: CKD and glaucoma history were significant risk factors for developing ocular NV after CRAO, particularly among patients that underwent dialysis. [ABSTRACT FROM AUTHOR]

Purpose: To evaluate the therapeutic benefit of a novel peptide, ALM201, in ocular pathologic vascularization. Design: Experimental study in mouse, rat, and rabbit animal models. Participants: Ten-week-old Lister Hooded male rats, 8-week-old Brown Norway male rats, 9-day-old C57BL/6J mice, and 12-month-old New Zealand male rabbits. Methods: Corneal vascularization was scored for vessel density and vessel distance to suture in a rat corneal suture model. Ocular penetration and biodistribution were evaluated by matrix-assisted laser desorption/ionization mass spectrometry imaging after topical ALM201 application to rabbit eyes. A mouse choroidal sprouting assay, with aflibercept as positive control, was used to evaluate choroidal neovascularization (CNV) in the posterior segment tissue. Efficacy of topical ALM201 was assessed using a rat laser CNV model of neovascular age-related macular degeneration. Main Outcome Measures: Clinical scoring and histologic analysis of vascularized corneas, sprouting area, lesion size, and vessel leakiness in posterior segments. Results: Assessment of ALM201 treatment in the rat corneal suture model showed a significant decrease in vessel density (P = 0.0065) and vessel distance to suture (P = 0.021) compared with vehicle control (phosphate-buffered saline [PBS]). Infiltration of inflammatory cells into the corneal stroma also was reduced significantly compared with PBS (724.5 ± 122 cells/mm2 vs. 1837 ± 195.9 cells/mm2, respectively; P = 0.0029). Biodistribution in rabbit eyes confirmed ALM201 bioavailability in anterior and posterior ocular segments 1 hour after topical instillation. ALM201 treatment significantly suppressed choroid vessel sprouting when compared with PBS treatment (44.5 ± 14.31 pixels vs. 120.9 ± 33.37 pixels, respectively; P = 0.04) and was not inferior to aflibercept (65.63 ± 11.86 pixels; P = 0.7459). Furthermore, topical ALM201 significantly improved vessel leakiness (leakage scores: 2.1 ± 0.7 vs. 2.9 ± 0.1; P = 0.0274) and lesion size (144,729 ± 33,239 μm3 vs. 187,923 ± 28,575 μm3; P = 0.03) in the rat laser CNV model when compared with topical PBS vehicle. Conclusions: ALM201 is a promising novel molecule with anti-inflammatory and antivascularization activity and is a strong candidate to meet the clinical need of a new, topically delivered therapeutic agent for treating inflammation and pathologic vascularization in the anterior and posterior segments of the eye.

Ocular neovascularization often leads to severe vision loss. The role of bone marrow-derived cells (BMCs) in the development of ocular neovascularization, and its significance, is increasingly being recognized. In this review, we discuss their contribution and the potential mechanisms that mediate the effect of BMCs on the progression of ocular neovascularization. The sequence of events by which BMCs participate in ocular neovascularization can be roughly divided into four phases, i.e., mobilization, migration, adhesion and differentiation. This process is delicately regulated and liable to be affected by multiple factors. Cytokines such as vascular endothelial growth factor, granulocyte colony-stimulating factor and erythropoietin are involved in the mobilization of BMCs. Studies have also demonstrated a key role of cytokines such as stromal cell-derived factor-1, tumor necrosis factor-α, as well as vascular endothelial growth factor, in regulating the migration of BMCs. The adhesion of BMCs is mainly regulated by vascular cell adhesion molecule-1, intercellular adhesion molecule-1 and vascular endothelial cadherin. However, the mechanisms regulating the differentiation of BMCs are largely unknown at present. In addition, BMCs secrete cytokines that interact with the microenvironment of ocular neovascularization; their contribution to ocular neovascularization, especially choroidal neovascularization, can be aggravated by several risk factors. An extensive regulatory network is thought to modulate the role of BMCs in the development of ocular neovascularization. A comprehensive understanding of the involved mechanisms will help in the development of novel therapeutic strategies related to BMCs. In this review, we have limited the discussion to the recent progress in this field, especially the research conducted at our laboratory.

As a newly discovered endogenous regulator, fibroblast growth factor 21(FGF21)becomes hot topic in recent professional research. It plays an important role in diabetic retinopathy recently. Moreover, FGF21 has attracted more and more attention in recent years. This paper discussed the molecular structure of FGF 21, biological function, relationship between FGF21 and inflammatory response and role in the pathological process of diabetic retinopathy.

Ischemic and neovascular disease is one of the most difficult ocular diseases to deal with nowadays. Redundancy, poor visual acuity and decreased life quality are bothering patients and ophthalmologists for decades. After vascular endothelial growth factor (VEGF) was found to be a primary factor in promoting retinal angiogenesis, intravitreal injection of anti-VEGF drugs has been the first-line treatment. Whereas, some patients are refractory to this therapy and problems of economic burden, local complications and adverse effects promote researches into other possible targets. The vasohibin (VASH) family is a newly-investigated factor in modulating ocular angiogenesis. The family includes VASH1 and VASH2, which show opposite effects of inhibiting and accelerating angiogenesis respectively. Positive results have been reported in cellular and animal experiments. With further researches, it can be a promising future target of treating ocular neovascular diseases.

CRISPR-Cas13 nucleases are programmable RNA-targeting effectors that can silence gene expression in a transient manner. Recent iterations of Cas13 nucleases are compact for adeno-associated virus (AAV) delivery to achieve strong and persistent expression of various organs in a safe manner. Here, we report significant transcriptomic signatures of Cas13bt3 expression in retinal cells and show all-in-one AAV gene therapy with Cas13bt3 can effectively silence VEGFA mRNA in human retinal organoids and humanized VEGF transgenic mouse (trVEGF029, Kimba) models. Specifically, human embryonic stem cells (hESC)-derived retinal pigment epithelium cells show high expression of Cas13bt3 from virus delivery corresponding to a significant reduction of VEGFA mRNA. We further show that intravitreal delivery of Cas13bt3 by AAV2.7m8 can efficiently transduce mouse retinal cells for specific knockdown of human VEGFA in the Kimba mouse. Our results reveal important considerations for assessing Cas13 activity, and establish the Cas13bt3 RNA editing system as a potential anti-VEGF agent that can achieve significant control of VEGFA for the treatment of retinal neovascularization., Competing Interests: Competing interests statement:The authors declare no competing interest.

Purpose: To investigate the effects of a common dietary flavonoid apigenin on retinal endothelial cell proliferation, retinal morphological structure, and apoptotic cell death in an oxygen-induced retinopathy (OIR) mouse model to evaluate the possibility of the use of apigenin in the treatment of ocular neovascular diseases (ONDs). Methods: Ninety-six newborn C57BL/6J mice were included. Eight groups were randomized, each including 12 mice. Two negative control groups were kept in room air: the first without any injection and the second received intravitreal (IV) dimethyl sulfoxide (DMSO), which is the solvent we used. The OIR groups were exposed to 75% ± 2% oxygen from postnatal days (PD) 7 to 12. On PD 12, the mice were randomly assigned to 6 groups: 2 OIR control groups (1 received no injection, 1 received IV-DMSO), 2 IV-apigenin groups (10 and 20 μg/mL), and 2 intraperitoneal (IP)-apigenin groups (10 and 20 mg/kg). We quantified retinal endothelial cell proliferation by counting neovascular tufts in cross-sections and examined histological and ultrastructural changes through light and electron microscopy. We evaluated apoptosis by terminal deoxynucleotidyl transferase-mediated nick end-labeling (TUNEL). Results: We detected a significant increase in endothelial cell proliferation in the OIR groups. Groups receiving apigenin, both IP and IV, had significant decreases in endothelial cells, atypical mitochondrion count, and apoptotic cells compared with the groups receiving no injections. None of the apigenin-injected groups revealed cystic degeneration or cell loss. Conclusions: Apigenin suppresses neovascularization, has antiapoptotic and antioxidative effects in an OIR mouse model, and can be considered a promising agent for treating OND. Clinical trial (Project number: DA15/19). [ABSTRACT FROM AUTHOR]

Artemisinin, also named qinghaosu, is a family of sesquiterpene trioxane lactone originally derived from the sweet wormwood plant (Artemisia annua), which is a traditional Chinese herb that has been universally used as anti-malarial agents for many years. Evidence has accumulated during the past few years which demonstrated the protective effects of artemisinin and its derivatives (artemisinins) in several other diseases beyond malaria, including cancers, autoimmune disorders, inflammatory diseases, viral and other parasite-related infections. Recently, this long-considered anti-malarial agent has been proved to possess anti-oxidant, anti-inflammatory, anti-apoptotic and anti-excitotoxic properties, which make it a potential treatment option for the ocular environment. In this review, we first described the overview of artemisinins, highlighting the activity of artemisinins to other diseases beyond malaria and the mechanisms of these actions. We then emphasized the main points of published results of using artemisinins in targeting ocular disorders, including uveitis, retinoblastoma, retinal neurodegenerative diseases and ocular neovascularization. To conclude, we believe that artemisinins could also be used as a promising therapeutic drug for ocular diseases, especially retinal vascular diseases in the near future.

Abstract Currently, biological drug therapy for ocular angiogenesis treatment is based on the administration of anti‐VEGF agents via intravitreal route. The molecules approved with this purpose for ocular use include pegaptanib, ranibizumab, and aflibercept, whereas bevacizumab is commonly off‐label used in the clinical practice. The schedule dosage involves repeated intravitreal injections of anti‐VEGF agents to achieve and maintain effective concentrations in retina and choroids, which are administrated as solutions form. In this review article, we describe the features of different anti‐VEGF agents, major challenges for their ocular delivery and the nanoparticles in development as delivery system of them. In this way, several polymeric and lipid nanoparticles are explored to load anti‐VEGF agents with the aim of achieving sustained drug release and thus, minimize the number of intravitreal injections required. The main challenges were focused in the loading the molecules that maintain their bioactivity after their release from nanoparticulate system, followed the evaluation of them through studies of formulation stability, pharmacokinetic, and efficacy in in vitro and in vivo models. The analysis was based on the information published in peer‐reviewed published papers relevant to anti‐VEGF treatments and nanoparticles developed as ocular anti‐VEGF delivery system.

Currently, biological drug therapy for ocular angiogenesis treatment is based on the administration of anti-VEGF agents via intravitreal route. The molecules approved with this purpose for ocular use include pegaptanib, ranibizumab, and aflibercept, whereas bevacizumab is commonly off-label used in the clinical practice. The schedule dosage involves repeated intravitreal injections of anti-VEGF agents to achieve and maintain effective concentrations in retina and choroids, which are administrated as solutions form. In this review article, we describe the features of different anti-VEGF agents, major challenges for their ocular delivery and the nanoparticles in development as delivery system of them. In this way, several polymeric and lipid nanoparticles are explored to load anti-VEGF agents with the aim of achieving sustained drug release and thus, minimize the number of intravitreal injections required. The main challenges were focused in the loading the molecules that maintain their bioactivity after their release from nanoparticulate system, followed the evaluation of them through studies of formulation stability, pharmacokinetic, and efficacy in in vitro and in vivo models. The analysis was based on the information published in peer-reviewed published papers relevant to anti-VEGF treatments and nanoparticles developed as ocular antiVEGF delivery system. [ABSTRACT FROM AUTHOR]

The mitogen-inducible gene 6 (MIG6) is an adaptor protein widely expressed in vascular endothelial cells. However, it remains unknown thus far whether it plays a role in angiogenesis. Here, using comprehensive in vitro and in vivo model systems, we unveil a potent anti-angiogenic effect of MIG6 in retinal development and neovascularization and the underlying molecular and cellular mechanisms. Loss of function assays using genetic deletion of Mig6 or siRNA knockdown increased angiogenesis in vivo and in vitro, while MIG6 overexpression suppressed pathological angiogenesis. Moreover, we identified the cellular target of MIG6 by revealing its direct inhibitory effect on vascular endothelial cells (ECs). Mechanistically, we found that the anti-angiogenic effect of MIG6 is fulfilled by binding to SHC1 and inhibiting its phosphorylation. Indeed, SHC1 knockdown markedly diminished the effect of MIG6 on ECs. Thus, our findings show that MIG6 is a potent endogenous inhibitor of angiogenesis that may have therapeutic value in anti-angiogenic therapy.

Background: Ocular neovascularization is a hallmark of retinal diseases including neovascular age-related macular degeneration and diabetic retinopathy, two leading causes of blindness in adults. Neovascularization is driven by the interaction of soluble vascular endothelial growth factor (VEGF) ligands with transmembrane VEGF receptors (VEGFR), and inhibition of the VEGF pathway has shown tremendous clinical promise. However, anti-VEGF therapies require invasive intravitreal injections at frequent intervals and high doses, and many patients show incomplete responses to current drugs due to the lack of sustained VEGF signaling suppression. Methods: We synthesized insights from structural biology with molecular engineering technologies to engineer an anti-VEGF antagonist protein. Starting from the clinically approved decoy receptor protein aflibercept, we strategically designed a yeast-displayed mutagenic library of variants and isolated clones with superior VEGF affinity compared to the clinical drug. Our lead engineered protein was expressed in the choroidal space of rat eyes via nonviral gene delivery. Results: Using a structure-informed directed evolution approach, we identified multiple promising anti-VEGF antagonist proteins with improved target affinity. Improvements were primarily mediated through reduction in dissociation rate, and structurally significant convergent sequence mutations were identified. Nonviral gene transfer of our engineered antagonist protein demonstrated robust and durable expression in the choroid of treated rats one month post-injection. Conclusions: We engineered a novel anti-VEGF protein as a new weapon against retinal diseases and demonstrated safe and noninvasive ocular delivery in rats. Furthermore, our structure-guided design approach presents a general strategy for discovery of targeted protein drugs for a vast array of applications. [ABSTRACT FROM AUTHOR]

The insulin-like growth factor 1 (IGF-1) stimulates expression and secretion of vascular endothelial growth factor-A (VEGF-A), the main actor in ocular neovascularization, by RPE cells. Activity of IGF-1 is regulated by interaction between its receptor and Caveolin-1 (Cav-1), the main component of caveolae. The aim of this study was to investigate whether modulation of Cav-1 expression affects synthesis and secretion of VEGF-A. ARPE-19 cells were transfected with small interfering RNA for Cav-1 (si-Cav-1) and with control siRNA (si-CTR) and stimulated with IGF-1. We found that down-regulation of Cav-1 did not affect activation of IGF-1R but regulated in an opposite manner the phosphorylation of Akt and Erk1/2. Moreover, we found that IGF-1 increased mRNA levels of VEGF-A in both si-CTR and in si-Cav-1 ARPE-19 cells and that Cav-1 silencing significantly reduced basal and IGF-1-stimulated VEGF-A release. Then we investigated the response of the microvascular endothelial cell line HMEC-1 to secretory products of ARPE-19 cells by evaluating wound healing closure, finding that conditioned media from si-Cav-1-ARPE-19 cells reduced endothelial cell migration rate. These data demonstrate that Cav-1 regulates secretion of VEGF-A, and that the depletion of Cav-1 reduces IGF-1 induced VEGF-A secretion in ARPE-19 cells and the migratory potential of their secretory products.

Abstract Retinal and choroidal neovascularization are a major cause of significant visual impairment, worldwide. Understanding the various factors involved in the accompanying physiopathology is vital for development of novel treatments, and most important, for preserving patient vision. The intraocular use of anti-vascular endothelial growth factor therapeutics has improved management of the retinal and choroidal neovascularization but some patients do not respond, suggesting other vascular mediators may also contribute to ocular angiogenesis. Several recent studies examined possible new targets for future anti-angiogenic therapies. Potential targets of retinal and choroidal neovascularization therapy include members of the platelet-derived growth factor family, vascular endothelial growth factor sub-family, epidermal growth factor family, fibroblast growth factor family, transforming growth factor-β superfamily (TGF-β1, activins, follistatin and bone morphogenetic proteins), angiopoietin-like family, galectins family, integrin superfamily, as well as pigment epithelium derived factor, hepatocyte growth factor, angiopoietins, endothelins, hypoxia-inducible factors, insulin-like growth factors, cytokines, matrix metalloproteinases and their inhibitors and glycosylation proteins. This review highlights current antiangiogenic therapies under development, and discusses future retinal and choroidal pro- and anti-angiogenic targets as wells as the importance of developing of new drugs.

Neovascular age-related macular degeneration (nAMD) is a leading cause of vision loss in older adults. nAMD is treated with biologics targeting vascular endothelial growth factor; however, many patients do not respond to the current therapy. Here, a small molecule drug, griseofulvin (GRF), is used due to its inhibitory effect on ferrochelatase, an enzyme important for choroidal neovascularization (CNV). For local and sustained delivery to the eyes, GRF is encapsulated in microparticles based on poly(lactide-co-glycolide) (PLGA), a biodegradable polymer with a track record in long-acting formulations. The GRF-loaded PLGA microparticles (GRF MPs) are designed for intravitreal application, considering constraints in size, drug loading content, and drug release kinetics. Magnesium hydroxide is co-encapsulated to enable sustained GRF release over >30 days in phosphate-buffered saline with Tween 80. Incubated in cell culture medium over 30 days, the GRF MPs and the released drug show antiangiogenic effects in retinal endothelial cells. A single intravitreal injection of MPs containing 0.18 µg GRF releases the drug over 6 weeks in vivo to inhibit the progression of laser-induced CNV in mice with no abnormality in the fundus and retina. Intravitreally administered GRF MPs prove effective in preventing CNV, providing proof-of-concept toward a novel, cost-effective nAMD therapy., (© 2023 The Authors. Small published by Wiley-VCH GmbH.)

Objective · To study the effect of inhibitor of differentiation 1 (ID1) on ocular neovascularization. Methods · The oxygen-induced retinal neovascularization (OIR) , laser-induced choroidal neovascularization (CNV) and over-expression of vascular endothelial growth factor (VEGF) (Rho-VEGF) transgenic mice were established. The localization and mRNA level of ID1 in retina of OIR mice and Rho-VEGF transgenic mice were determined by immunofluorescence staining and quantitative real-time PCR. Mice deficient in ID1 (ID1-/-) were used to induce retinal neovascularization in accordance with the above three models, and to compare the changes of ID1 on the number of retinal, subretinal and choroidal neovascularization areas. In order to explore the role ID1 in neovascularization, the numbers and areas of retinal, subretinal and choroidal neovascularization in the mice models with or without ID1 deficiency were compared. Its effect on the related factors, i.e. hypoxia-inducible factor-1α (HIF-1α) , VEGF and vascular endothelial growth factor receptor 1/2 (VEGFR1/2) were also observed. Results · Mice deficient in ID1 showed a significant reduction in the area of neovascularization in these three models (P<0.05) . Mice lacking ID1 showed reduced levels of HIF-1α, VEGF and VEGFR 1. Conclusion · ID1 promotes the expression of HIF-1α, VEGF and VEGFR1 in the retina and choroidal neovascularization during hypoxia and oxidative injury. [ABSTRACT FROM AUTHOR]

Bevacizumab (BVZ), an anti-VEGF antibody, has demonstrated reliable outcomes in the treatment of irritating ocular neovascularization. Frequent intravitreal injections are necessitated due to rapid clearance and short local accessibility. We recruited liposome as a highly prevailing drug delivery system to enhance drug availability. Two liposome formulations were characterized and their in vitro stability was analyzed. The toxicity of the formulations on some ocular cell lines was also evaluated. In addition, the anti-angiogenic effects of formulations were examined. Drug permeation was measured across ARPE-19 and HCE cell lines as in vitro cellular barrier models. Results revealed that NLP-DOPE-BVZ acquired high stability at 4 °C, 24 °C, and 37 °C for 45 days. It also showed more capacity to entrap BVZ in NLP-DOPE-BVZ (DEE% 69.1 ± 1.4 and DLE% 55.66 ± 1.15) as compared to NLP-BVZ (DEE% 43.57 ± 14.64, and DLE% 37.72 ± 12.01). Although both formulations inhibited the migration and proliferation of HUVECs, NLP-DOPE-BVZ was more effective at inhibiting angiogenesis. Furthermore, NLP-DOPE-BVZ better crossed our established barrier cellular models. Based on the findings, the inclusion of DOPE in NLPs has significantly enhanced the features of drug carriers. This makes them a potential candidate for treating ocular neovascularization and other related ailments. [Display omitted] • Dipalmitoylphosphatidylcholine and Dioleoylphosphatidylethanolamine are safe. • Dioleoylphosphatidylethanolamine improves cell fusion and ocular barrier permeation. • Dioleoylphosphatidylethanolamine enhances stability of liposomes. • Dioleoylphosphatidylethanolamine improves bevacizumab loading in liposomes. • Dioleoylphosphatidylethanolamine increases ocular neovascularization inhibition. [ABSTRACT FROM AUTHOR]

Pathological vessel growth harms vision and may finally lead to vision loss. Anti-angiogenic gene therapy with viral vectors for ocular neovascularization has shown great promise in preclinical studies. Most of the studies have been conducted with different adeno-associated serotype vectors. In addition, adeno- and lentivirus vectors have been used. Therapy has been targeted towards blocking vascular endothelial growth factors or other pro-angiogenic factors. Clinical trials of intraocular gene therapy for neovascularization have shown the treatment to be safe without severe adverse events or systemic effects. Nevertheless, clinical studies have not proceeded further than Phase 2 trials.

Background/Aims: The degree of neovascularization determines the aggressiveness of ocular hemangiomas (OH). So far, the anti-angiogenic treatments using either antagonists against vascular endothelial growth factor A (VEGF-A), or endostatin, do not always lead to satisfactory therapeutic outcome. Methods: We examined the VEGF receptor 1 (VEGFR1) levels in the OH specimen. We compared the effects of anti-PLGF, endostatin, as well as their combined treatments on the growth of OH in a mouse model, using bioluminescence imaging in living animals. We also examined vascularization by CD31 expression. Results: We detected higher VEGFR1 levels in the OH, compared to paired normal tissue. Thus, we hypothesize that as a major ligand for VEGFR1, placental growth factor (PLGF) may also play a role in the neovascularization and tumorigenesis of OH. In an implanted OH model in mice, we found that both anti-PLGF and endostatin significantly decreased OH growth as well as vascularization, while combined treatments had a significantly more pronounced effect. Conclusion: Our data suggest that combined anti-PLGF and endostatin may be a more effective therapy for inhibition of ocular vascularization and the tumor growth in OH.

Angiogenesis is the biological process in which existing blood vessels generate new ones and it is essential for body growth and development, wound healing, and granulation tissue formation. Vascular endothelial growth factor receptor (VEGFR) is a crucial cell membrane receptor that binds to VEGF to regulate angiogenesis and maintenance. Dysregulation of VEGFR signaling can lead to several diseases, such as cancer and ocular neovascular disease, making it a crucial research area for disease treatment. Currently, anti-VEGF drugs commonly used in ophthalmology are mainly four macromolecular drugs, Bevacizumab, Ranibizumab, Conbercept and Aflibercept. Although these drugs are relatively effective in treating ocular neovascular diseases, their macromolecular properties, strong hydrophilicity, and poor blood-eye barrier penetration limit their efficacy. However, VEGFR small molecule inhibitors possess high cell permeability and selectivity, allowing them to traverse and bind to VEGF-A specifically. Consequently, they have a shorter duration of action on the target, and they offer significant therapeutic benefits to patients in the short term. Consequently, there is a need to develop small molecule inhibitors of VEGFR to target ocular neovascularization diseases. This review summarizes the recent developments in potential VEGFR small molecule inhibitors for the targeted treatment of ocular neovascularization diseases, with the aim of providing insights for future studies on VEGFR small molecule inhibitors. [Display omitted] • Abnormal expression of VEGFR is associated with the occurrence and progression of ocular neovascular diseases. • The effect of VEGFR small molecule inhibitors in the ocular neovascular diseases is "rapid, accurate and aggressive". • The SAR and pharmacological activities of VEGFR small molecule inhibitors in ocular neovascular diseases were discussed. [ABSTRACT FROM AUTHOR]

Epigalloccatechin-3-gallate (EGCG) is the main polyphenol component of green tea (leaves of Camellia sinensis). EGCG is known for its antioxidant, anti-inflammatory, antiviral, and anti-carcinogenic properties. Here, we identify EGCG as a new inhibitor of ocular angiogenesis and its vascular permeability. Matrix metalloproteinases (MMPs) and vascular endothelial growth factor (VEGF) play a key role in the processes of extracellular matrix (ECM) remodeling and microvascular permeability during angiogenesis. We investigated the inhibitory effects of EGCG on ocular neovascularization and vascular permeability using the retina oriented cells and animal models induced by VEGF and alkaline burn. EGCG treatment significantly decreased mRNA and protein expression levels of MMP-9 in the presence of 12-O-tetradecanoylphorbol-13-acetate (TPA) and tumor necrosis factor alpha (TNF-α) in human retinal pigment epithelial cells (HRPECs). EGCG also effectively protected ARPE-19 cells from cell death and attenuated mRNA expressions of key angiogenic factors (MMP-9, VEGF, VEGF Receptor-2) by inhibiting generation of reactive oxygen species (ROS). EGCG significantly inhibited proliferation, vascular permeability, and tube formation in VEGF-induced human retinal microvascular endothelial cells (HRMECs). Furthermore, EGCG significantly reduced vascular leakage and permeability by blood-retinal barrier breakdown in VEGF-induced animal models. In addition, EGCG effectively limited upregulation of MMP-9 and platelet endothelial cell adhesion molecule (PECAM/CD31) on corneal neovascularization (CNV) induced by alkaline burn. Our data suggest that MMP-9 and VEGF are key therapeutic targets of EGCG for treatment and prevention of ocular angiogenic diseases such as age-related macular degeneration, diabetic retinopathy, and corneal neovascularization.

Ocular neovascularization is a common pathological feature in diabetic retinopathy and neovascular age-related macular degeneration that can lead to severe vision loss. We evaluated the therapeutic efficacy of a novel endogenous inhibitor of angiogenesis, the calreticulin anti-angiogenic domain (CAD180), and its functional 112-residue fragment, CAD-like peptide 112 (CAD112), delivered using a self-complementary adeno-associated virus serotype 2 (scAAV2) in rodent models of oxygen-induced retinopathy and laser-induced choroidal neovascularization. The expression of CAD180 and CAD112 was elevated in human umbilical vein endothelial cells transduced with scAAV2-CAD180 or scAAV2-CAD112, respectively, and both inhibited angiogenic activity in vitro. Intravitreal gene delivery of scAAV2-CAD180 or scAAV2-CAD112 significantly inhibited ischemia-induced retinal neovascularization in rat eyes (CAD180: 52.7% reduction; CAD112: 49.2% reduction) compared to scAAV2-mCherry, as measured in retinal flatmounts stained with isolectin B4. Moreover, the retinal structure and function were unaffected by scAAV2-CAD180 or scAAV2-CAD112, as measured by optical coherence tomography and electroretinography. Moreover, subretinal delivery of scAAV2-CAD180 or scAAV2-CAD112 significantly attenuated laser-induced choroidal neovascularization in mouse eyes compared to scAAV2-mCherry, as measured by fundus fluorescein angiography (CAD180: 62.4% reduction; CAD112: 57.5% reduction) and choroidal flatmounts (CAD180: 40.21% reduction; CAD112: 43.03% reduction). Gene delivery using scAAV2-CAD180 or scAAV2-CAD112 has significant potential as a therapeutic option for the management of ocular neovascularization. [ABSTRACT FROM AUTHOR]

Ocular neovascularization is a devastating pathology of numerous ocular diseases and is a major cause of blindness. Caveolin-1 (Cav-1) plays important roles in the vascular system. However, little is known regarding its function and mechanisms in ocular neovascularization. Here, using comprehensive model systems and a cell permeable peptide of Cav-1, cavtratin, we show that Cav-1 is a critical player in ocular neovascularization. The genetic deletion of Cav-1 exacerbated and cavtratin administration inhibited choroidal and retinal neovascularization. Importantly, combined administration of cavtratin and anti-VEGF-A inhibited neovascularization more effectively than monotherapy, suggesting the existence of other pathways inhibited by cavtratin in addition to VEGF-A. Indeed, we found that cavtratin suppressed multiple critical components of pathological angiogenesis, including inflammation, permeability, PDGF-B and endothelial nitric oxide synthase expression (eNOS). Mechanistically, we show that cavtratin inhibits CNV and the survival and migration of microglia and macrophages via JNK. Together, our data demonstrate the unique advantages of cavtratin in antiangiogenic therapy to treat neovascular diseases. [ABSTRACT FROM AUTHOR]

Purpose: To determine the time-based incidence of total blindness after central retinal artery occlusion (CRAO) with secondary ocular neovascularization (ONV). Methods: In this retrospective cohort study, electronic records were queried using ICD-9 and ICD-10 codes to identify patients with secondary ONV post-CRAO. Patients with possible alternative ONV etiologies, previous panretinal photocoagulation (PRP), and/or previous antivascular endothelial growth factor (anti-VEGF) therapy were excluded. Clinical data included demographics, medical comorbidities, ONV manifestations, medical/surgical management, and best-corrected visual acuity (BCVA). Kaplan-Meier analysis was performed with total blindness (defined as a BCVA of no light perception) as the outcome of interest. Results: Of 345 eyes with CRAO, 34 met the inclusion criteria with a mean (±SD) follow-up of 22.0 ± 26.2 months. ONV management included PRP (70.6%), glaucoma drainage implant surgery or transscleral cyclophotocoagulation (32.4%), and intravitreal anti-VEGF therapy (mean 2.8 ± 5.6 injections per patient). The cumulative incidence of total blindness was 49.4% (95% confidence interval, 27.2%-71.6%) at 24 months, with 53.3% of cases occurring within 4 months of ONV onset. Conclusions: Post-CRAO ONV is associated with a high risk for progression from severe vision loss to total blindness. Neovascular glaucoma can present up to 4 months after CRAO, challenging the paradigm of "30-day-glaucoma." Routine gonioscopy should extend through this period, while glaucoma surgery can delay further vision loss. These findings can be used to counsel patients on the importance of follow-up adherence., Competing Interests: The author(s) declared the following potential conflicts of interest with respect to the research, authorship, and/or publication of this article: Dr. Mazzulla is a consultant to Allergan PLC, Alimera Sciences Inc, and Apellis Pharmaceuticals unrelated to this study. None of the other authors reported financial disclosures related or unrelated to this study., (© The Author(s) 2023.)

Aberrant activation of the hypoxia inducible factor (HIF) pathway causing overexpression of angiogenic genes, like vascular endothelial growth factor (VEGF), is one of the underlying causes of ocular neovascularization (NV) and metastatic cancer. Consistently, along with surgical interventions, a number of anti-VEGF agents have been approved by FDA for the treatment of ocular neovascular diseases. These anti-VEGF agents, like ranibizumab/lucentis, have revolutionized the treatment in the past decade. However, substantial vision improvement is observed only in a subset of age-related macular degeneration patients receiving ranibizumab. Further, all current therapies are associated with limitations and side effects. For example, surgeries cause tissue destruction and inflammation while anti-VEGF therapies are expensive, require repeated administration, and offer temporary relief from vascular leakage. These factors impose significant cost and treatment burdens to both the patient and society. With an aging population in most western countries with a continually increasing number of patients on lifelong treatment for these retinal diseases, the focus of ocular drug development for neovascular diseases will be to improve efficacy while reducing treatment costs. Blocking the HIF pathway, a major regulator of ocular NV and cancer, offers an appealing therapeutic strategy. Therefore, this review summarizes HIF inhibitors that have been recently evaluated for the treatment of different cancers and ischemic retinopathies. [ABSTRACT FROM AUTHOR]

Interleukin-23 (IL-23) is a heterodimeric cytokine that consists of p19, a novel subunit, and p40, which is shared by IL-12. IL-23 has been demonstrated to play an important role in autoimmunity and tumor growth. However, the role of IL-23 in ocular neovascularization (NV) diseases remains unclear. In this study, we explored the role of IL-23 in the processing of retinal and choroidal neovascularization (RNV and CNV). We found a significantly higher expression of IL-23 in the retinas with oxygen-induced retinopathy (OIR), and after neutralizing IL-23, the mRNA and protein levels of the angiogenic factors vascular endothelial growth factor receptor (VEGFR)1/FLT-1, VEGFR2/FLK-1, placental growth factor (PIGF), endothelial-specific receptor tyrosine kinase (Tie2), inducible nitric-oxide synthase (iNOS), matrix metalloproteinase (MMP) 2 and MMP9 were significantly down regulated, while the opposite trend was found for the anti-angiogenic molecules chemokine (C-X-C motif) ligand (CXCL) 9 and CXCL10. IL-23 blockade caused less NV in both the RNV and CNV mouse models. In addition, our in vitro assay showed that IL-23 alone is able to increase the ability of endothelial cells to form tubes. Our findings suggest that targeting IL-23 could be a potential therapy for RNV and CNV diseases. [ABSTRACT FROM AUTHOR]

Neovascularization (NV), as a cardinal complication of several ocular diseases, has been intensively studied, and research has shown its close association with inflammation and immune cells. In the present study, the role of interleukin-17A (IL-17A) in angiogenesis in the process of ocular NV both in vivo and in vitro was investigated. Also, a paracrine role of IL-17A was demonstrated in the crosstalk between endothelial cells and macrophages in angiogenesis. In the retinas of mice with retinopathy of prematurity, the IL-17A expression increased significantly at postnatal day 15 (P15) and P18 during retinal NV. Mice given IL-17A neutralizing antibody (NAb) developed significantly reduced choroidal NV and retinal NV. Studies on vascular endothelial growth factor (VEGF) over-expressing mice suggested that IL-17A modulated NV through the VEGF pathway. Furthermore, IL-17A deficiency shifted macrophage polarization toward an M2 phenotype during retinal NV with significantly reduced M1 cytokine expression compared with wild-type controls. In vitro assays revealed that IL-17A treated macrophage supernatant gave rise to elevated human umbilical vascular endothelial cell proliferation, tube formation and VEGF receptor 1 and receptor 2 expression. Therefore, IL-17A could potentially serve as a novel target for treating ocular NV diseases. The limitation of this study involved the potential mechanisms, such as which transcription accounted for macrophage polarization and how the subsequent cytokines were modulated when macrophages were polarized. Further studies need to be undertaken to definitively determine the extent to which IL-17A neutralizing anti-angiogenic activity depends on macrophage modulation compared with anti-VEGF treatment. [ABSTRACT FROM AUTHOR]

Pathologic neovascularization of the retina is a major cause of substantial and irreversible loss of vision. Drugs are difficult to deliver to the lesions in the back of the eye and this is a major obstacle for the therapeutics. Current pharmacological approach involves an intravitreal injection of anti-VEGF agents to prevent aberrant growth of blood vessels, but it has limitations including therapeutic efficacy and side-effects associated with systemic exposure and invasive surgery. Nanotechnology provides novel opportunities to overcome the limitations of conventional delivery system to reach the back of the eye through fabrication of nanostructures capable of encapsulating and delivering small molecules. This review article introduces various forms of nanocarrier that can be adopted by ocular drug delivery systems to improve current therapy. The application of nanotechnology in medicine brings new hope for ocular drug delivery in the back of the eye to manage the major causes of blindness associated with ocular neovascularization. [ABSTRACT FROM AUTHOR]

Background/Aims: The degree of neovascularization determines the aggressiveness of ocular hemangiomas (OH). So far, the anti-angiogenic treatments using either antagonists against vascular endothelial growth factor A (VEGF-A), or endostatin, do not always lead to satisfactory therapeutic outcome. Methods: We examined the VEGF receptor 1 (VEGFR1) levels in the OH specimen. We compared the effects of anti-PLGF, endostatin, as well as their combined treatments on the growth of OH in a mouse model, using bioluminescence imaging in living animals. We also examined vascularization by CD31 expression. Results: We detected higher VEGFR1 levels in the OH, compared to paired normal tissue. Thus, we hypothesize that as a major ligand for VEGFR1, placental growth factor (PLGF) may also play a role in the neovascularization and tumorigenesis of OH. In an implanted OH model in mice, we found that both anti-PLGF and endostatin significantly decreased OH growth as well as vascularization, while combined treatments had a significantly more pronounced effect. Conclusion: Our data suggest that combined anti-PLGF and endostatin may be a more effective therapy for inhibition of ocular vascularization and the tumor growth in OH. [ABSTRACT FROM AUTHOR]

Purpose To verify the effect of vasostatin-1 ( VS-1), an anti-angiogenic fragment of chromogranin A, in the prevention of choroidal neovascularization ( CNV) in an established mouse model of laser-induced ocular neovascularization. Methods Bruch's membrane, the innermost layer of the choroid, was broken by laser photocoagulation in C57/Bl6 mice, to induce CNV. Mice were then treated daily for 14 days by intraperitoneal injection of VS-1 or vehicle (6 mice/group). CNV and vascular leakage were measured at three time-points (day 0, 7 and 14) in vivo by spectral domain optical coherence tomography ( OCT) and fluorescein angiography ( FA). Ex vivo analysis of CNV was also performed at day 14 by confocal microscopy analysis of dextran-perfused choroidal flat-mounts. Results In vivo analyses showed that VS-1 significantly reduced CNV at day 14 (p = 0.03) and vascular leakage at day 7 (p = 0.01) and 14 (p = 0.04). Ex vivo confocal microscopy analysis of CNV performed on dextran-perfused choroidal flat-mounts at day 14 confirmed the protective activity of VS-1 (p = 0.01). A significant correlation between the results of in vivo and ex vivo analyses of CNV was also observed (p = 0.001, R2 = 0.81). Conclusion The results indicate that VS-1 can prevent CNV and vascular leakage in a mouse model of ocular neovascularization, suggesting that this polypeptide might have therapeutic activity in human ocular diseases that are complicated by neovascularization or excessive vascular permeability. [ABSTRACT FROM AUTHOR]

Pathological angiogenesis related to neovascularization in the eye is mediated through vascular endothelial growth factors (VEGFs) and their receptors. Ocular neovascular-related diseases are mainly treated with anti-VEGF agents. In this study we evaluated the efficacy and safety of novel gene therapy using adeno associated virus 2 vector expressing a truncated form of soluble VEGF receptor-2 fused to the Fc-part of human IgG1 (AAV2-sVEGFR-2-Fc) to inhibit ocular neovascularization in laser induced choroidal neovascularization (CNV) in mice. The biological activity of sVEGFR-2-Fc was determined in vitro. It was shown that sVEGFR-2-Fc secreted from ARPE-19 cells was able to bind to VEGF-A 165 and reduce VEGF-A 165 induced cell growth and survival. A single intravitreal injection (IVT) of AAV2-sVEGFR-2-Fc (1 μl, 4.7 × 1012 vg/ml) one-month prior laser photocoagulation did not cause any changes in the retinal morphology and significantly suppressed fluorescein leakage at 7, 14, 21 and 28 days post-lasering compared to controls. Macrophage infiltration was observed after the injection of both AAV2-sVEGFR-2-Fc and PBS. Our findings indicate that AAV2 mediated gene delivery of the sVEGFR-2-Fc efficiently reduces formation of CNV and could be developed to a therapeutic tool for the treatment of retinal diseases associated with neovascularization. [Display omitted] • Novel antiangiogenic gene therapy for the treatment of neovascular associated retinal diseases. • AAV based fusion protein which combines the domains D1-D3 from human VEGFR-2. • Long-term suppression of laser induced choroidal neovascularization in mice after intravitreal injection. [ABSTRACT FROM AUTHOR]

Pathological neovascularization are the most prevalent causes of moderate or severe vision loss. Long non-coding RNAs (lncRNAs) have emerged as a novel class of regulatory molecules involved in numerous biological processes and complicated diseases. However, the role of lncRNAs in ocular neovascularization is still unclear. Here, we constructed a murine model of ocular neovascularization, and determined lncRNA expression profiles using microarray analysis. We identified 326 or 51 lncRNAs that were significantly either up-regulated or down-regulated in the vaso-obliteration or neovascularization phase, respectively. Based on Pearson correlation analysis, lncRNAs/mRNAs co-expression networks were constructed. GO enrichment analysis of lncRNAs-co-expressed mRNAs indicated that the biological modules were correlated with chromosome organization, extracellular region and guanylate cyclase activator activity in the vaso-obliteration phase, and correlated with cell proliferation, extracellular region and guanylate cyclase regulator activity in the neovascularization phase. KEGG pathway analysis indicated that MAPK signaling was the most significantly enriched pathway in both phases. Importantly, Vax2os1 and Vax2os2 were not only dynamically expressed in the vaso-obliteration and neovascularization phases, but also significantly altered in the aqueous humor of patients with neovascular age-related macular degeneration (AMD), suggesting a potential role of lncRNAs in the regulation of ocular neovascularization. Taken together, this study provided novel insights into the molecular pathogenesis of ocular neovascularization. The intervention of dysregulated lncRNA could become a potential target for the prevention and treatment of ocular vascular diseases. [ABSTRACT FROM AUTHOR]