Faricimab for neovascular age-related macular degeneration and diabetic macular edema: from preclinical studies to phase 3 outcomes.

Intravitreal anti-vascular endothelial growth factor (VEGF) therapy is the standard of care for diabetic macular edema (DME) and neovascular age-related macular degeneration (nAMD); however, vision gains and anatomical improvements are not sustained over longer periods of treatment, suggesting other relevant targets may be needed to optimize treatments. Additionally, frequent intravitreal injections can prove a burden for patients and caregivers. Angiopoietin-2 (Ang-2) has been explored as an additional therapeutic target, due to the involvement of Ang-2 in DME and nAMD pathogenesis. Recent evidence supports the hypothesis that targeting both VEGF and Ang-2 may improve clinical outcomes in DME and nAMD compared with targeting VEGF alone by enhancing vascular stability, resulting in reduced macular leakage, prevention of neovascularization, and diminished inflammation. Faricimab, a novel bispecific antibody that targets VEGF-A and Ang-2, has been evaluated in clinical trials for DME (YOSEMITE/RHINE) and nAMD (TENAYA/LUCERNE). These trials evaluated faricimab against the anti-VEGFA/B and anti-placental growth factor fusion protein aflibercept, both administered by intravitreal injection. In addition to faricimab efficacy, safety, and pharmacokinetics, durability was evaluated during the trials using a treat-and-extend regimen. At 1 year, faricimab demonstrated non-inferior vision gains versus aflibercept across YOSEMITE/RHINE and TENAYA/LUCERNE. In YOSEMITE/RHINE, faricimab improved anatomic parameters versus aflibercept. Reduction of central subfield thickness (CST), and absence of both DME and intraretinal fluid were greater in faricimab- versus aflibercept-treated eyes. In TENAYA/LUCERNE, CST reductions were greater for faricimab than aflibercept at the end of the head-to-head phase (0-12 weeks), and were comparable with aflibercept at year 1, but with less frequent dosing. CST and vision gains were maintained during year 2 of both YOSEMITE/RHINE and TENAYA/LUCERNE. These findings suggest that dual Ang-2/VEGF-A pathway inhibition may result in greater disease control versus anti-VEGF alone, potentially addressing the unmet needs and reducing treatment burden, and improving real-world outcomes and compliance in retinal vascular diseases. Long-term extension studies (RHONE-X, AVONELLE-X) are ongoing. Current evidence suggests that dual inhibition with faricimab heralds the beginning of multitargeted treatment strategies inhibiting multiple, independent components of retinal pathology, with faricimab providing opportunities to reduce treatment burden and improve outcomes compared with anti-VEGF monotherapy.

Inhibiting the NLRP3 inflammasome with MCC950 ameliorates retinal neovascularization and leakage by reversing the IL-1ß/IL-18 activation pattern in an oxygen-induced ischemic retinopathy mouse model.

Activation of the nucleotide-binding domain leucine-rich repeat and pyrin domain containing receptor 3 (NLRP3) inflammasome plays an important role in ocular neovascularization. In our study, we found that the expression and activation levels of NLRP3 inflammasome components, including NLRP3, an apoptosis-associated speck-like protein (ASC) containing caspase activation and recruitment domain (CARD) and caspase-1 (CAS1), were significantly upregulated. In addition, we found interleukin (IL)-1ß activity increased while IL-18 activity decreased in the retinas of oxygen-induced ischemic retinopathy (OIR) mice. MCC950, an inhibitor of NLRP3, reversed the IL-1ß/IL-18 activation pattern, inhibited the formation of retinal neovascularization (RNV), decreased the number of acellular capillaries and reduced leakage of retinal vessels. Moreover, MCC950 could regulate the expression of endothelial cell- and pericyte function-associated molecules, such as vascular endothelial growth factor (VEGF), VEGF receptor (VEGFR)1, VEGFR2, matrix metalloproteinase (MMP)2, MMP9, tissue inhibitor of metalloproteinases (TIMP)1, TIMP2, platelet-derived growth factor receptor-ß (PDGFR-ß), platelet-derived growth factor-B (PDGF-B), and angiopoietin2 (Ang2). In vitro, recombinant human (r)IL-18 and rIL-1ß regulated the expression of endothelial cell- and pericyte function-associated molecules and the proliferation and migration of endothelial cells and pericytes. We therefore determined that inhibiting the NLRP3 inflammasome with MCC950 can regulate the function of endothelial cells and pericytes by reversing the IL-1ß/IL-18 activation pattern to ameliorate RNV and leakage; thereby opening new avenues to treat RNV-associated ocular diseases.

Inhibiting NF-κB Signaling Activation Reduces Retinal Neovascularization by Promoting a Polarization Shift in Macrophages.

Purpose: Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling is involved in regulating tumor angiogenesis and metastasis; however, the exact mechanism of action in retinal neovascularization (RNV) remains unclear. The purpose of this study was to determine the role and underlying mechanism of NF-κB in regulating RNV in retinal neovascularization mice. Methods: Expression levels of NF-κB signaling were detected by immunofluorescence staining and western blotting in retinas of oxygen-induced retinopathy (OIR) mice. OIR mice were treated with either pyrrolidinedithiocarbamate (PDTC), a NF-κB signaling inhibitor, or PBS, and retinal flat-mounts were performed to quantify the area of RNV and the recruitment of retinal macrophages by immunofluorescence staining. Macrophage polarization detected by flow cytometric analysis and the expression of macrophage polarization-associated genes were evaluated by immunofluorescence staining, quantitative RT-PCR, and western blotting. Results: Expression levels of phosphorylated IκBα (p-IκBα) and p-p65 increased in OIR mice. Inhibiting NF-κB signaling activation by PDTC significantly reduced RNV. After treatment with PDTC, a reduction in the quantity of macrophages was observed: M1 polarized macrophages decreased, and M2 polarized macrophages increased; the expression of M1 macrophage-associated cytokines decreased and M2 macrophage-associated cytokines increased in the retinas of OIR mice. Conclusions: Blocking activation of NF-κB signaling reduces RNV by promoting polarization of M1 macrophages to M2 macrophages in OIR mice.

Neutralization of Bombina variegata peptide 8 suppresses retinal neovascularization in two different murine models: The oxygen-induced retinopathy model and the rhodopsin promoter/VEGF transgenic mouse model.

Bombina variegata 8 (Bv8), also known as prokineticin-2 (PK-2), is a potent pro-angiogenic factor. However, its role in retinal neovascularization (RNV) remains unknown. In this study, we explored the role of Bv8 in the pathogenesis of RNV. We found that the expression of Bv8 was significantly increased in two different models of retinal neovascularization: the oxygen-induced retinopathy (OIR) mouse model and the rhodopsin promoter (rho)/VEGF transgenic mouse model. Neutralization of Bv8 by intravitreal injections of its antibody, not only inhibited retinal and subretinal neovascularization but also decreased the mRNA and protein levels of several pro-angiogenic factors. Our in vitro assay showed that recombinant human Bv8 (RhBv8) protein promoted human retinal microvascular endothelial cells (HRECs) tube-formation, cell proliferation and vascular endothelial growth factor receptor 1 (VEGFR1) and receptor 2 (VEGFR2) expression. Our findings suggest that Bv8 could be used as a novel target for the treatment of RNV-related ocular diseases.

Retinal nerve fiber layer changes based on historic CD4 nadir among HIV positive patients undergoing glaucoma evaluation.

AIM: To determine relationships between retinal nerve fiber layer (RNFL) thickness and nadir CD4 cell count in human immunodeficiency virus (HIV) positive patients evaluated for glaucoma suspicion. METHODS: Data were reviewed for 329 HIV positive patients evaluated for glaucoma suspicion. High-definition optical coherence tomography (OCT) RNFL measurements were obtained at least 6mo apart. Analyses were performed to identify relationships between nadir CD4 count and RNFL thickness. RESULTS: Totally 110 eyes of 55 patients met inclusion criteria, of which 46 eyes were from subjects with nadir CD4<200 cells/mm3 and 64 had nadir CD4≥200 cells/mm3. Patients with nadir CD4<200 cells/mm3 had significantly thicker superior (119.7±18.6 µm) and temporal (63.8±11.7 µm) quadrants at time of initial OCT compared to the superior (112.8±16.8 µm, P=0.048) and temporal (57.1±11.9 µm, P=0.004) quadrants of patients with higher nadir CD4. This trend toward thicker RNFL among subjects with lower nadir CD4 cell counts persisted at the time of follow up OCT where participants with nadir CD4<200 cells/mm3 showed average RNFL thickness in the superior and temporal quadrants of 117.9±18.3 µm and 63.8±12.8 µm, respectively, compared to a superior thickness of 110.5±16.9 µm (P=0.034) and temporal thickness of 57.3±11.6 µm (P=0.007) among those with higher nadir CD4. CONCLUSION: Patients with lower nadir CD4 cell counts have thicker RNFL in the superior and temporal quadrants compared to those with higher nadir CD4 counts. RNFL thickness in HIV positive patients may be affected by historic HIV disease control and should be considered when evaluating HIV positive patients for glaucoma.

ATN-161 as an Integrin α5ß1 Antagonist Depresses Ocular Neovascularization by Promoting New Vascular Endothelial Cell Apoptosis.

BACKGROUND ATN-161 (Ac-PHSCN-NH2), an antagonist of integrin α5ß1, has shown an important influence in inhibiting tumor angiogenesis and metastasis of other tumor types. However, the mechanism of action of ATN-161 and whether it can inhibit ocular neovascularization (NV) are unclear. This study investigated the role of ATN-161 in regulating ocular angiogenesis in mouse models and explored the underlying signaling pathway. MATERIAL AND METHODS An oxygen-induced retinopathy (OIR) mouse model and a laser-induced choroidal neovascularization (CNV) mouse model were used to test integrin a5b1 expression and the effect of ATN-161 on ocular NV by immunofluorescence staining, Western blot analysis, and flat-mount analysis. The activation of nuclear factor-κB (NF-κB), matrix metalloproteinase-2/9 (MMP-2/9), and cell apoptosis were detected by immunofluorescence staining, Western blot, real-time RT-PCR, and terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL). The cell proliferation was detected by BrdU labeling. RESULTS In OIR and CNV mice, the protein expression level of integrin α5ß1 increased compared with that in age-matched controls. The mice given ATN-161 had significantly reduced retinal neovascularization (RNV) and CNV. Blocking integrin a5b1 by ATN-161 strongly inhibited nuclear factor-κB (NF-κB) activation and matrix metalloproteinase-2/9 (MMP-2/9) expression and promoted cell apoptosis, but the effect of ATN-161 on proliferation in CNV mice was indirect and required the inhibition of neovascularization. Inhibiting NF-κB activation by ammonium pyrrolidinedithiocarbamate (PDTC) reduced RNV and promoted cell apoptosis in ocular NV. CONCLUSIONS Blocking integrin α5ß1 by ATN-161 reduced ocular NV by inhibiting MMP-2/MMP-9 expression and promoting the cell apoptosis of ocular NV.

Flt3 Regulation in the Mononuclear Phagocyte System Promotes Ocular Neovascularization.

Fms-like tyrosine kinase 3 (Flt3), a tyrosine kinase receptor expressed in CD34+ hematopoietic stem/progenitor cells, is important for both normal myeloid and lymphoid differentiation. It has been implicated in mice and humans for potential multilineage differentiation. We found that mice deficient in Flt3 or mice that received an Flt3 inhibitor (AC220) showed significantly reduced areas of ischemia-induced retinal neovascularization (RNV) and laser-induced choroidal NV (CNV) (P < 0.05). Increased Flt3 expression at the protein level was detected in retinas of oxygen-induced retinopathy (OIR) mice at P15 and P18 during retinal NV (RNV) progression. We subsequently found that macrophages (Mphi) polarization was regulated at the site of CNV in Flt3-deficient mice. Flow cytometry analysis demonstrated that Flt3 deficiency shifted Mphi polarization towards an M2 phenotype during RNV with significant reduction in M1 cytokine expression when compared to the wild-type controls (P < 0.05). Based on the above findings, we concluded that Flt3 inhibition alleviated ocular NV by promoting a Mphi polarization shift towards the M2 phenotype. Therapies targeting Flt3 may provide a new approach for the treatment of ocular NV.

Inhibition of integrin α5ß1 ameliorates VEGF-induced retinal neovascularization and leakage by suppressing NLRP3 inflammasome signaling in a mouse model.

PURPOSE: To assess the effect of inhibiting integrin α5ß1 by ATN-161 on vascular endothelial growth factor (VEGF)-induced neovascularization (NV) and leakage causing retinal detachment in adult Tet/opsin/VEGF transgenic mice, and characterize the underlying mechanism of its function. METHOD: Retinas from adult Tet/opsin/VEGF transgenic mice and human retinal endothelial cells (HRECs) exposed to VEGF (treated with ATN-161 or PBS) were used to carry out immunofluorescence, RT-PCR and western blot to examine expression levels of integrin α5ß1 and the NACHT, LRR, and PYD domains-containing protein 3 (NLRP3) inflammasome. Retinal frozen section analysis was used to assess NV and leakage causing retinal detachment. RESULTS: In comparison to normal-treated mice, doxycycline-treated Tet/opsin/VEGF transgenic mice showed severe retinal detachment and higher integrin α5ß1 expression. Furthermore, the retinal detachment was inhibited significantly by ATN-161. Additionally, ATN-161 treatment was associated with a conspicuous reduction in NLRP3, apoptosis-associated speck-like protein containing a CARD (ASC), cleaved caspase-1, and mature interleukin-1ß expression levels in the retinas of Tet/opsin/VEGF transgenic mice treated with doxycycline as well as in HRECs exposed to VEGF. CONCLUSION: ATN-161, an antagonist of integrin α5ß1, is a promising treatment for retinal neovascularization (RNV), and its retinal protection role appears to take effect through inhibition of NLRP3 inflammasome activity.

Endocan Blockade Suppresses Experimental Ocular Neovascularization in Mice.

Purpose: Ocular neovascularization (NV) is a pathologic process characterized by the proliferation and infiltration of various types of cells such as RPE, glial, and endothelial cells, which interact with proangiogenic factors and inflammatory cytokines. Endocan is known to be enriched in retinal endothelial tip cells under hypoxia, but the effect of endocan on ocular NV progression is largely unknown. In this study, we investigated the role of endocan in the ocular NV pathologic process and the possible mechanisms involved. Methods: In the eyes of mice with oxygen-induced retinopathy (OIR); choroidal NV (CNV); and rhodopsin promoter (rho)/VEGF transgenic mice, endocan expression was assessed by quantitative real-time PCR (RT-PCR) and Western blot. In vivo, a specific functional antibody was used to neutralize endocan and ocular NV levels were evaluated by RT-PCR, Western blot and immunostaining of flat-mounts. In vitro, the effect of endocan on human retinal microvascular endothelial cell (HREC) tube formation was observed using a routine method. Results: Endocan was significantly elevated in these three experimental mice models. Endocan blockade with the neutralizer intravitreal injection not only suppressed the area of retinal, choroidal and subretinal NV, but also resulted in a decrease in several angiogenesis-associated molecules. Recombinant endocan protein (rhEndocan) was found to induce tube formation on HRECs directly. Conclusions: The current data suggest that endocan is a potential therapeutic or an additional target for retinal and subretinal NV diseases.

Interleukin-17A neutralization alleviated ocular neovascularization by promoting M2 and mitigating M1 macrophage polarization.

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.

One Year of Glaucoma Research in Review-2013 to 2014.

PURPOSE: The purpose of this study was to provide the practicing clinical ophthalmologist with an update on relevant glaucoma literature published from 2013 to 2014. DESIGN: This study is a literature review. METHODS: The authors conducted a 1-year (October 1, 2013, to September 30, 2014) English-language glaucoma literature search on PubMed of articles containing "glaucoma" or "glaucomatous" with title/abstract as a filter. Medical subject headings filtered searching was not performed because of the newness of the reviewed material. RESULTS: Literature search yielded 2314 articles, after which we excluded reviews and letters to the editor. We highlighted articles featuring new or updated approaches to the pathophysiology, diagnosis, or treatment of glaucoma and gave preference to human research. CONCLUSIONS: This review features literature that is of interest to ophthalmologists in practice and also highlights studies that may provide insight on future developments applicable to clinical ophthalmology.

Structural glaucomatous progression before and after occurrence of an optic disc haemorrhage.

BACKGROUND: An optic disc haemorrhage (DH) has been associated with subsequent structural glaucoma progression, but it is unknown if there is structural progression prior to a DH. We evaluated a cohort of patients to determine whether structural progression occurs before a DH, after a DH or is simply associated with a DH. METHODS: Eyes meeting inclusion criteria were placed into two groups. Group 1 included eyes that each had a baseline photograph of the optic nerve and a photograph with a DH at follow-up. Group 2 included eyes that each had a photograph of the optic nerve with a DH at baseline and a photograph at follow-up. Flicker images were created and graded by two ophthalmologists for structural glaucomatous change. We compared the proportion of structural progressors between Groups 1 and 2. Patient characteristics were also compared between the two groups. RESULTS: 49 patients and 51 unique eyes were included. Groups 1 and 2 had 28 and 38 sets of photographs, respectively. The proportion of global progression in Groups 1 and 2 were 21.4% and 39.5%, respectively (p=0.12). No significant differences in any structural progression feature and patient characteristics (besides age at time of DH (p=0.04) between the two groups were found. CONCLUSIONS: Patients show structural glaucomatous progression before and after the event of a disc haemorrhage without significant differences. This suggests that a DH is an ongoing structural progression in glaucoma and may not be a discrete event that leads to subsequent progression.

Juvenile iridoschisis and incomplete plateau iris configuration.

INTRODUCTION: Iridoschisis is a rare condition in which there is localized cleavage of the iris stroma into 2 layers. It can be associated with plateau iris and more often with either angle closure or open-angle glaucoma. It usually presents in later life as a bilateral progressive condition, however, in this article we present a young adult with bilateral iridoschisis and incomplete plateau iris configuration. CASE PRESENTATION: We report the case of a 22-year-old male with bilateral sensorineural hearing loss who presented to clinic for general ophthalmology evaluation. He had no prior ocular history. Examination findings revealed bilateral iridoschisis with incomplete plateau iris configuration. His ocular examination was otherwise unremarkable except for bilateral retinal pigment epithelium mottling. His work-up for glaucoma including optic disc evaluation, visual field testing, and imaging was all within normal limits. CONCLUSIONS: This case is unusual in its presentation of a young patient with bilateral iridoschisis and incomplete plateau iris configuration. Iridoschisis is more commonly a senile process and the presence of iridoschisis in a young adult prompts a review of congenital causes and associations. Although this patient had no evidence of glaucoma at this time, given the associated risk, it is important that he continue to have regular follow-up at this time.

One Year of Glaucoma Research in Review: 2012 to 2013.

PURPOSE: The objective of this study was to provide the practicing clinical ophthalmologist with an update of pertinent glaucoma literature published from 2012 to 2013. DESIGN: Literature review. METHODS: The authors conducted a 1-year (July 1, 2012, to September 30, 2013) English-language glaucoma literature search on PubMed using the following terms: glaucoma, automated perimetry, optic nerve imaging, optical coherence tomography, glaucoma structure and function, intraocular pressure, central corneal thickness, glaucoma medical therapy, neuroprotection, glaucoma laser treatment, secondary glaucoma, glaucoma surgery, and miscellaneous topics in glaucoma. RESULTS: Of 2659 articles on glaucoma published during our time frame, this review selected original and review articles that reflect novel aspects and updates in the field of glaucoma, while excluding letters to the editor, unpublished works, and abstracts. Preference was given to human research. CONCLUSIONS: This review focuses on literature that is applicable to ophthalmologists in practice and also highlights studies that may enhance the diagnosis and management of glaucoma.

Intraglandular injection of botulinum toxin a reduces tear production in rabbits.

PURPOSE: To develop an animal model and investigate the dose-dependent effect of an intraglandular injection of botulinum toxin A (BTX-A) on tear production. METHODS: In a volume of 0.1-ml, 0.625-, 1.25-, or 2.5-U BTX-A was injected transconjunctivally in the superolateral lobe of the lacrimal gland of adult New Zealand white female rabbits. In the contralateral lacrimal gland, 0.1 ml of 0.9% sodium chloride was injected. Prior to injection and at 1-week postinjection, photographs were taken to evaluate pre- and postoperative eyelid position. Fluorescein and Rose Bengal stain were used to evaluate the corneal surface, and Schirmer test was used to assess tear production. RESULTS: Glands injected with the intermediate (1.25 U) and the highest (2.5 U) doses of BTX-A displayed a statistically significant decrease in tear production (p = 0.002 and 0.007, respectively) compared with the contralateral saline-injected glands at 1 week. No corneal pathologic factors from excessive dryness were observed following the injection. While postinjection ptosis was observed (p = 0.025), no difference was seen between BTX-A and saline-injected eyes. CONCLUSIONS: In rabbits, intraglandular injection of BTX-A resulted in decreased tear production at 1 week. No additional reduction in tear production was seen with a BTX-A dose greater than 1.25 U, suggesting glandular receptor saturation at this dose. Despite suppression of tear production, no corneal pathologic factors were observed. Further studies are needed to refine this animal model with the ultimate goal of determining optimum delivery route and concentration to reduction in tear production while minimizing side effects in patients.

Bilateral internal carotid artery dissection presenting as isolated unilateral Horner syndrome.

A 52-year-old man developed a unilateral Horner syndrome following a skiing accident. He was otherwise asymptomatic. Neuroimaging with magnetic resonance revealed bilateral internal carotid artery dissections, and he was urgently treated with anticoagulation. Immediate neuroimaging should be performed in any patient with Horner syndrome following trauma, regardless of other symptoms or signs.

Isolated unilateral congenital lacrimal gland agenesis presenting as filamentary keratopathy in a child.

Congenital lacrimal gland agenesis is extremely rare, and there are only a few cases reported in the literature. These cases involve bilateral lacrimal gland agenesis associated, in some instances, with salivary gland agenesis and abnormalities of the lacrimal puncta and canaliculi. This report, to our knowledge, presents the first documented case of unilateral lacrimal gland agenesis, resulting in unilateral filamentary keratopathy.

Trans-scleral delivery of antiangiogenic proteins.

PURPOSE: In this study, we investigated the penetration of various proteins into the mouse eye after a periocular injection of the protein or an adenoviral vector (Ad) expressing the protein. METHODS: At several time points after the injection, the retina, retinal pigmented epithelium/choroid, and sclera were dissected and enzyme-linked immunosorbent assays were performed. RESULTS: After a periocular injection of AdsFlt-1.10, AdTGFbeta.10, or AdPEDF.11, choroidal levels of pigment epithelium-derived factor (PEDF) and transforming growth factor-beta (TGF-beta) were not significantly different from scleral levels, and choroidal levels of sFlt-1 (soluble Flt-1 or soluble VEGF receptor 1) were only moderately reduced from scleral levels, indicating that each of these proteins penetrate the sclera well. In contrast, retinal levels of each of the three proteins were low compared to choroidal levels, suggesting poor penetration into the retina. Levels of PEDF in the choroid peaked 2 h after a periocular injection of PEDF protein and returned to baseline between 6 and 24 h, and peak levels in the retina were 8.6% of peak choroidal levels. Levels of green fluorescent protein, a protein unlikely to have any binding sites in mouse tissues, peaked in the choroid 2 h after the periocular injection and were undetectable by 4 h, while peak levels in the retina were 64.3% of peak choroidal levels. CONCLUSIONS: These data suggest that size and binding characteristics of proteins are likely to influence their ability to penetrate the eye from the periocular space, but in general, proteins as large as 50-75 kDa penetrate well into the choroid, but not into the retina. Periocular injections are feasible for the treatment of choroidal neovascularization with proteins or vectors that express them, but additional investigations are needed before they can be considered for treatment of retinal diseases.