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Neovascular age-related macular degeneration (nvAMD) is the leading cause of severe vision loss in the elderly in the developed world. While the introduction of therapies targeting vascular endothelial growth factor (VEGF) has provided the first opportunity to significantly improve vision in patients with nvAMD, many patients respond inadequately to current anti-VEGF therapies. It was recently demonstrated that expression of a second angiogenic mediator, angiopoietin-like 4 (ANGPTL4), synergizes with VEGF to promote choroidal neovascularization (CNV) in mice and correlates with reduced response to anti-VEGF therapy in patients with nvAMD. Here, we report that expression of ANGPTL4 in patients with nvAMD increases following treatment with anti-VEGF therapy and that this increase is dependent on accumulation of hypoxia-inducible factor (HIF)-1a in response to inhibition of VEGF/KDR signaling in the retinal pigment epithelium (RPE). We therefore explored HIF-1 inhibition with 32-134D, a recently developed pharmacologic HIF-inhibitor, for the treatment of nvAMD. 32-134D prevented the expression of both VEGF and ANGPTL4 and was at least as effective as aflibercept in treating CNV in mice. Moreover, by preventing the increase in HIF-1a accumulation in the RPE in response to anti-VEGF therapy, combining 32-134D with aflibercept was more effective than either drug alone for the treatment of CNV. Collectively, these results help explain why many patients with nvAMD respond inadequately to anti-VEGF therapy and suggest that the HIF inhibitor 32-134D will be an effective drug--alone or in combination with current anti-VEGF therapies--for the treatment of patients with this blinding disease. [ABSTRACT FROM AUTHOR]

Many patients with diabetic eye disease respond inadequately to anti-VEGF therapies, implicating additional vasoactive mediators in its pathogenesis. We demonstrate that levels of angiogenic proteins regulated by HIF-1 and -2 remain elevated in the eyes of people with diabetes despite treatment with anti-VEGF therapy. Conversely, by inhibiting HIFs, we normalized the expression of multiple vasoactive mediators in mouse models of diabetic eye disease. Accumulation of HIFs and HIF-regulated vasoactive mediators in hyperglycemic animals was observed in the absence of tissue hypoxia, suggesting that targeting HIFs may be an effective early treatment for diabetic retinopathy. However, while the HIF inhibitor acriflavine prevented retinal vascular hyperpermeability in diabetic mice for several months following a single intraocular injection, accumulation of acriflavine in the retina resulted in retinal toxicity over time, raising concerns for its use in patients. Conversely, 32-134D, a recently developed HIF inhibitor structurally unrelated to acriflavine, was not toxic to the retina, yet effectively inhibited HIF accumulation and normalized HIF-regulated gene expression in mice and in human retinal organoids. Intraocular administration of 32-134D prevented retinal neovascularization and vascular hyperpermeability in mice. These results provide the foundation for clinical studies assessing 32-134D for the treatment of patients with diabetic eye disease., Introduction Identification of factors that trigger the development of nonproliferative diabetic retinopathy (NPDR) remains under debate; these factors include advanced glycosylation end products (AGEs), oxidative stress, and inflammation, all leading [...]

BACKGROUND. Studies assessing the efficacy of therapies for neovascular age-related macular degeneration (nvAMD) have demonstrated that aflibercept may have a longer treatment interval than its less-expensive alternative, bevacizumab. However, whether this benefit justifies the additional cost of aflibercept remains under debate. We have recently reported that a treat-and-extend-pause/monitor approach can be used to successfully wean 31% of patients with nvAMD off anti-VEGF therapy. Here, we examined whether the choice of therapy influences the outcomes of this approach. METHODS. In this retrospective analysis, 122 eyes of 106 patients with nvAMD underwent 3 consecutive monthly injections with either aflibercept (n = 70) or bevacizumab (n = 52), followed by a treat-and-extend protocol, in which the decision to extend the interval between treatments was based on visual acuity, clinical exam, and the presence or absence of fluid on optical coherence tomography. Eyes that remained stable 12 weeks from their prior treatment were given a 6-week trial of holding further treatment, followed by quarterly monitoring. Treatment was resumed for worsening vision, clinical exam, or optical coherence tomography findings. RESULTS. At the end of 1 year, eyes receiving bevacizumab had similar vision but required more injections (8.7 [+ or -] 0.3 treatments vs. 7.2 [+ or -] 0.3 treatments) compared with eyes receiving aflibercept. However, eyes treated with aflibercept were almost 3 times more likely to be weaned off treatment (43% vs. 15%) compared with eyes treated with bevacizumab at the end of 1 year. CONCLUSION. These observations expose an advantage of aflibercept over bevacizumab and have important clinical implications for the selection of therapy for patients with nvAMD. FUNDING. This work was supported by the National Eye Institute, NIH grants R01EY029750 and R01EY025705, Research to Prevent Blindness, the Alcon Young Investigator Award from the Alcon Research Institute, and the Branna and Irving Sisenwein Professorship in Ophthalmology., Introduction Aflibercept (Eylea) and bevacizumab (Avastin) are the two most frequently used therapies for the treatment of neovascular age-related macular degeneration (nvAMD) (1). Both therapies target the vasoactive mediator VEGF. [...]

Background: Angiopoietin‐like 4 is a molecular hallmark that correlates with the growth and metastasis of head and neck squamous cell carcinoma, one of the most prevalent cancers worldwide. However, the molecular mechanisms by which angiopoietin‐like 4 promotes head and neck squamous cell carcinoma tumorigenesis are unclear. Methods: Using well‐characterized cell lines of head and neck squamous cell carcinoma development, including human normal oral keratinocytes, dysplastic oral keratinocytes, oral leukoplakia‐derived oral keratinocytes, and head and neck squamous cell carcinoma cell lines, HN13, HN6, HN4, HN12, and CAL27, we investigated the signaling pathways upstream and downstream of angiopoietin‐like 4‐induced head and neck squamous cell carcinoma tumorigenesis. Results: We found that both epidermal growth factor receptor ligands, epithelial growth factor, and amphiregulin led to angiopoietin‐like 4 upregulation in normal oral keratinocytes and dysplastic oral keratinocytes and cooperated with the activation of hypoxia‐inducible factor‐1 in this effect. Interestingly, amphiregulin and angiopoietin‐like 4 were increased in dysplastic oral keratinocytes and head and neck squamous cell carcinoma cell lines, and amphiregulin‐induced activation of cell proliferation was dependent on angiopoietin‐like 4. Although both p38 mitogen‐activated protein kinases (p38 MAPK) and protein kinase B (AKT) were activated by angiopoietin‐like 4, only pharmacological inhibition of p38 MAPK was sufficient to prevent head and neck squamous cell carcinoma cell proliferation and migration. We further observed that angiopoietin‐like 4 promoted the secretion of interleukin 11 (IL‐11), interleukin 12 (IL‐12), interleukin‐1 alpha (IL‐1α), vascular endothelial growth factor, platelet‐derived growth factor (PDGF), and tumour necrosis factor alpha (TNF‐α), cytokines and chemokines previously implicated in head and neck squamous cell carcinoma pathogenesis. Conclusion: Our results demonstrate that angiopoietin‐like 4 is a downstream effector of amphiregulin and promotes head and neck squamous cell carcinoma development both through direct activation of p38 kinase as well as paracrine mechanisms. [ABSTRACT FROM AUTHOR]

The majority of patients with diabetic macular edema (DME), the most common cause of vision loss in working-age Americans, do not respond adequately to current therapies targeting VEGFA. Here, we show that expression of angiopoietin-like 4 (ANGPTL4), a HIF-1-regulated gene product, is increased in the eyes of diabetic mice and patients with DME. We observed that ANGPTL4 and VEGF act synergistically to destabilize the retinal vascular barrier. Interestingly, while ANGPTL4 modestly enhanced tyrosine phosphorylation of VEGF receptor 2, promotion of vascular permeability by ANGPTL4 was independent of this receptor. Instead, we found that ANGPTL4 binds directly to neuropilin 1 (NRP1) and NRP2 on endothelial cells (ECs), leading to rapid activation of the RhoA/ROCK signaling pathway and breakdown of EC-EC junctions. Treatment with a soluble fragment of NRP1 (sNRP1) prevented ANGPTL4 from binding to NRP1 and blocked ANGPTL4-induced activation of RhoA as well as EC permeability in vitro and retinal vascular leakage in diabetic animals in vivo. In addition, sNRP1 reduced the stimulation of EC permeability by aqueous fluid from patients with DME. Collectively, these data identify the ANGPTL4/NRP/RhoA pathway as a therapeutic target for the treatment of DME., Introduction Human angiopoietin-like (ANGPTL) proteins are a family of 8 related gene products that are structurally similar to the angiopoietins (ANGPTs) and that play a role in a wide array [...]

Wilmer Eye Institute, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA. 'Department of Oncology and Diagnostic Sciences, School of Dentistry and Department of Pathology, School of Medicine, Greenebaum Cancer Center, University of Maryland, Baltimore, Maryland, USA. 'Department of Pathology, Johns Hopkins University, Baltimore, Maryland, USA. BACKGROUND. To reduce the treatment burden for patients with neovascular age-related macular degeneration (nvAMD), emerging therapies targeting vascular endothelial growth factor (VEGF) are being designed to extend the interval between treatments, thereby minimizing the number of intraocular injections. However, which patients will benefit from longer- acting agents is not clear. METHODS. Eyes with nvAMD (n = 122) underwent 3 consecutive monthly injections with currently available anti-VEGF therapies, followed by a treat-and-extend protocol. Patients who remained quiescent 12 weeks from their prior treatment entered a treatment pause and were switched to pro re nata (PRN) treatment (based on vision, clinical exam, and/or imaging studies). Proteomic analysis was performed on aqueous fluid to identify proteins that correlate with patients' response to treatment. RESULTS. At the end of 1 year, 38 of 122 eyes (31%) entered a treatment pause ([greater than or equal to] 0 weeks). Conversely, 21 of 122 eyes (17%) failed extension and required monthly treatment at the end of year 1. Proteomic analysis of aqueous fluid identified proteins that correlated with patients' response to treatment, including proteins previously implicated in AMD pathogenesis. Interestingly, apolipoprotein-B100 (ApoB100), a principal component of drusen implicated in the progression of nonneovascular AMD, was increased in treated patients who required less frequent injections. ApoB100 expression was higher in AMD eyes compared with controls but was lower in eyes that develop choroidal neovascularization (CNV), consistent with a protective role. Accordingly, mice overexpressing ApoB100 were partially protected from laser-induced CNV. FUNDING. This work was supported by the National Eye Institute, National Institutes of Health grants R01EY029750, R01EY025705, and R01 EY27961; the Research to Prevent Blindness, Inc.; the Alcon Research Institute; and Johns Hopkins University through the Robert Bond Welch and Branna and Irving Sisenwein professorships in ophthalmology. CONCLUSION. Aqueous biomarkers could help identify patients with nvAMD who may not require or benefit from long-term treatment with anti-VEGF therapy., Introduction Neovascular (nv) age-related macular degeneration (AMD) is the leading cause of severe vision loss in elderly Americans (1). If left untreated, patients with nvAMD suffer dramatic and irreversible vision [...]

Anti–vascular endothelial growth factor therapy has had a substantial impact on the treatment of choroidal neovascularization (CNV) in patients with neovascular age-related macular degeneration (nAMD), the leading cause of vision loss in older adults. Despite treatment, many patients with nAMD still develop severe and irreversible visual impairment because of the development of subretinal fibrosis. We recently reported the anti-inflammatory and antiangiogenic effects of inhibiting the gene encoding adenosine receptor 2A (Adora2a), which has been implicated in cardiovascular disease. Here, using two mouse models of subretinal fibrosis (mice with laser injury–induced CNV or mice with a deficiency in the very low–density lipoprotein receptor), we found that deletion of Adora2a either globally or specifically in endothelial cells reduced subretinal fibrosis independently of angiogenesis. We showed that Adora2a-dependent endothelial-to-mesenchymal transition contributed to the development of subretinal fibrosis in mice with laser injury–induced CNV. Deficiency of Adora2a in cultured mouse and human choroidal endothelial cells suppressed induction of the endothelial-to-mesenchymal transition. A metabolomics analysis of cultured human choroidal endothelial cells showed that ADORA2A knockdown with an siRNA reversed the increase in succinate because of decreased succinate dehydrogenase B expression under fibrotic conditions. Pharmacological inhibition of ADORA2A with a small-molecule KW6002 in both mouse models recapitulated the reduction in subretinal fibrosis observed in mice with genetic deletion of Adora2a. ADORA2A inhibition may be a therapeutic approach to treat subretinal fibrosis associated with nAMD. Editor's summary: Retinal fibrosis reversal age-related macular degeneration (AMD) is driven by neovascularization and is associated with subretinal fibrosis. Working in mice with genetically induced AMD or with a laser-induced retinal injury, Yang et al. show that deleting the gene encoding adenosine receptor 2A (Adora2a) prevented subretinal fibrosis in both mouse models. Specifically, deleting Adora2a in endothelial cells was sufficient to reverse subretinal fibrosis by preventing endothelial-to-mesenchymal transition. In human choroidal endothelial cells, the authors showed that endothelial-to-mesenchymal transition was associated with succinate accumulation due to decreased succinate dehydrogenase subunit B expression. Treatment with an ADORA2A antagonist reversed subretinal fibrosis in both mouse models, suggesting that targeting ADORA2A in endothelial cells could be a therapeutic strategy to treat AMD. —Brandon Berry [ABSTRACT FROM AUTHOR]

Therapies targeting VEGF have proven only modestly effective for the treatment of proliferative sickle cell retinopathy (PSR), the leading cause of blindness in patients with sickle cell disease. Here, we shift our attention upstream from the genes that promote retinal neovascularization (NV) to the transcription factors that regulate their expression. We demonstrated increased expression of HIF-1[alpha] and HIF-2[alpha] in the ischemic inner retina of PSR eyes. Although both HIFs participated in promoting VEGF expression by hypoxic retinal Muller cells, HIF-1 alone was sufficient to promote retinal NV in mice, suggesting that therapies targeting only HIF-2 would not be adequate to prevent PSR. Nonetheless, administration of a HIF-2-specific inhibitor currently in clinical trials (PT2385) inhibited NV in the oxygen-induced retinopathy (OIR) mouse model. To unravel these discordant observations, we examined the expression of HIFs in OIR mice and demonstrated rapid but transient accumulation of HIF-1[alpha] but delayed and sustained accumulation of HIF- 2[alpha]; simultaneous expression of HIF-1[alpha] and HIF-2[alpha] was not observed. Staggered HIF expression was corroborated in hypoxic adult mouse retinal explants but not in human retinal organoids, suggesting that this phenomenon may be unique to mice. Using pharmacological inhibition or an in vivo nanoparticle-mediated RNAi approach, we demonstrated that inhibiting either HIF was effective for preventing NV in OIR mice. Collectively, these results explain why inhibition of either HIF-1[alpha] or HIF-2[alpha] is equally effective for preventing retinal NV in mice but suggest that therapies targeting both HIFs will be necessary to prevent NV in patients with PSR., Introduction Several multicenter randomized controlled clinical trials have demonstrated the benefit of monthly (or bimonthly) injections with biological molecules directed against the vasoactive mediator VEGF to treat macular edema in [...]

It has previously been reported that antioxidant vitamins can help reduce the risk of vision loss associated with progression to advanced age-related macular degeneration (AMD), a leading cause of visual impairment among the elderly. Nonetheless, how oxidative stress contributes to the development of choroidal neovascularization (CNV) in some AMD patients and geographic atrophy (GA) in others is poorly understood. Here, we provide evidence demonstrating that oxidative stress cooperates with hypoxia to synergistically stimulate the accumulation of hypoxia-inducible factor (HIF)-1a in the retinal pigment epithelium (RPE), resulting in increased expression of the HIF-1-dependent angiogenic mediators that promote CNV. HIF-1 inhibition blocked the expression of these angiogenic mediators and prevented CNV development in an animal model of ocular oxidative stress, demonstrating the pathological role of HIF-1 in response to oxidative stress stimulation in neovascular AMD. While human-induced pluripotent stem cell (hiPSC)-derived RPE monolayers exposed to chemical oxidants resulted in disorganization and disruption of their normal architecture, RPE cells proved remarkably resistant to oxidative stress. Conversely, equivalent doses of chemical oxidants resulted in apoptosis of hiPSC-derived retinal photoreceptors. Pharmacologic inhibition of HIF-1 in the mouse retina enhanced--while HIF-1 augmentation reduced--photoreceptor apoptosis in two mouse models for oxidative stress, consistent with a protective role for HIF-1 in photoreceptors in patients with advanced dry AMD. Collectively, these results suggest that in patients with AMD, increased expression of HIF-1a in RPE exposed to oxidative stress promotes the development of CNV, but inadequate HIF-1a expression in photoreceptors contributes to the development of GA. [ABSTRACT FROM AUTHOR]

Background and Purpose: Pathological angiogenesis is a major cause of irreversible blindness in individuals with neovascular age-related macular degeneration (nAMD). Macrophages and microglia (MΦ) contribute to aberrant ocular angiogenesis. However, the role of glucose metabolism of MΦ in nAMD is still undefined. Here, we have investigated the involvement of glycolysis, driven by the kinase/phosphatase PFKFB3, in the development of choroidal neovascularization (CNV).Experimental Approach: CNV was induced in mice with laser photocoagulation. Choroid/retinal pigment epithelium (RPE) complexes and MΦ were isolated for analysis by qRT-PCR, western blot, flow cytometry, immunostaining, metabolic measurements and angiogenesis assays.Key Results: MΦ accumulated within the CNV of murine nAMD models and expressed high levels of glycolysis-related enzymes and M1/M2 polarization markers. This phenotype of hyper-glycolytic and activated MΦ was replicated in bone marrow-derived macrophages stimulated by necrotic RPE in vitro. Myeloid cell-specific knockout of PFKFB3, a key glycolytic activator, attenuated pathological neovascularization in laser-induced CNV, which was associated with decreased expression of MΦ polarization markers and pro-angiogenic factors, along with decreased sprouting of vessels in choroid/RPE complexes. Mechanistically, necrotic RPE increased PFKFB3-driven glycolysis in macrophages, leading to activation of HIF-1α/HIF-2α and NF-κB, and subsequent induction of M1/M2 markers and pro-angiogenic cytokines, finally promoting macrophage reprogramming towards an angiogenic phenotype to facilitate development of CNV. The PFKFB3 inhibitor AZ67 also inhibited activation of HIF-1α/HIF-2α and NF-κB signalling and almost completely prevented laser-induced CNV in mice.Conclusions and Implications: Modulation of PFKFB3-mediated macrophage glycolysis and activation is a promising strategy for the treatment of nAMD. [ABSTRACT FROM AUTHOR]

PRAGMatic myeloid cells: Ocular angiogenesis is a major cause of vision impairments and blindness. Microglia/macrophages have been shown to contribute to pathological angiogenesis; however, their phenotype and role in retinal angiogenesis has not been completely elucidated. Now, Liu et al. show that in pathological retinal angiogenesis, microglia/macrophages are highly glycolytic and acquire a pathological phenotype, characterized by high expression of proinflammatory and proangiogenic cytokines. They called these cells pathological retinal angiogenesis–associated glycolytic macrophages/microglia (PRAGMs). Glycolytic activation triggered PRAGMs, suggesting that strategies targeting metabolic changes could be effective in treating pathological retinal angiogenesis. The coordination of metabolic signals among different cellular components in pathological retinal angiogenesis is poorly understood. Here, we showed that in the pathological angiogenic vascular niche, retinal myeloid cells, particularly macrophages/microglia that are spatially adjacent to endothelial cells (ECs), are highly glycolytic. We refer to these macrophages/microglia that exhibit a unique angiogenic phenotype with increased expression of both M1 and M2 markers and enhanced production of both proinflammatory and proangiogenic cytokines as pathological retinal angiogenesis–associated glycolytic macrophages/microglia (PRAGMs). The phenotype of PRAGMs was recapitulated in bone marrow–derived macrophages or retinal microglia stimulated by lactate that was produced by hypoxic retinal ECs. Knockout of 6-phosphofructo-2-kinase/fructose-2, 6-bisphosphatase (PFKFB3; Pfkfb3 for rodents), a glycolytic activator in myeloid cells, impaired the ability of macrophages/microglia to acquire an angiogenic phenotype, rendering them unable to promote EC proliferation and sprouting and pathological neovascularization in a mouse model of oxygen-induced proliferative retinopathy. Mechanistically, hyperglycolytic macrophages/microglia produced large amount of acetyl–coenzyme A, leading to histone acetylation and PRAGM-related gene induction, thus reprogramming macrophages/microglia into an angiogenic phenotype. These findings reveal a critical role of glycolytic metabolites as initiators of reciprocal activation of macrophages/microglia and ECs in the retinal angiogenic niche and suggest that strategies targeting the metabolic communication between these cell types may be efficacious in the treatment of pathological retinal angiogenesis. [ABSTRACT FROM AUTHOR]

The recent success of therapies directly targeting the angiogenic mediator, vascular endothelial growth factor (VEGF), for the treatment of proliferative diabetic retinopathy has encouraged clinicians to extend the use of anti-VEGF therapies for the treatment of another ischemic retinal vascular disease, proliferative sickle cell retinopathy (PSR), the most common cause of irreversible blindness in patients with sickle cell disease. However, results from case reports evaluating anti-VEGF therapies for PSR have been mixed. This highlights the need to identify alternative therapeutic targets for the treatment of retinal neovascularization in sickle cell patients. In this regard, angiopoietin-like 4 (ANGPTL4) is a novel angiogenic factor regulated by the transcription factor, hypoxia-inducible factor 1, the master regulator of angiogenic mediators (including VEGF) in ischemic retinal disease. In an effort to identify alternative targets for the treatment of sickle cell retinopathy, we have explored the expression of ANGPTL4 in the eyes of patients with PSR. To this end, we examined expression and localization of ANGPTL4 by immunohistochemistry in autopsy eyes from patients with known PSR (n = 5 patients). Complementary studies were performed using enzyme-linked immunosorbent assays in aqueous (n = 8; 7 patients, 2 samples from one eye of same patient) and vitreous (n = 3 patients) samples from a second group of patients with active PSR. We detected expression of ANGPTL4 in neovascular tissue and in the ischemic inner retina in PSR, but not control, eyes. We further observed elevated expression of ANGPTL4 in the aqueous and vitreous of PSR patients compared to controls. These results suggest that ANGPTL4 could contribute to the development of retinal neovascularization in sickle cell patients and could therefore be a therapeutic target for the treatment of PSR. [ABSTRACT FROM AUTHOR]

The transcriptional response promoted by hypoxia-inducible factors has been associated with metastatic spread of uveal melanoma. We found expression of hypoxia-inducible factor 1α (HIF-1α) protein in well-vascularized tumor regions as well as in four cell lines grown in normoxia, thus this pathway may be important even in well-oxygenated uveal melanoma cells. HIF-1α protein accumulation in normoxia was inhibited by rapamycin. As expected, hypoxia (1% pO2) further induced HIF-1α protein levels along with its target genes VEGF and LOX. Growth in hypoxia significantly increased cellular invasion of all 5 uveal melanoma lines tested, as did the introduction of an oxygen-insensitive HIF-1α mutant into Mel285 cells with low HIF-1α baseline levels. In contrast, HIF-1α knockdown using shRNA significantly decreased growth in hypoxia, and reduced by more than 50% tumor invasion in four lines with high HIF-1α baseline levels. Pharmacologic blockade of HIF-1α protein expression using digoxin dramatically suppressed cellular invasion both in normoxia and in hypoxia. We found that Notch pathway components, including Jag1-2 ligands, Hes1-Hey1 targets and the intracellular domain of Notch1, were increased in hypoxia, as well as the phosphorylation levels of Erk1-2 and Akt. Pharmacologic and genetic inhibition of Notch largely blocked the hypoxic induction of invasion as did the pharmacologic suppression of Erk1-2 activity. In addition, the increase in Erk1-2 and Akt phosphorylation by hypoxia was partially reduced by inhibiting Notch signaling. Our findings support the functional importance of HIF-1α signaling in promoting the invasive capacity of uveal melanoma cells in both hypoxia and normoxia, and suggest that pharmacologically targeting HIF-1α pathway directly or through blockade of Notch or Erk1-2 pathways can slow tumor spread. [ABSTRACT FROM AUTHOR]

Age-related macular degeneration (AMD) is the leading cause of blindness in developed countries; with the aging population, the negative health impacts and costs of the disease will increase dramatically over the next decade. Although the exact cause of AMD is unknown, genetic studies have implicated the complement system as well as other immune responses in disease pathogenesis and severity. Furthermore, histologic studies have shown the presence of macrophages, lymphocytes, and mast cells, as well as fibroblasts, in both atrophic lesions and with retinal neovascularization. This review summarizes discussions from the fifth annual conference of the Arnold and Mabel Beckman Initiative for Macular Research by the Inflammation and Immune Response Task Force. These deliberations focused on the role of inflammatory immune responses, including complement, inflammasomes, adaptive immune responses, and para-inflammation, unanswered questions and studies to address these questions, and potential immune-related therapeutic targets for AMD. [ABSTRACT FROM AUTHOR]

Background: Kaposi's sarcoma (KS) is a vascular neoplasm characterized by the dysregulated expression of angiogenic and inflammatory cytokines. The driving force of the KS lesion, the KSHV-infected spindle cell, secretes elevated levels of vascular endothelial growth factor (VEGF), essential for KS development. However, the origin of VEGF in this tumor remains unclear. Methodology/Principal Findings: Here we report that the KSHV G protein-coupled receptor (vGPCR) upregulates VEGF in KS through an intricate paracrine mechanism. The cytokines secreted by the few vGPCR-expressing tumor cells activate in neighboring cells multiple pathways (including AKT, ERK, p38 and IKKβ) that, in turn, converge on TSC1/2, promoting mTOR activation, HIF upregulation, and VEGF secretion. Conditioned media from vGPCR-expressing cells lead to an mTOR-dependent increase in HIF-1α and HIF-2α protein levels and VEGF upregulation. In a mouse allograft model for KS, specific inhibition of the paracrine activation of mTOR in non-vGPCR-expressing cells was sufficient to inhibit HIF upregulation in these cells, and abolished the ability of the vGPCR-expressing cells to promote tumor formation in vivo. Similarly, pharmacologic inhibition of HIF in this model blocked VEGF secretion and also lead to tumor regression. Conclusions/Significance: Our findings provide a compelling explanation for how the few tumor cells expressing vGPCR can contribute to the dramatic amplification of VEGF secretion in KS, and further provide a molecular mechanism for how cytokine dysregulation in KS fuels angiogenesis and tumor development. These data further suggest that activation of HIF by vGPCR may be a vulnerable target for the treatment of patients with KS. [ABSTRACT FROM AUTHOR]

Abstract: It has been nearly a century since Jules Gonin performed the first intervention for rhegmatogenous retinal detachment, trans-scleral cautery, achieving successful outcomes in close to 50% of his cases. With the introduction of alternative surgical approaches in the last half-century, including Charles Schepens'' scleral buckle technique and Robert Machemer''s pars plana vitrectomy, the surgical success rates have risen to close to 90%. Nonetheless, despite dramatic progress in the success of reattachment surgeries, reasonable disagreement exists as to which approach (or combination of approaches) is the best form of surgical intervention for patients with rhegmatogenous retinal detachments. In this review, the authors summarize the current knowledge of retinal detachment, and examine emerging results from the first large scale, prospective, randomized, controlled clinical trials addressing the efficacy of these surgical approaches for retinal detachment, with the hope of identifying the most appropriate (evidence-based) therapeutic intervention for the treatment of rhegmatogenous retinal detachment. [Copyright &y& Elsevier]

Viruses use a surprising diversity of approaches to hijack G-protein-coupled receptors and harness their activated intracellular signalling pathways. All of these approaches ultimately function to ensure viral replicative success and often contribute to their pathogenesis. Indeed, a single virus might deploy a repertoire of these strategies to regulate key intracellular survival, proliferative and chemotactic pathways. Understanding the contribution of these biochemical routes to viral pathogenesis might facilitate the development of effective target-specific therapeutic strategies against viral diseases. [ABSTRACT FROM AUTHOR]

In 1994, the Kaposi's sarcoma-associated herpesvirus (KSHV/HHV-8) was identified as the etiologic agent of Kaposi's sarcoma (KS). KSHV has since been associated with two additional AIDS-related malignancies: primary effusion lymphomas (PEL) and multicentric Castleman's disease (MCD). Although molecular characterization of the KSHV genome has revealed several candidate oncogenes, infection with KSHV alone is not sufficient to cause KS, suggestive of an accomplice in KS initiation. Recent experimental evidence supports a key role for a particular KSHV gene, a constitutively-active G-protein-coupled receptor (vGPCR), in the development of KS. However, it is unclear how a lytic gene expressed in cells destined to die can cause cancer. Here we propose that dysregulation of the viral gene program may lead to nonlytic vGPCR expression. Several candidate cofactors (e.g., HIV-1 Tat, inflammation, aborted lytic cycle progression) are identified that may trigger vGPCR dysregulation, enabling oncogenic signaling pathways up-regulated by vGPCR, combined with the paracrine secretions from vGPCR-expressing cells, to promote the initiation of KS. If KS is indeed dependent on vGPCR dysregulation, then the development of new therapeutic modalities specifically targeting this viral protein or its downstream targets may ultimately prove to be the most effective treatment strategy for this enigmatic disease. [ABSTRACT FROM AUTHOR]

The article presents a case study of a 55-year old man with a medium-size choroidal melanoma who was treated with plaque radiotherapy using iodine-125. He was fashioned with a partial peritomy at the inferotemporal and temporal quadrant and the plaque was inspected and found to contain all its seeds. The patient was observed to have Visual Acuity (VA) of 20/100 +1 in the right eye, full extraocular movements and was orthophoric. It discusses transillumination, peritomy and enucleation.

Uveal melanoma (UM) is an aggressive intraocular malignancy derived from melanocytes in the uvea tract of the eye. Up to 50% of patients with UM develop distant metastases which is usually fatal within one year; preventing metastases is therefore essential. Metabolic reprogramming plays a critical role in UM progression and metastasis. However, the metabolic phenotype of UM cells in the hypoxic tumor is not well understood. Here, we report that hypoxia-induced BNIP3 reprograms tumor cell metabolism, promoting their survival and metastasis. In response to hypoxia, BNIP3-mediated mitophagy alleviates mitochondrial dysfunction and enhances mitochondrial oxidative phosphorylation (OXPHOS) while simultaneously reducing mitochondrial reactive oxygen species (mtROS) production. This, in turn, impairs HIF1A/HIF-1α protein stability and inhibits glycolysis. Inhibition of mitophagy significantly suppresses BNIP3-induced UM progression and metastasis in vitro and in vivo. Collectively, these observations demonstrate a novel mechanism whereby BNIP3 promotes UM metabolic reprogramming and malignant progression by mediating hypoxia-induced mitophagy and suggest that BNIP3 could be an important therapeutic target to prevent metastasis in patients with UM.Abbreviations: AOD: average optical density; BNIP3: BCL2/adenovirus E1B interacting protein 3; CQ: chloroquine; CoCl2: cobalt chloride; GEPIA: Gene Expression Profiling Interactive Analysis; HIF1A: hypoxia inducible factor 1, alpha subunit; IHC: immunohistochemistry; mtROS: mitochondrial reactive oxygen species; NAC: N-acetylcysteine; OCR: oxygen consumption rate; OXPHOS: oxidative phosphorylation; ROS: reactive oxygen species; TCGA: The Cancer Genome Atlas; UM: uveal melanoma. [ABSTRACT FROM AUTHOR]

Many patients with diabetic eye disease respond inadequately to anti-VEGF therapies, implicating additional vasoactive mediators in its pathogenesis. We demonstrate that levels of angiogenic proteins regulated by HIF-1 and -2 remain elevated in the eyes of people with diabetes despite treatment with anti-VEGF therapy. Conversely, by inhibiting HIFs, we normalized the expression of multiple vasoactive mediators in mouse models of diabetic eye disease. Accumulation of HIFs and HIF-regulated vasoactive mediators in hyperglycemic animals was observed in the absence of tissue hypoxia, suggesting that targeting HIFs may be an effective early treatment for diabetic retinopathy. However, while the HIF inhibitor acriflavine prevented retinal vascular hyperpermeability in diabetic mice for several months following a single intraocular injection, accumulation of acriflavine in the retina resulted in retinal toxicity over time, raising concerns for its use in patients. Conversely, 32-134D, a recently developed HIF inhibitor structurally unrelated to acriflavine, was not toxic to the retina, yet effectively inhibited HIF accumulation and normalized HIF-regulated gene expression in mice and in human retinal organoids. Intraocular administration of 32-134D prevented retinal neovascularization and vascular hyperpermeability in mice. These results provide the foundation for clinical studies assessing 32-134D for the treatment of patients with diabetic eye disease. [ABSTRACT FROM AUTHOR]

Background: Studies assessing the efficacy of therapies for neovascular age-related macular degeneration (nvAMD) have demonstrated that aflibercept may have a longer treatment interval than its lesser-expensive alternative, bevacizumab. However, whether this benefit justifies the additional cost of aflibercept remains under debate. We have recently reported that a "treat-and-extend-pause/monitor" (TEP/M) approach can be used to successfully wean 31% of nvAMD patients off anti-VEGF therapy. Here we examine whether the choice of therapy influences the outcomes of this approach.Methods: In this retrospective analysis, 122 eyes of 106 patients with nvAMD underwent 3 consecutive monthly injections with either aflibercept (n=70) or bevacizumab (n=52) followed by a treat-and-extend protocol in which the decision to extend the interval between treatments was based on visual acuity, clinical exam, and the presence or absence of fluid on optical coherence tomography (OCT). Eyes that remained stable 12 weeks from their prior treatment were given a 6-week trial of holding further treatment, followed by quarterly monitoring. Treatment was resumed for worsening vision, clinical exam, or OCT findings.Results: At the end of one year, eyes receiving bevacizumab had similar vision but required more injections (8.7 ±0.3 vs. 7.2 ±0.3) compared to aflibercept. However, eyes treated with aflibercept were almost 3-times more likely to be weaned off treatment (43% vs. 15%) compared to eyes treated with bevacizumab at the end of one year.Conclusions: These observations expose a previously unappreciated advantage of aflibercept over bevacizumab and have important clinical implications for the selection of therapy for patients with nvAMD. [ABSTRACT FROM AUTHOR]

The article hypothesized that vascular cells within neovascular tissue secrete autocrine/paracrine angiogenic factors that promote disease progression. It discusses that multiplex ELISA angiogenesis arrays was performed on aqueous fluid from proliferative diabetic retinopathy patients who responded inadequately to anti-vascular endothelial growth factor therapy and/or n pan-retinal scatter laser photocoagulation and identified plasminogen activator inhibitor-1 (PAI-1).

BackgroundTo reduce the treatment burden for patients with neovascular age-related macular degeneration (nvAMD), emerging therapies targeting vascular endothelial growth factor (VEGF) are being designed to extend the interval between treatments, thereby minimizing the number of intraocular injections. However, which patients will benefit from longer-acting agents is not clear.MethodsEyes with nvAMD (n = 122) underwent 3 consecutive monthly injections with currently available anti-VEGF therapies, followed by a treat-and-extend protocol. Patients who remained quiescent 12 weeks from their prior treatment entered a treatment pause and were switched to pro re nata (PRN) treatment (based on vision, clinical exam, and/or imaging studies). Proteomic analysis was performed on aqueous fluid to identify proteins that correlate with patients' response to treatment.ResultsAt the end of 1 year, 38 of 122 eyes (31%) entered a treatment pause (≥30 weeks). Conversely, 21 of 122 eyes (17%) failed extension and required monthly treatment at the end of year 1. Proteomic analysis of aqueous fluid identified proteins that correlated with patients' response to treatment, including proteins previously implicated in AMD pathogenesis. Interestingly, apolipoprotein-B100 (ApoB100), a principal component of drusen implicated in the progression of nonneovascular AMD, was increased in treated patients who required less frequent injections. ApoB100 expression was higher in AMD eyes compared with controls but was lower in eyes that develop choroidal neovascularization (CNV), consistent with a protective role. Accordingly, mice overexpressing ApoB100 were partially protected from laser-induced CNV.FundingThis work was supported by the National Eye Institute, National Institutes of Health grants R01EY029750, R01EY025705, and R01 EY27961; the Research to Prevent Blindness, Inc.; the Alcon Research Institute; and Johns Hopkins University through the Robert Bond Welch and Branna and Irving Sisenwein professorships in ophthalmology.ConclusionAqueous biomarkers could help identify patients with nvAMD who may not require or benefit from long-term treatment with anti-VEGF therapy. [ABSTRACT FROM AUTHOR]