Selective Inhibition of Retinal Angiogenesis by Targeting PI3 Kinase Yolanda Alvarez1*, Olaya Astudillo1, Lasse Jensen2, Alison L. Reynolds1, Nora Waghorne1, Derek P. Brazil3, Yihai Cao2, John J. O’Connor1, Breanda´ n N. Kennedy1* 1 UCD Schools of Biomolecular and Biomedical Sciences, UCD Conway Institute, University College Dublin, Dublin, Ireland, 2 Institute for Microbiology Tumor and Cell Biology, Karolinska Institute, Stockholm, Sweden, 3 Centre for Vision and Vascular Science, School of Medicine, Dentistry and Biomedical Science, Queen’s University Belfast, Belfast, United Kingdom Abstract Ocular neovascularisation is a pathological hallmark of some forms of debilitating blindness including diabetic retinopathy, age related macular degeneration and retinopathy of prematurity. Current therapies for delaying unwanted ocular angiogenesis include laser surgery or molecular inhibition of the pro-angiogenic factor VEGF. However, targeting of angiogenic pathways other than, or in combination to VEGF, may lead to more effective and safer inhibitors of intraocular angiogenesis. In a small chemical screen using zebrafish, we identify LY294002 as an effective and selective inhibitor of both developmental and ectopic hyaloid angiogenesis in the eye. LY294002, a PI3 kinase inhibitor, exerts its anti-angiogenic effect in a dose-dependent manner, without perturbing existing vessels. Significantly, LY294002 delivered by intraocular injection, significantly inhibits ocular angiogenesis without systemic side-effects and without diminishing visual function. Thus, targeting of PI3 kinase pathways has the potential to effectively and safely treat neovascularisation in eye disease. Citation: Alvarez Y, Astudillo O, Jensen L, Reynolds AL, Waghorne N, et al. (2009) Selective Inhibition of Retinal Angiogenesis by Targeting PI3 Kinase. PLoS ONE 4(11): e7867. doi:10.1371/journal.pone.0007867 Editor: Rafael Linden, Universidade Federal do Rio de Janeiro (UFRJ), Instituto de Biof´ısica da UFRJ, Brazil Received August 10, 2009; Accepted October 9, 2009; Published November 17, 2009 Copyright: 2009 Alvarez et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported by grants: SFI 04/IN3/B559 (BK), SFI 06/RFP/GEN030 (BK), HRB/MRCG FB06KEN (BK), and HRB Postdoctoral Fellowship PD2006- 32 (YA). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: yolanda.alvarez@ucd.ie (YA); brendan.kennedy@ucd.ie (BNK) Introduction Ocular Diseases and Neovascularisation Functional damage to existing blood vessels and inappropriate development of new blood vessels are hallmarks of prominent forms of blindness. In diabetic retinopathy (DR), which affects ,150 million patients, neovascularisation of vessels in the adult retina occurs [1]. In age-related macular degeneration (AMD), which is estimated to affect ,8 million US patients and ,20 million patients in US plus Europe, neovascularisation of capillaries in the choroid layer adjacent to the retina occurs [2]. Ocular neovascularisation associated with retinopathy of prema- turity (ROP) arises when premature infants are placed into high oxygen to alleviate respiratory distress. As a consequence, concomitant regression of the lens-associated hyaloid vasculature and growth of the nascent retinal vasculature ceases. When returned to normoxia, the infant retina becomes hypoxic leading to neovascularisation, persistent hyaloid vasculature and eventual blindness [3]. Thus, a therapeutic goal for DR and AMD is to develop agents that inhibit ocular neovascularisation and/or stabilise existing vessels, without diminishing visual function. Current Ocular Angiogenesis Treatments Current interventions for resolving inappropriate growth of new vessels in the eye include laser treatment or molecular therapies targeted to vascular endothelial cell growth factor (VEGF). Although the therapeutic mechanism of action is not fully understood, laser photocoagulation surgery can halt neovascularisation and reduce macular oedema [1,4]. Howev- er, success is partial, requires multiple treatments and side effects include cataracts, hemorrhage, retinal detachment and visual field loss. Variations of VEGF molecular therapy to inhibit ocular neovascularisation include antibodies, siRNAs or small molecule inhibitors [5–10]. Those in clinical use are antibodies targeted to VEGF (e.g. bevacizumab (AvastinH) and randizumab (LucentisH)) which delay the growth of new vessels and delay vision loss. Treatment also requires intraocular injection by retinal specialists, repeat administrations and patient sedation. Concerns regarding the development of siRNAs to VEGF have also arisen as they may act non-specifically [7]. More general concerns regarding the strategy of solely targeting VEGF include, the observed compen- satory up-regulation of other pro-angiogenic factors which decrease therapeutic efficacy, and the potential to cause retinal neurodegeneration by inhibiting VEGF’s neuroprotectant func- tion [11,12]. Alternative Anti Angiogenic Targets Targeting angiogenic molecules other than, or in addition to VEGF, may reveal more effective and safer inhibitors of intraocular neovascularisation. Potential targets include growth factors (e.g. angiopoietin, FGF, HGF, IGF-1, PDGF-B, PlGF), chemokines (e.g. IL8, SDF1, G-CSF), receptors (e.g. CXCR1, FGF- R, PlGFR, PDGFR, Tie-receptors), intracellular mediators (e.g. c-kit kinase, PI3 kinase, PKC) and extracellular mediators (e.g. integrins, cadherins) [13]. PLoS ONE | www.plosone.org 1 November 2009 | Volume 4 | Issue 11 | e7867