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AAV-mediated gene delivery of the calreticulin anti-angiogenic domain inhibits ocular neovascularization

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Abstract

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.

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Acknowledgements

This work was supported by grants from The National Health and Medical Research Council of Australia (#1061912), The Natural Science Foundation of Guangdong Province, China (2015A030310158 and 2014A030313359), The Science and Technology Planning Project of Guangdong Province, China (2015B020226003), The Scientific and Cultivation Foundation of the First Affiliated Hospital of Jinan University (2015201), The fundamental Research Funds for the Central Universities (21611446), The Ophthalmic Research Institute of Australia, The Angior Family Foundation and The Rebecca L. Cooper Medical Research Foundation. J.H.W. received a R.B. McComas Research Scholarship in Ophthalmology, a Gordon P. Castles Scholarship and a Melbourne Research Scholarship. G.J.D. received a Principal Research Fellowship from NHMRC. The Centre for Eye Research, Australia, received Operational Infrastructure Support from the Victorian Government.

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Authors and Affiliations

  1. Department of Ophthalmology, The First Affiliated Hospital of Jinan University, Guangzhou, Guangdong, China

    Leilei Tu, Jingxiang Zhong & Guei-Sheung Liu

  2. Centre for Eye Research Australia, Royal Victorian Eye and Ear Hospital, East Melbourne, VIC, Australia

    Leilei Tu, Jiang-Hui Wang, Jia Hui Lee, Gregory J. Dusting & Guei-Sheung Liu

  3. Ophthalmology, Department of Surgery, University of Melbourne, Melbourne, VIC, Australia

    Jiang-Hui Wang, Gregory J. Dusting & Guei-Sheung Liu

  4. Translational Pre-clinical Model Platform, Singapore Eye Research Institute, Singapore National Eye Centre, Singapore, Singapore

    Veluchamy A. Barathi

  5. Department of Ophthalmology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore

    Veluchamy A. Barathi

  6. Ophthalmology and Visual Sciences Academic Clinical Program, DUKE-NUS Graduate Medical School, Singapore, Singapore

    Veluchamy A. Barathi

  7. Department of Optometry and Vision Sciences, University of Melbourne, Parkville, VIC, Australia

    Selwyn M. Prea, Zheng He & Bang V. Bui

  8. Wicking Dementia Research and Education Centre, University of Tasmania, Hobart, TAS, Australia

    James Bender & Anna E. King

  9. Gene Therapy Research Unit, Children’s Medical Research Institute and Sydney Children’s Hospitals Network, University of Sydney, Sydney, NSW, Australia

    Grant J. Logan & Ian E. Alexander

  10. Discipline of Child and Adolescent Health, University of Sydney, Westmead, NSW, Australia

    Ian E. Alexander

  11. Department of Ophthalmology, Kaohsiung Veterans General Hospital, Kaohsiung, Taiwan

    Youn-Shen Bee

  12. Institute of Biomedical Sciences, National Sun Yat-Sen University, Kaohsiung, Taiwan

    Ming-Hong Tai

  13. Menzies Institute for Medical Research, University of Tasmania, Hobart, TAS, Australia

    Guei-Sheung Liu

  14. Liverpool St, Hobart, TAS, 7000, Australia

    Guei-Sheung Liu

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  1. Leilei Tu
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  2. Jiang-Hui Wang
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  3. Veluchamy A. Barathi
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  4. Selwyn M. Prea
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  16. Guei-Sheung Liu
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Contributions

Conceptualization, L.T., J.Z., and G.S.L. Methodology, L.T., V.A.B., M.H.T., B.V.B., and G.S.L. Formal analysis, L.T., J.H.W., V.A.B., B.V.B., and G.S.L. Investigation, L.T., J.H.W., V.A.B., S.M.P., Z.H., J.B., B.V.B., and G.S.L. Resources, A.E.K., G.J.L., I.E.A., and M.H.T. Data curation, L.T., B.V.B., J.Z., and G.S.L. Writing–original draft, L.T., J.Z., and G.S.L. Writing–review & editing, J.H.W., V.A.B., J.H.L., A.E.K., I.E.A., Y.S.B., G.J.D., and B.V.B. Visualization, L.T., and G.S.L. Supervision, G.J.D., B.V.B., J.Z., and G.S.L. Project administration, L.T., and G.S.L. Funding acquisition, G.J.D., J.Z., and G.S.L.

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Correspondence to Jingxiang Zhong or Guei-Sheung Liu.

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Tu, L., Wang, JH., Barathi, V.A. et al. AAV-mediated gene delivery of the calreticulin anti-angiogenic domain inhibits ocular neovascularization. Angiogenesis 21, 95–109 (2018). https://doi.org/10.1007/s10456-017-9591-4

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