Abstract

Volume.120 Number.3

To Protect Corneal Transparency against Diseases
Tomohiko Usui
Department of Ophthalmology, The University of Tokyo, Graduate School of Medicine and Faculty of Medicine

To protect corneal transparency, we tried to develop a new therapeutic strategy for corneal neovascularization, corneal scar, and TGFBI-related corneal dystrophy using nucleic acid drug.
1. The expression of angiopietin-like protein 2 (Angptl2) markedly increased in the neovascularized corneas compared to the normal cornea, and Angptl2 was a potent inducer of inflammatory corneal neovascularization. We have produced a single-stranded proline-modified short hairpin anti-Angptl2 ribonucleric acid interference (RNAi) molecule that is carried in a lipid nano-particle for topical application. We have found this agent can penetrate all layers of the cornea. Angptl2 mRNA expression and corneal neovascularization were inhibited in a mouse alkari injury model by topical application of this agent. Thus, this modified RNAi agent is a new topical formulation for use against corneal neovascularization and scar.
2. Human umbilical vein endothelial cells (HUVECs) were cultured with human corneal keratocytes under serum-free conditions. We performed microarray gene-expression analysis in the co-culture system and selected angiopoietin-like protein 7 (Angptl7). In vivo, intrastromal injections of an anti-Angptl7 RNAi agent into the avascular corneal stroma of mice resulted in the growth of blood vessels. Further, we examined the effects of Angptl7 on corneal nerves using culture rat trigeminal cells and this molecule had neurotrophic property on the cornea. Thus, Angptl7 is a unique molecule, which contain its bilateral character (anti-angiogenic and neurotrophic) in the cornea; an agonistic nucleic acid drug for Angptl7 may be a new therapeutic tool for protecting corneal transparency.
3. We examined local gene editing for TGFBI-related corneal dystrophy using CRISPR-Cas9 mediated homology directed repair (HDR). Cultured corneal keratocytes were obtained from a patient of R124H granular dystrophy. The R124H gene arrangement was corrected by a tranfection of guide RNA and HDR repair template single strand DNA in vitro. Thus, CRISPR-Cas9 mediated HDR could be a future radical treatment for TGFBI-related corneal dystrophy.
Nippon Ganka Gakkai Zasshi (J Jpn Ophthalmol Soc) 120: 246-263, 2016.

Key words
Cornea, Nucleic acid drug, Angiopoietin-like protein 2 (Angptl2), Angiopoietin-like protein 7 (Angptl7), Gene therapy, CRISPR-Cas9
Reprint requests to
Tomohiko Usui, M.D., Ph.D. Department of Ophthalmology, The University of Tokyo, Graduate School of Medicine and Faculty of Medicine. 7-3-1 Hongo, Bunkyo-ku, Tokyo 161-0033, Japan