Stable retinal gene expression in nonhuman primates via subretinal injection of SIVagm-based lentiviral vectors.
Abstract Gene therapy may hold promise as a therapeutic approach for the treatment of intractable ocular diseases, including retinitis pigmentosa (RP). Gene transfer vectors that are able to show long-lasting transgene expression in vivo are highly desirable to treat RP; however, there is a dearth of information regarding long-term transgene expression in the eyes of large animals. We previously reported that the simian immunodeficiency virus from African green monkeys (SIVagm)-based lentiviral vector showed efficient, stable, and safe retinal gene transfer, resulting in significant prevention of retinal degeneration by gene transfer of a neurotrophic factor, human pigment epithelium-derived factor (hPEDF), in rodents. Before applying this strategy in a clinical setting, we here assessed the long-lasting transgene expression of our third-generation SIVagm-based lentiviral vectors in the retinal tissue of nonhuman primates. Approximately 20-50 mul of SIV-EGFP (enhanced green fluorescent protein) or SIV-hPEDF was injected into the subretinal space via a glass capillary tube. To detect EGFP expression in the retina, we used a fluorescence fundus camera at various time points after gene transfer. Human PEDF expression was assessed by immunohistochemical analysis, Western blot assay, and enzyme-linked immunosorbent assay. The retinas demonstrated frequent EGFP expression that was preserved for at least 4 years without significant decline. The expression of hPEDF was stable, and occurred mainly in the retinal pigment epithelium. The secreted protein was detected in vitreous and aqueous humor. We thus propose that SIVagm-mediated stable gene transfer might be significantly useful for ocular gene transfer in a clinical setting.
Ikeda
Yasuhiro
Y
Department of Ophthalmology, Graduate School of Medical Sciences, Kyushu University, Fukuoka 812-8582, Japan. ymocl@pathol1.med.kyushu-u.ac.jp
Yonemitsu
Yoshikazu
Y
Miyazaki
Masanori
M
Kohno
Ri-Ichiro
R
Murakami
Yusuke
Y
Murata
Toshinori
T
Tabata
Toshiaki
T
Ueda
Yasuji
Y
Ono
Fumiko
F
Suzuki
Toshimichi
T
Ageyama
Naohide
N
Terao
Keiji
K
Hasegawa
Mamoru
M
Sueishi
Katsuo
K
Ishibashi
Tatsuro
T
D000818:Animals; D001082:Aqueous Humor / Q000378:metabolism; D015153:Blotting, Western; D002460:Cell Line; D002522:Chlorocebus aethiops / Q000235:genetics* / Q000821:virology*; D005136:Eye Proteins / Q000378:metabolism; D005786:Gene Expression Regulation; D005822:Genetic Vectors / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D007267:Injections; D016086:Lentivirus / Q000235:genetics*; D009414:Nerve Growth Factors / Q000378:metabolism; D012160:Retina / Q000166:cytology / Q000378:metabolism*; D015843:Serpins / Q000378:metabolism; D015302:Simian Immunodeficiency Virus / Q000502:physiology*; D014822:Vitreous Body / Q000378:metabolism; D000099174:Pigment Epithelium-Derived Factor
D005136:Eye Proteins; D049452:Green Fluorescent Proteins; D009414:Nerve Growth Factors; D015843:Serpins; D000099174:Pigment Epithelium-Derived Factor; C485184:enhanced green fluorescent protein