Correction of the Disease Phenotype in the Mouse Model of Stargardt Disease by Lentiviral Gene Therapy

Abstract

Abstract: : Purpose: Stargardt disease (STGD) is a macular dystrophy caused by mutations in the ABCA4 (ABCR) gene. The most recognized phenotypic feature of STGD patients and the mouse model, the Abca4^–/– mice, is lipofuscin (A2E) accumulation in retinal pigment epithelium (RPE), considered to be both the earliest sign and the cause of RPE cell death at late stages of the disease. Since there is no treatment for STDG, we tested whether delivery of the normal (wt) human ABCA4 gene to the subretinal space of the Abca4^–/– mice via lentiviral vectors would correct the disease phenotype, i.e., reduce A2E accumulation. Methods: Human immunodeficiency virus (HIV) and equine infectious anemia virus (EIAV)–derived lentiviral vectors were constructed, with the human ABCA4 gene under the control of constitutive (CMV, EF1α) or photoreceptor–specific (Rho) promoters. Similar vectors were made for the reporter gene (LacZ) to determine the transduction efficiency of retinal cells. Abca4^–/– mice were injected subretinally with 1–3x10⁵ IU of each virus at P4–5 in one eye; the mock–injected contralateral eye served as a control. Mice were sacrificed at 4, 6, and 8 months after injection and the A2E content was determined by HPLC. Results: Subretinal injections of ∼10⁵ IU of the HIV–LacZ effectively transduced RPE cells, but less effectively (<5%) photoreceptors (PR). In comparison, the same amounts of EIAV–LacZ vectors were more effective in transducing PR in both mice (10–20%) and rabbits (∼100%). Most importantly, a single subretinal injection of 3x10⁵ IU of HIV–EF1a–ABCA4 to Abca4^–/– mice practically eliminated A2E accumulation compared to mock–injected controls. ABCA4–injected eyes of Abca4^–/– mice accumulated 4–6 pmol/eye of A2E at 4, 6 and 8 months after treatment, amounts similar to wt controls of the same genetic background. By comparison, mock–treated eyes had 6–8 times more A2E (22–35 pmol/eye). Conclusions: The excessive A2E (lipofuscin) accumulation in the mouse model of STGD disease was eliminated by lentivirus–based gene therapy. While extrapolations to humans require caution and further evaluation, our results suggest that lentiviral gene therapy is a potentially efficient tool for treating ABCA4–associated diseases.

Keywords: gene transfer/gene therapy • retinal degenerations: hereditary • retina