1. ¹Jules Stein Eye Institute, Molecular Biology Institute, UCLA School of Medicine, Los Angeles, CA, USA
2. ²Departments of Pharmacology and Neurosciences, UCSD School of Medicine, La Jolla, CA, USA

Correspondence: Dr DS Williams, Department of Pharmacology, UCSD School of Medicine, Mail code 0912, 9500 Gilman Drive, La Jolla, CA 92093-0912, USA. E-mail: dswilliams@ucsd.edu; Dr X-J Yang, Jules Stein Eye Institute, UCLA School of Medicine, 100 Stein Plaza, Los Angeles, CA 90095, USA. E-mail: yang@jsei.ucla.edu

³These authors contributed equally to this work.

Received 20 July 2006; Revised 25 October 2006; Accepted 25 October 2006; Published online 1 February 2007.

Abstract

One of the most disabling forms of retinal degeneration occurs in Usher syndrome, since it affects patients who already suffer from deafness. Mutations in the myosin VIIa gene (MYO7A) cause a major subtype of Usher syndrome, type 1B. Owing to the loss of function nature of Usher 1B and the relatively large size of MYO7A, we investigated a lentiviral-based gene replacement therapy in the retinas of MYO7A-null mice. Among the different promoters tested, a CMV-MYO7A chimeric promoter produced wild-type levels of MYO7A in cultured RPE cells and retinas in vivo. Efficacy of the lentiviral therapy was tested by using cell-based assays to analyze the correction of previously defined, MYO7A-null phenotypes in the mouse retina. In vitro, defects in phagosome digestion and melanosome motility were rescued in primary cultures of RPE cells. In vivo, the normal apical location of melanosomes in RPE cells was restored, and the abnormal accumulation of opsin in the photoreceptor connecting cilium was corrected. These results demonstrate that a lentiviral vector can accommodate a large cDNA, such as MYO7A, and mediate correction of important cellular functions in the retina, a major site affected in the Usher syndrome. Therefore, a lentiviral-mediated gene replacement strategy for Usher 1B therapy in the retina appears feasible.