Gene therapy for choroideremia using an adeno-associated viral (AAV) vector

Abstract

Choroideremia is an outer retinal degeneration with a characteristic clinical appearance that was first described in the nineteenth century. The disorder begins with reduction of night vision and gradually progresses to blindness by middle age. The appearance of the fundus in sufferers is recognizable by the characteristic pale color caused by the loss of the outer retina, retinal-pigmented epithelium, and choroidal vessels, leading to exposure of the underlying sclera. Choroideremia shows X-linked recessive inheritance and the choroideremia gene (CHM) was one of the first to be identified by positional cloning in 1990. Subsequent identification and characterization of the CHM gene, which encodes Rab escort protein 1 (REP1), has led to better comprehension of the disease and enabled advances in genetic diagnosis. Despite several decades of work to understand the exact pathogenesis, no established treatments currently exist to stop or even slow the progression of retinal degeneration in choroideremia. Encouragingly, several specific molecular and clinical features make choroideremia an ideal candidate for treatment with gene therapy. This work describes the considerations and challenges in the development of a new clinical trial using adeno-associated virus (AAV) encoding the CHM gene.

Figure 1.

A fundus photograph of a patient with advanced choroideremia. Patient eye (left) compared with a normal eye (right). In the choroideremia eye, there is a residual island of healthy tissue in the central macula and fovea.

Figure 2.

A schematic of the cellular function of Rab escort protein (REP1). REP1 binds Rabs (member of the Ras subfamily of small GTPases, ∼70 members) as they are made by ribosomes (R) on the endoplasmic reticulum (ER). REP1 present newly synthesized, unprenylated Rabs to a catalytic Rab geranylgeranyltransferase (GGTase) subunit. Rabs undergo prenylation, which involves the covalent attachment of one or more hydrophobic prenyl groups (geranylgeranyl, GG shown here) to carboxy-terminal cysteine residues. REP1 also escorts prenylated Rabs to specific destination membranes in the Golgi apparatus (GA) and of diverse cellular vesicles (Vesicle, represents various cellular vesicles, e.g., lysosomes or transport vesicles).

Figure 3.

Design of the AAV2/2-REP1 vector used for choroideremia gene therapy. ITR, AAV2 inverted terminal repeats; CMV, cytomegalovirus enhancer element; CBA, chicken β actin promoter; Exon1, chicken β-actin exon 1; Intron, hybrid chicken β-actin and rabbit β-globin intron; Exon2, rabbit β-globin exon 3 fragment (creates an artificial splice site) and human REP1 cDNA (REP1) containing Kozak consensus sequence and start codon initiating translation (Kozak/start) and stop codon terminating translation (stop); WPRE*, modified woodchuck hepatitis virus posttranscriptional regulatory element; bGH pA, bovine growth hormone polyadenylation signal.