Exchange of surface proteins impacts on viral vector cellular specificity and transduction characteristics: the retina as a model

Abstract

Recombinant vectors based on adeno-associated virus (AAV) or human immunodeficiency 1 (lentivirus) are promising tools for long term in vivo gene delivery. Their design allows the exchange of capsids or envelopes, respectively, theoretically providing the opportunity to transduce a range of cell types. We constructed AAV vectors encoding enhanced green fluorescent protein (EGFP) within an AAV serotype 2 (AAV2) genome contained in an AAV2, five or one capsid (called AAV2/2, AAV2/5 and AAV2/1, respectively). Similarly we produced lentiviral vectors, encoding the same expression cassette present in the AAV vectors, pseudotyped with proteins from vesicular stomatitis virus glycoprotein (VSVG) or Mokola envelopes. Transduction characteristics of these vectors were evaluated in the murine retina following subretinal or intravitreal administration. The time of onset of transgene expression and the targeted cell types differed between the various recombinants. Onset of transgene expression was 3-4 days for lentiviral vectors and AAV2/1. In contrast, onset was at 2-4 weeks for AAV2/5 and AAV2/2, respectively. After subretinal injection, both lenti-VSVG and AAV2/5 transduced the retinal pigment epithelium (RPE) and photoreceptors efficiently whereas transgene expression was restricted to RPE cells using lenti with the Mokola envelope or AAV2/1. After intravitreal administration, only AAV2/2 and lenti-VSVG transduced the inner retina. Vector-mediated fluorescence was detected in the retina for over 12 weeks for all of the vectors. We conclude that pseudotyping provides a useful means to manipulate viral vector cell targeting specificity as well as retinal transduction characteristics of vectors containing the same genome.

Recombinant AAV-mediated gene transfer to the retina: gene therapy perspectives

“…In contrast, onset of transgene expression occurs at 2-4 weeks for AAV-2/2 and AAV-2/ 5 with both vectors transducing RPE and photoreceptors cells. 24 No transduced cells are detected following intravitreal injection of AAV-2/1 and AAV-2/5. Subret-inal injection of AAV-2/1, -2/2, -2/3, -2/4 and -2/5 in rats results in a hierarchy in the levels of transgene expression, with rAAV-2/4 and -2/5 capsids being the most efficient.…”

Cellular tropism and transduction properties of seven adeno-associated viral vector serotypes in adult retina after intravitreal injection

“…2 In the eye, rAAV has been used in experimental studies aimed at treating retinal degenerative diseases or for cell rescue following trauma. [3][4][5][6][7][8][9][10][11][12][13][14] On the basis of the success of some of these animal studies, clinical trials with AAV vectors have recently been initiated in humans with genetically linked ophthalmic diseases. 15,16 Human AAV serotype 2 was originally found as a contaminant in adenovirus preparations 17 and has not been associated with any human pathogenicity.…”

“…23 Variation in capsid structure affects viral tropism and expression kinetics in terms of the onset and expression levels of therapeutic genes, and the use of different serotypes may allow for a more cellspecific gene transfer. 3,6,8,12,24,25 Robust photoreceptor transduction has been reported after subretinal injection of many vector types; however, photoreceptors are only infrequently transduced after intravitreal injection of the serotypes tested to date. 3,4,[6][7][8]11,12,26 Intravitreal vector delivery is a less invasive technique compared with subretinal delivery.…”

“…28 rAAV2 vectors transduce RGCs [8][9][10][11][12] but not with absolute selectivity, 11 which may complicate gene therapy treatment of ophthalmic disease. It is, therefore, important to evaluate the tropism and transduction efficiency of other AAV serotypes to determine if there are even more efficient and specific vectors that target RGCs.…”

“…Retinas were analyzed 10 weeks post-injection, a time point at which all vector types should maintain a strong gene expression. 6,8,12,24 For each serotype, the number, morphology and laminar distribution of GFP-positive ( + ) cells were quantified using confocal and fluorescence microscopy. Phenotype was also assessed by double immunolabelling with various markers for retinal cell types.…”