Adeno-associated Virus Tropism in Hematopoietic Cells

Abstract

<p>Adeno-associated virus (AAV) is a small, nonenveloped, helper-dependent virus with a single stranded DNA genome. AAV is an attractive vector for gene therapy due to its lack of pathogenicity, low immunogenicity, and persistent transgene expression. However, a major limitation for AAV gene transfer is the specific tropism demonstrated by each serotype. Although there are over 100 known AAV serotypes, only serotypes 2 and 5 have been investigated in hematopoietic cells, both of which show low transduction efficiency. Thus, AAV is not currently being considered for gene transfer to blood cells. We hypothesized that other serotypes may demonstrate improved transduction over AAV2 or 5, and that transduction efficiency may be blood cell type specific. To test this, several hematopoietic cell lines (promyeloblast (HL60), monocyte (THP-lL and T lymphocyte (H9) cells) were infected with recombinant AAV serotypes 1, 2, 4, 5, 8, or 9, encoding the gene for green fluorescent protein (AAV-GFP) at various MOL Little to no toxicity was observed, as measured by trypan blue exclusion. Transduction of all serotypes in all cell types was low, with AAV5 showing the greatest transduction in HL60 cells, as determined by fluorescence microscopy and flow cytometry. Surprisingly, expression lasted for over 3 weeks despite rapid proliferation in all hematopoietic cell lines. Many AAV use cell surface polysaccharides (e.g. heparan sulfate or sialic acid) as receptors, thus, infection efficiency was correlated with fluorescentlylabeled lectin binding. Interestingly, each cell type showed distinct lectin staining that did not correlate with viral infection, suggesting postbinding barriers. Although none of the AAV serotypes investigated demonstrated transduction efficiency sufficient to achieve a clinically relevant therapeutic index, other serotypes or novel methods to modify tropism may yield vectors suitable for gene delivery in disease-associated leukocytes.</p><p>This presentation occurred at the Wright State University Campus-Wide Celebration of Research, Scholarship and Creative Activities on April 8, 2011</p>