Nontargeted stable integration of recombinant adeno-associated virus into human leukemia and lymphoma cell lines as evaluated by fluorescence in situhybridization

A number of studies on human epithelial cells of varying origin have demons trated integration of recombinant adeno-associated virus (AAV) vectors into a variety of chromosomes compared with the site-specific integration on ch romosome 19 predominantly observed for wild-type (wt) AAV, We have construc ted a recombinant AAV (rAAV) vector and tested the integration into hematop oietic cells, using the human acute myeloid leukemia cell line AML5 and the human non-Hodgkin's lymphoma cell line OCI-LY18 as targets. The integratio n sites were visualized by fluorescence in sial hybridization (FISH). Posit ive signals were observed for chromosomes 1, 2, 3, 8, 14, 15, 19, and Y, Th e majority of cells demonstrated integration into one specific site. A mino rity showed simultaneous integration into more than one chromosome. The fre quency of observed integrations was not uniformly distributed among chromos omes; for instance, in AML5 chromosome 2 seemed to be favored. Colony-deriv ed AML5 clones bore unique integration patterns indicating successful trans duction of clonogenic progenitor cells with high proliferative potential. T he integration was stable and observed for more than 12 months after transd uction, FISH has been shown to be a powerful tool for detailed analyses of rAAV integration patterns and can be used to evaluate targets and transduct ion conditions.