High efficiency transduction of chronic lymphocytic leukemia with adenovirus type 35

Abstract

6612 Background: Adenovirus (Ad) vectors have been used extensively in genetic manipulation of cultured cells and gene therapy, with the group C Ad serotypes 2 and 5 being most widely used. The group C viruses have been shown to infect target cells in a two-step process. There is an initial high-affinity step in which Ad fiber protein binds to the the coxsackie/adenovirus receptor (CAR), increasing the local concentration of the virus at the cell surface. In the second step, the Ad penton base protein binds to integrins on the target cell surface and the virus is internalized. CLL cells to not express CAR and so require a very high concentration of Ad5 for transduction. Group B adenoviruses differ from other groups, having been shown to require CD46 (the measles virus receptor) to infect target cells. We are investigating Ad35 for use as a gene therapy vector and a laboratory tool for manipulating CLL cells ex vivo. Methods: We used a modified viral vector (Shayakhmetov et al., 2000 J Virol (74) pp 2567–83) with first 64 amino acids from the Ad5 fiber fused to the last 279 amino acids from the Ad35 fiber substituted into the Ad5 fiber ORF. This vector was used to assess the relative efficiency of the chimeric Ad5/35 vector compared to wild-type Ad5. Results: Primary CLL cells were transduced by the standard Ad5 and chimeric Ad5F35 vectors. Ad5F35 vector is about 100-fold more potent at transducing CLL cells compared to an Ad5 vector. Twelve CLL, PBMC and HeLa samples were tested for CD46 expression by flow cytometry. Each CLL population was 90–99% bright for CD46. Primary CLL cells and normal PBMCs were transduced by Ad5F35-GFP at an MOI of 10. The CLL cells are more transducible than normal PBMCs (p<0.05). This difference in transducibility is not a result of low CD46 expression. Conclusions: Adenovirus Type 35 is a promising vector for use in delivering gene therapy agents to CLL cells at significantly lower viral doses and more favorable therapeutic index compared with traditional Ad5 vectors. No significant financial relationships to disclose.