1. Efficient gene transfer of CD40 ligand into primary B-CLL cells using recombinant adeno-associated virus (rAAV) vectors.
- Author
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Wendtner CM, Kofler DM, Theiss HD, Kurzeder C, Buhmann R, Schweighofer C, Perabo L, Danhauser-Riedl S, Baumert J, Hiddemann W, Hallek M, and Büning H
- Subjects
- Adult, B7-1 Antigen immunology, Cancer Vaccines, Cytotoxicity, Immunologic, Female, Genetic Therapy, Genetic Vectors, Green Fluorescent Proteins, HeLa Cells, Humans, Immunotherapy, In Vitro Techniques, Leukemia, Lymphocytic, Chronic, B-Cell immunology, Luminescent Proteins, Male, Middle Aged, Transduction, Genetic, CD40 Ligand genetics, Dependovirus, Leukemia, Lymphocytic, Chronic, B-Cell genetics
- Abstract
B cells of chronic lymphocytic leukemia (B-CLL) are resistant to transduction with most currently available vector systems. Using an optimized adenovirus-free packaging system, recombinant adeno-associated virus (rAAV) vectors coding for the enhanced green fluorescent protein (AAV/EGFP) and CD40 ligand (AAV/CD40L) were packaged and highly purified resulting in genomic titers up to 3 x 10(11)/mL. Cells obtained from 24 patients with B-CLL were infected with AAV/EGFP or AAV/CD40L at a multiplicity of infection (MOI) of 100 resulting in transgene expression in up to 97% of cells as detected by flow cytometry 48 hours after infection. Viral transduction could be specifically blocked by heparin. Transduction with AAV/CD40L resulted in up-regulation of the costimulatory molecule CD80 not only on infected CLL cells but also on noninfected bystander leukemia B cells, whereas this effect induced specific proliferation of HLA-matched allogeneic T cells. Vaccination strategies for patients with B-CLL using leukemia cells infected ex vivo by rAAV vectors now seems possible in the near future.
- Published
- 2002