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Blood, 1 September 2002, Vol. 100, No. 5, pp. 1655-1661
GENE THERAPY
Efficient gene transfer of CD40 ligand into primary B-CLL cells
using recombinant adeno-associated virus (rAAV) vectors
Clemens-Martin Wendtner,
David M. Kofler,
Hans D. Theiss,
Christian Kurzeder,
Raymund Buhmann,
Carmen Schweighofer,
Luca Perabo,
Susanne Danhauser-Riedl,
Jens Baumert,
Wolfgang Hiddemann,
Michael Hallek, and
Hildegard Büning
From the Medical Clinic III, University Hospital
Grosshadern and Gene Center, Ludwig-Maximilians-University; KKG Gene
Therapy, GSF, National Research Center for Environment and Health; IBE,
Institute for Biometry and Epidemiology, Ludwig-Maximilians-University,
Munich, Germany.
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 × 1011/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.
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