Efficient gene transfer of CD40 ligand into primary B-CLL cells using recombinant adeno-associated virus (rAAV) vectors.
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.
Wendtner
Clemens-Martin
CM
Medical Clinic III, University Hospital Grosshadern and Gene Center, Ludwig-Maximilians-University, Munich, Germany.
Kofler
David M
DM
Theiss
Hans D
HD
Kurzeder
Christian
C
Buhmann
Raymund
R
Schweighofer
Carmen
C
Perabo
Luca
L
Danhauser-Riedl
Susanne
S
Baumert
Jens
J
Hiddemann
Wolfgang
W
Hallek
Michael
M
Büning
Hildegard
H
D000328:Adult; D018122:B7-1 Antigen / Q000276:immunology; D023201:CD40 Ligand / Q000235:genetics*; D019496:Cancer Vaccines; D003602:Cytotoxicity, Immunologic; D000229:Dependovirus; D005260:Female; D015316:Genetic Therapy; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D007167:Immunotherapy; D066298:In Vitro Techniques; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000235:genetics* / Q000276:immunology; D008164:Luminescent Proteins; D008297:Male; D008875:Middle Aged; D014161:Transduction, Genetic
D018122:B7-1 Antigen; D019496:Cancer Vaccines; D008164:Luminescent Proteins; D023201:CD40 Ligand; D049452:Green Fluorescent Proteins