Efficient gene transfer of CD40 ligand into primary B-CLL cells using recombinant adeno-associated virus (rAAV) vectors

• Published in: Blood Vol. 100; no. 5; pp. 1655 - 1661
• Main Authors: Wendtner, Clemens-Martin, Kofler, David M., Theiss, Hans D., Kurzeder, Christian, Buhmann, Raymund, Schweighofer, Carmen, Perabo, Luca, Danhauser-Riedl, Susanne, Baumert, Jens, Hiddemann, Wolfgang, Hallek, Michael, Büning, Hildegard
• Format: Journal Article
• Language: English
• Published: Washington, DC Elsevier Inc 01.09.2002; The Americain Society of Hematology
• Subjects: Adult; Anesthesia. Intensive care medicine. Transfusions. Cell therapy and gene therapy; Applied cell therapy and gene therapy; B7-1 Antigen - immunology; Biological and medical sciences; Cancer Vaccines; CD40 Ligand - genetics; Cytotoxicity, Immunologic; Dependovirus; Female; Genetic Therapy; Genetic Vectors; Green Fluorescent Proteins; HeLa Cells; Hematologic and hematopoietic diseases; Humans; Immunotherapy; In Vitro Techniques; Leukemia, Lymphocytic, Chronic, B-Cell - genetics; Leukemia, Lymphocytic, Chronic, B-Cell - immunology; Leukemias. Malignant lymphomas. Malignant reticulosis. Myelofibrosis; Luminescent Proteins; Male; Medical sciences; Middle Aged; Transduction, Genetic; Transfusions. Complications. Transfusion reactions. Cell and gene therapy; Antineoplastic agent; Human; Adenoviridae; Malignant hemopathy; B-Lymphocyte; Vaccine; Dependovirus; Costimulatory molecule; Transgene; Genetic transfer; Virus; Chronic; Chronic lymphocytic leukemia; Parvoviridae; Lymphoproliferative syndrome; CD40 ligand; Transduction; Technique; Gene therapy; Vector; Parvovirinae
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood.V100.5.1655.h81702001655_1655_1661

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.